Abstract 3953: BET inhibitor increases DNA damage, modulates Wnt signaling, and suppresses osteosarcoma growth in naïve and metastatic disease models
Authors: Riyahi Niknam , Pandya Pankita H. , Bailey Barbara J. , Dobrota Erika A. , Young Courtney , Shannon Harlan E. , Barghi Farinaz , Malko Rada , Bijangi-Vishehsaraei Khadijeh , Trowbridge Melissa A. , Coy Kathy , Kennedy Felicia M. , Sinn Anthony L. , Angus Steve , Ferguson Michael J. , Saadatzadeh M. Reza , Pollok Karen E. 
Published: March 2024

Abstract
Osteosarcoma (OS) is an aggressive bone cancer in pediatric adolescent and young adult patients. Survival rate for metastatic and relapsed OS patients remains dismal at &lt;30%. Additionally, no effective standardized salvage therapy currently exists for these patients, in part, due to genomic complexities arising from moderate levels of replication stress (RS). Bromodomain and extra-terminal domain protein inhibitors (BETi) are an underexplored option to target RS for BETi creates an imbalance between transcription-replication kinetics resulting exacerbation of oncogenic RS and cell death. BET proteins are epigenetic readers that play a role in regulating gene expression networks as well as DNA replication and repair. We tested the hypothesis that BET inhibition leads to decreased OS cell growth and potentiates the efficacy of salvage therapy via gene dysregulation and increased DNA damage and RS. Combination index and Bliss independence analyses of bivalent BET inhibitor (BETi) AZD5153 or PROTAC ARV825 in combination with salvage agents demonstrated additive-to-synergistic cell growth inhibition in OS lines. Increased apoptotic-mediated cell death was observed following AZD5153+topotecan. In addition, γ-H2AX levels and comet assays demonstrated that BETi+topotecan induces its effect, in part through increased RS in-vitro. AZD5153 monotherapy significantly suppressed OS tumor growth in patient-derived xenografts (PDXs) derived from both naïve (PDX96) and metastatic (TT2) OS compared to vehicle (p&lt;0.05). Moreover, AZD5153 reduced lung metastatic lesions and increased survival in the 143B OS lung colonization model. In-vivo mechanisms of drug-induced tumor response were evaluated. Anti-tumor effect correlated with increased γ-H2AX following AZD5153 exposure in PDX96, indicative of increased RS. RNA-seq analysis and protein validation from vehicle versus BETi-treated PDX96 highlighted dysregulation of several key genes involved in DNA damage response including elevated TXNIP, a tumor suppressor which can induce DNA damage and apoptosis. Downregulation of TCF7, a downstream effector of the Wnt pathway, was also observed following BETi. Global kinome profiling and validation experiments revealed increased activity and expression of EphA2 and EphA4 receptors both implicated in development of drug resistance. Furthermore, in BETi-treated PDX96 tumors, Wnt/β-catenin pathway activation emerged as tumors rebounded once BETi treatment was stopped. TT2 PDX was resistant to commonly used salvage agents ifosfamide and irinotecan; however, combination BETi+ topotecan increased the probability of survival compared to each agent alone (p&lt;0.05) and was well tolerated. These data collectively suggest that BET inhibition alone or in combination with low-dose salvage therapy holds promise as novel treatment strategies in aggressive OS.
Citation Format: Niknam Riyahi, Pankita H. Pandya, Barbara J. Bailey, Erika A. Dobrota, Courtney Young, Harlan E. Shannon, Farinaz Barghi, Rada Malko, Khadijeh Bijangi-Vishehsaraei, Melissa A. Trowbridge, Kathy Coy, Felicia M. Kennedy, Anthony L. Sinn, Steve Angus, Michael J. Ferguson, M. Reza Saadatzadeh, Karen E. Pollok. BET inhibitor increases DNA damage, modulates Wnt signaling, and suppresses osteosarcoma growth in naïve and metastatic disease models [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 3953.