Abstract 1636: Novel CDK12/13 inhibitor CTX-439 downregulates MCL1 via transcriptional mRNA read-through and synergistically acts with BCL-XL inhibitors for cancer therapy

Abstract Combination therapeutic strategies for cancer use multiple agents targeting distinct pathways to enhance the treatment efficacy and minimize resistance. By simultaneously disrupting key cancer cell survival mechanisms, these strategies improve the outcomes better than monotherapies, effectively combating cancer complexity and adaptability. Cyclin-dependent kinase (CDK)-12/13 regulates RNA transcription by phosphorylating RNA polymerase II, promoting the elongation of transcripts involved in the DNA damage response, such as BRCA1/2. As an emerging drug target, the potential of CDK12 to enhance the poly (ADP ribose) polymerase (PARP) inhibitor effects has been re-evaluated. CDK12/13 and PARP inhibitors exert synergistic effects. CDK12/13 inhibitor is currently undergoing clinical trial. However, therapeutic potential of CDK12/13 inhibitor combinations with other factors beyond PARP remains unclear, necessitating further investigations to expand their clinical applications. We successfully developed CTX-439, a novel orally bioavailable ATP-competitive CDK12/13 small-molecule inhibitor (Yamakawa et al., AACR2022/2024). CTX-439 exhibited anti-tumor activity alone and in combination with PARP inhibitors. To identify other factors synergistic with CTX-439, we performed a CRISPR activation screening using the SUM149 breast cancer cell line and identified four genes, whose overexpression conferred CTX-439 resistance. Anti-apoptotic factors BCL-XL and BCL2 were among the top candidates, whose inhibition enhanced CTX-439 efficacy. Moreover, dual inhibition of CTX-439 and AZD4320, which targets BCL-XL, synergistically induced rapid apoptosis in vitro and significantly suppressed tumor growth in vivo. Mechanistically, CTX-439 downregulated the protein levels of anti-apoptotic factor MCL1, but not BCL-XL, thereby shifting the dependency to BCL-XL for cell survival, suggesting BCL-XL inhibition as a critical apoptosis mechanism in CTX-439 combination strategies. Notably, CTX-439 significantly increased the read-through transcription of the MCL1 mRNA, leading to its nuclear retention. Furthermore, lack of mature mRNA supply resulted in the rapid turnover of cytosolic MCL1 mRNA due to its short half-life, leading to the downregulation of MCL1 protein levels. In summary, previously reported combination strategies with CDK12/13 inhibitors focused on regulating long mRNAs, particularly those associated with the DNA damage response. Interestingly, our study revealed a novel CDK12/13 inhibition mechanism targeting short mRNAs, such as MCL1, that can be used to establish new therapeutic combination strategies with BCL-XL inhibitors. Additionally, our study provides a new avenue to improve cancer therapy outcomes. Citation Format: Hiroko Yamakawa, Raghda Khatab, Yusuke Tarumoto, Mariko Takano, Seiichi Sugino, Shafiqul Islam, Shunsuke Ebara, Kosuke Kawaguchi, Masakazu Toi, Akio Mizutani, Midori Sugiyama, Misaki Yoshida, Daisuke Morishita, Kosuke Yusa. Novel CDK12/13 inhibitor CTX-439 downregulates MCL1 via transcriptional mRNA read-through and synergistically acts with BCL-XL inhibitors for cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1636.