NOVEL AZOLE-BASED HEME OXYGENASE INHIBITORS: IN VIVO CHARACTERIZATION AND THERAPEUTIC APPLICATION IN CANCER
Dercho, Ryan Albert
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Heme degradation is catalyzed through the action of heme oxygenases (HO), present as inducible (HO-1) and constitutive (HO-2) isoforms, resulting in the endogenous production of carbon monoxide (CO), biliverdin and iron. Numerous studies attempting to elucidate the physiological roles of these products have relied on metalloporphyrin inhibitors of HO; however, they have limited utility, as they are known to inhibit other hemoproteins. Previous work in our laboratory has led to the identification of a number of azole-based compounds that inhibit HO in vitro, including several that have demonstrated selectivity for HO-1. The principal goal of this project was to characterize lead compounds in vivo and investigate their therapeutic potential as anti-cancer agents. The first objective was to develop a method to screen novel azole-based HO inhibitors (abHOi) in vivo. To this end, we describe a non-invasive method of measuring CO exhalation (VeCO) as a measure of in vivo HO activity. Using the VeCO method, we found that the inhibition of HO by abHOi was dose- and time-dependent, and was comparable to the effectiveness of the classical metalloporphyrin HO inhibitor, zinc protoporphyrin. Moreover, abHOi inhibited HO by a reversible, non-competitive mechanism and had no effect on HO-1 or HO-2 protein expression. Interestingly, structurally similar azole antifungal drugs were found to inhibit HO activity at therapeutically relevant doses, which raises the possibility that some of the anti-mycotic and anti-tumourigenic effects of drugs like ketoconazole may be mediated, in part, by the inhibition of HO. Our last objective was to investigate the effectiveness of abHOi in in vitro and in vivo models of tumour angiogenesis, growth and metastasis. We found that the HO-1-selective abHOi, QC-15, decreased cancer cell viability and inhibited endothelial cell sprouting from aortic rings and capillary-like endothelial cell tube formation in vitro. In an orthotopic breast cancer model, treatment of tumour-bearing mice with QC-15 reduced primary tumour volume and lung metastasis, but not tumour angiogenesis. These results demonstrate that abHOi are valuable pharmacological tools in the elucidation of the physiological roles of HO and its products and, barring serious toxicity, promising therapeutic entities for the treatment of metastatic breast cancer.