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Bartolome, Amelita, Anna Almazan, Salina Abusali, Stanley Tam, Eric Lee, Amogh Changavi, Wendy Trinh et al. "1089. Analytical Performance of an Ultrasensitive Immunoassay for Detection of Clostridium difficile Toxins in Stool". Open Forum Infectious Diseases 5, suppl_1 (novembre 2018): S326. http://dx.doi.org/10.1093/ofid/ofy210.924.
Testo completoAbstract (sommario):
Abstract Background Clostridium difficile infection (CDI) is the main cause for nosocomial diarrhea. Currently available assays for the diagnosis of CDI show deficits in sensitivity, specificity, and/or turnaround time. The Singulex Clarity® C. diff toxins A/B assay, in development for the Singulex Clarity® system, was designed to provide an accurate and automated detection of C. difficile toxins A (TcdA) and B (TcdB) in stool. Here, the analytical performance of the assay is reported. Methods Limits of detection (LoD) for TcdA and TcdB in stool and buffer was determined, and a preliminary cutoff, as compared with cell cytotoxicity neutralization assay (CCNA), was established. Analytical reactivity against 38 toxigenic and nontoxigenic C. difficile strains of eight different toxinotypes was determined. Cross-reactivity against 53 other gastrointestinal pathogens and potential interference by 11 endogenous and exogenous substances were determined. Reproducibility was tested with triplicate samples (n = 85), and stability was evaluated in samples stored at room temperature, refrigerated, and frozen conditions, and subjected to three freeze-thaw cycles. Results The LoDs for TcdA and TcdB were 0.8 and 0.3 pg/mL in buffer, and 2.0 and 0.7 pg/mL in stool, respectively. Using a preliminary cutoff, the assay demonstrated 96.3% sensitivity and 96.1% specificity compared with CCNA. The Singulex Clarity® C. diff toxins A/B assay detected toxins from all tested strains and toxinotypes. No cross-reactivity or interference were detected. The repeatability was 99%, and samples for C. difficile toxin testing were stable up to 8 hours in room temperature, 1 week in 2–8°C, 6 months in −70°C, and up to three freeze–thaw cycles. Conclusion The Singulex Clarity C. diff toxins A/B assay (in development) can detect TcdA and TcdB at very low concentrations and it has high sensitivity and specificity compared with CCNA. The assay demonstrates reactivity to common C. difficile strains, does not show cross-reactivity to common gastrointestinal pathogens, is robust against common interferents, allows for toxin detection in both fresh and frozen stool samples and up to three freeze–thaw cycles, and provides results with high reproducibility. Disclosures A. Bartolome, Singulex, Inc.: Employee, Salary. A. Almazan, Singulex, Inc.: Employee, Salary. S. Abusali, Singulex, Inc.: Employee, Salary. S. Tam, Singulex, Inc.: Employee, Salary. E. Lee, Singulex, Inc.: Employee, Salary. A. Changavi, Singulex, Inc.: Employee, Salary. W. Trinh, Singulex, Inc.: Employee, Salary. K. Chau, Singulex, Inc.: Employee, Salary. J. Estis, Singulex, Inc.: Employee, Salary. B. Noland, Singulex, Inc.: Employee, Salary. J. Bishop, Singulex, Inc.: Employee, Salary.
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Gomperts, Edward D., Shashikant Apte, Utpal Chaudhuri, Joseph M. John, Vijay Ramanan, Ri Liesner, Amy D. Shapiro, Bonnie J. Mills e Martin Lee. "IB1001, an Investigational Recombinant Factor IX, Pharmacokinetics in Pediatric Patients with Hemophilia B." Blood 120, n. 21 (16 novembre 2012): 2225. http://dx.doi.org/10.1182/blood.v120.21.2225.2225.
Testo completoAbstract (sommario):
Abstract Abstract 2225 Introduction IB1001 is a recombinant factor IX product being investigated for the treatment and prevention of bleeding in individuals with hemophilia B. Pharmacokinetics (PK) in adults (>12 years) demonstrated that IB1001 had results similar to the currently available recombinant FIX with respect to parameters such as terminal phase half-life and incremental recovery. We report the interim findings from a PK assessment in children <12 years, with severe hemophilia B (FIX <2%), >50 prior exposure days to FIX, and no history of or currently detectable inhibitor to FIX. Methods Non-randomized, open-label PK study with patients receiving 75±5 IU/kg of IB1001 following a washout period of ≥4 days from a previous FIX infusion. Factor IX levels were determined pre-infusion and at 15–30 minutes, 4–6, 24–26, and 68–72 hours post-infusion. Additional samples could be drawn at 1–3 and 10–14 hours. Calculated PK parameters were: half-life (β-phase t1/2, determined using a robust regression approach [Lee ML et al. XVIth ISTH Congress, Florence, Italy, 1997]) but generally assuming a single compartmental model because of the small number of points, maximum plasma concentration (Cmax), in vivo recovery (IVR) and AUC(0-∞) (determined by the trapezoidal rule). In addition, the AUC(0-t) and mean residence time (MRT) were calculated. Results When compared to the findings previously reported with IB1001 in adult (≥12 years of age) subjects (Martinowitz U et al. Haemophilia, 18, 2012), the results in pediatric patients demonstrate a more rapid metabolism of factor IX as is indicated by the shorter terminal half-life (mean±SD of 19.3±7.8 h versus 29.6±18.2 h in adults) and the smaller AUC0-∞ (mean±SD of 1059±264 versus 1668±598 in adults). In addition, the in vivo recovery was lower (mean±SD of 0.69±0.21) versus that seen in adults (mean±SD of 0.98±0.22). These results are similar to those reported by Berntorp et al (Haemophilia, 7, 2001) with nonacog alfa. Conclusions The pharmacokinetics of IB1001 has previously been shown to be non-inferior to nonacog alfa, another recombinant factor IX, in hemophilia B individuals >12 years of age. The current study is intended to provide information on children <12 and, particularly, <6 years of age. IB1001 is metabolized faster and has a lower recovery than the comparable findings in patients >12 years of age. Although the study is ongoing, these may represent important implications for the potential use of IB1001 in pediatric patients. Disclosures: Gomperts: Inspiration Biopharmaceuticals Inc: Consultancy. Apte:Inspiration Biopharmacauticals Inc: Research Funding. Chaudhuri:Inspiration Biopharmaceuticals Inc: Research Funding. John:Inspiration Biopharmaceuticals Inc: Research Funding. Ramanan:Inspiration Biopharmaceuticals Inc: Research Funding. Liesner:Inspiration Biopharmaceuticals Inc: Research Funding. Shapiro:Inspiration Biopharmaceuticals Inc: Honoraria, Research Funding. Mills:Inspiration Biopharmaceuticals Inc: Employment. Lee:Inspiration Biopharmaceuticals Inc: Employment.
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Karasiewicz, Kathy, Shuyang He, Mary Ng, Kristina Tess, Weifang Ling, Gunnar F. Kaufmann, Jerome B. Zeldis, Henry Ji, Robert Hariri e Xiaokui Zhang. "Preclinical Evaluation of Human Placental-Derived Allogeneic CD19 CAR-T Cells Against B Cell Malignancies". Blood 134, Supplement_1 (13 novembre 2019): 3222. http://dx.doi.org/10.1182/blood-2019-130782.
Testo completoAbstract (sommario):
Celularity, Inc. is developing a CD19 CAR-T Cell therapy using an allogeneic platform derived from postpartum human placental cells. T cells isolated from placenta/ umbilical cord blood and genetically modified to express CD19 chimeric antigen receptor (CAR), termed Placental-derived (P-) CD19 CAR T cells, are in development for the treatment of B cell malignancies. Unlike adult peripheral blood mononuclear cell (PBMC)-derived T cells, P-T cells are mostly naïve (CD45RA+) and can be readily expanded while maintaining an earlier differentiation phenotype such as greater expression of naïve/ memory markers, lower expression of effector/ exhaustion markers, allowing for greater proliferative potential of these cells ex vivo. These cells are also known to have greater immune tolerance to HLA mismatch and display impaired allogeneic activation, contributing to lower incidences of severe graft-verse-host disease (GvHD) (Barker, et. al. Blood, 2001; Chen, et al. Biology of Blood and Marrow Transplantation, 2006), making them an attractive cell population for use as an allogeneic, adoptive cell therapy. A robust process for the isolation, transduction, and expansion of placental-derived T cells to generate "off-the-shelf" allogeneic P-CD19 CAR T cells was developed. Twenty-One day expanded, non-modified P-T cells (N=3) were compared to adult PBMCs for their allo-reactivity in a Xenogeneic GvHD model in NCG mice. P-T cells did not induce xeno-GvHD whereas PBMCs did, as evidenced by significant weight loss and death of all mice (N=5) by Day 28 post infusion. Despite expanded P-T cells demonstrating lack of in vivo GvHD, current manufacture of P-CD19 CAR T cells does include a CRISPR-mediated T-cell receptor a constant (TRAC) knockout (KO) step as an additional risk-mitigation strategy to circumvent any potential GvHD stemming from expression of endogenous T cell receptor. CD19 CAR transduction using a retrovirus provided by Sorrento Therapeutics, Inc., followed by TRAC knockout with CRISPR results in both high efficiency of CD19 CAR expression (~30% CD19 Fc+) and TCR KO (>96% CD3-/ TCR a/b-). In vitro, the functional activity of P-CD19 CAR-TRAC KO T cells against CD19+ Burkitt's Lymphoma (Daudi) and Acute lymphoblastic Leukemia (NALM6) cell lines was assessed in cytotoxicity and cytokine release assays. P-CD19 CAR T cells specifically lyse CD19+ Daudi/ Nalm6 targets in both 4-hour endpoint FACS and ACEA kinetic cytotoxicity assays, and in most cases at levels equivalent to or greater than PBMC-derived CD19 CAR T cells. When P-CD19 CAR T cells were co-cultured with CD19+ Daudi/ Nalm6 target cells for 24-hours, they secreted pro-inflammatory cytokines and effector proteins in an antigen-specific manner. In vivo, the anti-tumor activity of P-CD19 CAR T cells was assessed using a disseminated lymphoma xenograft model in NSG mice. Luciferase expressing Daudi cells (3×106) were intravenously (IV) injected on Day 0, followed by IV injection of P-CD19 CAR T cells (14×106) on Day 7. Bioluminescence Imaging (BLI) and survival were used as primary study endpoints. P- CD19 CAR T cells were well tolerated and safe. P-CD19 CAR T cells significantly reduced tumor burden, and improved survival. Four weeks after treatment, the vehicle group had a 100% mortality rate, while all animals from P-CD19 CAR T-treated group (N=5) remained alive without clinical symptoms including weight loss or changes in their fur. In summary, Celularity has defined a robust process for the generation and expansion of CD19 CAR T cells from human placenta. These cells exhibit potent anti-tumor activity both in vitro and in vivo with little evidence of acute GvHD induction, highlighting their potential as an allogeneic, adoptive cell therapeutic agent. Future in vivo GvHD studies will include assessment of both CD19 CAR and TRAC KO genetically modified P-T cells. Disclosures Karasiewicz: Celgene: Equity Ownership; Celularity, Inc.: Employment, Equity Ownership, Patents & Royalties: Patent Inventor. He:Celularity Inc: Employment. Ng:Celularity, Inc.: Employment. Tess:Celularity, Inc.: Employment. Ling:Celularity Inc: Employment. Kaufmann:Sorrento Therapeutics, Inc.: Employment, Equity Ownership, Patents & Royalties. Zeldis:Sorrento Therapeutics Inc: Employment, Equity Ownership. Ji:Celularity, Inc.: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Sorrento Therapeutics Inc: Employment, Equity Ownership, Patents & Royalties. Hariri:Celularity Inc: Employment. Zhang:Celularity Inc: Employment.
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Coyne-Beasley, Tamera, Joseph Bocchini, Alejandro Cane, Cindy Burman, Maria J. Tort e Jessica Presa. "01. Serum Bactericidal Activity Against Circulating and Reference Strains of Meningococcal Serogroup B in the United States: A Review of Meningococcal Serogroup B (MenB) Vaccines in Adolescents and Young Adults". Open Forum Infectious Diseases 8, Supplement_1 (1 novembre 2021): S124—S125. http://dx.doi.org/10.1093/ofid/ofab466.204.
Testo completoAbstract (sommario):
Abstract Background US adolescents and young adults are at particular risk of invasive meningococcal disease (IMD). In 2018, menincococcal serogroup B was responsible for 36% of IMD cases in the US overall and for 66% of cases in adolescents and young adults. This age group is at high risk of IMD during outbreaks, which result in significant response-related costs. MenB vaccine efficacy against IMD relies on its ability to provide broad protection against diverse disease-causing strains. MenB-FHbp (Trumenba) and MenB-4C (Bexsero) are MenB vaccines licensed in the US as 2-dose series with an interval of 6 mo or 1 mo, respectively, recommended in healthy adolescents and young adults. We review available data on vaccine coverage of serogroup B strains. Methods A literature review identified relevant information from peer-reviewed publications, congress presentations, and ClinicalTrials.gov. Previously presented but unpublished data from phase 2/3 studies were included. Results After 2 MenB-FHbp doses, percentages of adolescents and young adults achieving serum bactericidal activity assay using human complement (hSBA) titers ≥1:8 were 79%–99% for 4 heterologous representative test strains and 71%–97% for 10 additional strains, confirming cross-protection against a diverse strain panel (Figure 1; unpublished data). These 14 heterologous strains collectively represent ~80% of disease-causing strains in the US and Europe. In a published study with limited sample size, 44%–78% of subjects had hSBA titers ≥1:8 against strains from 4 US college outbreaks after 2 MenB-FHbp doses. After 2 MenB-4C doses, percentages of 10–25-year-olds achieving hSBA titers ≥1:5 against 3 reference strains homologous to the vaccine antigen were 82%–93% (published data); 15%–100% of adolescents achieved hSBA titers ≥1:4 against a panel of 14 strains (unpublished data). Of college students who received 2 MenB-4C doses, 53%–93% achieved hSBA titers ≥1:4 against 5 US outbreak strains (4/5 strains had antigenic similarity to MenB-4C; published data). Conclusion MenB-FHbp and MenB-4C protect against various serogroup B strains. As for the breadth of coverage provided by these vaccines, available data show that MenB-FHbp elicits robust immune responses to a wide variety of disease-causing strains prevalent in the US (Figure 2). Disclosures Tamera Coyne-Beasley, MD, MPH, Pfizer Inc and GlaxoSmithKline (Advisor or Review Panel member) Joseph Bocchini, MD, Pfizer Inc and Dynavax (Advisor or Review Panel member) Alejandro Cane, M.D., Pfizer Inc (Employee, Shareholder) Cindy Burman, PharmD, Pfizer Inc (Employee, Shareholder) Maria J. Tort, PhD, Pfizer Inc (Employee, Shareholder) Jessica Presa, MD, Pfizer Inc (Employee, Shareholder)
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Lasater, Elisabeth A., An D. Do, Luciana Burton, Yijin Li, Erin Williams, Jason Oeh, Luciana Molinero et al. "Resistance of Natural Killer and T Cells to Venetoclax Allows for Combination Treatment with Cancer Immunotherapy Agents". Blood 132, Supplement 1 (29 novembre 2018): 1118. http://dx.doi.org/10.1182/blood-2018-99-116405.
Testo completoAbstract (sommario):
Abstract Introduction: Intrinsic apoptosis is regulated by the BCL-2 family of proteins, which consists of both anti-apoptotic (BCL-2, BCL-XL, MCL-1) and pro-apoptotic (BIM, BAX, BAK, BAD) proteins. Interaction between these proteins, as well as stringent regulation of their expression, mediates cell survival and can rapidly induce cell death. A shift in balance and overexpression of anti-apoptotic proteins is a hallmark of cancer. Venetoclax (ABT-199/GDC-0199) is a potent, selective small molecule BCL-2 inhibitor that has shown preclinical and clinical activity across hematologic malignancies and is approved for the treatment of chronic lymphocytic leukemia with 17p deletion as monotherapy and in combination with rituximab. Objective: To investigate the effects of BCL-2 inhibition by venetoclax on viability and function of immune-cell subsets to inform combinability with cancer immunotherapies, such as anti-PD-L1. Methods and Results: B cells, natural killer (NK) cells, CD4+ T cells, and CD8+ T cells in peripheral blood mononuclear cells (PBMCs) from healthy donors (n=3) were exposed to increasing concentrations of venetoclax that are clinically achievable in patients, and percentage of live cells was assessed by flow-cytometry using Near-IR cell staining. B cells were more sensitive to venetoclax (IC50 of ~1nM) than CD8+ T cells (IC50 ~100nM), NK cells (IC50 ~200nM), and CD4+ T cells (IC50 ~500nM) (Figure A). CD8+ T-cell subset analysis showed that unstimulated naive, but not memory cells, were sensitive to venetoclax treatment (IC50 ~30nM and 240nM, respectively). Resistance to venetoclax frequently involves compensation by other BCL-2 family proteins (BCL-XL and MCL-1). As assessed by western blot in PBMCs isolated from healthy donors (n=6), BCL-XL expression was higher in NK cells (~8-fold) and CD4+ and CD8+ T cells (~2.5-fold) than in B cells (1X). MCL-1 protein expression was higher only in CD4+ T cells (1.8-fold) relative to B cells. To evaluate the effect of venetoclax on T-cell function, CD8+ T cells were stimulated ex vivo with CD3/CD28 beads, and cytokine production and proliferation were assessed. Venetoclax treatment with 400nM drug had minimal impact on cytokine production, including interferon gamma (IFNg), tumor necrosis factor alpha (TNFa), and IL-2, in CD8+ effector, effector memory, central memory, and naïve subsets (Figure B). CD8+ T-cell proliferation was similarly resistant to venetoclax, as subsets demonstrated an IC50 >1000nM for venetoclax. Taken together, these data suggest that survival of resting NK and T cells in not impaired by venetoclax, possibly due to increased levels of BCL-XL and MCL-1, and that T-cell activation is largely independent of BCL-2 inhibition. To evaluate dual BCL-2 inhibition and PD-L1 blockade, the syngeneic A20 murine lymphoma model that is responsive to anti-PD-L1 treatment was used. Immune-competent mice bearing A20 subcutaneous tumors were treated with clinically relevant doses of venetoclax, murine specific anti-PD-L1, or both agents. Single-agent anti-PD-L1 therapy resulted in robust tumor regression, while single-agent venetoclax had no effect. The combination of venetoclax and anti-PD-L1 resulted in efficacy comparable with single-agent anti-PD-L1 (Figure C), suggesting that BCL-2 inhibition does not impact immune-cell responses to checkpoint inhibition in vivo. These data support that venetoclax does not antagonize immune-cell function and can be combined with immunotherapy targets. Conclusions: Our data demonstrate that significant venetoclax-induced cell death at clinically relevant drug concentrations is limited to the B-cell subset and that BCL-2 inhibition is not detrimental to survival or activation of NK- or T-cell subsets. Importantly, preclinical mouse models confirm the combinability of BCL-2 and PD-L1 inhibitors. These data support the combined use of venetoclax and cancer immunotherapy agents in the treatment of patients with hematologic and solid tumor malignancies. Figure Figure. Disclosures Lasater: Genentech Inc: Employment. Do:Genentech Inc: Employment. Burton:Genentech Inc: Employment. Li:Genentech Inc: Employment. Oeh:Genentech Inc: Employment. Molinero:Genentech Inc: Employment, Equity Ownership, Patents & Royalties: Genentech Inc. Penuel:Genentech Inc: Employment. Sampath:Genentech Inc: Employment. Dail:Genentech: Employment, Equity Ownership. Belvin:CytomX Therapeutics: Equity Ownership. Sumiyoshi:Genentech Inc: Employment, Equity Ownership. Punnoose:Roche: Equity Ownership; Genentech Inc: Employment. Venstrom:Genentech Inc: Employment. Raval:Genentech Inc: Consultancy, Employment, Equity Ownership.
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He, Jie, Jose A. Bufill, Michelle K. Nahas, Kai Wang, Lauren E. Young, Kristina M. Knapp, Kristina W. Brennan et al. "Clinical Utility of Comprehensive Profiling of Genomic Alterations in Hematologic Malignancies". Blood 124, n. 21 (6 dicembre 2014): 1072. http://dx.doi.org/10.1182/blood.v124.21.1072.1072.
Testo completoAbstract (sommario):
Abstract Background: The clinical utility of mutation profiling in hematologic malignancies requires robust detection of all classes of genomic alterations in a single analysis across different tumor types. We developed a novel, hybrid capture-based NGS assay (FoundationOne® Heme) designed to comprehensively assess the genomic landscape of hematologic malignancies from archived FFPE, blood and bone marrow aspirate samples, sequencing both DNA and RNA to improve sensitivity for driver fusion events. We have analyzed the genomic alterations including substitutions, indels, copy number alterations and genomic rearrangements of 795 samples with various hematolymphoid malignancies including lymphoma, leukemia and multiple myeloma to demonstrate the diagnostic, therapeutic and prognostic utility of this test in routine clinical practice. Method: Genomic profiles of 746 out of 795 (94%) consecutive samples received in a CLIA-certified laboratory (Foundation Medicine) were successfully characterized by the FoundationOne® Heme test including 527 from bone marrow aspirates, 176 FFPE samples and 92 samples derived from blood. The clinical cohort is composed of 375 multiple myeloma cases, 185 leukemia cases, 150 lymphoma cases, 75 MDS/MPN cases, and 15 unknown hematologic disorder cases (Figure A). Adaptor-ligated sequencing libraries were captured by solution hybridization using a custom bait-set targeting 405 blood cancer-related genes and 31 frequently rearranged genes by DNA-seq, and 265 frequently-rearranged genes by RNA-seq. All captured libraries were sequenced to high depth (Illumina HiSeq), averaging 498x for DNA and ~7M on-target unique pairs for RNA, to enable the sensitive and specific detection of substitutions, indels, copy number alteration and gene rearrangements. Results: 705/746 (95%) patients had at least one alteration reported as a somatic driver mutation, 68% of cases harbored at least one alteration linked to a targeted therapy or would inform enrollment in a clinical trial consistent with therapeutic relevance. These potentially actionable alterations included mutations in KRAS (14% of cases), NRAS (13%), CDKN2A (8%), DNMT3A (6%), IDH1/2 (5%), BRAF (4%) and FLT3 (3%) (Figure B). In addition, 64% of cases harbored at least one alteration that have been shown to predict outcome and have prognostic relevance, including TP53 (14% of cases), TET2 (7%), ASXL1 (7%), CDKN2B (5%), CREBBP (5%), MLL (5%) and NPM1 (3%)(Figure B). We also detected 344 genomic rearrangements in 280/746 (38%) patients. Of the 344 reported rearrangements, 166 were known fusions in various disease types and 178 were novel rearrangements involving known oncogenes and tumor suppressor genes, including a novel RCSD1-ABL2 fusion in a patient with B-cell ALL who will derive benefit from kinase inhibitor therapy as part of their anti-leukemic regimen. Genomic rearrangements (detected in 38% of cases) can lead to various functional consequences: 260 (76%) rearrangements detected in this cohort are predicted to create activating fusions; 19 (6%) rearrangements are predicted to be activating intragenic rearrangements in MLL, FLT3 and CARD11; and 60 (17%) rearrangements are likely truncating events in tumor suppressors. Conclusions: We demonstrated that hybrid capture-based targeted DNA and RNA sequencing can be used to identify all classes of genomic alterations in genes known to be therapeutic targets or prognostic predictors in a broad spectrum of hematologic malignancies. Moreover, our data suggests that the FoundationOne® Heme comprehensive genomic profiling test can alter therapeutic strategy in routine clinical practice. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures He: Foundation Medicine: Employment, Equity Ownership. Nahas:Foundation Medicine, Inc: Employment. Wang:Foundation Medicine, Inc.: Employment, Equity Ownership. Young:Foundation Medicine: Employment, Equity Ownership. Brennan:Foundation Medicine, Inc: Employment. Donahue:Foundation Medicine: Employment. Young:Foundation Medicine, Inc: Employment. Ross:Foundation Medicine, Inc.: Employment, Equity Ownership. Morosini:Foundation Medicine, Inc. : Employment, Equity Ownership. van den Brink:Foundation Medicine, Inc: Consultancy. Intlekofer:Foundation Medicine, Inc: Consultancy. Dogan:Foundation Medicine, Inc: Consultancy. Armstrong:Epizyme, Inc: Consultancy. Yelensky:Foundation Medicine, Inc.: Employment, Equity Ownership. Otto:Foundation Medicine, Inc.: Employment. Lipson:Foundation Medicine, Inc.: Employment, Equity Ownership. Stephens:Foundation Medicine, Inc.: Employment, Equity Ownership. Miller:Foundation Medicine, Inc: Employment. Levine:Foundation Medicine, Inc: Consultancy.
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Ge, Jianning, Chunhui Yang, Jing Sun, Jiao Chen, Shuyi Qiu, Wenjie Yin, Lianjun Shen, Martina Sersch, Wei Cao e Xinxin Wang. "Preclinical Results of an Allogeneic, Universal CD19/CD7-Targeting CAR-T Cell Therapy (GC502) for B Cell Malignancies". Blood 138, Supplement 1 (5 novembre 2021): 1722. http://dx.doi.org/10.1182/blood-2021-148500.
Testo completoAbstract (sommario):
Abstract Background Autologous CD19 CAR-T therapies show very promising clinical efficacy, but are limited in their applicability by several factors including cost, time to manufacture, and other factors involving patients own T-cell qualities. GC027, a CD7 targeting allogeneic, universal CAR-T (UCAR-T) currently in development for the treatment of T-cell acute lymphoblastic leukemia (T-ALL) has demonstrated robust expansion and anti-leukemia efficacy with a manageable safety profile in an investigator-initiated trial in China. These data suggest that, a single CD7 targeting CAR-T therapy is able to generate a therapeutic window by suppressing host vs graft (HvG) rejection of UCAR-T cells by patients' own NK and T cells, and achieve efficacy in patients with T-ALL. Based on these findings we developed GC502, a CD19/CD7 dual-targeting, allogeneic CAR-T therapy for B-cell malignancies, in which the CD19 CAR moiety targets malignant cells while CD7 CAR moiety suppresses HvG in variety of preclinical models. Methods GC502 was manufactured using leukopaks from HLA-unmatched healthy donors. It contains a 4-1BB based 2 nd-generation dual targeting CAR comprising an anti-CD19 and an anti-CD7 single-chain variable fragments (scFvs). TRAC and CD7 loci were disrupted to avoid graft vs host disease and fratricide, respectively. To select the leading CAR candidate, CAR expression and functionalities of CAR constructs with different heavy-light (H-L) chain orientations of the dual CAR were analyzed via in vitro assays and mouse xenograft models, in comparing to single CD19 CAR and CD7 CAR products. To achieve optimal anti-tumor efficacy, a T-cell enhancer was included in the CAR construct. Result Gene editing and dual CAR orientation selection TRAC and CD7 double knockout efficiencies were constantly above 97% across multiple donor pan T cells. Although CD19/CD7 CAR expression levels in different H-L chain orientations were similar, in the final CAR-T product as measured by flow cytometry (FCM) analysis, significant difference was observed in their cytotoxicity and in vitro expansion under repeated antigen stimulations by CD19+ B-cell acute lymphoblastic leukemia (B-ALL) cell line Nalm6 and CD7+ T-cell line CCRF-CEM. CAR candidates mediated the strongest cytotoxicity and most durable response were selected for further optimizations. CAR construct optimizations For the leading candidates, we first assessed the dual CAR efficacy after incorporation of an enhancer. While the IL-2, TNFα and IFNγ secretion levels were comparable, enhancer addition significantly improved tumor killing and CAR-T cell expansion under repeated stimulations by either CD19+ or CD7+ target cells. Anti-leukemia response under sub-optimal CAR-T cell dosages were also greatly enhanced as assessed by both B-ALL and T-ALL mouse xenograft models. GC502 CAR functionality comparison to single CAR products with proven clinical efficacies GC502 and GC027 were compared for their CD7 CAR function to assess their anti-HvG activities. GC502 and GC027 exhibited comparable toxicities towards pan T cells and similar efficacies in a highly malignant T-ALL mouse model. The CD19 CAR functionality of GC502 were evaluated and compared to a 2 nd generation CD19 CAR product comprising a FMC63 scFv and a 4-1BB-CD3ȥ signaling domain. In a Raji based B-ALL mouse xenograft model, both products rapidly eliminated cancer cells. While CD19 CAR treated mice showed signs of relapse at 2 weeks post CAR-T infusion, GC502 treatment group maintained "leukemia free" status till the end of study (Day28). Conclusion GC502 was optimized for CD19/CD7 dual CAR functionality and in vivo durability. It demonstrated robust anti-tumor efficacy and promising potentials to suppress HvG. This report presents an example that the dual CAR design of GC502 may serve as a novel "off-the-shelf" CAR-T technology. Disclosures Ge: GracellBiotechnologies Ltd: Current Employment, Current equity holder in publicly-traded company. Yang: GracellBiotechnologies Inc: Current Employment, Current equity holder in publicly-traded company. Sun: GracellBiotechnologies Inc: Current Employment, Current equity holder in publicly-traded company. Chen: GracellBiotechnologies Inc: Current Employment, Current equity holder in publicly-traded company. Qiu: GracellBiotechnologies Inc: Current Employment, Current equity holder in publicly-traded company. Yin: GracellBiotechnologies Inc: Current Employment, Current equity holder in publicly-traded company. Shen: GracellBiotechnologies Inc: Current Employment, Current equity holder in publicly-traded company. Sersch: GracellBiotechnologies Inc: Current Employment, Current equity holder in publicly-traded company. Cao: GracellBiotechnologies Inc: Current Employment, Current equity holder in publicly-traded company. Wang: GracellBiotechnologies Inc: Current Employment, Current equity holder in publicly-traded company.
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Lilley, Graham, Alden Ladd, Daniel Cossette, Laura Viggiano, Gregory Hopkins, John W. Evans, Mingli Li et al. "Characterization of Lentiviral Vector Derived Anti-Bcma CAR T Cells Reveals Key Parameters for Robust Manufacturing of Cell-Based Gene Therapies for Multiple Myeloma". Blood 126, n. 23 (3 dicembre 2015): 3243. http://dx.doi.org/10.1182/blood.v126.23.3243.3243.
