Inhibition of RNA polymerase II-activating CDK9 and CDK12/13, but not of cell cycle relevant CDKs, induces apoptosis by downregulating the short-lived Bcl-2 proteins Mcl1 and Bfl1/A1

Abstract Cyclin-dependent kinases (CDKs) play a crucial role in cell cycle (such as CDK1 and CDK4/6) and transcription (such as CDK7, CDK9, and CDK12/13). While CDK inhibitors are clinically effective in cancer therapy, their mechanisms of apoptosis induction remain incompletely understood. Here, we demonstrate that inhibition of cell cycle CDKs (CDK1, CDK4/6) or transcriptional initiation (CDK7) did not induce cytotoxicity. Only inhibition of CDK9 (by AZD4573 and atuveciclib) and of CDK12/13 (by SR4835 and THZ531) – which target the transcriptional elongation of RNA polymerase II (RNAPII) – exerted a strong apoptotic potential. Since CDK9 and CDK12/13 target different elongation factors associated with RNAPII (such as SPT6 and NELF for CDK9; CDC73, and LEO1 for CDK12/13), they cannot substitute for each other. Consequently, the combination of AZD4573 with SR4835 resulted in significant, synergistic cytotoxicity. Inhibiting CDK9 or CDK12/13 induced the rapid downregulation of the short-lived, anti-apoptotic Bcl-2 proteins Mcl1 and A1/Bfl1 in Jurkat leukemia cells and SUDHL1 lymphoma cells, whereas the expression of Bcl-2 and Bcl-xL remained unaffected. Since Mcl1 and A1 antagonize the pro-apoptotic Bcl-2 proteins Bak and Bax, apoptosis induction by AZD4573 or SR4835 was blocked in Bak- and Bax/Bak-deficient Jurkat cells and strongly reduced in Bax-deficient cells. Because Bcl-2 only inhibits Bax, but not Bak, AZD4573 and SR4835 were able to induce apoptosis in Jurkat cells overexpressing Bcl-2. As tumor cells frequently upregulate Bcl-2, inhibitors of CDK9 and CDK12/13 represent promising anticancer drugs.