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1
Tabata, Rikako, SungGi Chi, Junichiro Yuda, and Yosuke Minami. "Emerging Immunotherapy for Acute Myeloid Leukemia." International Journal of Molecular Sciences 22, no. 4 (February 16, 2021): 1944. http://dx.doi.org/10.3390/ijms22041944.
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Several immune checkpoint molecules and immune targets in leukemic cells have been investigated. Recent studies have suggested the potential clinical benefits of immuno-oncology (IO) therapy against acute myeloid leukemia (AML), especially targeting CD33, CD123, and CLL-1, as well as immune checkpoint inhibitors (e.g., anti-PD (programmed cell death)-1 and anti-CTLA4 (cytotoxic T-lymphocyte-associated protein 4) antibodies) with or without conventional chemotherapy. Early-phase clinical trials of chimeric antigen receptor (CAR)-T or natural killer (NK) cells for relapsed/refractory AML showed complete remission (CR) or marked reduction of marrow blasts in a few enrolled patients. Bi-/tri-specific antibodies (e.g., bispecific T-cell engager (BiTE) and dual-affinity retargeting (DART)) exhibited 11–67% CR rates with 13–78% risk of cytokine-releasing syndrome (CRS). Conventional chemotherapy in combination with anti-PD-1/anti-CTLA4 antibody for relapsed/refractory AML showed 10–36% CR rates with 7–24 month-long median survival. The current advantages of IO therapy in the field of AML are summarized herein. However, although cancer vaccination should be included in the concept of IO therapy, it is not mentioned in this review because of the paucity of relevant evidence.
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Cartellieri, Marc, Irene Michalk, Malte von Bonin, Thomas Krüger, Slava Stamova, Stefanie Koristka, Claudia Arndt, et al. "Chimeric Antigen Receptor-Engineered T Cells for Immunotherapy of Acute Myeloid Leukemia." Blood 118, no. 21 (November 18, 2011): 2618. http://dx.doi.org/10.1182/blood.v118.21.2618.2618.
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Abstract Abstract 2618 Adoptive transfer of antigen-specific T cells emerged as an attractive strategy to provide cancer patients with immune cells of a desired specificity. The efficacy of such adoptive transfers has been demonstrated in clinical studies. However, enrichment and expansion of tumor-specific T cells are time-consuming and often ineffective, due to the low frequency of tumor-specific precursors in vivo. Alternatively, polyclonal T cells can be genetically modified with chimeric antigen receptors (CARs) to provide these cells with new antigen specificities. CARs consist of a binding moiety specifically recognizing a tumor cell surface antigen fused to a signaling chain derived from a lymphocyte activating receptor. The chimeric receptor approach is able to bypass many of the mechanisms by which tumors avoid immunorecognition, such as MHC down-regulation, lack of expression of costimulatory molecules, and induction of T cell suppression. Acute myeloid leukemia (AML) is an intrinsically resistant disease and even though the majority of the patients initially respond to chemotherapy, the 3-year survival rate is still low. A promising target for immunotherapy of AML is CD33, which is absent on normal pluripotent hematopoietic stem cells and normal tissues, but is present on leukemic blasts in 85–90 % of adult and pediatric AML cases independent of the subtype of AML. Novel human CD33-specific CARs were constructed by fusing a CD33 specific scFv in series with CD3ζ chain plus an additional costimulatory sequence derived from CD28 (Fig. 1A). Both native human CD8+ and CD4+ T cells engrafted with CD33-specific CARs exhibited antigen-specific cytokine secretion, proliferation and target cell lysis (Fig. 1B, C). Moreover, AML blast derived from patients were efficiently killed by allogeneic CAR-engrafted T cells (Fig. 1D). Next, the CD33-specific scFv was fully humanized and afterwards incorporated into the CAR constructs. With this humanized CAR engrafted T cells from AML patients could be redirected against CD33+ cell lines and autologous AML blasts. Upon antigen-recognition, the modified T cells secreted high amount of inflammatory cytokines and efficiently killed the target cells.Fig. 1:Human CAR-engrafted T cells mediate effector functions against CD33+ target cells.A. Schematic representation of the CD33-specific CARs. VL: variable light chain; VH: variable heavy chain; E-Tag: linker epitope in between the VL and the VH chain.Fig. 1:. Human CAR-engrafted T cells mediate effector functions against CD33+ target cells. / A. Schematic representation of the CD33-specific CARs. VL: variable light chain; VH: variable heavy chain; E-Tag: linker epitope in between the VL and the VH chain.B. Cytotoxic effector functions of CAR engrafted human CD8+ and CD4+ T cells against the CD33+ blast line U937 were measured in a standard chromium release assay after 6h of co-incubation.B. Cytotoxic effector functions of CAR engrafted human CD8+ and CD4+ T cells against the CD33+ blast line U937 were measured in a standard chromium release assay after 6h of co-incubation.C. Killing of patient-derived AML blast by allogeneic CAR engrafted T cells was measured in a flow cytometer by exact cell count after 48h of co-cultivation. Three independent donor/patient pairings are shown.C. Killing of patient-derived AML blast by allogeneic CAR engrafted T cells was measured in a flow cytometer by exact cell count after 48h of co-cultivation. Three independent donor/patient pairings are shown. Until now, one major obstacle for an adoptive therapy of genetically modified T cells is the limited amount of T cells that can be isolated from AML patients and modified in vitro. For an efficient in vitro expansion restricted to CAR modified T cells from patients we developed a new method based on a novel CAR-mediated strategy. For this purpose, magnetic beads were coated with an antibody recognizing an epitope (Fig. 1A, E-Tag) which we included in the linker domain between the heavy and the light chain of the scFv portion of the CAR. Adding such magnetic beads to cultures of CAR modified T cells, the CAR engrafted T cells were expanded to similar extends as polyclonal T cell populations with anti-CD3/anti-CD28 coated beads. Furthermore, the antibody-coated beads can be used to isolate the CAR engrafted T cells after expansion and get rid of any contaminating non-modified cells. It may also be useful for elimination of CAR expressing T cells in vivo if necessary. The feasibility of the described method was not limited to CD33 specific CARs but was also functional for CARs equipped with scFvs of other specificities. Therefore, it might be universally applicable for the expansion and preparation of CAR modified T cell grafts in vitro before adoptive transfer back in patients. Taken together, we describe novel humanized CD33-specific CARs that (I) can be specifically expanded, (II) specifically eliminated, if necessary, and (III) may therefore become a novel potent treatment option for a cellular therapy of AML patients. Disclosures: No relevant conflicts of interest to declare.
3
Epperly, Rebecca, Stephen Gottschalk, and Mireya Paulina Velasquez. "Harnessing T Cells to Target Pediatric Acute Myeloid Leukemia: CARs, BiTEs, and Beyond." Children 7, no. 2 (February 17, 2020): 14. http://dx.doi.org/10.3390/children7020014.
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Outcomes for pediatric patients with acute myeloid leukemia (AML) remain poor, highlighting the need for improved targeted therapies. Building on the success of CD19-directed immune therapy for acute lymphocytic leukemia (ALL), efforts are ongoing to develop similar strategies for AML. Identifying target antigens for AML is challenging because of the high expression overlap in hematopoietic cells and normal tissues. Despite this, CD123 and CD33 antigen targeted therapies, among others, have emerged as promising candidates. In this review we focus on AML-specific T cell engaging bispecific antibodies and chimeric antigen receptor (CAR) T cells. We review antigens being explored for T cell-based immunotherapy in AML, describe the landscape of clinical trials upcoming for bispecific antibodies and CAR T cells, and highlight strategies to overcome additional challenges facing translation of T cell-based immunotherapy for AML.
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Hao, Fang, Christine Sholy, Chen Wang, Min Cao, and Xunlei Kang. "The Role of T Cell Immunotherapy in Acute Myeloid Leukemia." Cells 10, no. 12 (December 1, 2021): 3376. http://dx.doi.org/10.3390/cells10123376.
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Acute myeloid leukemia (AML) is a heterogeneous disease associated with various alterations in T cell phenotype and function leading to an abnormal cell population, ultimately leading to immune exhaustion. However, restoration of T cell function allows for the execution of cytotoxic mechanisms against leukemic cells in AML patients. Therefore, long-term disease control, which requires multiple therapeutic approaches, includes those aimed at the re-establishment of cytotoxic T cell activity. AML treatments that harness the power of T lymphocytes against tumor cells have rapidly evolved over the last 3 to 5 years through various stages of preclinical and clinical development. These include tissue-infiltrated lymphocytes (TILs), bispecific antibodies, immune checkpoint inhibitors (ICIs), chimeric antigen receptor T (CAR-T) cell therapy, and tumor-specific T cell receptor gene-transduced T (TCR-T) cells. In this review, these T cell-based immunotherapies and the potential of TILs as a novel antileukemic therapy will be discussed.
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Boyiadzis, Michael M., Ivan Aksentijevich, Daniel A. Arber, John Barrett, Renier J. Brentjens, Jill Brufsky, Jorge Cortes, et al. "The Society for Immunotherapy of Cancer (SITC) clinical practice guideline on immunotherapy for the treatment of acute leukemia." Journal for ImmunoTherapy of Cancer 8, no. 2 (October 2020): e000810. http://dx.doi.org/10.1136/jitc-2020-000810.
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Acute leukemia is a constellation of rapidly progressing diseases that affect a wide range of patients regardless of age or gender. Traditional treatment options for patients with acute leukemia include chemotherapy and hematopoietic cell transplantation. The advent of cancer immunotherapy has had a significant impact on acute leukemia treatment. Novel immunotherapeutic agents including antibody-drug conjugates, bispecific T cell engagers, and chimeric antigen receptor T cell therapies have efficacy and have recently been approved by the US Food and Drug Administration (FDA) for the treatment of patients with acute leukemia. The Society for Immunotherapy of Cancer (SITC) convened a panel of experts to develop a clinical practice guideline composed of consensus recommendations on immunotherapy for the treatment of acute lymphoblastic leukemia and acute myeloid leukemia.
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Tasian, Sarah K. "Acute myeloid leukemia chimeric antigen receptor T-cell immunotherapy: how far up the road have we traveled?" Therapeutic Advances in Hematology 9, no. 6 (May 17, 2018): 135–48. http://dx.doi.org/10.1177/2040620718774268.
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Chemotherapy resistance and relapse remain significant sources of mortality for children and adults with acute myeloid leukemia (AML). Further intensification of conventional cytotoxic chemotherapy is likely not feasible due to the severity of acute and long-term side effects upon normal tissues commonly induced by these drugs. Successful development and implementation of new precision medicine treatment approaches for patients with AML, which may improve leukemia remission and diminish toxicity, is thus a major priority. Tumor antigen-redirected chimeric antigen receptor (CAR) T-cell immunotherapies have induced remarkable responses in patients with relapsed or chemorefractory B-lymphoblastic leukemia, and similar strategies are now under early clinical study in adults with relapsed/refractory AML. However, potential on target/off tumor toxicity of AML CAR T-cell immunotherapies, notably aplasia of normal myeloid cells, may limit broader implementation of such approaches. Careful selection of optimal target antigens, consideration of toxicity mitigation strategies, and development of methodologies to circumvent potential CAR T-cell resistance are essential for successful implementation of cellular immunotherapies for patients with high-risk AML.
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Pizzitola, Irene, Fernando Anjos-Afonso, Kevin Rouault-Pierre, Francois Lassailly, Sarah Tettamanti, Andrea Biondi, Ettore Biagi, and Dominique Bonnet. "Chimeric Antigen Receptor for Specific Targeting of Acute Myeloid Leukemia." Blood 120, no. 21 (November 16, 2012): 4225. http://dx.doi.org/10.1182/blood.v120.21.4225.4225.
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Abstract Abstract 4225 Despite the progress in the treatment of acute myeloid leukemia (AML) achieved in the last decades, a significant number of patients are still refractory to or relapse after conventional chemotherapy regimens. Therefore it is necessary to develop novel alternative approaches. Immunotherapy with T cells genetically modified to express chimeric antigen receptors (CARs) represent a valid option in this sense. CARs are artificial T-cell receptors constituted by a specific antigen-binding domain, and a signaling region, that, upon antigen recognition, leads to T-cell activation, and lysis of the target cells. AML is a potential optimal target for CAR strategy because of the over-expression of a number of surface antigens like CD33, CD123. Since CD33 is also expressed on normal hematopoietic stem/progenitors cells (HSPCs) resulting in a potential severe impairment of normal myelopoiesis, CD123 has recently emerged as new potential attractive molecules based on its differential expression pattern, being still wildly overexpressed by AML population, and at the same time less expressed on HSPCs. Here we describe the in vivo efficacy and the safety of this approach based on Cytokine-Induced-Killers (CIK) cells genetically modified to express CAR molecules specific for the CD33 or CD123 antigen. Once injected into low-level AML engrafted NSG mice (median of hCD45+CD33+ 0.6% before treatment), genetically modify T cells had a potent antitumor effect. Indeed, the bone marrow of control untreated animals or mice treated with un-manipulated CIK cells, was infiltrated by leukemic cells (86% and 81% leukemic engraftment), while in 7/8 anti-CD33-CD28-OX40-ζ and 8/10 anti-CD123-CD28-OX40-ζ treated mice we couldn't detect any AML cells. Similar results have been obtained when the treatment via T cell injection start when high AML burden has been obtained (median of hCD45+CD33+ 70% before treatment). One week after the last CIK's injection the level of AML engraftment was 96%, 87%, 0.35% and 0.34% for untreated mice, mice treated with un-manipulated CIK cells and with anti-CD33-CD28-OX40-ζ and anti-CD123-CD28-OX40-ζ transduced CIK-cells respectively. We performed secondary transplantation on the residual AML cells present in these animals and mice were treated again with transduced CIK cells. Residual AML cells were still sensitive to CARs approach, leading once again to an almost complete eradication of the disease (median level of hCD45+CD33+ engraftment was 98%, 0.02% and 0.04% respectively for untreated mice, anti-CD33-CD28-OX40-ζ and anti-CD123-CD28-OX40-ζ transduced CIK-cells). Furthermore, a fundamental issue was to determine the safety profile of such approach against normal hematopoietic precursors. In untreated mice injected with primary cord blood derived CD34+ cells the level of engraftment of hCD45 compartment was 42% whilst in mice treated with un-manipulated, anti-CD33-CD28-OX40-ζ or anti-CD123-CD28-OX40-ζ transduced CIK-cells the levels of human compartment was 40%, 11.7% and 26.3% respectively. Moreover when we consider specifically the CD34+CD38- compartment, enriched in HSC, the level of engraftment was 1.92%, 1.02%, 0.55% and 0.83%. Secondary transplantations are now ongoing to give a more complete profile about the remaining HSC repopulating capability after treatment. To more closely mimic a physiological context, similar experiments are ongoing using mice engrafted with normal adult bone marrow instead of umbilical cord blood. These experiments should offer relevant information concerning the efficacy and safety of the proposed strategy particularly in the context of minimal residual disease in high-risk transplanted AML patients. Moreover CAR approach could be potentially used to treat patients resistant to conventional chemotherapeutic approaches or for whom high dose chemotherapy treatment could not be proposed. Disclosures: No relevant conflicts of interest to declare.
