Investigating PLX4032 resistance in a human melanoma cell line: a question of metabolism

To date, the standard approach to metastatic melanoma with classic chemoterapeutic agents has been unsuccessful. In the last 15 years, he identification of mutations in BRAF genes, such as BRAFV600E, led to the development of BRAF mutant target therapies with small biological inhibitors (i.e. Vemurafenib, PLX4032) that specifically block the constitutively active MAPK-mediated pathway. Although patients respond positively at first, after six-months therapy most patients relapse due to acquired resistance to inhibitors. The heterogeneity of this tumor makes the study of melanoma acquired resistance more complex, pointed out the importance of directing research towards personalized medicine. Moreover, cancer cells undergo a multifaceted rewiring of cellular metabolism to support their biosynthetic needs, and the implications and clinical relevance in melanoma acquired resistance to PLX4032 has not been elucidated. For these purposes, in this thesis a human mutant BRAFV600E PLX4032 resistant subline was selected. It is important to note that this selection derives directly from tumor biopsies prior to therapy, and the treatment has been protracted for six months, in order to obtain a truthful model of resistance. In the first part of the work the acquisition of resistance, together with tumorigenicity, clonogenicity and invasiveness were assessed. It was also found that P-ERK and MITF, two known mediators of resistance in melanoma, are not overexpressed in melanoma resistant cell line and demonstrated to not mediate vemurafenib resistance in our resistant cell lines. The second part of the work was focused on the identification of metabolic changes, that could be involved in the acquisition of resistance to BRAF inhibitors. This thesis highlights some potential key metabolic and phenotypic markers mediating resistance to targeted therapies in cutaneous malignant melanoma, emphasizing the importance to study metabolic changes that could be used as adjuvant target to overcome or bypass acquired resistance.