Journal articles on the topic 'Prenol kinases'

A proposed treatment for malaria is a combination of fosmidomycin and clindamycin. Both compounds inhibit the methylerythritol 4-phosphate (MEP) pathway, the parasitic source of farnesyl and geranylgeranyl pyrophosphate (FPP and GGPP, respectively). Both FPP and GGPP are crucial for the biosynthesis of several essential metabolites such as ubiquinone and dolichol, as well as for protein prenylation. Dietary prenols, such as farnesol (FOH) and geranylgeraniol (GGOH), can rescue parasites from MEP inhibitors, suggesting the existence of a missing pathway for prenol salvage via phosphorylation. In this study, we identified a gene in the genome of P. falciparum, encoding a transmembrane prenol kinase (PolK) involved in the salvage of FOH and GGOH. The enzyme was expressed in Saccharomyces cerevisiae, and its FOH/GGOH kinase activities were experimentally validated. Furthermore, conditional knockout parasites (Δ-PolK) were created to investigate the biological importance of the FOH/GGOH salvage pathway. Δ-PolK parasites were viable but displayed increased susceptibility to fosmidomycin. Their sensitivity to MEP inhibitors could not be rescued by adding prenols. Additionally, Δ-PolK parasites lost their capability to utilize prenols for protein prenylation. Experiments using culture medium supplemented with whole/delipidated human plasma in transgenic parasites revealed that human plasma has components that can diminish the effectiveness of fosmidomycin. Mass spectrometry tests indicated that both bovine supplements used in culture and human plasma contain GGOH. These findings suggest that the FOH/GGOH salvage pathway might offer an alternate source of isoprenoids for malaria parasites when de novo biosynthesis is inhibited. This study also identifies a novel kind of enzyme related to isoprenoid metabolism.

Membrane fusion is catalyzed by conserved proteins R, Qa, Qb, and Qc SNAREs, which form tetrameric RQaQbQc complexes between membranes; SNARE chaperones of the SM, Sec17/αSNAP, and Sec18/NSF families; Rab-GTPases (Rabs); and Rab effectors. Rabs are anchored to membranes by C-terminal prenyl groups, but can also function when anchored by an apolar polypeptide. Rabs are regulated by GTPase-activating proteins (GAPs), activating the hydrolysis of bound GTP. We have reconstituted fusion with pure components from yeast vacuoles including SNAREs, the HOPS (homotypic fusion and vacuole protein sorting) tethering and SNARE-assembly complex, and the Rab Ypt7, bound to membranes by either C-terminal prenyl groups (Ypt7-pr) or a recombinant transmembrane anchor (Ypt7-tm). We now report that HOPS-dependent fusion occurs with Ypt7 anchored by either means, but only Ypt7-pr requires GTP for activation and is inactive either with bound GDP or without bound guanine nucleotide. In contrast, Ypt7-tm is constitutively active for HOPS-dependent fusion, independent of bound guanine nucleotide. Fusion inhibition by the GAP Gyp1-46 is not limited to Ypt7-tm with bound GTP, indicating that this GAP has an additional mode of regulating fusion. Phosphorylation of HOPS by the vacuolar kinase Yck3 renders fusion strictly dependent on GTP-activated Ypt7, whether bound to membranes by prenyl or transmembrane anchor. The binding of GTP or GDP constitutes a selective switch for Ypt7, but with Ypt7-tm, this switch is only read by HOPS after phosphorylation to P-HOPS by its physiological kinase Yck3. The prenyl anchor of Ypt7 allows both HOPS and P-HOPS to be regulated by Ypt7-bound guanine nucleotide.

Dimethylallyl diphosphate (DMAPP) is a key intermediate metabolite in the synthesis of isoprenoids and is also the prenyl donor for biosynthesizing prenylated flavonoids. However, it is difficult to prepare DMAPP via chemical and enzymatic methods. In this study, three promiscuous kinases from Shigella flexneri (SfPK), Escherichia coli (EcPK), and Saccharomyces cerevisiae (ScPK) and three isopentenyl phosphate kinases from Methanolobus tindarius (MtIPK), Methanothermobacter thermautotrophicus str. Delta H (MthIPK), and Arabidopsis thaliana (AtIPK) were cloned and expressed in Escherichia coli. The enzymatic properties of recombinant enzymes were determined. The Kcat/Km value of SfPK for DMA was 6875 s−1 M−1, which was significantly higher than those of EcPK and ScPK. The Kcat/Km value of MtIPK for DMAP was 402.9 s−1 M−1, which was ~400% of that of MthIPK. SfPK was stable at pH 7.0–9.5 and had a 1 h half-life at 65 °C. MtIPK was stable at pH 6.0–8.5 and had a 1 h half-life at 50 °C. The stability of SfPK and MtIPK was better than that of the other enzymes. Thus, SfPK and MtIPK were chosen to develop a one-pot enzymatic cascade for producing DMAPP from DMA because of their catalytic efficiency and stability. The optimal ratio between SfPK and MtIPK was 1:8. The optimal pH and temperature for the one-pot enzymatic cascade were 7.0 and 35 °C, respectively. The optimal concentrations of ATP and DMA were 10 and 80 mM, respectively. Finally, maximum DMAPP production reached 1.23 mM at 1 h under optimal conditions. Therefore, the enzymatic method described herein for the biosynthesis of DMAPP from DMA can be widely used for the synthesis of isoprenoids and prenylated flavonoids.

KRAS4b is a small guanosine triphosphatase (GTPase) protein that regulates several signal transduction pathways that underlie cell proliferation, differentiation, and survival. KRAS4b function requires prenylation of its C terminus and recruitment to the plasma membrane, where KRAS4b activates effector proteins including the RAF family of kinases. The Ca2+-sensing protein calmodulin (CaM) has been suggested to regulate the localization of KRAS4b through direct, Ca2+-dependent interaction, but how CaM and KRAS4b functionally interact is controversial. Here, we determined a crystal structure, which was supported by solution nuclear magnetic resonance (NMR), that revealed the sequestration of the prenyl moiety of KRAS4b in the hydrophobic pocket of the C-terminal lobe of Ca2+-bound CaM. Our engineered fluorescence resonance energy transfer (FRET)–based biosensor probes (CaMeRAS) showed that, upon stimulation of Ca2+ influx by extracellular ligands, KRAS4b reversibly translocated in a Ca2+-CaM–dependent manner from the plasma membrane to the cytoplasm in live HeLa and HEK293 cells. These results reveal a mechanism underlying the inhibition of KRAS4b activity by Ca2+ signaling pathways.

Breast cancer is one of the most prevalent malignancies and the leading cause of cancer-associated mortality in China. Icaritin (ICT), a prenyl flavonoid derived from the Epimedium Genus, has been proven to inhibit the proliferation and stemness of breast cancer cells. Our previous study demonstrated that IC2, a derivative of ICT, could induce breast cancer cell apoptosis by Stearoyl-CoA desaturase 1 (SCD1) inhibition. The present study further investigated the mechanism of the inhibitory effects of IC2 on breast cancer cells in vitro and in vivo. Our results proved that IC2 could stimulate autophagy in breast cancer cells with the activation of adenosine monophosphate (AMP)-activated protein kinase (AMPK) signaling and mitogen-activated protein kinase (MAPK) signaling. Combination treatment of the AMPK inhibitor decreased IC2-induced autophagy while it markedly enhanced IC2-induced apoptosis. In common with IC2-induced apoptosis, SCD1 overexpression or the addition of exogenous oleic acid (OA) could also alleviate IC2-induced autophagy. In vivo assays additionally demonstrated that IC2 treatment markedly inhibited tumor growth in a mouse breast cancer xenograft model. Overall, our study was the first to demonstrate that IC2 induced cytoprotective autophagy by SCD1 inhibition in breast cancer cells and that the autophagy inhibitor markedly enhanced the anticancer activity of IC2. Therefore, IC2 was a potential candidate compound in combination therapy for breast cancer.

Aims. In this work we studied cytodifferentiation effects of newly characterized prenyl flavonoid 4′-O-methylkuwanon E (4ME) isolated from white mulberry (Morus albaL.).Main Methods. Cell growth and viability were measured by dye exclusion assay; cell cycle and surface antigen CD11b were monitored by flow cytometry. For the cytodifferentiation of cells the NBT reduction assay was employed. Regulatory proteins were assessed by western blotting.Key Findings. 4ME induced dose-dependent growth inhibition of THP-1 cells, which was not accompanied by toxic effect. Inhibition of cells proliferation caused by 4ME was associated with the accumulation in G1 phase and with downregulation of hyperphosphorylated pRb. Treatment with 4ME led to significant induction of NBT-reducing activity of PMA stimulated THP-1 cells and upregulation expression of differentiation-associated surface antigen CD11b. Our results suggest that monocytic differentiation induced by 4ME is connected with up-regulation of p38 kinase activity.Significance. Our study provides the first evidence that 4ME induces the differentiation of THP-1 human monocytic leukemia cells and thus is a potential cytodifferentiating anticancer agent.

Tomato (Solanum lycopersicum) fruit maturation is associated with a developmental transition from chloroplasts (in mature green fruit) to chromoplasts (in red fruit). The hallmark red color of ripe tomatoes is due to carotenogenesis and accumulation of the red carotenoid lycopene inside chromoplasts. Plastoglobules (PG) are lipid droplets in plastids that are involved in diverse lipid metabolic pathways. In tomato, information on the possible role of PG in carotogenesis and the PG proteome is largely lacking. Here, we outline the role of PG in carotenogenesis giving particular attention to tomato fruit PG proteomes and metabolomes. The proteome analysis revealed the presence of PG-typical FBNs, ABC1K-like kinases, and metabolic enzymes, and those were decreased in the PG of tomato chromoplasts compared to chloroplasts. Notably, the complete β-carotene biosynthesis pathway was recruited to chromoplast PG, and the enzymes PHYTOENE SYNTHASE 1 (PSY-1), PHYTOENE DESATURASE (PDS), ZETA-CAROTENE DESATURASE (ZDS), and CAROTENOID ISOMERASE (CRTISO) were enriched up to twelvefold compared to chloroplast PG. We profiled the carotenoid and prenyl lipid changes in PG during the chloroplast to chromoplast transition and demonstrated large increases of lycopene and β-carotene in chromoplast PG. The PG proteome and metabolome are subject to extensive remodeling resulting in high accumulation of lycopene during the chloroplast-to-chromoplast transition. Overall, the results indicate that PGs contribute to carotenoid accumulation during tomato fruit maturation and suggest that they do so by functioning as a biosynthetic platform for carotenogenesis.

: The use of anti-diabetic drugs has been increasing worldwide and the evolution of therapeutics has been enormous. Still, the currently available anti-diabetic drugs do not present the desired efficacy and are generally associated with serious adverse effects. Thus, entirely new interventions, addressing the underlying etiopathogenesis of type 2 diabetes mellitus, are required. Chalcones, secondary metabolites of terrestrial plants and precursors of the flavonoids biosynthesis, have been used for a long time in traditional medicine due to their wide-range of biological activities, from which the anti-diabetic activity stands out. : This review systematizes the information found in literature about the anti-diabetic properties of chalcones, in vitro and in vivo. Chalcones are able to exert these properties by acting in different therapeutic targets: Dipeptidyl Peptidase 4 (DPP-4); Glucose Transporter Type 4 (GLUT4), Sodium Glucose Cotransporter 2 (SGLT2), α-amylase, α-glucosidase, Aldose Reductase (ALR), Protein Tyrosine Phosphatase 1B (PTP1B), Peroxisome Proliferator-activated Receptor-gamma (PPARγ) and Adenosine Monophosphate (AMP)-activated Protein Kinase (AMPK). Chalcones are, undoubtedly, promising anti-diabetic agents, and some crucial structural features have already been established. From the Structure-Activity Relationships analysis, it can generally be stated that the presence of hydroxyl, prenyl and geranyl groups in their skeleton improves their activity for the evaluated anti-diabetic targets.

Abstract Background Ras proteins are vital for normal T cell activation, and downstream effectors of Ras include the MEK/ERK, PI3-kinase/AKT, and NF-kB pathways. T cells from Rheumatoid Arthritis patients exhibit abnormal activation of the Ras/MEK/ERK pathway. The small molecule Farnesylthiosalicylic acid (FTS) blocks the interaction between Ras proteins and their prenyl binding chaperones, attenuating plasma membrane localization and signaling. Objectives To investigate the immunomodulatory effect of FTS alone or combined with methotrexate (MTX) in the DBA/1 mouse collagen type-II induced arthritis (CIA) model. Methods Arthritis was induced in 8–10 week old male DBA/1 mice by immunization with collagen type-II (CII) and complete Freund’s adjuvant (CFA). Animals were treated semi-prophylactically with once daily oral FTS (100 mg/kg); weekly i.p injection of MTX (0.5 mg/kg), FTS combined with MTX, or daily oral vehicle solution (control). Arthritis severity was scored daily from disease onset until study termination and multiple immunological biomarkers of inflammation were analyzed. Results Our data from the mouse CIA model show that the therapeutic efficacy of FTS was similar to MTX, and both drugs significantly reduced arthritis severity compared to CMC controls. Importantly, FTS significantly inhibited the production of pathogenic anti-CII autoantibodies and upregulation of serum IL-6 and IL-17A compared to control arthritic mice. The in depth, multiplex, analysis of the effect of FTS on the T cell cytokine response to CII, revealed strong suppression of IL-22, IL-17, IL-9, GM-CSF and TNF production. Importantly, FTS therapy positively correlated with reduced p-ERK1/2 and p-AKT levels in splenic lymphocytes.

