Radiofrequency-induced Cellular Hyperthermia: Water-soluble Fullerene as a New Cancer Therapeutic Agent
Cheney, Matthew Andrade
Wilson, Lon J.
Doctor of Philosophy
Due to the susceptibility of cancer to hyperthermia, abundant biomedical research is being conducted for the development of therapies using nanotechnology for noninvasive cancer hyperthermia. In this work, the water-soluble and neutrally-charged C60 fullerene, C60-ser, is presented as a new cancer therapeutic agent for the hyperthermia of liver cancer using radiofrequency (RF) energy. With liver cancer being the second leading cause of death in men and the sixth most in women worldwide, new and improved therapies are needed to combat the limitations associated with current cancer treatments. Using a RF generator operating at 13.56 MHz, aqueous heating of purified C60-ser within the RF field has been observed in a concentration- and aggregation-dependent manner. These findings now lay the foundation for a new cancer hyperthermia therapy using C60 fullerene. Though the exact mechanism of C60-ser heating within the RF field is still uncertain, evidence has shown that the six serinol malonate (ser) groups of C60-ser must be attached to C60 to produce any significant heating. C60 is known to play numerous roles in the biomedical field, including drug delivery, active cancer cell targeting, bone therapy, gene therapy, X-ray contrast enhancement, and now, through the present work, as a potential new cancer therapy by RF-induced hyperthermia. By attaching a fluorescent tag (PF) to C60-ser to form C60-serPF, passive and active internalization into Hep3B and Huh7 liver cancer cells has been observed, with evidence of internalization within the cytoplasm and, more surprisingly, the nucleus. Very few nanomaterials have been definitively proven to internalize within the nucleus of cells, raising the potential for both C60-ser and C60-serPF to be used for other biomedical purposes and not just solely for RF-induced hyperthermia effects on cellular DNA. In vivo biodistribution studies of C60-serPF also has showed its presence in all major organs, including the brain. Since C60-ser was shown to inhibit the internalization of C60-serPF in vitro, uptake by liver cancer cells of both nanomaterials proceed by similar pathways. Therefore, the ability of C60-serPF to traverse the blood brain barrier of mice suggests an array of opportunities for brain-related treatments and therapies using both C60-serPF and C60-ser. Taken together, the materials provide simultaneous diagnostic and therapeutic capabilities, making them a new class of theranostic agent. In vitro studies of C60-ser have shown that it is non-toxic at up to 1.0 mg/mL to liver cancer cells (Hep3B, SNU449, and HepG2), that it modulates cellular metabolic activity, that it negligibly influences the cell cycle, but that it does not promote radiofrequency-induced hyperthermia of individual liver cancer cells. However, when injected intratumorally in mice, C60-ser enhanced the RF heating of liver tumors by 89% and 26% when compared to PBS and water controls alone, respectively. This observed increase in heating led to significant necrosis for the C60-ser treated groups, while necrosis for the control groups was nonexistent. This work has shown in vivo efficacy of an RF/C60-ser combination therapy by hyperthermia for the treatment of liver cancer for the first time. Future directions must involve the incorporation of biological targeting groups conjugated to the C60 moiety to realize the full potential of a noninvasive and targeted approach to cancer cell hyperthermia for liver cancer and other types of cancer, as well.