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Abstract
Introduction/Background High-Grade Serous Ovarian Cancer (HGSOC) is the most dominant subtype of Epithelial Ovarian Cancer (EOC). Cytoreductive surgery and platinum-based chemotherapy are main treatment modalities for HGSOC. However, 25% of patients develop resistance within 6 months. Although poly (ADP-ribose) polymerase (PARP) inhibitors can be used as second-line therapy, a high recurrence rate (>%70) within 3 years emphasizes the need for new treatment strategies. microRNAs (miRNAs) play an important role in various cancer related-pathways as suppressors or promoters. This study aims to explore miRNA’s role in HGSOC and investigate possible combinatorial drug therapy strategies to overcome drug resistance.
Methodology Total 40 FFPE sample of patients diagnosed with HGSOC were included in this study in collaboration with Koç University Hospital, Gynecology, and Oncology Department. Differential miRNA expression levels between 20 sensitive and 20 resistant FFPE samples were determined by microarray and qRT-PCR assays. For in vitro studies, resistant HGSOC cell lines were generated by the stepwise dose-escalation method. miRNA-mimic molecules were transfected into resistant cells to test the synergistic effects of miRNAs and drugs. Downstream molecular investigations were performed by western blot and flow cytometry experiments.
Determination of IC50 values and resistant index of resistant OVCAR-3, CAOV-3 and OVSAHO cells by SRB Cell Viability Assay. Cells were treated with olaparib and carboplatin drugs for 1 week and results were normalized with DMSO control. Figure 1: Characterization of acquired drug resistance in OVSAHO (A), OVCAR-3 (B) and CAOV-3 (C) cells by clonogenic assay. DMSO was used as control. Figure 2: Differential miRNA gene expression levels were determined by microarray (A) and qRT-PCR assays in FFPE (B) and in vitro resistant cell samples (C, D, E). Fold change value >|1.5| was determined as significant. Figure 3: Combinational treatment with miRNA-mimic and conventional drugs analysis were examined by SRB cell viability assay in mimic transfected resistant cell samples (A, B, C). Results were compared with untransfected resistant cells. Significancy were determined using non-parametric t-test.
Results Depending on the differential expression analysis of miRNAs, 10 candidate miRNAs were selected in resistant FFPE and cell samples. In consideration of combinational drugs and miRNA-mimic treatment results, the synergistic effect of let7b-5p and 188–5p with olaparib and carboplatin drugs in resistant OVCAR-3 and CAOV-3 cells was shown. Most synergistic miRNA and drug combinations were selected for further downstream molecular assays to re-sensitize resistance cells to therapy.
Conclusion Within the scope of this study, role of miRNA’s in the context of treatment response and resistance in HGSOC is explored and synergistic effects of miRNA’s with combination therapy strategies with drugs used in the clinic is tested. This informations will shed light on the alternative treatment of HGSOC.
Disclosures Koç University Ethics Committee approved the use of human material (2019.257.IRB2.079)).