Testo completoAbstract (sommario):
Abstract T cells engineered with chimeric antigen receptors (CAR) specific to CD19 have caused rapid and durable clinical responses in ~90% of patients with acute lymphoblastic leukemia. These data support the development of additional CAR T cell products for the treatment of other hematological malignancies. Recently, B cell maturation antigen (BCMA) expression has been proposed as a marker for identification of malignant plasma cells in patients with multiple myeloma (MM). Nearly all MM and some non-Hodgkin's lymphoma tumor cells express BCMA, while normal tissue expression is restricted to plasma cells and a subset of mature B cells. Therefore, BCMA is an attractive CAR T cell target to treat patients with MM and some B cell lymphomas. To this end, using lentiviral vector technology, we successfully generated CAR T cells specific to BCMA that exhibit potent anti-tumor activity to both multiple myeloma and Burkitt's lymphoma in animal models. Manufacture of CAR T cells for individual patient treatment requires the establishment of a robust and reproducible process - since variability in manufacturing could impact the potency of each cell product. To begin to understand the parameters of the manufacturing process that might contribute to the activity of the final product, we first tested the impact of lentiviral vector (LVV) multiplicity of infection (MOI) on CAR T cell phenotype and function. Using a broad range of MOIs (0.625 to 40) across multiple independent PBMC donors we observed no differences in population doubling or cell size throughout the ~10 day manufacturing process, irrespective of the MOI used. As expected, the number of anti-BCMA CAR expressing cells, the level of CAR expression per cell and the average vector copy number (VCN) in the cell product increased proportionally with MOI. Similarly, T cell function, as determined by an IFNg cytokine release assay in response to BCMA-expressing K562 target cells, was also correlated with the LVV MOI. Notably, increased IFNg expression was readily observable at MOIs as low as 1.25 and reached a plateau with T cells generated using an MOI of 20 or more - highlighting the sensitivity of this functional assay. Analogous data demonstrating MOI dependent in vitro killing activity were obtained using a BCMA-expressing tumor cell cytotoxicity assay. Varying the LVV MOI used during transduction simultaneously alters both the amount of anti-BCMA CAR molecules expressed per cell as well as the number of T cells in the cell product that express anti-BCMA CAR. To evaluate each variable in isolation we generated T cell products containing the same frequency of anti-BCMA CAR T cells (26 ± 4% CAR+ T cells) but different levels of anti-BCMA expression per cell by diluting T cell products made with MOIs from 5 to 40 with donor-matched untransduced cells. While these populations had markedly different levels of CAR surface expression per cell (based on anti-BCMA CAR MFI levels measured by flow cytometry) both low and high expressing anti-BCMA CAR T cell products exhibited identical levels of cytotoxicity against BCMA-expressing tumor cells. These data suggest it is the number of CAR expressing cells that is the critical driver of higher functional activity (perhaps due to the efficiency of LVV mediated anti-BCMA CAR expression per transduced cell). Finally, using this information the variability in manufacturing of anti-BCMA CAR T cells was evaluated across 11 independent normal PBMC donors. All 11 products demonstrated very similar properties with respect to cell growth (population doublings, cell volume), and VCN. Importantly, using our standard MOI we obtained a consistent and high level of anti-BCMA CAR expressing T cells that resulted in robust IFNg cytokine release when co-cultured with BCMA-expression cells. Together, our data highlight the frequency of anti-BCMA CAR T cells per cell product as a key parameter for anti-tumor activity in vitro. Moreover, these data suggest that our LVV driven T cell engineering process can reproducibly generate robust anti-BCMA CAR expressing T cell products in a donor independent manner. A phase I clinical trial to evaluate this technology as a cell-based gene therapy for MM is under development. Disclosures Lilley: bluebird bio, Inc: Employment, Equity Ownership. Ladd:bluebird bio, Inc: Employment, Equity Ownership. Cossette:bluebird bio, Inc: Employment, Equity Ownership. Viggiano:bluebird bio, Inc: Employment, Equity Ownership. Hopkins:bluebird bio, Inc: Employment, Equity Ownership. Evans:bluebird bio, Inc: Employment, Equity Ownership. Li:bluebird bio, Inc: Employment, Equity Ownership. Latimer:bluebird bio: Employment, Equity Ownership. Miller:bluebird bio: Employment, Equity Ownership. Kuczewski:bluebird bio: Employment, Equity Ownership. Bakeman:bluebird bio, Inc: Employment, Equity Ownership. MacLeod:bluebird bio, Inc: Employment, Equity Ownership. Friedman:bluebird bio: Employment, Equity Ownership. Maier:bluebird bio, Inc: Employment, Equity Ownership. Paglia:bluebird bio, Inc: Employment, Equity Ownership. Morgan:bluebird bio: Employment, Equity Ownership. Angelino:bluebird bio, Inc: Employment, Equity Ownership.
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Tran, Thai Hong, Krupa Scott, Shyun-Shyun Lee, Reshma Singh, Shawn Cogan, Jennifer Bordeaux, Hummel Jennifer et al. "Quantitative Multiplexed Immunohistochemistry Assays for Exploring CAR Modified T Cells and Checkpoint Inhibitors in Lymphoma Trials". Blood 126, n. 23 (3 dicembre 2015): 2659. http://dx.doi.org/10.1182/blood.v126.23.2659.2659.
Testo completoAbstract (sommario):
Abstract Although there have been compelling advances in the cancer immunotherapy space recently in the form of chimeric antigen receptor (CAR) modified T-cells and checkpoint inhibitors, advanced tools to explore the therapeutic mechanisms of their combination are not adequately developed or widely available. To address this growing need, we developed a robust quantitative fluorescent immunohistochemistry platform using multiplex AQUA (Automated Quantitative Analysis) technology to evaluate checkpoint inhibitor expression, enumerate CAR T cells and determine the interaction between tumor cells and immune cells via novel co-localization algorithms. We explored utility of this method both in preclinical- and clinical model systems. In an immunodeficient mouse model of B-cell lymphoma, we evaluated homing of CAR T cells to malignant B-cells in primary lymphoid organs. We determined the phenotype and functional status of the CAR T cells via multiplex analyses of CD4, CD8, PD1 and FOXP3 expression. Additionally, to enable combination immunotherapies in Diffuse Large B-Cell Lymphoma (DLBCL) setting, we explored prevalence of adaptive immune resistance mechanisms in the form of PD1 and PD-L1 expression in immune- and tumor cell compartments via landmarks created by cytoplasmic and nuclear stains in both primary and secondary biopsies from DLBCL patients (n = 63). To support patient selection for CAR T trials, we quantified expression and prevalence of relevant tumor antigens that could not be scored reproducibly by traditional methods to yield objective cut points. We anticipate utilization of these quantitative multiplexed IHC methods for optimal selection of patients into upcoming novel combination immunotherapy trials Disclosures Tran: Genoptix: Employment. Scott:Genoptix: Employment. Lee:Genoptix: Employment. Singh:Novartis: Employment. Cogan:Novartis: Employment. Bordeaux:Genoptix: Employment. Jennifer:Genoptix: Employment. Lameh:Genoptix: Employment. Tribouley:Novartis: Employment. Kassim:Novartis: Employment. Tangri:Genoptix Inc., a Novartis company: Employment. Dakappagari:Genoptix Inc., a Novartis company: Employment.
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Totman, Jennifer, Dorothy Brach, Vinny Motwani, Selene Howe, Emily Deutschman, John Lampe, Thomas V. Riera et al. "Pharmacologic Inhibition of the Histone Methyltransferase SETD2 with EZM0414 As a Novel Therapeutic Strategy in Relapsed or Refractory Multiple Myeloma and Diffuse Large B-Cell Lymphoma". Blood 138, Supplement 1 (5 novembre 2021): 1142. http://dx.doi.org/10.1182/blood-2021-151147.
Testo completoAbstract (sommario):
Abstract Introduction: SETD2 is the only known histone methyltransferase (HMT) capable of catalyzing H3K36 trimethylation (H3K36me3) in vivo. It plays an important role in several biological processes including B cell development and maturation, leading to the hypothesis that SETD2 inhibition in these settings could provide anti-tumor effects. The normal process of B cell development/maturation renders B cells susceptible to genetic vulnerabilities that can result in a dysregulated epigenome and tumorigenesis, including in multiple myeloma (MM) and diffuse large B-cell lymphoma (DLBCL). For example, 15%-20% of MM harbors the high risk (4;14) chromosomal translocation, resulting in high expression of the multiple myeloma SET domain (MMSET) gene. MMSET is an HMT that catalyzes H3K36me1 and H3K36me2 formation and extensive scientific work has established overexpressed MMSET as a key factor in t(4;14) myeloma pathogenesis. To the best of our knowledge MMSET has eluded drug discovery efforts, however, since t(4;14) results in high levels of the H3K36me2 substrate for SETD2, inhibiting SETD2 offers promise for targeting the underlying oncogenic mechanism driven by MMSET overexpression in t(4;14) MM patients. In addition, SETD2 loss of function mutations described to date in leukemia and DLBCL are always heterozygous, suggesting a haploinsufficient tumor suppressor role for SETD2. This observation points to a key role for SETD2 in leukemia and lymphoma biology and suggests that therapeutic potential of SETD2 inhibition may also exist in these or similar settings. EZM0414 is a first-in-class, potent, selective, orally bioavailable small molecule inhibitor of the enzymatic activity of SETD2. We explored the anti-tumor effects of SETD2 inhibition with EZM0414 in MM and DLBCL preclinical studies to validate its potential as a therapy in these tumor types. Methods: Cellular proliferation assays determined IC 50 values of EZM0414 in MM and DLBCL cell line panels. Cell line-derived xenograft preclinical models of MM and DLBCL were evaluated for tumor growth inhibition (TGI) in response to EZM0414. H3K36me3 levels were determined by western blot analysis to evaluate target engagement. Combinatorial potential of SETD2 inhibition with MM and DLBCL standard of care (SOC) agents was evaluated in 7-day cotreatment in vitro cellular assays. Results: Inhibition of SETD2 by EZM0414 results in potent anti-proliferative effects in a panel of MM and DLBCL cell lines. EZM0414 inhibited proliferation in both t(4;14) and non-t(4;14) MM cell lines, with higher anti-proliferative activity generally observed in the t(4;14) subset of MM cell lines. The median IC 50value for EZM0414 in t(4;14) cell lines was 0.24 μM as compared to 1.2 μM for non-t(4;14) MM cell lines. Additionally, inhibitory growth effects on DLBCL cell lines demonstrated a wide range of sensitivity with IC 50 values from 0.023 μM to >10 μM. EZM0414 resulted in statistically significant potent antitumor activity compared to the vehicle control in three MM and four DLBCL cell line-derived xenograft models. In the t(4;14) MM cell line-derived xenograft model, KMS-11, robust tumor growth regressions were observed at the top two doses with maximal TGI of 95%. In addition, two non-t(4;14) MM (RPMI-8226, MM.1S) and two DLBCL xenograft models (TMD8, KARPAS422) demonstrated > 75% TGI; with two additional DLBCL models (WSU-DLCL2, SU-DHL-10) exhibiting > 50% TGI in response to EZM0414. In all models tested, the antitumor effects observed correlated with reductions in intratumoral H3K36me3 levels demonstrating on-target inhibition of SETD2 methyltransferase activity in vivo. In vitro synergistic antiproliferative activity was also observed when EZM0414 was combined with certain SOC agents for MM and DLBCL. Conclusions: Targeting SETD2 with a small molecule inhibitor results in significantly reduced growth of t(4;14) MM, as well as non-t(4;14) MM and DLBCL cell lines, in both in vitro and in vivo preclinical studies. In addition, in vitro synergy was observed with EZM0414 and certain SOC agents commonly used in MM and DLBCL, supporting the combination of SETD2 inhibition with current MM and DLBCL therapies. This work provides the rationale for targeting SETD2 in B cell malignancies such as MM, especially t(4;14) MM, as well as DLBCL, and forms the basis for conducting Phase 1/1b clinical studies to evaluate the safety and activity of EZM0414 in patients with R/R MM and DLBCL. Disclosures Totman: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Brach: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Motwani: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Howe: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Deutschman: Epizyme, Inc.: Divested equity in a private or publicly-traded company in the past 24 months, Ended employment in the past 24 months. Lampe: Epizyme, Inc.: Divested equity in a private or publicly-traded company in the past 24 months, Ended employment in the past 24 months. Riera: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Tang: Epizyme, Inc.: Divested equity in a private or publicly-traded company in the past 24 months, Ended employment in the past 24 months. Eckley: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Alford: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Duncan: Epizyme, Inc.: Divested equity in a private or publicly-traded company in the past 24 months, Ended employment in the past 24 months. Farrow: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Dransfield: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Raimondi: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Thomeius: Foghorn Therapeutics: Current Employment, Current equity holder in publicly-traded company. Cosmopoulos: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company. Kutok: Epizyme, Inc.: Current Employment, Current equity holder in publicly-traded company.
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Schomer, Nathan Thomas, Laurent Boissel, Karen Jiang, Hans Klingemann, John H. Lee e Patrick Soon-Shiong. "Providing a Homing Receptor for CAR Engineered NK Cells - Improving Cellular Immunotherapy for B-Cell Lymphoma". Blood 132, Supplement 1 (29 novembre 2018): 4547. http://dx.doi.org/10.1182/blood-2018-99-116890.
Testo completoAbstract (sommario):
Abstract Background Despite some robust initial responses, anti-CD19 Chimeric Antigen Receptor (CAR) T-cell therapy can be associated with significant short-term (cytokine release syndrome) and long-term (B-cell deficiency) toxicities. CAR-engineered natural killer (NK) cells potentially provide a safer alternative while maintaining efficacy. Activated Natural Killer (aNKTM) cells are a clinical grade cell line derived from the NK-92R cell line that has demonstrated potent cytotoxicity towards a broad spectrum of malignant cell lines as well as safety and efficacy in phase I trials. Variants of the aNKcell line are currently in Phase I/II clinical trials: a CAR-expressing aNK cell line and the haNKTM cell line, which have been engineered to carry a high-affinity version of the CD16/FcγRIII receptor to allow for combination therapy with monoclonal antibodies. haNK cells have also been genetically modified to express an endoplasmic reticulum-retained version of IL-2 (ERIL-2), which provides IL-2 independence and limits IL-2 secretion to sub-physiological, safe levels. A key factor for the efficacy of cellular immunotherapies against a given target is biodistribution, which affects the local effector to target ratio. Inability to reach the tumor cells, either by lack of homing or by the accumulation of extracellular matrix (ECM) surrounding a tumor, can be responsible for the clinical failure of even the most effective CAR. The chemokines CCL19 and CCL21 drive recruitment of CCR7-expressing immune cells to secondary lymphoid organs. Engineering aNK cells to express the CCR7 receptor is likely to improve their efficacy by increasing their targeted migration to lymphoma tumor sites. Methods and Results Clinical grade aNK cells were electroporated with a non-viral vector containing the CCR7 receptor, an anti-CD19 CAR, and a high affinity CD16 receptor. To assay the migration of these engineered cell lines, a modified Boyden Chamber assay was performed using Matrigel coated Transwells. K562 cells or modified K562 cells engineered to express CCL19 (K-19) were placed in the destination chamber and CFSE-stained effector cells were placed in the top well. After 24 hours, cells in the bottom well were analyzed by flow cytometry to measure the number of effectors which had migrated through the Matrigel (Fig 1a). The excellent activity of the CAR in stably transfected cells was confirmed against SUP-B15 cells (aNK-resistant), while the ADCC activity was tested against a SUP-B15 variant expressing CD20, but engineered to lack the CD19 antigen (Sup-B15 CD19-, CD20+). Migration towards human lymph node chemokine CCL19 was also tested in vivo in NSG mice with bilateral subcutaneous tumors - with parental K562 in one flank and K-19 tumors on the contralateral flank. CFSE-stained effector cells were delivered via tail vein injection once average tumor size reached 100mm3 and following randomization. Tumors were then harvested at multiple time points, dissociated, and the number of infiltrating effectors in each tumor compared by flow cytometric analysis. (Fig 1b). In testing, monoclonal cell lines expressing all components of the polycistronic system displayed preferential migration towards CCR7 chemokines both in vitro and in vivo, as well as robust cytotoxicity vs. K562 (92.4% +/- 2.4% at 5:1 E:T), Sup-B15(97% +/- 0.6% at 5:1 E:T), and Sup-B15(CD19-, CD20+) when pre-incubated with Rituximab(83.2% +/- 2.8% at 5:1 E:T) but not with control antibody Trastuzumab (22.3% +/- 1.1% at 5:1 E:T) in standard cytotoxicity and ADCC assays. Conclusion We show here that the incorporation of a CCR7 receptor into an off the shelf CAR engineered NK cell line improves their homing towards lymph node chemokines both in vitro and in vivo. This improved homing should result in a greater ratio of effector to target in lymphoid tissue, and maximize the immunogenic cell death. Disclosures Schomer: NantKwest, Inc.: Employment, Equity Ownership. Boissel:NantKwest, Inc.: Employment, Equity Ownership. Jiang:NantKwest, Inc.: Employment, Equity Ownership. Klingemann:NantKwest, INc.: Employment, Equity Ownership, Patents & Royalties. Lee:NantKwest, Inc.: Employment, Equity Ownership. Soon-Shiong:NantKwest, Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties.
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Chang, Kai-Hsin, Minerva Sanchez, Jack Heath, Edouard deDreuzy, Scott Haskett, Abigail Vogelaar, Kiran Gogi et al. "Comparative Studies Reveal Robust HbF Induction By Editing of HBG1/2 Promoters or BCL11A Erythroid-Enhancer in Human CD34+ Cells but That BCL11A Erythroid-Enhancer Editing Is Associated with Selective Reduction in Erythroid Lineage Reconstitution in a Xenotransplantation Model". Blood 132, Supplement 1 (29 novembre 2018): 409. http://dx.doi.org/10.1182/blood-2018-99-114780.
Testo completoAbstract (sommario):
Abstract Beta hemoglobinopathies resulting from dysfunctional or deficient adult beta-globin expression are some of the most prevalent inherited blood disorders in the world. Upregulation of developmentally-silenced fetal gamma-globin would replace adult beta-globin to ameliorate disease symptoms. One of the approaches to reactivate fetal globin expression in erythroid cells is through gene editing by zinc finger or CRISPR-Cas9 nucleases to disrupt the expression of a transcription factor BCL11A, which mediates fetal globin silencing. As BCL11A-deficiency leads to hematopoietic stem cells (HSCs) defects, the current editing approaches target the BCL11A erythroid-enhancer region located in intron 2 of the BCL11A gene to selectively reduce BCL11A expression in erythroid cells. Instead of targeting BCL11A, we sought to identify novel cis-regulatory elements at the beta-globin locus for targeted gene editing to achieve fetal globin reactivation. From a lenti-CRISPR mediated saturated mutagenesis screen covering the beta-globin locus using Human Umbilical Cord Blood-Derived Erythroid Progenitor (HUDEP)-2 cells, multiple fetal hemoglobin (HbF)-inducing genomic domains were identified. Most of the hits were concentrated at the gamma-globin (HBG1/2) promoters, clustered at known hereditary persistence of fetal hemoglobin (HPFH) mutation hotspots. In-depth genotype to phenotype analysis further defined the indels responsible for HbF induction in these subdomains. We interrogated multiple families of nucleases and guide RNA (gRNA) combinations with or without single-stranded oligodeoxynucleotides (ssODN) to guide editing outcome. gRNAs were selected based on their HbF induction potential (up to 40%) when introduced into mobilized peripheral blood (mPB) CD34+ hematopoietic stem and progenitor cells (HSPCs) as ribonucleoprotein (RNP) complexes. HSPCs transfected with RNPs targeting either the BCL11A erythroid-enhancer or the HBG1/2 proximal regions were then injected into NBSGW mice to study the editing in SCID-repopulating cells (SRC) and their multilineage reconstitution potential. All groups achieved high levels of human chimerism (>70% hCD45+/hCD45+mCD45) and comparable monocytes, granulocytes, B lymphocytes, and hCD34+ HSPCs reconstitution. However, BCL11A-edited cells showed selective reduction in erythroid lineage (CD235a+) output, up to 4-fold lower than untreated or HBG1/2 promoter-edited HSPCs. Sequencing analysis from lineage-specific sorted cells further revealed reduced editing levels at BCL11A erythroid-specific enhancer in the erythroid compartment compared to unfractionated bone marrow (BM) or other human lineages (70% in erythroid vs. 90% in all other lineages). Furthermore, the nonproductive fraction of indels that did not disrupt the BCL11A GATAA motif was significantly enriched in erythroid cells (22% in erythroid vs. 8% in other lineages). Ex vivo erythroid cultures suggests BCL11A erythroid-enhancer editing may lead to slightly increased apoptosis during erythroid differentiation. In contrast, HBG1/2 promoter-edited cells had similar editing levels and indel patterns across all lineages with no significant lineage skewing. When chimeric BM from HBG1/2 promoter-edited groups were cultured in erythroid conditions, ex vivo-derived erythroid cells had significantly elevated levels of HbF compared to controls. When CD235a+ cells were sorted without further culture from chimeric BM of mice engrafted with HBG1/2 promoter-edited cells, significantly increased levels of HbF were detected by UPLC (up to 30%) compared to controls (~6%). Thus, long-term HSCs have been edited productively at the HBG1/2 promoters. These cells were able to generate erythroid progenitors that harbor HbF-inducing indels, which in turn, gave rise to erythroid cells in vivo with a clinically-relevant levels of HbF in a xenotransplantation model. Together, our data suggest that BCL11A-edited cells have an erythroid differentiation defect or survival disadvantage in NBSGW mouse model that warrants further investigation. In contrast, editing of the HBG1/2 promoters in mPB CD34+ cells achieved sustained HbF expression in erythroid lineage while maintaining multilineage differentiation potential. Targeting of the HBG1/2 promoters in HSPCs may be an attractive strategy for the development of potential gene editing medicines for beta hemoglobinopathies. Disclosures Chang: Editas Medicine Inc.: Employment, Equity Ownership. Sanchez:Editas Medicine Inc.: Employment, Equity Ownership. Heath:Editas Medicine Inc.: Employment, Equity Ownership. deDreuzy:Editas Medicine Inc.: Employment, Equity Ownership. Haskett:Editas Medicine Inc.: Employment, Equity Ownership. Vogelaar:Editas Medicine Inc.: Employment. Gogi:Editas Medicine Inc.: Employment, Equity Ownership. Da Silva:Editas Medicine Inc.: Employment, Equity Ownership. Wang:Editas Medicine Inc.: Employment, Equity Ownership. Sadowski:Editas Medicine Inc.: Employment, Equity Ownership. Gotta:Editas Medicine Inc.: Employment, Equity Ownership. Siwak:Editas Medicine Inc.: Employment, Equity Ownership. Viswanathan:Editas Medicine Inc.: Employment, Equity Ownership. Loveluck:Editas Medicine Inc.: Employment, Equity Ownership. Chao:Editas Medicine Inc.: Employment, Equity Ownership. Tillotson:Editas Medicine Inc.: Employment, Equity Ownership. Chalishazar:Editas Medicine Inc.: Employment, Equity Ownership. Dass:Editas Medicine Inc.: Employment, Equity Ownership. Ta:Editas Medicine Inc.: Employment, Equity Ownership. Brennan:Editas Medicine Inc.: Employment, Equity Ownership. Tabbaa:Editas Medicine Inc.: Employment, Equity Ownership. Marco:Editas Medicine Inc.: Employment, Equity Ownership. Zuris:Editas Medicine Inc.: Employment, Equity Ownership. Reyon:Editas Medicine Inc.: Employment, Equity Ownership. Isik:Editas Medicine Inc.: Employment, Equity Ownership. Friedland:Editas Medicine Inc.: Employment, Equity Ownership. Ta:Editas Medicine Inc.: Employment, Equity Ownership. Harbinski:Editas Medicine Inc.: Employment, Equity Ownership. Giannoukos:Editas Medicine Inc.: Employment, Equity Ownership. Teixeira:Editas Medicine Inc.: Employment, Equity Ownership. Wilson:Editas Medicine Inc.: Employment, Equity Ownership. Albright:Editas Medicine Inc.: Employment, Equity Ownership. Jiang:Editas Medicine Inc.: Employment, Equity Ownership.
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Campbell, Veronica, Ryan Thompson, Vivian Villegas, Jennifer Proctor, Karen McGovern, Jeffery L. Kutok e Howard M. Stern. "The Potent PI3K-δ,γ Inhibitor IPI-145 Exhibits Differential Activity In Diffuse Large B-Cell Lymphoma (DLBCL) Cell Lines". Blood 122, n. 21 (15 novembre 2013): 1832. http://dx.doi.org/10.1182/blood.v122.21.1832.1832.
Testo completoAbstract (sommario):
Abstract Diffuse large B-cell lymphoma (DLBCL) is the most common form of non-Hodgkin lymphoma (NHL). Despite aggressive chemoimmunotherapy, ∼40% of patients die of their disease. Molecular differences in the tumor genome are likely to be major contributors to the heterogeneity of clinical response. Several investigators have identified molecular subtypes, including a “cell-of-origin” (COO) signature with activated B-cell-like (ABC) and germinal center B-cell-like (GCB) sub-types (Alizadeh, et al. Nature 2000), and a “consensus clustering” (CC) signature with oxidative phosphorylation, B-cell receptor (BCR)/proliferation, and host response sub-types (Monti, et al. 2005). Phosphoinositide-3 kinases (PI3Ks) are key cellular signaling proteins that act as a central node, relaying signals from cell surface receptors to downstream mediators such as AKT. PI3K-δ and PI3K-γ isoforms are preferentially expressed in normal and malignant leukocytes where they are critical for cell differentiation, migration, and proliferation. IPI-145 is a potent, oral PI3K-δ,γ inhibitor that has shown clinical activity in the Phase 1 trial (IPI-145-02) in patients with advanced hematologic malignancies (ClinicalTrials.gov NCT01476657). Activation of the PI3K pathway is an important component of normal BCR signaling and has been implicated in the pathogenesis of DLBCL. To further explore the role of PI3K signaling in DLBCL, a panel of more than 10 DLBCL cell lines of varying molecular profile was treated with IPI-145. PI3K-δ and PI3K-γ were expressed at varying levels across the DLBCL cell lines, without evidence of a correlation with molecular subtype. In a cellular growth inhibition assay, 5 cell lines including 4 GCB (SU-DHL-4, SU-DHL-6, OCI-LY-8, and WSU-DLCL-2) and 1 ABC (Ri-1) subtype were sensitive to IPI-145 treatment in the micromolar (μM) range or below, and 5 cell lines (OCI-LY-3, OCI-LY-7, Pfeiffer, Toledo and U2932) were insensitive to IPI-145 (IC50 >25μM). There was no evidence of a correlation between IPI-145 sensitivity and COO or CC molecular profile in this panel. IPI-145 sensitivity did correlate with evidence of PI3K pathway inhibition as measured by reduction in phospho (p)-AKT. To characterize the kinetics of pathway modulation, phosphorylation of AKT, PRAS40, and S6 was examined following a time-course of IPI-145 treatment in selected cell lines. P-AKT and p-PRAS40 were modulated by 30 minutes, whereas modulation of p-S6 was not detected until after 8 hours. Upon BCR stimulation via antibody-induced crosslinking, some cell lines exhibited enhanced AKT phosphorylation, which was inhibited with IPI-145. The GCB cell line OCI-LY-8 was moderately sensitive to IPI-145 without BCR crosslinking (low μM range) and exhibited enhanced sensitivity to IPI-145 with BCR crosslinking (sub-μM range). These results suggest that intact BCR pathway signaling contributes to IPI-145 sensitivity in DLBCL cell lines. Current efforts are focusing on identification of biomarkers that define IPI-145 responsive DLBCL subsets. IPI-145 activity was also explored in combination with ibrutinib, an irreversible inhibitor of Bruton agammaglobulinemia tyrosine kinase (BTK). Interestingly, in the setting of BCR crosslinking, OCI-LY-8 cells exhibited a robust increase in p-AKT which was completely inhibited by IPI-145 but only partially inhibited by ibrutinib. In other cell lines, such as SU-DHL-4, robust inhibition of p-BTK was observed with ibrutinib treatment but not with IPI-145. These findings suggest a potential mechanistic rationale for combination of PI3K-δ,γ and BTK inhibition. Supporting this hypothesis, a combination effect was observed in a cellular growth inhibition assay with IPI-145 plus ibrutinib in the SU-DHL-4 cell line and in the OCI-LY-8 cell line with BCR crosslinking. Studies are ongoing to explore this combination in additional DLBCL cell lines as well as in in vivo DLBCL models. Taken together, these findings suggest that a subset of DLBCL patients might benefit fromPI3K-δ,γ inhibition. Additional work is necessary to determine how to prospectively identify IPI-145-sensitive DLBCL. In addition, preliminary evidence suggests that there may be opportunities to improve targeted therapy options for DLBCL patients with the combination of IPI-145 and ibrutinib. Disclosures: Campbell: Infinity Pharmaceuticals, Inc.: Employment. Thompson:Infinity Pharmaceuticals, Inc.: Employment. Villegas:Infinity Pharmaceuticals, Inc.: Employment. Proctor:Infinity Pharmaceuticals, Inc.: Employment. McGovern:Infinity Pharmaceuticals, Inc.: Employment. Kutok:Infinity Pharmaceuticals, Inc.: Employment. Stern:Infinity Pharmaceuticals, Inc.: Employment.
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Toudjarska, Ivanka, Zuhua Cai, Tim Racie, Julia Hettinger, Stuart Milstein, Brian R. Bettencourt, Dinah WY Sah, David Bumcrot, John Maraganore e Akshay Vaishnaw. "RNAi-Mediated Inhibition of Activated Protein C- A New Approach for Hemophilia Treatment". Blood 118, n. 21 (18 novembre 2011): 1204. http://dx.doi.org/10.1182/blood.v118.21.1204.1204.