Abstract Introduction The major driver for pancreatic ductal adenocarcinoma (PDAC) is oncogenic KRAS. However, adult acinar cells, a probable origin of PDAC, are largely refractory to KrasG12D-mediated oncogenic transformation. With the concomitant loss of transcription factors that regulate acinar cell differentiation, such as Pdx1 (Pancreatic and Duodenal Homeobox 1), acinar cells undergo a rapid cell identity switch, known as acinar-to-ductal metaplasia (ADM). How loss of cell identity cooperates with oncogenic Kras to induce pancreatic transformation is largely unclear. Methods To elucidate mechanisms responsible for the accelerated cellular reprogramming in KrasG12D;Pdx1f/f animals, single-cell ATAC-seq (Assay for Transposase-Accessible Chromatin using sequencing) from frozen pancreatic bulk tissue was performed. Chromatin accessibility states were captured at early stages of carcinogenesis and correlated to RNA-seq data. Differentially regulated genes were validated by multiplex RNAscope and immunohistochemistry staining and functionally studied in pancreatic cancer cell lines. Results Single-cell ATAC-seq proved a powerful tool for defining cell-type identity, cellular reprogramming and target genes in early metaplastic transformation of pancreatic tissue. Notably, acinar cells of KrasG12D;Pdx1f/f animals as well as a proportion of metaplastic lesions in both, KrasG12D and KrasG12D;Pdx1f/f mice, showed elevated accessibility and expression of the Ror2 (Receptor Tyrosine Kinase Like Orphan Receptor 2) gene. As a receptor tyrosine kinase, Ror2 controls noncanonical Wnt signaling and other essential signaling pathways, such as PI3K/AKT or Ras-MAPK. Genetic ablation of Ror2 in a mouse model of pancreatic neoplasia resulted in a shift in ADM cell identity, enriching ADM lesions with a senescent duct cell phenotype. In PDAC, ROR2 expression correlates with the more aggressive basal-like subtype. Overexpression of ROR2 in pancreatic cancer cell lines with a classical differentiation induced epithelial-to-mesenchymal transition, characterized by the downregulation of multiple epithelial markers and upregulation of mesenchymal genes. Knockout of ROR2 in pancreatic cancer cells significantly decreased cell proliferation. Conclusions Our in-depth sequencing data revealed that expression of KrasG12D with the concomitant loss of Pdx1 leads to vast alterations of acinar cell identity. We identified the receptor kinase Ror2 as a regulator of pancreatic cancer initiation and driver of pancreatic cancer cell aggressiveness. Citation Format: Simone Benitz, Malak Nasser, Alexander Steep, Jonathan Preall, Ujjwal Mahajan, Ian Loveless, Erick Davis, Hui-Ju Wen, Daniel Long, Michaela Louw, Samuel Zwernik, Donald Rempinski, Jacee Moore, Daniel Salas-Escabillas, Thomas Metzler, Ling Huang, Nina Steele, Ivonne Regel, Filip Bednar, Howard Crawford. Single-cell epigenomic analysis reveals an important role of the receptor kinase Ror2 in the erosion of cellular identity during pancreatic carcinogenesis [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr A109.

e12527 Background: FUL is the recommended 2L treatment for patients whose HR+ ABC progressed after aromatase inhibitor (AI) therapy. In first line ABC adding targeted therapy, eg. cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) palbociclib or ribociclib, or mammalian target of rapamycin inhibitor (mTORi) everolimus (EVE), to endocrine therapy (ET) has shown superior efficacy vs ET alone. The use of similar strategies to delay disease progression on ET in the 2L setting is an area of active research. Methods: PubMed and ClinicalTrials.gov were searched for trials investigating FUL + targeted therapies in 2L HR+ ABC. Search terms: (advanced OR metastatic) AND (breast cancer) AND (FUL OR faslodex) AND (2L OR relapse OR refractory OR resistant OR progression). Efficacy, adverse events (AEs) and quality of life were assessed. Results: 28 studies of FUL + targeted therapies in 2L ABC were found. Key randomized trials include 8 studies exploring FUL + CDK4/6i: palbociclib, ribociclib or abemaciclib. Palbociclib + FUL significantly improved progression-free survival (PFS) vs FUL in 2L HR+ ABC (p < 0.0001; PALOMA-3), AEs were manageable. Assessment of FUL + ribociclib (MONALEESA-3) or FUL + abemaciclib (MONARCH-2) in 2L HR+ ABC is ongoing. Ten studies are evaluating FUL + phosphatidylinositol 3-kinase (PI3K)/AKT/mTORi in 2L HR+ ABC. FUL + EVE significantly prolonged PFS vs FUL (p = 0.02; PrECOG 0102). Two trials evaluated buparlisib (pan-PI3K inhibitor [PI3Ki]) + FUL in HR+ ABC post-AI (BELLE-2) and post-mTORi (BELLE-3). In both trials, buparlisib + FUL improved PFS vs FUL in patients with PIK3CA-mutated tumors, FUL alone led to a poor response in this subgroup. However, pictilisib (pan-PI3Ki) + FUL did not improve PFS vs FUL even in the PIK3CA-mutated subgroup (FERGI). Ongoing phase 3 trials are exploring FUL + alpelisib (α-specific PI3Ki; SOLAR-1) or FUL + taselisib (β-sparing PI3Ki; SANDPIPER) in PIK3CA-mutant HR+ ABC. Pts who have progressed on AI, CDK4/6i or (neo)adjuvant chemotherapy are eligible for these studies. Data on FUL + other targeted therapies will also be discussed. Conclusions: Addition of targeted therapy to FUL demonstrates promising efficacy beyond first line.

Abstract Introduction: Reprogramming of pancreas cell fate drives development of pancreatic ductal adenocarcinoma (PDAC). Acinar cells, the most probable origin of pancreatic cancer, undergo a rapid cell identity switch towards a duct-like phenotype upon KrasG12D expression and when combined with pancreatitis or the loss of acinar differentiation factors. Metaplastic and dysplastic duct-like cells are heterogeneous with a proportion acquiring features reminiscent of gastric lineages. While some gastric signatures are maintained in the classical PDAC subtype, they are eroded in the more aggressive, basal-like PDAC. Since subtype identity has a major impact on prognosis and therapeutic targetability, druggable targets that regulate cellular reprogramming in pancreatic cancer can be exploited to increase sensitivity to therapy. Methods: To elucidate mechanisms responsible for early reprogramming, pancreatic tissue of mice with conditional KrasG12D expression and loss of Pdx1 was analyzed by single-nucleus ATAC-seq. Expression of identified target genes was studied in precancerous lesions and PDAC by using multiplex RNAscope and IHC staining. Computational analyses of publicly available sequencing data were used to establish correlation to PDAC subtype identity. Genes were functionally studied in PDAC cell lines. Results: By performing snATAC-seq and RNA-seq of early transformed pancreatic tissue, we discovered that acinar cells with the combined expression of KrasG12D and loss of Pdx1 activate expression of a gastric metaplastic gene signature accompanied by elevated levels of the receptor kinase Ror2. Ror2 is also highly expressed in distinct subpopulations of metaplastic and dysplastic cells, associated with a gastric neck cell phenotype and enhanced proliferative capacity. In contrast, Ror2Low lesions are characterized by a gastric pit cell-like and a senescent phenotype. Genetic ablation of Ror2 resulted in a shift in lesion identity, enriching those with a pit cell and senescent identity. In PDAC, we found that Ror2 anti-correlates with a similar gastric pit cell phenotype that is maintained in the classical subtype PDAC but is strongly associated with the more aggressive basal-like subtype. Overexpression of ROR2 in human PDAC cell lines with a classical differentiation induced loss of the classical gene signature as well as epithelial-to-mesenchymal transition. Moreover, ROR2 enforces a strong dependency on AKT signaling, causing increased vulnerability of ROR2-expressing cells to AKT inhibition, but increased resistance to the KRAS inhibitor MRTX1133. Conclusions: We discovered Ror2 as a critical determinant of precancerous lesion as well as PDAC subtype identity. Its role in driving an aggressive PDAC phenotype that is inherently resistant to Kras inhibition suggests that inhibiting this receptor tyrosine kinase will enhance sensitivity to the new generation of targeted therapies. Citation Format: Simone Benitz, Alexander Steep, Malak Nasser, Jonathan Preall, Ujjwal Mahajan, Ian Loveless, Holly McQuithey, Erick Davis, Hui-Ju Wen, Daniel Long, Thomas Metzler, Samuel Zwernik, Daniel Salas-Escabillas, Ling Huang, Nina Steele, Ivonne Regel, Filip Bednar, Howard Crawford. Ror2, a novel key regulator driving cell fate decisions throughout pancreatic tumor progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3939.

Abstract Gilteritinib (Gilt), a FLT3 tyrosine kinase inhibitor (TKI), is approved for the treatment of relapsed/refractory (R/R) FLT3-mutated (FLT3 +)acute myeloid leukemia (AML). However, long-term survival is limited by the development of drug resistance mutations in persistent FLT3 + clones. Combination regimens may deepen response and improve outcomes. Venetoclax (Ven), a BCL-2 inhibitor, is approved in combination with hypomethylating agents for newly diagnosed AML not suitable for standard induction therapy. FLT3 + AMLhas been associated with clinical resistance to Ven. Still, FLT3 TKIs + Ven have demonstrated synthetic lethality in preclinical models, prompting this multicenter, open-label, phase 1b trial (NCT03625505) to evaluate Ven + Gilt for R/R AML. Here, we report final response and survival endpoints, and molecular clearance among patients (pts) treated at the recommended phase two dose (RP2D). The study design has been previously described (Daver, et al. ASH 2020, Abstract 333). Pts in the dose expansion cohort received Ven 400 mg + Gilt 120 mg (RP2D) daily in 28-day cycles, following Ven ramp-up. The primary endpoint was mCRc (complete response [CR] + CR with incomplete platelet recovery [CRp] + CR with incomplete blood count recovery [CRi] + morphologic leukemia-free state) to align with the ADMIRAL phase III trial (Perl, et al. NEJM 2019). Baseline co-mutations and serial FLT3 internal tandem duplications (ITD) allelic burden were assessed using the MyAML panel (Invivoscribe, San Diego, CA) and FLT3-ITD MRD assay with sensitivities of 5% and 0.001%, respectively. As of the data cut off of January 31, 2021, 54 pts were treated at the RP2D. Fifty-two pts (as assessed locally) had FLT3 +AML; 41 had FLT3-ITD only, 8 had tyrosine kinase domain only, 3 had both mutations, and 2 were FLT3 wild type (wt). Additional baseline characteristics are in Table 1. Grade 3 or 4 AEs occurred in 51 (94.4%) pts, and 39 (72.2%) pts had serious AEs. Grade 3 or 4 cytopenias occurred in 43 (79.6%) pts. AEs of special interest included tumor lysis syndrome (2 [3.7%]) and QT prolongation (1 [1.9%]). AEs leading to dosing interruptions (Ven, 27 [50%]; Gilt, 26 [48.1%]), reductions (Ven, 3 [5.6%]; Gilt, 4 [7.4%]), and discontinuation (Ven, 7 [13%]; Gilt, 7 [13%]) were reported for both study drugs. There was 1 treatment-emergent death of typhlitis. Among FLT3 + pts with post-baseline assessments (51/52), mCRc was achieved by 38 pts (74.5%; CR/CRp/CRi, 19 [37.3%] pts), with a median follow-up time of 12 mo (range: 0.8 - 20.1). The mCRc in pts with prior TKI (32) or prior Ven (10) use were 78.1% and 60%, respectively. The mOS among all FLT3 +pts was 10 mo (95%CI: 6.6, NE), with a median DoR (mDoR) of 5.4 mo (95% CI: 3.3, 6.6). Pts with FLT3-ITD had a mOS of 10.5 mo (95%CI: 6.8, NE), and a mDoR of 5.6 mo (95% CI: 3.3, 8.3). For the 14 pts who had a transplant, the mOS was not reached (95%CI: 10.0, NE) vs 6.89 mo (95%CI: 3.0, 10.5) for those who did not have a transplant (37). The mOS for pts with prior TKI or Ven was 9.6 (95% CI: 4.3, NE) and 10.5 mo (95% CI: 2.0, NE), respectively, with a mDoR of 9.6 (95% CI: 4.3, NE) and 6.2 mo (95% CI: 2.6, NE), respectively. The 60-day mortality rate was 11.8% (95% CI: 4.4, 23.9). The mCRc for NPM1 + and NPM1wt were 92.3% and 61.1%, respectively. mCRc for DNMT3A + and DMNT3A wt pts were 82.4% and 64.3%, respectively. WT1 + and WT1 wt mCRc were 66.7% and 77.3%, respectively. The mCRc for NPM1 + and DNMT3A + co-mutated pts was 100%. In a post hoc analysis of the 30 analyzable mCRc pts with at least one follow-up MRD assessment, 17 (56.7%) achieved molecular clearance defined as FLT3 allelic burden &lt; 10 -2. The mOS of pts achieving mCRc with FLT3 allelic burden of &lt; 10 -2 vs mCRc with FLT3 allelic burden ≥ 10 -2 (Figure 1) was not reached (95% CI: 9.76, NE) vs 6.83 mo (4.21, NE). Ven + Gilt achieved high mCRc in patients with R/R FLT3 + AML, with or without prior TKI exposure, and an encouraging mOS. FLT3 mutation clearance was seen in a majority of patients and associated with longer OS. Encouraging remission rates were observed across many genotypes and were particularly high among pts with NPM1 + +/- DNMT3A co-mutation. Cytopenias were common but manageable with appropriate Ven or Gilt dosing modifications. Serial NGS molecular data, as well as updated survival data will be presented at the meeting. Figure 1 Figure 1. Disclosures Daver: Hanmi: Research Funding; Abbvie: Consultancy, Research Funding; Trillium: Consultancy, Research Funding; Trovagene: Consultancy, Research Funding; Novimmune: Research Funding; FATE Therapeutics: Research Funding; Glycomimetics: Research Funding; Amgen: Consultancy, Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Bristol Myers Squibb: Consultancy, Research Funding; Astellas: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Gilead Sciences, Inc.: Consultancy, Research Funding; Sevier: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; ImmunoGen: Consultancy, Research Funding; Novartis: Consultancy; Jazz Pharmaceuticals: Consultancy, Other: Data Monitoring Committee member; Dava Oncology (Arog): Consultancy; Celgene: Consultancy; Syndax: Consultancy; Shattuck Labs: Consultancy; Agios: Consultancy; Kite Pharmaceuticals: Consultancy; SOBI: Consultancy; STAR Therapeutics: Consultancy; Karyopharm: Research Funding; Newave: Research Funding. Perl: Actinium: Consultancy; Genentech: Consultancy; Roche: Consultancy; Astellas: Consultancy, Research Funding; Forma: Consultancy; Syndax: Consultancy; Loxo: Consultancy; Fujifilm: Research Funding; Arog: Research Funding; BMS/Celgene: Consultancy; AbbVie: Consultancy, Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Onconova: Consultancy; Sumitomo Dainippon: Consultancy. Levis: Amgen, Astellas Pharma, Daiichi-Sankyo, FujiFilm, and Menarini: Honoraria; Astellas and FujiFilm: Research Funding; BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria; Jazz: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria. Ritchie: Astellas: Consultancy, Research Funding; Novartis: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Takeda: Consultancy, Honoraria; Celgene/BMS: Consultancy, Other: travel support, Speakers Bureau; Bristol Myers Squibb: Consultancy, Research Funding; NS Pharma: Research Funding; Incyte: Consultancy, Honoraria, Speakers Bureau; Abbvie: Consultancy, Honoraria; Jazz: Consultancy, Research Funding; Protaganist: Consultancy, Honoraria; ARIAD Pharmaceuticals: Ended employment in the past 24 months, Speakers Bureau; Pfizer: Consultancy, Research Funding. Litzow: Astellas: Research Funding; Omeros: Other: Advisory Board; Pluristem: Research Funding; AbbVie: Research Funding; Amgen: Research Funding; Actinium: Research Funding; Jazz: Other: Advisory Board; Biosight: Other: Data monitoring committee. McCloskey: Jazz: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; BMS: Consultancy, Honoraria; Amgen: Consultancy, Honoraria. Smith: AbbVie: Research Funding; Revolutions Medicine: Research Funding; Daiichi Sankyo: Consultancy; FUJIFILM: Research Funding; Astellas Pharma: Consultancy, Research Funding; Amgen: Honoraria. Schiller: Takeda: Research Funding; Trovagene: Research Funding; Tolero: Research Funding; Ono: Consultancy; Novartis: Consultancy, Research Funding; ASH foundation: Other: Chair-unpaid; Pfizer: Current equity holder in publicly-traded company, Research Funding; Incyte: Consultancy; Sanofi: Honoraria, Research Funding, Speakers Bureau; Ariad: Research Funding; Stemline Therapeutics, Inc.: Honoraria, Research Funding, Speakers Bureau; Ono-UK: Consultancy, Research Funding; Karyopharm: Research Funding; Kite/Gilead: Honoraria, Research Funding, Speakers Bureau; Onconova: Research Funding; Mateon: Research Funding; Sangamo: Research Funding; Samus: Research Funding; Regimmune: Research Funding; PrECOG: Research Funding; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; AstraZeneca: Consultancy; Kaiser Permanente: Consultancy; Cyclacel: Research Funding; MedImmune: Research Funding; Ambit: Research Funding; Agios: Consultancy, Research Funding, Speakers Bureau; Amgen: Consultancy, Current equity holder in publicly-traded company, Honoraria, Research Funding, Speakers Bureau; Jazz: Consultancy, Honoraria, Research Funding, Speakers Bureau; Elevate: Research Funding; Bio: Research Funding; Pharma: Consultancy; Johnson & Johnson: Current equity holder in publicly-traded company; Biomed Valley Discoveries: Research Funding; Eli Lilly: Research Funding; Sellas: Research Funding; Geron: Research Funding; Genentech-Roche: Research Funding; Gamida Cell Ltd.: Research Funding; FujiFilm: Research Funding; Forma: Research Funding; Delta-Fly: Research Funding; Deciphera: Research Funding; Daiichi-Sankyo: Research Funding; Constellation Pharmaceuticals: Research Funding; Celator: Research Funding; BMS/Celgene: Consultancy, Current equity holder in publicly-traded company, Research Funding, Speakers Bureau; Astellas: Honoraria, Research Funding, Speakers Bureau; Arog: Research Funding; Actuate: Research Funding; Actinium Pharmaceuticals, Inc: Research Funding; Abbvie: Research Funding; Leukemia & Lymphoma Society: Research Funding; Bluebird Bio: Research Funding; Boehringer-Ingleheim: Research Funding; Cellerant: Research Funding; CTI Biopharma: Research Funding; Janssen: Research Funding; Kura Oncology: Research Funding; Pharmacyclics: Honoraria, Speakers Bureau; Millennium: Research Funding; National Marrow Donor Program: Research Funding; NIH: Research Funding; Onyx: Research Funding; Pharmamar: Research Funding; UC Davis: Research Funding; UCSD: Research Funding; Evidera: Consultancy; NCI: Consultancy; Novartis: Speakers Bureau. Bradley: AbbVie: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees. Tiu: Astellas Pharma: Current Employment; Eli Lilly and Company: Current equity holder in publicly-traded company, Ended employment in the past 24 months. Naqvi: Genentech/Roche: Current Employment, Current holder of stock options in a privately-held company. Dail: Genentech/Roche: Current Employment, Current equity holder in publicly-traded company. Siddani: AbbVie: Current Employment, Current holder of stock options in a privately-held company. Wang: AbbVie: Current Employment, Current holder of stock options in a privately-held company. Chyla: AbbVie: Current Employment, Current equity holder in publicly-traded company. Lee: AbbVie: Current Employment, Current holder of stock options in a privately-held company. Altman: Astellas: Consultancy, Other: Advisory Board, Research Funding; AbbVie: Consultancy, Other: Advisory Board, Research Funding; Biosight: Consultancy, Other: Travel fees, Research Funding; Daiichi Sankyo: Consultancy; Kura Oncology: Consultancy; Syros: Consultancy; Theradex: Consultancy, Other: Advisory boards; GlycoMimetics: Other: Participation on an advisory board; ALZ Oncology: Research Funding; Amgen: Research Funding; Aprea: Research Funding; BMS: Research Funding; Boehringer Ingelheim: Research Funding; Fujifilm: Research Funding; Immunogen: Research Funding; Kartos: Research Funding; Kura: Research Funding. OffLabel Disclosure: Venetoclax is a B-cell lymphoma 2 inhibitor approved in combination with azacitidine, or decitabine, or low-dose cytarabine for the treatment of newly-diagnosed acute myeloid leukemia (AML) in adults who are age 75 years or older, or who have comorbidities that preclude use of intensive induction chemotherapy. Gilteritinib is a kinase inhibitor indicated for the treatment of adult patients who have relapsed or refractory acute myeloid leukemia (AML) with an FLT3 mutation as detected by an FDA-approved test.

