Abstract 363: Genetic or pharmacological inactivation of CREBBP sensitizes B-cell acute lymphoblastic leukemia to ferroptotic cell death upon BCL2 inhibition
Authors: Gimenez Alicia Garcia , Ditcham Jonathan , Azazi Dhoyazan M. , Meduri Eshwar , Asby Ryan , Sakakini Nathalie , Lopez Cecile K. , Narayan Nisha , Bagri Jaana , Beinortas Tumas , Singh Shuchi Agrawal , Giotopoulos George , Murphy Michael P. , Horton Sarah J. , Huntly Brian J. , Richardson Simon E. 
Published: March 2024

Abstract
B-cell acute lymphoblastic leukemia (B-ALL) is an aggressive hematological malignancy of B lineage progenitors. It remains a leading cause of death in childhood, while outcomes in adults are dismal. There is therefore a need to better understand drivers of high-risk B-ALL and to develop novel therapeutic approaches targeting these challenging patient cohorts. Loss-of-function mutations affecting CREBBP are recurrent second-hit mutations across multiple genetic subtypes of B-ALL and are associated with adverse features, including high-risk genetic subtypes and persistent measurable residual disease. In addition, they have been mechanistically associated with chemoresistance and are more frequently found in relapse. We sought to identify novel treatment options for CREBBP-mutated high-risk B-ALL. CREBBP-mutated isogenic human B-ALL cell lines were genome-engineered to provide a platform for synthetic lethal drug screening. We subjected these cell lines to a targeted drug screen focused on clinically-actionable drugs in classes that have either been implicated or hypothesized to show differential sensitivity patterns in published models of B cell lymphoma and other CREBBP-mutated malignancies. Unexpectedly, CREBBP-mutated cells were not differentially sensitive to traditional cytotoxic chemotherapy, and paradoxically showed a degree of sensitization to the glucocorticoid Dexamethasone, used in current ALL induction regimens. As anticipated, and validating our screen design, inhibitors of the CREBBP paralogue EP300 (the CREBBP/EP300-specific bromodomain inhibitor Inobrodib and the CREBBP/EP300 acetylase inhibitor A485) exhibited synthetic lethality, consistent with previous reports in B-cell lymphoma. The most potent hit was the BCL2 inhibitor Venetoclax, which we show acts through a non-canonical, but BCL2-dependent mechanism resulting in ferroptotic programmed cell death. CREBBP-mutated cell lines were transcriptionally and functionally characterized, revealing underlying differences in cell-cycle, metabolism and response to oxidative stress. Acquisition of resistance to Venetoclax further dysregulated these pathways and resulted in a transcriptionally-convergent state. Lastly, we demonstrate that small-molecule inhibition of CREBBP sensitizes B-ALL cells, regardless of genotype, to Venetoclax-induced ferroptosis in vitro and in vivo, providing a potential novel drug combination for broader clinical translation in B-ALL. In summary, we have identified a number of actionable compounds that specifically target CREBBP-mutated high-risk B-ALL, demonstrate a novel mechanism-of-action for the BCL2 inhibitor Venetoclax in B-ALL and propose CREBBP-inhibitors and Venetoclax as a novel treatment combination for B-ALL across genotypes.
Citation Format: Alicia Garcia Gimenez, Jonathan Ditcham, Dhoyazan M. Azazi, Eshwar Meduri, Ryan Asby, Nathalie Sakakini, Cecile K. Lopez, Nisha Narayan, Jaana Bagri, Tumas Beinortas, Shuchi Agrawal Singh, George Giotopoulos, Michael P. Murphy, Sarah J. Horton, Brian J. Huntly, Simon E. Richardson. Genetic or pharmacological inactivation of CREBBP sensitizes B-cell acute lymphoblastic leukemia to ferroptotic cell death upon BCL2 inhibition [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 363.