Background: There is a significant rise in the prevalence of colon cancer worldwide. A variety of therapeutic approaches have been tried, but none have been successful due to the aggressive spread of the tumor. Natural products derived from plants have traditionally been used in the medical field. Objective: The aim of the current study was to investigate the effect of Cassia Fistula cytotoxicity on a colon cancer cell line and the synergy between Cassia fistulae extract and cisplatin significantly to induce apoptosis through upregulation of p53 and Msh2. Material and methods: Real-Time polymerase chain reaction indicated that gene expression proteins were likely involved in the pathway when the cytotoxicity potential of the plant extract on colon cancer cells was assessed using the (MTT) test. The database search techniques used with colon cancer cells identified the p53 gene and Msh2 gene's apoptotic function. Results: Aqueous extract of the plant was most toxic to cancer cells. Further, p53 gene expression and Msh2 were observed to be high in cells treated with the plant extracts of C. fistula and IC₅₀of cisplatin (CDDP) +Cassia Fistula suggesting an involvement of mitochondrial proteins in the colon cancer cell apoptosis. Conclusion: The results of the present study indicate that Cassia fistula extracts significantly regulate p53 and Msh2 expression to induce apoptosis. According to its cytotoxic and apoptotic action, Cassia fistula can be considered as a new discovery for the treatment of colon cancer.