DNA binding, cleavage and cytotoxicity studies of three mononuclear Cu(II) chloro-complexes containing N-S donor Schiff base ligands.

Abstract

We report the biological activity of three Cu(II) complexes [Cu(pabt)Cl] (1), [Cu(pma)Cl] (2), and [Cu(pdta)Cl]Cl (3) (pabt = N-(2-mercaptophenyl)-2'-pyridylmethylenimine, pma = N-(2-pyridylmethyl)-2-mercaptoaniline, pdta = 2,2'-di(pyridyl-2-methyleneimine)diphenyl disulfide). 1-3 display four-line EPR multiplet in solution at RT suggesting that these are mononuclear. DNA-binding studies using spectrophotometric titration of these complexes with calf thymus DNA showed binding through intercalation mode which was found to be consistent with the observation of increased viscosity of DNA and quenching of fluorescence of ethidium bromide bound DNA in the presence of these complexes. All three complexes were found to be efficient in bringing about oxidative and hydrolytic cleavage of DNA. The proposed mechanism of hydrolytic DNA cleavage has been discussed. MTT assay showed remarkable cytotoxicity on cervical cancer HeLa cell line and the IC50 values were 1.27, 4.13, and 3.92μM for 1, 2 and 3, respectively, as compared to the IC50 value (13 μM) reported for cisplatin in HeLa cells. AO/PI and Annexin-V/PI assay suggest the induction of cell death primarily via apoptotic pathway. Nuclear staining using DAPI was used to assess changes in nuclear morphology during apoptotic cell death. The role of reactive oxygen species (ROS) for induction of apoptotic cell death was studied using H2DCF-DA assay and the result suggests that the generation of ROS by the complexes may be a possible cause for their antiproliferative activity. TUNEL assay showed DNA fragmentation in apoptotic cells. Cell cycle analysis using flow cytometry showed significant increase in the G2/M phase in HeLa cells by the compounds 1-3. Mononuclear Cu(II) complexes display remarkable cytotoxicity against cervical cancer HeLa cell line. The generation of ROS by the complexes may be a cause of their antiproliferative activity. Fluorescent images from DAPI staining assay revealed that the cells undergoing apoptosis displayed typical features like cell shrinkage, membrane blebbing, chromatin condensation and nuclear fragmentation. TUNEL assay showed DNA fragmentation in apoptotic cells.