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Maucher, Marius, Micha Srour, Sophia Danhof, Hermann Einsele, Michael Hudecek, and Ibrahim Yakoub-Agha. "Current Limitations and Perspectives of Chimeric Antigen Receptor-T-Cells in Acute Myeloid Leukemia." Cancers 13, no. 24 (December 7, 2021): 6157. http://dx.doi.org/10.3390/cancers13246157.
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Adoptive transfer of gene-engineered chimeric antigen receptor (CAR)-T-cells has emerged as a powerful immunotherapy for combating hematologic cancers. Several target antigens that are prevalently expressed on AML cells have undergone evaluation in preclinical CAR-T-cell testing. Attributes of an ‘ideal’ target antigen for CAR-T-cell therapy in AML include high-level expression on leukemic blasts and leukemic stem cells (LSCs), and absence on healthy tissues, normal hematopoietic stem and progenitor cells (HSPCs). In contrast to other blood cancer types, where CAR-T therapies are being similarly studied, only a rather small number of AML patients has received CAR-T-cell treatment in clinical trials, resulting in limited clinical experience for this therapeutic approach in AML. For curative AML treatment, abrogation of bulk blasts and LSCs is mandatory with the need for hematopoietic recovery after CAR-T administration. Herein, we provide a critical review of the current pipeline of candidate target antigens and corresponding CAR-T-cell products in AML, assess challenges for clinical translation and implementation in routine clinical practice, as well as perspectives for overcoming them.
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Campillo-Davo, Diana, Sébastien Anguille, and Eva Lion. "Trial Watch: Adoptive TCR-Engineered T-Cell Immunotherapy for Acute Myeloid Leukemia." Cancers 13, no. 18 (September 8, 2021): 4519. http://dx.doi.org/10.3390/cancers13184519.
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Despite the advent of novel therapies, acute myeloid leukemia (AML) remains associated with a grim prognosis. This is exemplified by 5-year overall survival rates not exceeding 30%. Even with frontline high-intensity chemotherapy regimens and allogeneic hematopoietic stem cell transplantation, the majority of patients with AML will relapse. For these patients, treatment options are few, and novel therapies are urgently needed. Adoptive T-cell therapies represent an attractive therapeutic avenue due to the intrinsic ability of T lymphocytes to recognize tumor cells with high specificity and efficiency. In particular, T-cell therapies focused on introducing T-cell receptors (TCRs) against tumor antigens have achieved objective clinical responses in solid tumors such as synovial sarcoma and melanoma. However, contrary to chimeric antigen receptor (CAR)-T cells with groundbreaking results in B-cell malignancies, the use of TCR-T cells for hematological malignancies is still in its infancy. In this review, we provide an overview of the status and clinical advances in adoptive TCR-T-cell therapy for the treatment of AML.
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Allison, Michaela, Joel Mathews, Taylor Gilliland, and Stephen O. Mathew. "Natural Killer Cell-Mediated Immunotherapy for Leukemia." Cancers 14, no. 3 (February 8, 2022): 843. http://dx.doi.org/10.3390/cancers14030843.
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Leukemia is a malignancy of the bone marrow and blood resulting from the abnormal differentiation of hematopoietic stem cells (HSCs). There are four main types of leukemia including acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), chronic myeloid leukemia (CML), and chronic lymphocytic leukemia (CLL). While chemotherapy and radiation have been conventional forms of treatment for leukemia, these therapies increase infection susceptibility, adverse side effects and immune cell inactivation. Immunotherapies are becoming promising treatment options for leukemia, with natural killer (NK) cell-mediated therapy providing a specific direction of interest. The role of NK cells is critical for cancer cell elimination as these immune cells are the first line of defense against cancer proliferation and are involved in both recognition and cytolysis of rapidly dividing and abnormal cell populations. NK cells possess various activating and inhibitory receptors, which regulate NK cell function, signaling either inhibition and continued surveillance, or activation and subsequent cytotoxic activity. In this review, we describe NK cells and NK cell receptors, functional impairment of NK cells in leukemia, NK cell immunotherapies currently under investigation, including monoclonal antibodies (mAbs), adoptive transfer, chimeric antigen receptor-NKs (CAR-NKs), bi-specific/tri-specific killer engagers (BiKEs/TriKEs) and future potential targets of NK cell-based immunotherapy for leukemia.
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Zhang, Jiachang. "Newly Developed Treatments for Acute Lymphoblastic and Acute Myeloid Leukemia." SHS Web of Conferences 144 (2022): 01007. http://dx.doi.org/10.1051/shsconf/202214401007.
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Chemotherapy has been dominating the field of cancer treatment for a long time, however, its limitations have been revealed over time. Therefore, several antigen proteins and chimeric antigen receptorT cells (CAR-T) involved in the immunotherapy of acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML) have been introduced. This paper provides details on the mechanisms, implemented investigations, and drawbacks of the immunotherapy for ALL and AML. Current studies have shown that CAR-T cell therapy can eliminate pediatric ALL relapse along with treating B cell ALL. With the appearance of CAR-T cell therapy, especially CD19-, CD20-, and CD22-directed CAR-T cells, aggressive acute lymphomas involving ALL become treatable. Studies have also shown that AML can be treated with FLT3 inhibitors and immunotherapy including monoclonal antibodies (mbA) and CD33-, CD123-directed CAR-T cells. Anti-CD33 monoclonal antibodies can combine with calicheamicin, a cytotoxic agent in DNA strand cleavage, and monotherapy of gemtuzumab ozogamicin (GO), an antibody-drug conjugate, and this combination has been proved to extend the overall survival of both newly treated patients and R/R AML patients who are unable to tolerate standard chemotherapy.
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Mahalleh, Mehrdad, Mahsima Shabani, Elham Rayzan, and Nima Rezaei. "Reinforcing the primary immunotherapy modulators against acute leukemia; monoclonal antibodies in AML." Immunotherapy 11, no. 18 (December 2019): 1583–600. http://dx.doi.org/10.2217/imt-2019-0043.
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Recent therapeutic advances in cancer treatment recruit immune system potentiation against malignant cells. Numerous ongoing clinical trials on immunotherapy methods, either monotherapy or combination therapy, are investigating the impeding factors on the way of acute myeloid leukemia (AML) treatment. Due to the genetic diversity in AML progenitors, combining various strategies is more likely to be useful for improving patient outcomes. This review describes the details of applying monoclonal antibodies against AML, focusing on CD33, CD123, FLT3, CD45 and CD66 targeting. Furthermore, it clarifies the importance of immunotoxins, bispecific antibodies, chimeric antigen receptor (CAR)-T cells and T cell receptor-modified cells as reinforcing agents for monoclonal antibodies.
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Dutour, A., V. Marin, I. Pizzitola, S. Valsesia-Wittmann, D. Lee, E. Yvon, H. Finney, et al. "In VitroandIn VivoAntitumor Effect of Anti-CD33 Chimeric Receptor-Expressing EBV-CTL againstCD33+Acute Myeloid Leukemia." Advances in Hematology 2012 (2012): 1–10. http://dx.doi.org/10.1155/2012/683065.
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Genetic engineering of T cells with chimeric T-cell receptors (CARs) is an attractive strategy to treat malignancies. It extends the range of antigens for adoptive T-cell immunotherapy, and major mechanisms of tumor escape are bypassed. With this strategy we redirected immune responses towards the CD33 antigen to target acute myeloid leukemia. To improvein vivoT-cell persistence, we modified human Epstein Barr Virus-(EBV-) specific cytotoxic T cells with an anti-CD33.CAR. Genetically modified T cells displayed EBV and HLA-unrestricted CD33 bispecificityin vitro. In addition, though showing a myeloablative activity, they did not irreversibly impair the clonogenic potential of normal CD34+hematopoietic progenitors. Moreover, after intravenous administration into CD33+human acute myeloid leukemia-bearing NOD-SCID mice, anti-CD33-EBV-specific T cells reached the tumor sites exerting antitumor activityin vivo. In conclusion, targeting CD33 by CAR-modified EBV-specific T cells may provide additional therapeutic benefit to AML patients as compared to conventional chemotherapy or transplantation regimens alone.
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Chien, Christopher Daniel, Christopher Tor Sauter, Kazusa Ishii, Sang Minh Nguyen, Feng Shen, Sarah K. Tasian, Weizao Chen, Dimiter S. Dimitrov, and Terry J. Fry. "Preclinical Development of FLT3-Redirected Chimeric Antigen Receptor T Cell Immunotherapy for Acute Myeloid Leukemia." Blood 128, no. 22 (December 2, 2016): 1072. http://dx.doi.org/10.1182/blood.v128.22.1072.1072.
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Abstract Background: Outcomes for adults and children with acute myeloid leukemia (AML) are dismal with 20-40% and 60% 5-year event-free survival, respectively. Alternative therapeutic strategies for AML are thus needed to improve outcomes. Chimeric antigen receptor (CAR) T cell immunotherapy has induced remarkable clinical responses in multiple phase 1 clinical trials for patients with relapsed or chemorefractory B cell leukemias, encouraging great interest in developing similar approaches for AML. Prior studies have demonstrated efficacy of CD33 or CD123-redirected CAR T cells in AML models, although the genetic heterogeneity of AML will likely require identification of additional therapeutic targets. In the current studies, we report preliminary in vitro and in vivo efficacy of new CAR T cells targeting the FMS-like tyrosine kinase 3 (FLT3) in human AML. FLT3 mutations via internal tandem duplication or kinase domain point mutations occur in approximately 25% of AML and result in FLT3 surface protein overexpression, suggesting potential efficacy of FLT3-targeting therapies. Both types of FLT3 alterations induce ligand-independent activation of FLT3 signaling, further demonstrating a critical role of FLT3 in AML pathogenesis. Hypothesis: FLT3 is a promising target for CAR T cell immunotherapy based treatment of AML. Results: Quantitative flow cytometric analysis of human AML cell lines demonstrated FLT3 surface expression ranging from 1338 (MOLM-13), 2594 (MOLM-14), and 2710 (MV4;11) receptors/cell versus 623 receptors/cell on negative control U937 cells. We first generated FLT3-redirected CAR construct consisting of a single chain variable fragment (scFv) derived from a well-characterized anti-human FLT3 antibody coupled to T cell 4-1BB (CD137) costimulatory and CD3-zeta activation domains. CD33 CAR T cells based on Gemtuzumab created by identical methodologies were also used as AML CAR T cell controls. In vitro studies verified that human T cells transduced with the FLT3 CAR construct induced interferon-gamma and interleukin-2 production after co-culture with AML cell lines MOLM-13, MOLM-14, and MV4;11. One dose of FLT3 CAR T cells inhibited leukemia proliferation in vivo in NOD-SCID-IL2Rγc-/- (NSG) mice engrafted with FLT3-mutant MOLM-13 or MOLM14 cell lines. These first data demonstrate potent preclinical activity of FLT3 CAR T cells and warrant further study in additional AML models. However, on target/off tumor toxicities can occur with AML antigen-targeted immunotherapies, as previously reported in studies of CD33 and CD123 CAR T cells. Normal expression of FLT3 has been mainly described on CD34+ hematopoietic progenitor stem cell populations, and FLT3-targeted therapies have potential to induce aplastic anemia. To address this question of hematologic toxicity of FLT3 CAR T cells, we created normal human hematopoiesis xenograft models in NOD scid gamma Il3-GM-SF (NSGS) mice engrafted with CD34+ cord blood cells for treatment with anti-AML CAR T cells. No difference in human granulocyte numbers was observed in marrows of engrafted mice treated with FLT3 CAR T cells, CD33 CAR T cells, or non-transduced T cells. A significant reduction in monocytes was observed in FLT3 CAR T cell-treated animals, however (p<0.05 by t test). To determine potential for increased hematologic toxicity in the presence of greater target antigen levels, we injected MOLM-14 into CD34+ cell-engrafted mice, then treated animals with control or anti-AML CAR T cells. We surprisingly found no decrement in defined hematopoietic stem cell (HSC) or granulocyte macrophage progenitor (GMP) populations, but did observe increased multipotent and common myeloid progenitor (MPP, CMP) cell numbers and an increase in total human cell engraftment 5 days after FLT3 CAR treatment in comparison to non-transduced T cell-treated animals. Relative to CD33 CAR T cells, FLT3 CAR T cells induced less toxicity to HSCs and MPPs and equivalent toxicity to CMPs and GMPs, indicating lower hematologic toxicity with FLT3 targeting. Conclusions: Taken together, these initial data demonstrate potent in vitro and in vivo anti-AML activity with limited hematopoietic toxicity of FLT3 CAR T cell immunotherapy. Future studies are focused on testing the effectiveness on other AML cell lines with varying expression of FLT3. Disclosures No relevant conflicts of interest to declare.
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Qin, Haiying, Lila Yang, John A. Chukinas, Nirali Shah, Samiksha Tarun, Marie Pouzolles, Christopher D. Chien, et al. "Systematic preclinical evaluation of CD33-directed chimeric antigen receptor T cell immunotherapy for acute myeloid leukemia defines optimized construct design." Journal for ImmunoTherapy of Cancer 9, no. 9 (September 2021): e003149. http://dx.doi.org/10.1136/jitc-2021-003149.