Abstract Background : Gilteritinib, an oral FMS-like tyrosine kinase 3 (FLT3) inhibitor, is approved for the treatment of adults with FLT3-mutated (FLT3mut+) relapsed or refractory (R/R) acute myeloid leukemia (AML) in the United States and many other countries/regions. However, not all respond to treatment and most patients eventually develop recurrent disease. Combining gilteritinib with other agents may improve response. Atezolizumab (840 mg intravenous [IV] every 2 weeks [Q2W]) + azacitidine demonstrated an overall response rate of 62% in hypomethylating agent-naive patients with higher-risk myelodysplastic syndrome (Gerds AT, et al. Blood. 2018;132[suppl 1]:466). Therefore, the safety and efficacy of combination therapy with gilteritinib and atezolizumab was investigated in an ongoing phase 1, open-label, single-arm, dose-escalation study (ClinicalTrials.gov identifier: NCT03730012) in adult patients with FLT3mut+ R/R AML. Methods : This phase 1 dose-escalation study enrolled adults with FLT3mut+ AML and Eastern Cooperative Oncology Group performance status of ≤2 who were refractory to ≥1 cycle of induction chemotherapy or relapsed after achieving remission with a prior therapy. Key exclusion criteria included AML secondary to prior chemotherapy for other neoplasms (except for myelodysplastic syndrome) and patients who have relapsed after allogeneic hematopoietic stem cell transplantation. Patients received gilteritinib 120 mg/day combined with atezolizumab 420 mg or 840 mg via IV infusion Q2W in 28-day cycles. Herein, we present safety and tolerability (dose-limiting toxicities [DLT] and treatment-emergent adverse events [TEAE]; primary end points). Decisions regarding DLTs and recommended phase 2 dose are determined by a dose evaluation committee. Composite complete remission (CRc) rate (primary end point), best response rate (secondary end point), and gilteritinib trough plasma concentrations (C trough; secondary end point) were also evaluated. Results : As of 13 June 2021, 3 patients received gilteritinib 120 mg/day + atezolizumab 420 mg Q2W (cohort 1) and 8 patients received gilteritinib 120 mg/day + atezolizumab 840 mg Q2W (cohort 2). Median (range) age was 82.0 (68-84) and 66.5 (20-87) years in cohorts 1 and 2, respectively. No patients received prior second-generation FLT3 inhibitor therapy. The median duration of gilteritinib exposure was 107.0 and 49.5 days in cohorts 1 and 2, respectively. Seven patients received ≥2 cycles of atezolizumab. No DLTs were reported in cohort 1; 2 patients (25%) reported DLTs in cohort 2 (1 occurrence each of increased alanine aminotransferase and encephalopathy). The most common TEAEs (≥30%) across both cohorts were febrile neutropenia (72.7%); fatigue (54.5%); dyspnea, muscular weakness, and decreased platelet count (45.5% each); and anemia, decreased appetite, diarrhea, dizziness, epistaxis, fall, and pyrexia (36.4% each; Table). Serious TEAEs were reported in 10 patients (91.0%). Treatment-related adverse events (TRAEs) were reported in 10 patients (90.9%), of which 9 patients (81.8%) reported serious TRAEs (only event reported in &gt;1 patient was febrile neutropenia [54.5%]). Study treatment was withdrawn for 8 patients (72.7%) due to TEAEs, of which TEAEs were considered TRAEs in 6 patients (54.5%). TEAEs lead to death in 3 patients (27.3%). Efficacy results will be provided at the time of presentation. Pharmacokinetic analyses suggested that C trough of gilteritinib in combination with atezolizumab (420 mg or 840 mg Q2W) was similar to single-agent gilteritinib in patients with R/R AML. Cohorts 1 and 2 showed similar gilteritinib C trough values. Conclusions: In this phase 1 dose-escalation study, the combination of gilteritinib and atezolizumab had an acceptable safety profile with no new safety signals identified for either agent. Owing to a strategic decision, the expansion phase of the study will not be conducted. The combinatorial prospect of gilteritinib observed in this study supports future studies of gilteritinib combination therapy. Figure 1 Figure 1. Disclosures Altman: Kuro Oncology: Consultancy; Syros: Consultancy; Daiichi Sankyo: Consultancy; Astellas: Consultancy; AbbVie: Consultancy; Glycomimetics: Membership on an entity's Board of Directors or advisory committees; BioSight: Consultancy, Other: Travel fees to attend an advisory meeting (I did not accept payment for the advisory board); Theradex: Consultancy. Bhatnagar: Sumitomo Dainippon Pharma: Research Funding; Novartis: Honoraria; Karyopharm Therapeutics Inc.: Honoraria, Research Funding; Astellas: Honoraria; Pfizer: Honoraria; Kite: Honoraria; Cell Therapeutics: Honoraria, Research Funding; Celgene: Honoraria. Abedin: Agios: Honoraria; AltruBio: Research Funding; Actinium: Research Funding; Amgen: Honoraria; Helsinn: Research Funding; Pfizer: Research Funding; Astellas Pharma Inc.: Research Funding. Przespolewski: Jazz: Research Funding. Schiller: Agios: Consultancy, Research Funding, Speakers Bureau; Kaiser Permanente: Consultancy; Leukemia & Lymphoma Society: Research Funding; Bio: Research Funding; Tolero: Research Funding; Forma: Research Funding; Takeda: Research Funding; Delta-Fly: Research Funding; Actuate: Research Funding; Jazz: Consultancy, Honoraria, Research Funding, Speakers Bureau; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Elevate: Research Funding; Deciphera: Research Funding; Cyclacel: Research Funding; MedImmune: Research Funding; Johnson & Johnson: Current equity holder in publicly-traded company; Genentech-Roche: Research Funding; Gamida Cell Ltd.: Research Funding; Sanofi: Honoraria, Research Funding, Speakers Bureau; Eli Lilly: Research Funding; Actinium Pharmaceuticals, Inc: Research Funding; Abbvie: Research Funding; Arog: Research Funding; Biomed Valley Discoveries: Research Funding; Pharma: Consultancy; ASH foundation: Other: Chair-unpaid; Novartis: Consultancy, Research Funding; Sellas: Research Funding; Regimmune: Research Funding; PrECOG: Research Funding; Samus: Research Funding; Pfizer: Current equity holder in publicly-traded company, Research Funding; Onconova: Research Funding; Sangamo: Research Funding; Mateon: Research Funding; Karyopharm: Research Funding; Kite/Gilead: Honoraria, Research Funding, Speakers Bureau; FujiFilm: Research Funding; Ambit: Research Funding; Stemline Therapeutics, Inc.: Honoraria, Research Funding, Speakers Bureau; Geron: Research Funding; Amgen: Consultancy, Current equity holder in publicly-traded company, Honoraria, Research Funding, Speakers Bureau; Trovagene: Research Funding; Ono-UK: Consultancy, Research Funding; Ono: Consultancy; Incyte: Consultancy; Ariad: Research Funding; AstraZeneca: Consultancy; Daiichi-Sankyo: Research Funding; Constellation Pharmaceuticals: Research Funding; Celator: Research Funding; BMS/Celgene: Consultancy, Current equity holder in publicly-traded company, Research Funding, Speakers Bureau; Astellas: Honoraria, Research Funding, Speakers Bureau; Bluebird Bio: Research Funding; Boehringer-Ingleheim: Research Funding; Cellerant: Research Funding; CTI Biopharma: Research Funding; Janssen: Research Funding; Kura Oncology: Research Funding; Pharmacyclics: Honoraria, Speakers Bureau; Millennium: Research Funding; National Marrow Donor Program: Research Funding; NIH: Research Funding; Onyx: Research Funding; Pharmamar: Research Funding; UC Davis: Research Funding; UCSD: Research Funding; Evidera: Consultancy; NCI: Consultancy; Novartis: Speakers Bureau. Gill: Astellas Pharma Global Development: Current Employment. Patel: Astellas Pharma Global Development: Current Employment. Fan: Astellas Pharma Global Development: Current Employment. Tiu: Astellas Pharma Global Development: Current Employment. Strickland: Sunesis: Research Funding; AbbVie: Other: Advisory Board; ArcherDx: Other: Advisory Board; Astellas: Other: Advisory Board; Genentech: Other: Advisory Board; Incyte: Other: Advisory Board; Jazz: Other: Advisory Board; Kite: Other: Advisory Board; Kura Oncology: Other: Advisory Board; Novartis: Other: Advisory Board; Pfizer: Other: Advisory Board; Syros: Other: Advisory Board. OffLabel Disclosure: New indication for atezolizumab

