A novel VEGFR2 mutation has oncogenic potential in BRAF wild type melanoma

Tyrosine kinase receptors (RTKs) play central roles in tumor progression and represent an important class of drug targets. Nevertheless, clinical use of RTK inhibitors shows limited response and rapid resistance, which may be due to the presence of cancer cells expressing mutant forms of RTKs. Melanoma is an aggressive cancer which partially responds to existing therapies. Recently identified driver mutations (BRAFV600E) have allowed the development of effective targeted inhibitors. However, 30% of melanoma tumors lack activating mutations in BRAF or other drivers. VEGFR2 is a pro-angiogenic RTK which is also expressed in melanoma cells where it is implicated in tumor progression. Nevertheless, the possibility that activating vegfr2 gene mutations drive BRAFWT melanoma progression and response to TK inhibitors remains unexplored. In this study we used a new computational tool for non-synonymous low frequency mutation analysis (LowMACA) to simultaneously study the pattern of low frequency mutations conserved in the TK domain of various RTKs (PFAM: 07714) in human cancer samples. Heterozygous mutation of position 255, located in the activation loop of the TK domain, was found in VEGFR2, FGFR4, FLT4, PDGFRA and TIE1 in lung, breast and skin cancer. Of note, mutation of position 255 of VEGFR2-KD showed to be mutually exclusive with BRAFV600E in melanoma patients. Functional characterization demonstrated that mutation of residue 255 of VEGFR2-KD affects receptor activation/deactivation mechanism, thus allowing for ligand independent receptor activation. VEGFR2255 has a reduced lateral membrane motility when compared with VEGFR2wt as it is expected for signaling receptors. In BRAFWT melanoma, when co-expressed with endogenous VEGFR2WT to mimic heterozygosity, mutated VEGFR2255 modulates multiple phospho-kinases involved in cell metabolism. In vitro and in vivo analysis revealed that mutated VEGFR2255 confers to melanoma cells a strong pro-oncogenic phenotype. These activities, may be explained, at least in part, by its capacity to modulate VEGFR2wt through heterodymerization as shown by FLIM-FRET experiments. In-dept studies could be useful to understand the pro-oncogenic advantage conferred by mutated VEGFR2 and to design ad-hoc TK therapy.