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BackgroundSuccessful development of chimeric antigen receptor (CAR) T cell immunotherapy for children and adults with relapsed/refractory acute myeloid leukemia (AML) is highly desired given their poor clinical prognosis and frequent inability to achieve cure with conventional chemotherapy. Initial experiences with CD19 CAR T cell immunotherapy for patients with B-cell malignancies highlighted the critical impact of intracellular costimulatory domain selection (CD28 vs 4-1BB (CD137)) on CAR T cell expansion and in vivo persistence that may impact clinical outcomes. However, the impact of costimulatory domains on the efficacy of myeloid antigen-directed CAR T cell immunotherapy remains unknown.MethodsIn this preclinical study, we developed six CAR constructs targeting CD33, a highly expressed and validated AML target, comprised of one of three single-chain variable fragments with CD3ζ and either CD28 or 4-1BB costimulatory domains. We systematically compared the preclinical in vitro and in vivo efficacy of T cells lentivirally transduced with CD33 CAR constructs (CD33CARTs) against human AML.ResultsWe observed potent in vitro cytokine production and cytotoxicity of CD33CARTs incubated with human CD33+ AML cell lines, as well as robust in vivo antileukemia activity in cell line and childhood AML patient-derived xenograft (PDX) models. Gemtuzumab-based CD33CARTs were unexpectedly toxic in vivo in animal models despite observed in vitro anti-leukemia activity. CD28-based CD33CARTs consistently induced more robust inhibition of leukemia proliferation in AML cell line and PDX models than did 4-1BB-based CD33CARTs. A ‘best-in-class’ lintuzumab-CD28/CD3ζ CAR construct was thus selected for clinical translation.ConclusionsCD33 is a critical antigen for potential immunotherapeutic targeting in patients with AML. Based on this rigorous preclinical evaluation, our validated clinical grade lintuzumab-CD28/CD3ζ CD33CART immunotherapy is now under evaluation in a first-in-child/first-in-human phase 1 clinical trial for children and adolescents/young adults with relapsed/refractory AML.Trial registration numberclinicaltrials.gov; NCT03971799.
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Mo, George, Hao-Wei Wang, Aimee C. Talleur, Shilpa A. Shahani, Bonnie Yates, Haneen Shalabi, Michael G. Douvas, et al. "Diagnostic approach to the evaluation of myeloid malignancies following CAR T-cell therapy in B-cell acute lymphoblastic leukemia." Journal for ImmunoTherapy of Cancer 8, no. 2 (November 2020): e001563. http://dx.doi.org/10.1136/jitc-2020-001563.
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Immunotherapeutic strategies targeting B-cell acute lymphoblastic leukemia (B-ALL) effectively induce remission; however, disease recurrence remains a challenge. Due to the potential for antigen loss, antigen diminution, lineage switch or development of a secondary or treatment-related malignancy, the phenotype and manifestation of subsequent leukemia may be elusive. We report on two patients with multiply relapsed/refractory B-ALL who, following chimeric antigen receptor T-cell therapy, developed myeloid malignancies. In the first case, a myeloid sarcoma developed in a patient with a history of myelodysplastic syndrome. In the second case, two distinct events occurred. The first event represented a donor-derived myelodysplastic syndrome with monosomy 7 in a patient with a prior hematopoietic stem cell transplantation. This patient went on to present with lineage switch of her original B-ALL to ambiguous lineage T/myeloid acute leukemia. With the rapidly evolving field of novel immunotherapeutic strategies, evaluation of relapse and/or subsequent neoplasms is becoming increasingly more complex. By virtue of these uniquely complex cases, we provide a framework for the evaluation of relapse or evolution of a subsequent malignancy following antigen-targeted immunotherapy.
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Nwanze, Chiadika, Sabina Kratzmeier, and Rosandra Kaplan. "Immunotherapy for solid tumors: Chimeric antigen receptor (CAR) T-cells and the tumor microenvironment." Journal of Immunology 204, no. 1_Supplement (May 1, 2020): 239.36. http://dx.doi.org/10.4049/jimmunol.204.supp.239.36.
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Abstract Patients with solid tumors continue to experience decreased survival rates, with metastasis being the leading cause of mortality in these patients. This has prompted the development of novel immunotherapies, with cellular immunotherapy giving rise to the development of Chimeric Antigen Receptor (CAR) T-cells, an approach combining specificity for tumor antigens and cytotoxicity independent of major histocompatibility complex molecule co-stimulation. This modality has proven efficacious in Acute Lymphoblastic Leukemia and B-cell Lymphoma. However, progress has remained limited in solid tumors despite targeting of various antigens, one of such being disialoganglioside (GD2) which has proven a viable target in neuroblastoma. This study investigates the efficacy of GD2 as an antigen target for CAR T-cell therapy in osteosarcoma. Surface flow cytometric analysis revealed the presence of GD2 on human 143B and Hu09H3 osteosarcoma cell lines. Of note, the tumor microenvironment offers an additional potential therapeutic target, as we observed increases in myeloid-derived suppressor cells alongside decreases in CD4+ T cells in primary tumor and metastatic tissues. In summary, we report the utility of GD2 as a target for cellular immunotherapy in osteosarcoma and the role of the tumor microenvironment in anti-tumor therapeutic modalities, a therapeutic combination that could be applicable to optimizing potent therapeutic modalities for solid tumors.
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Aureli, Anna, Beatrice Marziani, Tommaso Sconocchia, Maria Ilaria Del Principe, Elisa Buzzatti, Gianmario Pasqualone, Adriano Venditti, and Giuseppe Sconocchia. "Immunotherapy as a Turning Point in the Treatment of Acute Myeloid Leukemia." Cancers 13, no. 24 (December 13, 2021): 6246. http://dx.doi.org/10.3390/cancers13246246.
Full textAbstract:
Acute myeloid leukemia (AML) is a malignant disease of hematopoietic precursors at the earliest stage of maturation, resulting in a clonalproliferation of myoblasts replacing normal hematopoiesis. AML represents one of the most common types of leukemia, mostly affecting elderly patients. To date, standard chemotherapy protocols are only effective in patients at low risk of relapse and therapy-related mortality. The average 5-year overall survival (OS) is approximately 28%. Allogeneic hematopoietic stem cell transplantation (HSCT) improves prognosis but is limited by donor availability, a relatively young age of patients, and absence of significant comorbidities. Moreover, it is associated with significant morbidity and mortality. However, increasing understanding of AML immunobiology is leading to the development of innovative therapeutic strategies. Immunotherapy is considered an attractive strategy for controlling and eliminating the disease. It can be a real breakthrough in the treatment of leukemia, especially in patients who are not eligible forintensive chemotherapy. In this review, we focused on the progress of immunotherapy in the field of AML by discussing monoclonal antibodies (mAbs), immune checkpoint inhibitors, chimeric antigen receptor T cells (CAR-T cells), and vaccine therapeutic choices.
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Chitre, Sneha, Joop Gaken, Andrea Venuso, and Ghulam J. Mufti. "Single and Dual Targeting Chimeric Antigen Receptor T-Cell Therapy in Acute Myeloid Leukemia." Blood 136, Supplement 1 (November 5, 2020): 25. http://dx.doi.org/10.1182/blood-2020-142013.
Full textAbstract:
BACKGROUND: Discovery of chimeric antigen receptors (CARs) specific for tumour-associated antigens are emerging to be an effective form of immunotherapy for cancer treatment in recent years. However, the lack of a compelling acute myeloid leukemia (AML)-specific cell surface antigen and the safety concerns for myeloid-directed CAR T therapies causing prolonged myeloablation/aplasia which necessitate bone marrow transplantation make it challenging to develop CARTs for AML. In spite of these challenges, the high relapse rate of the disease i.e. 43% and 18% patients never attaining clinical remission (CR) with front-line induction treatments highlight an unmet need for developing improved CAR T vectors with enhanced specificity towards leukemic blasts in refractory/resistant cases with poor cytogenetics. Therefore, we aim to develop improved CAR T vectors for AML that produce safe and consistent responses in patients with high-risk disease. Herein, we demonstrate the in-vitro data for (a) efficacy of a second-generation CAR expressing six single-chain variable fragments (scFv) with different affinities for CD123 [interleukin three receptor alpha (IL3RA; CD123), a molecule over expressed on AML blasts and leukemic stem cells (LSC)] and (b) evaluate the cytotoxic effects of a dual targeting CARCD123/CD33 (against CD123 and CD33; an important myeloid marker specifically expressed on bulk AML disease) to enhance specificity towards leukemic cells therefore reducing "on-target off-organ effects". METHOD: Six lentiviral vectors with CAR against CD123 were constructed i.e. two high affinity (4nM kD & 4nM kD with a point mutation resulting in amino acid change K136Q), two moderate (56nM kD & 56nM kD with mutation at A105G) and two low affinity vectors (101nM kD & 101nM kD with mutation at V24G). To improve the specificity of the single targeting CARCD123, the high affinity (4nM kD, K136Q) vector was utilized to generate two dual targeting (CARCD123/CD33) constructs containing the activation domain (CD3ζ) directed against CD33 and the costimulatory domain (either CD28 or 41BB) directed against CD123. All constructs were transduced (MOI 1:5) into peripheral blood mononuclear cells (PBMCs) from healthy donors or AML patients and their cytotoxicity was examined by flowcytometry on leukemic cell lines; Kg1, U937, K562 [Fig:1a], Ramos wild type (CD19+, CD123-), artificially engineered Ramos cells (transduced by lentiviral vectors with CD123 and/or CD33 cDNA) i.e. Ramos 123 (CD123+), Ramos 33 (CD33+), Ramos 123/33 (CD123+ and CD33+) and AML mononucelar cells (MNCs). RESULTS: Flowcytometric analysis confirmed the expansion of T cells from PBMCs and cytotoxicity of the eight CAR constructs against target cells in in-vitro co-culture assay. High affinity CARCD123 (4nM kD & 4nM kD K136Q) T cells demonstrated enhanced cytotoxicity [Fig 1a] compared to moderate (56nM kD, 56nM kD A105G) CARCD123 in all leukemic cell lines while the low affinity (101nM kD, 101nM kD V24G) vectors had no effect. Efficacy of the high affinity CARCD123 constructs was validated on Kg1 [Fig 1b] and Ramos 123+ target cells by the increasing effector: target ratios (1:2, 1:4 & 1:10). Similar cytotoxic effects were also consistently observed against autologous AML MNCs (target cells) [n=4] and allogenic (effector cells are PBMCs from healthy donors) AML MNCs [Fig 1c] [n=3]. T cell activation was confirmed by ELISA and showed increased IFN-γ (500-2000 fold) and TNF-α (150-200 fold) levels after 24hr co-culture. Furthermore, we also elucidated the exclusive cytotoxicity of the two dual targeting CARCD123/CD33 in Ramos 123+/33+ cells [Fig 1d] with no effect being observed on Ramos 123+, Ramos 33+ and Ramos wild type (123-/33-) cell lines. This confirmed the absence of non-specific targeting, validated the improved specificity of the CARs towards leukemic cells and demonstrated a potential to reduce deleterious "on-target but off organ effects". CONCLUSION: In summary, we illustrate in-vitro data establishing the importance of scFv on CAR T cell cytotoxicity and exemplify for the first time an improved specificity of CARTs by targeting two antigens simultaneously in AML. Future work will involve examining the in-vivo dynamics of CAR CD123 and CAR CD123 CD33 on the hematopoietic system and on disease pathogenesis with an aim to proceed to phase I clinical trial. Figure 1 Disclosures Mufti: BMS, Novartis: Research Funding; Abbvie, Novartis: Consultancy.
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Winer, Eric S., and Richard M. Stone. "Novel therapy in Acute myeloid leukemia (AML): moving toward targeted approaches." Therapeutic Advances in Hematology 10 (January 2019): 204062071986064. http://dx.doi.org/10.1177/2040620719860645.
Full textAbstract:
Acute myeloid leukemia (AML) is a heterogenous and complex disease characterized by rapid cellular proliferation, an aggressive clinical course, and generally high mortality. While progress has been made in the understanding of the genetic and molecular biology of the disease, the standard of care for patients had only changed minimally over the past 40 years. Recently, rapid movement of potentially useful agents from bench to bedside has translated into new therapies either recently approved or in clinical trials. These therapies include improved chemotherapies, mutationally targeted inhibitors, pro-apoptotic agents, microenvironment targeting molecules, cell cycle checkpoint inhibitors, and epigenetic regulators. Furthermore, advances in immunotherapy employ monoclonal and bispecific antibodies, chimeric antigen receptor (CAR) T cells, checkpoint inhibitors, and vaccines provide an alternative pathway for AML treatment. In this review, we discuss the recent results of completed or ongoing clinical trials with these novel therapeutic agents in AML.
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Gurney, Mark, and Michael O’Dwyer. "Realizing Innate Potential: CAR-NK Cell Therapies for Acute Myeloid Leukemia." Cancers 13, no. 7 (March 29, 2021): 1568. http://dx.doi.org/10.3390/cancers13071568.
Full textAbstract:
Next-generation cellular immunotherapies seek to improve the safety and efficacy of approved CD19 chimeric antigen receptor (CAR) T-cell products or apply their principles across a growing list of targets and diseases. Supported by promising early clinical experiences, CAR modified natural killer (CAR-NK) cell therapies represent a complementary and potentially off-the-shelf, allogeneic solution. While acute myeloid leukemia (AML) represents an intuitive disease in which to investigate CAR based immunotherapies, key biological differences to B-cell malignancies have complicated progress to date. As CAR-T cell trials treating AML are growing in number, several CAR-NK cell approaches are also in development. In this review we explore why CAR-NK cell therapies may be particularly suited to the treatment of AML. First, we examine the established role NK cells play in AML biology and the existing anti-leukemic activity of NK cell adoptive transfer. Next, we appraise potential AML target antigens and consider common and unique challenges posed relative to treating B-cell malignancies. We summarize the current landscape of CAR-NK development in AML, and potential targets to augment CAR-NK cell therapies pharmacologically and through genetic engineering. Finally, we consider the broader landscape of competing immunotherapeutic approaches to AML treatment. In doing so we evaluate the innate potential, status and remaining barriers for CAR-NK based AML immunotherapy.
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Roex, Gils, Tom Feys, Yves Beguin, Tessa Kerre, Xavier Poiré, Philippe Lewalle, Peter Vandenberghe, Dominique Bron, and Sébastien Anguille. "Chimeric Antigen Receptor-T-Cell Therapy for B-Cell Hematological Malignancies: An Update of the Pivotal Clinical Trial Data." Pharmaceutics 12, no. 2 (February 24, 2020): 194. http://dx.doi.org/10.3390/pharmaceutics12020194.
