Suppression of Spermatogenesis by Bisdichloroacetyldiamines Is Mediated by Inhibition of Testicular Retinoic Acid Biosynthesis

Abstract

The bisdichloroacetyldiamine WIN 18,446 reversibly inhibits spermatogenesis in many species, including humans; however, the mechanism by which WIN 18,446 functions is unknown. As retinoic acid is essential for spermatogenesis, we hypothesized that WIN 18,446 might inhibit retinoic acid biosynthesis from retinol (vitamin A) within the testes by inhibiting the enzyme aldehyde dehydrogenase 1a2 (ALDH1a2). We studied the effect of WIN 18,446 on ALDH1a2 enzyme activity in vitro, and on spermatogenesis and fertility in vivo, in mature male rabbits for 16 weeks. WIN 18,446 markedly inhibited ALDH1a2 enzyme activity in vitro with an IC50 of 0.3 μM. In vivo, the oral administration of 200 mg/kg WIN 18,446 to male rabbits for 16 weeks significantly reduced intratesticular concentrations of retinoic acid, severely impaired spermatogenesis, and caused infertility. Reduced concentrations of intratesticular retinoic acid were apparent after only 4 weeks of treatment and preceded the decrease in sperm counts and the loss of mature germ cells in tissue samples. Sperm counts and fertility recovered after treatment was discontinued. These findings demonstrate that bisdichloroacetyldiamines such as WIN 18,446 reversibly suppress spermatogenesis via inhibition of testicular retinoic acid biosynthesis by ALDH1a2. These findings suggest that ALDH1a2 is a promising target for the development of a reversible, nonhormonal male contraceptive.

Inhibition of Retinoic Acid Biosynthesis by the Bisdichloroacetyldiamine WIN 18,446 Markedly Suppresses Spermatogenesis and Alters Retinoid Metabolism in Mice

“…Analysis of Spermatogenesis-Using techniques reported previously (13), testes and epididymides were dissected, cleaned of fat, and processed for histologic and quantitative analysis of spermatogenesis. In each experiment, five mice per group were assigned for this analysis.…”

“…We have reported previously that WIN 18,446 strongly inhibits ALDH1A2 in vitro (13). It is known that WIN 18,446 also inhibits another aldehyde dehydrogenase, ALDH2 (the main enzyme involved in alcohol metabolism), causing the "disulfiram reaction" when a subject consumes alcohol while taking WIN 18,446 that is the basis for aversion therapy in treatment of alcoholism (14).…”

“…Previously, we have reported that the administration of the bisdichloroacetyldiamine WIN 18,446 abrogates spermatogenesis through its action as an inhibitor of aldehyde dehydroge-nase 1A2 (ALDH1A2 2 ; also known as RALDH2), one of the enzymes responsible for the oxidation of retinaldehyde to retinoic acid in tissues (13). The effect of WIN 18,446 on spermatogenesis is reversible in that cessation of drug administration results in complete recovery of spermatogenesis (13).…”

“…The effect of WIN 18,446 on spermatogenesis is reversible in that cessation of drug administration results in complete recovery of spermatogenesis (13). Although the reversible nature of WIN 18,446 inhibition of spermatogenesis is promising for its use as an oral contraceptive (14), its broad inhibitory effects on other isozymes of aldehyde dehydrogenase, such as ALDH2, the enzyme important for the metabolism of alcohol, prevents it from being used for such a purpose (13).…”

Metabolic Cooperation in Testis as a Pharmacological Target: From Disease to Contraception

“…Notably, the testis are enriched in several metaboliz-ing enzymes of vitamin A [184]. Besides the possible relevance of these enzymes [185], it has also been highlighted that signaling proteins involved in these processes are also possible relevant targets to develop a male contraceptive [186][187][188][189]. Nevertheless, research is still needed to disclose retinoic acid role on germ cells development, as well as its metabolism in testis and how can drugs specifically target testicular retinoic acid metabolism.…”