Prognostic Role of BRAF V600E Cellular Localization in Melanoma

Abstract

Background: Approximately half of cutaneous melanoma tissues harbor BRAF V600E mutations, resulting in a constitutive activation of the mitogen-activated protein kinase (MAPK) pathway. Nuclear-cytoplasmic transport machinery is dysregulated in neoplastic cells and alters the key regulatory proteins that can lead to tumor progression and drug resistance. The significance of nuclear localization of BRAF V600E has not been fully understood. We examined the clinical significance of intracellular localization of BRAF V600E in cutaneous melanoma. Study Design: Immunohistochemical analysis of BRAF V600E was performed on formalin-fixed, paraffin-embedded specimens of cutaneous melanoma (n = 91). Staining intensity was graded in a blinded manner. Correlations to clinical factors were analyzed by Fisher's exact test and 2-tailed t-test. Localization of BRAF V600E was determined in melanoma cells, and we investigated their resistance to BRAF V600E -specific inhibitor according to nuclear localization in both in vitro and in vivo models. Results: We included 91 patients, of whom 32% (29 of 91) had cytoplasmic BRAF V600E . Nuclear BRAF V600E was observed in 30% (27 of 91). Overall, BRAF V600E expression correlated with TNM stage (p = 0.011), mitotic activity (p = 0.010), and ulceration (p = 0.045). Nuclear BRAF V600E expression correlated with overall clinical stage (p < 0.001), tumor size (p < 0.001), regional lymph node (p < 0.017), depth of invasion (p = 0.005), Clark level (p < 0.001), mitotic activity (p < 0.001), ulceration (p < 0.001), and margin status (p = 0.017). On a cellular level, BRAF V600E was identified in the nucleus, and its translocation was serum dependent. Our in vitro and in vivo data revealed sequestration of BRAF V600E in the cytosol-sensitized resistant cells to vemurafenib; nuclear retention of BRAF V600E was associated with aggressiveness and drug resistance. Conclusions: Nuclear localization of BRAF V600E is associated with melanoma aggressiveness. Further multi-institutional studies are warranted to confirm the clinical relevance of nuclear localization of BRAF V600E .