Blood Cancer Discovery最新文献

{"title":"Abstract P04: Role of glucocorticoid receptor in normal and malignant germinal center B cells","authors":"Clarissa Corinaldesi, A. Holmes, Elodie Bal, L. Pasqualucci, K. Basso, R. Dalla-Favera","doi":"10.1158/2643-3249.bcdsymp24-p04","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p04","url":null,"abstract":"\u0000 We have reported that super-enhancers (SEs) are specifically hypermutated by Activation-Induced Deaminase (AID)-induced Aberrant Somatic Hypermutation in >90% of diffuse large B cell lymphoma (DLBCL) cases (Bal et al. Nature 2022). Analysis of 122 primary DLBCL biopsies and cell lines identified more than 80 SEs that are recurrently hypermutated, with an average of 12 hypermutated SE/case. Hypermutated SEs are predominantly linked to genes encoding regulators of B cell development and well-known lymphoma oncogenes, including BCL6, BCL2 and CXCR4. We showed that specific hotspot mutations prevent binding and transcriptional downregulation by transcriptional repressors leading to target gene dysregulation. Among these, hypermutation of the SEs linked to BCL2 and CXCR4 abrogates the binding of the glucocorticoid receptor (GR)/transcription factor encoded by NR3C1. As a result, BCL2 and CXCR4 escape GR-mediated transcriptional repression leading to their de-regulated expression in germinal center (GC) B cells. Together with the observation that the NR3C1 gene is genetically inactivated in a small fraction of DLBCL, and the relevance of glucocorticoid-based therapy in DLBCL, these results prompted a comprehensive analysis of the role of GR in normal and malignant B cells. We report that GR is detectable in the nucleus of the great majority of GC B cells, suggesting that it is active as a transcription factor. Similarly, DLBCL primary cases (including GCB-, ABC- and unclassified DLBCL) display GR nuclear expression. Analysis of a conditional GC-specific Nr3c1 knock-out (KO) mouse model showed that GC B cells form normally in the absence of Nr3c1. However, the transcriptional profile of Nr3c1-KO GC B cells was significantly altered compared to wild-type cells. By integrating the GR binding profile obtained in normal human GC B cells with the transcripts differentially expressed in Nr3c1-KO GC B cells, we identified a set of about 2,000 genes that are directly modulated by GR during the GC reaction. Pathway enrichment analysis showed that GR controls several signaling pathways, suggesting a modulatory role on the BCR and CXCR4 pathways. In addition, GR targets display a significant overlap with the BCL6 transcriptional network, suggesting a cooperative role in GC B cell development. Notably, GR was shown to regulate pathways involved in plasma cell differentiation, while repressing targets associated with memory B cell development. In DLBCL cases, several NR3C1 binding sites are recurrently targeted by mutations, suggesting that genetic impairment of GR activity is heterogeneous and extends beyond CXCR4 and BCL2. Together, these data support a role for the glucocorticoid pathway in GC physiology. Recurrent inactivation of the NR3C1 gene or part of its transcriptional regulatory program may contribute to DLBCL pathogenesis, with implications for presently unknown specific roles of glucocorticoids in the therapeutic regimens for DLBCL.\u0000 Citation Format: ","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140080212","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract P06: Characterization of the Immune Tumor Microenvironment in a Diverse Cohort of Patients with Multiple Myeloma","authors":"Suganti Shivaram, Hongwei Tang, Huihuang Yan, Shulan Tian, Guyu Qin, Emilie I. Anderson, Neeraj Sharma, Abiola Bolarinwa, Cinthya Zepeda-Mendoza, Stacey Lehman, Felicia Gomez, A. Mitra, Surendra Dasari, T. Kourelis, S. Kumar, Linda Baughn","doi":"10.1158/2643-3249.bcdsymp24-p06","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p06","url":null,"abstract":"\u0000 Multiple myeloma (MM) is the most common blood cancer in African American (AA) individuals. AAs have an increased risk of developing Monoclonal Gammopathy of Undetermined Significance (MGUS) and MM compared to non-Hispanic White (NHW) individuals. While several studies have shown a pivotal role for the immune tumor microenvironment (iTME) in MM pathogenesis, the composition of the iTME in AAs with MM is under investigated. Viably