Testo completoAbstract (sommario):
Abstract Abstract 1204 Protein C is a major anticoagulation factor that serves as a key regulator of the clotting cascade. Activated protein C (APC), which is formed by limited proteolysis of the zymogen protein C by the thrombin–thrombomodulin complex, modulates thrombin generation. The anticoagulant effect of APC is due to subsequent inactivation of both activated factors V (FVa) and VIII (FVIIIa). The important role of APC in coagulation is highlighted in FV Leiden (FVL), the most common inherited form of thrombophilia. FVL is caused by a mutation in the FV gene at the initial APC cleavage site, which results in FVL being inactivated at approximately one tenth the rate of normal FVa, leading to high thrombin levels that create a procoagulant state. Several clinical studies have suggested that the severity or onset of bleeding phenotype in hemophilia patients is substantially reduced in association with impaired inactivation of FVa by APC in the presence of the FVL mutation. Transgenic animal studies also show that hemophilic mice, either heterozygous or homozygous for FVL, have improved clotting times with the ability to form clots at sites of laser-induced injury in a microvascular bed injury model, supporting the role of the FVL mutation in enhancing hemostasis. Therapeutic strategies aimed at reducing the level of protein C and activated protein C, thereby increasing levels of FVa and thrombin could thus prove efficacious in hemophilia A and B. Here we investigate the systemic administration of lipid nanoparticle (LNP) formulated siRNA directed against Protein C (encoded by Proc). We have designed potent siRNA sequences against human, cynomologous monkey, dog, mouse and rat Proc mRNA. The lead human/cynomologous candidate has an IC50 of 5pM. The lead mouse/rat candidate has an IC50 of 30 pM and an ED50 of 20 ug/kg. We demonstrate robust and durable inhibition of Proc mRNA in normal mouse liver- a single injection of 0.3 mg/kg resulted in 90% silencing of Proc mRNA and reduction of more than 75% which was sustained for more than two weeks. We also demonstrate significant reduction of circulating protein as measured by Western blot. Further testing using genetic models of hemophilia is being conducted to provide proof-of-concept for developing a Proc siRNA as a novel therapeutic agent in hemophilia management. Disclosures: Toudjarska: Alnylam Pharmaceuticals, Inc.: Employment. Cai:Alnylam Pharmaceuticals, Inc.: Employment. Racie:Alnylam Pharmaceuticals, Inc.: Employment. Hettinger:Alnylam Pharmaceuticals, Inc.: Employment. Milstein:Alnylam Pharmaceuticals, Inc.: Employment. Bettencourt:Alnylam Pharmaceuticals, Inc.: Employment. Sah:Alnylam Pharmaceuticals, Inc.: Employment. Bumcrot:Alnylam Pharmaceuticals, Inc.: Employment. Maraganore:Alnylam pharmaceuticals, Inc.: Employment. Vaishnaw:Alnylam Pharmaceuticals, Inc.: Employment.
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Storhoff, James, Brett Wallden, Rita M. Braziel, Catherine Thieblemont, Tressa Hood, Harini Ravi, Shannon Dennis et al. "Analytical Performance of Nanostring's Companion Diagnostic (CDx) Lymphoma Subtyping Test (LST) with Diffuse Large B-Cell Lymphoma Core Needle Biopsy Samples". Blood 126, n. 23 (3 dicembre 2015): 1481. http://dx.doi.org/10.1182/blood.v126.23.1481.1481.
Testo completoAbstract (sommario):
Abstract Background: Diffuse large B-cell lymphoma (DLBCL) is an aggressive non-Hodgkin's lymphoma with two distinct molecular cell-of-origin (COO) subtypes known as germinal center B-cell (GCB) or activated B-cell (ABC). DLBCL subtypes have been reported to be prognostic and potentially predictive of treatment benefit, underscoring the need for a precise and accurate CDx test (Roschewski, Nat Rev Clin Oncol, 2014). NanoString's LST was developed to enable identification of the COO subtypes on the nCounter® Dx Analysis System using formalin fixed paraffin embedded (FFPE) tissue specimens based on the previously developed Lymph2Cx gene expression profiling assay (Scott, Blood 2014). The test assigns the COO subtypes based on a Linear Predictor Score (LPS) that is calculated using a weighted sum of the gene expression. The LST is currently being used as a CDx in a phase III global clinical validation trial (clinicaltrials.gov NCT02285062) to identify newly diagnosed DLBCL patients of the ABC type who may preferentially respond to lenalidomide. The LST was previously reported to be a highly precise and accurate method of determining DLBCL subtypes (ABC and GCB) when using FFPE excisional/surgical biopsy tissue specimens as a test input (Wallden, JCO 2015). Although excisional biopsies are recommended for DLBCL diagnosis by current ESMO and NCCN guidelines, recent studies have reported excellent sensitivity and specificity in diagnosing malignant lymphomas by core needle biopsy (CNB) leading to increased usage of this method (Hu Q, Am J Clin Pathol, 2013, de Kerviler, Best Pract Res Cl Ha, 2012). The aim of the current study was to assess the analytical performance of the LST when using CNB samples as a test input. Methods: Recently archived CNB and small biopsy DLBCL samples were procured from Oregon Health & Science University (Portland, OR) (n=23), Hôpital Saint-Louis (Paris, France) (n=19), and Asterand Bioscience (n=12). Both nodal and extra-nodal tissue sites were included. Pathology review was performed on an H&E stained slide from each tissue sample to identify the area of DLBCL. Unstained slide mounted FFPE tissue sections (5 µm) were prepared from each sample for assay processing. The RNA extraction and LST procedures were the same as reported previously for surgical biopsy samples (Wallden, JCO 2015). The number of sections used for each RNA extraction was based on the tumor surface area (TSA) measured on the slide (typically 4-6 slides for TSA<10 mm2; 2-4 slides for TSA≥10 mm2). Multiple extractions were performed and subsequently tested independently for each sample (n=148 total). Additional variables tested in this study were user (n=2), reagent lot (n=2), and instrument (n=2). The reproducibility of the test outputs were evaluated using a linear mixed model (LMM) for LPS variance and concordance for subtype call. Principal component analysis (PCA) was performed on the gene expression profile to determine the major source(s) of variability. Results: Of the 54 total samples tested, 51 provided test results (failures were due to RNA degradation). With the 51 passing samples, an assay pass rate of 97% (144/148) was achieved. The total tissue area input into the test was correlated to RNA yield (R2 =0.54). The LPS was highly reproducible across sample replicates (Figure 1) with a total assay standard deviation of 59 LPS units (<2% of the LPS range). This estimate was similar to surgical biopsy samples (76 LPS units). The subtype concordance was estimated as 96-98% (between users) where 2 of 51 samples with LPS scores close to the thresholds moved from ABC or GCB to unclassified, or vice versa. A PCA of the gene expression data demonstrated that the first principle component is highly correlated to the LPS providing further evidence that the biology underlying the LST is the major driver of algorithm gene expression in CNB. Conclusions: In this study we demonstrated robust performance of the LST using FFPE DLBCL CNB/small biopsy samples. Our data support that sufficient RNA yield and gene expression quality can be achieved from biopsy samples with measured TSA of 2 mm2 or greater. We also show that the analytical performance of the LST is comparable between CNB/small biopsy samples and surgical biopsies and that the principal component of gene expression variation is LPS. The LST sample input requirements for the ongoing phase III trial will be updated to include CNB/small biopsy samples to accommodate clinical practice. Disclosures Storhoff: NanoString Technologies, Inc.: Employment, Other: Stock . Wallden:NanoString Technologies, Inc.: Employment, Other: Stock. Braziel:Oregon Health & Sciences University: Employment. Thieblemont:St. Louis Hospital, Paris, France: Employment. Hood:NanoString Technologies, Inc.: Employment, Other: Stock. Ravi:NanoString Technologies, Inc.: Employment, Other: Stock. Dennis:NanoString Technologies, Inc.: Employment, Other: Stock. Dowidar:NanoString Technologies, Inc.: Employment, Other: Stock. Danaher:NanoString Technologies, Inc.: Employment, Other: Stock. Dunlap:Oregon Health & Sciences University: Employment. Briere:St. Louis Hospital, Paris, France: Employment. de Kerviler:St. Louis Hospital, Paris, France: Employment. Ferree:NanoString Technologies, Inc.: Employment, Other: Stock, Patents & Royalties: NanoString Technologies, Inc..
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Coiffier, Bertrand, Juan-Manuel Sancho, Wojciech Jurczak, Jin Seok Kim, Raj V. Nagarkar, Edvard Zhavrid, Jose Angel Hernandez Rivas et al. "Pharmacokinetic and Safety of CT-P10, a Biosimilar Candidate to the Rituximab Reference Product, in Patients with Newly Diagnosed Advanced Stage Follicular Lymphoma (AFL)". Blood 128, n. 22 (2 dicembre 2016): 1807. http://dx.doi.org/10.1182/blood.v128.22.1807.1807.
Testo completoAbstract (sommario):
Abstract Background: CT-P10 is a biosimilar candidate to the reference rituximab product, EU-approved MabThera® and US-licensed Rituxan®. CT-P10 has an identical amino acid sequence and highly similar physicochemical and in vitro functional properties to its reference drug. In patients with rheumatoid arthritis, CT-P10 has demonstrated compelling similarity in pharmacokinetics (PK), pharmacodynamics (PD), efficacy, safety and immunogenicity (Yoo DH, et al. Arthritis Rheum. 2013;65(10):1736). Objective: The goal of this study was to demonstrate PK similarity of CT-P10 to rituximab, each administered in combination with cyclophosphamide, vincristine, and prednisone (CVP) in patients with newly diagnosed Advanced Follicular Lymphoma (AFL) (NCT02162771) (Kim WS et al. Blood. 2015;126(23): 5111). The results of PK, PD, safety and immunogenicity up to Core Cycle 4 (12 weeks) are presented here from this ongoing study. Methods: Patients with AFL were randomized 1:1 to receive infusion (375mg/m2) of either CT-P10 or rituximab, at a 3-week interval, in combination with CVP. PK analysis was conducted in terms of AUCtau and Cmax at steady state, Core Cycle 4, as primary PK endpoints. PK parameter values considered as outliers determined by robust regression outlier testing were excluded from the pharmacokinetic primary analysis. Results: In total, 121 patients were randomly assigned to receive either CT-P10 (n=59) or rituximab (n=62) in combination with CVP. Result of CT-P10 PK at Core Cycle 4 was similar to that of rituximab. The ratios (90% CI) of geometric least square means (CT-P10 to rituximab treatment group) were 102.3% (94.1%-111.2%) for AUCtau and 100.7% (93.8%-108.0%) for CmaxSS at Core Cycle 4. The 90% CIs of ratio of geometric LS means for both AUCtau and CmaxSS were entirely contained within the equivalence margin of 80% to 125% (Table 1 and Figure 1). Mean serum concentrations of the study drug were highly similar for the 2 treatment groups at each time point (Core Cycle 1 to 4). The B-cell kinetics was similar up to Core Cycle 4 in the 2 treatment groups. Median number of B-cells decreased to below the lower limit of quantification (LLoQ) (20 cells/μL) 1 hour after the end of infusion at Core Cycle 1 and remained below the LLoQ at each subsequent cycle, up to and including Core Cycle 4. The proportion of patients with a positive anti-drug antibody up to Core Cycle 4 at post-treatment visits was similar between the 2 treatment groups; 3/59 (5.1%) patients and 2/62 (3.2%) patients in the CT-P10 and rituximab groups, respectively. In addition, CT-P10 was well tolerated and the safety profile of CT-P10 up to Core Cycle 4 was similar to that of rituximab. The number of patients who experienced at least 1 treatment emergent adverse event (TEAE) was 43 (72.9%) patients and 41 (66.1%) patients in CT-P10 and rituximab treatment groups, respectively. The proportion of patients who experienced at least 1 treatment emergent serious adverse event considered by the investigator to be related to the study treatment was similar between the 2 treatment groups; 2/59 (3.4%) patients and 2/62 (3.2%) patients in the CT-P10 and rituximab groups, respectively. The frequencies of adverse events special interest (AESI) were similar between the 2 treatment groups (Table 2). Conclusion: This study demonstrated similarity of PK in terms of AUCtau and CmaxSS between CT-P10 and rituximab in AFL patients. The B-cell kinetics and immunogenicity were comparable between the two treatment groups. CT-P10 was well tolerated with a safety profile comparable to that of rituximab up to and including Core Cycle 4 (12 weeks). Table 1 Statistical Analysis of Rituximab Pharmacokinetic Primary Endpoints for Core Cycle 4 at Steady State: Pharmacokinetic Population Table 1. Statistical Analysis of Rituximab Pharmacokinetic Primary Endpoints for Core Cycle 4 at Steady State: Pharmacokinetic Population Figure 1 Mean (±SD) Serum Concentration of Rituximab Versus Time for Cycle 4 at Steady State (Linear Scale): Pharmacokinetic Population Figure 1. Mean (±SD) Serum Concentration of Rituximab Versus Time for Cycle 4 at Steady State (Linear Scale): Pharmacokinetic Population Table 2 The Incidence Rates of Adverse Events Special Interest: Safety Population Table 2. The Incidence Rates of Adverse Events Special Interest: Safety Population Disclosures Coiffier: Celltrion, Inc.: Consultancy, Honoraria. Sancho:Gilead: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Celltrion, Inc: Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees; Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Jurczak:Celltrion, Inc: Research Funding; Gilead Sciences: Research Funding; Acerta: Research Funding; Bayer: Research Funding; Janssen: Research Funding. Kim:Celltrion, Inc.: Research Funding. Nagarkar:Celltrion, Inc.: Research Funding. Zhavrid:Celltrion, Inc.: Research Funding. Hernandez Rivas:Celltrion, Inc.: Research Funding. Prokharau:Celltrion, Inc.: Research Funding. Zodelava:Celltrion, Inc.: Research Funding. Osmanov:Celltrion, Inc.: Research Funding. Ogura:SymBio Pharmaceuticals: Consultancy, Honoraria; Celltrion, Inc.: Consultancy, Honoraria. Buske:Celltrion, Inc.: Consultancy, Honoraria. Kwak:Celltrion, Inc.: Consultancy. Kim:Celltrion, Inc.: Consultancy, Honoraria.
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Ganesan, Harish, Vipul Kumar, Sujata M. Bhavnani, David Melnick e Christopher M. Rubino. "1105. Population Pharmacokinetic Analyses for Tebipenem After the Administration of Tebipenem Pivoxil Hydrobromide". Open Forum Infectious Diseases 8, Supplement_1 (1 novembre 2021): S644—S645. http://dx.doi.org/10.1093/ofid/ofab466.1299.
Testo completoAbstract (sommario):
Abstract Background Tebipenem pivoxil hydrobromide (TBP-PI-HBr) is an oral prodrug that is converted to tebipenem (TBP), the active moiety. TBP is a carbapenem with activity against multidrug-resistant Gram-negative pathogens, including extended-spectrum-β-lactamase-producing Enterobacterales and is being developed for treating complicated urinary tract infections (cUTI) and acute pyelonephritis (AP). Data from three Phase 1 studies and one Phase 3 study in patients with cUTI/AP were used to develop a population pharmacokinetic (PPK) model for TBP and identify covariates that described the variability in TBP pharmacokinetics (PK). Methods The PPK model was developed using TBP plasma and urine concentration-time data from the above-described Phase 1 and 3 studies. TBPPI-HBr doses, which ranged from 100 to 900 mg, were administered as single or multiple doses every 8 hours. After development of the structural model, stepwise forward and backward selection procedures were used to identify significant covariate relationships. The robustness of the final PPK model was assessed using a prediction-corrected visual predictive check (PC-VPC). Results The final dataset included 3448 plasma concentrations from 99 Phase 1 subjects and 647 Phase 3 patients and urine concentrations from 128 Phase 1 subjects. A two-compartment model with linear, first-order elimination and transit compartments to describe the rate of drug absorption after oral administration of TBP-PI-HBr best described TBP PK. The most clinically significant covariate effect, which would warrant dose adjustment, was the relationship between apparent oral clearance and creatinine clearance. In contrast, age, body size, sex, and fed status each had a minimal impact on TBP exposure. The PC-VPC showed good agreement between median simulated plasma concentrations based on the final PPK model and the median observed plasma concentrations for the pooled dataset (Figure 1). Figure 1. Prediction-corrected visual predictive check plot for the final population PK model using the pooled analysis dataset Conclusion A robust description of TBP plasma PK in subjects and patients with cUTI/AP was achieved, such that derived measures of TBP plasma exposure are expected to be both accurate and precise. The population PK model was considered appropriate for model-based simulations and the assessment of PK-PD relationships for TBP. Disclosures Harish Ganesan, M.S., 3-V Biosciences (Grant/Research Support)Achogen (Grant/Research Support)Amplyx Pharmaceuticals, Inc. (Grant/Research Support)Arixa Pharmaceuticals (Grant/Research Support)Arsanis Inc. (Grant/Research Support)B. Braun Medical Inc. (Grant/Research Support)Basilea Pharmaceutica (Grant/Research Support)BLC USA (Grant/Research Support)Boston Pharmaceuticals (Grant/Research Support)Bravos Biosciences, LLC (Grant/Research Support)Cidara Therapeutics Inc. (Grant/Research Support)Cipla, USA (Grant/Research Support)Corcept Therapeutics (Grant/Research Support)Cumberland Pharmaceuticals (Grant/Research Support)Debiopharm International SA (Grant/Research Support)Discuva Limited (Grant/Research Support)Emerald Lake Technologies (Grant/Research Support)Enhanced Pharmacodynamics (Grant/Research Support)Entasis Therapeutics (Grant/Research Support)E-Scape Bio (Grant/Research Support)Genentech (Grant/Research Support)Geom Therapeutics, Inc. (Grant/Research Support)GlaxoSmithKline (Grant/Research Support)Hoffmann-La Roche (Grant/Research Support)Horizon Orphan LLC (Grant/Research Support)ICPD Biosciences, LLC (Grant/Research Support)Indalo Therapeutics (Grant/Research Support)Insmed Inc. (Grant/Research Support)Institute for Clinical Pharmacodynamics (Employee)Iterum (Grant/Research Support)KBP Biosciences USA (Grant/Research Support)Kyoto Biopharma, Inc. (Grant/Research Support)Matinas (Grant/Research Support)Meiji Seika Pharma Co., Ltd. (Grant/Research Support)Melinta Therapeutics (Grant/Research Support)Menarini Ricerche S.p.A. (Grant/Research Support)Merck & Co., Inc (Grant/Research Support)Mutabilis (Grant/Research Support)Nabriva Therapeutics AG (Grant/Research Support)Naeja-RGM Pharmaceuticals (Grant/Research Support)Nosopharm SAS (Grant/Research Support)Novartis Pharmaceuticals Corp. (Grant/Research Support)NuCana Biomed (Grant/Research Support)Paratek Pharmaceuticals, Inc. (Grant/Research Support)Polyphor, Ltd. (Grant/Research Support)Prothena Corporation (Grant/Research Support)PTC Therapeutics (Grant/Research Support)Rempex Pharmaceuticals (Grant/Research Support)Roche TCRC (Grant/Research Support)Sagimet (Grant/Research Support)scPharmaceuticals Inc. (Grant/Research Support)Scynexis (Grant/Research Support)Spero Therapeutics (Grant/Research Support)TauRx Therapeutics (Grant/Research Support)Tetraphase Pharmaceuticals (Grant/Research Support)Theravance Biopharma Pharmaceutica (Grant/Research Support)USCAST (Grant/Research Support)VenatoRx (Grant/Research Support)Vical Incorporated (Grant/Research Support)Wockhardt Bio AG (Grant/Research Support)Zavante Therapeutics (Grant/Research Support)Zogenix International (Grant/Research Support) Vipul Kumar, PhD, Spero Therapeutics (Employee, Shareholder) Sujata M. Bhavnani, Pharm.D., M.S., FIDSA, 3-V Biosciences (Grant/Research Support)Achogen (Grant/Research Support)Amplyx Pharmaceuticals, Inc. (Grant/Research Support)Arixa Pharmaceuticals (Grant/Research Support)Arsanis Inc. (Grant/Research Support)B. Braun Medical Inc. (Grant/Research Support)Basilea Pharmaceutica (Grant/Research Support)BLC USA (Grant/Research Support)Boston Pharmaceuticals (Grant/Research Support)Bravos Biosciences, LLC (Grant/Research Support, Other Financial or Material Support, member/owner)Cidara Therapeutics Inc. (Grant/Research Support)Cipla, USA (Grant/Research Support)Corcept Therapeutics (Grant/Research Support)Cumberland Pharmaceuticals (Grant/Research Support)Debiopharm International SA (Grant/Research Support)Discuva Limited (Grant/Research Support)Emerald Lake Technologies (Grant/Research Support)Enhanced Pharmacodynamics (Grant/Research Support)Entasis Therapeutics (Grant/Research Support)E-Scape Bio (Grant/Research Support)Genentech (Grant/Research Support)Geom Therapeutics, Inc. (Grant/Research Support)GlaxoSmithKline (Grant/Research Support)Hoffmann-La Roche (Grant/Research Support)Horizon Orphan LLC (Grant/Research Support)ICPD Biosciences, LLC (Grant/Research Support, Other Financial or Material Support, member/owner)Indalo Therapeutics (Grant/Research Support)Insmed Inc. (Grant/Research Support)Institute for Clinical Pharmacodynamics (Employee)Iterum (Grant/Research Support)KBP Biosciences USA (Grant/Research Support)Kyoto Biopharma, Inc. (Grant/Research Support)Matinas (Grant/Research Support)Meiji Seika Pharma Co., Ltd. (Grant/Research Support)Melinta Therapeutics (Grant/Research Support)Menarini Ricerche S.p.A. (Grant/Research Support)Merck & Co., Inc (Grant/Research Support)Mutabilis (Grant/Research Support)Nabriva Therapeutics AG (Grant/Research Support)Naeja-RGM Pharmaceuticals (Grant/Research Support)Nosopharm SAS (Grant/Research Support)Novartis Pharmaceuticals Corp. (Grant/Research Support)NuCana Biomed (Grant/Research Support)Paratek Pharmaceuticals, Inc. (Grant/Research Support)Polyphor, Ltd. (Grant/Research Support)Prothena Corporation (Grant/Research Support)PTC Therapeutics (Grant/Research Support)Rempex Pharmaceuticals (Grant/Research Support)Roche TCRC (Grant/Research Support)Sagimet (Grant/Research Support)scPharmaceuticals Inc. (Grant/Research Support)Scynexis (Grant/Research Support)Spero Therapeutics (Grant/Research Support)TauRx Therapeutics (Grant/Research Support)Tetraphase Pharmaceuticals (Grant/Research Support)Theravance Biopharma Pharmaceutica (Grant/Research Support)USCAST (Grant/Research Support)VenatoRx (Grant/Research Support)Vical Incorporated (Grant/Research Support)Wockhardt Bio AG (Grant/Research Support)Zavante Therapeutics (Grant/Research Support)Zogenix International (Grant/Research Support) David Melnick, MD, Spero Therapeutics (Employee, Shareholder) Christopher M. Rubino, Pharm.D., 3-V Biosciences (Grant/Research Support)Achogen (Grant/Research Support)Amplyx Pharmaceuticals, Inc. (Grant/Research Support)Arixa Pharmaceuticals (Grant/Research Support)Arsanis Inc. (Grant/Research Support)B. Braun Medical Inc. (Grant/Research Support)Basilea Pharmaceutica (Grant/Research Support)BLC USA (Research Grant or Support)Boston Pharmaceuticals (Grant/Research Support)Bravos Biosciences, LLC (Grant/Research Support, Other Financial or Material Support, member/owner)Cidara Therapeutics Inc. (Grant/Research Support)Cipla, USA (Grant/Research Support)Corcept Therapeutics (Grant/Research Support)Cumberland Pharmaceuticals (Grant/Research Support)Debiopharm International SA (Grant/Research Support)Discuva Limited (Grant/Research Support)Emerald Lake Technologies (Grant/Research Support)Enhanced Pharmacodynamics (Grant/Research Support)Entasis Therapeutics (Grant/Research Support)E-Scape Bio (Grant/Research Support)Genentech (Grant/Research Support)Geom Therapeutics, Inc. (Grant/Research Support)GlaxoSmithKline (Grant/Research Support)Hoffmann-La Roche (Grant/Research Support)Horizon Orphan LLC (Grant/Research Support)ICPD Biosciences, LLC (Grant/Research Support, Other Financial or Material Support, member/owner)Indalo Therapeutics (Grant/Research Support)Insmed Inc. (Grant/Research Support)Institute for Clinical Pharmacodynamics (Employee)Iterum (Grant/Research Support)KBP Biosciences USA (Grant/Research Support)Kyoto Biopharma, Inc. (Grant/Research Support)Matinas (Grant/Research Support)Meiji Seika Pharma Co., Ltd. (Grant/Research Support)Melinta Therapeutics (Grant/Research Support)Menarini Ricerche S.p.A. (Grant/Research Support)Merck & Co., Inc (Grant/Research Support)Mutabilis (Grant/Research Support)Nabriva Therapeutics AG (Grant/Research Support)Naeja-RGM Pharmaceuticals (Grant/Research Support)Nosopharm SAS (Grant/Research Support)Novartis Pharmaceuticals Corp. (Grant/Research Support)NuCana Biomed (Grant/Research Support)Paratek Pharmaceuticals, Inc. (Grant/Research Support)Polyphor, Ltd. (Grant/Research Support)Prothena Corporation (Grant/Research Support)PTC Therapeutics (Grant/Research Support)Rempex Pharmaceuticals (Grant/Research Support)Roche TCRC (Grant/Research Support)Sagimet (Grant/Research Support)scPharmaceuticals Inc. (Grant/Research Support)Scynexis (Grant/Research Support)Spero Therapeutics (Grant/Research Support)TauRx Therapeutics (Grant/Research Support)Tetraphase Pharmaceuticals (Grant/Research Support)Theravance Biopharma Pharmaceutica (Grant/Research Support)USCAST (Grant/Research Support)VenatoRx (Grant/Research Support)Vical Incorporated (Grant/Research Support)Wockhardt Bio AG (Grant/Research Support)Zavante Therapeutics (Grant/Research Support)Zogenix International (Grant/Research Support)
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Merrill, J. T., J. Guthridge, D. Zack, P. Foster, B. Burington, L. Tran, M. Smith e J. A. James. "SAT0187 DISCRIMINATION OF SYSTEMIC LUPUS (SLE) PATIENTS WITH CLINICAL RESPONSE TO OBEXELIMAB (XMAB®5871) BASED ON A PATTERN OF IMMUNOLOGIC MARKERS". Annals of the Rheumatic Diseases 79, Suppl 1 (giugno 2020): 1035–36. http://dx.doi.org/10.1136/annrheumdis-2020-eular.2972.
Testo completoAbstract (sommario):
Background:We recently reported Phase 2 SLE trial results of obexelimab, an FcγRIIb agonist (suppressor of B cell activation). Obexelimab did not meet the primary endpoint (% of patients without flare at Day 225) (p=0.183) but other endpoints such as time to flare (p=0.025) were met.Objectives:1. To assign SLE patients to phenotypic subsets based on patterns of gene expression in immune-related pathways.12. To explore the association of immune patterns and clinical response to obexelimab.Methods:This analysis included 71 of the 104 participants in the obexelimab study, those who either completed the protocol or terminated for disease flare, if there were adequate blood samples (biomarker subset). At screening, patients were assigned to clusters based on 41 SLE co-expression signature modules from the Human Immune Phenotyping Consortium via unsupervised random forest and K-means clustering.2Other markers of SLE disease were also examined. TheBOLD3study design mandated withdrawal of background immunosuppressants, supporting less ambiguous pharmacodynamic analysis as the trial progressed.Results:Immune pathway expression patterns of 7 patient clusters (Fig 1a) confirmed our prior characterization of 200 non-overlapping SLE patients.2The biomarker subset retained a trend of longer time to first flare in patients receiving obexelimab (n=38) vs placebo (n=33) (Fig1b, HR 0.61, p=0.11). A smaller set of only Clusters 3 and 6 demonstrated marked increased time to flare in the obexelimab group (n=13) compared to placebo (n=14) (Fig 1c, HR 0.22, p=0.025). Obexelimab had no effect on other clusters (Fig 1d). The responder clusters shared low expression of inflammation modules (p < 0.001) compared to other clusters and high expression of T Cell, immune response, cell cycle, mitochondrial modules (all p < 0.001) and B Cell modules (p=0.006). We therefore sorted patients by these specific features regardless of cluster assignment. Figure 2 shows significant impact of obexelimab on time to flare in 64 patients with B Cell pathway activation (HR 0.5, p=0.038), although less robust by itself than found in Clusters 3 and 6. In a group with high B or plasma cell modules but low inflammation (n=46), treatment effect increased (HR 0.35, p=0.019). Between Screening and Baseline, as brief steroids were given and background treatments withdrawn, expression of B Cell and Plasma Cell pathways increased. Both then decreased after treatment with obexelimab but not placebo (p< 0.0001 and p< 0.001 respectively), an effect not seen with other immune pathway modules.Conclusion:Precision medicine for SLE has been hampered by heterogenous immune signals with variable expression. Clustering of patients by gene co-expression pathways enabled an efficient, hierarchical array that reduplicated results of a prior SLE cohort, suggesting these are not random phenotypes. Of these 7 reproducible SLE subsets, the combination of clusters 3 and 6 distinguished an obexelimab responder population of 27 out of 71 subjects (38%) with high expression of B and T Cell modules and cell activation pathways. Focus on the defining features shared by these clusters revealed specific factors associated with response, enabling inclusion of some patients from other clusters in an optimized responder population of 46/71 (65% of subjects). B Cell and Plasma Cell pathways demonstrated mechanism-related pharmacodynamic effects of obexelimab. Lack of responders with high expression of inflammation modules could implicate inhibitory factors to obexelimab within inflammatory pathways, potentially targetable by complementary treatments.References:[1]Banchereau Cell 165:1548 2016[2]Lu ACR Abstract #2977 2017[3]Merrill Arthritis Rheumatol 69: 1257 2017Disclosure of Interests: :Joan T Merrill Grant/research support from: Xencor, Bristol Myers Squibb, Glaxo Smith Kline, Consultant of: Xencor, Abbvie, UCB, Glaxo Smith Kline, EMD Serono, Astellas, Remegen, Celgene/Bristol Myers Squibb, Exagen, Astra Zeneca, Amgen, Jannsen, Servier, ILTOO, Daitchi Sankyo, Lilly, Paid instructor for: Abbvie, Bristol Myers Squibb, Joel Guthridge Grant/research support from: Xencor, Bristol Myers Squibb, DXterity, Debra Zack Shareholder of: Xencor, Employee of: Xencor, Paul Foster Shareholder of: Xencor Inc, Employee of: Xencor Inc, Bart Burington Shareholder of: Xencor Inc, Employee of: Xencor Inc, Ly Tran: None declared, Miles Smith: None declared, Judith A. James Grant/research support from: Progentec Diagnostics, Inc, Consultant of: Abbvie, Novartis, Jannsen
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Dias, Lizalynn, Venkata Thodima, Geetu Mendiratta, Guttapalli Asha, Camille Gonzalez, Jocelyn C. Maragulia, Andrew D. Zelenetz, Julie Teruya Feldstein, Raju S. K. Chaganti e Jane Houldsworth. "Cross-Platform Assessment Of Genomic Imbalance In Diffuse Large B-Cell Lymphoma Identifies Novel Candidate Loci and Genes With Prognostic Value and Roles In Lymphomagenesis". Blood 122, n. 21 (15 novembre 2013): 4259. http://dx.doi.org/10.1182/blood.v122.21.4259.4259.
