Novel Retinoic Acid Receptor Alpha-Selective Antagonists

新型视黄酸受体α选择性拮抗剂

• 批准号: 9414227
• 负责人: DEBRA J. WOLGEMUTH
• 金额: $ 15.36万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9414227
• 关键词: Adverse effects; Aftercare; All-Trans-Retinol; Amides; Animals; Behavioral Research; Binding; Binding Sites; Bioavailable; Biological Assay; Biological Availability; Callithrix jacchus jacchus; Calorimetry; Carboxylic Acids; Collaborations; Contraceptive Agents; Contraceptive methods; Defect; Development; Diet; Dose; Drug usage; Exhibits; Failure; Fathers; Fertility; Fertility Study; Future; Gene Targeting; Genetic; Genetic study; Goals; Hormonal; Hormone Receptor; Hydrophobicity; In Vitro; Infertility; Lead; Luciferases; Male Contraceptive Agents; Male Sterility; Mass Spectrum Analysis; Metabolism; Modeling; Monkeys; Morphology; Mus; Nuclear; Nuclear Receptors; Oral; Oral Administration; Permeability; Pharmaceutical Preparations; Pharmacology; Pharmacotherapy; Phosphorylation; Physiological; Physiological Processes; Primates; Property; Protein Isoforms; Proteome; Pyrroles; Rattus; Receptor Signaling; Regimen; Reporter; Reporting; Reproductive Health; Research; Retinoic Acid Receptor; Retinoids; Scanning; Scientist; Signal Transduction; Spermatogenesis; Sterility; Surface Plasmon Resonance; Testing; Testis; Time; Titrations; Toxic effect; Toxicology; Treatment Efficacy; Tretinoin; Vitamin A; Withholding Treatment; Work; X-Ray Crystallography; base; design; drug candidate; in vitro activity; in vivo; male; men; mouse model; nonhuman primate; novel; pre-clinical trial; receptor; research and development; restoration; retinoic acid receptor alpha

PROJECT SUMMARY Animals deprived of vitamin A in their diet (VAD) exhibit male sterility. All trans retinoic acid (ATRA), an active metabolite of vitamin A, functions by binding to the nuclear retinoic acid receptors (RARs), of which there are three isoforms α, β, and γ. We and others have shown the importance of ATRA signaling via the RARα receptor in particular during spermatogenesis by gene targeting. Rara-/- mice are viable and healthy but the males are sterile, with defects in spermatogenesis similar to that seen in mice maintained on a VAD diet. Following up on observations of ‘testicular toxicity’ in rats treated with a pan-RAR antagonist, we have shown that indeed, spermatogenesis is inhibited in male mice after treatment with this orally bioavailable compound, and importantly, that the induced sterility is reversible. Our toxicology studies did not reveal any detectable side effects and normal progeny were sired after restoration of fertility. This suggests that antagonizing RAR signaling and RARα in particular, has potential as a novel and non-steroidal hormone-based approach to male contraception. The objective in this application is to identify retinoids with that selectively antagonize RAR α and to evaluate their therapeutic efficacy and reversibility. In Specific Aim 1, we propose to design, synthesize, and evaluate the in vitro activity, binding, and ADMET properties of RARα-selective antagonists. Binding studies will utilize isothermal titration calorimetry, differential scanning fluorimetry, surface plasmon resonance, microscale thermophoresis, and X-ray crystallography; activity studies will use a luciferase reporter assay; and pharmacological profiles will be assessed. Specific Aim 2 will then assess the in vivo effects of the most promising RARα-selective antagonists on spermatogenesis. After the initial morphological analysis, we will conduct fertility studies and develop dosing regimens for the most promising candidate molecules to determine the lowest doses and longest dosing periods at which efficient induction of sterility with complete reversibility could be achieved. Specific Aim 3 will then move to testing the lead candidate drug in a non-human primate, the common marmoset, model in a small scale trial to set the stage for future pre-clinical trials. Finally, we have observed in both our genetic and pharmacological models that one of the physiological processes in spermatogenesis that is extremely sensitive to altered retinoid signaling is spermiation. In Specific Aim 4, we therefore propose to determine both the quantitative global proteome and the quantitative phospho-proteome in particular during spermiation using dissected segments of testicular tubules and analysis by mass spectrometry.

项目摘要 饮食中剥夺了维生素A的动物（VAD）暴露于雄性不育。所有反式视网膜酸（ATRA），活性 维生素A的代谢产物，通过与核视黄酸受体（RARS）结合而起作用，其中存在 三种同工型α，β和γ。我们和其他人已经通过RARα受体显示了ATRA信号的重要性 特别是在基因靶向的精子发生过程中。 rara - / - 小鼠可行且健康，但男性是 无菌，精子发生中的缺陷类似于VAD饮食中维持的小鼠。跟进 在用泛射拮抗剂治疗的大鼠中观察到“睾丸毒性”，我们已经表明，确实 用这种口服生物利用化合物治疗后，雄性小鼠的精子发生受到抑制，重要的是 诱导的无菌性是可逆的。我们的毒理学研究没有发现任何可检测的副作用和正常 恢复生育能力后，后代被航行。这表明在拮抗RAR信号传导和RARα中 特别是作为一种新型和非甾体激素的男性违规方法具有潜力。 本应用中的目的是鉴定具有选择性拮抗RARα并评估其评估的类视黄素 治疗效率和可逆性。在特定目标1中，我们建议设计，合成和评估IN RARα选择性拮抗剂的体外活性，结合和ADMET特性。绑定研究将使用 等温滴定量热法，差分扫描荧光法，表面等离子体共振，显微镜 嗜热和X射线晶体学；活动研究将使用荧光素酶报告基因测定；和 将评估药理特征。然后，特定的目标2将评估最大的体内影响 有希望的RARα选择性拮抗剂对精子发生。经过最初的形态分析后，我们将 进行生育研究并为最有前途的候选分子开发剂量方案以确定 最低剂量和最长的剂量期，在该期间有效诱导不育具有完全可逆性 然后，特定的目标3将转移到非人类灵长类动物中测试铅候选药物， 常见的marmoset，在小型试验中为未来的临床前试验奠定了基础的模型。最后，我们有 在我们的遗传和药物模型中都观察到，这是一个物理过程之一 对改变类维生素的信号传导极为敏感的精子发生是精子。在特定的目标4中，我们 因此，建议确定定量全局蛋白质组和定量磷酸化蛋白质组 特别是在精子期间使用测试管的解剖片段和质谱分析。