Inhibition of RUNX1 promotes cisplatin-induced apoptosis in ovarian cancer cells.

Runt-related transcription factor 1 (RUNX1), one subunit of core-binding factors in hematopoiesis and leukemia, was highly expressed in ovarian cancer (OC), but the role of RUNX1 in OC is largely unknown. Since we found that high expression of RUNX1 is correlated with poor survival in patients with OC through bioinformatic analysis of TCGA database, we developed RUNX1-knockout clones by CRISPR/Cas9 technique and discovered that RUNX1 depletion could promote cisplatin-induced apoptosis in OC cells, which was further confirmed by RUNX1 knockdown and overexpression. We also proved that RUNX1 could elevate the expression of BCL2. We then examined a total of 32 candidate miRNAs that might mediate the regulation between RUNX1 and BCL2, of which three miRNAs from the miR-17~92 cluster were found to be negatively regulated by RUNX1. Consistently, our analysis of data from TCGA database revealed the negative correlation between RUNX1 and the cluster. We further confirmed that miR-17~92 cluster could enhance cisplatin-induced apoptosis by directly targeting BCL2 3'UTR. Since rescue experiments proved that RUNX1 could repress cisplatin-induced apoptosis by up-regulating BCL2 via miR-17~92 cluster, combining RUNX1 inhibitor Ro5-3335 and cisplatin showed synergic effect in triggering OC cell apoptosis. Collectively, these findings show for the first time that combinational treatment of cisplatin and RUNX1 inhibitor could be used to potentiate apoptosis of ovarian cancer cells, and reveal the potential of targeting RUNX1 in ovarian cancer chemotherapy.

miR-142-5p enhances cisplatin-induced apoptosis in ovarian cancer cells by targeting multiple anti-apoptotic genes.

Chemotherapy is the preferred treatment for advanced ovarian cancer, but the 5-year survival rate remains low partly because of the development of drug resistance. Although it has been reported that X-linked inhibitor of apoptosis (XIAP) causes more severe chemoresistance in ovarian cancer cells and is highly expressed in chemoresistant ovarian cancer, the molecular mechanism underlying this dysregulation is unknown. The purpose of this study was to identify microRNAs (miRNAs) that bind to the 3' untranslated region (3'UTR) of XIAP and have a role in chemoresistance in ovarian cancer. Using in silico analysis and literature review, a panel of miRNAs dysregulated in chemoresistant ovarian cancer was generated from hundreds of miRNAs that were predicted to target the XIAP 3'UTR. Using a dual luciferase reporter assay and cellular co-transfection of a miRNA expression vector and a luciferase reporter fused to the XIAP 3'UTR cognate miRNA binding site, we identified three miRNAs of which miR-142-5p had the greatest inhibitory effect. We found that overexpression of miR-142-5p suppressed XIAP expression by binding to its 3'UTR in OVCAR3 and SKOV3 cells. Using a chemosensitivity assay, we found that in OVCAR3, SKOV3, and primary epithelial ovarian cancer (EOC) cells, overexpression or inhibition of miR-142-5p increased or suppressed their sensitivities to cisplatin respectively. In contrast, introducing XIAP without a 3'UTR counteracted the effect of overexpressed miR-142-5p on cisplatin-induced apoptosis in OVCAR3 ovarian cancer cells. Furthermore, we found a negative correlation between miR-142-5p expression and XIAP protein levels in clinical samples from patients with EOC. Using clinical and miRNA expression data of more than 200 ovarian cancer patients treated with platinum-based chemotherapy from The Cancer Genome Atlas (TCGA) database, we found ovarian cancer patients with higher expression levels of miR-142-5p had longer median progression-free survival as compared to patients with lower miR-142-5p levels. We demonstrated that miR-142-5p also targeted four other anti-apoptotic genes, baculoviral IAP repeat-containing 3 (BIRC3), B-cell lymphoma-2 (BCL2), BCL2 like 2 (BCL2L2), and myeloid cell leukemia sequence 1 (MCL1) specifically. Transcriptome sequencing shed light on the essential apoptosis-related pathway in which miR-142-5p may be involved. To conclude, our findings illustrate that miR-142-5p sensitizes ovarian cancer cells to cisplatin-induced apoptosis by targeting multiple anti-apoptotic genes including XIAP, and may also suggest the therapeutic potential of miR-142-5p in ovarian cancer treatment.