MIF como possível alvo terapêutico do câncer gástrico

Abstract

Gastric cancer (GC) is the fifth most common and lethal neoplasm worldwide, characterized by its heterogeneity and multifactorial nature. In most cases, diagnosis occurs at advanced stages, making malignancy treatment challenging. The therapeutic approach is defined based on the tumor’s preoperative staging, with chemotherapy combined with surgical resection being the primary treatment choice, though it is restrictive to cases where the tumor is localized. Despite its widespread use, conventional chemotherapy faces significant limitations, such as severe adverse effects and the occurrence of chemoresistance, which restricts its efficacy. In this context, targeted therapy has gained prominence in recent years, aiming to overcome chemoresistance and minimize side effects, offering more specific and effective treatments. A promising strategy for identifying new biomarkers and therapeutic targets is target fishing, which involves exploring new targets from previously developed drugs, enabling the discovery of unexplored targets with relevant clinical potential. Thus, the main objective of this study was to identify a new molecular target as a therapeutic alternative for GC treatment. To achieve this, a panel of 28 molecular inhibitors was tested in two GC cell lines, selecting the molecular inhibitor with the best cytotoxic action in both models. The selected inhibitor was BTZO-1, a selective inhibitor of macrophage migration inhibitory factor (MIF). Subsequently, we evaluated MIF’s differential gene expression using the TCGA transcriptomic dataset for GC, MIF gene expression in gastric tumor cell lines, MIF inhibition after treatment with BTZO-1, and its effects on membrane integrity, cell migration, cell cycle alterations, caspase-3/7 activation, and protein expression. Bioinformatics analyses of MIF in GC showed that MIF is overexpressed in GC, particularly in the intestinal subtype. Furthermore, increased MIF expression correlated with patient survival, tumor size, absence of metastasis in regional lymph nodes, and the presence of distant metastases. Gene expression analysis revealed that the MIF gene is upregulated in all three evaluated gastric neoplastic cell lines (AGP-01, ACP-02, and ACP-03). The AGP-01 cell line, derived from a metastasis of the intestinal subtype, showed the highest MIF expression and was the most sensitive to the BTZO-1 inhibitor (IC50 = 1.1 μM). In contrast, the ACP-03 cell line, originating from a primary tumor of the same subtype, exhibited the lowest MIF expression and was three times more resistant to the inhibitor (IC50 = 3.1 μM). MIF inhibition in the AGP-01 cell line resulted in high levels of membrane integrity impairment, reduced cell migration after 6 hours of treatment, accumulation of cells in the G2/M phase of the cell cycle, and a significant impact on essential biological processes, as demonstrated by proteomic analysis. The main biological processes affected included mRNA metabolic regulation, RNA splicing and its regulation, and protein folding. The functional evaluation of differentially expressed proteins revealed that many act as structural chromatin constituents and chaperones involved in protein folding. Additionally, the BTZO-1 inhibitor proved to be a promising candidate as a therapeutic strategy for this neoplasm, further reinforcing the need for additional studies to validate its efficacy. The findings of this study indicate that MIF is a potential therapeutic target for GC, given its clinical significance and role in tumor progression. Its inhibition affected different biological processes essential for tumor development.