Abstract Background: Despite the demonstrated efficacy of ruxolitinib (potent and selective Janus kinase [JAK] 1 and JAK2 inhibitor) in patients with MF, inadequate responses or loss of response to ruxolitinib may occur, possibly due to persistent activation of the phosphatidylinositol 3-kinase (PI3K) pathway with chronic ruxolitinib therapy. Parsaclisib (INCB050465) is a potent and highly selective next-generation PI3Kδ inhibitor. We previously demonstrated preliminary efficacy in a phase 2 trial (INCB 50465-201, NCT02718300) of parsaclisib added to stable doses of ruxolitinib for patients with MF who experienced a suboptimal response to ruxolitinib. JAK inhibitors, including ruxolitinib, are associated with thrombocytopenia; therefore, patients with low platelet counts (PC) are traditionally more difficult to treat. We present the efficacy and safety subgroup analysis of this ongoing study in patients grouped by baseline PC. Methods: Eligible adults had primary or secondary (post-polycythemia vera or post-essential thrombocythemia) MF with suboptimal response (palpable spleen &gt;10 cm below left subcostal margin, or palpable splenomegaly 5-10 cm below left subcostal margin and presence of 1 symptom score ≥5 or 2 symptom scores ≥3 each using the Screening Symptom Form) after ≥6 months of ruxolitinib monotherapy (5-25 mg twice daily, stable dose for ≥8 weeks). Patients remained on their last stable ruxolitinib dose and received add-on parsaclisib 10 mg or 20 mg once-daily (QD) for 8 weeks and the same dose once-weekly thereafter (daily to weekly group), or parsaclisib 5 mg or 20 mg QD for 8 weeks and 5 mg QD thereafter (all daily group). For the subgroup analysis, patients were grouped by baseline PC (≥100×10 9/L, higher PC or 50-&lt;100×10 9/L, low PC). Key objectives were to evaluate the impact of baseline PC on spleen volume (SV) and total symptom score (TSS) as assessed by Myelofibrosis-Symptoms Assessment Form (MFSAF) v3.0 daily diary at week 12 and week 24, and safety. Results: At data cutoff (Aug 27, 2020), 67 patients were enrolled, 21 with low PC and 46 with higher PC. Median age of both groups was 68 years, and median prior duration of ruxolitinib use was 34.7 months for low PC and 14.9 months for higher PC. Patients with low PC had higher baseline symptoms (MFSAF-TSS median [range], 21.4 [0.6-47]) than patients with higher PC (MFSAF-TSS, 10 [0-43]). Responder analysis for SV reduction (SVR) is summarized in Table 1. At week 12, slightly more patients with low PC achieved ≥10% SVR compared with patients with higher PC, whereas, at week 24, responses were similar between the 2 groups. Of patients with at least 10% SVR at week 24, 4 of 6 with low PC and 9 of 13 with higher PC were receiving all daily dose regimens. Median (range) percentage change in MFSAF-TSS was −20.5 (−56.6 to +17.1) and −22.2 (−100 to +500) at week 12, and −26.1 (−54.7 to +2.4) and −23.1 (−91.3 to +222.5) at week 24, for patients with low PC and higher PC, respectively. Overall for both baseline platelet groups, nonhematologic treatment-emergent adverse events (TEAEs) were primarily grade 1/2. Most common (≥20%) TEAEs were dyspnea (33%), falls (33%), peripheral edema (29%), and nasal congestion (24%) for patients in the low PC group, and diarrhea (28%), nausea (24%), abdominal pain (24%), cough (20%), and fatigue (20%) for patients with higher PC. Interruption of parsaclisib due to thrombocytopenia was observed in 9 of the 21 patients (43%) with low PC and 3 of the 46 patients (7%) with higher PC. One patient with low PC had thrombocytopenia leading to ruxolitinib interruption. Conclusion: Add-on parsaclisib demonstrated promising preliminary efficacy in patients with MF experiencing suboptimal response to ruxolitinib monotherapy. In a subgroup analysis by baseline PC, responses for SV and MFSAF-TSS reduction were similar in both groups, indicating that patients with low PC can also tolerate and benefit from this treatment combination. Phase 3 trials in ruxolitinib-experienced and ruxolitinib-naive patients are underway to further assess the combination of JAK and PI3K inhibitors. Figure 1 Figure 1. Disclosures Yacoub: Agios: Membership on an entity's Board of Directors or advisory committees; Acceleron Pharma: Membership on an entity's Board of Directors or advisory committees; CTI Biopharma: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Incyte: Speakers Bureau. Borate: Takeda: Membership on an entity's Board of Directors or advisory committees; incyte: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Astellas: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; Blueprint Medicine: Membership on an entity's Board of Directors or advisory committees; Genentech: Membership on an entity's Board of Directors or advisory committees, Other: Advisory Board; Jazz Pharma: Research Funding; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Rampal: Membership on an entity's Board of Directors or advisory committees; Galecto, Inc.: Consultancy; Promedior: Consultancy. Rampal: Blueprint: Consultancy; Jazz Pharmaceuticals: Consultancy; Disc Medicine: Consultancy; Constellation: Research Funding; Stemline: Consultancy, Research Funding; Kartos: Consultancy; BMS/Celgene: Consultancy; Novartis: Consultancy; Sierra Oncology: Consultancy; CTI: Consultancy; Abbvie: Consultancy; Memorial Sloan Kettering: Current Employment; Incyte: Consultancy, Research Funding; Pharmaessentia: Consultancy. Ali: CTI BioPharma: Membership on an entity's Board of Directors or advisory committees; Incyte: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; BMS: Speakers Bureau. Wang: Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees; Daiichi Sankyo: Consultancy, Honoraria, Other: Advisory board; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Stemline Therapeutics: Consultancy, Honoraria, Other: Advisory board, Speakers Bureau; Genentech: Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy, Honoraria, Other: Advisory board; Jazz Pharmaceuticals: Consultancy, Honoraria, Other: Advisory Board; Kite Pharmaceuticals: Consultancy, Honoraria, Other: Advisory Board; Rafael Pharmaceuticals: Other: Data safety monitoring committee; BMS/Celgene: Membership on an entity's Board of Directors or advisory committees; GlaxoSmithKline: Consultancy, Honoraria, Other: Advisory Board; DAVA Oncology: Consultancy, Speakers Bureau; Takeda: Consultancy, Honoraria, Other: Advisory board; Pfizer: Consultancy, Honoraria, Other: Advisory Board, Speakers Bureau; Mana Therapeutics: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Other: Advisory Board; Kura Oncology: Consultancy, Honoraria, Other: Advisory board, steering committee, Speakers Bureau; PTC Therapeutics: Consultancy, Honoraria, Other: Advisory board; Genentech: Consultancy; MacroGenics: Consultancy. Gerds: CTI BioPharma: Research Funding; Sierra Oncology: Consultancy; AbbVie: Consultancy; Celgene/Bristol Myers Squibb: Consultancy; Constellation: Consultancy; PharmaEssentia Corporation: Consultancy; Novartis: Consultancy. Hobbs: Incyte Corporation: Research Funding; AbbVie.: Consultancy; Celgene/Bristol Myers Squibb: Consultancy; Novartis: Consultancy; Merck: Research Funding; Constellation Pharmaceuticals: Consultancy, Research Funding; Bayer: Research Funding. Kremyanskaya: Chimerix: Research Funding; Constellation: Research Funding; Protagonist Therapeutics: Consultancy, Research Funding; Incyte: Research Funding; Bristol Myers Squibb: Research Funding; Astellas: Research Funding; Astex: Research Funding. Winton: Samus Therapeutics: Research Funding; Incyte Corporation: Research Funding; Blueprint Medicines: Research Funding. O'Connell: Pfizer: Consultancy; Bristol Myers Squibb: Consultancy; Genentech: Research Funding; Astex Pharmaceuticals: Consultancy, Research Funding; Shionogi: Consultancy. Oh: Novartis: Consultancy; Kartos Therapeutics: Consultancy; Incyte Corporation: Consultancy; Geron: Consultancy; Disc Medicine: Consultancy; CTI BioPharma: Consultancy; Constellation: Consultancy; Celgene/Bristol Myers Squibb: Consultancy; Blueprint Medicines: Consultancy; Abbvie: Consultancy; PharmaEssentia: Consultancy; Sierra Oncology: Consultancy. Schiller: Kura Oncology: Research Funding; Pharmacyclics: Honoraria, Speakers Bureau; Evidera: Consultancy; NCI: Consultancy; UCSD: Research Funding; Eli Lilly: Research Funding; Onyx: Research Funding; UC Davis: Research Funding; Ambit: Research Funding; MedImmune: Research Funding; Bluebird Bio: Research Funding; ASH foundation: Other: Chair-unpaid; Ono-UK: Consultancy, Research Funding; Takeda: Research Funding; Leukemia & Lymphoma Society: Research Funding; Kaiser Permanente: Consultancy; Samus: Research Funding; Boehringer-Ingleheim: Research Funding; Millennium: Research Funding; Ariad: Research Funding; Cellerant: Research Funding; Janssen: Research Funding; Agios: Consultancy, Research Funding, Speakers Bureau; Stemline Therapeutics, Inc.: Honoraria, Research Funding, Speakers Bureau; Trovagene: Research Funding; Bio: Research Funding; National Marrow Donor Program: Research Funding; Sellas: Research Funding; Ono: Consultancy; Cyclacel: Research Funding; CTI Biopharma: Research Funding; Tolero: Research Funding; Pharma: Consultancy; AstraZeneca: Consultancy; NIH: Research Funding; Sanofi: Honoraria, Research Funding, Speakers Bureau; Novartis: Consultancy, Research Funding; Johnson & Johnson: Current equity holder in publicly-traded company; Sangamo: Research Funding; Biomed Valley Discoveries: Research Funding; Incyte: Consultancy; Amgen: Consultancy, Current equity holder in publicly-traded company, Honoraria, Research Funding, Speakers Bureau; Regimmune: Research Funding; Pharmamar: Research Funding; Jazz: Consultancy, Honoraria, Research Funding, Speakers Bureau; Elevate: Research Funding; PrECOG: Research Funding; Pfizer: Current equity holder in publicly-traded company, Research Funding; Onconova: Research Funding; Mateon: Research Funding; Kite/Gilead: Honoraria, Research Funding, Speakers Bureau; Karyopharm: Research Funding; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Geron: Research Funding; Genentech-Roche: Research Funding; Gamida Cell Ltd.: Research Funding; FujiFilm: Research Funding; Forma: Research Funding; Delta-Fly: Research Funding; Deciphera: Research Funding; Daiichi-Sankyo: Research Funding; Constellation Pharmaceuticals: Research Funding; Celator: Research Funding; BMS/Celgene: Consultancy, Current equity holder in publicly-traded company, Research Funding, Speakers Bureau; Astellas: Honoraria, Research Funding, Speakers Bureau; Arog: Research Funding; Actuate: Research Funding; Actinium Pharmaceuticals, Inc: Research Funding; Abbvie: Research Funding; Novartis: Speakers Bureau. Assad: Incyte: Current Employment, Current equity holder in publicly-traded company. Erickson-Viitanen: Incyte: Current Employment, Current equity holder in publicly-traded company. Zhou: Incyte: Current Employment, Current equity holder in publicly-traded company. Daver: Pfizer: Consultancy, Research Funding; Bristol Myers Squibb: Consultancy, Research Funding; Sevier: Consultancy, Research Funding; Astellas: Consultancy, Research Funding; Jazz Pharmaceuticals: Consultancy, Other: Data Monitoring Committee member; Novartis: Consultancy; Gilead Sciences, Inc.: Consultancy, Research Funding; Trovagene: Consultancy, Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Abbvie: Consultancy, Research Funding; ImmunoGen: Consultancy, Research Funding; Syndax: Consultancy; Agios: Consultancy; Dava Oncology (Arog): Consultancy; FATE Therapeutics: Research Funding; Novimmune: Research Funding; Amgen: Consultancy, Research Funding; Celgene: Consultancy; Trillium: Consultancy, Research Funding; Glycomimetics: Research Funding; Shattuck Labs: Consultancy; Hanmi: Research Funding; Genentech: Consultancy, Research Funding; Kite Pharmaceuticals: Consultancy; SOBI: Consultancy; STAR Therapeutics: Consultancy; Karyopharm: Research Funding; Newave: Research Funding.

