Radiosensitising effect of electrochemotherapy with bleomycin in LPB sarcoma cells and tumors in mice

BACKGROUND: Bleomycin is poorly permeant but potent cytotoxic and radiosensitizing drug. The aim of the study was to evaluate whether a physical drug delivery system – electroporation can increase radiosensitising effect of bleomycin in vitro and in vivo. METHODS: LPB sarcoma cells and tumors were treated either with bleomycin, electroporation or ionizing radiation, and combination of these treatments. In vitro, response to different treatments was determined by colony forming assay, while in vivo, treatment effectiveness was determined by local tumor control (TCD(50)). Time dependence of partial oxygen pressure in LPB tumors after application of electric pulses was measured by electron paramagnetic oxyimetry. RESULTS: Electroporation of cells in vitro increased radiosensitising effect of bleomycin for 1.5 times, in vivo radiation response of tumors was enhanced by 1.9 fold compared to response of tumors that were irradiated only. Neither treatment of tumors with bleomycin nor application of electric pulses only, affected radiation response of tumors. Application of electric pulses to the tumors induced profound but transient reduction of tumor oxygenation. Although tumor oxygenation after electroporation partially restored at the time of irradiation, it was still reduced at the level of radiobiologically relevant hypoxia. CONCLUSION: Our study shows that application of electric pulses to cells and tumors increases radiosensitising effect of bleomycin. Furthermore, our results demonstrate that the radiobiologically relevant hypoxia induced by electroporation of tumors did not counteract the pronounced radiosensitising effect of electrochemotherapy with bleomycin

Vascular disrupting action of electroporation and electrochemotherapy with bleomycin in murine sarcoma

Electrochemotherapy has a direct cytotoxic effect on tumour cells, and presumably, a vascular disrupting effect. In this study, on the basis of the prediction of the mathematical model, histological evaluation and physiological measurements of the tumours were carried out to confirm that electroporation and electrochemotherapy of tumours have a vascular disrupting action. In the study, SA-1 solid subcutaneous sarcoma tumours in A/J mice were treated by bleomycin (BLM) given intravenously (1 mg kg−1), application of electric pulses (8 pulses, 1040 V, 100 μs, 1 Hz) or a combination of both – electrochemotherapy. The vascular effect was determined by laser Doppler flowmetry, power Doppler ultrasonographic imaging and Patent blue staining. The extent of tumour hypoxia was determined immunohistochemically by hypoxia marker pimonidazole and partial pressure of oxygen (pO2) in tumours by electron paramagnetic resonance oximetry. Electrochemotherapy with BLM induced good antitumour effect with 22 days, tumour growth delay and 38% tumour cures. The application of electric pulses to the tumours induced instant but transient tumour blood flow reduction (for 70%) that was recovered in 24 h. During this tumour blood flow reduction, we determined an increase in hypoxic tumour area for up to 30%, which was also reflected in reduced tumour oxygenation (for 70%). According to the described mathematical model, endothelial cells lining in tumour blood vessels are exposed to a ∼40% higher electric field than the surrounding tumour cells, and therefore easily electroporated, allowing access of high BLM concentration to the cytosol. Consequently, electrochemotherapy has, besides the immediate vascular disrupting action, also a delayed one (after 24 h), as a consequence of endothelial cell swelling and apoptosis demonstrated by extensive tumour necrosis, tumour hypoxia, prolonged reduction of tumour blood flow and significant tumour growth delay, and tumour cures. Our results demonstrate that in addition to the well-established direct cytotoxic effect on tumour cells, electrochemotherapy also has an indirect vascular disrupting action resulting altogether in extensive tumour cell necrosis leading to complete regression of tumours

Increase in fluidity in the membrane of MT3 breast cancer cells correlates with enhanced cell adhesion in vitro and increased lung metastasis in NOD/SCID mice

To study whether membrane fluidity of tumor cells have an influence on metastasis, MT3 breast cancer cells harvested during exponential growth and under confluent conditions were compared. Electron paramagnetic resonance (EPR) data revealed that, in comparison to growing cells, confluent cells have a significant higher fluidity in their membrane related to a higher relative portion of disordered domains and a reduced portion of the most ordered domains. Further, sialyl Lewis X and/or A ligand-mediated adhesion of these cells was 2-fold enhanced. Confocal laser scanning microscopy further demonstrated a higher motility of ligands in the membrane of confluent cells, together with an accumulation of these ligands in distinct areas. Both facts are suggested to be responsible for an enhanced cell adhesion observed. Finally, an increased number of large distinct metastatic foci was registered in lungs of mice after i.v. inoculation of confluent cells. The results indicates that domain organization and fluidity of the cell membrane affect tumor cell adhesion and can have in this way also an impact on the malignancy of breast cancer cells

Performance of Waterborne Cu(II) Octanoate/Ethanolamine Wood Presevatives

Various aqueous wood preservative solutions containing Cu(II) in the form of copper(II) sulphate or copper(II) octanoate, ethanolamine and in one case octanoic acid were investigated by spectrophotometry, polarography and Electron Paramagnetic Resonance (EPR) spectroscopy. Results have shown the same coordination environment around Cu(II) in all solutions with ethanolamine. Computer simulation of the EPR spectra also revealed that the coordination in the first coordination sphere of copper is the same at low concentration of ethanolamine. At 20% ethanolamine concentration, a mixture of two complexes (one with two nitrogens and the other with three) could be detected. The active compound in the investigated ethanolamine containing solutions is the same when previously synthesised copper(II) octanoate was used, or when copper(II) sulphate and octanoic acid were utilized instead. Fungicidal and leaching experiments with the treated wood resulted in the same conclusion: it is not necessary to use pre-synthesised copper(II) octanoate for the preparation of waterborne copper/ethanolamine wood preservatives. Preservative preparation time and costs can be reduced by simply dissolving copper(II) sulphate and octanoic acid in aqueous ethanolamine solutions