Hyperthermia as a method to treat cancer, either by just its thermal effects or in combination with other agents (e.g., radiative cancer treatments) and has been explored for over 30 years See Hildebrandt, B, Wust, P., Ahlers, O., Dieing, A., Sreenivasa, G., Kerner, T., Felix, R., and Riess, H., 2002, “The cellular and molecular basis of hyperthermia,” Critical Reviews in Oncology Hematology, 43, pp. 33-56; Cetas, T. C and Roemer, R. B, 1984, “Status and future developments in the physical aspects of hyperthermia,” Cancer Research, 44, pp. 4894-4901). In such treatment methods, the temperature range generally does not exceed 42° C., but studies up to almost 50° C. have been performed. See, e.g., Dickson, J. A. and Calderwood, S. K., 1980, “Temperature Range and Selective Sensitivity of Tumors to Hyperthermia: A Critical Review,” Annals New York Academy of Sciences, Vol. 335, pp. 180-205. Further, it is known that by increasing the temperature it is possible to reduce the exposure time. For example, Dickson and Calderwood (1980) have reported that hyperthermia can be obtained by increasing the local body temperature to about 50° C. Under such conditions, an exposure time on the order of 0.1 hours (6 minutes) was required to provide an effective treatment.
It is also known, that short pulses from milliseconds to nanoseconds can be used to initiate cell death. For example, melanoma tumors in mice have been successfully treated with 300 ns pulsed electrics fields with electric field strengths up to 60 kV/cm. See R. Nuccitelli, U. Pliquett, X. Chen, W. Ford, J. Swanson, S. J. Beebe, J. F. Kolb, and K. H. Schoenbach, “Nanosecond pulsed electric fields cause melanomas to self-destruct,” Biochem. Biophys. Res. Commun., vol. 343, no. 2, pp. 351-360, 2006). These pulses were delivered to the tumor with needle electrodes, or with plate electrodes surrounding the tumor. In contrast to hyperthermia treatments, such pulse treatments are based on non-thermal effects.
Studies combining these two effects, and controlling them independently of each other, has been limited to the studies of bacteria. See S. Jayaram, G. S. P. Castle, and A. Margaritis, “Effects of High Electric Field Pulses on Lactobacillus Brevis at Elevated Temperatures,” Conference Record of the 1991 IEEE Industry Applications Society Annual Meeting, vol. 1, pp. 674-681, October 1991, T. Ohshima, K. Okuyama, M. Sato, “Effect of culture temperature on high voltage pulse sterilization of Escherichia coli,” Journal of Electrostatics, vol. 55, pp. 227-235, 2002. In such studies, the pulse durations were on the order of microseconds. Further, these studies indicated some enhancement at 60° C. compared to 30° C. In particular, the initial live cell count was 109 cells/cm3 at 30° C. The live cell count then decreased to 107 cells/cm3 when the temperature was increased to 60° C. for ten seconds. When exposed to 60 pulses (on the order of microseconds) of an amplitude of about 30 kV/cm, the live cell count decreased to 104 cells/cm at 30° C. and the live cell count was negligible at 60° C.
The cause of cell death of bacteria due to the applied electric fields was assumed to be due to electroporation. The increase in temperature causes an increase in the conductivity of the medium and cytoplasm of the cell, as well as a decrease in the viscosity of the membrane. Based on the increase in membrane and cytoplasm conductivity, it was concluded that the observed increase in cell death at higher temperature is due to a more rapid increase in membrane potential, leading to an earlier breakdown of the membrane. Similar results were obtained with E. coli in which an increase in bacteria death was observed when pulses at elevated temperatures were applied. Here, it was also assumed that this effect is due to a decrease in membrane viscosity.
It is known that cells exposed to certain pulsed electric fields will have increased permeability and enable nonpermeant molecules to enter the cell. This has been demonstrated for a variety of molecules including chemotherapeutic agents (Mir L M, Banoun H, Paoletti C. Introduction of definite amounts of nonpermeant molecules into living cells after electropermeabilization: direct access to the cytosol. Exp Cell Res 1988; 175:15-25) and nucleic acids (Heller, L and Heller R, In vivo electroporation for gene therapy, Human Gene Therapy, 2006; 17:890-897) among other molecules. Pulsed electric fields can also manipulate molecules to enable cell fusion as well as insertion of molecules into cell membranes. Combination of heat and electric pulses will facilitate these interactions due to the decrease in viscosity and increased fluidity of the cell membrane.