Dados do Resumo

Título

GENES OF NUCLEOTIDE SYNTHESIS PATHWAY TYMS AND MTHFD1 AS TARGETS OF CLINIC INTEREST IN GASTRIC CANCER THERAPY

Introdução

Gastric cancer (GC) is among the most common and lethal neoplasms in the world. The nucleotide biosynthetic pathway has great importance for maintaining tumor replicative potential. Therefore, drugs targeting the modulation and inhibition of these pathways are important to the clinic. The antihelmintic Mebendazole (MBZ) has demonstrated antitumor activity related to metabolic modulation, however, its mechanism in nucleotide metabolism pathways remains poorly elucidated.

Objetivo

Therefore, this study aims to analyze the potential modulator of genes in the purine and pyrimidine pathways by MBZ, as well as to evaluate whether this modulation is associated with a decrease in the proliferative profile of the AGP-01 lineage after exposure to MBZ.

Métodos

The mRNA expression and its relation with the overall survival (OS) rate of two nucleotide synthesis pathway genes (TYMS and MTHFD1) was evaluated by GEPIA and Kaplan Meyr plotter GEPIA (http://gepia. cancer-pku.cn/) and Kaplan Meier plotter (https://kmplot.com/ analysis/) softwares, respectively. Alamar Blue assay was assessed to determine the IC50 of MBZ and 5-FU in metastatic GC and non-tumoral cell lines, AGP-01 and MNP-01. Next, RT-qPCR was used to compare the expression levels of TYMS and MTHFD1 in AGP-01 and MNP-01, as well as, to analyze if the treatment with MBZ led to a reduction in the transcripts levels of these genes. Molecular docking was performed to analyze the interaction of MBZ with TYMS and MTHFD1 proteins. Finally, the clonogenic assay and cell cycle analyses were conducted to visualize the potential of MBZ in reducing the proliferative profile and leading to cell cycle arrest of the AGP-01 cell line.

Resultados

Results showed TYMS and MTHFD1 were shown to be overexpressed in patients with GC, as well as when highly expressed are related to reduced OS rate of patients (p<0.05). MBZ evidenced high cytotoxic and selective potential, with IC50 of approximately 0.15 µM and 1.37 µM for the AGP-01 and MNP-01 cell lines, respectively. Both genes (TYMS and MTHFD1) were also upregulated in the AGP-01 cell line and MBZ was shown to reduce their expression after 24 hours of treatment (p<0.01). Interestingly, MBZ also seems to highly interact with both proteins, leading to conformational modification. The reduction in mRNA synthesis and the structural modification caused by MBZ in TYMS and MTHFD1 enzymes were linked to the antiproliferative potential of MBZ, once our results evidenced that MBZ treatment led to a reduction in cell proliferation and cell cycle arrest in G0/G1 after 48 hours of treatment (p<0.001).

Conclusões

The study suggests that MBZ modulates the nucleotide synthesis pathway by reducing TYMS and MTHFD1 gene expression and interacting with their proteins, potentially linked to its antiproliferative effects on AGP-01 GC cells. Further studies are needed to quantify the biosynthetic intermediates post-MBZ treatment. This work aids in exploring and validating new pharmacological targets for GC treatment.

Financiador do resumo

Cearense Foundation for Scientific and Technological Development Support (FUNCAP), Coordination for the Improvement of Higher Education Personnel (CAPES), and National Council for Scientific and Technological Development (CNPq).

Palavras Chave

Metabolic reprogramming; Pharmacological target; Tumor malignancy