Fenbendazole as a Potential Anticancer Drug

Fenbendazole, [5-(phenylthio)-1H-benzimidazol-2-yl]carbamic acid methyl ester, is widely used to treat pinworms, other helminthes, and a variety of parasitic infections in laboratory animals, livestock, companion animals, and people (1–3). We became interested in fenbendazole when our university veterinarians recommended that all experimental rodents, including uninfected colonies such as ours, be treated with a fenbendazole-containing diet because of pinworm infections in some colonies. Our research uses tumors in rats and mice to evaluate the effects of new regimens for treating solid tumors with radiation and/or anticancer drugs (4–10). As we researched this proposed prophylactic treatment, we became concerned that the fenbendazole-containing chow might compromise our experiments, but also became intrigued by the possibility that this drug might have antineoplastic effects, by disrupting the tubulin microtubule equilibrium, or by altering the viability or radiosensitivity of cells in the hypoxic environments found in solid tumors. Fenbendazole acts on helminthes primarily by binding to tubulin and disrupting the tubulin microtubule equilibrium; its utility as an antiparasitic drug results from differences in the structures of tubulin in mammalian cells and in lower organisms, which lead to its greater binding to tubulin, and therefore greater inhibition of polymerization, in the parasites (11–13). In addition, the limited absorption of fenbendazole from the intestine results in low levels of the drug and its active metabolites in tissue relative to the levels within the gut, to which the targeted parasites are exposed (1, 14, 15). Several widely used anticancer drugs produce their antineoplastic effects by disrupting either microtubule formation (vincristine; vinblastine) or microtubule depolymerization (paclitaxel; docetaxel) (13, 16), suggesting that fenbendazole could have antitumor effects. Some data in the literature support this hypothesis (13). A Fenbendazole-containing diet combined with supplemental high dose of vitamins was reported by Gao et al. to inhibit growth of a human lymphoma xenografted into scid mice (17); it was unclear whether this reflected a direct effect of the drug on tumor cells or stimulation of host immune responses. Bai et al. reported that fenbendazole reduced the engraftment of brain tumors in nude mice (18). Chung et al. reported in a meeting presentation (19) that high doses of fenbendazole, albendazole and mebendazole inhibited the growth of paclitaxel-resistant tumors. Solid tumors develop regions of severe hypoxia very early in their development, at diameters of less than 1 mm (5, 6). As tumors grow, they concomitantly elicit the development of their vascular beds through angiogenesis, neovascularization, and co-option of vessels from normal tissues. However, tumor vascular beds lack the organization and regulation found in the vasculature supporting healthy normal tissues. The vessels are tortuous and irregular, and lack the musculature that normally regulates blood flow. Blind ends and shunting are common, and vessels frequently have microscopic and macroscopic holes that permit plasma or blood to leak into surrounding tissue. In addition, the growing tumors often invade or compress blood and lymphatic vessels, further compromising perfusion. As a result, solid tumors contain regions, where temporary interruptions in blood flow through individual vessels or persistent regional deficiencies in perfusion produce transient and chronic hypoxia. Because molecular oxygen is a potent chemical radiosensitizer (4–6, 20), hypoxic tumor cells are resistant to radiation, can survive radiation regimens that would eradicate fully aerobic cell populations, and can cause tumors to recur after radiation therapy. Intensive efforts in our laboratory and many others, have therefore been devoted to improving the outcome of radiation therapy by combining radiation with drugs that improve tumor oxygenation, selectively sensitize hypoxic cells to radiation, or preferentially kill hypoxic cells (4–9, 16, 20, 21). Several compounds related to fenbendazole have selective effects on hypoxic cells. Some nitroimidazoles and benzimidazoles act as hypoxic cell radiosensitizers, by replacing oxygen in the chemical reactions that lead to the production of DNA damage by ionizing radiation (4, 6, 20, 21). These drugs therefore increase the cytotoxic effects of ionizing radiation in hypoxia, while have no effect on the radiation response of aerobic cells (which are already maximally radiosensitive because of the oxygen levels in their environment). Several modified benzimidazoles synthesized by Gupta et al. were effective hypoxic cell radiosensitizers (21). Moreover, several series of substituted bis-benzimidazoles bind in the minor groove of DNA at specific DNA sequences, thereby inhibiting DNA helicase activity and cell proliferation (22, 23); some bis-benzimidazoles undergo bioreductive activation in hypoxic conditions (24), which could produce selective cytotoxicity toward hypoxic cells (5, 6, 24). Many benzimidazoles alter glucose uptake and carbohydrate metabolism (1, 3, 14), which could produce cytotoxic effects under the conditions of hypoxia, low pH, and nutritional inadequacy occurring in solid tumors. Because of these considerations, we tested for two possible effects of the fenbendazole-containing therapeutic rodent chow on our primary tumor model system: i) alteration of tumor growth, through a cytotoxic effect of the drug on cells within these solid tumors, and ii) alteration of tumor radiosensitivity (10). We saw no evidence that either effect occurred when our mice were fed the fenbendazole-containing diet, supporting the idea that this prophylactic intervention would not compromise our studies. However, cell culture data from that project also showed that fenbendazole given continuously at doses above those expected in the tissues of rodents fed the therapeutic diet, dramatically inhibited the growth of EMT6 tumor cells in vitro. This prompted us to extend our previously published studies to examine the effects of higher doses of fenbendazole on EMT6 tumor cells in vitro and EMT6 tumors in vivo and to more rigorously define the interactions of this drug with radiation. These studies are reported here.