Full textAbstract:
Chimeric antigen receptor (CAR)-T-cell therapy is an innovative form of adoptive cell therapy that has revolutionized the treatment of certain hematological malignancies, including B-cell non-Hodgkin lymphoma (NHL) and B-cell acute lymphoblastic leukemia (ALL). The treatment is currently also being studied in other B-cell neoplasms, including multiple myeloma (MM) and chronic lymphocytic leukemia (CLL). CD19 and B-cell maturation antigen (BCMA) have been the most popular target antigens for CAR-T-cell immunotherapy of these malignancies. This review will discuss the efficacy and toxicity data from the pivotal clinical studies of CD19- and BCMA-targeted CAR-T-cell therapies in relapsed/refractory B-cell malignancies (NHL, ALL, CLL) and MM, respectively.
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Tang, Thao T., Lindsey F. Call, Sommer Castro, Cynthia Nourigat-Mckay, LaKeisha Perkins, Laura Pardo, Amanda R. Leonti, Soheil Meshinchi, and Quy Le. "Therapeutic Targeting of Mesothelin with Chimeric Antigen Receptor Natural Killer Cell Therapy in Acute Myeloid Leukemia." Blood 138, Supplement 1 (November 5, 2021): 1712. http://dx.doi.org/10.1182/blood-2021-153156.
Full textAbstract:
Abstract Effective immunotherapy for acute myeloid leukemia (AML) has been limited by the lack of AML-restricted targets (expression in AML but silent in normal hematopoiesis). Current immunotherapies targeting lineage markers CD33 and CD123 (if effective) would lead to prolonged myelosuppression or myeloablation, requiring stem cell transplantation to restore hematopoiesis after treatment. In search for AML-restricted targets, we interrogated the AML transcriptome of nearly 3000 cases in pediatric and young adults and contrasting it to normal bone marrow and peripheral blood CD34+ samples. This extensive discovery effort has identified over 200 AML-restricted targets with mesothelin (MSLN) emerged as one of the highest expressing AML-restricted targets and highly enriched in the KMT2A-rearranged AML subtype. We previously validated cell surface expression of MSLN on both AML blasts and leukemic stem cells (Le et. al. 2021). We further showed efficacy targeting MSLN in AML using antibody-drug conjugates (Kaeding et. al. 2021) and chimeric antigen receptor (CAR) T cells (Le et. al. 2021). Given that natural killer (NK) cells are potent immune effector cells and generally have a more favorable toxicity profile than CAR T cells (i.e without cytokine release syndrome), we developed CAR NK cells targeting MSLN and evaluated their efficacy in AML preclinical models. From primary patient samples, we verified MSLN cell surface expression and showed high correlation between cell surface expression (mean fluorescence intensity, MFI) and transcript expression (TPM, R = 0.72, p = 2.1x10 -8, Figure 1A) . Importantly, MSLN expression was restricted to AML blasts and entirely absent in normal lymphocytes and myeloid cells in individual patients (Figure 1B, C). Having confirmed cell surface and AML-restricted expression of MSLN, we developed CAR NK cells targeting MSLN. The VH and VL sequences from immunotoxin SS1P were used to create the single-chain variable fragment domain of the standard CAR (41-BB and CD3Zeta, Figure 1D). CAR NK cells were generated by transducing NK-92 cells with the MSLN CAR construct. Cytotoxicity of CAR NK cells was evaluated against Nomo-1 AML cell line, which expresses endogenous level of MSLN; MV4;11 and Kasumi-1 cell lines engineered to express MSLN with a lentivirus construct (MV4;11 MSLN+ and Kasumi-1 MSLN+). We initially tested the target specificity of MSLN-directed CAR NK cells against MSLN-positive (Nomo-1, MV4;11 MSLN+ and Kasumi-1 MSLN+) and MSLN-negative (Nomo-1 MSLN KO, MV4;11 and Kasumi-1) cells. CAR NK cells exhibited enhanced cytolytic activity against MSLN-positive but not MSLN-negative AML cells after 12 hours of co-incubation at the indicated effector: target (E:T) ratios (Figure 1E). We next assessed the in vivo efficacy of CAR NK cells. Nomo-1 cells transduced with GFP/Luciferase were transplanted into NSG mice at 1x10 6 cells/mouse. Unmodified or CAR NK cells were infused 1 week following leukemic cell injection at 1x10 7 cells/mouse. Monitoring leukemia burden by bioluminescence IVIS imaging showed that after 4 days post NK injection, the leukemia was significantly reduced in Nomo-1 xenograft mice treated with CAR NK cells compared to mice that received unmodified NK cells (Figure 1F), suggesting highly potent anti-leukemia activity of CAR NK cells. In this study, we demonstrate that the cell surface expression of MSLN is restricted to AML blasts but is entirely silent in normal hematopoietic subsets in individual patients. Previous and current clinical trials utilizing NK-92 cells have demonstrated safety and efficacy in variety of cancers, including AML. Here, we demonstrate that NK-92 cells genetically modified with a CAR to redirect their specificity against MSLN-positive AML cells exhibit potent, target-dependent anti-leukemia activity in vitro and in vivo. These results provide compelling data to evaluate MSLN-directed CAR NK cell therapy in clinical trials for refractory/relapsed AML, especially for high-risk KMT2A-rearranged leukemia where majority of patients express MSLN at diagnosis and relapse. Figure 1 Figure 1. Disclosures Pardo: Hematologics, Inc.: Current Employment.
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Schultz, Liora, and Rebecca Gardner. "Mechanisms of and approaches to overcoming resistance to immunotherapy." Hematology 2019, no. 1 (December 6, 2019): 226–32. http://dx.doi.org/10.1182/hematology.2019000018.
Full textAbstract:
Abstract Immunotherapies have been successfully developed for the treatment of B-cell acute lymphoblastic leukemia (B-ALL) with FDA approval of blinatumomab, inotuzumab, and tisagenlecleucel for relapsed or refractory patients. These agents target either CD19 or CD22, which are both expressed on the surface of the leukemic blasts in the majority of patients. The use of these agents has greatly transformed the landscape of available treatment, and it has provided curative therapy in some patients. As the field has matured, we are learning that for most patients, the currently available immunotherapies are not curative. Leukemic resistance to both CD19 and CD22 pressure has been described and is a major component of developed resistance to these therapies. Patients with B-ALL have developed CD19- or CD22-negative B-ALL, and in more rare cases, they have undergone lineage switch to acute myeloid leukemia. Current efforts are focusing on overcoming antigen escape, either by forced antigen expression or by dual-targeting therapies. A functional immune system is also required for maximal benefit of immunotherapy, particularly with chimeric antigen receptor (CAR) T-cell therapies. Data are now being produced that may allow for the prospective identification of patients whose immune deficits may be identified up front and predict failure. Preclinical work is focusing on additional engineering of CAR T cells to overcome these inherent immune deficits. Last, with improved knowledge of which patients are likely to benefit from immunotherapy as definitive treatment, those patients who are predicted to develop resistance may be prospectively recommended to undergo a consolidative hematopoietic cell transplant to lessen the recurrence risk.
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Song, Degang, Michael H. Swartz, Steve G. Biesecker, Fernando Borda, Rutul R. Shah, Peter Emtage, William G. Wierda, Laurence J. N. Cooper, and Tim Chan. "Chimeric Antigen Receptor-Modified T Cells for the Treatment of Acute Myeloid Leukemia Expressing CD33." Blood 128, no. 22 (December 2, 2016): 4058. http://dx.doi.org/10.1182/blood.v128.22.4058.4058.
Full textAbstract:
Abstract Relapsed acute myeloid leukemia (AML) is an aggressive disease with very poor outcomes. Redirection of T-cell specificity via chimeric antigen receptor (CAR) has shown promising anti-tumor activity in clinical trials, particularly for B cell linage malignancies. CD33 is a transmembrane protein expressed on normal and malignant myeloid-derived cells as well (as on subsets of activated T cells and NK cells). Since this protein is commonly expressed on AML cells, we sought to evaluate the efficacy of targeting AML with CD33-specific CAR-T cells. We generated a lentiviral construct to co-express CD33-specific CAR and a kill switch based on a tag derived from the epidermal growth factor receptor. The latter allows for the conditional elimination of CAR-T cells in vivo. Following transduction of primary T cells, we confirmed CAR and kill switch co-expression by flow cytometry and western blot analyses. Elimination of genetically modified T cells was demonstrated using the clinically-available antibody, cetuximab. CD33 CAR-T cells demonstrated specific cytotoxicity to CD33+ target cell lines. CD33 CAR-T cells were also activated to produce IFNg, TNF, and IL-2 cytokines in response to CD33+ target cells. Furthermore, adoptive transfer of CD33 CAR-T in immunocompromised (NSG) mice bearing established CD33+(CD19neg) AML (MOLM-13) tumor resulted in reduction of tumor burden and improvement of overall survival, compared to control mice receiving CD19 CAR-T cells or no immunotherapy (Figure). Sampling of blood demonstrated the persistence of the CD33 CAR-T cells with no detection of AML (MOLM-13) tumor cells. These pre-clinical data demonstrate the effectiveness of CD33 CAR-T cells in targeting CD33+ AML tumor cells and provide a rationale for future clinical evaluation in AML patients with unmet medical need. Disclosures Song: Intrexon Corporation: Employment, Equity Ownership. Swartz:Intrexon Corporation: Employment, Equity Ownership. Biesecker:Intrexon Corporation: Employment, Equity Ownership. Borda:Intrexon Corporation: Employment. Shah:Intrexon Corporation: Employment, Equity Ownership. Wierda:Genentech: Research Funding; Gilead: Research Funding; Abbvie: Research Funding; Novartis: Research Funding; Acerta: Research Funding. Cooper:MD Anderson Cancer Center: Employment; Intrexon: Equity Ownership; Sangamo BioSciences: Patents & Royalties; Targazyme,Inc.,: Equity Ownership; City of Hope: Patents & Royalties; ZIOPHARM Oncology: Employment, Equity Ownership, Patents & Royalties; Miltenyi Biotec: Honoraria; Immatics: Equity Ownership. Chan:Intrexon Corporation: Employment, Equity Ownership.
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Kittel-Boselli, Enrico, Karla Elizabeth González Soto, Liliana Rodrigues Loureiro, Anja Hoffmann, Ralf Bergmann, Claudia Arndt, Stefanie Koristka, et al. "Targeting Acute Myeloid Leukemia Using the RevCAR Platform: A Programmable, Switchable and Combinatorial Strategy." Cancers 13, no. 19 (September 24, 2021): 4785. http://dx.doi.org/10.3390/cancers13194785.
Full textAbstract:
Clinical translation of novel immunotherapeutic strategies such as chimeric antigen receptor (CAR) T-cells in acute myeloid leukemia (AML) is still at an early stage. Major challenges include immune escape and disease relapse demanding for further improvements in CAR design. To overcome such hurdles, we have invented the switchable, flexible and programmable adaptor Reverse (Rev) CAR platform. This consists of T-cells engineered with RevCARs that are primarily inactive as they express an extracellular short peptide epitope incapable of recognizing surface antigens. RevCAR T-cells can be redirected to tumor antigens and controlled by bispecific antibodies cross-linking RevCAR T- and tumor cells resulting in tumor lysis. Remarkably, the RevCAR platform enables combinatorial tumor targeting following Boolean logic gates. We herein show for the first time the applicability of the RevCAR platform to target myeloid malignancies like AML. Applying in vitro and in vivo models, we have proven that AML cell lines as well as patient-derived AML blasts were efficiently killed by redirected RevCAR T-cells targeting CD33 and CD123 in a flexible manner. Furthermore, by targeting both antigens, a Boolean AND gate logic targeting could be achieved using the RevCAR platform. These accomplishments pave the way towards an improved and personalized immunotherapy for AML patients.
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Galetto, Roman, Celine Lebuhotel, Agnes Gouble, Cecile Schiffer-Mannioui, and Julianne Smith. "Allogenic T-Cells Targeting CD123 for Adoptive Immunotherapy of Acute Myeloid Leukemia (AML)." Blood 124, no. 21 (December 6, 2014): 1116. http://dx.doi.org/10.1182/blood.v124.21.1116.1116.
Full textAbstract:
Abstract Adoptive immunotherapy using autologous T-cells endowed with chimeric antigen receptors (CARs) has emerged as a promising new approach to treating cancer. However, a limitation of this approach is that CAR T-cells must be generated on a bespoke basis. To overcome this limitation, we have developed an allogeneic based platform for large scale production of “off-the-shelf” CAR T-cells from unrelated 3rd party donors. This platform utilizes Transcription Activator-Like Effector Nuclease (TALENTM) gene editing technology to inactivate the TCRα constant (TRAC) gene, eliminating the potential for T-cells bearing alloreactive TCR’s to mediate Graft versus Host Disease (GvHD). We have previously demonstrated that editing of the TRAC gene can be achieved at high frequencies, obtaining up to 80% of TCRαβ negative cells. This allows us to efficiently produce TCR-deficient T-cells that have been shown to no longer mediate alloreactivity in a xeno-GvHD mouse model. Furthermore, the capacity to perform efficient multiplex genome editing using TALENTM offers the possibility of simultaneously rendering cells resistant to standard chemotherapy or to tumor evasion mechanisms. In this work we present the adaptation of this allogeneic platform to the production of T cells targeting CD123, the transmembrane alpha chain of the interleukin-3 receptor, which is expressed in tumor cells from the majority of patients with Acute Myeloid Leukemia (AML). In a first step, we have screened human primary T-cells harboring CARs with different antigen recognition domains in the context of multiple CAR architectures in order to identify candidates displaying specific activity against cell lines expressing variable levels of the CD123 antigen. To provide proof of concept for the general applicability of the allogeneic approach we have manufactured a TCR-deficient CD123 CAR T-cell (UCART123) and demonstrated that this product maintains a potent anti-tumoral activity in vitro. Experiments in an orthotopic AML mouse model using UCART123 cells are currently ongoing, in order to establish the absence of alloreactivity and the anti-tumoral activity in vivo. The ability to carry out large scale manufacturing of allogeneic, non alloreactive CD123 specific T Cells from a single healthy donor could thus offer the possibility of an off-the-shelf treatment that would be immediately available for administration to a large number of AML patients. Disclosures Galetto: Cellectis SA: Employment. Lebuhotel:Cellectis SA: Employment. Gouble:Cellectis SA: Employment. Schiffer-Mannioui:Cellectis SA: Employment. Smith:Cellectis SA: Employment.
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Winters, Amanda, and Lia Gore. "Moving immunotherapy into the front line in ALL." Hematology 2019, no. 1 (December 6, 2019): 209–17. http://dx.doi.org/10.1182/hematology.2019000017.