frozen whole bone marrow (BM) aspirates from 128 individuals (53 AA and 75 NHW) with either MM (n=65), MGUS (n=28), smoldering MM (SMM, n=13), or healthy controls (n=22) were evaluated by single cell RNA-sequencing (scRNA-seq) or mass cytometry (CyTOF). We characterized 34 (14 AA and 20 NHW) by scRNA-seq and 94 (39 AA and 55 NHW) by CyTOF. CyTOF samples were stained with Maxpar Direct Immune Profiling Assay Kit (Fluidigm), acquired on a Helios mass cytometer and analyzed in Cytobank. Data were analyzed by logistic equivalent regression model with p<0.05 for significance. ScRNA-seq samples were processed as in Yao, 2022. Data were normalized using scvi-tools and clustered by Uniform manifold approximation and projection (UMAP). Differentially expressed markers between clusters were identified using Model-based Analysis of Single-cell Transcriptomics (MAST) in Seurat R package at FDR < 0.05 and log2 fold change > 0.55. ScRNA-seq of the CD45+ non-tumor lymphocytes revealed 17 immune populations (HSC, erythroblast, MDSC, GMP, CD1c+ DC, CD4+ and CD8+ T cell, CD14+ M1 macrophage, CD16+monocyte, monocyte, pDC, naïve B, pre-B, pro-B, memory B, NK/MAIT). CD3E/D+cells were clustered into naïve/central memory (CCR7+, LEF1+, SELL+), memory (GZMK+), effector (GZMB+, GZMH+, FGFBP2+), g/d (TRDC+), MAIT (SLC4A10+, KLRB1+) and IFN CD8+ T cells (IFIT1+, IFIT2+,IFIT3+). Disease progression was associated with increases in median CD4+ and CD8+ GZMK+ T cells, memory B cells and a reduction in all monocyte clusters. Within the diseased cohort in comparison to NHW patients, AA patients had increased CD3+ T cells (CD4 and CD8+) (AA 57.5% vs. NHW 44.3%) and a reduction in overall monocytes (AA 10.5% vs. 26.5%). These results were validated by CyTOF, which confirmed an increase in CD3+ T cells (AA 53.7% vs. NHW 35.7%, p<0.001) including increases in CD8+ (AA 7.0% vs. NHW 3.2%, p<0.05) and CD4+ terminal effector T cells (AA 5.7% vs. NHW 2.7%, p<0.05) and a reduction in monocytes (AA 18.3% vs. NHW 26.6%) among AAs. After adjusting for age and sex, AAs retained an increase in T cells (OR 1.59 (95%CI 1.2-2.01, p=0.00023)) and a reduction in monocytes (OR 0.88 (95%CI 0.78-1, p=0.045)). Despite the increase in CD8+ T cells in AA patients, we identified reduced GZMK+ T cells and reductions in exhaustion markers (TIGIT, LAG3, TOX) in CD8+ T cells in comparison to NHWs. Nevertheless, the distribution of the immune populations was similar between healthy AA and NHW individuals. Our multiomics approach to characterize the iTME identified significant diffe","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140079755","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract P34: Bayesian networks modeling identifies a reliance of TP53 mutant AML on NF kappa B signaling","authors":"Daniel Chang, Klara E Noble-Orcutt, Marie L Antony, Yoonkyu Lee, Fiona He, Karen Sachs, Chad L Myers, Z. Sachs","doi":"10.1158/2643-3249.bcdsymp24-p34","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p34","url":null,"abstract":"\u0000 In acute myeloid leukemia (AML), TP53 mutations ( TP53Mut ) confer the worst prognosis. Leukemia stem cells (LSCs) are rare AML cells endowed with self-renewal capacity, allowing them to recapitulate leukemia after therapy and cause disease progression. In human AML, signaling pathways associated with inflammation have been implicated in the pathogenesis of TP53Mut AML, but the pathways that mediate self-renewal in human TP53Mut AML are not well-known. To define the signaling pathways that are activated in TP53Mut human LSCs, we used CyTOF, a form of flow cytometry that can measure up to 40 proteins simultaneously at single-cell resolution. We compared the levels of activated signaling molecules in the LSC-enriched (CD34+CD38−) subset of seven TP53Mut and seven TP53 wild-type ( TP53WT ) AML samples. Phosphorylated NF kappa B (pNFkB), pSTAT1, and pP38 are significantly higher in TP53Mut AMLs. In contrast, Ki67 and pMAPKAPKII are higher in TP53WT AML. These data demonstrate a unique signaling activation state in TP53Mut AML. The influence of signaling molecules on downstream targets can vary by cellular context. Therefore, we asked whether the influence and interaction of the activated signaling molecules vary between TP53Mut and TP53WT LSCs. To model the signaling architecture, we performed Bayesian networks modeling, a machine learning