Abstract
5248
B-Raf transversion T1799A has been reported in melanoma (30-60% of cases), papillary thyroid carcinoma (30-50%) and colorectal cancer (5-20%). Mutant B-RAF V600E shows higher kinase activity compared to wild type and determines hyperactivation of MAPK pathway. B-RAF silencing induces regression of melanoma xenografts, indicating an essential role of B-RAF for cell survival. We characterized in vitro a new small molecule compound, PLX4032, in different thyroid carcinoma cell lines, in particular NPA and ARO carrying B-RAF mutation and TPC1, carrying RET/PTC1. A375 melanoma cell line was used as a control. PLX4032 showed submicromolar IC50 in proliferation assay in B-RAF V600E positive cells, while it was inactive in TPC1 cells. Growth arrest was accompanied by little cell death in NPA cell line and G2/M decrease in ARO cells; by contrast, significant apoptosis was observed in melanoma cell lines. Re-expression of the thyroid differentiation marker, sodium iodide symporter (NIS), was observed after long-term inhibition. We set up an inducible siRNA system to verify the role of B-RAF in ARO thyroid carcinoma cells, as compared to melanoma. While B-RAF knock-down led to apoptosis in the melanoma cell line A375, anaplastic thyroid carcinoma cells ARO showed growth arrest upon silencing, with little or no cell death. This different behaviour was associated with the ability of cell lines to upregulate p21 expression levels in response to the lack of B-RAF mitogenic signal. We confirmed these results using PLX4032. Cell survival could be partially restored by overexpressing p21 using an adenoviral construct, in A375 cell lines that undergo apoptosis in the absence of B-RAF activity. These data suggest the activation of a B-RAF- and p53-independent mechanism of cell survival. However, in ARO cells, p21 silencing determined an increased G2/M as in untreated cells, indicating a relationship between B-RAF and p21 in G2/M control.
98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA
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