Unveiling Tetrahydroberberine-1,3,4-Oxadiazole hybrids as potent anticancer agents and MMP-9 inhibitors with comprehensive insights from in silico, in vitro, and ex vivo permeation studies

Overexpression of the MMP9 enzyme is a critical biomarker in colorectal Cancer (CRC) and a key promoter of metastasis, contributing to mortality rates. Thus, a novel series of Tetrahydroberberine-1,3,4-oxadiazole series was synthesized to halt CRC progression via MMP9 inhibition. The designed molecules were incorporated with pharmacophoric features observed in MMP9 inhibitors. All the newly synthesized hybrids (10a-i) and (12a-e) were subjected to antiproliferative activity using the MTT assay against different Cancer cell lines. Among them, compound 10 g was found to be 16-fold selective towards the HCT-116 Cancer cell line with an IC₅₀ value of 1.54 ± 0.06 μM compared to a non-cancerous cell line, i.e., HEK293. Compound 10 g mediated Apoptosis was observed in cellular and nuclear morphology of cancerous cells. Quantitative analysis was performed using PI staining to validate its apoptotic potential. Target-based study for compound 10 g revealed excellent inhibition of MMP9, as evidenced by Western blotting, Immunofluorescence study and MMP9 enzyme inhibition assay. In-silico studies, including molecular docking and molecular dynamics simulations, corroborated the experimental results by demonstrating strong binding interaction and stability at the MMP9 active site. Additionally, solubility and ex-vivo permeation studies concluded the improved pharmacokinetic properties of compound 10 g over berberine. Collectively, these finding highlights compound 10 g as a promising lead candidate for further development as an Anticancer agent targeting MMP9 in colorectal Cancer.