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Dana Faingold, Silvin Bakalian, Vasco Bravo-Filho, Henry A Wood, Maria Eugenia Orellana, Emilia Antecka, Miguel N Burnier; The effect of 17-AAG on focal adhesion kinase expression in uveal melanoma cell lines. Invest. Ophthalmol. Vis. Sci. 2014;55(13):5063.
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We have previously reported that focal adhesion kinase (FAK) was overexpressed in primary uveal melanoma (UM) and that phosphorylated FAK (pFAK) was detected in the most aggressive UM cell lines. The aim of this study is to compare the immunohistochemical profile of FAK and pFAK in UM to normal ocular tissues and to determine the effect of an Hsp90 inhibitor, 17-AAG, on the expression of FAK and pFAK in UM cells.
Immunohistochemical expression of FAK was evaluated in 39 UM specimens, pFAK expression was assessed in 51 UM specimens, and both FAK and pFAK expression were assessed in the uveal tract of 20 normal eyes. Immunohistochemical staining was assessed based on staining extent and intensity. For extent, specimens were scored “1” when less than 50% of the tumor cells stained, “2” when 50% to 80% of tumor cells stained, and “3” when more than 80% of tumor cells stained. Intensity was scored as follows: weak staining, “1”; moderate staining, “2”; and intense staining, “3”. The cases were grouped into low FAK expression when combined intensity and extent scores were between 1 and 4; and high FAK expression when total score was 5 and 6. Five UM cell lines (92.1, OCM-1, MKT-BR, SP6.5, and UW-1) were incubated with 10 µmol/L 17-AAG for 24, 48, and 72 hours. The expression of FAK and pFAK following treatment was determined by Western blot.
FAK was expressed in 87.2% of UM specimens, and 69.2% (n=27) were classified as having high FAK expression. pFAK was expressed in 90% of UM specimens and 60.8% (n=31) of the cases had high expression. We observed low FAK expression in non-tumor structures, such as choroidal melanocytes and the ciliary body, adjacent to UM tumors. Counterpart structures in normal eyes, including choroidal and ciliary body melanocytes, displayed negative or low cytoplasmic FAK and pFAK immunostaining. FAK and pFAK protein levels in all five cell lines decreased following treatment with 10 µmol/L of 17-AAG at 24, 48 and 72 hours.
Higher expression of FAK and pFAK was observed in uveal melanomas compared to normal ocular melanocytes. Hsp90 inhibition downregulated FAK and pFAK expression in UM cell lines. Therefore, we hypothesize that there is a role of high expression of FAK in uveal melanoma pathogenesis. Future studies are needed to explore the use of Hsp90 inhibitors for modulating FAK in uveal melanoma.
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