Abstract 5485: Discovery and preclinical evaluation of a novel highly selective and potent CDK12 inhibitor
Authors: Yamakawa Hiroko , Mizutani Akio , Arikawa Yasuyoshi , Ebara Shunsuke , Satoh Yoshihiko , Morishita Daisuke 
Published: June 2022

Abstract
Recent research highlights that RNA processing is systematically altered in cancer, demonstrating the pivotal influence of RNA deregulation on tumorigenesis, growth and progression. Based on our expanding knowledge of RNA biology, RNA deregulation has drawn much attention from the perspective of cancer therapy. Indeed, small molecules that attack the RNA maturation processes and produce aberrant RNA are currently under development.
The first step in gene expression is transcription, which involves copying a DNA sequence to make an RNA. Transcription is performed by enzymes called RNA polymerase (Pol II) that links nucleotides to form RNA strand. The C-terminal domain (CTD) of Pol II comprises heptapeptide Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7 repeats and is dynamically post-translationally phosphorylated to regulate the distinct stages of transcription initiation, elongation and termination steps. Particularly, Ser2 and Ser5 phosphorylation have the closest association with the regulation of transcription. Therefore, we focus on the kinases that carry out this modification.
Cyclin-dependent kinase 12 (CDK12) belongs to the cyclin-dependent kinase (CDK) family of serine/threonine protein kinases. CDK12 regulates the elongation step of RNA transcription by phosphorylation of Ser2 on the CTD. CDK12 complexes with cyclin K to promote the elongation of transcripts, such as BRCA1 and BRCA2, involved in DNA damage responses. It is expected that the inhibition of CDK12 have a synergistic effect with PARP inhibitors and chemotherapeutic reagents.
Here we present data on CRD-1835439, orally available, selective, and first in class CDK12 inhibitor with optimized drug-properties. Biochemically, CRD-1835439 is more than 300-fold selective over other CDK isoforms such as CDK7 and CDK9. Importantly, cell based assays showed CRD-1835439 has selective inhibitory activity for phosphorylation of Ser2 on the CTD but not for other Ser5 and Ser7 phosphorylation. Comprehensive analysis by PolyA-seq, Chip-seq and RNA-seq revealed that CRD-1835439 inhibits transcriptional elongation on DNA damage response genes including BRCA1 and BRCA2. The effects on DNA damage and repair were assessed by immunofluorescence staining for γH2AX and RAD51 proteins. In vivo, oral treatment with CRD-1835439 in cell line derived xenograft models, resulted in increase of DNA damage biomarkers, induction of apoptosis and tumor regressions in a dose-dependent fashion. Besides the efficacy as a single reagent, the efficacy was augmented when CRD-1835439 was combined with PARP inhibitors in in vitro and in vivo.
In summary, the novel small-molecule CDK12 inhibitor CRD-1835439 demonstrated preclinical efficacy along with target engagement. Our results underscore the preclinical therapeutic potential of CRD-1835439 as a single-agent or in combination with PARP inhibitors for the treatment of intractable cancers.
Citation Format: Hiroko Yamakawa, Akio Mizutani, Yasuyoshi Arikawa, Shunsuke Ebara, Yoshihiko Satoh, Daisuke Morishita. Discovery and preclinical evaluation of a novel highly selective and potent CDK12 inhibitor [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 5485.