Abstract P5-03-07: Molecular characterization of CDK 4/6 inhibitors (palbociclib, abemaciclib, and ribociclib) in hormone receptor-positive (HR+) breast cancer cell lines

Abstract

Abstract Background: Palbociclib (Ab) in combination with letrozole or fulvestrant (ful) has been shown to significantly improve disease-free survival in patients with HR+ breast cancer, in first and second line metastatic settings. Although Ab and two other CDK4/6 inhbitors [Abemaciclib (Ab) and Ribociclib (Ri)] are evaluated in multiple clinical trials, predictive biomarkers for these inhibitors have not been identified. To elucidate their molecular mechanisms in endocrine-responsive and -resistant cancers and identify potential predictive biomarkers, a series of preclinical experiments were conducted. Methods: All three inhibitors, each as a single agent and combined with AI or ful, were used to treat cell lines: MCF-7aro (ER+/aromatase+/ endocrine responsive), Let-R (letrozole-resistant), LTEDaro (long-term estrogen deprived line of MCF-7aro), and Rad-R (Everolimus-resistant). Molecular pathways were investigated through reverse phase protein array analysis (RPPA). Western blot analyses were performed to validate key results from the RPPA. Results and Discussion: Ab, Pa, and Ri inhibit proliferation of MCF-7aro with IC50 values: 24nM, 77nM, and 234nM, respectively. These inhibitors and AIs inhibit androgen (converted to estrogen by the expressed aromatase)-dependent proliferation of MCF-7aro in a synergistic manner. Our protein expression analyses confirm why ER+ breast cancer is most sensitive to treatment with CDK4/6 inhibitors. In MCF-7aro, 17β-estradiol up-regulates expression of RB, cyclin D, and FoxM1. The expression of these proteins can be suppressed by ful/AI + CDK4/6 inhibitors in a synergistic manner. Moreover, ful has been found to induce the expression of p21, an inhibitor of cyclin D, possibly by the elimination of ER-antagonized p53 activation. Since levels of pRB (S807/S811) are most sensitive to treatment with CDK4/6 inhibitors + ful, we hypothesize that pRB could be a predictive biomarker for this therapy. Ab, Pa, and Ri inhibit proliferation of Let-R (IC50 values: 170nM, 530nM, and 5730nM, respectively). Notably, our analysis has revealed that ful + CDK4/6 inhibitors effectively reduce the expression of Aurora-A, Plk1, and CHK1. Since these proteins play important roles in endocrine resistance, our molecular studies may explain why these drugs improve disease-free survival in cancer progression after one endocrine therapy. Also, Rad-R (IC50 values: 57nM, 136nM, and 1140nM, respectively) results suggest the potential utility of CDK4/6 inhibitors against Everolimus resistance. Unexpectedly, LTEDaro was found to be significantly more sensitive to CDK4/6 inhibitors as a single agent. Ab, Pa, and Ri inhibit proliferation of LTEDaro with IC50 values: 6.3nM, 10.9nM, and 63nM, respectively. If LTEDaro is considered to be a model of late stage endocrine resistance, our results suggest CDK4/6 inhibitors could be valuable for heavily treated patients. Conclusion: Preclinical studies using our endocrine-responsive and –resistant cell lines on three CDK4/6 inhibitors have produced valuable information about molecular characteristics associated with their treatments. These results will be verified in tumor biopsies of patients receiving CDK 4/6 inhibitors. Citation Format: Petrossian K, Lo C, Yuan Y, Kanaya N, Wu SV, Synold T, Huang C-S, Chen S. Molecular characterization of CDK 4/6 inhibitors (palbociclib, abemaciclib, and ribociclib) in hormone receptor-positive (HR+) breast cancer cell lines. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P5-03-07.