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posted on 2025-04-03, 07:21 authored by Adam P. Dommer, Vishnu Kumarasamy, Jianxin Wang, Thomas N. O’Connor, Michelle Roti, Sidney Mahan, Karen McLean, Erik S. Knudsen, Agnieszka K. Witkiewicz Figure S6. P16INK4A is a key determinant of response to BLU-222. A, Heatmap analysis from the cancer dependency map (DepMap) database displaying relative expression of CCNE1 and several endogenous CDK inhibitor genes in a panel of established ovarian and breast cancer cell lines. B, Western blot of RB1-E2F pathway activity in MDA-MB-157 cells following overexpression (OE) of CDK4 with and without BLU-222 treatment. C, Live cell proliferative monitoring of CDK4 WT (left) and overexpressing (OE, right) of MDA-MB-157 cells treated with serial concentrations of BLU-222 [n = 3–4 per group; two-way ANOVA, significance (shown as #) comparing CDK4 OE versus CDK4 WT at 62.5 nM BLU-222 dose; error bars represent SEM]. D, Western blot of RB1-E2F pathway activity in MDA-MB-157 cells following overexpression (OE) of CDK6 with and without BLU-222 treatment. E, Live cell proliferative monitoring of CDK6 WT (left) and overexpressing (OE, right) MDA-MB-157 cells treated with serial concentrations of BLU-222 [n = 3–4 per group; two-way ANOVA, significance (shown as #) comparing CDK6 OE versus CDK6 WT at 250 nM BLU-222 dose; error bars represent SEM]. F, Table summarizing effect of CDK4 or CDK6 overexpression (OE) or CDKN2A knockdown on IC50 to BLU-222 in Kuramochi and MDA-MB-157 cells. G, Hematoxylin and Eosin (H&E) staining and of tumors excised from MDA-MB-157 xenografts in NOD scid gamma (NSG) mice treated with either vehicle or BLU-222; arrows indicate cells with enlarged morphology (scale bar = 100 µm). #, P < 0.05.
Funding
National Cancer Institute (NCI)
United States Department of Health and Human Services
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ARTICLE ABSTRACT
Cyclin-dependent kinase 2 (CDK2) inhibitors have recently been developed and have entered clinical trials. Combination approaches can help broaden the use of therapeutic agents and establish more effective treatments. Here, we evaluated the selective CDK2 inhibitor BLU-222 for mechanisms of response in the context of ovarian and breast cancer models. Sensors of cellular CDK activity indicated that sensitivity to either CDK4/6 or CDK2 inhibition was related to the differential dependence on a single CDK for G1–S transition. Unlike CDK4/6 inhibitors, BLU-222 was able to robustly inhibit proliferation through cell-cycle inhibition in both G1 and G2 phases. However, it remained possible for cells to reenter the cell cycle upon drug withdrawal. The antiproliferative strength and impact on G1–S versus G2–M accumulation was mediated by the RB tumor suppressor. To broaden the sensitivity to CDK2 inhibition, combinatorial drug screens were performed that identified both synergistic (e.g., CDK4/6 inhibitors) and antagonistic (e.g., WEE1 inhibitors) relationships. Models that were exceptionally sensitive to CDK2 inhibition displayed coordinate expression of cyclin E1 and P16INK4A, an endogenous CDK4/6 inhibitor. Functional studies demonstrated that P16INK4A and CDK4/6 activity were key mediators of sensitivity to BLU-222. Clinical gene and protein expression analyses revealed a positive correlation between cyclin E1 and P16INK4A and identified that ∼25% of ovarian cancers exhibited coordinate expression of cyclin E, P16INK4A, and RB, indicative of strong sensitivity to CDK2 inhibition. Together, this work advances a precision strategy for the use of CDK2 inhibitors in the context of ovarian and breast cancers.Significance: The CDK2-specific inhibitor BLU-222 shows preclinical efficacy in breast and ovarian cancer with select determinants of response and holds promise in combinatorial strategies.See related article by House and colleagues, p. 1297