Abstract Background: In vitro studies and emerging clinical data suggest that inhibition of spleen tyrosine kinase may have an antileukemic effect in human AML. Pts with AML and TP53 mutations (TP53m) are commonly associated with older age (≥60 years) and complex karyotype (CK) and respond poorly to standard 7 + 3 induction (IND) chemotherapy with &lt;10% 1-year overall survival (OS). DEC has been reported to improve response in TP53m AML. Here, we report the results of a Phase 2 sub-study of the Beat AML Master Trial that assessed the efficacy of ENTO + DEC combination treatment (Tx) in ND AML patients (pts) with TP53m or CK aged ≥60 years. [ClinicalTrials.gov: NCT03013998] Methods: This multicenter (13 sites), open-label, Phase 2 combination Tx study utilized Simon's 2-stage Phase 2 design and enrolled AML pts with TP53m (identified molecularly) ± CK (Cohort A) or CK (≥3 metaphase abnormalities) without TP53m (Cohort B). Pts initially received 5 days of ENTO lead-in (which was later discontinued), followed by ENTO + DEC and those who achieved CR/CRh/CRi/MLFS with up to 3 cycles of IND proceeded to consolidation (CON) Tx for up to 11 cycles (Figure 1). Pts with CRi/MLFS after IND were allowed up to 6 cycles (IND + CON) to achieve CR/CRh or stayed on Tx if they got clinical benefit or went off Tx. CON was followed by maintenance (MTN) Tx for up to 2 years from start of study Tx. Pts were eligible if aged ≥60 years, ND, and had ECOG performance status 0 - 2. Pts received ENTO 400 mg orally twice daily for 5 days during ENTO lead-in, and then every 28 days during IND, CON, and MTN + DEC 20 mg/m 2 IV days 1-10 (IND) or days 1-5 (CON) every 28 days. Response was assessed using modified 2017 ELN AML criteria. The primary endpoint was composite complete remission (CCR) rate (CR + CRh) with up to 3 cycles of IND, and CRi/MLFS that achieved CR/CRh by up to 6 cycles (IND + CON). Beyond stage 1, pt accrual to Cohort A was allowed based on pts with CRi and to Cohort B was stopped early for futility. Results: Between Oct 2017 and Feb 2020, of the 63 pts enrolled (Cohort A = 48; Cohort B = 15), pts with confirmed eligibility who started study Tx were included in the analyses (Cohort A = 45; Cohort B = 13). During lead-in, 27 pts in Cohort A and 6 pts in Cohort B received ENTOm for 5 days. All pts received ENTO + DEC except 1 pt in Cohort A who withdrew consent (WOC). Median ages of the pts were 70 years (range 60 - 84) in Cohort A and 74 years (range 65 - 86) in Cohort B. Median time (range) on Tx was 2.2 mos and 4.8 mos in Cohort A and B, respectively. Most common reasons for Tx discontinuation were adverse event (AE; 27%), Tx failure (TF; 27%) and WOC (18%) in Cohort A; TF (31%), disease progression and relapse (each 15%) in Cohort B. In each cohort, 1 pt discontinued Tx due to death from leukemia and 1 pt in Cohort A in CRh due to development of an additional genetic abnormality. Four pts (9%) in Cohort A and 1 pt (8%) in Cohort B proceeded to transplant. The CCR (CR + CRh) rates with up to 6 cycles of Tx for Cohort A and B were 13.3% and 30.8%, respectively; overall CR + CRh rates were 17.8% and 38.5% (Table 1). In Cohort A, with a median follow-up of 11.5 months, 0% were 1-year disease-free and median OS (mOS) was 6.5 months. In Cohort B, with a median follow-up of 15.1 months, 25% were 1-year disease-free and mOS was 11.5 months. Deaths within 7-, 30-, and 60-days of Tx were 0, 3 and 11 in Cohort A and 0, 0 and 2 in Cohort B. Most common treatment-related Grade ≥3 AEs in Cohort A and B were febrile neutropenia (31% and 39%) and anemia (22% and 31%) (Table 2). Overall, 83 serious AEs (SAEs) were reported in 33 pts in Cohort A and 12 SAEs in 6 pts in Cohort B; most common SAEs in Cohort A were pneumonia (18%) and respiratory failure (11%), and in Cohort B sepsis, dehydration and acute kidney injury (each 15%). Most common treatment-related grade ≥3 laboratory abnormalities in Cohort A and B were neutrophils decreased (27% and 31%), WBC count decreased (20% and 23%), and lymphocyte count decreased (18% and 15%). Conclusions: ENTO + DEC demonstrated activity in ND AML pts aged ≥60 years with TP53m ± CK and CK without TP53 but induced low CR/CRh rates and short OS consistent with previously published poor CR rates and OS in these pts. Our results differ from the high remission rate and longer OS previously reported for DEC monotherapy in AML pts with TP53m. ENTO + DEC was safe and acceptably tolerated. Novel Tx strategies that can benefit AML pts with these most adverse risk factors are urgently needed. Figure 1 Figure 1. Disclosures Ruppert: Telios Pharma: Consultancy. Mims: Leukemia and Lymphoma Society's Beat AML clinical study: Consultancy, Research Funding; Aptevo: Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Glycomemetics: Research Funding; Kartos Pharmaceuticals: Research Funding; Xencor: Research Funding; Genentech: Consultancy; Abbvie: Consultancy; BMS: Consultancy; Kura Oncology: Consultancy; Syndax Pharmaceuticals: Consultancy; BMS: Consultancy; Jazz Pharmaceuticals: Consultancy; Aptevo: Research Funding. Borate: Jazz Pharma: Research Funding; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Blueprint Medicine: Membership on an entity's Board of Directors or advisory committees; Genentech: Membership on an entity's Board of Directors or advisory committees, Other: Advisory Board; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; Astellas: Membership on an entity's Board of Directors or advisory committees; incyte: Membership on an entity's Board of Directors or advisory committees, Research Funding; Rampal: Membership on an entity's Board of Directors or advisory committees; Galecto, Inc.: Consultancy; Promedior: Consultancy. Stein: Abbvie: Consultancy; Janssen Pharmaceuticals: Consultancy; Gilead Sciences, Inc.: Consultancy; Foghorn Therapeutics: Consultancy; Syros Pharmaceuticals, Inc.: Consultancy; Daiichi Sankyo: Consultancy; Blueprint Medicines: Consultancy; PinotBio: Consultancy; Genentech: Consultancy; Jazz Pharmaceuticals: Consultancy; Bristol Myers Squibb: Consultancy; Celgene: Consultancy; Agios Pharmaceuticals, Inc: Consultancy; Novartis: Consultancy; Astellas: Consultancy; Syndax Pharmaceuticals: Consultancy. Stock: Pfizer: Consultancy, Honoraria, Research Funding; amgen: Honoraria; agios: Honoraria; jazz: Honoraria; kura: Honoraria; kite: Honoraria; morphosys: Honoraria; servier: Honoraria; syndax: Consultancy, Honoraria; Pluristeem: Consultancy, Honoraria. Kovacsovics: AbbVie: Research Funding; Janssen Pharmaceuticals: Research Funding; Amgen Inc.: Research Funding; Novartis: Research Funding; Stemline: Honoraria; Jazz Pharmaceutials: Honoraria. Blum: Amerisource Bergen; Abbvie, Syndax: Honoraria; Forma Therapeutics, Xencor; Celyad: Research Funding. Arellano: KITE Pharma, Inc: Consultancy; Syndax Pharmaceuticals, Inc: Consultancy. Schiller: Actinium Pharmaceuticals, Inc: Research Funding; Gamida Cell Ltd.: Research Funding; Pfizer: Current equity holder in publicly-traded company, Research Funding; Forma: Research Funding; Agios: Consultancy, Research Funding, Speakers Bureau; Constellation Pharmaceuticals: Research Funding; MedImmune: Research Funding; Ambit: Research Funding; Karyopharm: Research Funding; Daiichi-Sankyo: Research Funding; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Kaiser Permanente: Consultancy; Abbvie: Research Funding; AstraZeneca: Consultancy; Genentech-Roche: Research Funding; Delta-Fly: Research Funding; Cyclacel: Research Funding; Mateon: Research Funding; Actuate: Research Funding; Onconova: Research Funding; Geron: Research Funding; Sangamo: Research Funding; Arog: Research Funding; Ariad: Research Funding; Stemline Therapeutics, Inc.: Honoraria, Research Funding, Speakers Bureau; Takeda: Research Funding; Trovagene: Research Funding; Ono-UK: Consultancy, Research Funding; Tolero: Research Funding; BMS/Celgene: Consultancy, Current equity holder in publicly-traded company, Research Funding, Speakers Bureau; Celator: Research Funding; Kite/Gilead: Honoraria, Research Funding, Speakers Bureau; Astellas: Honoraria, Research Funding, Speakers Bureau; Incyte: Consultancy; Novartis: Consultancy, Research Funding; Deciphera: Research Funding; FujiFilm: Research Funding; Samus: Research Funding; Regimmune: Research Funding; PrECOG: Research Funding; Amgen: Consultancy, Current equity holder in publicly-traded company, Honoraria, Research Funding, Speakers Bureau; Bio: Research Funding; Elevate: Research Funding; Jazz: Consultancy, Honoraria, Research Funding, Speakers Bureau; Biomed Valley Discoveries: Research Funding; Ono: Consultancy; Eli Lilly: Research Funding; Sellas: Research Funding; ASH foundation: Other: Chair-unpaid; Leukemia & Lymphoma Society: Research Funding; Bluebird Bio: Research Funding; Sanofi: Honoraria, Research Funding, Speakers Bureau; Pharma: Consultancy; Johnson & Johnson: Current equity holder in publicly-traded company; Boehringer-Ingleheim: Research Funding; Cellerant: Research Funding; CTI Biopharma: Research Funding; Janssen: Research Funding; Kura Oncology: Research Funding; Pharmacyclics: Honoraria, Speakers Bureau; Millennium: Research Funding; National Marrow Donor Program: Research Funding; NIH: Research Funding; Onyx: Research Funding; Pharmamar: Research Funding; UC Davis: Research Funding; UCSD: Research Funding; Evidera: Consultancy; NCI: Consultancy; Novartis: Speakers Bureau. Olin: Astellas: Honoraria, Research Funding; Actinium: Honoraria; Amgen: Honoraria; Abbvie: Honoraria; Cellectis: Research Funding; Daiichi Sankyo: Research Funding; Genentech: Research Funding; Pfizer: Research Funding. Foran: certara: Honoraria; pfizer: Honoraria; syros: Honoraria; taiho: Honoraria; boehringer ingelheim: Research Funding; servier: Honoraria; revolution medicine: Honoraria; trillium: Research Funding; takeda: Research Funding; abbvie: Research Funding; novartis: Honoraria; bms: Honoraria; OncLive: Honoraria; gamida: Honoraria; sanofi aventis: Honoraria; aptose: Research Funding; actinium: Research Funding; kura: Research Funding; h3bioscience: Research Funding; aprea: Research Funding; sellas: Research Funding; stemline: Research Funding. Litzow: AbbVie: Research Funding; Omeros: Other: Advisory Board; Astellas: Research Funding; Pluristem: Research Funding; Jazz: Other: Advisory Board; Actinium: Research Funding; Amgen: Research Funding; Biosight: Other: Data monitoring committee. Lin: AbbVie, Aptevo Therapeutics, Astellas Pharma, Bio-Path Holdings, Celgene, Celyad, Genentech-Roche, Gilead Sciences, Incyte, Jazz Pharmaceuticals, Novartis, Ono Pharmaceutical, Pfizer, Prescient Therapeutics, Seattle Genetics, Tolero, Trovagene: Research Funding. Cogle: Aptevo therapeutics: Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees. Vergilio: Foundation Medicine: Current Employment; Roche: Current equity holder in publicly-traded company. Gana: Bausch: Current holder of individual stocks in a privately-held company; The Leukemia & Lymphoma Society: Consultancy. Druker: The RUNX1 Research Program: Membership on an entity's Board of Directors or advisory committees; Aileron: Membership on an entity's Board of Directors or advisory committees; Novartis Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Recludix Pharma, Inc.: Consultancy; Third Coast Therapeutics: Membership on an entity's Board of Directors or advisory committees; Vincerx Pharma: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; VB Therapeutics: Membership on an entity's Board of Directors or advisory committees; GRAIL: Current equity holder in publicly-traded company; Iterion Therapeutics: Membership on an entity's Board of Directors or advisory committees; ALLCRON: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Aptose Therapeutics: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Blueprint Medicines: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Research Funding; EnLiven: Consultancy, Research Funding; Merck & Co: Patents & Royalties; Pfizer: Research Funding; Nemucore Medical Innovations, Inc.: Consultancy; Cepheid: Consultancy, Membership on an entity's Board of Directors or advisory committees; Vivid Biosciences: Membership on an entity's Board of Directors or advisory committees. Byrd: Vincerx Pharmaceuticals: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Novartis, Trillium, Astellas, AstraZeneca, Pharmacyclics, Syndax: Consultancy, Honoraria; Newave: Membership on an entity's Board of Directors or advisory committees. Levine: Isoplexis: Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria; Zentalis: Membership on an entity's Board of Directors or advisory committees; Imago: Membership on an entity's Board of Directors or advisory committees; Ajax: Membership on an entity's Board of Directors or advisory committees; Lilly: Honoraria; Prelude: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria; Janssen: Consultancy; Celgene: Research Funding; Roche: Honoraria, Research Funding; Auron: Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy; Astellas: Consultancy; Morphosys: Consultancy; QIAGEN: Membership on an entity's Board of Directors or advisory committees; Mission Bio: Membership on an entity's Board of Directors or advisory committees. OffLabel Disclosure: Off-label use of entospletinib and decitabine.

