Melanoma and chemoresistance: in vitro and in silico models to exploit therapeutic targets

Abstract

Melanoma is a highly invasive and metastatic cancer, with high rates of mortality and chemoresistance. The MAPK pathway is constitutively activated, and there are already available powerful target-specific chemotherapeutics, such as BRAF inhibitors (vemurafenib and dabrafenib). However, resistance to BRAF inhibitors is observed after about 7 months of treatment, primarily because of the high degree of phenotypic, genetic and epigenetic intra-tumor heterogeneity. Although, combination therapies benefit patients, and immunotherapy has shown highly promising results, overcoming resistance remains major challenge. In silico screening studies from our laboratory using the TCGA and Gene Expression Omnibus (GEO) databases identified a collection of genes as being differentially expressed between invasive melanoma when compared with nevi (non-malignant lesion). Understanding the role and regulation of specifics genes should provide insights into the molecular mechanisms underpinning melanoma progression and ultimately resistance to therapy. In this study we will characterize the action of TOP1, ATP6V0B, ADK, PEMT and SINB3 genes in humans and vemurafenib-resistant Melanomas. This thematic project aim to complex process of the microenvironment, aging and metabolic process, and IDO and imune response. The objective is to identify target genes and mechanisms that contribute to vemurafenib resistance. To achieve this goal, the gene expression will be explored in a broad panel of human Melanomas at different stages of progression and mutation status, as well as in vemurafenib-sensitive and resistant cell lines. We will employ both monolayer culture and organotypic models that simulate invasion in reconstructed skin containing Melanoma, and explore histological samples from patients coming from a partnership with the Barretos Cancer Hospital. We will dissect the molecular pathways associated with these gene expression, as well as IDO, using gene manipulating tools (shRNA/CRISPR CAS9), and evaluate the impact of them on proliferation, migration, invasion, angiogenesis, and cell death induction. This study, like others already developed in our laboratory, can characterize genes that generate tumor subpopulations of resistant cells and aims at identifying pharmacologically tractable targets that may be used in combination to enhance therapeutic response. (AU)