Abstract A38: Members of the noncanonical WNT pathway confer resistance to the MEK 1/2 inhibitor AZD6244 in colorectal cancer cell lines

Abstract

Abstract Background: The ERK pathway is constitutively activated in several human cancers including CRC. The MAPK/ERK kinase, MEK, occupies a central role in the ERK pathway and therefore, MEK inhibitors constitute a promising class of novel targeted cancer therapies. AZD6244 (ARRY-142886) is a potent, selective MEK1/2 inhibitor with nanomolar potency against cell lines and activity in several in vivo models of CRC. Despite initial enthusiasm for this class of agents in tumors with a high incidence of RAS or BRAF mutations, clinical studies indicate that resistance to MEK inhibitors remains a problem, and therefore, improved patient selection strategies are needed. The goal of this study was to identify and target predictive markers of resistance to AZD6244 in order to develop a rational basis for patient selection and combination therapy in CRC. Methods and Results: In order to identify predictive markers of responsiveness to AZD6244, 27 CRC cell lines were exposed to the agent, of which five highly sensitive (S) or resistant (R) cell lines were selected with IC50 values of <0.1 and >1.0 µM, respectively, and subjected to Affymetrix U133 plus 2.0 gene array analysis. ANOVA comparison of gene expression profiles between S and R cell lines revealed over 100 differentially expressed genes with a global p-value of <0.04, whereas 64 transcripts met the p<0.001 criteria. Of note, two genes, frizzled homolog 2 (FZD2) and wingless integrated 5B (WNT5B) were overrepresented in AZD6244 R cell lines by 17 and 7.5-fold, versus the S cells, respectively. These two genes comprise a receptor-ligand pair that has been shown to signal via the non-canonical, β-catenin-independent WNT signaling pathway. To determine the functional significance of over-expression of the non-canonical WNT pathway in resistance to AZD6244, we silenced FZD2 or WNT5B expression with stably transfected shRNAs and assessed the responsiveness of SW480 R cells to AZD6244. We observed a greater than 50% reduction in the IC50 to AZD6244 in the FZD2 knock-down cells and greater than 65% reduction in the WNT5B knock-down compared to wild-type and scrambled shRNA controls. Finally, we tested the effects of combining cyclosporine A (CsA) and tacrolimus, inhibitors of the non-canonical WNT pathway, with AZD6244. We observed synergistic inhibition of proliferation of SW480 cells treated with the combination of either inhibitor with AZD6244. Conclusions: Members of the non-canonical WNT pathway confer functional resistance to AZD6244 and thereby may represent not only predictive biomarkers of responsiveness to AZD6244, but also the basis for rational combination partners. These biomarkers and combination strategies are currently being validated in vivo with cell line and patient-derived CRC xenograft models. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A38.