Abstract Background: GILT is an oral potent selective FLT3 kinase inhibitor approved for marketing for the treatment (Tx) of patients (pts) with relapsed/refractory FLT3 mutated (FLT3m) AML but efficacy in older ND FLT3m AML pts is unknown. Furthermore, FLT3m can be present as a dominant or subclone and impact of FLT3 inhibitor therapy in this setting is uncertain. Here we report the results of a Phase 2/1b sub-study of the Beat AML Master Trial to assess the efficacy of GILT monotherapy (GILTm) in ND FLT3m AML pts aged ≥60 years with high and low VAF and the subsequent response-driven addition of DEC Tx. (ClinicalTrials.gov NCT03013998) Methods: The study was an open-label multicenter (15 sites), 3-outcome, 2-stage Phase 2 design that assigned pts to either Dominant FLT3/Group 1 (GP1) or Non-Dominant FLT3/Group 2 (GP2) as shown in Figure 1. Key eligibility criteria included ND FLT3m AML pts with high and low VAF and/or ITD ratio, aged ≥60 years, and ECOG performance status 0-2. In the Phase 2 study, all pts received GILTm 120 mg/day on days 1 - 28. Pts without CR/CRi after cycle 2 were transferred to the Phase 1b study to receive GILT + DEC (Figure 1). Phase 1b study utilized a standard 3+3 design to evaluate the safety/tolerability of concurrent GILT + DEC. Pts received GILT (dose level 1 [DL1] = 80 mg/day or dose level 2 [DL2] = 120 mg/day on days 1-28) + DEC 20 mg/m 2 IV on days 1-10 or 1-5 every 28 days. Primary endpoint was CR+CRi rate (Phase 2). Response was assessed using modified 2017 ELN AML criteria. The non-dominant GP2 was stopped for futility, GP1 was stopped early to modify trial to include venetoclax. Results: Phase 2 - Between 9/10/2018 to 2/11/2020, 19 / 20 enrolled pts (GP1: n = 9; GP2: n = 10) received GILTm and were included in analyses. Baseline pt characteristics are shown in Table 1. Median (range) time on GILTm was 3 cycles (1 - 18) in GP1 and 1 cycle (1 - 9) in GP2. Most common reasons for discontinuing Tx were Tx failure (TF; 44%) and relapse (33%) in GP1 and TF (70%) and disease progression (PD; 20%) in GP2. Overall CR+CRi was achieved in 4 pts (44%) in GP1 and 1 pt (10%) in GP2. Response duration are shown in Table 2. After median follow-up of 14.3 months (mos) and 19 mos in GP1 and GP2, respectively, 1-year OS was 56% and 76%. Most common Grade ≥3 adverse events (AEs) were febrile neutropenia and colitis (each 25%) in GP1; anemia and low platelet count in GP2 (each 30%). Overall, 7 pts had 15 serious AEs (SAEs) and all SAEs occurred in GP1 pts; most common SAE was colitis (25%) and 1 pt (13%) had a Tx-related Grade 3 SAE of tumor lysis syndrome. In GP2, 1 pt (10%) had Tx-related Grade 2 AE of differentiation syndrome. In GP1, 2 pts died within 60 days of Tx and none in GP2. Phase1b - After up to 2 cycles of GILTm, 12 pts with no CR/CRi (GP1: n = 4; GP2: n = 8) were transferred to receive GILT + DEC (Figure 1). At the time of this report, 1 pt with CRh remained on Tx. Median total time on Tx (including GILTm) was 4 cycles and median time on GILT + DEC Tx was 3 cycles (Table 2). Most common reasons for discontinuing Tx were PD (33%) and TF (25%); and 2 pts (17%) stopped Tx due to an AE. Pts were treated with DL1 GILT + DEC (n = 3), then DL2 GILT + DEC (n = 9); only 1 pt had dose-limiting toxicity (DLT) at DL2 (Grade 3 hyperbilirubinemia and pneumonitis requiring steroid therapy), hence, DL2 GILT + DEC was considered the MTD. CR+CRi rate was 25% in 3 pts, all at DL2 (Table 2). After a median follow-up of 17.8 mos, the 1-year OS from start of GILT + DEC Tx was 57%. Most common Grade ≥3 Tx-related AEs were anemia, febrile neutropenia and low WBC count (each 22%). Overall, 6 pts had 12 SAEs; 1 pt with SAE of Grade 4 sepsis died. Three GILT-related SAEs occurred in 1 pt - Grade 3 hyperbilirubinemia, and pneumonitis and Grade 1 transaminases increased. One pt died within 30 days and a second within 60 days of Tx. No difference was observed in GILT pharmacokinetics (PK) with or without DEC, however steady state Ctrough values were 1.4 to 2.3-fold greater than in relapsed/refractory AML pts (Admiral trial). Conclusions: In ND pts ≥60 years old with dominant FLT3 AML, GILTm induced a high 44% CR+CRi rate and long median OS (21.7 mos). Pts with non-dominant FLT3 had low 10% CR+CRi rate. GILTm was generally safe and was associated with differentiation syndrome in 1 pt. Concurrent GILT + DEC was acceptably tolerated, only 1 pt had a DLT, and the MTD was 120 mg/day GILT + DEC. A subset of pts with no CR/CRi during GILTm achieved remission with addition of DEC. Based on these results, a triple combination Tx study with venetoclax is currently enrolling. Figure 1 Figure 1. Disclosures Traer: Genentech: Membership on an entity's Board of Directors or advisory committees; Schrodinger: Research Funding; Incyte: Research Funding; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Servier/Agios: Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees; ImmunoGen: Membership on an entity's Board of Directors or advisory committees. Mims: Glycomemetics: Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Aptevo: Research Funding; Leukemia and Lymphoma Society's Beat AML clinical study: Consultancy, Research Funding; Genentech: Consultancy; Xencor: Research Funding; Kartos Pharmaceuticals: Research Funding; Abbvie: Consultancy; BMS: Consultancy; Kura Oncology: Consultancy; Syndax Pharmaceuticals: Consultancy; BMS: Consultancy; Jazz Pharmaceuticals: Consultancy; Aptevo: Research Funding. Stein: Agios Pharmaceuticals, Inc: Consultancy; Novartis: Consultancy; Astellas: Consultancy; Syndax Pharmaceuticals: Consultancy; Daiichi Sankyo: Consultancy; Syros Pharmaceuticals, Inc.: Consultancy; PinotBio: Consultancy; Celgene: Consultancy; Bristol Myers Squibb: Consultancy; Jazz Pharmaceuticals: Consultancy; Foghorn Therapeutics: Consultancy; Blueprint Medicines: Consultancy; Gilead Sciences, Inc.: Consultancy; Abbvie: Consultancy; Janssen Pharmaceuticals: Consultancy; Genentech: Consultancy. Stock: Pfizer: Consultancy, Honoraria, Research Funding; amgen: Honoraria; agios: Honoraria; jazz: Honoraria; kura: Honoraria; kite: Honoraria; morphosys: Honoraria; servier: Honoraria; syndax: Consultancy, Honoraria; Pluristeem: Consultancy, Honoraria. Kovacsovics: AbbVie: Research Funding; Jazz Pharmaceutials: Honoraria; Janssen Pharmaceuticals: Research Funding; Amgen Inc.: Research Funding; Stemline: Honoraria; Novartis: Research Funding. Blum: Xencor: Research Funding; Abbvie: Honoraria; Nkarta: Research Funding; Celyad Oncology: Research Funding; AmerisourceBergen: Honoraria; Forma Therapeutics: Research Funding; Leukemia and Lymphoma Society: Research Funding; Syndax: Honoraria. Arellano: Syndax Pharmaceuticals, Inc: Consultancy; KITE Pharma, Inc: Consultancy. Schiller: Actinium Pharmaceuticals, Inc: Research Funding; Mateon: Research Funding; Tolero: Research Funding; Geron: Research Funding; Regimmune: Research Funding; Kite/Gilead: Honoraria, Research Funding, Speakers Bureau; Celator: Research Funding; Sangamo: Research Funding; Stemline Therapeutics, Inc.: Honoraria, Research Funding, Speakers Bureau; Takeda: Research Funding; PrECOG: Research Funding; Pfizer: Current equity holder in publicly-traded company, Research Funding; Karyopharm: Research Funding; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Gamida Cell Ltd.: Research Funding; FujiFilm: Research Funding; Samus: Research Funding; Trovagene: Research Funding; Daiichi-Sankyo: Research Funding; Constellation Pharmaceuticals: Research Funding; BMS/Celgene: Consultancy, Current equity holder in publicly-traded company, Research Funding, Speakers Bureau; Abbvie: Research Funding; Actuate: Research Funding; Arog: Research Funding; Delta-Fly: Research Funding; Amgen: Consultancy, Current equity holder in publicly-traded company, Honoraria, Research Funding, Speakers Bureau; Agios: Consultancy, Research Funding, Speakers Bureau; Novartis: Consultancy, Research Funding; Jazz: Consultancy, Honoraria, Research Funding, Speakers Bureau; Elevate: Research Funding; Ono-UK: Consultancy, Research Funding; Onconova: Research Funding; Deciphera: Research Funding; Astellas: Honoraria, Research Funding, Speakers Bureau; Forma: Research Funding; Genentech-Roche: Research Funding; Bio: Research Funding; Sanofi: Honoraria, Research Funding, Speakers Bureau; Pharma: Consultancy; Johnson & Johnson: Current equity holder in publicly-traded company; Biomed Valley Discoveries: Research Funding; Eli Lilly: Research Funding; ASH foundation: Other: Chair-unpaid; Sellas: Research Funding; Ono: Consultancy; Incyte: Consultancy; Ariad: Research Funding; AstraZeneca: Consultancy; Kaiser Permanente: Consultancy; Cyclacel: Research Funding; MedImmune: Research Funding; Ambit: Research Funding; Leukemia & Lymphoma Society: Research Funding; Bluebird Bio: Research Funding; Boehringer-Ingleheim: Research Funding; Cellerant: Research Funding; CTI Biopharma: Research Funding; Janssen: Research Funding; Kura Oncology: Research Funding; Pharmacyclics: Honoraria, Speakers Bureau; Millennium: Research Funding; National Marrow Donor Program: Research Funding; NIH: Research Funding; Onyx: Research Funding; Pharmamar: Research Funding; UC Davis: Research Funding; UCSD: Research Funding; Evidera: Consultancy; NCI: Consultancy; Novartis: Speakers Bureau. Olin: Astellas: Honoraria, Research Funding; Daiichi Sankyo: Research Funding; Genentech: Research Funding; Pfizer: Research Funding; Cellectis: Research Funding; Amgen: Honoraria; Abbvie: Honoraria; Actinium: Honoraria. Foran: taiho: Honoraria; syros: Honoraria; kura: Research Funding; boehringer ingelheim: Research Funding; sanofi aventis: Honoraria; trillium: Research Funding; aptose: Research Funding; abbvie: Research Funding; pfizer: Honoraria; gamida: Honoraria; actinium: Research Funding; takeda: Research Funding; certara: Honoraria; OncLive: Honoraria; bms: Honoraria; revolution medicine: Honoraria; servier: Honoraria; novartis: Honoraria; h3bioscience: Research Funding; aprea: Research Funding; sellas: Research Funding; stemline: Research Funding. Litzow: AbbVie: Research Funding; Jazz: Other: Advisory Board; Pluristem: Research Funding; Amgen: Research Funding; Omeros: Other: Advisory Board; Actinium: Research Funding; Astellas: Research Funding; Biosight: Other: Data monitoring committee. Lin: AbbVie, Aptevo Therapeutics, Astellas Pharma, Bio-Path Holdings, Celgene, Celyad, Genentech-Roche, Gilead Sciences, Incyte, Jazz Pharmaceuticals, Novartis, Ono Pharmaceutical, Pfizer, Prescient Therapeutics, Seattle Genetics, Tolero, Trovagene: Research Funding. Patel: BMS-Celgene, Agios: Membership on an entity's Board of Directors or advisory committees; Peerview: Honoraria; Aptevo Therapeutics: Research Funding. Foster: Bellicum Pharmaceuticals: Research Funding; Macrogenics: Research Funding; Rafael Pharmaceuticals: Research Funding; Macrogenics: Consultancy; Daiichi Sankyo: Consultancy; Agios: Consultancy. Cogle: Aptevo therapeutics: Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees. Vergilio: Foundation Medicine: Current Employment. Gana: The Leukemia & Lymphoma Society: Consultancy; Bausch: Current holder of individual stocks in a privately-held company. Druker: VB Therapeutics: Membership on an entity's Board of Directors or advisory committees; The RUNX1 Research Program: Membership on an entity's Board of Directors or advisory committees; GRAIL: Current equity holder in publicly-traded company; Third Coast Therapeutics: Membership on an entity's Board of Directors or advisory committees; EnLiven: Consultancy, Research Funding; Iterion Therapeutics: Membership on an entity's Board of Directors or advisory committees; Recludix Pharma, Inc.: Consultancy; Pfizer: Research Funding; Merck & Co: Patents & Royalties; Cepheid: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Nemucore Medical Innovations, Inc.: Consultancy; Bristol-Myers Squibb: Research Funding; Blueprint Medicines: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Aptose Therapeutics: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Amgen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; ALLCRON: Consultancy, Membership on an entity's Board of Directors or advisory committees; Aileron: Membership on an entity's Board of Directors or advisory committees; Vincerx Pharma: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Vivid Biosciences: Membership on an entity's Board of Directors or advisory committees. Byrd: Novartis, Trillium, Astellas, AstraZeneca, Pharmacyclics, Syndax: Consultancy, Honoraria; Vincerx Pharmaceuticals: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Newave: Membership on an entity's Board of Directors or advisory committees. Levine: Auron: Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy; Ajax: Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Isoplexis: Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy; Celgene: Research Funding; Roche: Honoraria, Research Funding; Zentalis: Membership on an entity's Board of Directors or advisory committees; Prelude: Membership on an entity's Board of Directors or advisory committees; Mission Bio: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria; Janssen: Consultancy; Gilead: Honoraria; Morphosys: Consultancy; Imago: Membership on an entity's Board of Directors or advisory committees; QIAGEN: Membership on an entity's Board of Directors or advisory committees; Lilly: Honoraria. Borate: Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Blueprint Medicine: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Genentech: Membership on an entity's Board of Directors or advisory committees, Other: Advisory Board; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; Jazz Pharma: Research Funding; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; incyte: Membership on an entity's Board of Directors or advisory committees, Research Funding; Rampal: Membership on an entity's Board of Directors or advisory committees; Galecto, Inc.: Consultancy; Promedior: Consultancy.

