Modulation of genes and proteins related to metastatic processes in retinoic acid-sensitive and resistant breast cancer cells.

VANDERHOEVEN, F.; CASTRO C. ; REDONDO A.; SANCHEZ A.M; FLAMINI M.I.

San Luis

Congreso; XL Reunión Científica Anual. Sociedad de Biología de Cuyo; 2023

Sociedad de Biología de Cuyo

Breast cancer (BC) is women's most frequent malignant neoplasia and has a high mortality rate. All-trans retinoic acid (RA) is a vitamin A-derived pleiotropic signaling molecule that regulates critical genetic programs. Although RA has demonstrated a potent anticarcinogenic activity, its use in solid tumors is limited. Previous studies showed that MDA-MB-231, MDA-MB-468, T-47D andSK-BR3, BC cell lines, have different RA sensitivity and RA-growth inhibitory effects. We have previously demonstrated that RA decreases cell migration in RA-sensitive BC cells by reducing the expression of movement-fundamental proteins. Furthermore, we determined how RA-pathway phosphorylates/activates and redistributes these proteins, ultimately inhibiting BC cell migration.Metalloproteases (MMPs) are proteins involved in the degradation of the extracellular matrix, promoting invasion and metastasis. MMPs are closely related to E-Cadherin and Vimentin, protein markers of Epithelial/Mesenchymal Transition (EMT), and their levels affect cell adhesion and migration. We propose to characterize target gene expression profiles in RA-treated BC cell lines using the Gene Expression Omnibus (GEO) public repository and to determine the effect of RA in RA-sensitive or resistant BC cells on cell viability and proliferation. We also propose to determine the role in the EMT process. We performed bioinformatics analysis using GEO (GSE103426) to determine differential gene expression in RA-resistant cell lines by volcano plots and in-vitro experiments to assess the effect of RA on cell viability by MTT assay and protein expression by Western Blot in RA-sensitive and RA-resistant cell lines. Differential gene expression analysis of controls vs. treated with RA (100 nM, 18 h) MDA-MB-231 and MDA-MB-468 cells showed that RA modulates migration-related gene expression similarly in both lines. Interestingly, genes related to proliferation were not affected. The viability of MDA-MB-231 cells was not reduced after different doses of RA, demonstrating their resistance to RA. In contrast, RA decreased the viability of T-47D cells, confirming their sensitivity to RA. We also observed that retinoic acid receptors (RARα, RARβ and RARγ) were expressed at different levels within BC cells. This fact coincides with RA-sensitivity reported for BC cells, where T-47D and SK-BR3 are considered RA-sensitive and MDA-MB-231 RA-resistant based on their high and low expression of RARs, respectively. Finally, RA treatment modulated EMT- related proteins, E-Cadherin, Vimentin, and metalloproteases MMP2 and MMP9 in both cell lines. In conclusion, despite the absence of RAR receptors, RA affects and modulates mRNA and protein levels in RA-resistant cells and affects the expression of proteins involved in the migratory process in sensitive and resistant cells. These results could be significant as a potential therapy for metastatic breast cancers without specific treatment.