Control of Oocyte Maturation

项目总结(见说明)： 这项研究的总体目的是开发一种基于配子的、非激素的女性避孕药。针对卵母细胞特异性的非激素避孕药，最实用的方法包括靶向磷酸二酯酶(PDE)调节的环磷酸腺苷(CAMP)代谢[M期促进因子(MPF)上游的减数分裂恢复的中央控制器]，或直接靶向MPF的激活剂或抑制物。卵泡内cAMP的主要调节因子是PDE3。环磷酸鸟苷(CGMP)[产生于卵泡的卵丘颗粒细胞，并通过缝隙连接转运到卵母细胞]是PDE3的内源性抑制物。PDE5是卵泡细胞中主要的cGMP降解酶，PDE9是卵母细胞中唯一的cGMP降解酶。卵母细胞特有的蛋白激酶WEE2是MPF的重要调节因子，是恢复减数分裂和退出减数分裂所必需的 受精后中期II。我们的研究计划是在非人类灵长类动物身上开发和测试这些靶标的抑制剂，并最终在猕猴身上进行避孕试验。具体目标是：(1)开发新的PDE和WEE2抑制剂，并确定选定的药物是否能扰乱猕猴卵母细胞的适时减数分裂成熟；(2)评估现有药物的药效学和药效学。 以及新的PDE抑制剂和WEE2抑制剂；以及(3)确定PDE和WEE2抑制剂是否可以在群体交配情况下定期循环的猕猴中作为避孕剂发挥作用。实验设计将包括表征药物与靶点的相互作用、合理的抑制剂设计、高通量筛选和药物化学(目标1)；通过孵化、体外受精、猕猴卵母细胞胞质内精子注射等方法测试候选药物的体外活性(目标1)；通过优化给药系统(如口服、阴道环、植入物)和监测非靶标效应，评估候选抑制剂在体内的药代动力学和药效学效应(目标2)；然后在社会饲养的雌性猕猴中进行避孕实验(目标3)。这种方法是 有望确定PDE和WEE2抑制剂作为女性避孕药的潜力。

PROJECT SUMMARY (See instructions): The overall purpose of this research is to develop a gamete-based, nonhormonal female contraceptive. The most practical approach to an oocyte-specific nonhormonal contraceptive involves targeting of the phosphodiesterase (PDE)-regulated metabolism of cyclic adenosine monophosphate (cAMP) [a central controller of meiotic resumption upstream of M-phase Promoting Factor (MPF)], or direct targeting of activators or inhibitors of MPF. The principal regulator of cAMP in the follicle is PDE3. Cyclic guanosine monophosphate (cGMP) [produced in cumulus granulosa cells of the follicle, and transported to the oocyte via gap junctions] is an endogenous inhibitor of PDE3. PDE5 is the principal cGMP-degrading enzyme in follicle cells, and PDE9 is the only cGMP-degrading enzyme in the oocyte. The oocyte-specific protein kinase WEE2 is an essential regulator of MPF necessary for resumption of meiosis and for exit from metaphase II following fertilization. Our research plan is to develop and test inhibitors of these targets in nonhuman primates and to ultimately perform a contraceptive trial in macaques. The Specific Aims are: (1) To develop novel PDE and WEE2 inhibitors and determine if selected agents can disrupt timely meiotic maturation of the macaque oocyte; (2) To evaluate the pharmacoklnetics and pharmacodynamics of existing and novel PDE inhibitors and WEE2 inhibitors; and (3) To determine whether PDE and WEE2 inhibitors can function as contraceptive agents in regularly cycling macaques in group-mating situations. Experimental designs will include characterization of drug-target interactions, rational inhibitor design, high throughput screening and medicinal chemistry (Aim 1); testing of candidate agents for activity in vitro using incubation, in vitro fertilization, and intracytoplasmic sperm injection of macaque oocytes (Aim 1); Assessment of pharmacokinetic and phamacodynamic effects of candidate inhibitors in macaques in vivo with optimization of drug delivery (e.g. oral, vaginal ring, implant) systems and monitoring of non-target effects (Aim 2); and then conducting a contraceptive experiment in socially-housed female macaques (Aim 3). This approach is expected to establish the potential for PDE and WEE2 inhibitors as contraceptive agents for women.