Evaluation of the roles of caveolin-1 protein and focal adhesion complex proteins in nitric oxide-mediated resistance to death by anoikis in human melanoma cells

Abstract

Cancer is one of the major global public health problems, being the second leading cause of death worldwide. It is defined as a set of diseases that in general are characterized by mutations in genes that have the function of regulating cell proliferation, differentiation and death. In Brazil, more than 625 thousand new cases are estimated for each year of the 2020-2022 period, which are mainly related to skin, prostate, lung, colon, stomach and breast cancers. Still in Brazil, melanoma stands out for its not so high incidence, corresponding to an estimated risk of 4.03 new cases for every 100 thousand men and 3.94 for every 100 thousand women, but with high mortality rates. Its 5-year survival rate is less than 5% for patients with metastatic melanomas for lymph nodes. A successful metastasis process is associated with the ability to resist death due to loss of adhesion to the substrate (anoikis), which melanomas and other tumors acquire during their development. The tumor microenvironment can be described as a non-healing wound, directly associated with chronic inflammation regulated by specialized cells, inflammatory cytokines and chemically reactive species such as reactive oxygen species (ROS) and nitric oxide (NO) that participate directly in the cellular signaling processes that in turn will act on the epithelial-mesenchymal - EMT and mesenchymal-epithelial - MET transitions thus regulating tumor progression. The cells of breast cancer, colon and melanomas, developed a mechanism to survive and adapt to oxidative and nitrosative stress, this mechanism involves an adjustment of the intracellular concentrations of NO and ROS, thus guaranteeing tumor progression. Recently, our group showed the positive effects of increasing NO concentrations on resistance to death by anoikis, demonstrating activation of Src kinase protein in relatively high concentrations of NO in HeLa cells (cervical cancer) and in murine melanoma cells, kept in suspension. Under these conditions, induction of Caveolina-1 (Cav-1) expression was also observed. Cav-1 is a membrane-associated protein with a molecular weight of 22 kDa located in the caveoles. In cells maintained adhered to a surface, Cav-1 interacts with activated Src kinase and plays an important role in the organization dynamics of the focal adhesion complex, which in turn is important for cell survival. Thus, in developing this project, we intend to evaluate the roles of Cav-1 protein and proteins of the focal adhesion complex, Src, FAK, p130Cas and PTPa and paxillin, in NO-mediated resistance to death by anoikis in cell lines human melanoma from two stages of development and in two experimental conditions: (1) Cells of the A375 strain obtained from malignant human melanoma of the primary site maintained in suspension and subsequent re-injury; (2) SK-MEL-28 lineage cells obtained from human melanoma with metastasis to lymph nodes maintained in suspension and subsequent re-injury. The results achieved in this study may help to describe a cell signaling pathway mediated by NO and associated with resistance to anoikis in melanoma. (AU)