Received for publication February 23, 2006.
Revised June 26, 2006.
Accepted for publication June 26, 2006.

Inhibition of the enzymatic activity of heme oxygenases by azole-based antifungal drugs

Abstract

Ketoconazole (KTZ) and other azole antifungal agents are known to have a variety of actions beyond the inhibition of sterol synthesis in fungi. These drugs share structural features with a series of novel heme oxygenase (HO) inhibitors designed in our laboratory. Accordingly, we hypothesized that therapeutically used azole-based antifungal drugs are effective HO inhibitors. Using gas chromatography to quantify carbon monoxide (CO) formation in vitro and in vivo, we have shown that azole-containing antifungal drugs are potent HO inhibitors. Terconazole, sulconazole and ketoconazole (KTZ) were the most potent drugs with IC₅₀ values of 0.41 ± 0.01 µM, 1.1 ± 0.4 µM and 0.3 ±0.1 µM for rat spleen microsomal HO activity respectively. Kinetic characterization revealed that KTZ was a non-competitive HO inhibitor. In the presence of KTZ (2.5 µM and 10 µM), Km values for both rat spleen and brain microsomal HO were not altered but a significant decrease in the catalytic capacity (V_max) was observed (P < 0.005). KTZ was also found to weakly inhibit nitric oxide synthase with an IC₅₀ of 177 ± 2 µM, but had no effect on the enzymatic activity of NADPH cytochrome P450 reductase. Because these drugs were effective within the concentration range observed in humans, it is possible that inhibition of HO may play a role in some of the pharmacological actions of these antimycotic drugs.

Key words: antifungal, azoles, enzyme inhibition, heme oxygenase, ketoconazole, terconazole