algorithm that has been validated as a method to discover statistical correlations and dependencies between signaling molecules. We found a similar global signaling network structure and hierarchy in TP53Mut and TP53WT AMLs. However, TP53Mut model showed NFkB have strong influences on phosphorylated Histone H3 (pHIS3) and Ki67, suggesting that proliferation depends on NFkB levels in TP53Mut LSCs. Furthermore, NFkB levels were highly dependent on pSTAT1 in TP53Mut LSCs. Next, we compared the transcriptional profiles of TP53Mut and TP53WT samples in the BEAT AML dataset and found that TP53Mut AML displayed significant enrichment of all the NFkB gene sets (n=19) in the molecular signatures database, providing more evidence for a unique reliance of TP53Mut human AML on NFkB signaling. Therefore, we used lentivirus transduced shRNA constructs to determine the influence of mutant p53 on signaling in human AML. We found that knockdown of TP53Mut reduced NFkB protein levels in Kasumi AML cells (which are TP53R248Q/ − ) but not in MOLM13 AML cells (which are TP53WT). RNA sequencing of transduced Kasumi cells showed that TP53Mut knockdown led to loss of NFkB and LSC self-renewal signatures. Knockdown of TP53Mut in primary human TP53Mut AMLs (S127F/- and S241Y/-) led to a similar loss of NFkB and LSC signatures in these primary samples as well. Colony formation in Kasumi cells and TP53Mut primary human AML were also abrogated after TP53Mut knockdown. These data show TP53Mut promotes NFkB activation in human AML and suggests that human TP53Mut LSCs may be dependent on NFkB for self-renewal. These data implicate NFkB as a pos","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140080060","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract P21: VISTA a potential new Immuno-oncology therapeutic target to treat human Acute Myeloid Leukemia","authors":"Thierry Guillaudeux, S. Sridhar, E. Frazier, Yulia Ovechkina, Shawn Iadonato","doi":"10.1158/2643-3249.bcdsymp24-p21","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p21","url":null,"abstract":"\u0000 V-domain Immunoglobulin Suppressor of T cell Activation (VISTA/PD-1H) is a co-inhibitor molecule of the B7 family member highly expressed on circulating and intra-tumoral myeloid cells. VISTA is a negative checkpoint protein that inhibits anti-tumor T cell responses. VISTA expression has been associated with poor overall survival in multiple solid tumors and is also described as a potential mediator of resistance to anti-CLTA-4 and anti-PD1 therapies. VISTA expression was also investigated in hematological tumors and described as highly expressed on blasts including leukemic stem cells from AML patients. Therefore, VISTA is a potentially new therapeutic target for cancer immunotherapy in this indication. Kineta has developed a fully human monoclonal antibody targeting VISTA, KVA12123, currently evaluated in a Phase 1/2 clinical trial alone and in combination with pembrolizumab in cancer patients with advanced solid tumors. Supported with previous published data on VISTA expression and function in myeloid leukemia, we also believe that our lead therapeutic anti-VISTA antibody could be a promising drug for AML patients. Therefore, we have tested in vivo, in a Kasumi-3 AML disseminated tumor model, the efficacy of our antibody alone or in combination with standard of care therapy, cytarabine and doxorubicin. The Kasumi-3 cell line exhibits a high level of VISTA expression on its cell surface, like leukemic cells in AML patients. Female NOD-scid mice were first inoculated with Kasumi cells via tail vein injection, and tumor engraftment was evaluated at different time points in the blood, spleen and bone marrow to monitor engraftment using CD34, CD33, CD45 and HLA-DR cell surface expression markers. Then, when tumor burden was high, standard of care treatment (5 doses of 16mg/kg AraC by IV injections and 3 doses of 0.5mg/kg Dox by IV injections) was initiated in indicated groups for 5 cycles, before starting treatment with KVA12 for the rest of the study. Tumor engraftment was tested after chemotherapy alone to evaluate the efficacy before starting treatment with our antibody. Our results showed that untreated mice had higher number of Kasumi-3 cells in the blood when compared to chemotherapy treated mice. Two backbone versions of our antibody were then tested, a human IgG1 – KVA12123 and a human IgG4 - KVA12402, at 30mg/kg IP injections twice a week for 5 weeks. Our data showed that KVA12123 and KVA12402 as single agents and in combination with chemotherapy reduced Kasumi-3 tumor load in the blood, spleen and bone marrow within the groups and significantly improved survival when compared to IgG1 control groups. These data indicate that anti-VISTA targeted therapy should be evaluated in myeloproliferative disorders like AML alone or in combination with standard of care.