hsa-miR-34a-5p enhances temozolomide anti-tumoral effects on glioblastoma: in-silico and in-vitro study.

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This study investigated the effect of hsa-miR-34a-5p and its combination with temozolomide on GBM, the related molecular mechanisms, and the signaling pathway using in-silico and in-vitro approaches. The in-silico tumor bulk and single-cell RNA sequencing analyses were done on TCGA-GTEx, CGGA, GSE13276, GSE90603, and GSE182109 datasets. After selecting the A172 cell line, hsa-miR-34a-5p mimics were transfected, and the cell viability, migration, cell cycle, clonogenicity, and apoptosis of studied groups were studied using MTT, scratch, flow cytometry, colony formation, and Annexin V/PI assays. The mRNA expression of CASP9, CASP3, CASP8, MMP2, CD44, CDK6, CDK4, CCND1, RAF1, MAP2K1, MET, SRC, and CD274 was studied using qRT-PCR method. hsa-miR-34a-5p downregulated RAF1 expression, as the signaling factor of the MAPK pathway. The combined treatment significantly downregulated the expression of MET, SRC, and MAP2K1, leading to the inhibition of the MET/MAPK pathway compared to temozolomide. Besides exerting anti-tumoral effects on the cell viability, migration, cell cycle, apoptosis, and clonogenicity of A172 cells, its combination with temozolomide enhanced temozolomide anti-tumoral effect. Compared to temozolomide, the combined treatment significantly decreased CDK4, CDK6, CCND1, and MMP2 expression. hsa-miR-34a-5p targets RAF1, as the signaling factor of the MAPK pathway, and potentiates the temozolomide anti-tumoral effect on A172 cells.", "claims": [{"id": "claim-method", "statement": "To conduct statistical comparisons between more than two groups, the one-way ANOVA was used.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:38655096", "locator": "sections[17] \"Statistical analyses\" paragraph 2", "quote": "To conduct statistical comparisons between more than two groups, the one-way ANOVA was used."}], "state": "complete"}, {"id": "claim-data-access", "statement": "The correlational study between hsa-miR-34a-5p and RAF1 was done based on the TCGA-GBM dataset from cBioPortal, and the prognostic studies were conducted based on the CGGA dataset (Cerami et al., 2012[8]; Zhao et al., 2021[44]).", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:38655096", "locator": "sections[5] \"Bioinformatic tools\" paragraph 2", "quote": "The correlational study between hsa-miR-34a-5p and RAF1 was done based on the TCGA-GBM dataset from cBioPortal, and the prognostic studies were conducted based on the CGGA dataset (Cerami et al., 2012[8]; Zhao et al., 2021[44])."}], "state": "complete"}, {"id": "claim-target-result", "statement": "hsa-miR-34a-5p significantly arrests the A172 cell cycle at the sub-G1 phase and potentiates temozolomide-mediated sub-G1 arrest.", "kind": "quantitative", "target": {"value": 0.0001, "unit": "p-value", "tolerance": null, "source": "It has been found that hsa-miR-34a-5p significantly arrests the cell cycle at the sub-G1 phase in A172 cells (P-value < 0.0001) (Figure 10A, B(Fig. 10))."}, "evidence": [{"asset": "source:pmid:38655096", "locator": "sections[28] \"hsa-miR-34a-5p ectopic expression increases the anti-tumoral effect of temozolomide on the A172 cell cycle\" paragraph 2", "quote": "It has been found that hsa-miR-34a-5p significantly arrests the cell cycle at the sub-G1 phase in A172 cells (P-value < 0.0001) (Figure 10A, B(Fig. 10))."}, {"asset": "source:pmid:38655096", "locator": "sections[28] \"hsa-miR-34a-5p ectopic expression increases the anti-tumoral effect of temozolomide on the A172 cell cycle\" paragraph 2", "quote": "Also, hsa-miR-34a-5p significantly increases the temozolomide-mediated cell cycle arrests at the sub-G1 of A172 cells (P-value < 0.0001) (Figure 10A, B(Fig. 10))."}], "state": "complete"}], "gaps": []}, "experiment": {"state": "complete", "objective": "hsa-miR-34a-5p significantly arrests the A172 cell cycle at the sub-G1 phase and potentiates temozolomide-mediated sub-G1 arrest.", "steps": [{"id": "read_published_estimate", "order": 1, "name": "Read the published estimate from the source", "kind": "other", "state": "complete", "blocked_on": null, "description": "The claim, method and evidence span published with the source article.", "duration_hours": null, "parameters": {}, "inputs": ["source:pmid:38655096"], "outputs": ["the published estimate and its evidence"], "instruments": [], "reagents": []}, {"id": "obtain_source_data", "order": 2, "name": "Obtain the source data from the named repository", "kind": "data-preparation", "state": "complete", "blocked_on": null, "description": "The GEO series accession resolves and its sample accessions are listed. 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