Testo completoAbstract (sommario):
Abstract Diffuse large B-cell lymphomas (DLBCL) display marked clinical, pathologic, and genetic heterogeneity. With current frontline immunotherapy (RCHOP), only about 40% of patients are cured, with most relapses occurring within the first 2-3 years. Patients are currently risk-stratified based primarily on clinical features where the inclusion of molecular biomarkers into risk assessment could impact the potential to identify those patients most likely to have refractory disease or have an early relapse. Various cytogenomic studies have revealed the prognostic significance of genomic gain/loss in DLBCL, but their lack of utility and reproducibility across datasets can be attributed to not only different patient populations, but also the use of disparate platforms and analytical methods. The goal of the present study was to use a common analytical approach across different clinical datasets, to identify genomic loci with robust prognostic value in DLBCL. To this end, GISTIC was applied to copy number data of newly-diagnosed DLBCL (GSE11318 [n=170], E-MEXP-3463 [n=49]) and together with another similarly analyzed published dataset (PMID: 22975378 n= [180]). Regions of known copy number variations were excluded from the analysis. A total of 35 overlapping minimal common regions (MCRs) and 21 overlapping peaks of genomic gain/loss with well-defined calling parameters were identified in at least 2 of the 3 datasets with a minimum occurrence of 5%. Overlapping peaks mapped to loci of genes with known involvement in DLBCL such as TP53, CDKN2A, and TNFAIP3 confirming the validity of the approach. Individual MCRs were tested for association with overall survival using the Kaplan-Meier method and log-rank statistic in three RCHOP datasets: E-MEXP-3463, GSE15127 [N=124], and an in-house dataset for which array-CGH was performed using a targeted oligonucleotide (Agilent Technologies) with DNA extracted from 1-3 CD20+ enriched cores of formalin–fixed paraffin-embedded de novo DLBCL; IH [n=46]. Nine MCRs were found to significantly associate (P ≤0.05) with poor outcome: gain of 3q, 9q, 19p, and loss of 1p, 8p, 9p, 13q, 15q, and 17p. Four remained significant after multiple testing correction: gain of chr3:187,651,865-196,853,350 (P<0.001), chr9:138,543,735-140,878,804 (P=0.0110), and loss of chr15:40,295,857-46,224,648 (P=0.012), chr17:1,000,000-16,936,602 (P=0.050). Genomic complexity assessed as previously described (PMID:22975378), failed to significantly correlate with outcome in any of the three RCHOP datasets tested. Correlation of gene expression levels to copy number change was evaluated in the (GSE11318 [n=162]) dataset. A total of 397 differentially expressed genes (P ≤0.05, FDR) showing positive correlation with copy number status were identified within 16 MCRs of which, five mapped to three peak MCRs. Genes mapped to the peak MCRs were found to have roles in NFKB activation, iron transport, phosphatidylcholine biosynthesis and regulation of transcription all of which represent novel therapeutic targets in DLBCL. For the nine MCRs associated with poor outcome, 69 genes exhibited positive correlation with copy number and enrichment of the KEGG pathway using these genes in the DAVID 6.7 program identified the endocytosis, apoptosis, and glycosylphosphatidylinositol (GPI)-anchor biosynthesis pathways. In summary, using this platform-agnostic approach, common and novel loci of genomic imbalance in DLBCL were identified, of which some were found to have clinical significance and could be included, with additional validation, in patient risk assessment. This analysis also afforded the identification of novel genes with possible roles in lymphomagenesis, representing potential therapeutic targets in DLBCL. Disclosures: Dias: Cancer Genetics, Inc.: Employment. Thodima:Cancer Genetics, Inc.: Employment. Mendiratta:Cancer Genetics, Inc.: Employment. Asha:Cancer Genetics, Inc.: Employment. Feldstein:Memorial Slaon-Kettering Cancer Center: Employment. Chaganti:Cancer Genetics, Inc.: Consultancy, Equity Ownership, Patents & Royalties. Houldsworth:Cancer Genetics, Inc.: Employment, Equity Ownership.
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Hsieh, Y. T., C. Hubeau, V. Massa, W. LI, S. Frei, B. Capraro, A. Umana et al. "OP0316 EMERGING BEST-IN-CLASS IL-2 VARIANT HIGHLIGHTS TREG-DIRECTED THERAPY FOR AUTOIMMUNE DISEASE". Annals of the Rheumatic Diseases 79, Suppl 1 (giugno 2020): 195.1–195. http://dx.doi.org/10.1136/annrheumdis-2020-eular.1999.
Testo completoAbstract (sommario):
Background:Impairment or deficiency of regulatory T cells (Treg) is associated with chronic inflammation and autoimmune diseases. Interleukin 2 (IL-2) is a cytokine indispensable for Treg expansion and immunosuppressive function. However, expansion of cytotoxic effector T (Teff) and NK cells and the associated vascular leakage side effect limit the use of IL-2 in autoimmune diseases [1].Objectives:Cugene developed a long-acting IL-2 variant with high Treg specificity and low toxicity to restore immune homeostasis and self-tolerance, and potentially cure autoimmune and inflammatory diseases.Methods:IL-2 variants were generated based on the quaternary structure of IL-2 and IL-2Rαβγ (alpha, beta, gamma) complex. Biological activity was determined by examining differential signaling activity in induction of STAT5 phosphorylation in defined lymphocyte populations of human PBMC using flow cytometry. Binding activity was evaluated by ELISA. Pharmacokinetics, pharmacodynamics, safety and tolerability were assessed in mice and cynomolgus monkeys. Treg suppressive function was determinedin vivo/ex vivo,and anti-inflammatory and anti-antibody production efficacy were determined in delayed-type hypersensitivity (DTH) and T-cell-dependent antibody response (TDAR) models.Results:Structure-based rational design and activity-guided fine-tuning generated an optimized IL-2 variant, CUG252. It demonstrated a strong and near wild-type IL-2 ability to stimulate STAT5 phosphorylation in IL-2Rαβγ dominant Treg cells but abolished activities in IL-2Rβγ dominant effector CD4, CD8 and NK cells. This was a result of biased binding activity to IL-2Rα while dramatically attenuated binding to IL-2Rβγ complex. In mice and monkeys, administration of CUG252 resulted in dose-dependent increases in Treg proliferation and expansion by more than 10- and 30-fold, respectively, with largely abolished activities in CD4+ T conventional, cytotoxic CD8+ Teff and NK cells. The ratio of Treg/Teff cells achieved was as high as 0.4 in mice and 1.2 in monkeys. Both CD4+ and CD8+ Tregs were expanded with preferential increases in memory over naïve subsets. A substantial increase in Treg-suppressive capacity over T effector cells was corroborated by enhanced expression of functional and inhibitory markers, including CD25, Foxp3, PD-1, CTLA-4, Tim3 and ICOS. In DTH and TDAR models, CUG252 strongly inhibited antigen-driven inflammation, B cell maturation, and antibody production. The sustained PK/PD profile supports monthly dosing or better in humans. CUG252 was well-tolerated and no changes in body weight, body temperature, clinical pathology or signs of vascular leakage were observed. Moreover, CUG252 demonstrated superior manufacturability.Conclusion:CUG252 demonstrates an emerging best-in-class profile among IL-2 variants. It displayed exquisite Treg-selectivity while retaining potency comparable to wild-type IL-2. It showed strong anti-inflammatory and anti-antibody production efficacy with significantly improved therapeutic index and manufacturability. Its favorable drug-like property and robust preclinical efficacy warrant further evaluation in patients with a variety of inflammation and autoimmune diseases.References:[1]Tahvildari M. et al. Low-Dose IL-2 Therapy in Transplantation, Autoimmunity, and Inflammatory Diseases. J Immunol. 2019; 203: 2749-2755Disclosure of Interests:Yao-te Hsieh Employee of: Cugene INC., CEDRIC HUBEAU Employee of: Cugene INC., Virginia MASSA Employee of: Cugene INC., WEN Li Employee of: Cugene INC., SANDRA FREI Employee of: Cugene INC., BEN CAPRARO Employee of: Cugene INC., ANDREA UMANA Employee of: Cugene INC., ANDREW AHERRERA Employee of: Cugene INC., YUESHENG LI Employee of: Cugene INC., JING XU Employee of: Cugene INC., LINGYUN RUI Employee of: Cugene INC.
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Harrington, Deirdre M., Sarahjane Belton, Tara Coppinger, Muireann Cullen, Alan Donnelly, Kieran Dowd, Teresa Keating et al. "Results from Ireland’s 2014 Report Card on Physical Activity in Children and Youth". Journal of Physical Activity and Health 11, s1 (gennaio 2014): S63—S68. http://dx.doi.org/10.1123/jpah.2014-0166.
Testo completoAbstract (sommario):
Background:Physical activity (PA) levels are a key performance indicator for policy documents in Ireland. The first Ireland Report Card on Physical Activity in Children and Youth aims to set a robust baseline for future surveillance of indicators related to PA in children and youth.Methods:Data collected between 2003−2010 on more than 35,000 7- to 18-year-old children and youth were used and graded using a standardized grading system for 10 indicators.Results:Grades assigned for the indicators were as follows: overall physical activity levels, D-; sedentary behavior (TV viewing), C-; organized sport participation, C-: physical education, D-; active play, inconclusive (INC); active transportation, D; school, C-, community and the built environment, B; family, INC; and government, INC.Conclusions:PA recommendations exist in Ireland but this Report Card has shown that participation is still low. A number of promising policies, programs and services are in place but these require thorough evaluation and adequate resourcing. Agreement and implementation of a common framework for the systematic surveillance of indictors related to PA of children and youth is necessary to monitor change over time and ensure the impact of promising work is captured.
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Riley, Brigit E., Jeff Boonsripisal, Alicia Goodwin, Dominique Cartier, Eudean Garces, Kate Nesbitt, Courtney Dwyer et al. "Development of an Optimized rAAV2/6 Human Factor 8 cDNA Vector Cassette for Hemophilia a Gene Therapy". Blood 128, n. 22 (2 dicembre 2016): 1173. http://dx.doi.org/10.1182/blood.v128.22.1173.1173.
Testo completoAbstract (sommario):
Abstract Hemophilia A, which is caused by a mutation in the Factor 8 (F8) gene resulting in a deficiency or lack of the Factor VIII (FVIII) protein, is the most common inherited bleeding disorder in humans with an estimated worldwide incidence of half a million people. The disorder is X-linked and occurs in approximately 1 in 5,000 males; however there is also a growing appreciation of the impact on carrier females having a single mutant allele, with at least 10% of hemophilia A female carriers having less than normal clotting activity. Even modest increases in Factor V III activity (>1% of normal) can have a positive impact on patient lives, thus making the disease an ideal candidate for liver-directed gene therapy. Recombinant AAV (rAAV) has been used extensively for nearly 20 years as a gene therapy vector in preclinical and clinical studies where rAAV delivery to non-dividing tissues such as liver, brain and muscle affords stable, long-term transgene expression. However, there has been a lag in the clinical translation of a rAAV gene therapy approach for Hemophilia A/human F8 (hF8) compared to Hemophilia B/human Factor 9 due to poor yields of rAAV encoding a F8 transgene at clinical scale, and a requirement for large doses of rAAV F8 vector to achieve therapeutically relevant levels of circulating human FVIII (hFVIII), with the attendant risk of inducing an AAV-directed immune response requiring transient immunosuppression. To address these issues we optimized a rAAV F8 cDNA vector cassette to improve both virus yields and liver-specific hFVIII expression. The rAAV F8 cDNA vector cassette optimization required multi-factorial modifications to the liver-specific promoter module, hF8 transgene, synthetic polyadenylation signal and vector backbone sequence. This iterative process resulted in improved vector yields at research scale and greater than five-fold improvement in vector yields at clinical scale using our proven manufacturing process. Administration of the optimized rAAV hF8 cDNA packaged in serotype AAV2/6 at a dose of ~7.2E+12 vg/kg to both wild type and Hemophilia A mice resulted in robust circulating hFVIII levels and activity (levels in wild type mice were 241.6% of normal, and activity in Hemophilia A mice were 330.9% of normal). An analysis of hF8 mRNA levels in different tissues following dosage with our optimized vector demonstrated that hF8 expression from the modified promoter module was restricted to the liver. Notably there was a striking impact on hemostasis in the Hemophilia A mice treated with the optimized rAAV hF8 cDNA, with a reduction in bleeding time from 38.3 minutes to 2.5 minutes in treated mice (n = 12, p-value < 0.0001), which is in line with bleeding times in wild type mice. Initial studies in non-human primates (NHPs) resulted in supraphysiological levels of circulating hFVIII with mean peak values of 400-600% of normal levels. A follow up dose-ranging study was performed in NHPs with a rAAV2/6 F8 cDNA vector manufactured using our GMP clinical manufacturing process. Administration of vector doses ranging from 6E+11 vg/kg to 6E+12 vg/kg resulted in therapeutic circulating levels of hFVIII that were 5% - 229% of normal levels. The peak circulating hFVIII levels achieved in this dose-ranging study using GMP clinical-scale vector exceeds the levels previously reported in NHPs by several fold on an AAV vector dose basis. The high potency of this enhanced rAAV F8 cDNA cassette could significantly reduce the dose required to achieve therapeutically relevant levels in human subjects and reduce the potential of developing immune responses to AAV capsid requiring immunosuppression. Disclosures Riley: Sangamo BioSciences Inc: Employment. Boonsripisal:Sangamo BioSciences Inc: Employment. Goodwin:Sangamo BioSciences Inc: Employment. Garces:Sangamo BioSciences Inc: Employment. Ballaron:Sangamo BioSciences Inc: Employment. Tran:Sangamo BioSciences Inc: Employment. Kang:Sangamo BioSciences Inc: Employment. Zhang:Sangamo BioSciences Inc: Employment. Meyer:Sangamo BioSciences Inc: Employment. Greengard:Sangamo BioSciences Inc: Employment. Surosky:Sangamo BioSciences Inc: Employment. Ando:Sangamo BioSciences Inc: Employment. Lillicrap:bayer: Research Funding; biogen: Research Funding; CSL: Research Funding; Octapharma: Research Funding; Sangamo Biosciences Inc: Research Funding. Nichol:Sangamo BioSciences Inc: Employment. Holmes:Sangamo BioSciences Inc: Employment.
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Dias, Lizalynn, Venkata Thodima, Asha Guttapalli, Geetu Mendiratta, Sergei I. Syrbu e Jane Houldsworth. "Robust Assessment of Genomic Imbalance in Diffuse Large B-Cell Lymphoma Confirms Inferior Outcome Is Associated with Genomic Complexity and Identifies Potential Therapeutic Pathway Targets". Blood 124, n. 21 (6 dicembre 2014): 2993. http://dx.doi.org/10.1182/blood.v124.21.2993.2993.
Testo completoAbstract (sommario):
Abstract Despite excellent initial responses of diffuse large B-cell lymphoma (DLBCL) patients to current frontline immunotherapy (RCHOP), only about 40% of patients are ultimately cured, with most relapses occurring within the first 2-3 years. Apart from clinical features, very few biomarkers are available and validated for risk stratification of DLBCL patients, other than cell-of-origin (COO) subtyping and MYC rearrangement. Genomic copy number changes identified using a variety of genomic profiling technologies including massively parallel sequencing, have variously been reported to have prognostic value in DLBCL. Our previous studies used a common analytical approach across three different clinical datasets to identify and define 32 common regions (CR) of overlapping genomic imbalance (17 gain, 15 loss), comprising 36 minimal common regions (MCR) observed in newly-diagnosed DLBCL. We now report the establishment of standardized calling criteria for a total of 50 aberrations within the 32 CR, taking into consideration GISTIC-defined peaks based on copy number data from three publicly available datasets: IS-172 (GSE11318, n=170), IS-51HR (E-MEXP-3463, n=51 with high IPI), and one similarly analyzed dataset IS-180 (GSE34171, n=180). As a form of validation of the scoring criteria, the frequencies of the aberrations detected in IS-172 in the known GCB and ABC COO subtypes were compared with those previously reported (PMID:18765795). The relative imbalance of the aberrations between the subtypes were re-capitulated including six occurring at higher frequency in the GCB subtype and five in the ABC subtype. Integration of the expression profiles of 162 samples of IS-172 by univariate t-test with the 50 aberrations revealed 569 unique positively-correlated Refseqs mapping to 24 of the MCRs with at least 1.2 fold change in expression (Univariate t-test with P ≤ 0.05 after Benjamini-Hochberg false discovery rate [FDR]). Of these, 27 were located within overlapping peaks of gain/loss. Ingenuity Pathway Analysis determined five significant pathways (P ≤ 0.001, FDR<0.10): p53 signaling, PKC theta signaling in T lymphocytes, geranylgeranyl-diphosphate biosynthesis, B-cell receptor signaling, and RANK signaling in osteoclasts. Association of each of the defined 50 aberrations with overall survival (OS) was performed with the Kaplan-Meier method and log-rank statistic in three RCHOP datasets: IS-124 (GSE15127, n=124), a subset of 70 of IS-180 (IS-70), and a dataset of 41 for which array-CGH was performed in-house using a targeted oligonucleotide (Agilent Technologies) with DNA extracted from sections of formalin–fixed paraffin-embedded de novo DLBCL (IH-41). Ten aberrations significantly associated (P ≤0.05) with poor outcome in at least one of the datasets: gain of 12q (12q13 and 12q14), 16q24, 19q13, and loss of 2q24, 2 sites at 6q21, 8p22, 9p21, 15q15, and 17p13. Of these 10 aberrations, loss of 17p significantly associated with gain of 16q, and losses of 6q, 8p, and 15q (P <0.05), while loss of 9p21 correlated with loss of 2q, 6q, and 8p as evaluated using the exact t-test. Gain of the 2 sites on 12q did not significantly correlate with any of the other 8 aberrations. Genomic complexity was assessed by two methods. In the first, the median number of MCR aberrations was determined to be 1, across three datasets (IS-172, IS-180, and IS-124). Specimens in RCHOP-treated datasets IS-124 and IS-70 were then called “complex” if 2 or more MCR aberrations were detected: 31% and 67% respectively. In both datasets, a complex genome was significantly associated with overall unfavorable outcome (P=0.037, 0.006 respectively). Specimens in IS-172 and IS-70 were additionally classified as “complex” based on the presence of at least one of nine aberrations involving the CDKN2A-TP53-RB-E2F axis (PMID:22975378) where 33% and 57% of cases were scored as complex respectively. Using this approach, significant association with outcome was only found for IS-70 (P<0.001). Of note, between “complex” scoring approaches, 11% and 13% discordance was evident. In summary, establishment of robust scoring criteria for copy number changes afforded correlative analyses with clinical features to reveal that genomic complexity is indeed associated with overall inferior survival in DLBCL and that integrated expression analysis identified biological pathways in DLBCL that may represent potential therapeutic targets. Disclosures Dias: Cancer Genetics,Inc: Employment. Thodima:Cancer Genetics,Inc: Employment, Stock option holder Other. Guttapalli:Cancer Genetics, Inc: Employment, Stock option holder Other. Mendiratta:Cancer Genetics,Inc: Employment. Houldsworth:Cancer Genetics, Inc.: Employment, Stock and stock option holder Other.
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Anguela, Xavier M., Rajiv Sharma, Yannick Doyon, Sunnie Y. Wong, David E. Paschon, Hojun Li, Virginia Haurigot et al. "In Vivo Genome Editing of Liver Albumin for Therapeutic Gene Expression: Rescue of Hemophilic Mice Via Integration of Factor 9". Blood 120, n. 21 (16 novembre 2012): 751. http://dx.doi.org/10.1182/blood.v120.21.751.751.
Testo completoAbstract (sommario):
Abstract Abstract 751 Gene correction using zinc finger nuclease (ZFN) technology can be applied to target virtually any locus in the human genome. Beyond correcting mutated genes causative of disease, ZFNs can also be utilized to target transgene insertion into genomic “safe harbors.” Ideally, specific gene targeting to such “safe harbor” sites would (i) ensure therapeutically relevant levels of transgene expression and (ii) tolerate transgene addition without deleterious effect on the host organism. For liver-derived protein replacement, albumin represents an attractive target locus. Firstly, albumin is very highly expressed exclusively in the liver, thus targeting of a relatively small percentage of alleles should yield therapeutically relevant levels of liver-specific transgene expression. Second, the reduction or complete absence of albumin in animals and even humans (analbuminemia) produces surprisingly few symptoms. Here, we sought to investigate whether ZFN-mediated targeted insertion of a promoter-less copy of the human F9 cDNA at the mouse albumin locus could result in human Factor IX production and successfully correct the hemophilic phenotype in mice. To address this question, we constructed an AAV vector encoding a pair of ZFNs targeting intron 1 of the mouse albumin locus (AAV8-mAlb-ZFN) and a donor AAV vector (AAV8-Donor) harboring a partial cDNA cassette containing exons 2–8 of the wild-type human F9 gene flanked by sequences lacking significant homology to the mouse genome. Co-delivery of 1e11 vg of AAV8-mAlb-ZFN along with 5e11vg of AAV8-Donor resulted in stable (>12wk) circulating F.IX levels of 1600–3200 ng/mL (32–64% of normal). As a control, mice injected with the AAV8-Donor along with an AAV vector encoding a ZFN pair targeting an unrelated locus exhibited background F.IX levels (∼50 ng/mL). A dose-response study was performed by administering a fixed dose of donor (5e11 vg/mouse) with decreasing doses of AAV8-mAlb-ZFN (1e11, 1e10 and 1e9 vg/mouse). Human F.IX levels increased as a function of ZFN dose in the range tested (3260±480, 225±43 and 31±4 ng/mL at the high, medium and low dose, respectively). Importantly, these results showed that donor homology to the target site is not required to achieve robust levels of gene addition to the albumin locus in adult mice, thus permitting the design of donor vectors harboring corrective copies of transgenes up to the maximum AAV packaging capacity of ∼4.7 Kb. Albumin and factor IX are both synthesized as pre-propeptides and turned into propeptides after the signal peptide is removed. Expression of human F9 exons 2–8 spliced with mouse albumin exon 1 is expected to yield a chimeric propeptide. The first 2 N-terminal amino acids would originate from proalbumin, followed by a Val to Leu mutation at position −17 of the hF.IX propeptide and 16 aa encoded by human F9. To evaluate whether this chimeric human F.IX derived from gene addition to the albumin locus would be processed correctly and normalize the prolonged clotting times in hemophilia B (HB) mice, we injected 1e11 vg of AAV8-mAlb-ZFN and 5e11vg of AAV8-Donor into HB animals. Two weeks post-treatment, hF.IX antigen levels were in the range of 20% of normal and activated partial thromboplastin time, a measurement of clot formation, was corrected to wild-type levels (42 seconds), from an average of 70 seconds pre-treatment. Thus expression of a therapeutic protein (F.IX) from the albumin locus is shown to correct the HB disease phenotype in vivo. In summary, these data provide the first demonstration of ZFN-mediated in vivo genome editing of a safe harbor locus for therapeutic protein production. While we provide here a proof of principle establishing phenotypic correction of hemophilia B, appropriately designed donors could expand this strategy. Most importantly the magnitude of albumin expression (>15 g / day) should enable production of a diverse range of transgenes at therapeutically consequential levels. Disclosures: Anguela: The Children's Hospital of Philadelphia: Patents & Royalties. Sharma:The Children's Hospital of Philadelphia: Patents & Royalties. Doyon:Sangamo BioSciences, Inc.: Employment. Wong:Sangamo BioSciences, Inc.: Employment. Paschon:Sangamo BioSciences, Inc.: Employment. Gregory:Sangamo BioSciences, Inc.: Employment. Holmes:Sangamo BioSciences, Inc.: Employment. Rebar:Sangamo BioSciences, Inc.: Employment. High:Shire Pharmaceuticals: Consultancy; Sangamo Biosciences, Inc: Collaborator, Collaborator Other; Novo Nordisk: Visiting Professor, Visiting Professor Other; Genzyme, Inc: Membership on an entity's Board of Directors or advisory committees; The Children's Hospital of Philadelphia: Patents & Royalties; Bluebird Bio, Inc: Membership on an entity's Board of Directors or advisory committees.
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Timmerman, John M., Kristopher K. Steward, Reiko E. Yamada, Patricia A. Young, Dena M. Minning, Raj K. Sachdev, Michael J. Gresser, Sanjay D. Khare e Sherie L. Morrison. "Antibody-Interferon-Alpha Fusion Protein Therapy for the Treatment of B-Cell Non-Hodgkin Lymphoma: Enhanced ADCC, Inhibition of Proliferation, and In Vivo Eradication of CD20+ Human Lymphomas". Blood 126, n. 23 (3 dicembre 2015): 2762. http://dx.doi.org/10.1182/blood.v126.23.2762.2762.
Testo completoAbstract (sommario):
Abstract Background: Interferon-alpha (IFNα) is a pleiotrophic cytokine with direct anti-tumor and immunostimulatory effects. Currently IFNα is approved for the treatment of multiple hematologic malignancies, including non-Hodgkin lymphoma (NHL). However, its clinical utility has been hindered by dose-limiting toxicitiy due to systemic activation of the interferon receptor. To overcome this limitation, we engineered anti-tumor antibody-IFNα fusion proteins to selectively increase delivery of IFN to the tumor site and reduce systemic toxicity. We previously reported that IGN002, an anti-CD20-IFNα fusion protein, exhibits enhanced complement-dependent cytotoxicity (CDC) compared to rituximab, and inhibits proliferation and induces apoptosis of human B-cell NHL (Yamada et al, ASCO 2013). We now extend these previous findings and show that IGN002 possesses enhanced antibody-dependent cell-mediated cytotoxicity (ADCC) effector function and superior in vivo anti-tumor activity against B-cell NHL, compared to rituximab. Methods: IGN002 was evaluated against a panel of human Burkitt and diffuse large B-cell lymphoma (DLBCL) cell lines. Proliferation was measured by [3 H]-thymidine incorporation, STAT1 activation by flow cytometry, ADCC by lactate dehydrogenase release using human PBMC effectors, and IFN bioactivity by encephalomyocarditis (EMC) viral protection assay. NHL xenografts were grown in SCID mice. Results: IGN002 more potently inhibited the growth of NHL cell lines expressing CD20 than rituximab or unfused IFNα. Intrinsic IFNα activity of IGN002 was reduced in viral protection and anti-proliferation assays using cells lacking CD20 expression. STAT1 activation by IGN002 was enhanced on cells expressing the target antigen, whereas a control antibody-IFNα fusion protein showed reduced STAT activation activity compared to unfused IFNα. Together, these results indicate that fusion of IFNα to the antibody results in reduced IFN effects on cells not bearing the tumor antigen target. IGN002 exhibited enhanced ADCC activity compared to rituximab against Daudi, Ramos, and Raji NHL cells in long-term (overnight incubation) assays, demonstrating both higher potency and higher maximal cytotoxicity. This result is possibly due to activation of the effector cell populations by the fused IFNα moiety, as IFN is known to activate both NK cells and monocytes. The in vivo anti-tumor efficacy of IGN002 was compared to rituximab and a control antibody-IFNα fusion protein against 10-day established Raji NHL xenografts. IGN002 was superior to both rituximab and the control fusion protein, achieving a longer median survival and higher long-term survival rate (p = 0.0015 and < 0.0001 vs. rituximab and control fusion protein, respectively). The in vivo anti-tumor efficacy of IGN002 was also compared to rituximab at three equimolar dose levels (5 mg/kg, 1 mg/kg, and 0.2 mg/kg antibody) against 10-day established Daudi NHL xenografts. IGN002 showed superior efficacy compared to rituximab at all doses (p < 0.001), achieving tumor eradication (100% long-term survival) in all mice treated at all three dose levels, whereas rituximab only delayed tumor progression. Conclusions: IGN002 demonstrated more robust direct anti-proliferative and antibody effector functions than rituximab against human NHL cells in vitro, and also showed the ability to eradicate established NHL xenografts in vivo. Against cells expressing the CD20 target antigen, IGN002 exhibited greater anti-proliferative potency than unfused IFNα. In contrast, the anti-proliferative and anti-viral potency of IGN002 was reduced against cells lacking CD20, compared to unfused IFNα. These findings support the hypothesis that tumor antigen-targeted IFN therapeutics may possess a broader therapeutic index than unfused IFNα, inhibiting tumor growth by multiple mechanisms while reducing systemic toxicity. These results support the further development of IGN002 for the treatment of B-cell NHL, and a first-in-human phase I clinical study will begin later this year in the United States. Disclosures Timmerman: Janssen: Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; Valor Biotherapeutics: Research Funding. Steward:ImmunGene, Inc.: Employment. Minning:Valor Biotherapeutics, LLC: Consultancy. Sachdev:ImmunGene, Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Gresser:ImmunGene, Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Valor Biotherapeutics: Membership on an entity's Board of Directors or advisory committees. Khare:Valor Biotherapeutics: Membership on an entity's Board of Directors or advisory committees; ImmunGene, Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Morrison:ImmunGene, Inc.: Membership on an entity's Board of Directors or advisory committees, Research Funding.