\u0000 Citation Format: Thierry Guillaudeux, Shaarwari Sridhar, Emily Frazier, Yulia Ovechkina, Shawn Iadonato. VISTA a potential new Immuno-oncology therapeutic target to treat human Acut","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140266100","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract P13: Genetic and epigenetic characterization of B-cell chronic lymphocytic leukemia with IG::BCL3-translocation provides evidence for a distinct biological entity","authors":"Cosima Drewes, Cristina López, Nnamdi Okeke, S. Hillebrecht, Petra Schütz, Anja Fischer, Anja Mottok, S. Bens, C. Schneider, S. Stilgenbauer, E. Tausch, Reiner Siebert","doi":"10.1158/2643-3249.bcdsymp24-p13","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p13","url":null,"abstract":"\u0000 IG translocations occur in various B-cell neoplasms and result in the deregulation of (onco)genes due to their juxtaposition with regulatory elements in the immunoglobulin loci, predominantly the immunoglobulin heavy chain (IGH) locus (14q32). Some of these IGH translocations are pathognomonic for distinct disease entities, like those involving MYC, BCL2 or CCND1 in Burkitt, Follicular or Mantle Cell Lymphomas, respectively. In Chronic Lymphocytic Leukemia (CLL), the BCL3 gene (19q13), a transcriptional coactivator of the nuclear factor-kappaB (NF-κB) family, is a known partner gene in these translocations. Two subsets of IG::BCL3 translocation-positive B-cell neoplasms have been described that exhibit differences in chromosomal aberrations, IGHV mutation status, and histopathology [Martín-Subero et al. 2007]. In CLL, IG::BCL3 translocation is linked to a younger age at diagnosis, a more aggressive clinical course, and an atypical tumor cell phenotype [Michaux et al. 1997; Au et al. 2002]. A total of 89 B-cell neoplasms, primarily diagnosed as CLL, displaying IG::BCL3 translocation detected by fluorescence in situ hybridization (FISH), were examined. Analysis of DNA methylation and copy number aberrations (CNA) was conducted for 86 IGH::BCL3-translocated B-cell neoplasms using 850K EPIC BeadChip array data. Evaluation of chromosomal aberrations typical for CLL was moreover performed by FISH and the IGHV mutation status was determined. IGH::BCL3 breakpoint junctions were sequenced in 23 samples using targeted capture and whole-genome sequencing approaches. RNA expression of BCL3 was assessed using the HTG mRNA pan B-cell panel. In UMAP analysis of DNA methylation, CLLs with IG::BCL3 translocation exhibited segregation from other CLLs, irrespective of IGH translocations. Supervised analysis revealed global hypomethylation of B-cell neoplasms with IG::BCL3 translocation compared to other CLL. A total of 66/69 CLLs with IG::BCL3 translocation showed an unmutated IGHV status (96%). Copy number determination using BeadChip data (n=86) and FISH analysis (n=85) showed a significantly higher incidence of trisomy 12 in IG::BCL3-translocated cases compared to the general CLL population (p=0.011). Breakpoint sequencing indicated IG::BCL3 junctions to be recurrently associated with aberrant class-switch recombination at the IGH locus, involving IGHA (n=11/23), IGHG (n=5/23) and IGHM segments (n=5/23). Breakpoints at the BCL3 locus displayed two groups. 80 cases (all CLL) were located upstream of BCL3 and two other B-cell neoplasms (NHL, DLBCL) were located downstream of BCL3. BCL3 expression analysis in 15 IG::BCL3-translocated CLLs confirmed its transcriptional upregulation compared to the general CLL population. Our genetic and epigenetic analyses support the view that CLL with IG::BCL3 translocation may constitute a genetically and epigenetically distinct subtype of B-cell neoplasms, diverging from the typical CLL.