Full textAbstract:
Abstract Although almost 90% of children with acute lymphoblastic leukemia (ALL) and ∼60% of children with acute myeloid leukemia are cured with frontline therapy, relapse and chemotherapy resistance are significant challenges that contribute to morbidity and mortality. Even with long-term survival, the acute and chronic burdens of therapy are major issues for patients and families. Long-term side effects occur, including cardiac, endocrinologic, neurcognitive, orthopedic, and psychosocial problems, and healthy survivorship is frequently compromised. With goals of minimizing relapse and/or decreasing traditional chemotherapy-associated toxicities, exploration of immunotherapeutic strategies has moved to the forefront in pediatric cancer. New immunotherapy approaches provide a major paradigm shift in oncology overall, often curing previously incurable patients. The past several years have yielded successful uses across a variety of malignancies, and enthusiasm continues to rise for applying these therapies more broadly. Herein we discuss current approaches incorporating the bispecific T-cell engager blinatumomab, the antibody-drug conjugate inotuzumab ozogamicin (InO), and CD19-directed chimeric antigen receptor T cells in children with relapsed/refractory B-cell ALL and discuss the potential for using these immunotherapies in the treatment of newly diagnosed children.
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Tettamanti, Sarah, Virna Marin, Irene Pizzitola, Chiara Francesca Magnani, Greta Maria Paola Giordano Attianese, Elisabetta Cribioli, Angel Lopez, Andrea Biondi, Dominique Bonnet, and Ettore Biagi. "Targeting of Acute Myeloid Leukemia by Cytokine-Induced Killer Cells Redirected with a Novel CD123-Specific Chimeric Antigen Receptor." Blood 120, no. 21 (November 16, 2012): 3010. http://dx.doi.org/10.1182/blood.v120.21.3010.3010.
Full textAbstract:
Abstract Abstract 3010 Current therapeutic regimens for Acute Myeloid Leukemia (AML) are still associated with high rates of relapse. In the last years, great interest has been focused on the identification of surface molecules that are preferentially expressed by AML cells and leukemic stem cells (LSCs), in order to selectively target the tumor population, whilst sparing the normal counterpart of hematopoietic stem/progenitor cells (HSPC), and possibly impeding disease recurrence. Immunotherapy with T-cells genetically modified to express chimeric antigen receptors (CARs) represents a valid and innovative cell therapy approach for hematological malignancies. In this study we developed a new CAR molecule specific for the IL-3Rα (CD123) target antigen, which is overexpressed on AML blasts, CD34+ leukemic progenitors, and leukemic stem cells (AML-LSCs) compared to normal hematopoietic stem/progenitor cells (HSPCs), and whose overexpression is associated with poor prognosis. Cytokine Induced Killer (CIK) cells, ex-vivo expanded T cells with spontaneous antitumoral activity, were transduced with an SFG-retroviral vector encoding an anti-CD123.CAR and CAR functionality has been evaluated by short-term cytotoxicity assay. Transduced CIK cells strongly killed CD123+ THP-1 cell line (60%±5.4%, Effector:Target –E:T- ratio of 5:1, n=3), as well as primary AML blasts (59%±5.4%, E:T ratio of 3:1, n=4). With the aim to better characterize the ability of anti-CD123.CAR+CIK cells to kill leukaemia cells over time we performed long-term cytotoxicity assay, observing a leukemic cell recovery for THP-1 of 3.5%±1.5% (n=5) and for primary AML cells of 2.4%±1.4% (n=3) when co-cultured with CIK cells expressing anti-CD123.CAR, compared to an average target survival of up to 80%, when co-cultured with unmanipulated (NT) CIK cells. Interestingly, secondary colonies experiments after co-culture of healthy donor cord blood-derived HSPCs (Lin-) with anti-CD123.CAR+CIK cells demonstrated that this newly generated CAR molecule better preserved the normal haematopoietic reconstitution in contrast to a previously generated anti-CD33.CAR (total number of colonies of 146.8±6.6, 66.4±5.1, 117.6±4.6, for Lin- cells co-cultured with NT CIK cells, anti-CD33.CAR+CIK cells, anti-CD123.CAR+CIK cells respectively, n=4), while keeping identical cytotoxicity profile towards AML. Furthermore, a limited killing of normal CD123 expressing monocytes and CD123-low expressing endothelial cells was measured, accompanied by a lesser release of stimulatory cytokines such as IFN-gamma, TNF-alfa and TNF-beta when compared to the levels released after stimulation with CD123+ leukemic cells (THP-1 and AML), thus indicating a low toxicity profile of the anti-CD123.CAR. Taken together, our results indicate that CD123-specific CAR strongly enhances anti-leukemic CIK functions towards AML, while sparing HSPCs and normal CD123-expressing tissues, paving the way for the development of novel immunotherapy approaches for the treatment of resistant forms of AML, particularly for a precocious intervention in presence of minimal residual disease, in the context of early relapse after HSCT. Disclosures: No relevant conflicts of interest to declare.
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Shimizu, Kanako, Tomonori Iyoda, Satoru Yamasaki, Norimitsu Kadowaki, Arinobu Tojo, and Shin-ichiro Fujii. "NK and NKT Cell-Mediated Immune Surveillance against Hematological Malignancies." Cancers 12, no. 4 (March 28, 2020): 817. http://dx.doi.org/10.3390/cancers12040817.
Full textAbstract:
Recent cancer treatment modalities have been intensively focused on immunotherapy. The success of chimeric antigen receptor T cell therapy for treatment of refractory B cell acute lymphoblastic leukemia has pushed forward research on hematological malignancies. Among the effector types of innate lymphocytes, natural killer (NK) cells show great importance in immune surveillance against infectious and tumor diseases. Particularly, the role of NK cells has been argued in either elimination of target tumor cells or escape of tumor cells from immune surveillance. Therefore, an NK cell activation approach has been explored. Recent findings demonstrate that invariant natural killer T (iNKT) cells capable of producing IFN-γ when optimally activated can promptly trigger NK cells. Here, we review the role of NKT and/or NK cells and their interaction in anti-tumor responses by highlighting how innate immune cells recognize tumors, exert effector functions, and amplify adaptive immune responses. In addition, we discuss these innate lymphocytes in hematological disorders, particularly multiple myeloma and acute myeloid leukemia. The immune balance at different stages of both diseases is explored in light of disease progression. Various types of innate immunity-mediated therapeutic approaches, recent advances in clinical immunotherapies, and iNKT-mediated cancer immunotherapy as next-generation immunotherapy are then discussed.
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Sauer, Tim, Kathan Parikh, Sandhya Sharma, Bilal Omer, David Sedloev, Qian Chen, Linus Angenendt, et al. "CD70-specific CAR T cells have potent activity against acute myeloid leukemia without HSC toxicity." Blood 138, no. 4 (March 15, 2021): 318–30. http://dx.doi.org/10.1182/blood.2020008221.
Full textAbstract:
Abstract The prognosis of patients with acute myeloid leukemia (AML) remains dismal, highlighting the need for novel innovative treatment strategies. The application of chimeric antigen receptor (CAR) T-cell therapy to patients with AML has been limited, in particular by the lack of a tumor-specific target antigen. CD70 is a promising antigen to target AML, as it is expressed on most leukemic blasts, whereas little or no expression is detectable in normal bone marrow samples. To target CD70 on AML cells, we generated a panel of CD70-CAR T cells that contained a common single-chain variable fragment (scFv) for antigen detection, but differed in size and flexibility of the extracellular spacer and in the transmembrane and the costimulatory domains. These CD70scFv CAR T cells were compared with a CAR construct that contained human CD27, the ligand of CD70 fused to the CD3ζ chain (CD27z). The structural composition of the CAR strongly influenced expression levels, viability, expansion, and cytotoxic capacities of CD70scFv-based CAR T cells, but CD27z-CAR T cells demonstrated superior proliferation and antitumor activity in vitro and in vivo, compared with all CD70scFv-CAR T cells. Although CD70-CAR T cells recognized activated virus-specific T cells (VSTs) that expressed CD70, they did not prevent colony formation by normal hematopoietic stem cells. Thus, CD70-targeted immunotherapy is a promising new treatment strategy for patients with CD70-positive AML that does not affect normal hematopoiesis but will require monitoring of virus-specific T-cell responses.
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Tasian, Sarah K., Saad S. Kenderian, Feng Shen, Yong Li, Marco Ruella, William C. Fix, Miroslaw Kozlowski, et al. "Efficient Termination of CD123-Redirected Chimeric Antigen Receptor T Cells for Acute Myeloid Leukemia to Mitigate Toxicity." Blood 126, no. 23 (December 3, 2015): 565. http://dx.doi.org/10.1182/blood.v126.23.565.565.
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Abstract Background. Many children and adults with acute myeloid leukemia (AML) relapse or are incurable with current treatment modalities, highlighting a need for alternative therapies. Chimeric antigen receptor (CAR) T cells targeting CD123 have demonstrated potent anti-leukemia activity in murine xenograft models of human AML. However, CD123-redirected T cell treatment of mice engrafted with normal human hematopoietic cells resulted in profound myeloablation, raising concerns for hematologic toxicity in patients with AML who may be treated with such therapies. We hypothesized that T cell deletion after CD123-redirected T cell-induced eradication of AML could minimize this bystander toxicity without impairing leukemia control, thereby increasing the therapeutic window of anti-AML CAR T cell immunotherapy. Methods. To test this hypothesis, we compared three termination strategies in human AML xenograft models: (1) T cell ablation with the anti-CD52 antibody alemtuzumab after treatment with T cells lentivirally-transduced with anti-CD123-41BB-CD3ζ (CART123), (2) T cell ablation with the anti-CD20 antibody rituximab after treatment with CART123 engineered to co-express CD20 (CART123/CD20), and (3) treatment with "biodegradable" anti-CD123 mRNA-electroporated CAR T cells (RNA-CART123). Mice engrafted with luciferase-expressing human AML cell lines (MOLM14, MOLM13, U937) or primary AML specimens (n=3) were treated with CD123-redirected CAR T cells as above. For T cell depletion studies, alemtuzumab 1 or 5 mg/kg was injected intraperitoneally (IP) at 1-4 weeks after 1x105-106 CART123 to determine optimal dosing and timing of T cell ablation. In subsequent studies, rituximab 10 mg/kg was injected IP 4 weeks after 1x105 -106 CART123/CD20, or 1x107 RNA-CART123 were injected intravenously at 5, 9, and 16 days after AML engraftment. Mice were followed by weekly bioluminescent imaging and/or quantitative flow cytometry analyses of blood, spleen, and/or bone marrow. Results. CART123 treatment of CD123+ AML xenografts induced marked T cell expansion and leukemia eradication in vivo, resulting in long-term animal survival (p<0.0001 vs untransduced T cell-treated controls). Minimal xenogeneic graft-versus-host effects were observed. One dose of alemtuzumab rapidly eliminated T cells in all tested models with best efficacy of 5 mg/kg dosing at 4 weeks post-CART123. CART123/CD20 inhibited AML proliferation with similar kinetics to those of CART123, and 1 dose of rituximab at 4 weeks post-CART123/CD20 infusion rapidly eliminated T cells while preserving leukemia remission. Alemtuzumab or rituximab alone did not inhibit AML proliferation in non-CART123-treated xenograft models vs AML-only controls (p=1.00). Mice with CART123- or CART123/CD20-induced AML remission at time of T cell ablation remained leukemia-free for ≥12 weeks, and animal survival did not differ from that of CD123-redirected CAR T cell-treated mice that did not undergo T cell depletion (p=1.00). In contrast, early depletion of T cells with alemtuzumab at 1, 2, or 3 weeks post-CART123 resulted in failure to eliminate AML with subsequent leukemia progression and animal death. Furthermore, AML rechallenge of animals with alemtuzumab- or rituximab-ablated T cells resulted in rapid leukemia proliferation without T cell re-expansion, confirming completeness of T cell depletion. Non-ablated mice demonstrated CAR T cell re-expansion with rejection of CD123+ leukemia rechallenge (p<0.0001). RNA-CART123 rapidly eliminated AML and facilitated long-term animal survival, although RNA-CART123 had expectedly shorter persistence in vivo than did CART123 or CART123/CD20. Conclusions. Alemtuzumab and rituximab completely eliminated CD123-redirected CAR T cells in human AML xenograft models. Sustained leukemia remission required CART123 or CART123/CD20 persistence for 4 weeks prior to T cell termination via alemtuzumab or rituximab post-CART123 or CART123/CD20, respectively. Ongoing studies are investigating efficacy of T cell elimination in additional xenograft models and against other anti-AML CAR T cell immunotherapies. Results from these studies may help to identify promising T cell termination strategies that will augment efficacy of CAR T cell therapy in patients with AML, particularly prior to stem cell transplantation. RNA-CART123 trials in patients with relapsed/refractory AML will open soon. Disclosures Off Label Use: depletion of CAR T cells with alemtuzumab or rituximab. Kenderian:Novartis: Patents & Royalties, Research Funding. Ruella:Novartis: Patents & Royalties, Research Funding. Aplenc:Sigma Tau: Consultancy. June:Novartis: Patents & Royalties, Research Funding. Grupp:Novartis: Consultancy, Research Funding. Gill:Novartis: Patents & Royalties, Research Funding.
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Luo, Yi, Lung-Ji Chang, Yongxian Hu, Lujia Dong, Guoqing Wei, and He Huang. "First-in-Man CD123-Specific Chimeric Antigen Receptor-Modified T Cells for the Treatment of Refractory Acute Myeloid Leukemia." Blood 126, no. 23 (December 3, 2015): 3778. http://dx.doi.org/10.1182/blood.v126.23.3778.3778.
