Abstract A71: Basal- and treatment-induced activation of the PI3K-AKT pathway are critical determinants of the efficacy of the MEK inhibitor AZD6244 (ARRY-142886) in melanoma

Abstract

Abstract Over 80% of cutaneous melanomas have constitutive activation of the RAS-RAF-MEK-MAPK signaling pathway, which is a result of activating mutations in BRAF (>60%) or NRAS (∼20%). AZD6244 is a highly selective MEK1/2 inhibitor, which was well tolerated, induced marked reduction of tumor P-MAPK expression and resulted in ∼15% clinical responses in melanoma patients with BRAF mutations. The discordance between the prevalence of activated RAS-RAF-MEK-MAPK pathway and clinical efficacy of a potent pathway inhibitor suggests that other pathways may determine efficacy. Improved understanding of such pathways may lead to improved patient selection for AZD6244, and rational combinatorial approaches. We tested 17 molecularly characterized human melanoma cell lines for growth inhibition by AZD6244. Cell lines demonstrated a wide spectrum of growth inhibition; AZD6244 produced a dose- and time-dependent induction of cell death in sensitive cell lines, while resistant lines showed a G1 cell cycle arrest only. The mutational status of BRAF and NRAS did not predict sensitivity. We characterized the activation of signaling pathways in the cells by reverse phase protein array (RPPA) analysis for >100 proteins, including activation-specific markers of multiple critical signaling pathways. Baseline protein expression in the BRAF-mutant cell lines failed to show correlation between activation of the MAPK signaling pathway and sensitivity. Whilst BRAF-mutant cells with activation of the PI3K-AKT pathway tended to show increased resistance, we identified two resistant BRAF-mutant cell lines (SKMEL5, MEL624) with normal levels of p-AKT. Using RPPA, we characterized the time- and dose-dependent effects of AZD6244 on protein networks in these cells, and compared the results to two sensitive BRAF-mutant cell lines (WM35, A375). RPPA analysis showed potent and durable inhibition of MEK activity with AZD6244 treatment in all 4 cell lines. However, the treatment resulted in increased levels of p-AKT after 24–48 hours in the resistant cell lines; sensitive cell lines failed to activate AKT but instead showed an increase in PTEN expression (negative regulator of AKT activation). Parallel transcriptional profiling revealed that AZD6244 treatment resulted in increased PTEN mRNA in the sensitive WM35 cells, but not the resistant SKMEL5. Inhibition of AKT or IGF1R by siRNA resulted in synergistic cell death induction with AZD6244 in the resistant cell lines, but not in the sensitive cell lines. These studies support the critical nature of cross-talk between signaling pathways to the efficacy AZD6244, demonstrate a novel induction of PTEN expression in sensitive melanoma cell lines following MEK inhibition, and demonstrate synergy for combined inhibition of AKT and IGF1R with AZD6244 in cells that exhibit treatment-induced activation of the PI3K pathway. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A71.