Targeting Na,K-ATPase alpha4 for male contraception

靶向 Na,K-ATPase α4 的男性避孕

• 批准号: 10618848
• 负责人: V GUSTAVO BLANCO
• 金额: $ 53.04万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-15 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10618848
• 关键词: ATPase inhibitory protein; Affect; Affinity; Animals; Antihistamines; Appearance; Area; Binding; Binding Proteins; Biochemical; Biological; Biological Availability; Biological Markers; Cell membrane; Cells; Characteristics; Chemicals; Clinical; Clinical Research; Contraceptive Agents; Contraceptive methods; Cytoplasm; Data; Drug Kinetics; Drug Targeting; Effectiveness; Ejaculation; Epididymis; Ethers; Female; Female genitalia; Fertility; Fertilization in Vitro; Future; Gland; Goals; Human; Hyperactivity; In Vitro; Infertility; Investigational Drugs; Ions; K ATPase; Knockout Mice; Lead; Male Contraceptive Agents; Male Infertility; Maximum Tolerated Dose; Meiosis; Membrane Potentials; Metabolic; Metabolism; Mus; Mutation; Oral; Oral Administration; Oral Contraceptives; Ouabain; Partner in relationship; Permeability; Pharmaceutical Chemistry; Pilot Projects; Plasma; Population; Property; Protein Isoforms; Proteins; Public Health; Research; Safety; Series; Site; Solubility; Source; Specificity; Sperm Capacitation; Sperm Motility; Sperm Tail; Spermatocytes; Spermatogenesis; Sterility; Surface; System; Testing; Testis; Toxic effect; Validation; Woman; absorption; analog; birth control; cell motility; druggable target; efficacy study; experimental study; extracellular; improved; in silico; in vivo; inhibitor; male; male fertility; men; molecular size; monolayer; novel; pharmacologic; pre-clinical; preclinical study; progenitor; programs; reproductive tract; reversible contraceptive; scaffold; scale up; side effect; small molecule; sperm cell; sperm function; unintended pregnancy; virtual screening

PROJECT SUMMARY / ABSTRACT The rapidly growing world population and the high rate of unintended pregnancies make contraception a need and a priority for any public health program. While several contraceptive methods, with varying efficacy are currently available for women, a more comprehensive approach to birth control requires extending contraception to males. However, a safe, effective and reversible contraceptive for men is still unavailable. An attractive approach to develop male contraceptives consists in targeting proteins that are specifically expressed in sperm and are required for sperm fertility. We have shown that Na,K-ATPase α4 (NKAα4), a plasma membrane ion transporter which exchanges cytoplasmic Na+ for extracellular K+, is a validated target for male contraception. NKAα4 is uniquely expressed in testis male germ cells after meiosis, is particularly abundant in the sperm flagellum, and is critical for sperm function. Deletion of NAKα4 in mice results in complete sterility of only the male but not the female animals. NKAα4 is essential for sperm motility and sperm capacitation. Its activity maintains sperm intracellular Na+ levels ([Na+]i) and several vital sperm parameters, including membrane potential (Vm), intracellular Ca+2 ([Ca2+]i) and pH. From a biochemical standpoint, NKAα4 has a particularly high affinity for ouabain, the specific inhibitor of Na,K-ATPase. We took advantage of this property to specifically target NKAα4 and block its function to achieve male infertility. We synthesized a series of small molecule compounds, which can selectively bind to the high ouabain affinity site of NKAα4. Some of these compounds inhibit NKAα4 and affect sperm motility both in vitro and after administration to mice. This provides strong evidence for the suitability of NKAα4 as a pharmacological target and our compounds as agents that can be used for the control of male fertility. However, before NKAα4 inhibitors can be moved forward into their application as male contraceptives, it is necessary that their efficacy, drug-target interaction, biomarkers for their in vitro and vivo specificity, side effects, mechanisms of action and pharmacokinetic parameters are identified and optimized for future clinical use. We will test this in two aims. In specific aim 1, we will develop compounds with the capacity to selectively inhibit NKAα4 and block sperm function, which will be ready for testing in mice. Then, during specific aim 2, we will perform studies to obtain preclinical validation to advance the NKAα4 inhibitors as male contraceptives. A series of rigorous approaches from the medicinal chemistry and biological areas will be used to identify the compounds which will have the characteristics that will be necessary for male contraception. This research will be essential to fulfill the highly desired unmet goal of obtaining a non-hormonal pharmacological agent that could be used as an oral, reversible agent for the control of male fertility.

