MiR-410-3p suppresses primary gastric cancer and exosomes regulate endogenous expression of miR-410-3p.

MicroRNAs play significant roles in cancer initiation and progression. Exosomes are important extracellular vesicles for transporting molecules to distant sites. This study aims to investigate the functional roles of miR-410-3p in primary gastric cancer, as well as the roles of exosomes in regulating expression of miR-410-3p. In this study, forty-seven pairs of human gastric cancer tissue samples were collected. Endogenous expression of miR-410-3p in tissue samples and cell lines, and expression of exosomal miR-410-3p in cell culture medium were evaluated by RT-qPCR. Functional assays including cell proliferation assay by MTT, cell migration and invasion assay by transwell, and cell adhesion assay were performed. Targets of miR-410-3p were screened. Cell culture medium of culturing cell lines established from stomach (AGS and BCG23) was applied for culturing cell lines established from other sites (MKN45 and HEK293T). It was found that miR-410-3p was significantly downregulated in gastric cancer. Overexpression of miR-410-3p inhibited gastric cancer cell proliferation, migration, and invasion. MiR-410-3p mimic enhanced cell adhesion. HMGB1 was a target of miR-410-3p in primary gastric cancer. Expression of exosomal miR-410-3p in cell culture medium was dramatically higher than its endogenous expression. Exosomes from cell culture medium of AGS or BCG23 regulated endogenous expression of miR-410-3p in MKN45. In conclusion, miR-410-3p functioned as a tumor suppressor in primary gastric cancer. MiR-410-3p was higher expressed in exosomes of cell culture medium than its endogenous expression in cells. Endogenous expression of miR-410-3p in a distant site could be regulated by exosomes from the original site.

Inhibition of EP2 receptor suppresses tumor growth and chemoresistance of gastric cancer.

Gastric cancer is one of the leading causes of cancer death in the world. Early diagnosis and effective chemotherapy are vital to reduce the overall mortality. Prostaglandin E2 (PGE2) has been implicated as an important factor in gastric cancer carcinogenesis. ECF based regimen (epirubicin, cisplatin, 5-fluorouracil) is the first-line chemotherapy for advanced gastric cancer. However, patients develop resistance after chemotherapy. The aim of this study is sought to investigate the role of EP2 receptor, a PGE2 receptor, and the antagonism of EP2 receptor in response to ECF treatment. Expression of EP2 receptor was evaluated in gastric cancer tissue samples and cell lines. Cell proliferation and cell apoptosis assays were performed in vitro and in vivo, upon knockdown of EP2 receptor, antagonist of EP2 receptor and/or ECF treatment. Western Blot was applied for evaluation of proteins relating to cell cycle, apoptosis and drug transporter. Next generation sequencing and ingenuity pathway analysis were applied for screening for downstream targets of EP2 receptor. Expressions of the targets of EP2 receptor were further evaluated in gastric cancer cells and tissues. In this study, we found that expression of EP2 receptor was significantly upregulated in gastric cancer. Inhibition of EP2 receptor reduced gastric cancer cell proliferation, induced cell cycle arrest proteins, and enhanced cell apoptosis. Moreover, knockdown of EP2 receptor by siRNA or antagonist sensitized gastric cancer cells to ECF. Silence of EP2 receptor also significantly abrogated gastric cancer growth in a mice model. Analysis revealed that CAV1 was a downstream target of EP2 receptor in gastric cancer. Our findings illustrated that blocking EP2 receptor reduced tumor growth and induced apoptosis in gastric cancer. This novel study unraveled CAV1 was a downstream target of EP2 receptor. Antagonizing EP2 receptor could be a potential therapeutic target in gastric cancer, in particular those with high EP2 receptor expression.

Methylation-associated silencing of miR-193a-3p promotes ovarian cancer aggressiveness by targeting GRB7 and MAPK/ERK pathways.

Human growth factor receptor-bound protein-7 (GRB7) is a pivotal mediator involved in receptor tyrosine kinase signaling and governing diverse cellular processes. Aberrant upregulation of GRB7 is frequently associated with the progression of human cancers. However, the molecular mechanisms leading to the upregulation of GRB7 remain largely unknown. Here, we propose that the epigenetic modification of GRB7 at the post-transcriptional level may be a crucial factor leading to GRB7 upregulation in ovarian cancers. Methods: The upstream miRNA regulators were predicted by in silico analysis. Expression of GRB7 was examined by qPCR, immunoblotting and immunohistochemical analyses, while miR-193a-3p levels were evaluated by qPCR and in situ hybridization in ovarian cancer cell lines and clinical tissue arrays. MS-PCR and pyrosequencing analyses were used to assess the methylation status of miR-193a-3p. Stable overexpression or gene knockdown and Tet-on inducible approaches, in combination with in vitro and in vivo tumorigenic assays, were employed to investigate the functions of GRB7 and miR-193a-3p in ovarian cancer cells. Results: Both miR-193a-3p and its isoform, miR-193b-3p, directly targeted the 3' UTR of GRB7. However, only miR-193a-3p showed a significantly inverse correlation with GRB7-upregulated ovarian cancers. Epigenetic studies revealed that methylation-mediated silencing of miR-193a-3p led to a stepwise decrease in miR-193a-3p expression from low to high-grade ovarian cancers. Intriguingly, miR-193a-3p not only modulated GRB7 but also ERBB4, SOS2 and KRAS in the MAPK/ERK signaling pathway to enhance the oncogenic properties of ovarian cancer cells in vitro and in vivo. Conclusion: These findings suggest that epigenetic silencing of miR-193a-3p by DNA hypermethylation is a dynamic process in ovarian cancer progression, and miR-193a-3p may be explored as a promising miRNA replacement therapy in this disease.