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Abstract Introduction: Adoptive immunotherapy using T-cells endowed with chimeric antigen receptors (CARs) has emerged as a promising new approach to treating CD19+ acute lymphoblastic leukemia (ALL). However,treatments for relapse/refractory acute myeloid leukemia (AML) have remained largely unchanged for nearly 50 years, and some AML patients have very poor prognosis. The interleukin-3 receptor alpha chain (CD123) has been identified as a potential immunotherapeutic target due to its overexpression in AML cells compared with normal hematopoietic stem cells. Antibodies targeting CD123 for the treatment of AML have demonstrated promising anti-leukemic activity in murine models but showed limited efficacy in clinical trials, suggesting that alternative and more potent therapies targeting CD123 are required. Methods: We have generated a 4th generation, apoptosis-inducible lentiviral CAR targeting CD123: CD123-scFv/CD28/CD137/CD27/CD3ζ-iCasp9 (4SCAR123), and demonstrated its high AML killing and AP1903-inducible apoptosis functions in ex vivo analyses. In a pilot trial of 4SCAR123, we enrolled a 47-year-old male patient with AML-M2 (FLT3/ITD+). The patient underwent allogeneic hematopoietic stem cell transplantation and relapsed. After 3 chemotherapies combined with sorafenib, his AML cells kept at 59% in bone marrow. He received CTX 250mg/kg/day for 3 days as conditioning regimen followed by 1.8x106/kg 4SCAR123 T cell infusion. Serum cytokine levels were measured by flow cytometric bead assay. Results: At day 1 after 4SCAR123 T infusion, the patient experienced rigorous chills and fevers, low blood pressure and hypoxemia. We detected elevated serum cytokine levels including interleukin-6 (2,500pg/ml) and tumor necrosis factor-α (33.8 pg/ml) at day 8, and the patient suffered from severe cytokine release syndrome (CRS) on day 4, which was controlled by one dose of Tocilizumab. BM examination detected a decrease of blasts from 59% to 45% 20 days after CAR-T therapy. Conclusion: Here we report a first-in-man pilot safety study of CD123 CAR-T therapy for AML patients. The 4SCAR123 exhibited potent cytotoxicity against AML in vitro, and in this pilot trial, the patient developed a rapid response consistent with CRS and achieved partial remission within 20 days. Importantly, although CD123 is universally expressed in myeloid and endothelial cells, we did not observe overt off-target cytotoxicity from the 4SCAR123 T cells, except for a controllable CRS. Our pilot study warrants further exploration of CD123 CAR for the management of refractory AML. Disclosures Dong: America Yuva Biomed: Consultancy.
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Abadir, Edward, Robin E. Gasiorowski, Pablo A. Silveira, Stephen Larsen, and Georgina J. Clark. "Is Hematopoietic Stem Cell Transplantation Required to Unleash the Full Potential of Immunotherapy in Acute Myeloid Leukemia?" Journal of Clinical Medicine 9, no. 2 (February 18, 2020): 554. http://dx.doi.org/10.3390/jcm9020554.
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From monoclonal antibodies (mAbs) to Chimeric Antigen Receptor (CAR) T cells, immunotherapies have enhanced the efficacy of treatments against B cell malignancies. The same has not been true for Acute Myeloid Leukemia (AML). Hematologic toxicity has limited the potential of modern immunotherapies for AML at preclinical and clinical levels. Gemtuzumab Ozogamicin has demonstrated hematologic toxicity, but the challenge of preserving normal hematopoiesis has become more apparent with the development of increasingly potent immunotherapies. To date, no single surface molecule has been identified that is able to differentiate AML from Hematopoietic Stem and Progenitor Cells (HSPC). Attempts have been made to spare hematopoiesis by targeting molecules expressed only on later myeloid progenitors as well as AML or using toxins that selectively kill AML over HSPC. Other strategies include targeting aberrantly expressed lymphoid molecules or only targeting monocyte-associated proteins in AML with monocytic differentiation. Recently, some groups have accepted that stem cell transplantation is required to access potent AML immunotherapy and envision it as a rescue to avoid severe hematologic toxicity. Whether it will ever be possible to differentiate AML from HSPC using surface molecules is unclear. Unless true specific AML surface targets are discovered, stem cell transplantation could be required to harness the true potential of immunotherapy in AML.
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Hattori, Norimichi, and Tsuyoshi Nakamaki. "Natural Killer Immunotherapy for Minimal Residual Disease Eradication Following Allogeneic Hematopoietic Stem Cell Transplantation in Acute Myeloid Leukemia." International Journal of Molecular Sciences 20, no. 9 (April 26, 2019): 2057. http://dx.doi.org/10.3390/ijms20092057.
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The most common cause of death in patients with acute myeloid leukemia (AML) who receive allogeneic hematopoietic stem cell transplantation (allo-HSCT) is AML relapse. Therefore, additive therapies post allo-HSCT have significant potential to prevent relapse. Natural killer (NK)-cell-based immunotherapies can be incorporated into the therapeutic armamentarium for the eradication of AML cells post allo-HSCT. In recent studies, NK cell-based immunotherapies, the use of adoptive NK cells, NK cells in combination with cytokines, immune checkpoint inhibitors, bispecific and trispecific killer cell engagers, and chimeric antigen receptor-engineered NK cells have all shown antitumor activity in AML patients. In this review, we will discuss the current strategies with these NK cell-based immunotherapies as possible therapies to cure AML patients post allo-HSCT. Additionally, we will discuss various means of immune escape in order to further understand the mechanism of NK cell-based immunotherapies against AML.
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Sivori, Simona, Raffaella Meazza, Concetta Quintarelli, Simona Carlomagno, Mariella Della Chiesa, Michela Falco, Lorenzo Moretta, Franco Locatelli, and Daniela Pende. "NK Cell-Based Immunotherapy for Hematological Malignancies." Journal of Clinical Medicine 8, no. 10 (October 16, 2019): 1702. http://dx.doi.org/10.3390/jcm8101702.
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Natural killer (NK) lymphocytes are an integral component of the innate immune system and represent important effector cells in cancer immunotherapy, particularly in the control of hematological malignancies. Refined knowledge of NK cellular and molecular biology has fueled the interest in NK cell-based antitumor therapies, and recent efforts have been made to exploit the high potential of these cells in clinical practice. Infusion of high numbers of mature NK cells through the novel graft manipulation based on the selective depletion of T cells and CD19+ B cells has resulted into an improved outcome in children with acute leukemia given human leucocyte antigen (HLA)-haploidentical hematopoietic transplantation. Likewise, adoptive transfer of purified third-party NK cells showed promising results in patients with myeloid malignancies. Strategies based on the use of cytokines or monoclonal antibodies able to induce and optimize NK cell activation, persistence, and expansion also represent a novel field of investigation with remarkable perspectives of favorably impacting on outcome of patients with hematological neoplasia. In addition, preliminary results suggest that engineering of mature NK cells through chimeric antigen receptor (CAR) constructs deserve further investigation, with the goal of obtaining an “off-the-shelf” NK cell bank that may serve many different recipients for granting an efficient antileukemia activity.
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Arcangeli, Silvia, Marco Bardelli, Sarah Tettamanti, Maria Caterina Rotiroti, Luca Simonelli, Chiara F. Magnani, Luca Varani, Andrea Biondi, and Ettore Biagi. "Unraveling the Efficacy and Safety Profiles of Anti-CD123 Chimeric Antigen Receptors (CARs) in a Model of Acute Myeloid Leukemia Immunotherapy By Investigating CAR Binding Affinity and Density Variables." Blood 126, no. 23 (December 3, 2015): 1359. http://dx.doi.org/10.1182/blood.v126.23.1359.1359.
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Abstract In the last years, adoptive cellular immunotherapy employing T lymphocytes genetically modified with Chimeric Antigen Receptors (CARs) has demonstrated impressive clinical results, particularly in the treatment of acute/chronic lymphoblastic leukemia and B-cell lymphoma, paving the way towards the possibility to translate this approach also to other hematological malignancies, such as Acute Myeloid Leukemia (AML). In the AML context, the CD123 antigen (IL-3 receptor alpha subunit) represents a good target antigen, being a poor prognosis over-expressed marker on AML cells and leukemic stem cells (LSCs), a rare population that plays a key role in perpetuating leukemia. However, CD123 is also expressed on the surface of healthy cells such as monocytes and endothelial cells, although at lower levels as compared to leukemic cells. The potential recognition of low antigen positive healthy tissues by CAR-redirected T cells, through the so called "on-target-off-organ" effect, limits a safe clinical employment of this immunotherapeutic approach. CARs are artificial receptors generated by joining the cytoplasmic TCR (T Cell Receptor) signaling modules to the heavy and light chain variable regions of a monoclonal antibody, whose affinity toward a target antigen is a variable capable of influencing the CAR-mediated functional responses. Therefore, in our study we investigated how the CAR affinity variable in the context of CD123 targeting, together with the CAR and CD123 target antigen density, could impact anti-CD123. CAR-redirected effector cells efficacy against leukemic cells and safety towards the healthy cells. To this aim, Cytokine-Induced Killer (CIK) effector cells have been genetically modified with four Chimeric Affinity Mutants (CAMs), CAM-1, CAM-2, CAM-3 and CAM-4, identified by means of a computational docking technique. In vitro cytotoxic assays, cytokine production and proliferation experiments have been performed in order to evaluate both the efficacy and safety profile of the CAR-redirected CIK cells, using un-manipulated CIK cells (NO DNA) and wild-type anti-CD123.CAR condition as controls. The functional characterization of all the CAMs revealed both the specificity and the effectiveness of CIK-CAR+ cells against the CD123+ THP-1 cell line and primary AML cells. However we observed that, at least in the context of CD123 targeting, a good CAR expression level is necessary for inducing effective later functions, such as proliferation and cytokine production, towards a high CD123+ target. When introducing leukemic cell lines with different CD123 density on their surface we observed different effector properties minimally influenced by the CAR affinity. In particular, in terms of killing activity, we noticed that a number of ≈1600 CD123 molecules is sufficient to induce a good cytotoxic response of all the CARs tested, with the CAM-2 (2-magnitude log lower in affinity) being less powerful. At the same time, this antigen density is not enough to determine a good proliferative capability which instead occurs with leukemic target cells expressing 5000 or more CD123 molecules. Considering the cytokine production (IL-2 and IFN-gamma), we observed that all CIK-CAR+ cells showed a cytokine release that is directly proportional to the target antigen density, with CAM-2 showing a reduced response towards low-CD123 expressing leukemic targets. When analyzing the safety profile of the CAMs against low-CD123+ endothelial target cells (≈1600 molecule/cell), we observed a lower functional activity of the CAMs as compared to the leukemic cell lines expressing the same level of CD123 molecules on their surface, with the low-affinity CAM-2 showing a major sparing capability in terms of killing activity (being the only one not statistically different from NO DNA). In conclusion, exploiting our model of affinity mutants we were able to in vitro characterize the role of the CAR density balanced with the affinity of the anti-CD123.CAR towards AML cells expressing different CD123 levels and CD123-low expressing normal tissues. In particular, the results obtained with CAM-2 suggested a potential threshold of affinity below which, even if the safety profile is preserved, the anti-leukemic efficacy would be impaired. Disclosures No relevant conflicts of interest to declare.
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Testa, Pelosi, and Castelli. "CD123 as a Therapeutic Target in the Treatment of Hematological Malignancies." Cancers 11, no. 9 (September 12, 2019): 1358. http://dx.doi.org/10.3390/cancers11091358.
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The interleukin-3 receptor alpha chain (IL-3R), more commonly referred to as CD123, is widely overexpressed in various hematological malignancies, including acute myeloid leukemia (AML), B-cell acute lymphoblastic leukemia, hairy cell leukemia, Hodgkin lymphoma and particularly, blastic plasmacytoid dendritic neoplasm (BPDCN). Importantly, CD123 is expressed at both the level of leukemic stem cells (LSCs) and more differentiated leukemic blasts, which makes CD123 an attractive therapeutic target. Various agents have been developed as drugs able to target CD123 on malignant leukemic cells and on the normal counterpart. Tagraxofusp (SL401, Stemline Therapeutics), a recombinant protein composed of a truncated diphtheria toxin payload fused to IL-3, was approved for use in patients with BPDCN in December of 2018 and showed some clinical activity in AML. Different monoclonal antibodies directed against CD123 are under evaluation as antileukemic drugs, showing promising results either for the treatment of AML minimal residual disease or of relapsing/refractory AML or BPDCN. Finally, recent studies are exploring T cell expressing CD123 chimeric antigen receptor-modified T-cells (CAR T) as a new immunotherapy for the treatment of refractory/relapsing AML and BPDCN. In December of 2018, MB-102 CD123 CAR T developed by Mustang Bio Inc. received the Orphan Drug Designation for the treatment of BPDCN. In conclusion, these recent studies strongly support CD123 as an important therapeutic target for the treatment of BPDCN, while a possible in the treatment of AML and other hematological malignancies will have to be evaluated by in the ongoing clinical studies.
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Simmons, Gary L., and Omar Castaneda Puglianini. "T-Cell-Based Cellular Immunotherapy of Multiple Myeloma: Current Developments." Cancers 14, no. 17 (August 31, 2022): 4249. http://dx.doi.org/10.3390/cancers14174249.
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T-cell-based cellular therapy was first approved in lymphoid malignancies (B-cell acute lymphoblastic leukemia and large B-cell lymphoma) and expanding its investigation and application both in hematological and non-hematological malignancies. Two anti-BCMA (B cell maturation antigen) CAR (Chimeric Antigen Receptor) T-cell therapies have been recently approved for relapsed and refractory multiple myeloma with excellent efficacy even in the heavily pre-treated patient population. This new therapeutic approach significantly changes our practice; however, there is still room for further investigation to optimize antigen receptor engineering, cell harvest/selection, treatment sequence, etc. They are also associated with unique adverse events, especially CRS (cytokine release syndrome) and ICANS (immune effector cell-associated neurotoxicity syndrome), which are not seen with other anti-myeloma therapies and require expertise for management and prevention. Other T-cell based therapies such as TCR (T Cell Receptor) engineered T-cells and non-genetically engineered adoptive T-cell transfers (Vγ9 Vδ2 T-cells and Marrow infiltrating lymphocytes) are also actively studied and worth attention. They can potentially overcome therapeutic challenges after the failure of CAR T-cell therapy through different mechanisms of action. This review aims to provide readers clinical data of T-cell-based therapies for multiple myeloma, management of unique toxicities and ongoing investigation in both clinical and pre-clinical settings.
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Karbowski, Christine, Rebecca Goldstein, Brendon Frank, Kei Kim, Chi-Ming Li, Oliver Homann, Kelly Hensley, et al. "Nonclinical Safety Assessment of AMG 553, an Investigational Chimeric Antigen Receptor T-Cell Therapy for the Treatment of Acute Myeloid Leukemia." Toxicological Sciences 177, no. 1 (June 26, 2020): 94–107. http://dx.doi.org/10.1093/toxsci/kfaa098.
