Aromatase inhibitors: past, present and future.

For the cellular physiology of sex steroid sensitive cells, the androgen/estrogen ratio may be more important than only one hormone action per se, in both sexes. This ratio is controlled in vertebrates by aromatase; its gene expression can be inhibited in different ways, and this is crucial for the treatment of estrogen-dependent diseases such as breast cancer, or gynecomastia in males for instance. To reach this goal, new steroidal and non-steroidal inhibitors are continuously being developed, and some of them are used as first or second line agents. Aromatase inhibition is also an essential tool for studying the role of estrogens in the adult, or during development. Aromatase inhibitors have shown in particular that estrogens are essential also in males for skeletal maturation and bone mineralization, development of masculine dendritic morphology in male brain linked to mating behaviour, and testicular function. Testosterone is often the prohormone converted in situ in active estrogens, at these levels. Several strategies can be used for aromatase inhibition. The first ones employed were blind screening or deductions from in vivo observations, which led for instance to the discovery of the role of aminoglutethimide in aromatase inhibition. Subsequently, in the years 1975-1990, the molecular modeling of compounds to mimic the substrate shape of the enzyme constituted the major idea. Hundreds of chemicals were synthesized by numerous authors, ranging from the well-known and very efficient 4-OHA to complicated imidazole or indane derivatives tested by sophisticated comparative molecular field analyses. Reticulum-bound active aromatase has not as yet been X-ray analyzed. Thus, aromatase inhibitors were also used more recently to probe and understand the active site conformation of the enzyme and its modelization was obtained from comparisons with bacterial-related cytochromes. We developed a mammalian model considerably closer to human aromatase in order to study the active site shape with new potent aromatase non-steroidal inhibitors. This model is equine aromatase. This enzyme was biochemically characterized, purified, and cloned by our group. It allowed testing, by site-directed mutagenesis, predictive hypotheses in human aromatase which contributed to designing of new inhibitors. The understanding of the functioning of an essential member of the cytochrome P450 family, which is necessary for cellular detoxification, was also facilitated. Inhibition of aromatase activity has also been carried out with antibodies directed to the catalytic site and at the gene level by knock-out or by control of factor-specific promoters. This may result in different mRNA synthesized by alternative splicing. We have also obtained specific inhibition of aromatase activity in human cells with antisense stable phosphorothioate oligodeoxynucleotides directed against aromatase mRNA tertiary structures. Besides known steroidal and non-steroidal inhibitors, the antiaromatase effects of compounds found in our daily environment such as dietary flavonoids or xenobiotic pollutants have also been described. Finally, we underline that all these aromatase inhibitors, or methods of aromatase inhibition, can modulate the estrogenic balance essential not only for female, but also for male physiology, including gonadal function.