Abstract A037: CDK12 – A druggable dependency and potential therapeutic target of desmoplastic small round cell tumor

Abstract Desmoplastic small round cell tumor (DSRCT) is a pediatric tumor driven by the fusion oncoprotein EWSR1-WT1 formed as a result of a recurrent translocation event between chromosomes 11 and 22. It is a rare malignancy with a 5-year survival rate of 15-20%. Owing to the difficulties in developing targeted therapies for specific oncogenic transcription factors, there is a need to identify DSRCT-specific dependencies. We performed a CRISPR-Cas9 drop-out screen against the human genome and identified Cyclin-dependent Kinase CDK12 to be a selective DSRCT dependency. To validate this finding, doxycycline(dox)-inducible shRNA system was used to establish a stable knockdown of CDK12 expression in JN-DSRCT-1 and BER-DSRCT cell lines. Dox-dependent knockdown of CDK12 was validated by Western blot and quantitative RT-PCR. Consistent with our CRISPR screen, silencing CDK12 decreased DSRCT cell proliferation as indicated by colony formation assays. CDK12 complexes with Cyclin K to phosphorylate the C-terminal domain (CTD) of RNA Polymerase II, which is required for transcription initiation and elongation. THZ531, a CDK12/13 selective inhibitor, binds to the CDK12/Cyclin K complex to inhibit its activity. THZ531 treatment significantly reduced cell viability with IC50 values of 59.25 nM and 51.31 nM in JN-DSRCT-1 and BER-DSRCT, respectively. In contrast, the normal mesothelial cell line LP9 showed a much lower susceptibility to THZ531 with an IC50 of 183.8 nM. THZ531 led to a significant reduction in the phosphorylation of Ser2 of the CTD of RNA polymerase II after 6 hours of treatment, demonstrating effective inhibition of CDK12 activity. A prolonged treatment of THZ531 (24h) significantly increased PARP cleavage in JN-DSRCT-1 and BER-DSRCT cell lines and CDK12 inhibition causes downregulation of large DNA damage response genes. These results indicate that CDK12 inhibition decreases transcription initiation and elongation and induces apoptosis in DSRCT cell lines, making it a potential therapeutic target for DSRCT. Furthermore, co-treatment of DSRCT cell lines with THZ531 and DNAPK inhibitor KU-57788 synergistically reduced DSRCT cell growth and proliferation. These findings indicate that CDK12 is a critical dependency and therapeuic target in DSRCT suggesting that combinatorial inhibition strategies may enhance antitumor efficacy. Citation Format: Shruthi Sanjitha Sampath, Justin W Magrath, Dane A Flinchum, Sean B Lee. CDK12 – A druggable dependency and potential therapeutic target of desmoplastic small round cell tumor [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Discovery and Innovation in Pediatric Cancer— From Biology to Breakthrough Therapies; 2025 Sep 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2025;85(18_Suppl_2):Abstract nr A037.