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Abstract Feline McDonough Sarcoma-like tyrosine kinase 3 (FLT3), a tyrosine-protein kinase involved in hematopoiesis, is detectable on the cell surface of approximately 80% of leukemia isolates from adult patients with acute myeloid leukemia (AML). AMG 553 is an investigational chimeric antigen receptor (CAR) T-cell immunotherapy for the treatment of AML. FLT3 expression analysis and in vitro and in vivo studies were leveraged to evaluate the nonclinical safety of AMG 553. Cynomolgus monkeys administered autologous anti-FLT3 CAR T cells demonstrated no evidence of CAR T-cell-mediated toxicity, expansion, or persistence, likely due to restricted cell surface FLT3 protein expression in healthy animals. This highlights the limited value of such in vivo studies for safety assessment of the CAR T-cell modality when directed against a target with restricted expression. To complement these studies and directly evaluate the potential toxicities of eliciting T-cell-mediated cytotoxicity against cells with surface expression of FLT3 protein in vivo, data from cynomolgus monkey toxicology studies with 2 bispecific T-cell engager molecules targeting FLT3 were leveraged; findings were consistent with the targeted killing of bone marrow cells expressing cell surface FLT3. Potential AMG 553-induced cytotoxicity was assessed against a wide range of normal human primary cells and cell lines; cytotoxicity was observed against FLT3-positive AML cell lines and a percentage of primary bone marrow CD34+ cells. In conclusion, the nonclinical safety data suggest that AMG 553 can target FLT3 protein on AML cells, whereas only affecting a percentage of normal hematopoietic stem and progenitor cells, supporting clinical development.
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Galetto, Roman, Céline Lebuhotel, Agnès Gouble, Nuria Mencia-Trinchant, Cruz M. Nicole, Gail J. Roboz, Monica L. Guzman, and Julianne Smith. "TCRab Deficient CAR T-Cells Targeting CD123: An Allogeneic Approach of Adoptive Immunotherapy for the Treatment of Acute Myeloid Leukemia (AML)." Blood 126, no. 23 (December 3, 2015): 2555. http://dx.doi.org/10.1182/blood.v126.23.2555.2555.
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Abstract The remissions achieved using autologous T-cells expressing chimeric antigen receptors (CARs) in patients with advanced B cell leukemia and lymphomas have encouraged the use of CAR technology to treat different types of cancers by targeting distinct tumor-specific antigens. Since the current autologous approach utilizes CAR T-cells manufactured on a "per patient" basis, we propose an alternative approach based on the use of a standardized platform for manufacturing T-cells from third-party healthy donors to generate allogeneic "off-the-shelf" CAR T-cell-based frozen products. In the present work we have adapted this allogeneic platform to the production of T-cells targeting CD123, the transmembrane alpha chain of the interleukin-3 receptor, which is expressed on tumor cells from the majority of patients with Acute Myeloid Leukemia (AML). Multiple antigen recognition domains were screened in the context of different CAR architectures to identify candidates displaying activity against cells expressing variable levels of the CD123 antigen. The three lead candidates were tested in an orthotopic human AML cell line xenograft mouse model. From the three candidates that displayed comparable activity in vitro, we found two candidates capable of eradicating tumor cells in vivo with high efficiency. Subsequently, Transcription Activator-Like Effector Nuclease (TALEN) gene editing technology was used to inactivate the TCRα constant (TRAC) gene, eliminating the potential for engineered T-cells to mediate Graft versus Host Disease (GvHD). Editing of the TRAC gene can be achieved at high frequencies, and allows efficient amplification of TCR-deficient T-cells that no longer mediate alloreactivity in a xeno-GvHD mouse model. In addition, we show that TCR-deficient T-cells display equivalent in vitro and in vivo activity to non-edited T-cells expressing the same CAR. We have performed an initial evaluation of the expression of CD123 in AML patients and found an average cell surface expression of CD123 was of 67% in leukemic blasts (95% CI 48-82), 71% in CD34+CD38+ cells (95% CI 56-86), and 64% in CD34+CD38- (95% CI 41-87). Importantly, we have found that CD123 surface expression persists in CD34+CD38-CD90- cells after therapy in at least 20% of patients in remission (n=25), thus emphasizing the relevance of the target. Currently, the sensitivity of primary AML cells to CAR T-cells is being tested. Finally, we will also present our large scale manufacturing process of allogeneic CD123 specific T-cells from healthy donors, showing the feasibility for this off-the-shelf T-cell product that could be available for administration to a large number of AML patients. Disclosures Galetto: Cellectis SA: Employment. Lebuhotel:Cellectis SA: Employment. Gouble:Cellectis SA: Employment. Smith:Cellectis: Employment, Patents & Royalties.
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Ali, Muhammad, Eirini Giannakopoulou, Yingqian Li, Madeleine Lehander, Stina Virding Culleton, Weiwen Yang, Cathrine Knetter, et al. "T cells targeted to TdT kill leukemic lymphoblasts while sparing normal lymphocytes." Nature Biotechnology 40, no. 4 (December 6, 2021): 488–98. http://dx.doi.org/10.1038/s41587-021-01089-x.
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AbstractUnlike chimeric antigen receptors, T-cell receptors (TCRs) can recognize intracellular targets presented on human leukocyte antigen (HLA) molecules. Here we demonstrate that T cells expressing TCRs specific for peptides from the intracellular lymphoid-specific enzyme terminal deoxynucleotidyl transferase (TdT), presented in the context of HLA-A*02:01, specifically eliminate primary acute lymphoblastic leukemia (ALL) cells of T- and B-cell origin in vitro and in three mouse models of disseminated B-ALL. By contrast, the treatment spares normal peripheral T- and B-cell repertoires and normal myeloid cells in vitro, and in vivo in humanized mice. TdT is an attractive cancer target as it is highly and homogeneously expressed in 80–94% of B- and T-ALLs, but only transiently expressed during normal lymphoid differentiation, limiting on-target toxicity of TdT-specific T cells. TCR-modified T cells targeting TdT may be a promising immunotherapy for B-ALL and T-ALL that preserves normal lymphocytes.
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Sleiman, Sara, Olga Shestova, Francisco Santiago, Elina Shrestha, Raymond Liang, Ronen Ben Jehuda, Vladislav Sandler, and Saar Gill. "Anti-FLT3 CAR T Cells in Acute Myeloid Leukemia." Blood 138, Supplement 1 (November 5, 2021): 1703. http://dx.doi.org/10.1182/blood-2021-151196.
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Abstract INTRODUCTION In patients with AML who are eligible for intensive therapy, the goal of treatment is the achievement of complete response followed by consolidation chemotherapy (in favorable risk disease) or hematopoietic stem cell transplantation (in intermediate or adverse risk disease). Patients who do not attain this initial goal lack effective therapeutic options. Extensive experience with chimeric antigen receptor (CAR) T cells in B-ALL has shown that CART cells can deliver potent and durable antigen-specific leukemia control, and that targeting a single antigen (CD19 for B-ALL) is associated with antigen-negative relapse. In this context, we sought to expand the existing preclinical CART armamentarium in AML by developing FLT3-specific CART cells and comparing them to our existing gold standard CD123-specific CART cells. Since activating mutations in FLT3 occur commonly in AML, we reasoned that this molecule would serve as an "Achilles heel" in AML immunotherapy. METHODS Novel fully humanized anti-human FLT3 receptor single chain variable fragments (scFV) were fused to CD28 and CD137 (41BB) costimulatory molecules and the CD3zeta signaling domain and cloned into a lentiviral expression vector. Based on recently published data, we tested linker lengths ranging from 5 to 20 amino acids between the light and heavy chains of the CAR. We used a FLT3-ITD mutated AML cell line (MOLM14) expressing luciferase for in vitro function studies including an exhaustion assay. For in vivo function studies, we engrafted MOLM14 expressing luciferase into NSG mice and treated with CART-FLT3 or untransduced T cells (negative control). RESULTS All FLT3 and CD123-specific CART cells degranulated and produced the effector cytokines IL-2, INFg, TNF and GM-CSF in an antigen-specific manner, with some variability between the different linker lengths and with some superiority of the CAR123 likely resulting from the higher expression of CD123 compared with FLT3 in this model (p < 0.0001, one way ANOVA) (Figure 1). Short-term killing assays (24 hours) revealed that all CART cells killed MOLM14 with equivalent efficiency at low effector:target ratios (Figure 2A). Since short-term killing assays likely do not replicate the physiological situation in vivo wherein CART cells encounter cancer cells repeatedly over many days, we next developed an in vitro exhaustion assay. We incubated MOLM14 cells with CAR T cells at 1:10 E:T ratio and added MOLM 14 tumor cells along with fresh media every other day. Killing was quantified every 48 hours. Interestingly, all CAR constructs showed equivalently efficient cytotoxicity from days 5-15. However, after day 15 there was progressive dysfunction and loss of cytotoxic activity. This exhaustion "stress test" revealed some superiority of the FLT3 CAR 10AA construct (p = 0.042 on day 17, two way ANOVA) (Figure 2B). NOD/SCID gamma chain KO (NSG) mice were then engrafted with 1x10 6 luciferized MOLM14 cells and treated with 0.5x10 6 CAR T cells 7 days later, randomized to treatment groups based on tumor burden. CAR T cells expansion was monitored in peripheral blood by flow cytometry. (Fig 3A). Serial BLI revealed prompt and durable leukemia remissions and survival (Figure 3B,C). CONCLUSIONS We have developed CART-FLT3 for AML using novel human anti-FLT3 targeting domains and demonstrated preclinical efficacy similar to that of CART-123 in an AML model with substantially lower expression of FLT3 compared to CD123 (data not shown). Since inhibition of FLT3 leads to upregulation of surface FLT3 expression, future experiments will explore combinatorial FLT3 inhibition with CART-FLT3. If successful, these experiments will provide a strong rationale for a combination clinical trial in AML where leukemia control by small molecules is followed by a coup-de-grace delivered by CART cells. Figure 1 Figure 1. Disclosures Sleiman: Hemogenyx Pharmaceuticals LLC: Research Funding. Shestova: Hemogenyx Pharmaceuticals LLC: Research Funding. Santiago: Hemogenyx Pharmaceuticals LLC: Research Funding. Shrestha: Hemogenyx Pharmaceuticals LLC: Current Employment. Liang: Hemogenyx Pharmaceuticals LLC: Current Employment. Ben Jehuda: Hemogenyx Pharmaceuticals LLC: Current Employment. Sandler: Hemogenyx Pharmaceuticals LLC: Current Employment, Current equity holder in publicly-traded company. Gill: Novartis: Other: licensed intellectual property, Research Funding; Interius Biotherapeutics: Current holder of stock options in a privately-held company, Research Funding; Carisma Therapeutics: Current holder of stock options in a privately-held company, Research Funding.
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Friesen, Jeffrey, Vladimir Senyukov, Cecele J. Denman, Srinivas S. Somanchi, Simon Olivares, Laurence J. N. Cooper, and Dean A. Lee. "The Contribution of Signaling Endodomains to CD33-Mediated Killing of Acute Myeloid Leukemia by Natural Killer Cells Transiently Modified with Chimeric Antigen Receptor." Blood 114, no. 22 (November 20, 2009): 2463. http://dx.doi.org/10.1182/blood.v114.22.2463.2463.
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Abstract Abstract 2463 Poster Board II-440 Acute myeloid leukemia (AML) is an aggressive malignancy for which current therapy fails to provide durable remission in approximately half of cases. Natural killer (NK) cells, as a key component of innate immunty, have recently shown clinical potential for adoptive immunotherapy against AML, particular when the donor and recipient are KIR mismatched. In addition to patients who do not have a suitable related donor, approximately 30% of patients bear all three families of KIR ligands and therefor can not benefit from KIR mismatch. Thus, the major obstacles for adoptive NK cell immunotherapy are 1) obtaining sufficient numbers of NK cells for effective thereapy and 2) finding a related donor with predicted KIR mismatch. Clinical trials with humanized or engineered mAbs against CD33 have validated this antigen as a target for immunotherapy of AML, but are complicated by side effects such as a hepatotoxicity due to CD33 expression on normal hepatocytes. To address the first hurdle, we developed a method to expand CD3-CD56+ primary NK cells in vitro using artificial APCs expressing membrane-bound IL21, and have validated electroporation as an efficient method for gene modification of these NK cells. To address the second hurdle and expand the therapeutic potential of KIR-matched expanded NK cells, we hypothesized that gene transfer of CD33 Chimeric Antigen Receptor (CAR) could provide additional activation signal to increase the lysis of AML blasts by expanded NK cells, and sought to compare signaling endodomains for this purpose. CD3z is a signal adapter molecule for NKp30, NKp46, and CD16 in NK cells. We developed a CD33CAR composed of a CD33 single-chain variable fragment fused with the CD3z transmembrane domain expressed in Sleeping Beauty transposon vector system, and compared a first generation (CD3z only) endodomain with second generation endodomains (CD3z plus either CD28 or CD137). Transient gene transfer of the CD33CAR DNA into NK cells was accomplished using the Amaxa Nucleofector device. Functional expression of the CAR was determined by binding of a Siglec3-IgG fusion protein to the cell surface followed by secondary staining with anti-IgG-FITC. Cytotoxicity of the NK cells against CD33+ AML cells and CD33-transduced HEK293T cells was determined in a 4h lysis assay using Calcein-AM. While the maximum electroporation efficiency was only 15% at 24h, expression levels as low as 4% significantly increased the cytotoxic activity of NK cells compared to unelectroporated NK cells. Each of the CD33CAR constructs harboring different endodomains yielded an equivalent increase in target cell lysis (Figure). This data supports recent observations that signal transduction through CD3z is sufficient to activate cytotoxic activity in NK cells. However, to increase the percentage of CAR-expressing NK cells we are further evaluating the role of endodomain signaling in CAR-dependent proliferation of NK cells electroporated with both transposon and transposase. Disclosures: No relevant conflicts of interest to declare.
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Gill, Saar, Sarah K. Tasian, Marco Ruella, Olga Shestova, Yong Li, David L. Porter, Martin Carroll, et al. "Anti-CD123 Chimeric Antigen Receptor T Cells (CART-123) Provide A Novel Myeloablative Conditioning Regimen That Eradicates Human Acute Myeloid Leukemia In Preclinical Models." Blood 122, no. 21 (November 15, 2013): 143. http://dx.doi.org/10.1182/blood.v122.21.143.143.
