Abstract 1553: Discovery of CDK4/6 bifunctional degraders for ER+/HER2- breast cancer andtriple negative breast cancer
Authors: Majeski Hannah , Okano Akinori , Pasis Angela , Carlson Casey , Liu Qiao , Shakya Arvind , Huang Shenlin , Chourasia Aparajita Hoskote , Fung Leah 
Published: June 2023

CDK4/6 inhibitors (CDK4/6i) such as palbociclib and ribociclib are used to treatER+/HER2- breast cancer, but patients can develop resistance via mechanismsincluding the INK4-CDK6 complex which have been shown to limit the effectiveness ofCDK4/6i in ER+ breast cancer. Up to 20% patients exhibit innate resistance and up to70% patients develop acquired resistance after 3 years on therapy (Scheidemann,2021). To address this limitation, we utilized our PRODEGY platform of Cereblon(CRBN) binders to synthesize CRBN mediated CDK4/6 bifunctional degraders topotently inhibit tumor growth in CDK4/6i resistant tumors and in treatment naïveER+/HER2- breast cancer and triple negative breast cancer (TNBC). We examined theeffect of our degraders on multiple nodes of this pathway. Target degradation byimmunoblot analysis of TNBC cell line, MDA-MB-231, treated with our CDK4/6bifunctional degraders for 6 hours showed over 50% degradation of CDK4 and CDK6at 10-100nM. Active CDK4/6 phosphorylates the protein RB which releases thetranscription factor E2F, inducing the expression of genes which promote cell cycleprogression. Analysis of RB phosphorylation by in-cell western upon 24 hour ofCDK4/6 degrader treatment showed phospho-RB IC50s at &lt;50nM. Cell cycle analysisby staining with propidium iodide after 24 hours of treatment with CDK4/6 degradersinduced G0/G1 cell cycle arrest at concentrations as low as 10nM. As a readout forgrowth inhibition through cell cycle arrest, we examined the ability of these degradersto inhibit the proliferation of MDA-MB-231 cells in a 2D colony formation assay (CFA)over a 10-day period. Our CDK4/6 degraders showed potent inhibition of cellproliferation with CFA IC50s of &lt;50nM. We demonstrated that our CDK4/6 bifunctionaldegraders were significantly more potent in vitro than the CDK4/6i, palbociclib andribociclib, and the increased activity was due to CRBN mediated target degradation.Our CDK4/6 bifunctional degraders display excellent pharmacokinetic properties inmice with half-lives between 2-10 hours, oral bioavailability between 50-96% and arecurrently being evaluated for tumor growth inhibition against standard of caremolecules in MDA-MB-231 and MCF7 xenograft models. Clinical CDK4/6i are beingevaluated as combinations in TNBC, but the enhanced potency of CDK4/6 degradersin the TNBC cell lines shows that this approach is more promising for this aggressivetype of breast cancer.

Reference: Scheidemann, Erin R, and Ayesha N Shajahan-Haq. “Resistance toCDK4/6 Inhibitors in Estrogen Receptor-Positive Breast Cancer.” International journalof molecular sciences vol. 22,22 12292. 14 Nov. 2021, doi:10.3390/ijms222212292

Citation Format: Hannah Majeski, Akinori Okano, Angela Pasis, Casey Carlson, Qiao Liu, Arvind Shakya, Shenlin Huang, Aparajita Hoskote Chourasia, Leah Fung. Discovery of CDK4/6 bifunctional degraders for ER+/HER2- breast cancer andtriple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1553.