项目总结/摘要 快速增长的世界人口和高意外怀孕率使避孕成为一项重要任务。 这是任何公共卫生计划的优先事项。虽然有几种避孕方法，效果各异， 目前妇女可获得的，更全面的节育方法需要扩大 男性避孕然而，仍然没有一种安全、有效和可逆的男子避孕方法。一个 开发男性避孕药的有吸引力的方法在于靶向蛋白质， 在精子中表达，并且是精子生育力所必需的。我们已经证明，Na，K-ATPase α4（NKAα4）， 质膜离子转运蛋白是一个有效的靶点，它将细胞质中的Na+交换为细胞外的K+ 用于男性避孕。NKAα4在减数分裂后的睾丸生殖细胞中特异表达， 在精子鞭毛中含量丰富，对精子功能至关重要。在小鼠中缺失NAKα4导致 只有雄性动物而不是雌性动物的完全不育。NKAα4对精子活力和精子 获能其活性维持精子细胞内Na+水平（[Na+]i）和几个重要的精子参数， 包括膜电位（Vm）、细胞内Ca ~（2+）（[Ca ~（2+）]i）和pH。从生化角度来看，NKAα4 对哇巴因具有特别高的亲和力，哇巴因是Na，K-ATP酶的特异性抑制剂。我们利用了这一点 特异性靶向NKAα4并阻断其功能以实现男性不育。我们合成了一系列 小分子化合物，可以选择性地结合到NKAα4的高哇巴因亲和力位点。一些 这些化合物抑制NKAα4，并在体外和对小鼠给药后影响精子活力。这 为NKAα4作为药理学靶点的适用性和我们的化合物作为 可用于控制男性生育力的药剂。然而，在NKAα4抑制剂可以移动之前， 在其作为男性避孕药的应用中，有必要研究其功效，药物-靶点相互作用， 生物标志物的体外和体内特异性、副作用、作用机制和药代动力学 识别并优化参数以用于将来的临床使用。我们将在两个目标中对此进行测试。在具体目标1中， 我们将开发具有选择性抑制NKAα4和阻断精子功能能力的化合物， 准备好在老鼠身上进行测试。然后，在具体目标2中，我们将进行研究以获得临床前验证 促进NKAα4抑制剂作为男性避孕药。一系列严格的方法，从医学 化学和生物学领域将用于鉴定具有以下特性的化合物， 是男性避孕所必需的这项研究将是必不可少的，以实现高度期望的未实现的目标 获得一种非激素药理学试剂，其可用作口服可逆试剂， 控制男性生育能力。

项目成果

Design, Synthesis, and in Vitro and in Vivo Evaluation of Ouabain Analogues as Potent and Selective Na,K-ATPase α4 Isoform Inhibitors for Male Contraception.

• DOI: 10.1021/acs.jmedchem.7b00925
• 发表时间: 2018-03-08
• 期刊: Journal of medicinal chemistry
• 影响因子: 7.3
• 作者: Syeda SS;Sánchez G;Hong KH;Hawkinson JE;Georg GI;Blanco G
• 通讯作者: Blanco G

Na,K-ATPase α4, and Not Na,K-ATPase α1, is the Main Contributor to Sperm Motility, But its High Ouabain Binding Affinity Site is Not Required for Male Fertility in Mice.

Na,K-ATPase α4（而不是 Na,K-ATPase α1）是精子活力的主要贡献者，但其高哇巴因结合亲和力位点并不是小鼠雄性生育能力所必需的。

• DOI: 10.1007/s00232-021-00181-2
• 发表时间: 2021
• 期刊: The Journal of membrane biology
• 作者: McDermott,JeffP;Sánchez,Gladis;Mitra,Amrita;Numata,September;Liu,LijunCatherine;Blanco,Gustavo
• 通讯作者: Blanco,Gustavo