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Chekmasova, Alena A., Holly M. Horton, Tracy E. Garrett, John W. Evans, Johanna Griecci, Amanda Hamel, Howard J. Latimer et al. "A Novel and Highly Potent CAR T Cell Drug Product for Treatment of BCMA-Expressing Hematological Malignances". Blood 126, n. 23 (3 dicembre 2015): 3094. http://dx.doi.org/10.1182/blood.v126.23.3094.3094.
Testo completoAbstract (sommario):
Abstract Recently, B cell maturation antigen (BCMA) expression has been proposed as a marker for identification of malignant plasma cells in patients with multiple myeloma (MM). Nearly all MM and some lymphoma tumor cells express BCMA, while normal tissue expression is restricted to plasma cells and a subset of mature B cells. Targeting BCMA maybe a therapeutic option for treatment of patients with MM and some lymphomas. We are developing a chimeric antigen receptor (CAR)-based therapy for the treatment of BCMA-expressing MM. Our anti-BCMA CAR consists of an extracellular single chain variable fragment (scFv) antigen recognition domain derived from an antibody specific to BCMA, fused to CD137 (4-1BB) co-stimulatory and CD3zeta chain signaling domains. Selection of our development candidate was based on the screening of four distinct anti-BCMA CARs (BCMA01-04) each comprised of unique single chain variable fragments. One candidate, BCMA02 (drug product name bb2121) was selected for further studies based on the robust frequency of CAR-positive cells, increased surface expression of the CAR molecule, and superior in vitro cytokine release and cytolytic activity against the MM cell lines. In addition to displaying specific activity against MM (U226-B1, RPMI-8226 and H929) and plasmacytoma (H929) cell lines, bb2121 was demonstrated to react to lymphoma cell lines, including Burkitt's (Raji, Daudi, Ramos), chronic lymphocytic leukemia (Mec-1), diffuse large B cell (Toledo), and a Mantle cell lymphoma (JeKo-1). Based on receptor density quantification, bb2121 can recognize tumor cells expressing less than 1000 BCMA molecules per cell. The in vivo pharmacology of bb2121 was studied in NSG mouse models of human MM and Burkitt's lymphoma. NSG mice were injected subcutaneously (SC) with 107 RPMI-8226 MM cells. After 18 days, mice received a single intravenous (IV) administration of vehicle or anti-CD19Δ (negative control, anti-CD19 CAR lacking signaling domain) or anti-BCMA CAR T cells, or repeated IV administration of bortezomib (Velcade®; 1 mg/kg twice weekly for 4 weeks). Bortezomib, which is a standard of care for MM, induced only transient reductions in tumor size and was associated with toxicity, as indicated by substantial weight loss during dosing. The vehicle and anti-CD19Δ CAR T cells failed to inhibit tumor growth. In contrast, treatment with bb2121 resulted in rapid and sustained elimination of the tumors, increased body weights, and 100% survival. Flow cytometry and immunohistochemical analysis of bb2121 T cells demonstrated trafficking of CAR+ T cells to the tumors (by Day 5) followed by significant expansion of anti-BCMA CAR+ T cells within the tumor and peripheral blood (Days 8-10), accompanied by tumor clearance and subsequent reductions in circulating CAR+ T cell numbers (Days 22-29). To further test the potency of bb2121, we used the CD19+ Daudi cell line, which has a low level of BCMA expression detectable by flow cytometry and receptor quantification analysis, but is negative by immunohistochemistry. NSG mice were injected IV with Daudi cells and allowed to accumulate a large systemic tumor burden before being treated with CAR+ T cells. Treatment with vehicle or anti-CD19Δ CAR T cells failed to prevent tumor growth. In contrast, anti-CD19 CAR T cells and anti-BCMA bb2121 demonstrated tumor clearance. Adoptive T cell immunotherapy approaches designed to modify a patient's own lymphocytes to target the BCMA antigen have clear indications as a possible therapy for MM and could be an alternative method for treatment of other chemotherapy-refractory B-cell malignancies. Based on these results, we will be initiating a phase I clinical trial of bb2121 for the treatment of patients with MM. Disclosures Chekmasova: bluebird bio, Inc: Employment, Equity Ownership. Horton:bluebird bio: Employment, Equity Ownership. Garrett:bluebird bio: Employment, Equity Ownership. Evans:bluebird bio, Inc: Employment, Equity Ownership. Griecci:bluebird bio, Inc: Employment, Equity Ownership. Hamel:bluebird bio: Employment, Equity Ownership. Latimer:bluebird bio: Employment, Equity Ownership. Seidel:bluebird bio, Inc: Employment, Equity Ownership. Ryu:bluebird bio, Inc: Employment, Equity Ownership. Kuczewski:bluebird bio: Employment, Equity Ownership. Horvath:bluebird bio: Employment, Equity Ownership. Friedman:bluebird bio: Employment, Equity Ownership. Morgan:bluebird bio: Employment, Equity Ownership.
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Younes, Anas, Connie Lee Batlevi, Jonathon B. Cohen, Sven de Vos, Daniel J. Landsburg, Zaw Win Myint, Krish Patel et al. "A Multi-Center Dose-Finding Study to Assess Safety, Tolerability, Pharmacokinetics and Preliminary Efficacy of Fimepinostat (CUDC-907) in Combination with Venetoclax in Patients with Relapsed/Refractory (R/R) Lymphoma". Blood 134, Supplement_1 (13 novembre 2019): 4104. http://dx.doi.org/10.1182/blood-2019-128857.
Testo completoAbstract (sommario):
Background High-grade B-cell lymphoma (HGBL) with MYC and BCL2 and/or BCL6 rearrangements (double- and triple-hit lymphoma), as well as diffuse large B-cell lymphoma (DLBCL), NOS with increased expression of MYC and BCL2 protein (double-expressor lymphoma) are associated with a poor prognosis after front-line treatment with standard immunochemotherapy. As such, therapies targeting MYC and BCL2 alterations are urgently needed. Currently, there are no approved therapies that target MYC. Fimepinostat is an investigational small molecule dual inhibitor of phosphotidyl-inositol-3-kinases (PI3Ks) and Class I and II histone deacetylases (HDACs). Both of fimepinostat's dual mechanisms of action lead to decreased MYC protein: PI3K inhibition leads to enhanced ubiquitin-mediated MYC protein degradation, and HDAC inhibition leads to repression of MYC gene expression. PI3K and HDAC were also inhibited by fimepinostat in peripheral blood mononuclear cells collected from patients (pts) receiving fimepinostat therapy. When dosed in combination with venetoclax in non-clinical studies, fimepinostat demonstrated striking synergistic anti-tumor effects in vitro and in vivo with nearly 100% tumor growth inhibition in a double-hit lymphoma mouse xenograft model (Landsburg 2018a). In clinical studies, fimepinostat +/- rituximab was well tolerated with a favorable safety profile in pts with R/R lymphoma, and resulted in robust and durable objective response rates (ORR) in pts with R/R MYC-altered DLBCL with an ORR of 23% and a median duration of response (DOR) of 13.6 months (Landsburg 2018b). Study Design and Methods CUDC-907-101 is a Phase 1/2, multi-center, dose-finding study that was recently amended to evaluate fimepinostat in combination with other anti-cancer therapies, including venetoclax. Cohorts of patients will receive increasing dose levels of fimepinostat administered on a 5-days-on-2-days-off (5/2) schedule in combination with venetoclax in 21-day cycles (Table 1). The primary objectives are to determine the maximum tolerated dose, PK, safety and tolerability, and to assess preliminary efficacy, as measured by the ORR and DOR. Eligible pts must have a histologically-confirmed diagnosis of DLBCL or HGBL with or without MYC and/or BCL2 alterations, which is refractory to, or relapsed after, ≥1 prior lines of therapy. Patients must also have an ECOG performance status of 0 or 1, a life expectancy of ≥3 months, measurable disease per Lugano criteria, and have archived or fresh tumor tissue available. Approximately 12 pts in the Ph 1 dose escalation (3+3 design) and 30 pts in the Ph 2 expansion will be enrolled to receive fimepinostat + venetoclax treatment. Patients will be treated until progression or unacceptable toxicity. The Ph 2 expansion will be an estimation study for detecting an efficacy signal. Patients who receive ≥1 dose and have ≥1 post-baseline response evaluation will be included in the efficacy analysis set. Investigator-assessed ORR based on Lugano criteria will be summarized as the proportion of pts who achieve a best response of CR or PR for each combination, and the corresponding two-sided 95% confidence interval (CI, Clopper-Pearson) will be calculated. DOR will be summarized for pts who achieve response using the Kaplan-Meier (KM) product-limit method. The median DOR along with the two-sided 95% CI using the Brookmeyer and Crowley method will be calculated. PFS and OS will be estimated in pts using the KM product-limit method, along with the median and two-sided 95% CI. The first patient in this study was treated in July 2019, and enrollment is on-going. This new study represents the first clinical trial of the novel-novel combination of fimepinostat with venetoclax in pts with non-Hodgkin lymphoma harboring alterations of both MYC and BCL2. Clinical trial: NCT01742988. References a. Landsburg, D. J. et al., Durable Responses Achieved in Patients with MYC-altered Relapsed/Refractory Diffuse Large B-cell Lymphoma Treated with Fimepinostat (CUDC-907): Combined Results from a Phase 1 and Phase 2 Study. Poster presented at: Society of Hematologic Oncology annual meeting. September 12-15, 2018. b. Landsburg, D. J. et al., (2018). A Pooled Analysis of Relapsed/Refractory Diffuse Large B-Cell Lymphoma Patients Treated with the Dual PI3K and HDAC Inhibitor Fimepinostat (CUDC-907), Including Patients with MYC-Altered Disease. Blood,132(Suppl 1), 4184. Disclosures Younes: Epizyme: Consultancy, Honoraria; Roche: Consultancy, Honoraria, Research Funding; Janssen: Honoraria, Research Funding; AstraZeneca: Research Funding; Genentech: Research Funding; Biopath: Consultancy; Xynomics: Consultancy; Syndax: Research Funding; Curis: Honoraria, Research Funding; Merck: Honoraria, Research Funding; Abbvie: Honoraria; Celgene: Consultancy, Honoraria; HCM: Consultancy; BMS: Research Funding; Pharmacyclics: Research Funding; Takeda: Honoraria. Batlevi:Juno Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees. Cohen:Takeda Pharmaceuticals North America, Inc.: Research Funding; Genentech, Inc.: Consultancy, Research Funding; Bristol-Meyers Squibb Company: Research Funding; Seattle Genetics, Inc.: Consultancy, Research Funding; Gilead/Kite: Consultancy; LAM Therapeutics: Research Funding; UNUM: Research Funding; Hutchison: Research Funding; Astra Zeneca: Research Funding; Lymphoma Research Foundation: Research Funding; Janssen Pharmaceuticals: Consultancy; ASH: Research Funding. de Vos:Verastem: Consultancy; Portola Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Bayer: Consultancy. Landsburg:Triphase: Research Funding; Seattle Genetics: Speakers Bureau; Takeda: Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Curis, INC: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Curis, INC: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Seattle Genetics: Speakers Bureau; Triphase: Research Funding; Takeda: Research Funding. Patel:Sunesis: Consultancy; AstraZeneca: Consultancy, Research Funding, Speakers Bureau; Celgene: Consultancy, Speakers Bureau; Genentech: Consultancy, Speakers Bureau; Pharmacyclics/Janssen: Consultancy, Speakers Bureau. Phillips:Celgene: Membership on an entity's Board of Directors or advisory committees; Abbvie: Research Funding; Pharmacyclics: Consultancy, Research Funding; Genentech: Consultancy; Incyte: Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy; Seattle Genetics: Consultancy; Bayer: Consultancy. Smith:Portola Pharmaceuticals: Research Funding. Westin:Novartis: Other: Advisory Board, Research Funding; 47 Inc: Research Funding; MorphoSys: Other: Advisory Board; Kite: Other: Advisory Board, Research Funding; Genentech: Other: Advisory Board, Research Funding; Curis: Other: Advisory Board, Research Funding; Juno: Other: Advisory Board; Unum: Research Funding; Celgene: Other: Advisory Board, Research Funding; Janssen: Other: Advisory Board, Research Funding. Ma:Curis, Inc.: Employment. Grayson:Curis, Inc.: Employment. von Roemeling:Curis, Inc.: Employment. Barta:Takeda: Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees; Celgene: Research Funding; Mundipharma: Honoraria; Seattle Genetics: Honoraria, Research Funding; Bayer: Consultancy, Research Funding; Mundipharma: Honoraria; Merck: Research Funding; Celgene: Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees.
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Bjordahl, Ryan, Raedun Clarke, Svetlana Gaidarova, Brian Groff, Paul Rogers, Stacey Moreno, Ramzey Abujarour et al. "Off-the-Shelf Natural Killer Cell Immunotherapy for Enhanced Antibody Directed Cellular Cytotoxicity". Blood 128, n. 22 (2 dicembre 2016): 3363. http://dx.doi.org/10.1182/blood.v128.22.3363.3363.
Testo completoAbstract (sommario):
Abstract Natural Killer (NK) cells play a crucial role in immunosurveillance and form a first line of defense against cancer. In comparison to other lymphocytes, NK cells are unique in their capability to elicit tumoricidal responses without the need for antigen presentation or prior sensitization. Clinical data from bone marrow transplant and allogeneic NK immunotherapy suggest that MHC mismatch is advantageous in promoting graft-versus-leukemia without eliciting graft-versus-host, providing evidence that NK cells hold promisa as an allogeneic, universal immunotherapeutic. Further, the anti-tumor effect of many monoclonal antibodies is mediated through binding of the low-affinity Fc receptor CD16 on NK cells, which induces tumor cell killing through antibody-dependent cellular cytotoxicity (ADCC). Thus, NK cells represent a unique source of effector cells that can be combined with monoclonal antibodies, bispecific engagers or chimeric antigen receptors to direct tumor specificity and enhance cytotoxicity. Despite the significant potential of NK cell therapy, current clinical practices are limited by the need for large numbers of healthy NK cells, lack of in vivo persistence, and a burdensome manufacturing strategy that requires donor cell extraction, modulation, expansion and re-introduction per each patient. The ability to generate universally histocompatible and genetically-enhanced NK cells from continuously renewable human induced pluripotent stem cell (hiPSC) lines offers the potential to develop a true "off-the-shelf" cellular immunotherapy. While NK differentiation from hiPSC has been demonstrated, the clonal derivation of engineered hiPSCs to improve effector function has been challenging and the scalability and robustness of the differentiation method has been limited by skewed development towards primitive hematopoiesis and the cumbersome use of embryoid bodies. Here we highlight our "off-the-shelf" NK cell therapy preclinical program by demonstrating robust and highly scalable generation of functionally mature, genetically targeted and universally histocompatible NK cells. This program utilizes our previously described naïve hiPSC platform where we uniquely create clonal lines of precisely engineered, renewable hiPSCs and drive definitive hematopoiesis in a highly scalable manner. Because hiPSC differentiation is lineage directed, minimal cellular contamination is seen, including the lack of T and B cells, in the final product. Through precise genetic engineering of naïve hiPSC lines, we have engineered HLA-class I deficient NK cells uniformly expressing a high affinity, non-cleavable version of the Fc receptor CD16 (NcHaCD16-NK). The hiPSC-derived NcHaCD16-NKs display markers of maturity, including CD16, KIR, NCRs, and CD94. When compared to conventional cord blood and peripheral blood sourced NK cells, NcHaCD16-NKs exhibit superior cytotoxicity and production of effector cytokines in response to both solid and liquid tumor cell challenge in vitro. NcHaCD16-NKs exhibit augmented cytokine response following Fc-mediated stimulation, demonstrating function competence of the engineered CD16 construct. Because surface expression of CD16 is resistant to activation-induced shedding, NcHaCD16-NKs continuously maintain enhanced ADCC while retaining the capacity for general cytotoxicity. Importantly, the hiPSC-derived hematopoietic cells can be successfully cryopreserved and banked, serving as a highly-stable cell bank for subsequent therapeutic use. Preliminary data also shows NcHaCD16-NKs elicit preferred specificity for cancer stem cells as defined by expression of ALDH1 and surface markers such as CD24. In conclusion, the outlined preclinical data demonstrate the potential therapeutic utility of NK cells developed via precision genetic engineering of a renewable, scalable hiPSC platform, and highlights the therapeutic value of NcHaCD16-NKs as an ideal ADCC-mediated "off-the-shelf" NK cell-based immunotherapeutic product with augmented persistence, anti-tumor capacity and preclinical efficacy. Disclosures Bjordahl: Fate Therapeutics, Inc: Employment. Clarke:Fate Therapeutics: Employment. Gaidarova:Fate Therapeutics: Employment. Groff:Fate Therapeutics: Employment. Rogers:Fate Therapeutics, Inc: Employment. Moreno:Fate Therapeutics, Inc.: Employment, Equity Ownership. Abujarour:Fate Therapeutics, Inc.: Employment. Bonello:Fate Therapeutics, Inc.: Employment. Lee:Fate Therapeutics: Employment. Lan:Fate Therapeutics: Employment. Burrascano:Fate Therapeutics: Employment. Bauer:Fate Therapeutics: Employment. Robinson:Fate Therapeutics: Employment. Sasaki:Fate Therapeutics, Inc.: Employment. Kim:Fate Therapeutics, Inc.: Employment. Robbins:Fate Therapeutics: Employment, Equity Ownership. Rezner:Fate Therapeutics, Inc: Employment, Equity Ownership. Abbot:Fate Therapeutics: Employment. Wolchko:Fate Therapeutics: Employment. Shoemaker:Fate Therapeutics: Employment, Equity Ownership. Valamehr:Fate Therapeutics, Inc: Employment.
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Sommer, Cesar, Hsin-Yuan Cheng, Yik Andy Yeung, Duy Nguyen, Janette Sutton, Zea Melton, Julien Valton et al. "Preclinical Evaluation of ALLO-819, an Allogeneic CAR T Cell Therapy Targeting FLT3 for the Treatment of Acute Myeloid Leukemia". Blood 134, Supplement_1 (13 novembre 2019): 3921. http://dx.doi.org/10.1182/blood-2019-129025.
Testo completoAbstract (sommario):
Autologous chimeric antigen receptor (CAR) T cells have achieved unprecedented clinical responses in patients with B-cell leukemias, lymphomas and multiple myeloma, raising interest in using CAR T cell therapies in AML. These therapies are produced using a patient's own T cells, an approach that has inherent challenges, including requiring significant time for production, complex supply chain logistics, separate GMP manufacturing for each patient, and variability in performance of patient-derived cells. Given the rapid pace of disease progression combined with limitations associated with the autologous approach and treatment-induced lymphopenia, many patients with AML may not receive treatment. Allogeneic CAR T (AlloCAR T) cell therapies, which utilize cells from healthy donors, may provide greater convenience with readily available off-the-shelf CAR T cells on-demand, reliable product consistency, and accessibility at greater scale for more patients. To create an allogeneic product, the TRAC and CD52 genes are inactivated in CAR T cells using Transcription Activator-Like Effector Nuclease (TALEN®) technology. These genetic modifications are intended to minimize the risk of graft-versus-host disease and to confer resistance to ALLO-647, an anti-CD52 antibody that can be used as part of the conditioning regimen to deplete host alloreactive immune cells potentially leading to increased persistence and efficacy of the infused allogeneic cells. We have previously described the functional screening of a library of anti-FLT3 single-chain variable fragments (scFvs) and the identification of a lead FLT3 CAR with optimal activity against AML cells and featuring an off-switch activated by rituximab. Here we characterize ALLO-819, an allogeneic FLT3 CAR T cell product, for its antitumor efficacy and expansion in orthotopic models of human AML, cytotoxicity in the presence of soluble FLT3 (sFLT3), performance compared with previously described anti-FLT3 CARs and potential for off-target binding of the scFv to normal human tissues. To produce ALLO-819, T cells derived from healthy donors were activated and transduced with a lentiviral construct for expression of the lead anti-FLT3 CAR followed by efficient knockout of TRAC and CD52. ALLO-819 manufactured from multiple donors was insensitive to ALLO-647 (100 µg/mL) in in vitro assays, suggesting that it would avoid elimination by the lymphodepletion regimen. In orthotopic models of AML (MV4-11 and EOL-1), ALLO-819 exhibited dose-dependent expansion and cytotoxic activity, with peak CAR T cell levels corresponding to maximal antitumor efficacy. Intriguingly, ALLO-819 showed earlier and more robust peak expansion in mice engrafted with MV4-11 target cells, which express lower levels of the antigen relative to EOL-1 cells (n=2 donors). To further assess the potency of ALLO-819, multiple anti-FLT3 scFvs that had been described in previous reports were cloned into lentiviral constructs that were used to generate CAR T cells following the standard protocol. In these comparative studies, the ALLO-819 CAR displayed high transduction efficiency and superior performance across different donors. Furthermore, the effector function of ALLO-819 was equivalent to that observed in FLT3 CAR T cells with normal expression of TCR and CD52, indicating no effects of TALEN® treatment on CAR T cell activity. Plasma levels of sFLT3 are frequently increased in patients with AML and correlate with tumor burden, raising the possibility that sFLT3 may act as a decoy for FLT3 CAR T cells. To rule out an inhibitory effect of sFLT3 on ALLO-819, effector and target cells were cultured overnight in the presence of increasing concentrations of recombinant sFLT3. We found that ALLO-819 retained its killing properties even in the presence of supraphysiological concentrations of sFLT3 (1 µg/mL). To investigate the potential for off-target binding of the ALLO-819 CAR to human tissues, tissue cross-reactivity studies were conducted using a recombinant protein consisting of the extracellular domain of the CAR fused to human IgG Fc. Consistent with the limited expression pattern of FLT3 and indicative of the high specificity of the lead scFv, no appreciable membrane staining was detected in any of the 36 normal tissues tested (n=3 donors). Taken together, our results support clinical development of ALLO-819 as a novel and effective CAR T cell therapy for the treatment of AML. Disclosures Sommer: Allogene Therapeutics, Inc.: Employment, Equity Ownership. Cheng:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Yeung:Pfizer Inc.: Employment, Equity Ownership. Nguyen:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Sutton:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Melton:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Valton:Cellectis, Inc.: Employment, Equity Ownership. Poulsen:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Djuretic:Pfizer, Inc.: Employment, Equity Ownership. Van Blarcom:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Chaparro-Riggers:Pfizer, Inc.: Employment, Equity Ownership. Sasu:Allogene Therapeutics, Inc.: Employment, Equity Ownership.
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Richon, Victoria M., Christopher J. Sneeringer, Margaret Porter Scott, Kevin W. Kuntz, Sarah K. Knutson, Roy M. Pollock e Robert A. Copeland. "Lymphoma-Associated Mutations of EZH2 Result In a Change-of-Function". Blood 116, n. 21 (19 novembre 2010): 707. http://dx.doi.org/10.1182/blood.v116.21.707.707.
Testo completoAbstract (sommario):
Abstract Abstract 707 EZH2, the catalytic subunit of the polycomb repressive complex 2 (PRC2), catalyzes the mono- through tri-methylation of lysine 27 on histone H3 (H3K27). While overexpression of EZH2 and increased H3K27 methylation have generally been associated with both hematologic malignancies and solid tumors, inactivating somatic mutations of Tyr641 (Y641F, Y641N, Y641S and Y641H) of EZH2 were recently reported to be associated with follicular lymphoma (FL) and the GCB subtype of diffuse large B-cell lymphoma (DLBCL) (Morin, Nat Genet 2010; 42: 181). In all cases, occurrence of the mutant EZH2 gene was heterozygous, and expression of both wild type and mutant alleles was detected in the mutant samples profiled by transcriptome sequencing. Further, the mutant forms of EZH2 could be incorporated into the multi-protein PRC2 complex, but the resulting complexes lacked the ability to catalyze trimethylation of an unmethylated H3K27 peptide substrate. To explore further the role of EZH2 in lymphomagenesis, we have evaluated the catalytic activity of the mutant EZH2 proteins in greater detail. Recombinant PRC2 complexes were prepared with wild type and Tyr641 mutant EZH2 forms. As previously reported, the wild type enzyme demonstrated robust activity but none of the mutant enzymes displayed significant methyltransferase activity on an unmodified H3K27 peptide. We next evaluated the activity of the enzymes using native avian erythrocyte olignucleosomes as the substrate in the reaction. In contrast to the peptide result, we found that the wild type and all of the mutant enzymes were active methyltransferases against the native nucleosome substrate. Since native nucleosome represents an admixture of the unmodified and mono-, di- and tri-methylated H3K27 we next evaluated the activity of the wild type and mutant enzymes on unmodified, and mono- and di-methylated H3K27 peptide. We demonstrate that the wild type enzyme displays greatest catalytic efficiency (kcat/K) for the zero to mono-methylation reaction of H3K27, and diminished efficiency for subsequent (mono- to di- and di- to tri-methylation) reactions. In stark contrast, the disease-associated Y641 mutants display very limited ability to perform the first methylation reaction, but have enhanced catalytic efficiency for the subsequent reactions, relative to WT-enzyme. Catalytic coupling between the mutant EZH2 species and PRC2 complexes containing either wild type EZH2 or wild type EZH1 are predicted to augment H3K27 trimethylation and thus produce the malignant phenotype associated with mutant heterozygosity. To test this prediction, the level of H3K27 methylation was evaluated in lymphoma cell lines harboring only wild type EZH2 (OCI-LY-19) or heterozygous for EZH2 Y641N (DB, KARPAS and SU-DHL-6) or EZH2 Y641F (WSU-DLCL2) by immunoblotting. As predicted by simulations, the level of H3K27 trimethylation was elevated in all of the lymphoma cell lines harboring the mutant EZH2 relative the wild type. Additionally, we observe decreased H3K27 dimethylation and monomethylation in the cells harboring the mutated EZH2 relative to wild type enzyme; these reductions in di- and monomethylation are likewise consistent with expectations based on steady state kinetic simulations. The present results imply that the malignant phenotype of follicular lymphoma and diffuse large B cell lymphoma of the GCB subtype, associated with expression of mutant forms of EZH2, results from of an overall gain-of-function with respect to formation of the trimethylated form of H3K27. These data suggest that selective, small molecule inhibitors of EZH2 enzymatic function may form a rational underpinning for molecularly targeted therapeutics against mutant-harboring lymphomas and other malignancies in which EZH2 gain-of-function is pathogenic. Disclosures: Richon: Epizyme, Inc: Employment. Sneeringer:Epizyme: Employment. Porter Scott:Epizyme, Inc: Employment. Kuntz:Epizyme, Inc: Employment. Knutson:Epizyme, Inc.: Employment. Pollock:Epizyme, Inc: Employment. Copeland:Epizyme, Inc: Employment.
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Huard, Justin, Laurie Tatalick, Carl Walkey e Ryan Swanson. "NL-201, a De Novo Agonist of IL-2 and IL-15 Receptors, Demonstrates Synergistic Antitumor Activity with Anti-PD-1 Checkpoint Inhibitor Therapy in a Preclinical Non-Hodgkin Lymphoma Model". Blood 138, Supplement 1 (5 novembre 2021): 4560. http://dx.doi.org/10.1182/blood-2021-148885.
Testo completoAbstract (sommario):
Abstract NL-201 is a potent, selective, and long-acting computationally designed alpha-independent agonist of the IL-2 and IL-15 receptors that is being developed as an immunotherapy for cancer. NL-201 binds to the beta and gamma subunits, selectively stimulating dose-dependent expansion and tumor infiltration of cytotoxic CD8+ T cells and natural killer (NK) cells, thereby enhancing the immune response to the tumor. Absence of binding to the IL-2 alpha subunit reduces the undesirable effects of traditional IL-2 therapies, such as vascular leak syndrome and expansion of immunosuppressive regulatory T cells. In this abstract, we demonstrate that NL-201, alone or in combination, demonstrates robust antitumor activity in preclinical models of non-Hodgkin lymphoma (NHL). We have previously demonstrated that NL-201 has marked antitumor activity in multiple syngeneic tumor models, including the A20 lymphoma model. These observations in lymphoma have been extended to explore the effects of NL-201 in combination with anti-mPD-1 checkpoint inhibitor therapy in vivo. In this model, NL-201 and anti-mPD-1 demonstrated tumor growth inhibition and increased median survival (21 days each vs 17 days as observed in control) when given alone. In combination, NL-201 and anti-mPD-1 resulted in increased antitumor activity and significant prolongation of survival (>51 days). We have also demonstrated that NL-201 does not directly induce signaling or cell death in B cell−derived NHL, suggesting that the observed antitumor activity is due to activation of non-malignant host immune cells. Additional in vitro and in vivo NHL models are being tested to enhance understanding of the interaction between NL-201 and other approved therapies within the hematopoietic tumor microenvironment. These data will be used to design future clinical trials of NL-201 in novel regimens to treat hematological malignancies. Disclosures Huard: Neoleukin Therapeutics, Inc.: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company, Patents & Royalties. Tatalick: Neoleukin Therapeutics, Inc.: Consultancy, Current equity holder in publicly-traded company, Other: Independent paid nonclinical consultant for Neoleukin. Walkey: Neoleukin Therapeutics, Inc.: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Swanson: Neoleukin Therapeutics, Inc.: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties.
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Gyurkocza, Boglarka, Rajneesh Nath, Patrick J. Stiff, Edward Agura, Mark Litzow, Ben Tomlinson, Hannah K. Choe et al. "Re-Induction and Targeted Conditioning with Anti-CD45 Iodine (131I) Apamistamab [Iomab-B] Leads to High Rates of Transplantation and Successful Engraftment in Older Patients with Active, Relapsed or Refractory (rel/ref) AML after Failure of Chemotherapy and Targeted Agents: Preliminary Midpoint Results from the Prospective, Randomized Phase 3 Sierra Trial". Blood 134, Supplement_1 (13 novembre 2019): 5642. http://dx.doi.org/10.1182/blood-2019-122776.
