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Abstract 2017: Therapeutic induction of replication stress in the context of salvage therapy in osteosarcoma


 Osteosarcoma (OS) is an aggressive pediatric cancer with ~35% of patients developing metastasis over time. The survival rate for metastatic and relapsed OS patients is <30% and there is currently no standardized salvage therapy. Lack of efficacy is attributed to extensive genetic complexity present in OS that is partly due to moderate levels of replication stress (RS). While high levels of RS can induce cell death, moderate RS levels may cause genomic instability that contributes to progression of OS. Therefore, induction of RS to high levels, especially in genetically complex cancers like OS, could be a promising therapeutic strategy. Bromodomain and extra-terminal domain (BET) proteins (BRD2,3,4) are a family of epigenetic readers that not only regulate gene expression networks, but also regulate DNA replication and RS. BRD4 directly regulates major factors involved in DNA replication and checkpoint signaling. Thus, disruption of BRD4 function should exacerbate RS to levels that cause cell death. The objective of this study is to test the hypothesis that BET inhibition potentiates the efficacy of current salvage therapy through RS induction in aggressive OS. The effects of BET inhibitors (BETi), AZD5153 and OTX-015, as single agents and in combination with drugs used in salvage therapy such as topotecan were evaluated for effects on OS cell growth, PARP cleavage, and the DNA damage repair network. BET knockdown experiments were performed to evaluate target selectivity and dependency. In vivo efficacy and safety studies focused on patient-derived xenografts (PDXs) of relapsed OS. TT2-77 xenoline, Saos2, G292, and U2OS cell lines were selected for in vitro experiments. Combination index and Bliss independence analyses demonstrated additive to synergistic cell growth inhibition upon treatment with clinically relevant concentrations of BETi+topotecan. Significant increase in PARP cleavage was observed in the combination compared to single agent, indicating enhancement of apoptosis. Moreover, Western analyses demonstrated that BETi induces its effect, at least partly, via decreased CHK1 activation and increased DNA damage. Selective siRNA treatments illustrated that transient knockdown of individual BET proteins was not sufficient for potentiation of topotecan-induced cell death in OS cells, indicating that simultaneous knockdown of BETs may be required. Dose-finding studies of AZD5153 in relapsed OS PDXs that harbor replication stress signatures (TT2-77 and PDX96) indicated that daily doses of 1.25 or 2.5 mg/kg AZD5153 were well tolerated and effective in partially suppressing tumor growth compared to vehicle (p<0.05, Two-way ANOVA; Holm-Sidak). In vivo combination treatments of BETi+topotecan are in progress. These data collectively suggest that BET inhibition alongside salvage therapy holds promise as a novel treatment strategy for inducing RS-mediated cell death in aggressive OS.
 Citation Format: Niknam Riyahi, Pankita H. Pandya, M. Reza Saadatzadeh, Khadijeh Bijangi-Vishehsaraei, Barbara J. Bailey, Erika A. Dobrota, Courtney Young, Melissa A. Trowbridge, Kathy Coy, Henry Mang, Reagan K. Wohlford, Anthony L. Sinn, Emily S. Sims, Matt J. Repass, Nuri Damayanti, Farinaz Barghi, Harlan E. Shannon, Michael J. Ferguson, Jamie L. Renbarger, Karen E. Pollok. Therapeutic induction of replication stress in the context of salvage therapy in osteosarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2017.