Discovery of INCB123667, a potent and selective cyclin-dependent kinase 2 (CDK2) inhibitor for the treatment of cyclin E dysregulated cancers

Background: Dysregulation of the CDK/cyclin complex causes aberrant cell cycle entry and progression that can lead to cancer. For instance, the CDK2/ cyclin E complex governs the G1-S transition and overexpression of cyclin E has been associated with ovarian and breast cancers. Our initial studies have demonstrated that genetic depletion of CDK2 inhibits cell growth and survival in cancer cells overexpressing cyclin E. Furthermore, inhibition of CDK2 may be effective in cancers resistant to clinically approved CDK4/6 therapies as increased CDK2/cyclin E activity has been reported as a compensatory mechanism. Current CDK2 inhibitors are limited by off-target activity on other CDK family members, including CDK1. Here we report the identification and characterization of INCB123667, a potent and selective, orally available small molecule inhibitor of CDK2, for the treatment of cancers with high unmet medical need, such as cyclin E-amplified ovarian tumors. Materials and methods: Biochemical and cell-based assays were used to determine the potency and selectivity of INCB123667. Additionally, antitumor activity was assessed in cyclin E-overexpressing cancer cell lines, and in tumor xenograft models. Results: In biochemical assays, INCB123667 demonstrated sub-nanomolar inhibition of CDK2 activity and was not active against CDK1, CDK4, CDK6, CDK7, and CDK9. Selectivity of INCB123667 was confirmed across an extended panel of kinases, G protein-coupled receptors, ion channels, and transporters. In cell-based studies, INCB123667 inhibited phosphorylation of retinoblastoma (Rb), a CDK2/cyclin E substrate and key regulator of the G1-S transition. Consistent with its overall selectivity profile, INCB123667 inhibited the expression of cell cycle regulatory genes in cyclin E-overexpressed cancer cells. Furthermore, the transcriptional gene signature of INCB123667 was distinct from that reported for CDK4/6 inhibitors. In cellular assays, addition of INCB123667 resulted in G1 cell cycle arrest and cell senescence. In the OVCAR3 tumor xenograft model, which harbors cyclin E amplification, INCB123667 inhibited Rb phosphorylation and resulted in dose-dependent tumor growth inhibition along with minimal body weight loss. In the IGROV-1 model, in which cyclin E is not amplified, the same doses of INCB123667 were less effective in blocking Rb phosphorylation and inhibiting tumor growth. Notably, across in vitro and in vivo studies, cyclin E-overexpressing cancer cells were more sensitive to INCB123667 compared with cells with normal levels of cyclin E. Conclusions: Our data indicate that INCB123667 is a selective and potent CDK2 kinase inhibitor that has antitumor activity in cyclin E-overexpressing cancers and tumor xenograft models. These preclinical findings support the ongoing clinical evaluation of INCB123667 as a CDK2-selective inhibitor for the treatment of cancer. Conflict of interest: Ownership: Susan Wee, Yvonne Lo, Michael Hansbury, Michael Weber, Valerie Roman, Lu Huo, Katherine Drake, Kristine Stump, Jie Yang, Saswati Chand, Cynthia Timmers, Joshua Hummel, Guofeng Zhang, Yan-ou Yang, Maryanne Covington, Sunkyu Kim: Employment and stock ownership – Incyte Corporation. Min Ye, Niu Shin, Hollie Skaggs, Kanishk Kapilashrami, Yingda Ye, Liangxing Wu, Holly Koblish: Former employee and stock ownership – Incyte Corporation.