Testo completoAbstract (sommario):
Background: Patients ≥55 years of age with rel/ref AML who have failed to achieve complete remission (CR) after standard induction & salvage therapies do not routinely undergo allogeneic hematopoietic cell transplantation (HCT) due to inability to receive myeloablative conditioning and lack of efficacy. With the advent of recently approved targeted therapies (e.g. venetoclax, IDH inhibitors) more patients achieve CR, however durable responses in heavily pre-treated patient populations remain unsatisfactory with very few patients being transplanted. The SIERRA (Study of Iomab-B in Elderly Relapsed or Refractory AML) trial is a prospective, randomized, phase 3 trial designed to address this unmet need. We present the preliminary analysis of the first 50% of patients and analyze the proportion of patients who fail standard chemo and targeted therapies that can then successfully undergo HCT with Iomab-B-based conditioning. Study continues to enroll (N=150), primary end-point durable CR of ≥180 days. Methods: Eligible patients are ≥ 55 years with rel/ref AML (≥5% blasts), adequate organ function, and related/unrelated 8/8 HLA-matched donors. Patients are randomized (1:1) to receive Iomab-B followed 12-14 days later by reduced intensity conditioning (fludarabine(FLU)/TBI) and HCT, or to conventional care (CC). CC patients may receive investigator's choice of salvage therapy, including newly approved targeted agents such as venetoclax, IDH Inhibitors and may proceed to standard HCT if they achieve CR. If patients do not achieve CR, the study allows cross-over (CO) to Iomab-B with HCT. Results: Preliminary data are available from the first 75 patients enrolled on the SIERRA trial. Median age of 64 years (range: 55-77). All patients had active measurable disease (Table 1). Patients in this trial were heavily pretreated with 85% failing ≥2 induction therapies and 33% failing targeted therapies. Molecular and cytogenetics was 68% adverse risk. (NCCN, v1.2018) Following therapeutic dose of Iomab-B as a single agent, there was significant reduction of circulating blasts by day 3 (99%, p<0.0001), and peripheral blasts became undetectable by day 8 prior to FLU. All patients transplanted on the Iomab-B arm with data available showed median days to engraftment of ANC at 14 (range: 9-22) and platelets at 16 (range: 4-30) (Table 1). After randomization, 82% (31/38) patients in the CC arm failed salvage therapy and did not proceed to standard HCT. Significantly, 32% (12/38) of these patients received targeted therapy. 73% (8/11) of the patients receiving venetoclax with HMA or cytarabine did not adequately respond and did not proceed to standard HCT, 45% (5/11) crossed over to Iomab-B and received HCT. Following standard chemotherapies, 85% (22/26) were evaluated for CO. In total, 63% (19/30) received Iomab-B and HCT. The most common reason preventing crossover was disease progression. All crossover (Iomab-B) patients showed similar engraftment (Table 1). CC patients showed increased incidence of febrile neutropenia with grade 3 prior to CO evaluation or HCT, compared to Iomab-B patients prior to HCT (CC: 34%, Iomab-B: 8%). Iomab-B administration was generally well-tolerated with no grade 4 and 1 grade 3 infusion reaction. There were no Iomab-B-related deaths or 100 day non-relapse transplant related mortality deaths in patients on the Iomab-B arm (Table 1). Conclusion: 100% of the patients receiving HCT on the Iomab-B arm engrafted, despite a median of 29% BM blasts pre-Iomab-B. 82% of the patients on the CC arm failed to be eligible for standard of care transplant, including a high percentage of those receiving targeted therapies. On the CC arm, 63% of eligible patients crossed over to Iomab-B/HCT with robust engraftment despite a median of 35% BM blasts pre-Iomab-B. The incidence of grade 3 febrile neutropenia was higher in patients treated on the conventional care arm with salvage therapy vs. those treated on the Iomab-B arm prior to HCT. In conclusion, despite advanced age, active disease and heavily pre-treated patients, 68% (50/74) of all patients enrolled in the SIERRA trial were able to undergo HCT after reinduction and targeted conditioning with Iomab-B. These results show significant improvements in the current rates of transplantation in this patient population. This SIERRA trial is currently enrolling. For full study details see www.sierratrial.com or clinicaltrials.gov (NCT02665065). Disclosures Gyurkocza: Actinium Pharmaceuticals: Research Funding. Nath:Astellas: Consultancy; Actinium: Consultancy; Daiichi Sankyo: Consultancy. Stiff:Amgen: Research Funding; Gamida-Cell: Research Funding; Incyte: Research Funding; Unum: Research Funding; Cellectar: Research Funding; Gilead/Kite Pharma: Consultancy, Honoraria, Research Funding. Tomlinson:Actinium Pharmaceuticals, Inc: Other: Travel Support. Abhyankar:Incyte: Speakers Bureau; Therakos: Other: Consulting, Speakers Bureau. Hari:AbbVie: Consultancy, Honoraria; Cell Vault: Equity Ownership; Sanofi: Honoraria, Research Funding; Spectrum: Consultancy, Research Funding; Amgen: Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Research Funding; Janssen: Consultancy, Honoraria; Kite: Consultancy, Honoraria. Al-Kadhimi:Seattle Genetics: Other: Stocks; Celldex Biotech: Other: Stocks. Foran:Agios: Honoraria, Research Funding. Orozco:Actinium Pharmaceuticals: Research Funding. Van Besien:Miltenyi Biotec: Research Funding. Sabloff:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Astellas Pharma Canada: Honoraria, Membership on an entity's Board of Directors or advisory committees; ASTX: Membership on an entity's Board of Directors or advisory committees, Research Funding; Jazz Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees; Actinium Pharmaceuticals, Inc: Membership on an entity's Board of Directors or advisory committees; Novartis Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pfizer Canada: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sanofi Canada: Research Funding. Kebriaei:Pfizer: Honoraria; Kite: Honoraria; Jazz: Consultancy; Amgen: Research Funding; Pfizer: Honoraria; Kite: Honoraria; Jazz: Consultancy; Amgen: Research Funding. Levy:Takeda (Millennium Pharmaceuticals): Consultancy. Lazarus:Adaptive Biotechnologies: Consultancy, Honoraria; Seattle Genetics: Consultancy, Honoraria, Speakers Bureau; Actinium Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pharmacyclics: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Teva Pharmaceuticals: Speakers Bureau; Biosight: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Consultancy, Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; AstraZeneca: Honoraria, Speakers Bureau; Bristol Myers Squibb: Honoraria, Speakers Bureau; Pluristem Therapeutics, Inc: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; CSL Behring: Consultancy; Genentech: Speakers Bureau. Giralt:Novartis: Consultancy; Actinium: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Jazz Pharmaceuticals: Consultancy; Amgen: Consultancy, Research Funding; Kite: Consultancy; Miltenyi: Research Funding; Takeda: Consultancy, Research Funding; Johnson & Johnson: Consultancy, Research Funding; Spectrum Pharmaceuticals: Consultancy. Berger:Actinium Pharmaceuticals, Inc: Employment, Equity Ownership. Reddy:Actinium Pharmaceuticals: Employment. Pagel:Pharmacyclics: Consultancy; AstraZeneca: Consultancy; Gilead Sciences: Consultancy.
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Storms, Robert W., Congxiao Liu, Elisabeth T. Tracy, Tracy Gentry, Andrew Balber e Joanne Kurtzberg. "ALDHbr Hematopoietic Progenitors Promote Short-Term Engraftment in Experimental Models for Cord Blood Transplantation." Blood 114, n. 22 (20 novembre 2009): 2433. http://dx.doi.org/10.1182/blood.v114.22.2433.2433.
Testo completoAbstract (sommario):
Abstract Abstract 2433 Poster Board II-410 We previously developed robust methods to purify human hematopoietic progenitors based on their high expression of aldehyde dehydrogenase (ALDHbr cells). These cells are enriched with both short- and long-term NOD/SCID repopulating cells. One early clinical trial suggested that augmenting a conventional cord blood transplant (CBT) with ALDHbr cells accelerated both neutrophil and platelet engraftment. We now describe experimental models for these clinical transplants, performed in two immunologically disparate strains of NOD/SCID mice. In the clinical study, pediatric patients first received 80% of an unmanipulated cord blood (CB) graft. 4 hours later they received the ALDHbr cells purified from the remaining 20% of the graft. In the experimental model, the mice were divided among three cohorts. Some mice were transplanted with 4,000 purified ALDHbr cells, alone. Other mice were transplanted with total CB using a cell dose that contained 4,000 ALDHbr cells. In the final group, mice first received the same dose of unmanipulated CB and, after 4 hours, 4,000 purified ALDHbr cells were also administered. After 4 weeks, the mice were sacrificed to determine their levels of human hematopoietic chimerism. When these transplants were performed using NOD/SCID-IL2Rγnull (NSγ) mice, the ALDHbr cells demonstrated strong short-term engraftment to the bone marrow (12 ± 4.9%; n = 5) that was characterized by human CD19+ B cells and CD33+ myeloid cells. The latter included CD15+ cells that indicate neutrophil engraftment. In addition, the peripheral blood of these mice contained low levels of human CD41+ CD61+ platelets. Unmanipulated CB also engrafted the bone marrow of NSγ mice (7.4 ± 4.7%; n = 8); however, >95% of the human cells appeared to be mature CD3+ T cells. Engraftment by either B cells or myeloid cells was consistently low to undetectable. Similarly, human platelets were not detected in the peripheral blood. When NSγ mice were transplanted first with total CB and subsequently with purified ALDHbr cells, the level of engraftment to the bone marrow increased >2-fold over what had been observed in mice transplanted with CB alone (18.9 ± 9.3%; n = 10; P = 0.006). However, nearly all of the human cells present within the bone marrow were T cells, as had been observed in animals that received only unmanipulated CB. In addition, human platelets were not observed in the peripheral blood of NSγ mice that had received both CB and ALDHbr cells. When similar transplants were performed using NOD/SCID-β2-microglobulinnull (NSβ) mice, the ALDHbr cells again demonstrated strong short-term engraftment to the bone marrow (5.6 ± 4.7%; n = 5) that was characterized by human CD19+ B cells and CD33+ myeloid cells. The peripheral blood of these mice also contained low levels of human platelets. In contrast, total CB achieved only very low engraftment to the bone marrow of NSβ mice (0.32 ± 0.19%; n = 4) and human platelets were not detected in the peripheral blood. However, when NSβ mice first received total CB which was augmented 4 hours later with purified ALDHbr progenitors, the level of human hematopoietic chimerism in the bone marrow increased >10-fold (4.3 ± 1.9%; n = 5), with engraftment of human B cells and myeloid cells. The co-transplanted NSβ mice also demonstrated low-level engraftment of human platelets in the peripheral blood. Continuing studies will resolve the relative contributions of the ALDHbr cells and the unmanipulated CB by using two HLA-matched (6/6), but sex-mismatched, CBs. In total, these studies confirmed that ALDHbr progenitors by themselves provided efficient short-term myeloid engraftment in both NOD/SCID strains. This was in contrast to what was observed after transplantation of bulk CB, which by itself did not provide efficient myeloid engraftment in either strain. Finally, in both mouse strains, ALDHbr cells altered the outcome of CB transplants. In NSγ mice, ALDHbr progenitors appeared to facilitate either the engraftment or proliferation of mature CB T cells. Most importantly, the studies in NSβ mice strongly suggested that ALDHbr progenitors directly augment short-term myeloid and platelet engraftment by total CB. The latter studies in particular mirror the experience of early clinical CBT studies that use the same strategy. Disclosures: Storms: Aldagen, Inc: Equity Ownership, Patents & Royalties. Gentry:Aldagen, Inc: Employment. Balber:Aldagen, Inc: Employment, Equity Ownership. Kurtzberg:Aldagen, Inc: Research Funding.
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Rosen, David B., James A. Cordeiro, David M. Soper, Ying-Wen Huang, Donna E. Hogge, Roland B. Walter, Alessandra Cesano e Wendy J. Fantl. "Distinct Signaling Profiles of Gemtuzumab Ozogamicin Responsiveness and Refractoriness in Acute Myeloid Leukemia." Blood 114, n. 22 (20 novembre 2009): 2745. http://dx.doi.org/10.1182/blood.v114.22.2745.2745.
Testo completoAbstract (sommario):
Abstract Abstract 2745 Poster Board II-721 Background: Gemtuzumab Ozogamicin (GO, Mylotarg), a humanized CD33 monoclonal antibody linked to calicheamicin was approved by the US FDA for use as a monotherapy in patients older than 60 years with relapsed acute myeloid leukemia (AML) unfit to tolerate standard salvage therapy. GO is internalized rapidly after infusion, and calicheamicin, a potent enediyene, is subsequently released and acts as a cytotoxic agent by causing double strand DNA breaks. Currently GO is in multiple clinical trials as a single agent or in combination with other therapies for both induction and consolidation treatment of various clinical subgroups of AML. However, the mechanisms of action and resistance of GO are incompletely understood and it is unclear which patient subgroups benefit from GO-based therapy. Single cell network profiling (SCNP) has shown promise as a methodology wherein multiple signaling networks are measured after treatment with an exogenous modulator such as a growth factor, cytokine or therapeutic agent and the identified signaling profiles can be used as clinical and therapeutic enablement tools. Objectives: SCNP using multiparameter flow cytometry was used to identify intracellular pathways that were associated with responsiveness or refractoriness to in vitro GO exposure in both cancer cell lines and primary AML samples. Methods: Signaling pathways emphasizing DNA damage response, cell cycle, apoptosis and drug transporter activity were measured by SCNP after in vitro exposure of cell lines and AML primary samples to clinically relevant concentrations of GO. Samples were processed for cytometry by paraformaldehyde /methanol fixation and permeabilzation followed by incubation with fluorochrome-conjugated antibody cocktails that recognize cell surface proteins to delineate cell subsets and intracellular signaling molecules. Results: In cell lines, responsiveness to in vitro GO exposure was defined as a) induction of DNA Damage as measured by increased p-ATM, p-Chk2 and p-H2AX, b) cell cycle arrest at G2/M as measured by increased cyclin B1 and DNA content & c) induction of apoptosis as measured by cleaved PARP and viability dyes. Of note, inhibition of drug transporter activity in 2 MDR-1+ cell lines did not restore GO responsiveness, suggesting the presence of additional relevant resistance mechanisms in these cell lines. In primary AML diagnostic samples, DNA damage and apoptosis pathway readouts were able to identify responsiveness or refractoriness to GO exposure. In the GO responsive profile, induction of both DNA damage responses and apoptosis were seen. Within the refractory samples, two distinct profiles were observed: a) robust and early induction of DNA damage response without apoptosis and 2) delayed and attenuated DNA damage response without apoptosis. Conclusions: Characterization of intracellular Cell Cycle, DNA Damage, and Apoptosis networks in single cells after GO exposure distinguishes GO responsive from refractory AML cells. Further, these pathway signatures provide information about mechanisms of refractoriness. (e.g. a block between a successful DNA damage response and initiation of apoptosis versus a block in the initial induction of DNA damage after GO exposure). The ability of the same profiles to predict clinical responses to the drug will be tested in future studies. Disclosures: Rosen: Nodality, Inc.: Employment, Equity Ownership. Cordeiro:Nodality Inc.: Employment, Equity Ownership. Soper:Nodality Inc.: Employment, Equity Ownership. Huang:Nodality Inc.: Employment, Equity Ownership. Cesano:Nodality Inc.: Employment, Equity Ownership. Fantl:Nodality Inc.: Employment, Equity Ownership.
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Howell, Stephen B., Hongying Zhang e William G. Rice. "A Phase 1a/b Dose Escalation Study of Apto-253 in Patients with Relapsed or Refractory AML or High-Risk MDS". Blood 134, Supplement_1 (13 novembre 2019): 5148. http://dx.doi.org/10.1182/blood-2019-129685.
Testo completoAbstract (sommario):
INTRODUCTION: APTO-253 has captured attention because of its distinguishing mechanism of action as the only known a small molecule in clinical development to target a conserved structure in the promoter of the MYC oncogene and interrupt MYC gene expression. In preclinical studies of acute myeloid leukemia (AML) cell lines, APTO-253 was discovered to potently down-regulate MYC gene expression, reduce MYC mRNA and protein level, induce apoptosis, and deplete cells of the MYC oncoprotein. APTO-253 demonstrated broad killing of primary mononuclear cells isolated from the bone marrow of patients with AML, MDS or MPN and enhanced AML cell killing when combined with BET bromodomain inhibitors or FLT3 inhibitors (Kurtz, Blood 2017 126:1358). Because dysregulated MYC is considered a major oncogenic operator in AML and myelodysplasias and because APTO-253 has such a distinct mechanism to kill such cells, APTO-253 has been granted orphan drug designation for the treatment of AML by the US FDA and is currently in a Phase 1a/b clinical trial in patients with relapsed or refractory AML (R/R AML) or high-risk myelodysplasias (high-risk MDS). METHODS: APTO-253 is being evaluate in an ongoing clinical trial, entitled "A Phase I a/b Dose Escalation and Expansion, Multicenter, Open-label, Safety, Pharmacokinetic and Pharmacodynamic Study of APTO-253 in Patients with Relapsed or Refractory Acute Myelogenous Leukemia or High-Risk Myelodysplasia (NCT02267863)". The protocol provides for patients to be dosed once weekly on days 1, 8, 15, and 22 of each 28-day cycle with the intended dose levels of 20, 40, 66, 100, 140, 180 or 220 mg/m2. Eligible patients must be ≥18 years old, have a life expectancy of at least 2 months, and have acceptable hematologic, renal and liver functions and coagulation status parameters. Primary objectives are to determine the safety and tolerability of APTO-253, to determine the maximum tolerated dose (MTD) and the dose limiting toxicities (DLT), and to establish the recommended Phase 2 dose for patients with specific types of hematologic malignancies. Key secondary objectives are to seek evidence of antitumor activity by hematologic and bone marrow evaluations and to assess the impact of APTO-253 on the expression of pharmacodynamic biomarkers (including MYC and p21 gene expression). RESULTS: To date, R/R AML and high-risk MDS patients have been enrolled and treated at dose levels of 20, 40 and 66 mg/m2 APTO-253. As specified by the protocol, only one patient was required at each of the 20 and 40 mg/m2 doses. The AML patient on the lowest dose level completed the 28-day cycle and demonstrated approximately a 72% reduction in MYC expression levels in peripheral white blood cells during the cycle. The high-risk MDS patient who received 40 mg/m2 demonstrated a 79% reduction in MYC expression during the first cycle. To date (as of July 2019), no drug related TEAEs, SAEs or dose limiting toxicities have been reported. Two patients are receiving treatment with 66 mg/m2, and samples are being analyzed for a series of pharmacodynamic markers. CONCLUSIONS: APTO-253 is a potent molecule with a particularly interesting and novel mechanism of action that results in robust suppression of MYC expression. The Phase 1 a/b trial has successfully enrolled R/R-AML and HR-MDS patients into the first three cohorts at 20, 40 and 66 mg/m2. Clinical data to date suggest APTO-253 is generally well tolerated at the doses tested, and target engagement has been evidenced by the reduction in cellular MYC gene expression in whole blood samples from R/R AML and high-risk MDS patients. Recruitment and enrollment are continuing, and updated safety, PK and biomarker data will be presented at the meeting. Disclosures Howell: Aptose Biosciences, Inc: Consultancy, Research Funding. Zhang:Aptose Biosciences, Inc: Employment. Rice:Aptose Biosciences, Inc: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees.
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Steen, Chloe B., Bogdan A. Luca, Mohammad Shahrokh Esfahani, Barzin Y. Nabet, Brian Sworder, Farshad Farshidfar, Kiarash Shamardani et al. "An Atlas of Clinically-Distinct Tumor Cellular Ecosystems in Diffuse Large B Cell Lymphoma". Blood 134, Supplement_1 (13 novembre 2019): 655. http://dx.doi.org/10.1182/blood-2019-129461.
Testo completoAbstract (sommario):
Background: Diffuse large B cell lymphoma (DLBCL) exhibits significant clinical and biological heterogeneity, in part due to cell-of-origin subtypes, somatic alterations, and diverse stromal constituents within the tumor microenvironment (TME). Several immunologically-active lymphoma therapies are known to rely on innate and adaptive anti-tumor responses occurring within this dynamic TME, including agents that are approved (e.g., rituximab, lenalidomide, CART19, ibrutinib) or emerging (e.g., anti-CD47, checkpoint inhibitors). We hypothesized that a large-scale characterization of the cellular heterogeneity in DLBCL might reveal previously unknown biological variation in the TME linked to tumor subtypes and genotypes, therapeutic responses and clinical outcomes, with implications for future personalization of immunotherapy. Methods: Using a combination of lymphoma single-cell RNA sequencing (scRNA-seq) and bulk tumor transcriptome deconvolution (CIBERSORTx; Newman et al., Nat Biotech, 2019), we developed a new machine learning framework for identifying cellular states and ecosystems that reflect fundamental TME subtypes and distinctions in tumor biology (Fig. 1). Specifically, using CIBERSORTx, we purified the transcriptomes of B cells and 12 different TME cell types, including immune and stromal subsets, from 1,279 DLBCL tumor biopsies profiled in 3 prior studies (Reddy et al., Cell 2017; Schmitz et al., NEJM 2018; Chapuy et al., Nat Med 2018). Then, we defined distinct transcriptional states for each of the 13 cell types, which we validated at single-cell resolution, using a combination of two scRNA-seq techniques (Smart-Seq2 and 10x Chromium 5' GEP, BCR and TCR) to profile primary DLBCL, FL, and human tonsils, as well as leveraging multiple scRNA-seq datasets from previous studies. We identified robust co-associations between cell states that form tumor cellular ecosystems, which we validated in independent datasets of bulk DLBCL tumor gene expression profiles. Finally, we related TME ecosystems to defined tumor subtypes, including genotype classes, and to clinical outcomes. Results: By systematically characterizing the landscape of cellular heterogeneity in nearly 1,300 DLBCL tumors, we defined an atlas of 49 distinct transcriptional states across 13 major cell types. These novel cell states spanned diverse innate and adaptive immune effector cells of the lymphoid and myeloid lineages, as well as tumor-associated fibroblasts. Remarkably, 94% of these states (46 of 49) could be validated in a compendium of ~200,000 single-cell transcriptomes derived from lymphomas, healthy control tonsils, and other tissue types. Moreover, single cells from DLBCL, FL and tonsils best mirrored these newly discovered cell states. We next characterized the biology and potential clinical utility of each cell state. We observed clear distinctions in the transcriptional programs of immune and stromal elements between germinal center and activated B cell DLBCL, as well as between known mutational subtypes. Importantly, many cell states reflected novel phenotypic groupings, and the majority were significantly associated with overall survival (P<0.05). These findings were highly concordant both within and across 3 independent DLBCL cohorts. By identifying groups of DLBCL patients with similar communities of cellular states, we defined cohesive cellular ecosystems that collectively capture the landscape of transcriptional heterogeneity in DLBCL tumors. Patients whose tumors were assigned to these ecosystems exhibited striking variation in overall survival. Importantly, the ecosystems defined distinct subgroups that could not be fully recapitulated by known transcriptional and genetic subtypes. Moreover, several TME classes showed significant enrichments in canonical or novel tumor genotypes, suggesting an evolutionary interplay between the tumor and host microenvironment. Conclusion: We describe a novel computational framework to digitally dissect the DLBCL TME and an atlas of novel states for diverse cell types in these tumors. We show how cellular states form cohesive tumor ecosystems, which exhibit distinct clinical outcomes and novel somatic alterations. These results expand our understanding of cellular heterogeneity in DLBCL, with implications for the development of individualized immunotherapies. Disclosures Kurtz: Roche: Consultancy. Advani:Kura: Research Funding; Merck: Research Funding; Millennium: Research Funding; Pharmacyclics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Regeneron: Research Funding; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; Cell Medica, Ltd: Consultancy; Kyowa Kirin Pharmaceutical Developments, Inc.: Consultancy; Stanford University: Employment, Equity Ownership; Janssen: Research Funding; AstraZeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees; Seattle Genetics: Consultancy, Research Funding; Infinity Pharma: Research Funding; Bayer: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Research Funding; Celmed: Consultancy, Membership on an entity's Board of Directors or advisory committees; Forty-Seven: Research Funding; Roche/Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead Sciences, Inc./Kite Pharma, Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees; Autolus: Consultancy, Membership on an entity's Board of Directors or advisory committees; Agensys: Research Funding. Diehn:Roche: Consultancy; AstraZeneca: Consultancy; Novartis: Consultancy; BioNTech: Consultancy; Quanticell: Consultancy. Alizadeh:Janssen: Consultancy; Genentech: Consultancy; Pharmacyclics: Consultancy; Chugai: Consultancy; Celgene: Consultancy; Gilead: Consultancy; Roche: Consultancy; Pfizer: Research Funding.
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Stapleton, Ann, Aurelio Silvestroni, Pacita Roberts, Marsha Cox, Hillary Hayden, Mitchell Brittnacher, Samuel Miller e Thomas Hooton. "126. Robust and Persistent Vaginal Colonization with LACTIN-V Vaginal Lactobacillus crispatus Probiotic in a Double-Blind, Placebo-Controlled (DBPC) Phase 2b Trial to Prevent Recurrent UTI (rUTI)". Open Forum Infectious Diseases 5, suppl_1 (novembre 2018): S8. http://dx.doi.org/10.1093/ofid/ofy209.016.
Testo completoAbstract (sommario):
Abstract Background We investigated vaginal colonization using repetitive sequence PCR (repPCR) and 16S rRNA sequencing in a Phase 2b DBPC trial of a L. crispatus intravaginal suppository probiotic for prevention of rUTI in premenopausal women. Methods Twenty-four young women with a history of rUTI and current culture-confirmed symptomatic UTI were enrolled and treated (Visit 0), then randomized (Visit 1) to receive an intravaginal suppository containing L. crispatus CTV-05 (LACTIN-V®, Osel, Inc.) or placebo daily for 5 days, then once weekly for 2 months. Participants were followed up during the 2-month probiotic/placebo intervention (Visits 2 to 4; active intervention) and during 2 months following the intervention (Visits 5 and 6; post-intervention). At each visit, vaginal swabs were collected for repPCR to determine the presence or absence of the probiotic strain and the duration of its presence in the vagina and for 16S rRNA-based sequence analysis to determine relative abundance of any L. crispatus. Results LACTIN-V vaginal suppository induced selective and sustained colonization in the probiotic but not the placebo recipients, as follows. Pre-intervention: Probiotic lactobacillus strain, not found in vaginal specimens obtained from participants in either arm of study. Active intervention: (1) Probiotic lactobacillus strain, (a) Probiotic arm: 100% of participants positive at one or more visits and (b) Placebo arm: 0% of participants positive at any time. (2) L. crispatus relative abundance, (a) Probiotic arm: above 90%, all specimens, all visits and (b) Placebo arm: below 15%, all specimens, all visits. Post-intervention: (1) Probiotic lactobacillus strain, (a) Probiotic arm: 75% of participants positive at Visit 5, 58% at Visit 6 and (b) Placebo arm: 0% of participants positive at Visits 5 and 6. (2) L. crispatus relative abundance, (a) Probiotic arm: 70% to 100% and (b) Placebo arm: below 15%. Conclusion LACTIN-V L. crispatus vaginal probiotic achieved robust and persistent colonization throughout 2 months of weekly dosing and for 2 months after the last dose in most participants. Disclosures All authors: No reported disclosures.
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Smolen, J. S., F. Behrens, S. Liu Leage, C. Sapin, I. De La Torre, G. Meszaros, G. Schett et al. "AB0841 TARGET OUTCOMES IN PsA: SIMULTANEOUS ACHIEVEMENT of ACR50-PASI100 AND BEYOND: INSIGHTS FROM SPIRIT-H2H AT WEEK 24". Annals of the Rheumatic Diseases 79, Suppl 1 (giugno 2020): 1726.2–1727. http://dx.doi.org/10.1136/annrheumdis-2020-eular.2884.
