-Author name in bold denotes the presenting author
-Asterisk * with author name denotes a Non-ASH member
Clinically Relevant Abstract denotes an abstract that is clinically relevant.

PhD Trainee denotes that this is a recommended PHD Trainee Session.

Ticketed Session denotes that this is a ticketed session.

53 Failure to Execute Caspase Activation Drives Therapy Resistance in TP53 Mutated AML

Program: Oral and Poster Abstracts
Type: Oral
Session: 604. Molecular Pharmacology and Drug Resistance: Myeloid Neoplasms: Determinants of Venetoclax Resistance and Response
Hematology Disease Topics & Pathways:
Research, Fundamental Science, Acute Myeloid Malignancies, AML, Apoptosis, Translational Research, Diseases, Myeloid Malignancies, Biological Processes, Molecular biology
Saturday, December 7, 2024: 10:30 AM

Ahmed M Mamdouh1*, Fang Qi Lim2*, Yang Mi1*, Elyse A Olesinski3*, Yichen Wang1*, Shaista Shabbir Jasdanwala1*, Karanpreet Singh Bhatia1,4, Xiao Xian Lin5*, Valeriia Sapozhnikova, PhD6,7,8*, Philipp Mertins, PhD9*, Brandon D. Brown, MD10*, Yihua Qiu, MD11*, Anthony G. Letai12, Alexandre Puissant, PhD13, Steven M. Kornblau, MD11, Jan Krönke, MD6,14, Edward Ayoub, PhD15*, Koji Itahana, PhD16*, Michael Andreeff, MD, PhD15 and Shruti Bhatt, PhD1

1Department of Pharmacy and Pharmaceutical Sciences, National University of Singapore, Singapore, Singapore
2Department of Pharmacy and Pharmaceutical Sciences, National University of Singapore, Singapore, AL, Singapore
3Boston University Chobanian & Avedisian School of Medicine, Boston, MA
4Winship Cancer Institute, Emory University, Atlanta, GA
5Department of Pharmacy and Pharmaceutical Sciences, National University of Singapore, Singapore, Singapore, Singapore
6Department of Hematology, Oncology, and Cancer Immunology, Charité – Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt Universität zu Berlin, and Berlin Institute of Health, Berlin, Germany
7German Cancer Consortium (DKTK), German Cancer Research Center (DKFZ), Heidelberg, Germany
8Max Delbrück Center for Molecular Medicine, Berlin, Germany
9Core Unit Proteomics, Berlin Institute of Health at Charité, Universitätsmedizin Berlin and Max-Delbrück-Center for Molecular Medicine, Berlin, Germany
10Department of Pediatrics, MD Anderson Cancer Center, Houston, TX
11Department of Leukemia, The University of Texas MD Anderson Cancer Center, Houston, TX
12Department of Medical Oncology, Dana Farber Cancer Institute, Boston, MA
13Saint-Louis Research Institute, Paris, France
14German Cancer Consortium (DKTK) partner site Berlin and German Cancer Research Center (DKFZ), Heidelberg, Germany
15Section of Molecular Hematology and Therapy, Department of Leukemia, The University of Texas MD Anderson Cancer Center, Houston, TX
16Cancer & Stem Cell Biology Program, Duke-NUS Graduate Medical School Singapore, Singapore, Singapore

Among the spectrum of mutations driving malignancy, mutation in the TP53 gene is perhaps the most important event leading to therapy resistance. Only ~25% of patients with TP53-mutated acute myeloid leukemia (AML) respond to standard induction therapy, with dismal median survival of 4.2 months. This underscores a pressing need to characterize therapy resistance mechanisms for TP53-mutated AML and to identify novel therapeutic approaches.

To investigate TP53 mutation-driven chemoresistance, we generated isogenic AML cells harboring TP53 WT, mutant, or knockout (KO) variants. TP53 WT cells exhibited the highest sensitive to Venetoclax+Azacytidine (VenAza), while TP53-/mut cells displayed varying sensitivities, with TP53-/R248Q cells being the least responsive. This variability suggests that different mutations confer distinct levels of TP53 function loss. TP53-/mut and KO AML cells showed reduced sensitivity to cytarabine, etoposide, and VenAza-induced apoptosis without impairment of G1 arrest or cell cycle. To pinpoint the dysfunction in intrinsic apoptotic signaling allowing TP53 mutant cells to evade apoptosis, we performed Gene Set Enrichment Analysis (GSEA) on RNA-seq data. This analysis demonstrated significant enrichment of p53 signaling pathways in WT cells treated with VenAza, but not in TP53-/R248Q and KO cells. Further we observed that despite the loss of key pro-apoptotic regulators (BAX, PUMA, NOXA), upregulation inactivator protein BIM compensated for proapoptotic stimuli in TP53-/R248Q and KO after VenAza. While mitochondrial outer membrane permeabilization (MOMP) is typically considered a crucial step in apoptosis, BH3 profiling analysis revealed comparable MOMP (priming) at baseline between TP53-mutant and TP53-WT AML primary tumors (n=37) and cell lines. This suggests that MOMP is not the definitive "point-of-no-return" for TP53 mutants. Instead, we found a blockade in executioner caspase-3/7 activity in TP53-/mut and KO AML cells, following VenAza or cytotoxic chemotherapy, contrasting with WT cells. This finding was also recapitulated in TP53 null isogenic solid cancer cell lines, revealing a novel post-MOMP resistance mechanism in TP53-deficient cells across cancer types.

