Antiproliferation of MP12 derived from a fungus, Aphanomyces invadans virulence factor, cysteine-rich trypsin inhibitor on human laryngeal epithelial cells, and in vivo zebrafish embryo model

Peptide-based drug development is an emerging and promising approach in cancer therapeutics. The present study focuses on understanding the mechanism of MP12 peptide (MDNHVCIPLCPP) derived from cysteine-rich trypsin inhibitor protein of virulence factor of pathogenic fungus Aphanomyces invadans. MP12 is involved in antiproliferative activity against the human laryngeal epithelial cell (Hep-2), demonstrated in this study. MP12 sequence showed a significant binding score and has multiple hydrogen bond interactions with the proteins that play a vital role in apoptotic pathways such as Bcl-2, caspase-3, caspase-7, and XIAP. Based on the bioinformatics characterization and molecular docking result, further study was focused on MP12 antiproliferative activity. The peptide showed a dose-dependent inhibition against Hep-2 cell line proliferation, analyzed over MTT and neutral red uptake assays. The IC₅₀ value of the MP12 peptide was calculated based on the antiproliferative property (24.7 ± 0.34 μM). MP12 treated Hep-2 cells showed significant shrinkage in cell morphology compared to untreated cells, inhibiting the cell cycle. The gene expression analysis validated that the MP12 significantly upregulates the caspase-3, caspase-7, and caspase-9 genes. The developmental toxicity study using zebrafish embryos as in vivo model proved that the MP12 is nontoxic. Based on the obtained results, we proposed that the peptide MP12 derived from cysteine-rich trypsin inhibitor protein of virulence molecule of pathogenic fungus have a potential antiproliferative activity. However, further clinical trials need to be focused on the mechanism and therapeutic application against laryngeal cancer.