Introducción: El objetivo principal en el manejo de los pacientes receptores de trasplante renal, es mantener el estado de inmunosupresión adecuado para evitar la presentación de un rechazo inmunológico del injerto. El rechazo activo mediado por anticuerpos es una de las causas más frecuentes de disfunción del injerto en el periodo postrasplante temprano, además de representar una causa importante de disminución en su sobrevida, sin embargo, en México son escasos los reportes de la prevalencia de este evento, sobre todo en lo referente a la población pediátrica. Métodos: Estudio retrospectivo, transversal, descriptivo. Todos los niños con trasplante renal que hayan sido trasplantados en la UMAE Hospital de Pediatría CMNO. Se solicitó en el archivo médico el listado de pacientes con diagnóstico de “rechazo de injerto”, “rechazo humoral”, “rechazo agudo de injerto”, “rechazo agudo humoral” “rechazo mediado por anticuerpos” de Enero 2018 a Diciembre 2020, posteriormente, con el número de afiliación, se revisaron los expedientes electrónicos. En una hoja de captura se vaciaron los datos necesarios para la investigación, los cuales fueron analizados. Resultados: Se estudiaron 103 pacientes que recibieron un trasplante de riñón en un período de 3 años, de estos, 2 fueron excluidos, quedando 101 pacientes. De ellos, 15 pacientes presentaron rechazo agudo catalogado por biopsia de injerto renal, con 1 (6.6%) clasificado como rechazo celular, 3 (20%) clasificados como rechazo mixto y 11 (73.3%) catalogados como rechazo mediado por anticuerpos, representando un 10.9% de los pacientes trasplantados, de los cuales 8 (72.7%) recibieron trasplante de donador vivo relacionado y 3 (27.3%) lo recibieron de donador cadavérico. Se encontró una distribución equitativa por género, hubo un total de 6 (54.5%) pacientes del género masculino y 5 (45.4%) del género femenino. El rango de edad fue de 7 a 19 años, con una media de 13 años. Conclusión: La prevalencia de rechazo mediado por anticuerpos en pacientes pediátricos de la UMAE Hospital de Pediatría CMNO fue de 10.9%. Recibido: Agosto 01, 2022 Aceptado: Septiembre 30, 2022 Publicado: Septiembre 30, 2022 Editor: Dr. Franklin Mora Bravo. Introducción El rechazo mediado por anticuerpos es la mayor causa de disfunción y pérdida del injerto después del trasplante renal. Actuamente están descritos tres categorías de cambios de la función renal mediados por anticuerpo: Presencia del marcador C4d sin rechazo, rechazo agudo mediado por anticuerpos (RAMA) y rechazo crónico mediado por anticuerpos (RCMA) [1]. Para el diagnóstico de RAMA se requiere el diagnóstico de lesiones histológicas (glomerulitis, capilaritis peritubular, microangiopatía trombótica, necrosis tubular aguda, arteritis intimal), evidencia latente de interacción entre anticuerpos y endotelio (C4d peritubular, glomerulitis + capilaritis, activación endotelial) y la presencia de anticuerpos donante específico [2]. El RAMA se clasifica en 2 fenotipos : el rechazo precoz durante los primeros 3 mses posttrasplante y el recrhazo tradía que ocurre luego del primer año de trasplante, el primero ocurre en paciente con panel reactivo de anticuerpo positivos pretrasplante, usualmente C4D positivo y el segundo con anticuerpos anti HLA de novo usualmente C4D negativo, asociado a la falta de adherencia farmacológica con peor respuesta terapéutica [3]. El RCMA constituye una de las causas principales de pérdida de los injertos, y se asocia a mal pronóstico. La característica que permite el diagnóstico es el hallazgo histológico de la glomerulopatía del trasplante (GT) [4]. La GT se diagnostica en etapas avanzadas por microscopia óptica por la aparición de dobles contornos y expansión de la matriz mesangial. Los cambios preceden a cualquier manifestación clínica. En la microscopia electrónica se aprecia la multilaminación de la membrana basal capilar y/o engrosamiento y duplicación de la membrana basal glomerular. Desde el punto de vista clínico cursa en 2 etapas. Una etapa subclínica sin alteraciones en la función renal ni proteinuria, cuyo único hallazgo es la GT en las biopsias de protocolo y una segunda etapa clínica, caracterizada por disfunción crónica del injerto, con proteinuria e hipertensión arterial [5]. Los pacientes con rechazo crónico pueden asociar elementos de daño activo en la microcirculación mediada por anticuerpos, conocido como rechazo crónico activo, que se define por la presencia concomitante de: Evidencia histológica de daño tisular crónico como: GT, si no hay microangiopatía crónica; delaminación severa de la membrana basal de los capilares peritubulares (en la microscopia electrónica; fibrosis de la íntima arterial de inicio reciente, descartando otras causas; evidencia de interacción de anticuerpos (actual o reciente) dado por cualquiera de los siguientes: C4d en los capilares peritubulares, inflamación de la microcirculación moderada, marcadores moleculares (endothelial-associated transcripts), Evidencia de anticuerpos donantes específico (HLA y no HLA) [6, 7]. Aunque se ha referido una prevalencia baja de rechazo de injerto mediado por anticuerpos en pacientes no sensibilizados, ésta aumenta de forma muy importante en pacientes de alto riesgo, como pacientes previamente sensibilizados, en quienes puede alcanzar de un 10 a un 35% [6]. Esta cifra es alarmante, pues el rechazo de injerto mediado por anticuerpos es el principal factor de riesgo para pérdida del injerto renal en el primer año postrasplante, además de que todo rechazo mediado por anticuerpos no tratado culmina en la pérdida del injerto renal. En México existen escasos datos que aporten la prevalencia de este evento en la población pediátrica, por lo que el objetivo del presente estudio fue determinar la prevalencia de rechazo de injerto mediado por anticuerpos en pacientes pediátricos trasplantados de riñón en la UMAE Hospital de pediatría CMNO. Materiales y métodos Diseño del estudio El presente estudio es observacional, descriptivo, de tipo retrospectivo. Escenario El estudio se realizó en el departamento de nefrología en la Unidad Médica de Alta Especialidad, Hospital de Pediatría, Centro Médico Nacional de Occidente, Instituto Mexicano del Seguro Social, Guadalajara, Jalisco-México, durante el periodo de 1ro de enero del 2017 al 30 de diciembre del 2020. Participantes Se incluyeron pacientes pediátricos con el diagnóstico de enfermedad renal crónica estadio 1-3-T que requirieron hospitalización por necesidad de tratamiento de rechazo agudo, catalogado como mediado por anticuerpos. Se excluyeron paciente con anticuerpos pre-formados. Se eliminaron casos con datos incompletos para el análisis, con historias clínicas incompletas o sin seguimiento posterior al ingreso. Variables Las variables fueron: edad, sexo, tipo de trasplante, tiempo transcurrido del trasplante, grupo sanguíneo, fuente de donación, HLA receptor-donador, inducción, esquema de inducción, esquema de inmunosupresión, biopsia del injerto, dosis acumulada de timoglobulina, identificación de anticuerpos HLA donante específico. Fuentes de datos/mediciones La fuente fue indirecta, se revisó el expediente electrónico institucional, el registro de los servicios de estadística, nefrología y la unidad de trasplantes. Sesgos Con el fin de evitar posibles sesgos de entrevistador, de información y de memoria, los datos fueron custodiados durante todo el tiempo por el investigador principal con una guía y registros aprobados en el protocolo de investigación. El sesgo de observación y selección fueron evitados con la aplicación de los criterios de selección de los participantes. Se consignaron todas las variables clínicas y paraclínicas del periodo ya comentado. Dos investigadores de manera independiente analizaron cada uno de los registros por duplicado y se consignaron las variables en la base de datos una vez verificada su concordancia. Tamaño del estudio La muestra fue no probabilística, tipo censo, en donde se incluyeron todos los casos posibles del período en estudio. Variables cuantitativas Se utilizó estadística descriptiva. Se expresaron los resultados en escala en medias y desviación estándar. Los datos categóricos como el sexo se presentan en proporciones. Análisis estadístico Se utiliza estadística no inferencial. Se calculó la prevalencia de RAMA con intervalo de confianza para una proporción. El paquete estadístico utilizado fue SPSS 25.0 (IBM Corp. Released 2017. IBM SPSS Statistics for Windows, Version 25.0. Armonk, NY: IBM Corp.). Resultados Participantes Se estudiaron 103 pacientes que recibieron un trasplante de riñón en un período de 3 años, de estos, 2 fueron excluidos, uno por no contar con expediente clínico completo y el otro por tener anticuerpos preformados, quedando 101 pacientes que contaban con expedientes clínicos completos. Características basales de la población de estudio De los 101 pacientes trasplantados, 60 (59.4%) corresponden al sexo masculino y 41 (40.6%) corresponden al sexo mujer. La edad media de los pacientes al momento del trasplante fue 12 años. En la mayoría de los trasplantes se realizaron de injertos provenientes de donador vivo 80 (79.2%), y 21 (20.7%) pertenecían a donante cadavérico. Durante el periodo revisado, se encontró un total de 15 pacientes que presentaron rechazo agudo catalogado por biopsia de injerto renal (histopatología), con 1 (6.7%) clasificado como rechazo celular, 3 (20%) clasificados como rechazo mixto y 11 (73.3%) catalogados como rechazo mediado por anticuerpos. Rechazo mediado por anticuerpos Los 11 pacientes con rechazo mediado por anticuerpos representan un 10.9% de los pacientes trasplantados, de los cuales 8 (72.7%) recibieron trasplante de donador vivo relacionado y 3 (27.3%) lo recibieron de donador cadavérico. Del total de pacientes catalogados con rechazo mediado por anticuerpos, se encontró una distribución equitativa por género, ya que hubo un total de 6 (54.5%) pacientes del género masculino y 5 (45.4%) del género femenino. El rango de edad fue de 7 a 19 años, con una media de 13 años y una moda de 14 años de edad. En la figura 1, se presenta la distribución por edad de la población de pacientes con rechazo. Tomando en cuenta el tiempo transcurrido de la fecha del trasplante a la fecha de la presentación del rechazo, el período de tiempo más corto fue de un mes y el más largo fue de 23 meses, con un promedio de tiempo de presentación de 11 meses, como se muestra en la figura 2. Análisis secundarios La variable compatibilidad se definió como una variable dicotómica: < 3 antígenos y ≥3 antígenos. De los 11 pacientes que presentaron rechazo mediado por anticuerpos, sólo fue posible obtener los datos de compatibilidad de 8 de ellos, los cuales fueron en su totalidad trasplantes de donador vivo, los 8 pacientes compartían más de 3 antígenos. Descrito con más detalle, uno de ellos compartía dos haplotipos, cinco de ellos compartían un haplotipo y dos de ellos compartían únicamente antígenos de la clase II. No contamos con los HLA de los 3 pacientes cuyo trasplante fue de donador cadavérico. La inducción recibida durante el trasplante renal se realizó con Basiliximab en 10 casos (90.9%) y 1 (9%) con Timoglobulina. El único paciente que recibió inducción con Timoglobulina presentó rechazo a los 23 meses del trasplante renal. En cuanto al esquema de inmunosupresión de mantenimiento que recibían los pacientes al momento de la presentación del rechazo, la totalidad de ellos tenía triple esquema, teniendo corticosteroide (Prednisona) en común, sin embargo existieron variaciones en el uso de antimetabolitos e inhibidores de calcineurina como se presenta en el Gráfico 8. Se encontraron 6 pacientes en el grupo de Prednisona + Tacrolimus + MMF, 4 pacientes en el grupo de Prednisona + CsA + MMF y solo un paciente tenía Azatioprina como antimetabolito. El periodo de tiempo transcurrido del trasplante a la presentación del rechazo en el grupo de Prednisona + CsA+ MMF fue de 1 a 23 meses, en el grupo de Prednisona + Tacrolimus + MMF fue de 1 a 22 meses y en el paciente con Prednisona + Tacrolimus + Azatioprina fue de 16 meses. No existió diferencia significativa en el tiempo de presentación del rechazo (p= 0.66) al comparar los grupos de Prednisona + CsA+ MMF y Prednisona + Tacrolimus + MMF. Del total de los pacientes, 6 tenían mal apego a tratamiento confirmado por cuidadores y los pacientes mismos, sin embargo, incluso entre estos pacientes, el tiempo transcurrido del trasplante al rechazo fue muy variable, encontrando periodos tan cortos como 6 meses hasta períodos de 23 meses. Entre estos pacientes, se encuentra el que recibió inducción con Timoglobulina. De estos pacientes, 10 (90.9%) fueron tratado con plasmaféresis, en cantidad de 5 sesiones con inmunoglobulina endovenosa a dosis de 2g/kg/dosis, además del uso de anticuerpo monoclonal anti CD20 (Rituximab) al haber completado las 5 sesiones de plasmaféresis, uno de los 11 pacientes recibió Timoglobulina además del tratamiento mencionado. El paciente restante fue dado de alta por edad antes de poder recibir tratamiento. Discusión Para la realización de este estudio se estudiaron de forma retrospectiva los expedientes de pacientes que habían tenido diagnóstico de disfunción de injerto, rechazo agudo o rechazo de injerto. La prevalencia de rechazo de injerto mediada por anticuerpos encontrada en este estudio fue de 10.9%, lo que correlaciona con la prevalencia en adultos encontrada en estudios en Europa y Latinoamérica, como los realizados por Lorent [8] y Borroto Diaz [9], donde se reportó en 16.2% y 11.23% respectivamente. En México no encontramos reportes de este evento en edad pediátrica. El tiempo transcurrido desde la fecha del trasplante renal hasta la presentación del rechazo fue muy variable, ocurriendo desde 1 hasta 23 meses después. La inducción recibida en los pacientes trasplantado fue con un mayor porcentaje a base de Basiliximab, sin embargo, aunque no se cuenta con una muestra representativa de pacientes con inducción a base de Timoglobulina, si se observa que en estos pacientes el tiempo de presentación de rechazo fue mayor y la incidencia de rechazo agudo confirmado por biopsia fue significativamente más alta en el grupo que recibió inducción con Basiliximab. Aunque otros estudios no se reportado diferencias significativas en la supervivencia del injerto en los dos grupos de estudio [10]. En cuanto a la inmunosupresión de mantenimiento recibida al momento de la presentación del rechazo no se observaron diferencias al comparar los grupos (PDN+ Tacrolimus + MMF vs PDN + CsA + MMF) (P=0.66), sin embargo, el no apego al tratamiento de mantenimiento fue una variable importante en la presentación de rechazo; la no adherencia es un factor de riesgo significativo e independiente para la pérdida del injerto; por ello consideramos que la vigilancia del paciente pediátrico, sobre todo el adolescente, debe ser extremadamente estrecha por el alto riesgo de no apego, y en toda la población pediátrica trasplantada, esta vigilancia debe ser estricta con el objetivo de mantener sus niveles de inmunosupresión dentro de parámetros recomendados y con ello evitar la presentación de rechazo mediado por anticuerpos. Estos pacientes recibieron tratamiento en varias líneas, tanto plasmaféresis como Rituximab y utilización de gamaglobulina endovenosa; se plantea a futuro la realización de estudios posteriores los cuales pudiesen demostrar la eficacia de algunas de estas medidas. Se pretendió conocer la compatibilidad de los antígenos HLA, ya que en investigaciones anteriores, se ha evaluado y concluido que la compatibilidad HLA influye sobre la supervivencia del injerto y la mortalidad en el trasplante renal, por ejemplo, un estudio observacional y encontró que la incompatibilidad HLA en general se asoció significativamente con mayor riesgo de falla del injerto, incluso mayor mortalidad [11]. Un sesgo muy importante en este estudio es la falta de conocimiento de la compatibilidad en los antígenos HLA en los tres pacientes que presentaron rechazo mediado por anticuerpos, que habían recibido su injerto de donante cadavérico. Sin embargo, cabe mencionar que muchos de los órganos procurados de donante cadavérico provienen de otros estados y el envío de muestras para la realización de estudios así como el tiempo necesario para tener los resultados disponibles dificulta la realización de los mismos, por lo que estos pacientes deben de ser considerado como de alto riesgo independientemente de la edad y deberán ser inducidos a base de Timoglobulina. Los 8 pacientes restantes, recibieron su injerto de donador vivo y todos compartían más de 3 antígenos. Se decidió no excluir a los pacientes de donador cadavérico para contar con este grupo de pacientes y dar a conocer la evolución de los mismos, además de que, de haberlos eliminado, se habría reportado una menor prevalencia. Conclusiones De los 101 pacientes trasplantados, 60 (59.4%) corresponden al género masculino y 41 (40.5%) corresponden al género femenino. La prevalencia de rechazo mediado por anticuerpos fue de 10.9%. Del total de pacientes catalogados con rechazo mediado por anticuerpos, 6 (54.5%) pacientes del género masculino y 5 (45.4%) del género femenino. En los pacientes catalogados con rechazo mediado por anticuerpos, el rango de edad fue de 7 a 19 años, con una media de 13 años y una moda de 14 años de edad. El tiempo transcurrido de la fecha del trasplante a la fecha de la presentación del rechazo fue desde 1 hasta 23 meses. El tiempo de presentación del rechazo en pacientes de donante cadavérico, en quienes se desconoce la compatibilidad HLA, fue muy variable, abarcando desde 8 hasta 22 meses. No existió diferencia estadísticamente significativa al comparar el esquema de inducción recibido y la presencia de rechazo. El tiempo de presentación más largo fue en dos pacientes, que habían recibido Basiliximab y Timoglulina cada uno. No existió diferencia estadísticamente significativa en el tiempo de presentación del rechazo al comparar los distintos esquemas de inmunosupresión de mantenimiento recibidos. Sin embargo, se observó poca adherencia al tratamiento como variable interviniente. Abreviaturas ABMR: Rechazo mediado por anticuerpos (Antibodies mediated rejection) ADCC: Citotoxicidad dependiente de anticuerpos CAM: Complejo de ataque de membrana CD: Célula dendrítica. CDC: Citotoxicidad dependiente de complemento CMH: Complejo Mayor de Histocompatibilidad. CMNO: Centro Médico Nacional de Occidente. CPA: Célula presentadora de antígeno. CXCL: Ligando de quimiocina (Chemokine Ligand) DC: Donador cadavérico. DSA: Anticuerpos Donante Específicos (Donor Specific Antibodies). DVR: Donador vivo relacionado. ERC: Enfermedad renal crónica ERO: Especies reactivas de oxígeno. FDA: Administración de fármacos y alimentos (Food and drug administration). GT: Glomerulopatía del trasplante HLA: Antígeno Leucocitario Humano (Human Leucocitary Antigen) ICAM: Molécula de adhesión intercelular (Intercellular Adhesion Molecule 1) IFN: Interferón Ig: Inmunoglobulina IKK: Cinasa inhibidora del factor Kappa B (I Kappa B Kinase) IMSS: Instituto Mexicano del Seguro Social. LRA: Lesión renal aguda MAPK: Proteincinasa activada por mitógeno (mitogen-activated protein kinase) MMP: Metalopeptidasa de la matriz (Matrix Metallopeptidase) NLRs: Receptores tipo NOD (NOD like receptors). PMAD: Productos moleculares asociados a daño. PRA: Panel reactivo de anticuerpos rATG: Globulina antitimocito de conejo (rATG) RRP: receptores de reconocimiento de patrones TLRs: Receptores tipo Toll (Toll like receptors). TNF: Factor de necrosis tumoral (Tumoral necrosis factor) TRIF: Interferón-B inductor de adaptadores que contienen dominios de Tirosina UMAE: Unidad Médica de Alta Especialidad. VCAM-1: Molécula de adhesión celular vascular (vascular cell adhesion molecule 1) VEGF: Factor de crecimiento endotelial vascular (vascular endotelial grown factor). Información suplementaria Materiales suplementarios no han sido declarados. Agradecimientos No aplica. Contribuciones de los autores Mónica Verenice Cámara Carrillo: Conceptualización, Curación de datos, Análisis formal, Adquisición de fondos, Investigación, Metodología, Administración de proyecto, Recursos, Software, Escritura – borrador original. Santa Ramírez Godinez: Conceptualización, Supervisión, Validación, Visualización, Redacción: revisión y edición. Juan Carlos Barrera de León: Metodología, validación, supervisión, redacción: Revisión y edición. Todos los autores leyeron y aprobaron la versión final del manuscrito. Financiamiento Los autores proveyeron los gastos de la investigación. Disponibilidad de datos o materiales Los conjuntos de datos generados y analizados durante el estudio actual no están disponibles públicamente debido a la confidencialidad de los participantes, pero están disponibles a través del autor correspondiente a pedido académico razonable. Declaraciones Aprobación del comité de ética y consentimiento para participar Este estudio fue aprobado por el comité de ética en investigación de la Unidad Médica de Alta Especialidad Hospital de Pediatría CMNO, aprobación número R-2021-1302-077. Consentimiento para publicación No aplica cuando no se publican imágenes o fotografías del examen físico o radiografías/tomografías/resonancias de pacientes. Conflictos de interés Los autores reportan no tener conflictos de interés. Referencias Loupy A, Lefaucheur C. Antibody-Mediated Rejection of Solid-Organ Allografts. N Engl J Med. 2018 Sep 20;379(12):1150-1160. DOI: 10.1056/NEJMra1802677. PMID: 30231232. Zorn E, See SB. Is There a Role for Natural Antibodies in Rejection Following Transplantation? Transplantation. 2019 Aug;103(8):1612-1619. DOI: 10.1097/TP.0000000000002743. PMID: 30951015; PMCID: PMC6660357. Haas M, Mirocha J, Reinsmoen NL, Vo AA, Choi J, Kahwaji JM, Peng A, Villicana R, Jordan SC. Differences in pathologic features and graft outcomes in antibody-mediated rejection of renal allografts due to persistent/recurrent versus de novo donor-specific antibodies. Kidney Int. 2017 Mar;91(3):729-737. DOI: 10.1016/j.kint.2016.10.040. Epub 2017 Jan 16. PMID: 28104301. Chapman JR, Wavamunno M, O'Connell PJ, Nankivell BJ. Unravelling the connections between donor specific antibodies and renal allograft pathology. Clin Transpl. 2013:361-5. PMID: 25095530. Asante-Korang A, Jacobs JP, Ringewald J, Carapellucci J, Rosenberg K, McKenna D, McCormack J, Wilmot I, Gjeldum A, Lopez-Cepero M, Sleasman J. Management of children undergoing cardiac transplantation with high Panel Reactive Antibodies. Cardiol Young. 2011 Dec;21 Suppl 2:124-32. DOI: 10.1017/S1047951111001703. PMID: 22152539. Velez M, Johnson MR. Management of allosensitized cardiac transplant candidates. Transplant Rev (Orlando). 2009 Oct;23(4):235-47. DOI: 10.1016/j.trre.2009.07.001. PMID: 19778695; PMCID: PMC2796825. Jeong HJ. Diagnosis of renal transplant rejection: Banff classification and beyond. Kidney Res Clin Pract. 2020 Mar 31;39(1):17-31. DOI: 10.23876/j.krcp.20.003. PMID: 32164120; PMCID: PMC7105630. Lorent M, Foucher Y, Kerleau K, Brouard S, Baayen C, Lebouter S, Naesens M, Bestard Matamoros O, Åsberg A, Giral M; EKiTE consortium. The EKiTE network (epidemiology in kidney transplantation - a European validated database): an initiative epidemiological and translational European collaborative research. BMC Nephrol. 2019 Oct 11;20(1):365. DOI: 10.1186/s12882-019-1522-8. PMID: 31601177; PMCID: PMC6788117. Borroto Díaz G, Caballero Gonzalez M, Chong López A. Relación entre los resultados de biopsia del trasplante renal, según la clasificación de Banff del 2011, y el tiempo de vida del injerto. Revista Cubana de Medicina 2016;554(2):97-113. SCIELO: S0034-7523 Newland DM, Royston MJ, McDonald DR, Nemeth TL, Wallace-Boughter K, Carlin K, Horslen S. Analysis of rabbit anti-thymocyte globulin vs basiliximab induction in pediatric liver transplant recipients. Pediatr Transplant. 2019 Dec;23(8):e13573. DOI: 10.1111/petr.13573. Epub 2019 Sep 12. PMID: 31512802. Senev A, Coemans M, Lerut E, Van Sandt V, Kerkhofs J, Daniëls L, Driessche MV, Compernolle V, Sprangers B, Van Loon E, Callemeyn J, Claas F, Tambur AR, Verbeke G, Kuypers D, Emonds MP, Naesens M. Eplet Mismatch Load and De Novo Occurrence of Donor-Specific Anti-HLA Antibodies, Rejection, and Graft Failure after Kidney Transplantation: An Observational Cohort Study. J Am Soc Nephrol. 2020 Sep;31(9):2193-2204. DOI: 10.1681/ASN.2020010019. Epub 2020 Aug 6. PMID: 32764139; PMCID: PMC7461684. DOI: Digital Object Identifier. PMID: PubMed Identifier. Nota del Editor La REV SEN se mantiene neutral con respecto a los reclamos jurisdiccionales sobre mapas publicados y afiliaciones institucionales.

