HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) [Supplemental Data] All Versions of this Article: 78/6/1139    most recent biolreprod.107.062679v2 biolreprod.107.062679v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Google Scholar Articles by Tash, J. S. Articles by Georg, G. I. PubMed PubMed Citation Articles by Tash, J. S. Articles by Georg, G. I. Agricola Articles by Tash, J. S. Articles by Georg, G. I.

Gamendazole, an Orally Active Indazole Carboxylic Acid Male Contraceptive Agent, Targets HSP90AB1 (HSP90BETA) and EEF1A1 (eEF1A), and Stimulates Il1a Transcription in Rat Sertoli Cells¹

Department of Molecular and Integrative Physiology⁴ and Interdisciplinary Center for Male Contraceptive Research and Drug Development,⁵ University of Kansas Medical Center, Kansas City, Kansas 66160 Department of Medicinal Chemistry,⁶ University of Kansas, Lawrence, Kansas 66045-7582 Department of Molecular Genetics, Microbiology, and Immunology,⁷ UMDNJ Robert Wood Johnson Medical School, Piscataway, New Jersey 08854-5635

ABSTRACT

Gamendazole was recently identified as an orally active antispermatogenic compound with antifertility effects. The cellular mechanism(s) through which these effects occur and the molecular target(s) of gamendazole action are currently unknown. Gamendazole was recently designed as a potent orally active antispermatogenic male contraceptive agent. Here, we report the identification of binding targets and propose a testable mechanism of action for this antispermatogenic agent. Both HSP90AB1 (previously known as HSP90beta [heat shock 90-kDa protein 1, beta]) and EEF1A1 (previously known as eEF1A [eukaryotic translation elongation factor 1 alpha 1]) were identified as binding targets by biotinylated gamendazole (BT-GMZ) affinity purification from testis, Sertoli cells, and ID8 ovarian cancer cells; identification was confirmed by matrix-assisted laser desorption/ionization-time of flight mass spectrometry and Western blot analysis. BT-GMZ bound to purified yeast HSP82 (homologue to mammalian HSP90AB1) and EEF1A1, but not to TEF3 or HBS1, and was competed by unlabeled gamendazole. However, gamendazole did not inhibit nucleotide binding by EEF1A1. Gamendazole binding to purified Saccharomyces cerevisiae HSP82 inhibited luciferase refolding and was not competed by the HSP90 drugs geldanamycin or novobiocin analogue, KU-1. Gamendazole elicited degradation of the HSP90-dependent client proteins AKT1 and ERBB2 and had an antiproliferative effect in MCF-7 cells without inducing HSP90. These data suggest that gamendazole may represent a new class of selective HSP90AB1 and EEF1A1 inhibitors. Testis gene microarray analysis from gamendazole-treated rats showed a marked, rapid increase in three interleukin 1 genes and Nfkbia (NF-kappaB inhibitor alpha) 4 h after oral administration. A spike in II1a transcription was confirmed by RT-PCR in primary Sertoli cells 60 min after exposure to 100 nM gamendazole, demonstrating that Sertoli cells are a target. AKT1, NFKB, and interleukin 1 are known regulators of the Sertoli cell-spermatid junctional complexes. A current model for gamendazole action posits that this pathway links interaction with HSP90AB1 and EEF1A1 to the loss of spermatids and resulting infertility.

fertilization, male contraception, Sertoli cells, spermatogenesis, testis

³Current address: Department of Medicinal Chemistry, 717 Delaware Street SE, University of Minnesota, Minneapolis, MN 55414.

¹Supported by National Institutes of Health (NIH) U54 HD-055763 (to J.S.T.), NIH N01-HD-1–3313 (to G.I.G.), and U54 HD33994 Center for Reproductive Sciences, a National Institute of Child Health and Human Development (NICHD) Grant for Specialized Cooperative Centers Program in Reproductive Research (SCCPRR), at the University of Kansas Medical Center. Also supported by NICHD contract N01-HD-6–3259 awarded to BIOQUAL, Inc. Dedicated to the memory of Geoffrey M.H. Waites, Sc.D., Ph.D., and Thaddeus R.R. Mann, M.D., Ph.D., Sc.D., F.R.S.

Correspondence: ²FAX: 913 588 7180; e-mail, jtash{at}kumc.edu

This article has been cited by other articles:

				J. S. Tash, B. Attardi, S. A. Hild, R. Chakrasali, S. R. Jakkaraj, and G. I. Georg A Novel Potent Indazole Carboxylic Acid Derivative Blocks Spermatogenesis and Is Contraceptive in Rats after a Single Oral Dose Biol Reprod, June 1, 2008; 78(6): 1127 - 1138. [Abstract] [Full Text] [PDF]