Hypoxia-induced autophagy increased resistance to Vemurafenib treatment on sensitive, but not resistant melanoma cells

PEREZ CELIA; FALCÓN CRISTIAN ROBERTO; ALVAREZ SERGIO; CUELLO ORLANDI SERGIO FEDERICO

Mar del Plata

Congreso; LXIV Reunion anual de la Sociedad Argentina de Investigacion Clinica; 2019

SAIC

Melanoma is the most aggressive type of skin cancer. Although recent therapies have shown an impressive success, unfortunately patients develop resistance after a short period of disease control. Autophagy has been indicated as a mechanism of resistance in tumor cells. Autophagy is a process regulated by numerous factors, including hypoxia, a distinctive feature of the tumor microenvironment. Our hypothesis is that hypoxia or prolonged treatment with Vemurafenib (Vf, BRAF inhibitor) augment autophagy that in turn further increases Vf resistance. To this end, we cultured Lu1205 human melanoma cells with or without CoCl2 during 24h to induce hypoxic condition that was confirmed by HIF-1α expression. Certainly, CoCl2 treatment increased LC3BII level, a marker of autophagy, but it did not affect Lu1205 cell viability. To investigate the role of hypoxia in Vf resistance, Lu1205 cells were cultured in presence or absence of CoCl2 during 24h and then treated with Vf for another 24h. As assessed by MTT assay, cells that were pre-treated with CoCl2 were more resistant to Vf (5-10µM). Interestingly, the inhibition of autophagy with chloroquine or NH4Cl diminished this resistance. Hypoxia did not modify the ability of Vf to reduce pERK1/2 but decreased the pAKT levels induced by the inhibitor. Moreover, Lu1205 cells with acquired resistance to Vf generated in our lab showed increased survival and higher levels of LC3BII and pERK1/2 than sensitive cells when were treated with Vf. On the other hand, treatment of Vf resistant cells with CoCl2 did not induce significant changes in LC3BII, pERK1/2 and pAKT levels. Altogether, these results suggest that hypoxia promotes a pre-resistant cell state with increased autophagy that enhances the develop of Vf resistance. Moreover, this mechanism is exacerbated in cells with acquired Vf resistance by long period treatment.