Computational structural modelling of the SRCR region of the SCARA5 receptor and in vitro investigation of the cytotoxic effects of a receptor blocker developed by using computer-based drug design method in MFC7 human breast cancer cell line

The computer-aided drug design method uses three-dimensional structures of potential drug targets involved in the disease process to design new molecules. Cancer is a major health problem for both developed and developing countries. In the last decade, researchers have investigated small peptides with anticancer properties as a potential alternative approach for cancer therapy. Therefore, the aim of this thesis is to design a peptide to inhibit the interaction of Scavenger Receptor Class A member 5 (SCARA5), a candidate therapeutic target in cancer, with its ligand L-ferritin and to investigate the effect of this peptide on human breast cancer cell line MCF7 in vitro. The thesis study consists of two stages, in silico and in vitro. For this purpose, a computational structural model of the three-dimensional structure of the cysteine-rich scavenger receptor (SRCR), the ligand binding site of SCARA5, was developed in silico. The modelled structure was compared with the three-dimensional structure revealed by X-ray crystallographic method. Robetta_cm was selected as the most appropriate model and its interaction with its ligand L-Ferritin was revealed by protein-protein docking method. Amino acid residues 16-22 and 79 in this interaction region were identified as target sites for peptide design. In the in vitro stage of the thesis study, the effects of the peptide on the viability of MCF7 cells were determined by WST-1 and Sulforhodamine B assay. The in vitro stage was completed with ELISA and Western blot techniques. The peptide exhibited cytotoxic effects in MCF7 cells at different doses and incubation times. The half maximum inhibitory concentration (IC50) value of the peptide on MCF7 cells was calculated as 188 μg/ml. Protein levels of initiator caspase-9 and executioner caspase-7, which are important markers for determining whether the cytotoxic effects of the peptide are apoptosis-induced, were determined by ELISA test. Peptide doses were determined as 2X IC50, IC50 and 1/2X IC50. At the end of 24 hours incubation period, the amount of IC50 and 1/2X IC50 doses applied to the cells increased statistically. After 48 hours, an increase in caspase-9 was observed at 1/2X IC50 dose. The dose and time of increase in the amount of caspase-7 were IC50 and 48th hour incubation period. At the end of 72 hours of incubation, an increase in the amount of caspase-7 was observed in cells treated with 2X IC50. According to Western blot results, the peptide increased the amount of caspase-7 in MCF7 cells after 24 hours of incubation, while the amount of cleaved caspase-7 increased after 48 hours of incubation. The designed peptide triggered apoptotic cell death in MCF7 cells. KEYWORDS: Anticancer Peptide, Caspase, Drug Design, MCF7, Molecular Docking, Molecular Dynamic Simulation.