Investigation of the efficacy of encapsulated miR-3670, miR-8078 based treatment approach in immunotherapy of breast cancer

The aim of this study was to inhibit the expression of PD-L1, an immune checkpoint overexpressed in MCF-7 breast cancer cells, by using microRNA-3670 (miR-3670) and microRNA-8078 (miR-8078) and evaluate its immune therapeutic properties in breast cancer. is to research. As a result of our bioinformatic analysis, miR-3670 can bind to two different regions of PD-L1 mRNA with high binding scores (82.15 and 87.35), significantly targets PD-L1 ligand synthesis pathways and basic regulatory pathways of cancer cells, and inhibits PD-L1 expression in breast cancer. It has been found to effectively suppress. Cytotoxicity analysis determined that the 1nM dose of both miRNAs exhibited high toxicity in the MCF-7 cell line, while being less toxic in HME1, a healthy breast tissue cell line. Total RNA analyzes determined that there was a significant correlation between total RNA concentrations and cytotoxicity levels. While a decrease in the amount of total RNA was observed at high cytotoxic doses, an increase was determined at low cytotoxic doses. Based on our RT-PCR and ELISA analyses, 1 nM doses of miRNA-3670 and miRNA-8078 mimics were found to reduce both PD-L1 gene expression and sPD-L1 protein levels in MCF-7 cells. However, no significant effect on HME1 cells was detected. In MCF-7 cells, miR-3670 and miR-8078 reduced cell survival and triggered apoptosis, while it did not cause a significant difference in HME1 cells. According to our cell migration analysis, miRNAs had a strong inhibitory effect on cell migration in MCF-7 cells (81.47% and 99.8%, respectively) and exhibited anti-metastatic property. Conversely, it was found to be unable to inhibit cell migration in HME1 cells (22.8% and 6.2%, respectively). Based on morphology and bioinformatics analyzes (KEGG and GO), both miRNAs were identified to cause a significant decrease in the presence of lamellipodia, causing the transition from EMT to MET in MCF-7 cells. However, no visible morphological changes were observed in HME1 cells. According to ELISA analyzes performed on metastasis-related parameters, it was observed that 1nM doses of miR-3670 and miR-8078 significantly reduced free e-cadherin antigen levels in MCF-7 and HME1 cells. Additionally, miR-3670 and miR-8078 were found to cause a decrease in the sVEGF-A level and sTINAGL-1 antigen level in MCF-7 and HME1 cells. It was determined that miR-3670 and miR-8078 significantly inhibited colony formation in MCF-7 cells, with a decrease of 17.7% and 78.2%, respectively. It was found to inhibit 3D spheroid formation in the MCF-7 cell line by 90% and cause the development of amorphous spheroids (13% and 22%, respectively), as opposed to the typically ellipsoid-shaped 3D spheroids. It was determined that the size of the 3D spheroids created was below 100 µm and their volume was less than 1 mm3. 3D spheroid structure was not observed in the HME1 cell line. In conclusion; demonstrated that miRNAs (miR-3670, miR-8078) may have potential for cancer immunotherapy through redirection of immune responses in breast cancer cells. It may also provide a basis for future research into personalized cancer immunotherapy and targeted therapy, which is becoming increasingly important in current cancer treatment approaches in particular.