To identify targetable dependencies driving caspase blockade in TP53 defective cells, we conducted whole-genome CRISPR knockout screen in WT, TP53-/R248Q, and KO cells following VenAza treatment. Our findings confirmed that knockout of pro-apoptotic genes (BAX, PMAIP1, BCL2L11) conferred resistance across all genotypes. Notably, we discovered a novel functional selective dependence on BIRC5, a member of the inhibitor of apoptosis (IAP) family, in TP53-/R248Q cells exposed to VenAza. Mechanistically, BIRC5 deletion strongly sensitized TP53-mutant/KO cells to VenAza and other chemotherapy drugs by restoring caspase 3/7 activation. RNA-seq and global proteomic analyses of isogenic cells revealed upregulation of IAP family genes in TP53-mutant/KO cells. We validated translational implications of these findings by differential RNA-seq analysis comparing TP53-mutants (n = 63) and WT (n = 668) AML patient samples, and reverse phase protein array (RPPA) in TP53-mutants (n=153) as compared to TP53-WT (n=501) primary tumors. Both analyses identified BIRC5, encoding survivin, as one of the top overexpressed targets in TP53-mutant primary tumors. Extending beyond AML, TCGA data shows BIRC5 upregulation in TP53-mutant tumors across 19/25 cancers, indicating a pan-cancer dependency in p53-inactivated tumors.

To uncover targetable vulnerabilities, we conducted high-throughput drug screening (n=250 agents) in isogenic AML cells, with IAP inhibitors emerging as top hits against TP53-mutant/KO cells. In isogenic mice (CDX) models, a combination of VenAza with IAP inhibitor demonstrated strikingly durable leukemia blast inhibition in the bone marrow and spleen, with triple therapy showing superior efficacy in TP53-/R248Q mice compared to WT treated with VenAza alone (median survival 44.5 vs 26 days). Our research unveils a critical post-mitochondrial defect in TP53 mutant cells as a key driver of chemoresistance and shows IAP inhibition can reinstate the apoptotic potential of both cytotoxic chemotherapy and targeted therapies. This insight offers a potential therapeutic target for TP53-mutated cancers, spanning both hematological and solid tumors.

Disclosures: Letai: Flash Therapeutics: Membership on an entity's Board of Directors or advisory committees; Zentalis Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Dialectic Therapeutics: Membership on an entity's Board of Directors or advisory committees; AbbVie: Research Funding; Novartis: Research Funding; AstraZeneca: Research Funding. Krönke: Charité – Universitätsmedizin Berlin: Current Employment; Janssen, Takeda, Abbvie, Jazz Pharmaceuticals, Sanofi, Astra Zeneca, Pfizer: Consultancy; Janssen, Takeda, Abbvie, Jazz Pharmaceuticals, Sanofi, Astra Zeneca, Pfizer: Honoraria; Janssen, Takeda, Abbvie, Jazz Pharmaceuticals, Sanofi, Astra Zeneca, Pfizer: Speakers Bureau. Andreeff: Kintor Pharmaceutical: Research Funding; Ellipses: Research Funding; Oxford Biomedical: Research Funding; Daiichi-Sankyo: Research Funding; Oncolyze: Current holder of stock options in a privately-held company; Chimerix: Current holder of stock options in a privately-held company; Roivant: Honoraria; Paraza: Honoraria; Boehringer-Ingelheim: Honoraria; Sellas: Honoraria, Research Funding; Ona: Honoraria; Aptose: Honoraria; Glycomimetics: Honoraria; Syndax: Honoraria, Research Funding; SentiBio: Current holder of stock options in a privately-held company, Honoraria, Research Funding; Eterna: Current holder of stock options in a privately-held company, Honoraria, Research Funding.

*signifies non-member of ASH