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Mode of action studies on cytotoxic copper(II) and rhenium(I) salicylaldehyde semicarbazone complexes
Author
Lee, Wan Yen
Supervisor
Lee, Peter Peng Foo
Yan, Yaw Kai
Abstract
This thesis is concerned with the in vitro evaluation of the biological activities of five salicylaldehyde-N,N-disubstituted semicarbazones, HOC6H4CH=N-NHCONR2 (R = alkyl or aryl group) (abbreviated as H2R2), and their copper(II) and rhenium(I) tricarbonyl complexes, against four human cancer cell lines. The semicarbazones and complexes, [CuCl(HR2)] and [ReBr(CO)3(H2R2)], were first tested for their in vitro cytotoxicities on the MOLT-4 (leukemia), MCF-7 (mammary adenocarcinoma), A-549 (lung carcinoma) and SK-II (skin carcinoma) cell lines, and one cell line of benign origin derived from human foreskin fibroblasts. The IC50 values of the compounds were determined using the MTT assay. Results revealed that the Cu(II) complexes have higher in vitro activities than the Re(I) complexes and the reference drug, cisplatin, on all the tumor cell lines, with IC50 values at micromolar levels.
Next, apoptotic and cell cycle studies were conducted to determine the mode of cell death induced by four of the complexes, [CuCl(HBnz2)], [CuCl(HBu2)], [ReBr(CO)3(H2Bnz2)] and [ReBr(CO)3(H2Bu2)], on MOLT-4 cells. Results revealed that the complexes mediated cytotoxicity in MOLT-4 cells via apoptosis. The effect of the complexes on MOLT-4 cells was also characterized by a proteomic investigation, during which 33 proteins were identified. These proteins can be grouped functionally as molecular chaperones and cytoskeletal, apoptosisrelated, glucose metabolic, cell cycle-related, stress-responsive, DNA replication, tRNA processing, transcription and translation proteins. Similarities in the protein expression patterns were observed in cells treated with the two Cu(II) complexes. A smaller number of the identified proteins were differentially expressed following treatment with Re(I) complexes.
Finally, the DNA binding and cleavage properties of [CuCl(HBnz2)], [CuCl(HBu2)], [ReBr(CO)3(H2Bnz2)] and [ReBr(CO)3(H2Bu2)] were studied by spectrophotometric DNA titration, fluorescence-based ethidium bromide competitive assay and electrophoretic gel mobility shift assay. The Cu(II) complexes were found to bind to DNA more strongly than the Re(I) complexes through a partial intercalation mode with binding constants of 1.1 × 104 M −1 and 9.52 × 103 M −1. The Cu(II) complexes also displayed a more efficient DNA cleavage ability in the presence of ascorbic acid than the Re(I) complexes. This DNA cleavage activity was found to be mediated via hydroxyl radicals that are most likely generated from atmospheric oxygen via redox cycling between Cu(II) and Cu(I) states. The results obtained in this project suggest that Cu(II) complexes of salicylaldehyde semicarbazones are a potential class of antitumor agents.
Next, apoptotic and cell cycle studies were conducted to determine the mode of cell death induced by four of the complexes, [CuCl(HBnz2)], [CuCl(HBu2)], [ReBr(CO)3(H2Bnz2)] and [ReBr(CO)3(H2Bu2)], on MOLT-4 cells. Results revealed that the complexes mediated cytotoxicity in MOLT-4 cells via apoptosis. The effect of the complexes on MOLT-4 cells was also characterized by a proteomic investigation, during which 33 proteins were identified. These proteins can be grouped functionally as molecular chaperones and cytoskeletal, apoptosisrelated, glucose metabolic, cell cycle-related, stress-responsive, DNA replication, tRNA processing, transcription and translation proteins. Similarities in the protein expression patterns were observed in cells treated with the two Cu(II) complexes. A smaller number of the identified proteins were differentially expressed following treatment with Re(I) complexes.
Finally, the DNA binding and cleavage properties of [CuCl(HBnz2)], [CuCl(HBu2)], [ReBr(CO)3(H2Bnz2)] and [ReBr(CO)3(H2Bu2)] were studied by spectrophotometric DNA titration, fluorescence-based ethidium bromide competitive assay and electrophoretic gel mobility shift assay. The Cu(II) complexes were found to bind to DNA more strongly than the Re(I) complexes through a partial intercalation mode with binding constants of 1.1 × 104 M −1 and 9.52 × 103 M −1. The Cu(II) complexes also displayed a more efficient DNA cleavage ability in the presence of ascorbic acid than the Re(I) complexes. This DNA cleavage activity was found to be mediated via hydroxyl radicals that are most likely generated from atmospheric oxygen via redox cycling between Cu(II) and Cu(I) states. The results obtained in this project suggest that Cu(II) complexes of salicylaldehyde semicarbazones are a potential class of antitumor agents.
Date Issued
2011
Call Number
RC268 Lee
Date Submitted
2011