Abstract 2832: Synergistic effect of combined cdk4/6 inhibitor with docetaxel in lung cancer cell lines harboring KRAS mutations

Abstract

Abstract Background & Purpose LY2835219(LY), a novel CDK4/6 inhibitor, inhibits phosphorylation of RB and E2F activation, thereby arresting the cell cycle in the G1 phase and suppressing the cell proliferation and cell division. Docetaxel(DTX) is a cytotoxic anti-cancer drug, which inhibits mitotic cell division, thus induces G2/M arrest and apoptosis of various cancer cells. In present study, we evaluated the activities of CDK4/6 inhibitor alone or combined with docetaxel on the anti-proliferation, cell cycle and apoptosis in lung cancer cell lines harboring KRAS mutations. Methods We measured the anti-proliferative activities of LY or DTX single and their combinations (DTX+LY 72h and DTX 24h->LY 48h) on cell proliferation in A549(G12S) and H727(G12V) cells using CCK-8 asssy. We evaluated the expression of CDK2, CDK4, cPARP and caspase-3 by Western blot. The cell cycle distribution and apoptosis in subG1 phase were analyzed by flow cytometry. Results The IC50 values of the LY and DTX alone were 0.4 ± 0.2 uM and 0.9 ± 0.2 nM in A549 cells and 2.0 ± 0.7 uM and 3.1 ± 0.3 nM in H727 cells, respectively. The CI (Combination index) of the DTX->LY sequential and the DTX+LY simultaneous treatments were 0.8 and 0.9 (CI<1; synergism) in A549 cells, respectively. In H727 cells, DTX->LY showed decreased cell viability by 60% at very low concentration of DTX (3.0e−5 nM), but DTX+LY showed significantly decreased cell viability by 40% at higher concentration of DTX (IC50 values can not be measured). In A549 cells, after DTX treatment for 72h, the cell population in G2/M phase increased by 54.5% relative to control (24.6%). With LY alone, the fraction of A549 cells in G0/G1 phase increased compared to control (78.4% vs. 49.5%). In case of DTX+LY combination, the G2/M fraction of A549 cells significantly increased by 33% vs 24.6% in a dose-dependent manner. Meanwhile, in DTX->LY treatment, cell fractions in G2/M were increased (44.5% vs 24.6%), especially those in G0/G1 phases were remarkably reduced (14.7% vs 49.5%). In addition, the subG1 fraction was accumulated in response to individual drug treatments. In case of DTX single treatment, the subG1 fractions of both A549 and H727 cells increased in a dose-dependent manner (0.7% vs. 17.4% and 2.5% vs. 28.1%, respectively). We were not able to observe the change of subG1 fraction in DTX+LY combination treatment in A549 cells. However, in DTX->LY sequential treatment, the subG1 fraction increased in a dose-dependent manner by relative to control (23.3% vs 10.9%). The expression of c-PARP and caspase-3 was increased by the DTX alone and two different combination schedule and CDK2 and CDK4 expressions were decreased by LY alone and DTX->LY sequential treatment. Conclusions Taken together, our results suggest that DTX->LY sequential treatment has enhanced antitumor efficacy with synergistic interaction in lung cancer cell lines harboring KRAS mutations. Citation Format: Kyoung Hwa Son, Jung-Young Shin, Jeong-Oh Kim, JinHyoung Kang. Synergistic effect of combined cdk4/6 inhibitor with docetaxel in lung cancer cell lines harboring KRAS mutations. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2832.