\u0000 Citation Format: Cosima Drewes, Cristi","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140265498","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract P01: Impact of Germline and Somatic ATM Variants in Chronic Lymphocytic Leukemia (CLL): Clinical Implications and Response to PARP Inhibition","authors":"Kiyomi Mashima, Nicholas S. Moore, M. Mikhaleva, Anna Petráčková, Samantha Shupe, Stacey M Fernandes, A. Taylor-Weiner, R. Gillani, Hoyin Chu, Seunghun Han, S. Camp, Eric Kofman, G. Getz, Catherine J. Wu, S. Tyekucheva, M. Davids, E. V. Van Allen, S. AlDubayan, Jennifer R. Brown","doi":"10.1158/2643-3249.bcdsymp24-p01","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p01","url":null,"abstract":"\u0000 Background: Next-generation sequencing has revealed numerous Variants of Unknown Significance (VUS) in the ATM gene, important in DNA repair and chronic lymphocytic leukemia (CLL) pathogenesis. This study evaluates the effect of rare ATM germline variants, including VUS, on features of CLL, ATM kinase activity, and response to PARP inhibition. Methods:ATM variants were detected in 885 CLL patients (327 DFCI; 276 German CLL Study Group; 282 ICGC) using DeepVariant, and compared to 5310 ancestry-matched controls. Concurrently, 278 CLL patients with ATM aberrations from clinical sequencing were retrospectively analyzed, annotated by population allele frequency (JCO 2023, Lampson et al.). ATM kinase activity was measured by ATM and KAP1 phosphorylation following gamma-irradiation in primary CLL samples. The impact of talazoparib, a PARP inhibitor, on ATM aberrant and wild-type CLL cell proliferation was also assessed. Results: Among the 885 CLL cases, 130 ATM germline variants were identified by DeepVariant, including 87 missense variants. The top 5 most frequent rare germline missense variants with high odds ratio for association with CLL were ATM p.L2332P (AF cases vs controls, 0.328% vs 0.0188%, OR 17.46), p.L2307F (OR 16.88), p.F763L (OR 18.89), p.Y1442H (OR 35.39) and p.S99G (OR 16.28). In our retrospective clinical cohort, we identified 294 ATM variants with 146 unique variants. Our previously proposed algorithm for classifying germline vs somatic identified 74 germline, with 70 missense, and 72 somatic. Phosphorylation of ATM and KAP1 (pATM, pKAP1) in response to IR was reduced in 13 patients assessed with germline ATM variants (9 rare and 4 non-rare; ATM variant alone group) compared to those without any somatic or germline ATM aberrancy (WT group) (P<0.01). Phosphorylation levels were also significantly lower in 17 patients with rare germline ATM variants with concurrent del(11q) (del11q + ATM germline variant group) compared to 15 with del(11q) alone (del 11q group) (P<0.01). We obtained similar results from Western blot analysis. To assess the sensitivity of ATM deficient cells to talazoparib, we compared proliferation inhibition across groups cocultured with talazoparib for 14 days. The del11q with ATM-somatic group exhibited significantly greater sensitivity to talazoparib at lower dosages compared to ATM WT (talazoparib 0.1 μM, P<0.05; 0.5μM, P=0.098). The monoallelic ATM deficient groups (del11q alone or ATM-germline alone) tended to have lower proliferation, but not significantly. Higher concentrations of talazoparib (2.5μM) strongly inhibited proliferation of all CLLs including WT. Conclusion: This study highlights the prevalence of rare ATM germline variants in CLL and their impact on ATM function in CLL. Our findings also suggest that these variants affect the DNA damage response in primary CLL cells and that ATM biallelic deficiency influences sensitivity to PARP inhibitors. Our evidence could lead to more personalized therapeut","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140080275","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract P25: Identification of biological components and novel clinical subsets of NPM1-mutated AML using an integrated, multi-omics approach","authors":"Salma Abdelbaky, Kyoko Yamaguchi, Yue-Zhong Wu, Kevin R. Coombes, Lianbo Yu, Christopher C Oakes","doi":"10.1158/2643-3249.bcdsymp24-p25","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p25","url":null,"abstract":"\u0000 Acute myeloid leukemia is the most common acute leukemia in adults with a 5-year overall survival rate of only 28%. Relapse occurs in almost half of adults within 3 years. Due to a high degree of heterogeneity in biological and genetic features in AML tumor cells, current prognostic markers (including genetic aberrations) do not fully explain the ranges of