Testo completoAbstract (sommario):
Background:Psoriatic Arthritis (PsA) treatment should aim to achieve robust improvement of arthritis as well as control of extra-articular manifestations like the skin. SPIRIT-H2H evaluated the efficacy of ixekizumab (IXE) and adalimumab (ADA) in patients with active PsA and psoriasis, and naïve to biologic Disease-Modifying Anti-Rheumatic Drugs (bDMARDs). At week 24 (W24), IXE showed superiority to ADA in simultaneous achievement of ACR50 and PASI100 as well as significant improvement of treat-to-target and other extra-articular outcomes.Objectives:To examine and to compare PsA efficacy outcomes in patients beyond achievement of the primary endpoint of the SPIRIT-H2H trial at W24, irrespective of treatment allocation.Methods:All patients recruited had active PsA (defined as tender joint count ≥3/68, swollen joint count ≥3/66 and body surface area [BSA] ≥3%), and inadequate response to conventional synthetic (cs)-DMARDs. Patients were randomised 1:1 to open-label, assessor-blinded IXE or ADA. We conducted post-hoc analysis of SPIRIT-H2H (NCT03151551), categorizing patients into four independent groups based on the achievement of the primary outcome (ACR50 & PASI100), ACR50 only, PASI100 only or none of them after 24 weeks of treatment. Statistical analyses consisted of mixed model for repeated measurement and logistic regression models using non-response imputation.Results:At week 24, patients reaching simultaneously ACR50 and PASI100 had a statistically significant higher response in most treat-to-target endpoints than those meeting ACR50 only (p<0.05). In this latter group, a high response rate was observed in ACR70, MDA, DAPSA remission and PASI90 response (48.9%, 60.6%, 35.1%, 36.2%, respectively). In patients that did not achieve either ACR50 or PASI100, up to 1/3 of the patients did achieve ACR20, DAPSA score ≤14, or no physical impairment.Table.Efficacy Endpoints at W24ACR50 & PASI100ACR50 onlyPASI100 onlyNeither ACR50 nor PASI100n=181n=94n=121n=170ACR20100.0b,c100.053.734.7ACR7064.6a,b,c48.90.00.0MDA75.7a,b,c60.623.112.4VLDA32.6a,b,c13.83.31.8DAPSA LDA or Remission (≤14)92.3a,b,c81.943.028.8DAPSA Remission (≤4)44.8b,c35.16.62.4HAQ-DI score ≤0.575.7b,c64.930.627.4PASI75100.0a,c60.6100.037.1PASI90100.0a,c36.2100.014.7SF-36 PCS change from baseline§12.3±0.53b,c12.3±0.745.4±0.664.0±0.55Data are presented as %;§mean±standard error.ap<0.05 vs. ACR50 only;bp<0.05 vs. PASI100only;cp<0.05 vs. Neither ACR50 nor PASI100.ACR, American College of Rheumatology; DAPSA, Disease Activity in Psoriatic Arthritis; HAQ-DI, Health Assessment Questionnaire Disability Index; LDA, Low Disease Activity; MDA, Minimal Disease Activity; PASI, Psoriasis Area Severity Index; VLDA, Very Low Disease Activity.Nine patients with active PsO and BSA≥3% were assessed as PASI=0 at baseline, a medical inconsistency that was resolved using medical judgement. These patients were considered PASI100 responders if PASI=0 and BSA=0 at post baseline visits.Conclusion:Reflecting the complexity of PsA, different degrees of improvement were observed across all treat-to-target outcomes with greater improvements in patients that met ACR50 response regardless of skin resolution. These findings at week 24 need to be confirmed with a longer duration of treatment.Disclosure of Interests:Josef S. Smolen Grant/research support from: AbbVie, AstraZeneca, Celgene, Celltrion, Chugai, Eli Lilly, Gilead, ILTOO, Janssen, Novartis-Sandoz, Pfizer Inc, Samsung, Sanofi, Consultant of: AbbVie, AstraZeneca, Celgene, Celltrion, Chugai, Eli Lilly, Gilead, ILTOO, Janssen, Novartis-Sandoz, Pfizer Inc, Samsung, Sanofi, Frank Behrens Grant/research support from: Pfizer, Janssen, Chugai, Celgene, Lilly and Roche, Consultant of: Pfizer, AbbVie, Sanofi, Lilly, Novartis, Genzyme, Boehringer, Janssen, MSD, Celgene, Roche and Chugai, Soyi Liu Leage Shareholder of: Eli Lilly and Company, Employee of: Eli Lilly and Company, Christophe Sapin Shareholder of: Eli Lilly and Company, Employee of: Eli Lilly and Company, Inmaculada De La Torre Shareholder of: Eli Lilly and Company, Employee of: Eli Lilly and Company, Gabriella Meszaros Shareholder of: Eli Lilly and Company, Employee of: Eli Lilly and Company, Georg Schett Speakers bureau: AbbVie, BMS, Celgene, Janssen, Eli Lilly, Novartis, Roche and UCB, Laure Gossec Grant/research support from: Lilly, Mylan, Pfizer, Sandoz, Consultant of: AbbVie, Amgen, Biogen, Celgene, Janssen, Lilly, Novartis, Pfizer, Sandoz, Sanofi-Aventis, UCB, Andrew Ostor Consultant of: MSD, Pfizer, Lilly, Abbvie, Novartis, Roche, Gilead and BMS, Speakers bureau: MSD, Pfizer, Lilly, Abbvie, Novartis, Roche, Gilead and BMS, Bernard Combe Grant/research support from: Novartis, Pfizer, Roche-Chugai, Consultant of: AbbVie; Gilead Sciences, Inc.; Janssen; Eli Lilly and Company; Pfizer; Roche-Chugai; Sanofi, Speakers bureau: Bristol-Myers Squibb; Gilead Sciences, Inc.; Eli Lilly and Company; Merck Sharp & Dohme; Pfizer; Roche-Chugai; UCB, Filip van den Bosch Consultant of: AbbVie, Celgene Corporation, Eli Lilly, Galapagos, Janssen, Novartis, Pfizer, and UCB, Speakers bureau: AbbVie, Celgene Corporation, Eli Lilly, Galapagos, Janssen, Novartis, Pfizer, and UCB
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Fang, Riguo, Yingchi Zhang, Pengfei Yuan, Huihui Yang, Lingling Yu, Yongjian Zhang, Jia Shi et al. "Highly Efficient Gene Editing of Human Hematopoietic Stem Cells to Treat X-Linked Sideroblastic Anemia". Blood 136, Supplement 1 (5 novembre 2020): 1. http://dx.doi.org/10.1182/blood-2020-140208.
Testo completoAbstract (sommario):
X-linked sideroblastic anemia (XLSA) is an anemic disease caused by mutations in the gene encoding 5-aminolevulinate synthase 2 (ALAS2), which catalyzes the rate-limiting step of heme biosynthesis. With current interventions including pyridoxine treatment and allogeneic hematopoietic stem cell transplantation, severe unmet needs remain for patients with XLSA. Here, we used CRISPR/Cas9 technology to efficiently and functionally correct the pathogenic mutation in intron 1 of ALAS2 in CD34+ hematopoietic stem cell and progenitor cells (HSPCs) from two patients. Intriguingly, we found that the gene-editing efficiency of CD34+HSPCs from the elder patient was much lower than that from the younger patient, consistent with the poor hematopoiesis of older XLSA patients observed in clinical practice. Furthermore, we performed single cell RNA-sequencing (scRNA-seq) to investigate the causes of those phenomenon in-depth. Previously we and others identified the A>G mutation in the GATA1 binding region of intron 1 of ALAS2 in particular XLSA families and have demonstrated the key role of this site in regulating ALAS expression. Thus, we first designed a series of sgRNAs, along with single-stranded DNA oligonucleotide donors (ssODN). After co-electroporating with Cas9 mRNA into patient-derived hiPSCs, sgRNA-1 and ssODN were selected for further experiments based on optimal correction rate (43.93±3.43% via HDR). Next, using a well-established erythroid protocol, CD34+ HSPCs of control and gene-edited groups were differentiated into erythroid cells in vitro. Surprisingly, heme biosynthesis examined by benzidine staining showed that compared with the mock cells, the gene-corrected group significantly increased the frequency of benzidine-positive cells. To examine the multilineage differentiation potential of gene-corrected CD34+ HSCPs, we performed colony-forming unit (CFU) assays to quantify various types of colonies. Compared with mock cells, gene-edited group significantly enhanced the generation of total, CFU-GM and BFU-E colonies, suggesting higher clonogenic potential. Next, gene-corrected CD34+ HSPCs were transplanted into nonobese diabetic (NOD)/Prkdcscid/IL-2Rγnull (NPG) mice to evaluate the repopulating potential. All transplanted mice displayed engraftment in multiple organs at 10-16 weeks post transplantation, and the gene-corrected cells showed greater engraftment potential than mock group. In addition, hematopoietic reconstitution analysis indicated that the gene-corrected cells maintained normal lineage distribution, while the B cell development of mock group was impaired. Moreover, gene-editing efficiency analysis of bone marrow samples 16 weeks after transplantation exhibited high editing rate (34±7.18% via HDR), comparable to the in vitro efficiency. The specificity of the Cas9 mRNA-based gene editing system was examined using unbiased Digenome-Seq. In total, 32 potential off-target sites were identified and deeply interrogated via targeted PCR and NGS analysis of XLSA iPSCs electroporated with Cas9 mRNA and sgRNA. No off-target cleavage events were detected at these sites, suggesting a lack of detectable off-target events. Finally, scRNA-seq of CD34+ HSPCs from healthy donor and XLSA patients revealed more HSC/LMPP and erythroid progenitor cells in older XLSA patient. Further analysis showed that cell cycle and gene expression in older HSC/LMPP cells were significantly different from that from healthy donors and younger patients. Hence, we speculated that the compensatory differentiation of HSCs caused by long-term functional red blood cell deficiency caused the abnormal expansion of HSCs, which led to the poor hematopoiesis in elderly patients. Our study firstly uses CRISPR/Cas9 gene-editing technology to correct the disease mutation in patient's CD34+ HSPCs and rescues ALAS2 expression and heme biosynthesis, directly confirming that this mutation is the pathogenic factor for XLSA. In addition, we dissect the transcriptional profile of CD34+ HSCPs from XLSA patients at single cell resolution for the first time, shedding light on mechanistic insights into the XLSA pathogenesis. The robust gene-correction rates and significant function rescue in patient's CD34+ HSPCs further suggest a curable option of gene-edited HSC transplantation for the treatment of the patients with XLSA. Disclosures Fang: EdiGene Inc.: Current Employment. Yuan:EdiGene Inc.: Current Employment. Yang:EdiGene Inc.: Current Employment. Yu:EdiGene Inc.: Current Employment. Zhang:EdiGene Inc.: Current Employment. Shi:EdiGene Guangzhou Inc.: Current Employment. Qi:Novogene Co, Ltd: Current Employment. Wei:EdiGene Inc.: Current Employment.
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Lawrence, Jody, Nicholas Kitchin, Annaliesa S. Anderson, Michael W. Pride, Kathrin U. Jansen, William C. Gruber, Yahong Peng, Kevin Yi, Charles Knirsch e Chris Webber. "12. Randomized Studies of Two Clostridioides (Clostridium) difficile Vaccine Formulations". Open Forum Infectious Diseases 7, Supplement_1 (1 ottobre 2020): S29. http://dx.doi.org/10.1093/ofid/ofaa439.057.
Testo completoAbstract (sommario):
Abstract Background Two formulations of investigational bivalent Clostridioides (Clostridium) difficile vaccine (QS-21 adjuvanted toxoid and toxoid-alone) were assessed for safety and immunogenicity in randomized studies in healthy adults 50–85 years of age. Methods The Phase 1 study of QS-21 adjuvanted toxoid vaccine randomized subjects 3:1 to 100 μg QS-21-containing C difficile vaccine or placebo; 3 doses were given according to 2 different schedules: a shortened month (Months 0, 1, 3) or day (Days 1, 8, 30) regimen. The Phase 2 toxoid-alone vaccine study randomized subjects 3:3:1 to receive 100 or 200 μg unadjuvanted C difficile vaccine formulation or placebo in Stages 1 and 2 (sentinel cohorts of different age groups), and 3:1 to receive the selected dose of unadjuvanted C difficile vaccine formulation or placebo in Stage 3. Three doses were given on a day (Days 1, 8, 30) regimen. Safety was the primary outcome for both studies. Immunogenicity was determined by measuring serum toxin A– and B–specific neutralizing antibodies. Results In the day regimen, 10 reports across both studies of grade 3 injection site redness postdose 2 triggered predefined stopping rules. Local reactions in both studies were more common among vaccine versus placebo recipients. Injection site pain predominated and was generally mild in severity. Systemic events were infrequent and generally mild-to-moderate in severity. Adverse events were reported by 50.0%–75.0% and 16.7%–50.0% of subjects in the QS-21 and toxoid-alone studies, respectively. Immune responses peaked around Day 37 (shortened-month regimen) or between Day 15 and Month 2 (day regimen), and remained above baseline throughout follow-up. Conclusion Both formulations demonstrated robust immunogenicity. However, both studies stopped early due to grade 3 injection site redness postdose 2 of the day (Days 1, 8, 30) regimen; neither formulation progressed to later stage development. Instead, an aluminum hydroxide-containing formulation of the vaccine candidate administered at 0, 1, and 6 months, which was safe and immunogenic in phase 1 and 2 studies, advanced to phase 3 studies. Disclosures Jody Lawrence, MD, Pfizer, Inc (Employee) Nicholas Kitchin, MD, Pfizer, Inc (Employee) Annaliesa S. Anderson, PhD, Pfizer (Employee, Shareholder) Michael W. Pride, PhD, Pfizer (Employee, Shareholder) Kathrin U. Jansen, PhD, Pfizer (Employee, Shareholder) William C. Gruber, MD, Pfizer (Employee, Shareholder) Yahong Peng, PhD, Pfizer (Employee, Shareholder) Charles Knirsch, MD, Pfizer (Employee) Chris Webber, MD, Pfizer Inc (Employee, Shareholder)
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Loveday, Kenneth, Doriana Froim, Anna Pisania, Alissa Fernald, Mathew Spaits, R. Scott Pearsall, Ravi Kumar e Kathryn W. Underwood. "ACE-536, a Novel GDF-Trap, Increases Hemoglobin, Hematocrit, and Red Blood Cell Levels in Rats and Monkeys." Blood 114, n. 22 (20 novembre 2009): 3603. http://dx.doi.org/10.1182/blood.v114.22.3603.3603.
Testo completoAbstract (sommario):
Abstract Abstract 3603 Poster Board III-540 Anemia is a common and debilitating complication of several tumor types, especially multiple myeloma and myelodysplastic syndrome, chemotherapy, chronic kidney disease, and many other conditions. The vast majority of therapies approved to treat anemia target the erythropoietin (EPO) pathway; however, recent studies suggest an increased risk of mortality associated with this class of therapy, arising from exposure to high levels of EPO. The TGF-β protein superfamily has been described to play a role in RBC development, but signals through a pathway distinct from EPO. We have developed ACE-536, a novel, GDF-trap derived from the Type II activin receptor fused to the Fc region of IgG1, which binds to and inhibits several ligands in TGF-b protein superfamily. Here, we report the effects of ACE-536 on hematologic parameters (RBC, hemoglobin, and hematocrit) in Sprague-Dawley rats and cynomolgus monkeys. The study in rats examined the effect of dose on increases in hematology parameters. Ten male and ten female rats per group received two SC doses (Days 1 and 15) of vehicle, or 6, 20 or 60 mg/kg of ACE-536; hematology parameters were measured on Day 29. Significant increases (P < 0.05) in RBC count were seen at all three dose levels in both male and female rats, which correlated with increases in hemoglobin and hematocrit. The maximum hemoglobin concentration occurred at 60 mg/kg in both sexes (combined mean of 18.2 g/dL versus 15.7 g/dL in vehicle control rats). The corresponding mean hematocrit at 60 mg/kg was 55% versus 47% in vehicle control rats. Two studies were conducted in monkeys to evaluate the time course and the dose response for induction of RBC, hemoglobin, and hematocrit. In the first study, the time course of erythropoiesis was investigated in 4 male and 4 female cynomolgus monkeys, which received subcutaneous (SC) doses of ACE-536 (10 mg/kg) on Days 1 and 8. Hematology parameters were evaluated prior to dosing and on Days 3, 8, 15, 29, and 44. Increases in hematology parameters were noted 48 hours later (Day 3) in both males and females, and maximum increases were seen by Day 29. The mean increases in hemoglobin were 1.4 g/dL on Day 3 and 2.2 g/dL on Day 29. In contrast, mean hemoglobin concentration decreased on Day 3 in two vehicle control animals (mean of -0.85 g/dL). In the second study, the hematologic dose response was investigated in 5 male and 5 female cynomolgus monkeys per group, which received SC doses of vehicle, or 0.4, 2, 10 or 30 mg/kg of ACE-536 on Days 1 and 15. Hematology parameters were evaluated prior to dosing and on Days 13 and 28. A dose-related increase in hematology parameters was seen in animals treated with ACE-536. Hemoglobin concentration decreased during the study in vehicle control animals (-1.5 g/dL on Days 13 and 28), most likely due to frequent blood collections for clinical chemistry and pharmacokinetic evaluations. In contrast, animals treated with ACE-536 had changes in hemoglobin concentration on Day 13 of -0.6, +0.45, +0.57, and +0.52 g/dL at doses of 0.4, 2, 10, and 30 mg/kg, respectively. In conclusion, the administration of ACE-536, a novel, GDF-trap, promotes robust increases in hemoglobin, hematocrit, and RBC levels in animals. This provides evidence that targeting the TGF-β signaling pathway is a viable approach to the treatment of anemia, and that ACE-536 may prove to be an effective alternative to EPO-based therapies. Disclosures: Loveday: Acceleron Pharma Inc: Employment. Froim:Acceleron Pharma Inc: Employment. Pisania:Acceleron Pharma Inc: Employment. Fernald:Acceleron Pharma Inc: Employment. Spaits:Acceleron Pharma Inc: Employment. Pearsall:Acceleron Pharma: Employment. Kumar:Acceleron Pharma Inc: Employment. Underwood:Acceleron Pharma: Employment.
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Srinivasan, Surabhi, Nguyen Tan, Hsin-Yuan Cheng, Yi Zhang, Silvia Tacheva-Grigorova, Tom Van Blarcom, Cesar Sommer, Duy Nguyen, Barbra Sasu e Siler Panowski. "Investigation of ALLO-316: A Fratricide-Resistant Allogeneic CAR T Targeting CD70 As a Potential Therapy for the Treatment of AML". Blood 136, Supplement 1 (5 novembre 2020): 23. http://dx.doi.org/10.1182/blood-2020-142161.
Testo completoAbstract (sommario):
CD70, a member of the TNF superfamily, is a type II transmembrane glycoprotein that interacts with its receptor (CD27) to promote survival of primed T cells and leads to formation of effector and memory T cells. Expression of CD70 in normal tissues is restricted to activated T and B lymphocytes and mature dendritic cells. CD70 is also widely expressed in various malignancies, including renal cell carcinoma (RCC) and acute myeloid leukemia (AML). The restricted expression pattern of CD70 in normal tissues makes it an attractive target for cancer therapeutics. Adoptive transfer of T cells expressing chimeric antigen receptors (CARs) is an exciting new therapeutic modality showing great promise in hematologic malignancies. Approval of two CD19-targeting autologous CAR Ts, Kymriah® and Yescarta®, has been followed with promising results from BCMA autologous CAR T clinical trials, showing that activity can extend to other targets. We have previously described the functional screening of a library of anti-CD70 scFv-based CARs and the identification of lead CD70 allogeneic CAR T cells (AlloCAR TTM) with robust activity against RCC cell lines both in vitro and in vivo. Here, we evaluate the anti-tumor activity and safety of a lead CD70 AlloCAR T (ALLO-316) for the treatment of AML. CD70 expression was evaluated and detected on three AML cell lines and in six primary AML patient samples, with 5/6 patient samples showing expression on 24%-99% of cells. CD70 expression will be profiled in a broader subset of AML patients and preliminary data will be presented. Despite the expression of CD70 on activated T cells it was possible to generate CD70 AlloCAR T cells. No CD70 expression was observed on CAR T cells after generation, suggesting either cells are succumbing to fratricide or are being "masked" by the CAR. CD70 was also not detected on Jurkat cells expressing CARs and this data, in combination with results showing CAR expression is protective when overexpressed in RCC cells support the phenomenon of "masking". Cellectis' TALEN® gene-editing was used to inactivate the TRAC and CD52 loci with the intent to minimize the risk of graft-versus-host disease and to confer resistance to ALLO-647, an anti-CD52 antibody that can be used as part of the conditioning regimen to deplete host alloreactive immune cells potentially leading to increased persistence and efficacy of the infused allogeneic cells. ALLO-316 cells were highly effective at lysing CD70-expressing target cells and eliminated greater than 99% of cells at the high effector to target (E:T) ratio and were unable to lyse AML cells in which CD70 was knocked out. Moreover, ALLO-316 cells were able to kill primary AML blasts with CD70 expression ex vivo. An orthotopic in vivo model utilizing the AML cell line MV4-11 was developed and anti-tumor activity was observed. In addition to evaluating efficacy against AML cell lines and tumors we also explored potential toxicity liabilities related to ALLO-316 treatment. Previous studies have reported that certain AML tumor antigens can also be expressed on normal hematopoietic progenitors and such expression could potentially lead to toxicity with targeted therapeutics. No detectable CD70 was observed by flow cytometry on purified CD34+ cells from 14 healthy donors. Taken together, our results support clinical development of CD70 AlloCAR T therapy for the treatment of AML. Disclosures Srinivasan: Allogene Therapeutics: Current Employment. Tan:Allogene Therapeutics: Current Employment. Cheng:Allogene Therapeutics, Inc.: Current Employment, Current equity holder in publicly-traded company. Zhang:Allogene Therapeutics: Current Employment, Current equity holder in publicly-traded company. Tacheva-Grigorova:Allogene Therapeutics: Current Employment, Current equity holder in publicly-traded company. Van Blarcom:Allogene Therapeutics, Inc.: Current Employment, Current equity holder in publicly-traded company. Sommer:Allogene Therapeutics, Inc.: Current Employment, Current equity holder in publicly-traded company. Nguyen:Allogene Therapeutics, Inc.: Current Employment, Current equity holder in publicly-traded company. Sasu:Allogene Therapeutics, Inc.: Current Employment, Current equity holder in publicly-traded company. Panowski:Allogene Therapeutics, Inc.: Current Employment, Current equity holder in publicly-traded company.
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Peterson, James, Daniel Drazan, Hanna Czajka, Jason Maguire, Jean-Louis Pregaldien, Ilkka Seppa, Roger Maansson et al. "6. Pentavalent Meningococcal (MenABCWY) Vaccine is Safe and Well Tolerated With Immunogenicity Noninferior to Coadministered MenB-FHbp and MenACWY-CRM in a Phase 2 Study of Healthy Adolescents and Young Adults". Open Forum Infectious Diseases 7, Supplement_1 (1 ottobre 2020): S25—S26. http://dx.doi.org/10.1093/ofid/ofaa439.051.
Testo completoAbstract (sommario):
Abstract Background Meningococcal serogroups A, B, C, W and Y cause nearly all meningococcal disease globally. Vaccination is complicated by different dosing recommendations for serogroup B (MenB) and quadrivalent (MenACWY) vaccines, which could be solved with a single pentavalent vaccine. This study in adolescents and young adults evaluated a new pentavalent MenABCWY vaccine that combines 2 licensed vaccines, MenB-FHbp (Trumenba®; bivalent rLP2086) and MenACWY-TT (Nimenrix®), into a single vaccine. Methods In this ongoing, randomized, controlled, observer-blinded, multicenter study (NCT03135834), MenB vaccine-naive and MenACWY-naive or -experienced healthy 10–25-year-olds were randomized 1:2 to MenABCWY (Month 0,6) or MenB-FHbp (Month 0,6) and MenACWY-CRM (Month 0). Immune responses were measured by serum bactericidal activity assays with human complement (hSBA) against serogroup A, C, W and Y strains and 4 diverse, vaccine-heterologous MenB strains. Endpoints included percentages of subjects achieving ≥ 4-fold rises in titers from baseline. Noninferiority of immune responses was assessed at the 10% margin (95% CI lower limit > −10%). Safety was assessed. Results Following dose 2, high percentages of MenABCWY (n=543) and MenB-FHbp (n=1057) recipients achieved ≥ 4-fold rises against each of the 4 MenB strains (75.8−94.7% vs 67.4−95.0%) and titers reaching at least the lower limit of quantification against all 4 strains combined (79.9% vs 74.3%; Figure 1A). MenABCWY was noninferior to MenB-FHbp for all 5 endpoints. MenABCWY was also noninferior to a single MenACWY-CRM dose with 75.5−96.9% and 93.0−97.4% of MenABCWY recipients after dose 1 or 2, respectively, achieving ≥ 4-fold rises against serogroup A, C, W and Y depending on prior MenACWY experience (Figure 1B). Local reactions and systemic events after MenABCWY or MenB-FHbp were similarly frequent, mostly mild/moderate in severity (Figure 2), and unaffected by MenACWY experience. Figure 1. Immune Responses as Measured in hSBA to (A) MenB Test Strains at 1 Month After Dose 2 and (B) MenA, MenC, MenW, and MenY Test Strains at 1 Month After Doses 1 and 2 Figure 2. (A) Local Reactions and (B) Systemic Events Reported Within 7 Days After Any Dose Conclusion MenABCWY 4-fold immune responses from baseline were robust and noninferior to MenB-FHbp and MenACWY-CRM administered separately. Vaccination was safe and well tolerated. The favorable benefit-risk profile supports further MenABCWY development as a simplified alternative to current meningococcal vaccination practices. Funded by Pfizer. Disclosures James Peterson, MD, Pfizer (Scientific Research Study Investigator) Daniel Drazan, MD, Pfizer (Scientific Research Study Investigator) Hanna Czajka, MD, PhD, Pfizer (Scientific Research Study Investigator) Jason Maguire, MD, Pfizer (Employee, Shareholder) Jean-Louis Pregaldien, MS, Pfizer (Employee, Shareholder) Ilkka Seppa, MD, Pfizer (Scientific Research Study Investigator) Roger Maansson, MS, Pfizer (Employee, Shareholder) Robert O’Neill, PhD, Pfizer (Employee, Shareholder) Annaliesa S. Anderson, PhD, Pfizer (Employee, Shareholder) Paul Balmer, PhD, Pfizer Inc (Employee, Shareholder) Johannes Beeslaar, MD, Pfizer (Employee, Shareholder) John L. Perez, MD, MA, Pfizer Inc (Employee, Shareholder)
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Smith, Pascal, Evelyn Teh, Ken Chisholm, Susanne Penny, Devanand Pinto, Kelcey Patterson, Brennan Dirk et al. "Proteomic analysis of plasma exosomes as biomarkers of response to MVP-S based immunotherapy". Journal of Immunology 208, n. 1_Supplement (1 maggio 2022): 66.09. http://dx.doi.org/10.4049/jimmunol.208.supp.66.09.
Testo completoAbstract (sommario):
Abstract Ovarian cancer is a deadly gynaecological disease owing to its late-stage diagnosis. Recurrence and chemo-resistance develop despite initial treatment success. Maveropepimut-S (MVP-S), formerly DPX-Survivac, is a DPX-based immune educating therapy that elicits robust, targeted and, sustained specific anti-tumor T- and B-cell responses in advanced ovarian cancer patients. Exosomes are a subtype of extracellular vesicles (EVs) secreted by cells involved in intercellular communication. They contain bioactive molecules that can influence the tumor, the immune system, and the extracellular environment. Hence, circulating exosomes could provide real time assessment of disease evolution making them ideal minimally invasive biomarkers for monitoring response to immunotherapies. Plasma exosomes were isolated from advanced ovarian cancer patients (n=22) at pre- and on-treatment with MVP-S based therapy (DeCidE1 trial, NCT02785250). Subjects with clinical benefit (>10% tumor shrinkage) to therapy displayed a lower overall total EV protein concentration. Untargeted proteomic profiling by liquid chromatography tandem mass spectrometry identified 227 proteins in exosomes. Of these, 95 (42%) were likely from non-tumor exosomes and involved in immune modulation such as regulation of humoral response and B-cell immunity. When compared to the plasma proteome, 128 (56%) proteins were found to be exclusive to exosomes. Functional annotations of these proteins suggest their role in modulating immune pathways related to Fc receptor mediated signaling and T-cell migration. The results highlight the potential value of plasma exosome assays for monitoring disease and as a potential surrogate response biomarker. Supported by -IMV inc. -NRC-IRAP R&D Certificate Program.
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Cichocki, Frank, Ryan Bjordahl, Svetlana Gaidarova, Sajid Mahmood, Paul Rogers, Moyar Q. Ge, Dan S. Kaufman, Sarah Cooley, Bahram Valamehr e Jeffrey S. Miller. "iPSC-Derived NK Cells and Anti-PD-1 Antibody Synergize to Enhance T Cell Cytokine and Cytolytic Responses Against Multiple Tumors". Blood 132, Supplement 1 (29 novembre 2018): 730. http://dx.doi.org/10.1182/blood-2018-99-114893.
Testo completoAbstract (sommario):
Abstract The development of immunotherapeutic monoclonal antibodies targeting checkpoint inhibitory receptors (CIR), such as programmed death 1 (PD-1), has transformed the oncology landscape. However, many tumor subtypes are resistant to CIR-targeted therapy, and relapse remains a significant concern. Therefore, combination of novel immunotherapies with CIR targeting remains a promising and widely investigated approach to bolster anti-tumor responses and to overcome tumor resistance to CIR therapy. Natural killer (NK) cells mediate direct tumor cell lysis and are key regulators of T cell responses through the production of inflammatory cytokines and chemokines. In many cancers, NK cell numbers are low and their functional responses are sub-optimal. The use of allogeneic NK cell immunotherapy has shown significant clinical promise for the treatment of acute myelogenous leukemia (AML). However, this approach has inherent limitations with respect to the number of NK cells that can be isolated and variability in the quantity and quality of NK cells between donors. To overcome these barriers, we have developed a system for large scale expansion of NK cells derived from induced pluripotent stem cells (iPSCs) to be combined with CIR antibodies for multiple tumor types (Figure 1A). iPSC derive NK (iNK) cells (defined as CD45+CD3-CD56+) differentiated with high efficiency in this culture system (Figure 1B), and overall expansion from the hematopoietic progenitor stage to end of the protocol was approximately 1 million-fold (Figure 1C). iNK cells displayed a cell surface phenotype typical of primary peripheral blood NK cells, with high expression of the cytotoxicity molecules granzyme B and perforin. Functionally, iNK cells degranulated and produced the pro-inflammatory cytokines tumor necrosis factor (TNF) and interferon (IFN)-γ at high frequencies in response to K562 cells (a myeloid leukemia cell line) (Figure 1D). These cells also effectively killed a broad range of solid tumor targets, including ovarian cancer (SKOV-3), lung cancer (A549) and pancreatic cancer (PANC-1) cell lines, in 2D tissue culture assays (Figure 1E). To interrogate the ability of iPSC-derived NK cells to synergize with CIR therapy, we used an in vitro 3D tumor spheroid system to model the combinatorial effects of T cells, iNK cells, and checkpoint blockade in the context of anti-tumor function. Using SKOV-3 spheroids as targets in a 160-hour killing assay, we found that iNK cells could mediate significant, but not complete destruction of tumor spheroids (46% tumor reduction). Addition of twice as many activated T cells by themselves induced tumor spheroid destruction (58% tumor reduction). The combination of iNK and activated T cells led to robust target cell destruction (71% tumor reduction). Importantly, the combination of activated T cells, iNK cells and anti-PD-1 antibody led to a near complete elimination of tumor spheroid targets, with greater than 99% tumor reduction (Figure 1F). In addition to synergistic lysis of tumor spheroids, analyses of cytokine secretion in iNK, activated T cell and CIR spheroid cultures revealed synergistic production of TNF and IFN-g (Figure 1G). The results from our 3D spheroid assays led us to hypothesize that iNK cells could recruit T cells to promote tumor lysis. To test this hypothesis, we performed transwell migration assays to test the relative abilities of iNK cells and K562 cells to promote T cell migration. We found that, relative to media controls, both iNK cells and K562 cells promoted T cell migration across transwells, and the highest amount of degree of migration was observed in the presence of both iNK and K562 cells (Figure 1H). Finally, we tested iNK-mediated T cell recruitment in an in vivo setting where iNK cells and IL-2 were injected with or without K562 cells into the peritoneal cavities of immune-deficient NSG mice, and activated T cells were injected retro-orbitally into the blood. Mice were sacrificed 4 days later, and total T cells in the peritoneal cavities were determined. Compared to the T cell only control group, iNK cells promoted significant T cell recruitment to the peritoneum, which was amplified upon iNK cell activation with the addition of K562 cells (Figure 1I). Together, our data demonstrate that iNK cells can serve as an off-the-shelf source of NK cells and have the potential to synergize with anti-PD-1 CIR therapy to enhance anti-tumor T cell responses. Disclosures Cichocki: Fate Therapeutics Inc.: Consultancy, Research Funding. Bjordahl:Fate Therapeutics Inc.: Employment. Gaidarova:Fate Therapeutics Inc: Employment. Mahmood:Fate Therapeutics Inc.: Employment. Rogers:Fate Therapeutics Inc: Employment. Ge:Fate Therapeutics Inc.: Employment. Kaufman:Fate Therapeutics: Consultancy, Research Funding. Valamehr:Fate Therapeutics Inc.: Employment.
