ABSTRACT
BACKGROUND: Copper complex has been gaining much attention in anticancer research as a targeted agent since cancer cells uptake more copper than non-cancerous cells. Our group synthesised a ternary copper complex, which is composed of 1,10-phenanthroline and tyrosine [Cu(phen)(L-tyr)Cl].3H20. These two payloads have been designed to cleave DNA and inhibit protein degradation system (proteasome) concurrently in cancer cells, making this copper complex a dual-target compound. OBJECTIVE: The current study was carried out to investigate the mode of cell death and the role of autophagy induced by [Cu(phen)(L-tyr)Cl].3H20 in MCF-7 and MDA-MB-231 breast cancer cells. METHODS: Growth inhibition of [Cu(phen)(L-tyr)Cl].3H20 towards MDA-MB-231 and human non-cancerous MCF10A breast cells was determined by MTT assay. Annexin-V-FITC/PI and cell cycle analysis were evaluated by flow cytometry. The expression of p53, Bax, caspase-9, caspase-7, caspase-3 and LC3 was determined using western blot analysis. The cells were then co-treated with hydroxychloroquine to ascertain the role of autophagy induced by [Cu(phen)(L-tyr)Cl].3H20. RESULTS: [Cu(phen)(L-tyr)Cl].3H20 inhibited the growth of cancer cells dose-dependently with less toxicity towards MCF10A cells. Additionally, [Cu(phen)(L-tyr)Cl].3H20 induced apoptosis and cell cycle arrest towards MCF-7 and MDA-MB-231 breast cancer cells possibly via regulation of p53, Bax, caspase-9, caspase-3 and capase-7. The expression of LC3II was upregulated in both cancer cell lines upon treatment with [Cu(phen)(L-tyr) Cl].3H20, indicating the induction of autophagy. Co-treatment with autophagy inhibitor hydroxychloroquine significantly enhanced growth inhibition of both cell lines, suggesting that autophagy induced by [Cu(phen)(L-tyr) Cl].3H20 in both breast cancer cells promoted cell survival. CONCLUSION: [Cu(phen)(L-tyr)Cl].3H20 holds great potential to be developed for breast cancer treatment.
Subject(s)
Breast Neoplasms , Copper , Apoptosis , Autophagy , Breast Neoplasms/drug therapy , Caspase 3 , Caspase 9 , Cell Line, Tumor , Cell Proliferation , Copper/pharmacology , Female , Humans , Hydroxychloroquine , MCF-7 Cells , Tumor Suppressor Protein p53 , bcl-2-Associated X ProteinABSTRACT
BACKGROUND: The lack of specificity, severe side effects, and development of drug resistance have largely limited the use of platinum-based compounds in cancer treatment. Therefore, copper complexes have emerged as potential alternatives to platinum-based compounds. OBJECTIVE: Ternary copper (II) complex incorporated with 1-10-phenanthroline and L-tyrosine was investigated for its anti-cancer effects in HT-29 colorectal cancer cells. METHODS: Cytotoxic effects of ternary copper (II) complex in HT-29 cells was evaluated using MTT assay, Real-Time Cell Analysis (RTCA) and lactate dehydrogenase (LDH) assay. Cell cycle analysis was performed using flow cytometry. Apoptosis induction was studied by Annexin V-FITC/Propidium Iodide (PI) staining and mitochondrial membrane potential analysis (JC-10 staining) using flow cytometry. Intracellular Reactive Oxygen Species (ROS) were detected by DCFH-DA assay. The expression of proteins involved in the apoptotic signalling pathway (p53, caspases, and PARP-1) was evaluated by western blot analysis. RESULTS: Ternary copper (II) complex reduced the cell viability of HT-29 cells in a time- and dose-dependent manner, with IC50 of 2.4 ± 0.4 and 0.8 ± 0.04 µM at 24 and 48 hours, respectively. Cell cycle analysis demonstrated induction of S-phase cell cycle arrest. Morphological evaluation and Annexin V-FITC/PI flow cytometry analysis confirmed induction of apoptosis that was further supported by cleavage and activation of caspase-8, caspase-9, caspase-3, and PARP- 1. Mutant p53 was also downregulated in a dose-dependent manner. No LDH release, mitochondrial membrane potential disruption, and ROS production were observed. CONCLUSION: Ternary copper (II) complex holds great potential to be developed for colorectal cancer treatment.