phenotype and outcomes observed in AML patients. Recently, we characterized 13 DNA methylation signatures (epitypes) and a STAT hypomethylation signature (SHS) to be predictive of outcome and stable at relapse. Here, we used the integrative Multi-omics Factor Analysis (MOFA) approach to combine genetic, cytogenetic, transcriptomic, and our novel epigenetic information in an extensive multi-omics analysis to infer latent factors capable of explaining independent signatures comprising NPM1-mutated AML biology. The study used the well-annotated CALGB/Alliance AML cohort encompassing 581 NPM1-mutated patients. The analysis showed that RNA-sequencing contributed the most to explaining the variance between patients followed by RNA splicing, highlighting the depth of information present in the transcriptome. Surprisingly, genetic mutations contributed to only approximately 3% of the overall variance, while DNA methylation captured more than 15% of the total variance. We were able to infer 15 latent factors, 5 of which were associated with overall survival, event-free survival, and attainment of complete remission. All factors were significant after adjusting for established prognostic features, such as FLT3-ITD, sex, and age. Factors 2, 7 and 11 were associated with unfavorable outcomes, and variably included increased HOX signatures, suppressed TP53 activation, a stem cell-like phenotype, the epigenetic SHS signature, triple NPM1/FLT3-ITD/DNMT3A mutations, and alternative splicing, including NUP98. Factors 4 and 13 had a favorable prognostic value. Factor 13 uncovered a unique expression pattern of X-linked cancer testis antigen gene family members dividing NPM1 patients independent of sex, age, or common mutations. The factor was inversely associated with the expression of CD34, MN1 and BAALC – prognostic genes related to stemness. Analysis of immune cell subsets indicated differences in proportions of CD8 effector T cell populations between patients with high vs. low factor 13. This feature pattern and clinical significance were validated in an independent cohort (Beat-AML). Lastly, we clustered the 15 generated factors and identified 5 subsets of NPM1-mutated patients – 2 clusters with a favorable prognostic value, 2 clusters with unfavorable prognostic value, and one intermediate cluster. Overall, we identify 15 main sources of variance within NPM1-mutated patients, including 5 clinically-relevant factors associated with unique biological features, which together generate unique independent biomarker signatures not observed when individually considering separate omics data types.\u0000 Citation Form","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140080491","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract P17: Epigenomic fingerprinting of limited primary immune cells using automated CUT&RUN","authors":"Ellen N Weinzapfel, Keith E. Maier, M. Marunde, V. U. Kumary, Carolina P. Lin Windham, Danielle N. Maryanski, Liz Albertorio-Saez, Dughan J. Ahimovic, Michael J Bale, Juliana J Lee, Bryan J. Venters, Michael-Christopher Keogh","doi":"10.1158/2643-3249.bcdsymp24-p17","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p17","url":null,"abstract":"\u0000 Chromatin structure drives gene expression programs during immune cell development and in cancer, making it central to the advancement of precision medicine. Indeed, the study of chromatin regulation has provided valuable insight on effector cell differentiation and function, anti-tumor immune responses, and therapeutic resistance. Epigenomic features – such as histone post-translational modifications (PTMs) and chromatin-associated proteins – mark distinct genomic compartments (e.g., promoters, enhancers) and regulate chromatin function/gene expression. Mapping the genomic distribution of these may uncover new biomarkers or drug targets and provides a rich context for exploring immunology. Despite widespread interest, integration of epigenomics in large-scale immunotherapy and/or drug research has been hampered by the poor sensitivity, high background, and low throughput of traditional chromatin mapping technologies, most notably ChIP-seq. Instead, efforts to characterize chromatin dynamics have focused on DNA methylation (bisulfite-seq) or chromatin accessibility (ATAC-seq). However, these assays provide an incomplete view of the chromatin landscape, limiting their