STUDIES ON THE CONVERSION OF ANDROGENS TO ESTROGENS BY AROMATASE

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Aromatase (CYP19A1) is the enzyme responsible for the conversion of androgens to estrogens. This enzyme has received considerable attention over the last 30 years, with much of the recent effort directed at the development of inhibitors to be used in the treatment of estrogen dependent breast cancer. This laboratory is focusing on the mechanistic and regulatory aspects of aromatase. It has been shown that three molecules of molecular oxygen and six reducing equivalents from NADPH are consumed during estrogen formation. The androgen is first oxidized to the C19-hydroxyandrogen and then oxidized to the C19 gemdiol, or its dehydrated form, the C19 aldehyde. The mechanism of the third oxidation is still undefined, even though considerable information regarding the reaction has been established over the years. Experimental and theoretical studies are underway which investigate the mechanism of the third oxidation of aromatase and other recently described P450 mediated deformylation reactions. In addition to other regulatory mechanisms, aromatase activity may also be regulated by the levels of cytochrome P450 reductase present. We have shown that the relative amounts of estrogen intermediates is dependent on the aromatase:reductase ratio. We are using Chinese hamster ovary cells (CHO) that stably express the human aromatase gene. The influence of cytochrome P450 reductase or cytochrome b5 on rates of aromatase catalysis are investigated by transfection of CHO cells with vaccinia virus containing the appropriate gene constructs. This system enables us to develop a working model of aromatase regulation by variation in ratios of ancillary proteins.