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Zinzani, Pier Luigi, Vladimir Melnichenko, Krimo Bouabdallah, Jan Walewski, Alejandro Majlis, Laura Fogliatto, Dolores Caballero et al. "Pembrolizumab Monotherapy in Relapsed or Refractory Primary Mediastinal Large B-Cell Lymphoma (PMBCL): 3-Year Follow-up of the Keynote-170 Study". Blood 136, Supplement 1 (5 novembre 2020): 42–43. http://dx.doi.org/10.1182/blood-2020-137080.
Testo completoAbstract (sommario):
Introduction: No standard of care exists for patients (pts) with relapsed or refractory primary mediastinal B-cell lymphoma (rrPMBCL). As such, pts typically receive therapies recommended for diffuse large B-cell lymphoma, with poor prognosis. Similar to classical Hodgkin lymphoma, rrPMBCL tumors often overexpress the programmed death 1 (PD-1) ligands, PD-L1 and PD-L2. Data from the first full analysis of the phase 2 KEYNOTE (KN)-170 (NCT02576990) study showed that pembrolizumab provided effective and durable antitumor activity with a manageable safety profile in patients with rrPMBCL. These data led to the FDA approval of pembrolizumab for pts with rrPMBCL after ≥2 prior therapies. In this aggressive malignancy with few salvage options, duration of remission with PD-1 blockade remains a critical question. Here we present data from KN170 with an additional 24 months of follow-up in patients with rrPMBCL. Methods: Pts with rrPMBCL who had relapsed after or were ineligible for autologous stem cell transplant with ≥2 lines of prior therapy were enrolled in KN170 to receive 200 mg pembrolizumab IV Q3W until disease progression or toxicity, for up to 2 years. Tumor response was assessed Q12W with PET/CT scans by IWG 2007 criteria. The primary endpoint was objective response rate (ORR) by blinded independent central review (BICR). Secondary endpoints included duration of response (DOR), progression-free survival (PFS), overall survival (OS), and safety. Exploratory endpoints included response by Lugano 2014 criteria. The data cutoff date for this analysis was May 7, 2020. Results: At the data cutoff date, among all treated pts (N=53), 13 had completed 2 years of treatment and 40 had discontinued largely due to progression (n=30). The ORR was 45% (24/53; 95% CI 32-60) with a CR rate of 19% (10/53; 13/53 [25%] by Lugano criteria). No patient who achieved CR by BICR had received consolidation therapy or had progressed at data cutoff. After a median study follow-up of 43.1 months (range, 35.6-50.7) the median DOR was not reached (range, 1.1+ to 46.9+ mo); 76% of pts had a response duration ≥36 mo. Median PFS was 5.5 mo (95% CI, 2.7-15.1), with 36-mo PFS rate of 34%. The median OS was 22.3 mo (95% CI, 7.3 to not reached) with 36-mo OS rate of 45%. At data cutoff, 50 (94%) pts had at least one adverse event (AE), with 30 (57%) having a treatment-related AE. The most common treatment-related AEs were neutropenia (19%), asthenia (9%), hypothyroidism (8%), fatigue (6%), and pyrexia (6%). Grade ≥3 treatment-related AEs occurred in 12 (23%) pts. Six (11%) pts had an immune-mediated AE. There were no treatment-related grade 5 events. Conclusion: Results from the longer-term follow-up of KN170, the largest prospective clinical trial in rrPMBCL, shows that pembrolizumab provides robust and durable antitumor activity with a manageable safety profile in patients with rrPMBCL. Disclosures Zinzani: Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; TG Therapeutics, Inc.: Honoraria, Speakers Bureau; Servier: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Janssen: Consultancy, Honoraria, Speakers Bureau; Kirin Kyowa: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; EUSA Pharma: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Janssen-Cilag: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Kyowa Kirin: Consultancy, Speakers Bureau; Immune Design: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Immune Design: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; MSD: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Eusapharma: Consultancy, Speakers Bureau; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Verastem: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Sanofi: Consultancy, Membership on an entity's Board of Directors or advisory committees; Sandoz: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Portola: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Merck: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Gilead: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; ADC Therapeutics: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celltrion: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Bouabdallah:Roche: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Gilead Sciences: Consultancy, Honoraria. Walewski:Roche: Consultancy, Honoraria, Other: travel expenses, Research Funding; GSK/Novartis: Research Funding; Celgene: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Janssen-Cilag: Consultancy, Honoraria; Servier: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Bristol Myers Squibb: Consultancy. Caballero:Gilead: Other: travel; Roche: Other: travel; Janssen: Membership on an entity's Board of Directors or advisory committees, Other: travel; Celgene: Membership on an entity's Board of Directors or advisory committees, Other: travel; Kite: Membership on an entity's Board of Directors or advisory committees; Takeda: Other: travel; BMS: Other: travel. Christian:Acerta: Research Funding; Celgene: Research Funding; Genentech: Research Funding; Merck: Research Funding; Millenium: Research Funding; MorphoSys: Research Funding; F Hoffman-La Roche: Research Funding; Triphase: Research Funding; Seattle Genetics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Verastem: Membership on an entity's Board of Directors or advisory committees; AstraZenica: Membership on an entity's Board of Directors or advisory committees. Özcan:Takeda: Honoraria, Other: travel, Research Funding; Roche: Other: travel, Research Funding; Bayer: Research Funding; Abbvie: Other: travel, Research Funding; Archigen: Research Funding; Celgene: Research Funding; MSD: Research Funding; Novartis: Research Funding; Amgen: Honoraria, Other: travel; BMS: Other; Jazz: Other; Sanofi: Other; Abdi Ibrahim: Other; Janssen: Other: travel, Research Funding. Salles:Bristol Myers Squibb: Consultancy, Other; Takeda: Consultancy, Honoraria, Other; Gilead: Consultancy, Honoraria, Other: Participation in educational events; Debiopharm: Consultancy; Kite: Consultancy, Honoraria, Other; MorphoSys: Consultancy, Honoraria, Other; Janssen: Consultancy, Honoraria, Other: Participation in educational events; Novartis: Consultancy, Honoraria, Other; F. Hoffman-La Roche Ltd: Consultancy, Honoraria, Other; Celgene: Consultancy, Honoraria, Other: Participation in educational events; Epizyme: Consultancy; Karyopharm: Consultancy; Autolus: Consultancy; Amgen: Honoraria, Other: Participation in educational events; Abbvie: Consultancy, Honoraria, Other: Participation in educational events; Genmab: Consultancy. Shipp:Celgene: Honoraria; Ono Pharmaceutical: Honoraria; Bayer: Honoraria; Bristol Myers Squibb: Consultancy, Research Funding; Merck: Research Funding. Thompson:Merck Sharp & Dohme Corp.: Current Employment. Orlowski:Merck Sharp & Dohme Corp.: Current Employment. Marinello:Merck & Co., Inc., Kenilworth, NJ, USA: Other: Stock ownership; Merck & Co., Inc.: Other: Travel, accommodations, expenses; Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA: Current Employment. Armand:Pfizer: Consultancy; Merck & Co., Inc.: Consultancy, Honoraria, Research Funding; Adaptive: Consultancy, Research Funding; Affimed: Consultancy, Research Funding; Sigma Tau: Research Funding; Celgene: Consultancy; IGM: Research Funding; Otsuka: Research Funding; Tensha: Research Funding; Genentech: Research Funding; Infinity: Consultancy; ADC Therapeutics: Consultancy; Bristol-Myers Squibb: Consultancy, Honoraria, Research Funding; Roche: Research Funding.
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Nowakowski, Grzegorz, Annalisa Chiappella, Fangxin Hong, Randy D. Gascoyne, David W. Scott, William R. Macon, Rebecca L. King et al. "Potential Factors That Impact Lenalidomide/R-CHOP Efficacy in Previously Untreated Diffuse Large B-Cell Lymphoma in the ROBUST and ECOG-ACRIN 1412 Studies". Blood 134, Supplement_1 (13 novembre 2019): 4092. http://dx.doi.org/10.1182/blood-2019-123083.
Testo completoAbstract (sommario):
Background: Standard first-line treatment for patients diagnosed with diffuse large B-cell lymphoma (DLBCL) is the R-CHOP regimen. Despite numerous efforts to improve on this regimen, in all comers and specifically in those with activated B-cell (ABC)-type DLBCL, results from large randomized studies have failed to show a significant clinical advantage above this standard. Recently, 2 large clinical trials in previously untreated DLBCL evaluated the combination of lenalidomide/R-CHOP (R2-CHOP) vs standard R-CHOP and demonstrated discordant results. We analyzed data from these trials to identify differences that may have impacted the disparate trial outcomes while identifying patient characteristics associated with benefit from the addition of lenalidomide across the trials. Methods: The 2 trials included in this evaluation are ROBUST, an international, randomized, double-blinded, placebo-controlled, phase III study of R2-CHOP vs placebo/R-CHOP, and the Intergroup ECOG-ACRIN 1412 (E1412) trial, a US-only, randomized, open-label, phase II study of R2-CHOP vs R-CHOP. Additional key study design differences included dosing, and subtype timing and inclusion (both studies used gene expression profiling [GEP] by NanoString). R-CHOP dosing differed only for prednisone, which was a flat 100 mg dose, d1-5 in ROBUST and 100 mg/m2, d1-5 in E1412. The lenalidomide dose and schedule varied between studies: 15 mg/d, d1-14/21 for ROBUST (210 mg/cycle) and 25 mg/d, d1-10/21 (250 mg/cycle) for E1412. ROBUST enrolled only patients prospectively identified with the ABC subtype, whereas E1412 enrolled all otherwise eligible patients and retrospectively evaluated for ABC-type with prespecified ABC-type accrual and power parameters. Both studies enrolled patients with IPI status ≥ 2, ECOG PS 0-2, and Ann Arbor stage II-IV (E1412 stage II with mass > 10 cm). The primary endpoint in both was progression-free survival (PFS; central review for ROBUST and investigator-assessed for E1412). Secondary endpoints included event-free survival (EFS; key for ROBUST), overall and complete response rates, overall survival (OS), and safety. Results: ROBUST enrolled 570 ABC patients and E1412 enrolled 280 GCB, ABC, and unclassified DLBCL patients. Patient profiles in both studies (ROBUST [ABC]/E1412 [all COO]) were similar in terms of median ages (65 y/66 y), male sex (58%/61%), and ECOG PS 1-2 (55%/63%). Patients in E1412 were more high risk: IPI 2 (42% ROBUST/34% E1412), IPI ≥ 3 (58%/66%), and Ann Arbor stage III/IV disease (87%/97%). Importantly, there was a notable difference in the time from patient diagnosis to treatment: median 31 days for ROBUST and 21 days (22% > 31 d) for E1412. Only 6% of patients in ROBUST were treated within 2 weeks of diagnosis vs 30% in E1412. At a median follow-up of ~2.5 y for both studies, E1412 met its primary endpoint of improvement in PFS with 34% reduction in the risk of PD/death (2-y PFS: 76% vs 70%) for all patients, but did not reach statistical significance in ABC-DLBCL patients for 2-y PFS (71% vs 61%) or 2-y OS (88% vs 76%). ROBUST showed no significant improvement in the primary PFS endpoint (2-y PFS: 67% vs 64%) or 2-y OS (79% vs 80%) in ABC patients. Subgroup analyses of PFS comparing R2-CHOP vs R-CHOP control within each study suggested that certain baseline factors favor the R2-CHOP arm, including: female sex, high IPI score (≥ 3), non-bulky disease (< 7 cm), and high Ann Arbor stage IV disease. Conclusion: While the ROBUST study design integrated state-of-the-art, real-time GEP to prospectively identify ABC-DLBCL patients with rapid lab turnaround time, cases of logistical delays in tissue submission for central analysis and staging procedures may have resulted in accrual of patients with lower-risk DLBCL and prolonged time from diagnosis to treatment. In contrast, E1412 retrospectively stratified patients by COO through GEP and was able to accrue higher-risk patients irrespective of COO within a shorter time from diagnosis to treatment, which among other differences (eg, smaller population size, study phase), could contribute to the improvement over R-CHOP control. Our comparative findings have significant implications for the design of future biomarker-driven trials and provide momentum for studies of novel combinations with R-CHOP in DLBCL. Further investigation into other factors, including dose intensity and their correlation with outcomes, will be presented. Disclosures Nowakowski: Celgene: Consultancy, Research Funding; NanoString: Research Funding; MorphoSys: Consultancy, Research Funding; Genentech, Inc.: Research Funding; F. Hoffmann-La Roche Ltd: Research Funding; Selvita: Membership on an entity's Board of Directors or advisory committees; Curis: Research Funding; Bayer: Consultancy, Research Funding. Chiappella:Servier: Other: advisory board, Speakers Bureau; Roche: Speakers Bureau; Teva: Speakers Bureau; Celgene: Other: advisory board, Speakers Bureau; Janssen: Other: advisory board, Speakers Bureau. Hong:Dana Farber Cancer Institute: Employment; Merck: Consultancy. Scott:Roche/Genentech: Research Funding; Celgene: Consultancy; Janssen: Consultancy, Research Funding; NanoString: Patents & Royalties: Named inventor on a patent licensed to NanoSting [Institution], Research Funding. Amengual:Appia Pharmaceuticals: Research Funding; Epizyme: Speakers Bureau. Leonard:BeiGene: Consultancy; Miltenyi: Consultancy; Sandoz: Consultancy; Gilead: Consultancy; Celgene: Consultancy; ADC Therapeutics: Consultancy; MorphoSys: Consultancy; ADC Therapeutics: Consultancy; Epizyme, Inc: Consultancy; Miltenyi: Consultancy; Genentech, Inc./F. Hoffmann-La Roche Ltd: Consultancy; Karyopharm Therapeutics: Consultancy; Akcea Therapeutics: Consultancy; Merck: Consultancy; Nordic Nanovector: Consultancy; Akcea Therapeutics: Consultancy; Bayer Corporation: Consultancy; Sandoz: Consultancy; MorphoSys: Consultancy; Gilead: Consultancy; BeiGene: Consultancy; AstraZeneca: Consultancy; Nordic Nanovector: Consultancy; Sutro Biopharma: Consultancy; Merck: Consultancy. Friedberg:Bayer: Honoraria, Other: Data & Safety Monitoring Committee; Acerta: Other: Data & Safety Monitoring Committee. Kostakoglu:F. Hoffman-La Roche: Consultancy; Genentech: Consultancy. Jurczak:Morphosys: Research Funding; Bayer: Research Funding; Roche: Research Funding; Incyte: Research Funding; Gilead: Research Funding; Celgene Corporation: Research Funding; Sandoz: Membership on an entity's Board of Directors or advisory committees, Research Funding; AstraZeneca: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novo Nordisk: Research Funding; Servier: Research Funding; Takeda: Research Funding; TG Therapeutics: Research Funding; Celtrion: Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding; Loxo: Membership on an entity's Board of Directors or advisory committees, Research Funding. Yamamoto:Bristol-Myers Squibb: Consultancy, Honoraria; Bayer: Research Funding; Celgene Corporation: Consultancy, Honoraria, Research Funding; Mundipharma: Consultancy, Honoraria, Research Funding; Astra-Zeneca: Consultancy, Research Funding; Eisai: Consultancy, Honoraria, Research Funding; Meiji Seika Pharma: Consultancy, Honoraria; ARIAD: Research Funding; MSD: Consultancy, Honoraria; Gilead Sciences: Research Funding; Ono: Consultancy, Honoraria, Research Funding; SymBio: Research Funding; HUYA/IQVIA Services Japan: Consultancy, Honoraria; AbbVie: Consultancy, Research Funding; Chugai: Consultancy, Honoraria, Research Funding; Pfizer: Honoraria; Sumitomo Dainippon: Honoraria; Incyte: Research Funding; Janssen: Honoraria; Kyowa Kirin: Honoraria; Novartis: Honoraria, Research Funding; Otsuka: Honoraria; Sanofi: Honoraria; Solasia Pharma: Research Funding; Takeda: Consultancy, Honoraria, Research Funding. Czuczman:Celgene Corporation: Employment, Equity Ownership. Russo:Celgene Corporation: Employment, Equity Ownership. Hudak:Celgene Corporation: Employment, Equity Ownership; Novartis: Employment, Equity Ownership. Zhang:Celgene Corporation: Employment, Equity Ownership. Wade:Celgene: Employment, Equity Ownership. Kahl:ADC Therapeutics: Consultancy, Research Funding; BeiGene: Consultancy; TG Therapeutics: Consultancy; Seattle Genetics: Consultancy. Vitolo:Abbvie: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; F. Hoffmann-La Roche: Speakers Bureau; Juno Therapeutics: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Kite: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Gilead: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Janssen: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. OffLabel Disclosure: Yes, these trials discuss off-label clinical trial investigation of lenalidomide plus R-CHOP in patients with newly diagnosed diffuse large B-cell lymphoma.
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Martinowitz, Uri, Amy D. Shapiro, Doris V. Quon, Miguel Antonio Escobar, Christine L. Kempton, Peter Collins, Pratima Chowdary et al. "Pharmacokinetic Behavior of IB1001, An Investigational Recombinant Factor IX, in Patients with Hemophilia B: Repeat Pharmacokinetic Study and Subgroup Analysis". Blood 118, n. 21 (18 novembre 2011): 2267. http://dx.doi.org/10.1182/blood.v118.21.2267.2267.
Testo completoAbstract (sommario):
Abstract Abstract 2267 Introduction IB1001 is an investigational recombinant factor IX for the treatment and prevention of bleeding in individuals with hemophilia B. A randomized cross-over pharmacokinetic (PK) study demonstrated that IB1001 (75 IU/kg) compared with nonacog alfa (BeneFIX®). was non-inferior (lower bound of the 1-sided 95% confidence interval for the area under the concentration curve [AUC0–∞] was 90%) and was well tolerated. Here we report the findings from a repeat PK assessment, in which a subset of patients underwent a second PK evaluation with IB1001 only. In addition, we present the results of an exploratory analysis of IB1001 PK parameters to assess the relationship between the degree of sialylation and the pharmacokinetics of recombinant factor IX. [Griffith MJ et al. J Thromb Haemost 5 (Suppl 1), PM–043, 2007]. Methods In the randomized PK study, patients were assigned to receive 75 ± 5 IU/kg of IB1001 or nonacog alfa following a washout period of ≥5 days. Factor IX levels were determined pre-infusion and at 30 minutes, 1, 3, 6, 9, 12, 24, 36, 48, 60, and 72 hours post-infusion. The evaluation was repeated 5–28 days later, when a 75 ± 5 IU/kg dose of the alternate therapy was administered. Factor IX levels were assessed at the same time points. The repeat PK assessment was planned to include patients who had received 3–6 months of IB1001 prophylaxis following their initial PK assessment. Calculated PK parameters were identical to those determined during the randomized PK study: half-life (β-phase t1/2, determined using a robust regression approach [Lee ML et al. XVIth ISTH Congress, Florence, Italy, 1997]), maximum plasma concentration (Cmax) and AUC(0-∞) (determined by the trapezoidal rule). To explore the association between sialylation level and the PK behavior of IB1001, patients in the randomized PK study (n=32) were allocated to one of three subgroups based on the sialylation levels of the IB1001 lots used (see Table). Results Thirty-two evaluable patients were enrolled in the randomized PK study (Feb 2009–Aug 2010). Of these, 13 underwent repeat PK assessments with IB1001 after receiving 4–18 months of prophylaxis with IB1001. The results demonstrate the stability of PK parameters following up to 18 months of exposure to IB1001. No significant reduction in factor IX recovery or elimination half-life occurred in any patients over time. The sialylation subgroup analysis revealed that the use of IB1001 lots with the lowest sialylation levels (Group 1) resulted in slightly lower AUC levels when compared with nonacog alfa (see Table). When lots with intermediate or the highest sialylation levels were used, the AUC of IB1001 appeared similar (Group 2), or slightly higher (Group 3), than the corresponding nonacog alfa values. Although Cmax of IB1001 was lower in Group 1, it appears comparable with the nonacog alfa controls in all groups, suggesting that this was not an effect of sialylation but of individual biological variation. Conclusions The stability of IB1001 PK profile during prophylactic use was demonstrated in 13 patients and supported observations of the lack of inhibitor development over this period. The continued PK stability of IB1001 over time is of interest for the prophylactic treatment of hemophilia B.Evaluation of IB1001 sialylation levels was consistent with observations from previously reported nonclinical studies [Griffith MJ et al. J Thromb Haemost 5 (Suppl 1). Although the level of sialylation resulted in slightly different PK behavior, these differences may simply reflect the biological variation between individuals. Disclosures: Gomperts: Inspiration Biopharmaceuticals Inc: Consultancy. Lee:Inspiration Biopharmaceuticals Inc: Consultancy.
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Kirsch, Ilan, Zijun Yidan Xu-Monette, Thomas Snyder e Ken H. Young. "Defining Immunoglobulin Somatic Hypermutation in De Novo Diffuse Large B-Cell Lymphoma Patients: Potential Application for Prognosis and Risk Stratification". Blood 124, n. 21 (6 dicembre 2014): 1669. http://dx.doi.org/10.1182/blood.v124.21.1669.1669.
Testo completoAbstract (sommario):
Abstract Context Diffuse large B cell lymphoma (DLBCL) is a heterogeneous group of diseases with variable clinical presentation, morphologic features, genomics, gene expression signature and prognosis. Some of the variability in patient course and response to therapy is likely to represent a function of developmental stage and/or specific pathway of transformation. We have been engaged in a detailed investigation of the molecular and clinical features of a large cohort of patients with DLBCL at the MD Anderson Cancer Center. Through these analyses we have begun to subcategorize this patient population based on these distinctive clinical and biological parameters. In this current aspect of our investigation we have explored the prevalence of somatic hypermutation (SHM) of the immunoglobulin loci in these de novo DLBCL patients using the platform of multiplex PCR and high-throughput sequencing (immunoSEQ) developed by Adaptive Biotechnologies, Inc. It has previously been established that the presence or absence of somatic hypermutation is an independent prognostic factor in patients with chronic lymphocytic leukemia (CLL). The ultimate goal of this collaborative effort is to determine if a similar biological mechanism between somatic hypermutation and prognosis exists within the population of DLBCL patients or subset and to relate the presence of SHM to clinical, pathological, and molecular aspects of this disease. Objective In this study, we investigated whether the immunoSEQ (Adaptive Biotech) assay could be used to reliably discriminate dominant clones in diagnostic specimens from patients with DLBCL with regard to rearrangement status of the immunoglobulin heavy and light (kappa and lambda) chains and the presence or absence of SHM. Patients The study group consisted of 200 DLBCL patients treated with R-CHOP. Patients with primary mediastinal large B-cell lymphoma, primary cutaneous DLBCL, primary central nervous system DLBCL, and DLBCLs transformed from a low-grade B-cell lymphoma or associated with HIV infection were excluded. Methods Genomic DNA was extracted from FFPE sections of diagnostic lymph node specimens of patients with DLBCL. Immunoglobulin heavy and light chain sequences were then independently amplified using multiplex PCR with optimized primer sets. Following high-throughput sequencing, a bioinformatics pipeline clusters the sequences into distinct clonotypes to determine overall frequencies and to identify diagnostic clones. V, (D,) and J genes are also identified for each clonotype, and point mutations that are not known germ-line allele variants are assigned as somatic hypermutation events. Results Using both the IgH and IgL (kappa and lambda) we have been able to identify an index trackable sequence in 90%+ of the samples (we identify an index diagnostic sequence or sequences in about 70% of the cases using each assay individually). Using a definition of SHM as >2% point mutations in the observed V gene, the samples can be split into three distinct categories: 1, V(D)J or VJ rearranged with SHM (50-55%); 2, V(D)J or VJ rearranged without SHM (10-25%) and 3, DJ only evident (20-40%) The vast majority of complete V(D)J rearrangements are in-frame. Conclusions The IgH and IgL immunoSEQ assays are robust in their ability both to identify dominant sequences in diagnostic lymph node specimens from patients with DLBCL and to distinguish those clones in which evidence of somatic hypermutation is present. The distribution of SHM in these samples lends itself to potential correlative and stratifying analyses on this well-characterized patient cohort, and likely have significant application in other aggressive B-cell lymphoma patients. Disclosures Kirsch: Adaptive Biotechnologies: Employment, Equity Ownership. Snyder:Adaptive Biotechnologies, Inc: Employment, Equity Ownership.
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Minson, Katherine A., Madeline G. Huey, Amanda A. Hill, Irene Perez, Xiaodong Wang, Stephen Frye, H. Shelton Earp, Deborah DeRyckere e Douglas K. Graham. "Bone Marrow Stromal Cell Mediated Resistance to Mertk Inhibition in Acute Leukemia". Blood 128, n. 22 (2 dicembre 2016): 2819. http://dx.doi.org/10.1182/blood.v128.22.2819.2819.
Testo completoAbstract (sommario):
Abstract MERTK is a receptor tyrosine kinase of the TAM family (TYRO-3, AXL, MERTK) that is ectopically expressed in 30-50% of newly diagnosed pediatric acute lymphoblastic leukemia (ALL) patient samples and aberrantly expressed in 80-100% of pediatric and adult primary acute myeloid leukemia (AML) samples. MERTK inhibition mediated by shRNA or a small molecule inhibitor, MRX-2843, decreased colony-forming potential and induced apoptosis in leukemia cell cultures. Moreover, MERTK inhibition prolonged survival in mouse xenograft models of acute leukemia, but was not curative. In these models, treatment with MRX-2843 effectively reduced peripheral disease burden but was less effective in the bone marrow, suggesting a role for the bone marrow microenvironment in therapeutic resistance. Additionally, Gas6, a MERTK ligand, is a poor prognostic factor in AML, mediates increased resistance to cytotoxic chemotherapy in leukemia cells, and is expressed in the bone marrow. To determine the role of Gas6 produced by bone marrow stromal cells in mediating resistance to MERTK inhibition by MRX-2843, acute leukemia cell lines were cultured in the presence of a Gas6-producing fibroblast-like cell line (HS27) or bone marrow derived stromal cells (BMDSCs) from wild type or Gas6 knockout mice and induction of apoptosis and cell death was determined by flow cytometry after treatment with MRX-2843 or vehicle. Co-culture with the HS27 cell line significantly reduced cell death in Kasumi-1 AML cell cultures in response to treatment with 300nM MRX-2843 compared to leukemia cells alone (29.4% versus 60.5%, p=0.002). Similar results were observed in Nomo1 AML and 697 pre-B ALL cell cultures. To evaluate whether soluble factors mediated this protective effect, Kasumi-1 cells were cultured in HS27-conditioned medium in the presence or absence of MRX-2843. Interestingly, conditioned medium was not sufficient to provide protection from MRX-2843 induced apoptosis (86.0% vs. 85.5% in unconditioned medium). To more directly assess the role of Gas6, BMDSCs isolated from wild-type and Gas6 knockout C57Bl/6 mice were co-cultured with 697 leukemia cells and sensitivity to MRX-2843 was determined. BMDSCs from wild-type mice protected 697 leukemia cells from MRX-2843 induced cell death much more effectively than BMDSCs from Gas6 knockout mice (4.3% apoptotic and dead cells versus 72.4%, respectively). To investigate biochemical mechanisms of Gas6-mediated protection, Kasumi-1 AML cells were cultured with 200nM MRX-2843 or vehicle in the presence or absence of HS27 cells and expression and activation of MERTK, AXL, TYRO-3, and downstream signaling effectors STAT5, AKT, and ERK1/2 were determined by immunoblot. AXL was not expressed in Kasumi-1 cells with or without co-culture. Treatment with MRX-2843 mediated robust inhibition of MERTK activation indicated by reduced levels of phosphorylated protein in both the presence and absence of stromal cell co-culture. In contrast, activation of TYRO-3 was increased after treatment with MRX-2843 in leukemia cells co-cultured with HS27 stromal cells. Similarly, in the absence of co-culture MRX-2843 inhibited activation of STAT5, AKT, and ERK1/2. However, in the presence of HS27 cells there was robust activation of STAT5 that was sustained even after treatment with MRX-2843. In contrast, MRX-2843 inhibited activation of AKT and ERK1/2 in HS27 co-cultures, although higher doses were required. Together these data support a model whereby Gas6 produced by stromal cells mediates leukemia cell resistance to MERTK inhibition in the bone marrow by inducing activation of TYRO-3, thereby promoting downstream signaling and cell survival despite MERTK inhibition. Thus, combined treatment with MRX-2843 and a TAM ligand sink (eg MERTK-Fc), a TYRO-3 inhibitor, or a bone marrow mobilizing agent may be particularly effective therapeutic strategies. Disclosures Wang: Meryx, Inc: Equity Ownership, Patents & Royalties: MRX-2843. Frye:Meryx, Inc: Equity Ownership, Patents & Royalties: MRX-2843. Earp:Meryx, Inc: Equity Ownership, Patents & Royalties: targeting MERTK. DeRyckere:Meryx, Inc: Equity Ownership, Patents & Royalties: targeting MERTK. Graham:Meryx, Inc: Equity Ownership, Patents & Royalties: targeting MERTK.