ability to define cell differentiation pathways or derive mechanistic insight into disease etiology. Here, we present autoCUT&RUN, a high-throughput assay for rapid, ultra-sensitive profiling of epigenomic features from FACS-isolated primary cells or tissues. This workflow generates reliable profiles from ~10,000 cells per reaction, and is supported by a rigorous optimization strategy, high-quality antibodies, and quantitative spike-in controls. As part of a multi-site collaboration with the Immunological Genome Consortium, we used our autoCUT&RUN platform to build a comprehensive epigenomic database of primary mouse immune cells - composed of >1,500 epigenomic profiles from >100 different FACS-isolated immune cell types. These studies set the stage to leverage high-throughput epigenomics in immunotherapy and precision medicine applications.\u0000 Citation Format: Ellen N. Weinzapfel, Keith E. Maier, Matthew R. Marunde, Vishnu U. Sunitha Kumary, Carolina P. Lin Windham, Danielle N Maryanski, Liz Albertorio-Saez, Dughan J. Ahimovic, Michael J. Bale, Juliana J. Lee, Bryan J. Venters, Michael-Christopher Keogh, Immunological Genome Consortium. Epigenomic fingerprinting of limited primary immune cells using automated CUT&RUN [abstract]. In: Proceedings of the Blood Cancer Discovery Symposium; 2024 Mar 4-6; Boston, MA. Philadelphia (PA): AACR; Blood Cancer Discov 2024;5(2_Suppl):Abstract nr P17.","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140080629","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract P22: PVR (CD155) epigenetic status mediates adoptive cell therapy and anti-TIGIT response in Multiple Myeloma","authors":"L. Martinez-Verbo, L. Villanueva, P. Llinàs-Arias, C. García-Prieto, D. Piñeyro, A. Oliver-Caldés, A. García-Ortiz, A. Valeri, C. F. de Larrea, J. Martínez-López, Gerardo Ferrer, Manel Esteller","doi":"10.1158/2643-3249.bcdsymp24-p22","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p22","url":null,"abstract":"\u0000 DNA promoter methylation acts as a key regulator for gene expression and is deeply altered in tumorigenesis. In the context of tumor clearance, cytotoxic cells (i.e. T and NK cells) play a critical role. Their function is mainly controlled by immune checkpoint (IC) synapse events, where known inhibitory markers are detrimental for cytotoxic activation. TIGIT, an inhibitory receptor on immune cells, interacts with PVR on malignant cells, precluding recognition and clearance. Multiple Myeloma (MM) remains and incurable disease and represents 10% of all hematological malignancies. In this project, we have proved PVR is regulated by promoter methylation and the importance of PVR/TIGIT axis inhibition on current immunotherapeutic options for MM. Studying newly diagnosed MM patients (n=793) we observed that higher expression of PVR is associated with shorter overall survival (median overall survival of PVR high: 56 months; PVR low: not reached; Log-rank P<0.001). In silico analysis of the promoter region of PVR, suggested its regulation by DNA methylation, so we characterized the epigenetic regulation of PVR at DNA, RNA and protein levels on a panel of MM cell lines (two methylated: AMO-1, KMS-12_BM and four unmethylated: EJM, JJN-3, MM.1S and RPMI-8226) and validated its epigenetic regulation. We cocultured cytotoxic cells with PVR unmethylated (expressing) models as well as PVR depleted ones. This depletion was obtained by shRNA interference and at least four different donors were evaluated in all the coculture experiments. Three different cell lines (EJM, JJN-3 and RPMI-8226) were tested and all expressing controls showed resistance to cytotoxicity (Mann-Whitney P<0.05). To validate the PVR/TIGIT role, we incorporated 10μg/ml of neutralizing anti-TIGIT into the system. We observed an increase in cytotoxicity in the expressing models as observed in the depleted models (Mann-Whitney P<0.05). We then tested PVR/TIGIT effect on anti-BCMA/CD3 BiTE, anti-BCMA (ARI0002h) CAR-T, and anti-BCMA (ARI0002h) CAR-NK cells. In the presence of BiTE, PVR depletion had a significant impact on cytotoxicity (Mann-Whitney P<0.05) when compared to controls. Coculturing CAR-T cells and MM models showed similar effects (Mann-Whitney P<0.05). As for CAR-NK coculture, only depleting PVR on RPMI-8226 showed a significant increase in cytotoxicity (Mann-Whitney P=0.01) when compared with the control, whereas