In‐silico identification of small molecules targeting H‐Ras and in‐vitro cytotoxicity with caspase‐mediated apoptosis in carcinoma cells

H‐Ras oncogene plays a critical role in the transformation of normal cells to a malignant phenotype through constitutive activation of the GTP bound protein leading to uncontrolled cell proliferation in several human cancers. Thus, H‐Ras oncoprotein serves as an excellent target for anticancer drug discovery. To identify novel H‐Ras inhibitors, we performed structure‐based virtual screening of the Maybridge HitFinder™ library using Schrodinger suite. Thirty ligands from the chemical library were identified as they showed preferential in silico binding initially to H‐Ras proteins with Gly12Val, Gly13Asp, and Gly12Val‐Gly13Asp mutations. Absorption, distribution, metabolism, excretion, and toxicity profile confirmed drug‐like properties of the compounds. Three representative molecules were tested for antiproliferative effect on T24 urinary bladder carcinoma cell line, MCF‐7 breast cancer cell line and HDF‐7 normal dermal fibroblast cells using 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide assay. Two compounds (Cmpds) showed antiproliferative activity exclusively in the cancer cell lines with minimal effect on the control HDF‐7 cells. The effect of compound treatment on cell cycle progression, assessed by propidium iodide (PI) staining, depicted increased arrest of T24 cell line in the sub G1 phase. Further, Annexin‐V PI dual staining and pan caspase inhibitor Z‐VAD‐fmk indicated caspase‐dependent apoptotic activity of Cmpds 1 and 3. Our findings demonstrate caspase‐dependent apoptotic activity of Cmpds 1 and 3 selectively against Gly12Val mutated T24 cancer cell line implicating a potential for treatment of bladder cancer. We envisage that these molecules may be promising candidates with potential therapeutic value in H‐Ras mutation‐associated cancers. In the current study, we identified thirty small drug‐like molecules targeting the oncogenic H‐Ras mutants by structure‐based virtual screening. Two of the three tested compounds (Cmpds) show specific antiproliferative activity in carcinoma cell lines with marginal effect on normal cells. Annexin‐propidium iodide (PI) dual staining using pan‐caspase inhibitor elucidated caspase‐dependent apoptosis of Cmpds 1 and 3 specifically in H‐Ras mutated urinary bladder carcinoma T24 cell line. Our preliminary study implicates the anticancer potential of the specific compounds in H‐Ras mutation associated cancers.