Activating mutations in leucine-rich repeat kinase 2 (LRRK2) represent the most common cause of monogenic Parkinson's disease. LRRK2 is a large multidomain protein kinase that phosphorylates a specific subset of the ∼65 human Rab GTPases, which are master regulators of the secretory and endocytic pathways. After phosphorylation by LRRK2, Rabs lose the capacity to bind cognate effector proteins and guanine nucleotide exchange factors. Moreover, the phosphorylated Rabs cannot interact with their cognate prenyl-binding retrieval proteins (also known as guanine nucleotide dissociation inhibitors) and, thus, they become trapped on membrane surfaces. Instead, they gain the capacity to bind phospho-Rab-specific effector proteins, such as RILPL1, with resulting pathological consequences. Rab proteins also act upstream of LRRK2 by controlling its activation and recruitment onto membranes. LRRK2 signaling is counteracted by the phosphoprotein phosphatase PPM1H, which selectively dephosphorylates phospho-Rab proteins. We present here our current understanding of the structure, biochemical properties, and cell biology of LRRK2 and its related paralog LRRK1 and discuss how this information guides the generation of LRRK2 inhibitors for the potential benefit of patients.

Abstract Soy isoflavones have been shown to have anti-inflammatory properties; however, the anti-inflammatory effects of isoflavone metabolites produced during soybean germination remain unclear. We found that the daidzein and genistein derivatives, 8-prenyl daidzein (8-PD) and 8-prenyl genistein (8-PG), demonstrated a more potent effect than daidzein and genistein on repressing inflammatory responses in macrophages. Although IkB protein levels were unaltered, 8-PD and 8-PG repressed nuclear factor kappa B (NF-κB) activation, which was associated with reduced ERK1/2, JNK, and p38 MAPK activation and suppressed mitogen- and stress-activated kinase 1 phosphorylation. Inflammatory responses induced by the medium containing hypertrophic adipocyte secretions were successfully suppressed by 8-PD and 8-PG treatment. In the ex vivo study, 8-PD and 8-PG significantly inhibited proinflammatory C–C motif chemokine ligand 2 (CCL2) secretion from the adipose tissues of mice fed a long-term high-fat diet. The data suggest that 8-PD and 8-PG could regulate macrophage activation under obesity conditions.

Diabetes (DB) significantly exacerbates myocardial ischemia/reperfusion (MI/R) injury. Unfortunately, conventional pre-conditioning (PreCon) provides diminished cardioprotection during DB, due partially to impaired AMP-activated protein kinase (AMPK) signaling. The current study investigated whether PreCon by inhaled anesthetic sevoflurane (SF-PreCon) remains protective in DB, and if so, to dissect the involved mechanisms. Non-diabetic (ND) or high-fat diet-induced DB mice were subjected to MI/R and randomized into control and SF-PreCon (3 cycles of 15 minute-exposures to 2% sevoflurane prior to MI) groups. As expected, SF-PreCon significantly reduced MI/R injury in ND mice. Importantly, SF-PreCon also significantly reduced MI/R injury in DB mice, as evidenced by reduced apoptosis (-23.1±1.6, P<0.01), decreased infarct size (-21.2±2.3%, P<0.01), and augmented cardiac function (+24±3.0%, P<0.01). To determine the role of AMPK in SF-PreCon-mediated cardioprotection, the effect of SF-PreCon upon MI/R injury was determined in cardiac specific AMPKα2 dominant negative overexpression mice (AMPK-DN). We demonstrate SF-PreCon remained cardioprotective in AMPK-DN mice. To explore the responsible molecular mechanisms, multiple cell-survival signaling molecules were screened. Interestingly, SF-PreCon differentially regulated mitogen-activated protein kinase (MAPK) family members in the MI/R heart. In ND mice, SF-PreCon dramatically reduced (-81±7.2%) MI/R-induced activation of p38, a pro-death MAPK, without significantly altering ERK and JNK. In DB and AMPK-DN mice, the inhibitory effect of SF-PreCon upon p38 activation was significantly blunted (DB: -8±2.1%) or virtually abolished (AMPK-DN). However, SF-PreCon significantly increased (P<0.05) phosphorylation of ERK1/2, a pro-survival MAPK. Collectively, we demonstrate SF-PreCon protected the heart via AMPK-dependent inhibition of pro-death MAPK in ND mice. However, SF-PreCon exerts its cardioprotective actions via AMPK-independent activation of pro-survival MAPK in DB mice. These results suggest SF-PreCon may be a superior intervention over conventional PreCon in DB patients, where AMPK signaling is impaired.

AbstractDiabetes mellitus (DM) significantly increases myocardial ischemia/reperfusion (MI/R) injury. During DM, cardioprotection induced by conventional pre-conditioning (PreCon) is decreased due to impaired AMP-activated protein kinase (AMPK) signaling. The current study investigated whether PreCon with inhaled anesthetic sevoflurane (SF-PreCon) remains cardioprotective during DM, and identified the involved mechanisms. Normal diet (ND) and high-fat diet (HFD)-induced DM mice were randomized into control and SF-PreCon (3 cycles of 15-minute period exposures to 2% sevoflurane) groups before MI/R. SF-PreCon markedly reduced MI/R injury in DM mice, as evidenced by improved cardiac function (increased LVEF and ±Dp/dt), decreased infarct size, and decreased apoptosis. To determine the relevant role of AMPK, the effect of SF-PreCon was determined in cardiac-specific AMPKα2 dominant negative expressing mice (AMPK-DN). SF-PreCon decreased MI/R injury in AMPK-DN mice. To explore the molecular mechanisms responsible for SF-PreCon mediated cardioprotection in DM mice, cell survival molecules were screened. Interestingly, in ND mice, SF-PreCon significantly reduced MI/R-induced activation of p38, a pro-death MAPK, without altering ERK and JNK. In DM and AMPK-DN mice, the inhibitory effect of SF-PreCon upon p38 activation was significantly blunted. However, SF-PreCon significantly increased phosphorylation of ERK1/2, a pro-survival MAPK in DM and AMPK-DN mice. We demonstrate that SF-PreCon protects the heart via AMPK-dependent inhibition of pro-death MAPK in ND mice. However, SF-PreCon exerts cardioprotective action via AMPK-independent activation of a pro-survival MAPK member in DM mice. SF-PreCon may be beneficial compared to conventional PreCon in diabetes or clinical scenarios in which AMPK signaling is impaired.

Melanoma is an aggressive form of cancer with poor prognosis and survival rates and limited therapeutic options. Here, we report the anti-melanoma effect of 3-O-prenyl glycyrrhetinic acid (NPC-402), a derivative of glycyrrhtinic acid, from a reputed medicinal plant Glycyrrhiza glabra against B16F10 cells. We studied the cytotoxic effect of NPC-402 on melanoma cells and investigated the role of mitogen-activated protein (MAP) kinase, AKT axis, and endoplasmic reticulum (ER) stress/unfolded protein response (UPR)-mediated autophagy as the involved signaling cascade by studying specific marker proteins. In this study, 4-phenylbutyric acid (4PBA, a chemical chaperone) and small interference RNA (siRNA) knockdown of C/EBP Homologous Protein (CHOP)/growth arrest- and DNA damage-inducible gene 153(GAD153) blocked NPC-402-mediated autophagy induction, thus confirming the role of ER stress and autophagy in melanoma cell death. NPC-402 induced oxidative stress and apoptosis in melanoma cells, which were effectively mitigated by treatment with N-acetylcysteine (NAC). In vivo studies showed that intraperitoneal (i.p.) injection of NPC-402 at 10 mg/kg (5 days in 1 week) significantly retarded angiogenesis in the Matrigel plug assay and reduced the tumor size and tumor weight without causing any significant toxic manifestation in C57BL/6J mice. We conclude that NPC-402 has a high potential to be developed as a chemotherapeutic drug against melanoma.