Cisplatin, glutathione and the third wheel: a copper-(1,10-phenanthroline) complex modulates cisplatin-GSH interactions from antagonism to synergism in cancer cells resistant to cisplatin.

The antagonistic effect of glutathione (GSH) against the cytotoxicity of cisplatin was observed in both wild type and cisplatin-resistant human leukaemia and ovarian carcinoma cell lines. The simultaneous presence of the cytotoxic copper complex [Cu(phen)2(OH2)](ClO4)2 (C0) restored the sensitivity of the cells to cisplatin, and, at selected concentrations, led to strong synergistic effects. The C0-cisplatin-glutathione system showed a synergistic toxic effect even in the presence of 1000 µM GSH. The three-drug cocktail exerted a higher potency against leukemic cells than against freshly isolated lymphocytes from healthy donors. Compared to actively proliferating normal lymphocytes, leukaemia cells were much more susceptible to the cytocide effect of the three-drug combination and underwent the dying process(es) much faster. When the ovarian carcinoma cells were treated with cisplatin, alone or in combination with C0, late apoptotic effects were mainly observed, suggesting that DNA interactions with the C0-cisplatin complex trigger a process of programmed cell death. In contrast, the ternary combination induced apoptotic effects similar to that shown by C0 in single treatment, that is, early apoptosis. One possible explanation is that C0 and cisplatin compete for GSH-binding in the culture medium. GSH in combination with C0 and cisplatin caused a significant induction of the apoptotic process(es), through a pathway which does not compromise the integrity of the plasma membrane of cells.

Mass spectrometric discrimination of phospholipid patterns in cisplatin-resistant and -sensitive cancer cells.

RATIONALE: Development of therapy-resistant cancer is a major problem in clinical oncology, and there is an urgent need for novel markers identifying development of the resistant phenotype. Lipidomics represents a promising approach to discriminate lipid profiles of malignant phenotype cells. Alterations in phospholipid distribution or chemical composition have been reported in various pathologies including cancer. Here we were curious whether quantitative differences in phospholipid composition between cisplatin-resistant and -sensitive model cancer cell lines could be revealed by mass spectrometric means. METHODS: The phospholipid contents of cell membranes of the cancer cell lines CCRF-CEM and A2780, both responsive and resistant to cisplatin, were analyzed by solid-phase extraction (SPE) and electrospray ionization mass spectrometry (ESI-MS and tandem mass spectrometry (MS/MS)). Extracts were obtained by disruption of cells with a dounce tissue grinder set followed by centrifugation. To minimize the enzymatic activity, phospholipids were extracted from cell extracts by SPE immediately after the cell lysis and analyzed by MS. Both supernatant and pellet fractions of cell extracts were analyzed. RESULTS: A phospholipid profile specific for cell lines and their phenotypes was revealed. We have documented by quantitative analysis that phosphocholines PC P-34:0, PC 34:1, PC 20:2_16:0, LPC 18:1 and LPC 16:0 PLs were present in the 200-400 µM concentration range in CCRF-CEM cisplatin-responsive cells, but absent in their cisplatin-resistant cells. Similarly, PC 34:1, LPC 18:1 and LPC 16:0 were increased in cisplatin-responsive A2780 cells, and PC 20:2_16:0 was downregulated in cisplatin-resistant A2780 cells. CONCLUSIONS: In this work we showed that the ESI-MS analysis of the lipid content of the therapy-resistant and -sensitive cells can clearly distinguish the phenotypic pattern and determine the potential tumor response to cytotoxic therapy. Lipid entities revealed by mass spectrometry and associated with development of therapy resistance can thus support molecular diagnosis and provide a potential complementary cancer biomarker.

Mixed copper-platinum complex formation could explain synergistic antiproliferative effect exhibited by binary mixtures of cisplatin and copper-1,10-phenanthroline compounds: An ESI-MS study.

Cisplatin, cis-diammineplatinum(II) dichloride, is a metal complex used in clinical practice for the treatment of cancer. Despite its great efficacy, it causes adverse reactions and most patients develop a resistance to cisplatin. To overcome these issues, a multi-drug therapy was introduced as a modern approach to exploit the drug synergy. A synergistic effect had been previously found when testing binary combinations of cisplatin and three copper complexes in vitro, namely, Cu(phen)(OH2)2(OClO3)2, [Cu(phen)2(OH2)](ClO4)2 and [Cu(phen)2(H2dit)](ClO4)2,(phen=1,10-phenanthroline, H2dit=imidazolidine-2-thione), against the human acute T-lymphoblastic leukaemia cell line (CCRF-CEM). In this work [Cu(phen)2(OH2)](ClO4)2 was also tested in combination with cisplatin against cisplatin-resistant sublines of CCRF-CEM (CCRF-CEM-res) and ovarian (A2780-res) cancer cell lines. The tested combinations show a synergistic effect against both the types of resistant cells. The possibility that this effect was caused by the formation of new adducts was considered and mass spectra of solutions containing cisplatin and one of the three copper complexes at a time were measured using electrospray ionisation at atmospheric-pressure mass spectrometry (ESI-MS). A mixed complex was detected and its stoichiometry was assessed on the basis of the isotopic pattern and the results of tandem mass spectrometry experiments. The formed complex was found to be [Cu(phen)(OH)µ-(Cl)2Pt(NH3)(H2O)](+).