Abstract PO1-18-05: Combination treatment with CDK2 inhibitor (BLU-222) and either palbociclib or ribociclib is synergistic in pre-clinical models of CDK4/6 inhibitor-resistant breast cancer
Authors: Luo Linjie , Navarro-Yepes Juliana , Wang Yan , Bui Tuyen , Kettner Nicole , Tripathy Debu , Hunt Kelly , Faia Kerrie , Keyomarsi Khandan 
Published: May 2024

Abstract
Background:
Cyclin-dependent-kinase-4/6 inhibitors (CDK4/6 inhibitors) plus endocrine therapy (ET) is the standard of care first-line treatment for patients with hormone receptor (HR)-positive, HER2-negative metastatic breast cancer (MBC). However, since most patients progress following long-term treatment, resistance to CDK4/6 inhibitors plus ET remains a clinical problem with limited therapeutic options. Here we show that a potent and selective CDK2 inhibitor, BLU-222, while effective as a single agent, exhibited enhanced, synergistic activity when combined with palbociclib or ribociclib in both in vitro and in vivo (patient-derived xenografts-PDX) models of HR+/HER2- breast cancer resistant to CDK4/6 inhibitors. Methods:
MCF7 and T47D palbociclib-resistant (PR) breast cancer cell lines were generated by culturing cells in media supplemented with increasing concentrations of palbociclib, starting at 1.2 µM and reaching a final concentration of 4.8 µM in a stepwise manner over a 6-month period. The single agent activity of BLU-222 and the combination effect of BLU-222 plus palbociclib in vitro were evaluated using the highest single agent model by SynergyFinder. The efficacy of BLU-222 as a single agent or in combination with palbociclib or ribociclib was also assessed in two PDX models (PR 1 and 2) from patients with acquired resistance (i.e. progressed following &gt;6 months of treatment) to palbociclib plus letrozole treatment, and another two PDX models (PR 3 and 4) from patients who were intrinsically resistant (i.e. progressed following &lt; 3 months of treatment) to palbociclib plus fulvestrant or letrozole, respectively. Results:
In palbociclib-resistant MCF7 and T47D cells, protein levels of cyclin E and CDK2 were significantly upregulated, but Rb and p21 levels were downregulated compared to the parental cells. While these PR cells were completely refractory to palbociclib, they were significantly more sensitive to BLU-222. Moreover, combining BLU-222 with palbociclib demonstrated a strong synergistic effect in PR cells accompanied by enhanced apoptosis and cell cycle accumulation in G1 or G2/M phases. Consistent with these in vitro findings, combination treatment with BLU-222 (60 mg/kg; b.i.d.) and palbociclib (50 mg/kg; q.d.) or ribociclib (50 mg/kg; q.d.) exhibited significant antitumor activity in all four PDX models, as compared to either treatment alone. The combination of BLU-222 plus the CDK4/6 inhibitor induced durable tumor regression and prolonged survival, which continued unabated following drug removal. Mechanistically, treatment of PR cells and palbociclib-resistant PDX models with BLU-222 as a single agent, resulted in the induction of p21, which can sensitize the cells to palbociclib. Conclusions:
These results show robust activity of BLU-222 in combination with CDK4/6 inhibitors in breast cancer cell lines and PDX models resistant to CDK4/6 inhibitors, providing a strong rationale for advancing BLU-222 plus CDK4/6 inhibitor treatment to clinical development in HR-positive/HER2-negative breast cancer patients who have progressed on CDK4/6 inhibitors.
Citation Format: Linjie Luo, Juliana Navarro-Yepes, Yan Wang, Tuyen Bui, Nicole Kettner, Debu Tripathy, Kelly Hunt, Kerrie Faia, Khandan Keyomarsi. Combination treatment with CDK2 inhibitor (BLU-222) and either palbociclib or ribociclib is synergistic in pre-clinical models of CDK4/6 inhibitor-resistant breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-18-05.