Full textAbstract:
Abstract Engineering of T cells with chimeric antigen receptors (CARs) can impart novel T cell specificity for an antigen of choice, and anti-CD19 CAR T cells have been shown to effectively eradicate CD19+ malignancies. Most patients with acute myeloid leukemia (AML) are incurable with standard therapies and may benefit from a CAR-based approach, but the optimal antigen to target remains unknown. CD123, the IL3Rα chain, is expressed on the majority of primary AML specimens, but is also expressed on normal bone marrow (BM) myeloid progenitors at lower levels. We describe here in vitro and in vivostudies to evaluate the feasibility and safety of CAR-based targeting of CD123 using engineered T cells (CART123 cells) as a therapeutic approach for AML. Our CAR consisted of a ScFv derived from hybridoma clone 32716 and signaling domains from 4-1-BB (CD137) and TCR-ζ. Among 47 primary AML specimens we found high expression of CD123 (median 85%, range 6-100%). Quantitative PCR analysis of FACS-sorted CD123dim populations showed measurable IL3RA transcripts in this population, demonstrating that blasts that are apparently CD123dim/neg by flow cytometry may in fact express CD123. Furthermore, FACS-sorted CD123dimblasts cultured in methylcellulose up-regulated CD123, suggesting that anti-CD123 immunotherapy may be a relevant strategy for all AML regardless of baseline myeloblast CD123 expression. CART123 cells incubated in vitro with primary AML cells showed specific proliferation, killing, and robust production of inflammatory cytokines (IFN-α, IFN-γ, RANTES, GM-CSF, MIP-1β, and IL-2 (all p<0.05). In NOD-SCID-IL2Rγc-/- (NSG) mice engrafted with the human AML cell line MOLM14, CART123 treatment eradicated leukemia and resulted in prolonged survival in comparison to negative controls of saline or CART19-treated mice (see figure). Upon MOLM14 re-challenge of CART123-treated animals, we further demonstrated robust expansion of previously infused CART123 cells, consistent with establishment of a memory response in animals. A crucial deficiency of tumor cell line models is their inability to represent the true clonal heterogeneity of primary disease. We therefore engrafted NSG mice that are transgenic for human stem cell factor, IL3, and GM-CSF (NSGS mice) with primary AML blasts and treated them with CART123 or control T cells. Circulating myeloblasts were significantly reduced in CART123 animals, resulting in improved survival (p = 0.02, n=34 CART123 and n=18 control animals). This observation was made regardless of the initial level of CD123 expression in the primary AML sample, again confirming that apparently CD123dimAML may be successfully targeted with CART123 cells. Given the potential for hematologic toxicity of CART123 immunotherapy, we treated mice that had been reconstituted with human CD34+ cells with CART123 cells over a 28 day period. We observed near-complete eradication of human bone marrow cells. This finding confirmed our finding of a significant reduction in methylcellulose colonies derived from normal cord blood CD34+ cells after only a 4 hour in vitro incubation with CART123 cells (p = 0.01), and was explained by: (i) low level but definite expression of CD123 in hematopoietic stem and progenitor cells, and (ii) up-regulation of CD123 upon myeloid differentiation. In summary, we show for the first time that human CD123-redirected T cells eradicate both primary human AML and normal bone marrow in xenograft models. As human AML is likely preceded by clonal evolution in normal or “pre-leukemic” hematopoietic stem cells (Hong et al. Science 2008, Welch et al. Cell 2012), we postulate that the likelihood of successful eradication of AML will be enhanced by myeloablation. Hence, our observations support CART-123 as a viable therapeutic strategy for AML and as a novel cellular conditioning regimen prior to hematopoietic cell transplantation. Figure 1. Figure 1. Disclosures: Gill: Novartis: Research Funding; American Society of Hematology: Research Funding. Carroll:Leukemia and Lymphoma Society: Research Funding. Grupp:Novartis: Research Funding. June:Novartis: Research Funding; Leukemia and Lymphoma Society: Research Funding. Kalos:Novartis: Research Funding; Leukemia and Lymphoma Society: Research Funding.
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Atrash, Shebli, Kulsum Bano, Bradley Harrison, and Al-Ola Abdallah. "CAR-T treatment for hematological malignancies." Journal of Investigative Medicine 68, no. 5 (March 21, 2020): 956–64. http://dx.doi.org/10.1136/jim-2020-001290.
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Chimeric antigen receptor (CAR)-T-cell therapy has sparked a wave of optimism in hematological malignancies, reflected by the successful results of early clinical trials involving patients with pre-B-cell acute lymphoblastic leukemia, B-cell lymphomas and multiple myeloma. CAR-T-cell therapy is considered to be a novel immunotherapy treatment that has the potential for curing certain hematological cancers. However, as use of CAR-T-cell therapy has grown, new challenges have surfaced. These challenges include the process of manufacturing the CAR-T cells, the mechanisms of resistance that underlie disease relapse, adverse effects and cost. This review describes the published results of clinical trials and expected developments to overcome CAR-T resistance.
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Vishwasrao, Paresh, Gongbo Li, Justin C. Boucher, D. Lynne Smith, and Susanta K. Hui. "Emerging CAR T Cell Strategies for the Treatment of AML." Cancers 14, no. 5 (February 27, 2022): 1241. http://dx.doi.org/10.3390/cancers14051241.
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Engineered T cells expressing chimeric antigen receptors (CARs) on their cell surface can redirect antigen specificity. This ability makes CARs one of the most promising cancer therapeutic agents. CAR-T cells for treating patients with B cell hematological malignancies have shown impressive results. Clinical manifestation has yielded several trials, so far five CAR-T cell therapies have received US Food and Drug Administration (FDA) approval. However, emerging clinical data and recent findings have identified some immune-related toxicities due to CAR-T cell therapy. Given the outcome and utilization of the same proof of concept, further investigation in other hematological malignancies, such as leukemias, is warranted. This review discusses the previous findings from the pre-clinical and human experience with CAR-T cell therapy. Additionally, we describe recent developments of novel targets for adoptive immunotherapy. Here we present some of the early findings from the pre-clinical studies of CAR-T cell modification through advances in genetic engineering, gene editing, cellular programming, and formats of synthetic biology, along with the ongoing efforts to restore the function of exhausted CAR-T cells through epigenetic remodeling. We aim to shed light on the new targets focusing on acute myeloid leukemia (AML).
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Gillissen, Marijn A., Martijn Kedde, Greta de Jong, Etsuko Yasuda, Sophie E. Levie, Arjen Q. Bakker, Marie Jose Kersten, et al. "Tumor Specific Glycosylated CD43 Is a Novel and Highly Specific Target for Acute Myeloid Leukemia and Myelodysplastic Syndrome." Blood 128, no. 22 (December 2, 2016): 1646. http://dx.doi.org/10.1182/blood.v128.22.1646.1646.
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Abstract Background: Acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) are high-risk diseases with a poor prognosis. Even with intensive treatment regimens less than 50% of patients can be cured, and for the majority of patients - those over 65 years of age and/or patients with comorbidities - such intensive regimens are not feasible. Novel therapeutic approaches such as immunotherapy directed against a specific tumor target are highly needed. Aims: The aim of our study was to identify antibodies that are highly specific for AML and to discover novel tumor-specific antigens, widely expressed on AML and MDS but not on healthy hematopoietic and non-hematopoietic cells. Methods: Allogeneic bone marrow transplantation is an immunotherapy with proven therapeutic efficacy. We selected a patient with high-risk AML who remained disease free, now more than 5 years after receiving an allogeneic HSCT and therefore can be considered to have mounted a potent graft versus leukemia response. To study the antibody repertoire of this patient we isolated CD27+ IgG+ memory B lymphocytes, about 2 years after the transplant. These cells were transduced with Bcl-6 and Bcl-xL to generate plasmablast B cell clones that produce antibodies and express the B cell receptor on the cell surface. Supernatants of these B cell clones were used to screen for binding to surface antigens on the AML cell line THP-1. Results: We identified an donor derived IgG1 antibody, AT1413, that specifically bound to AML cell lines THP-1, MOLM-13, SH-2 and others, but not to normal bone marrow cells or non-hematopoietic cells. The antibody also interacted with AML blasts from the allogeneic HSCT patient from whom the antibody was derived, and with leukemic blasts isolated from newly diagnosed AML and MDS patients. Biochemical analysis revealed that AT1413 recognizes a sialylated epitope on CD43 which is specifically expressed on all types of AML and MDS cells, as illustrated by its reactivity with blasts of each of 60 randomly selected AML and MDS patients in our clinic. Since the target it is also expressed by CD34+ hematopoietic progenitor cells obtained from fetal liver and fetal bone marrow, but not by post-natal hematopoietic progenitor cells, it can be considered to be a oncofetal epitope. AT1413 induced antibody-dependent cell-mediated cytotoxicity and complement dependent cytotoxicity of AML cell lines and primary blasts. Summary and conclusion: We have identified oncofetal-sialylated CD43 (CD43os) as a novel tumor-specific target that is widely expressed on AML and MDS blasts. Antibodies against this target have therefore high potential as therapeutic antibodies, either as a naked antibody or manufactured into an antibody-drug conjugate, bispecific T cell engager or CAR (chimeric antigen receptor) T cell. Disclosures Kersten: Celgene: Research Funding; Amgen: Honoraria.
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Rotiroti, Maria Caterina, Silvia Arcangeli, Chiara Buracchi, Chiara Francesca Magnani, Claudia Cappuzzello, Ettore Biagi, Sarah Tettamanti, and Andrea Biondi. "Specific Targeting of Acute Myeloid Leukemia By the Use of Non-Virally Engineered CIK (Cytokine-Induced Killer) Cells Expressing the Anti-CD33 Chimeric Antigen Receptor (CAR)." Blood 132, Supplement 1 (November 29, 2018): 2201. http://dx.doi.org/10.1182/blood-2018-99-114572.
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Abstract Background Acute Myeloid Leukemia (AML) is still associated with high relapse rates when treated with conventional chemotherapeutic and hematopoietic transplantation regimens. Thus, new treatment options are urgently needed. Immunotherapy adopting T cells engineered to express tumor-directed Chimeric Antigen Receptors (CARs) has shown striking results particularly in the context of B-cell malignancies, sparking a keen interest in extending this approach also to other hematological malignancies such as AML. Among the surface molecules identified, the CD33 molecule represents so far one of the main validated target in AML and, being broadly expressed on AML blasts, represents a suitable antigen to be targeted with CAR-T cells. Objectives The aim of the present study is to preclinically evaluate the efficacy and safety profiles of CD33.CAR redirected Cytokine Induced Killer (CIK) cells alone and in combination with standard chemotherapeutic agents. Methods Donor derived- and autologous-CIK cells were stably or transiently transduced with a third generation anti-CD33.CAR by Sleeping Beauty transposon- or mRNA-mediated engineering. In vitro anti-AML activity has been assessed by means of Flow cytometry-based cytotoxicity (AnnV-7AAD staining), proliferation (Ki67 staining and CFSE dilution) and cytokine production (intracellular IFNg and IL2 detection) assays, upon challenge with AML samples. In vivo efficacy has been evaluated in NSG mice transplanted with MA9-NRas AML cell line or primary AML samples. Moreover, an already established xenograft chemotherapy model has been exploited to examine the potential benefit of combining CD33.CAR-CIK cells with standard AML induction therapy (Ara-C and doxorubicin). Results CD33.CAR stably expressing CIK cells were able to induce a potent anti-leukemic activity in vitro, in terms of specific killing either in short term (>70% at 4h, E:T ratio 5:1) and long term cytotoxic assays (>90% at 1 week, E:T ratio 1:10), with statistically significant differences as compared to the unmanipulated condition. Moreover, CD33.CAR-CIK cells were able to retain a significant cytotoxic activity when re-challenged with the CD33+ target following a previous stimulation (up to 65%). The proliferative response to AML target cells was also considerable and CAR-specific (up to 60% of Ki67+CAR-CIK cells and up to 70% of CFSE diluted CAR-CIK cells), as well as the cytokine production (up to 35% of IFN-γ producing CAR-CIK cells and up to 25% of IL-2 producing CAR-CIK cells). CIK cells transiently expressing the CD33.CAR were also effective towards the AML target. In vivo results showed that CD33.CAR-CIK cells were able to control the disease in MA9 grafted mice in all the districts analyzed (peripheral blood, bone marrow, spleen, liver and kidney), as compared to untreated mice. To evaluate the effect of CD33.CAR-CIK cell immunotherapy particularly on Leukemia Initiating Cells (LICs), CD33.CAR-CIK cells were administered as an early treatment approach, treating mice 5 days after i.v. injection of a secondary transplanted PDX sample. We observed a clear engraftment reduction in the treated cohort, nearly undetectable in 2 out 5 mice, while a high leukemic burden has been detected in untreated mice (up to 70% of engraftment in bone marrow). Furthermore, by exploiting CD33.CAR-CIK cell treatment in mice experiencing disease recurrence after the "5+3" chemotherapy-induction protocol, preliminary data showed that CD33.CAR-CIK cells were also capable to target chemotherapy resistant/residual AML cells. Conclusions Considering our in vivo preliminary results, we aim to further evaluate CD33.CAR-CIK cell immunotherapy efficacy, particularly against chemotherapy resistant/residual AML cells. Concerning the safety aspect, since the CD33 targeting raises concerns for a potential myelotoxicity, we will assess the potential long-term off-target effects of CD33.CAR-CIK cells (comparing stably with transiently expressed CD33.CARs) on normal hematopoietic stem/myeloid progenitor cells. Disclosures No relevant conflicts of interest to declare.
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Meiliana, Anna, Nurrani Mustika Dewi, and Andi Wijaya. "Prospect of Natural Killer Cells in Cancer Imunotherapy." Indonesian Biomedical Journal 10, no. 3 (December 28, 2018): 192–202. http://dx.doi.org/10.18585/inabj.v10i3.532.
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BACKGROUND: Current understanding in molecular character of natural killer (NK) cell, its function and mechanisms, send people the ideas to develop a NK-cell-based immunotherapeutic strategies against human cancer.CONTENT: Before being regards as a cell-based cellular therapy, NK cell have to be clinical proven. Early studies with NK cells infusions for acute myeloid leukemia and lung cancer showed a promising result. NK cells need simplified methods for enriching and expanding, in addition to its transfection with chimeric antigen receptors (CARs). NK-92 arise as an assuring effector cells to augment monoclonal and bispecific antibody therapy. Thus, NK cells show a potential opportunity for cell engineering, outstep the era of T cells.SUMMARY: It is believed that NK cells bring a bright hope for future cancer immunotherapies, either alone or in combination as a harmonious therapy.KEYWORDS: NK-cells, NK-92 cells, immunotherapy, CAR