BRAF inhibitor resistance is mediated by N‐Ras upregulation in melanoma (796.3)

Abstract

The clinical benefit of BRAF inhibitors therapies is limited by the emergence of drug resistance. Here, we investigated the molecular basis underlying acquired resistance to BRAF inhibitor by comparing the signaling pathway in the parental A375P cells and resistant sub‐line (A375P/Mdr). We demonstrate that MAPK re‐activation is not attributed to the mechanism of resistance to UAI‐201 in A375P/Mdr cells. Relative quantitative analysis with the 2‐ΔΔCt method revealed that BRAF inhibitor resistance observed in A375P/Mdr cells was not mediated through the overexpression of MDR proteins. We found that expression of N‐Ras was up‐regulated in BRAF inhibitor‐resistant A375P/Mdr cells compared to A375P cells. Actually, siRNA‐mediated N‐Ras knockdown partially conferred UAI‐201 sensitivity to A375P/Mdr cells, implying that acquired resistance to UAI‐201 developed by N‐Ras up‐regulation. Notably, the flow cytometric analysis revealed that UAI‐201 caused a significant accumulation of cells in G0/G1 phase in N‐Ras knock‐downed A375P/Mdr cells. However, PDGFRβ knockdown failed to sensitize A375P/Mdr cells to growth suppression by UAI‐201, although the remarkable increase in PDGFRβ was observed in A375P cells. Thus, the discovery of newly acquired genetic alterations that mediate resistance following exposure to BRAF targeted therapy would logically lead to sequential treatment strategies.