Abstract
Calcium electroporation is a novel anti-cancer treatment investigated in clinical trials. We explored cell sensitivity to calcium electroporation and electroporation with bleomycin, using viability assays at different time and temperature points, as well as heat calorimetry, lipidomics, and flow cytometry. Three cell lines: HT29 (colon cancer), MDA-MB231 (breast cancer), and HDF-n (normal fibroblasts) were investigated for; (a) cell survival dependent on time of addition of drug relative to electroporation (1.2 kV/cm, 8 pulses, 99 µs, 1 Hz), at different temperatures (37 °C, 27 °C, 17 °C); (b) heat capacity profiles obtained by differential scanning calorimetry without added calcium; (c) lipid composition by mass spectrometry; (d) phosphatidylserine in the plasma membrane outer leaflet using flow cytometry. Temperature as well as time of drug administration affected treatment efficacy in HT29 and HDF-n cells, but not MDA-MB231 cells. Interestingly the HT29 cell line displayed a higher phase transition temperature (approximately 20 °C) versus 14 °C (HDF-n) and 15 °C (MDA-MB231). Furthermore the HT29 cell membranes had a higher ratio of ethers to esters, and a higher expression of phosphatidylserine in the outer leaflet. In conclusion, lipid composition and heat capacity of the membrane might influence permeabilisation of cells and thereby the effect of calcium electroporation and electrochemotherapy.
Highlights
In electrochemotherapy tumor cells are permeabilised by electroporation thereby enhancing their uptake of chemotherapeutic drugs[1]
Each cell line was treated with calcium electroporation and bleomycin electroporation
Treatment with calcium alone resulted in 88–119% viability, treatment with calcium and bleomycin resulted in 86–113% viability, treatment with bleomycin alone resulted in 80–104% viability, and electroporation alone resulted in 70–88% viability, which correlate with previous studies on these cell lines[39]
Summary
In electrochemotherapy tumor cells are permeabilised by electroporation thereby enhancing their uptake of chemotherapeutic drugs (primarily bleomycin and cisplatin are used)[1]. The use of electrochemotherapy for treatment of internal tumors is currently being investigated in clinical trials[11,12,13,14,15,16]. As calcium electroporation may find importance in the clinic, we decided to investigate optimal temperature and time of administration of calcium in relation to electroporation, comparing with bleomycin electroporation as control. We compared calcium electroporation to bleomycin electroporation to investigate possible differences in the dependency of drug administration before and after electroporation as well as temperature. The combination of calcium and bleomycin was investigated to examine if addition of calcium would change the effect of bleomycin, as evidenced by alteration of the cytotoxic effect of bleomycin
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