JJN-3 remained unaffected. In summary, in the context of MM, PVR's expression is regulated by its promoter region's methylation status. Expression of PVR effectively reduces cytotoxic cell response, and the addition of anti-TIGIT validated that the PVR cytotoxic cell inhibition is mediated by TIGIT interaction. Furthermore, immunotherapies benefit from PVR depletion. Finally, PVR/TIGIT interaction seems to impact patients treated with current therapies since their outcome correlates with PVR expression. These results warrant further investigation on PVR as a biomarker and","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140265743","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract P29: Zotatifin, the eukaryotic translation initiation factor 4A (eIF4A) inhibitor, synergizes with Venetoclax to inhibit acute myeloid leukemia cell growth","authors":"Y. Lee, Jonathan Good, Noha Awais, Benjamin Eschle, Jenny Noel, Jean Acosta, Erica DePasquale, David Cucchi, Prafulla Gokhale, Fadi Najm, P. van Galen","doi":"10.1158/2643-3249.bcdsymp24-p29","DOIUrl":"https://doi.org/10.1158/2643-3249.bcdsymp24-p29","url":null,"abstract":"\u0000 New targeted therapies such as the selective and potent BCL-2 inhibitor Venetoclax have revolutionized the treatment of acute myeloid leukemia (AML). However, due to the cellular heterogeneity of the disease, drug-resistant cell types often emerge and drive relapse. We found that the eukaryotic translation initiation factor 4A (EIF4A) is highly expressed in multiple primitive AML cell types compared to their healthy counterparts, suggesting its potential as a therapeutic target. By unwinding complex structures in the 5’ UTR of transcripts, eIF4A promotes the translation of oncogene transcripts. A first-in-class drug, Zotatifin, inhibits the helicase activity of eIF4A and curtails tumor growth in receptor tyrosine kinase-driven tumors. Zotatifin is currently being evaluated in phase 1/2 trials across multiple solid tumors (NCT04092673). However, its effectiveness in blood cancers such as AML remains unexplored, and it is unknown whether Zotatifin may work synergistically with Venetoclax. Here, we treated AML cell lines in vitro and primary healthy or AML bone marrow ex vivo with Zotatifin or Venetoclax alone and in combination, and demonstrate synergistic killing of AML cells by the combination therapy (ZIP synergy score = 57.0 in MOLM-13, 19.3 in MV4-11, P < 0.0001). Mechanistically, we show that Zotatifin decreases eIF4A and AKT expression, and its combination with Venetoclax drives increased BAK and cleaved BAK expression, as well as increased cleavage of MCL-1 and BCL-2, consistent with drug synergy by convergence on the apoptotic pathway. In pilot colony-forming unit (CFU) assays, we found the combination to be tolerable in healthy CD34+ human hematopoietic stem and progenitor cells (HSPCs). To test drug efficacy in vivo, we generated a patient-derived xenograft (PDX) AML model from a patient who experienced relapse with complex cytogenetics, FLT3 and NPM1 mutations. Only the combination of Zotatifin and Venetoclax but not monotherapies effectively suppressed tumor burden as assessed by human CD45 engraftment in the peripheral blood. Further, the combination of Zotatifin and Venetoclax significantly prolonged survival of xenografted mice (median survival after treatment: vehicle 11 days, Venetoclax 13 days, Zotatifin 15 days, combination 42 days, P = 0.0002). Together, our data indicates that Zotatifin is synergistic with Venetoclax and may represent an effective treatment option for patients who are either primary refractory or have developed resistance to existing Venetoclax-containing regimens.\u0000 Citation Format: Yoke Seng Lee, Jonathan Good, Noha Awais, Benjamin Eschle, Jenny Noel, Jean Acosta, Erica DePasquale, David Cucchi, Prafulla Gokhale, Fadi Najm, Peter van Galen. Zotatifin, the eukaryotic translation initiation factor 4A (eIF4A) inhibitor, synergizes with Venetoclax to inhibit acute myeloid leukemia cell growth [abstract]. In: Proceedings of the Blood Cancer Discovery Symposium; 2024 Mar 4-6; Boston, MA. Philadelphia (PA): AACR; ","PeriodicalId":29944,"journal":{"name":"Blood Cancer Discovery","volume":null,"pages":null},"PeriodicalIF":11.2,"publicationDate":"2024-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140266282","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}