Model of the Human Testis for Reproductive Toxicology

用于生殖毒理学的人类睾丸模型

• 批准号: 8201518
• 负责人: Constance M John
• 金额: $ 25万
• 依托单位: MANDALMED, INC.
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-18 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8201518
• 关键词: 3-Dimensional; Adult; Animals; Aromatic Polycyclic Hydrocarbons; Asbestos; Asthma; Automobile Driving; BAX gene; Biological Assay; Blood-Testis Barrier; Cadmium; Cancer Survivor; Cells; Chemical Exposure; Chemicals; Clinical Trials; Data; Development; Drug Industry; Environment; Environmental Exposure; Fee-for-Service Plans; Fertility; Fertility Study; Fetus; Fiber; General Population; Generations; Genes; Germ Cells; Goals; Growth; Haploid Cells; Haploidy; Human; Human Experimentation; In Vitro; Incidence; Industry; Infertility; Investigation; Lead; Male Infertility; Malignant Neoplasms; Malignant neoplasm of testis; Marketing; Mediating; Meiosis; Mercury; Methods; Microfluidics; Modeling; Outcome; Overdose; Pathway interactions; Pesticides; Pharmaceutical Preparations; Phase; Physiological; Poison; Process; RNA-Binding Proteins; Reference Standards; Reporting; Reproduction; Research; Research Personnel; Seminal fluid; Spermatogenesis; Staging; Stem cells; Structure of primordial sex cell; System; Temperature; Testing; Testis; Toxic effect; Toxicant exposure; Toxicity Tests; Toxicology; Toxin; Vinyl Chloride; aromatic hydrocarbon receptor; base; bisphenol A; cell type; clinically relevant; cost; cost effective; developmental disease; drug development; exposed human population; human disease; human embryonic stem cell; improved; in vitro Model; in vivo; induced pluripotent stem cell; interest; lead exposure; leydig interstitial cell; male; men; novel; overexpression; phase 2 study; polycarbonate plastic; prospective; reproductive; sertoli cell; shear stress; sperm cell; toxicant

DESCRIPTION (provided by applicant): More than 87,000 chemicals have been developed and distributed over the past 50 years. The vast majority of these have not been tested for potential toxic effects in humans or animals. Most systematic toxicology research is conducted in animals and is very expensive. Although animal studies contribute important information, they may not closely approximate the exposure or the exposure effects in humans. There is an unmet need for cost-effective bioassay systems that mimic and assess human exposures and their reproductive effects for systematic, prospective investigation of potentially toxic substances. The overall goal of the research is to develop a functional in vitro human cell-based testis model of spermatogenesis as a clinically relevant product for reproductive toxicology. Development of the first generation product is enabled by our capability to isolate and propagate primary Sertoli cells from adult human donors (hSertCs) and to manipulate primordial germ cells (PGCs), human embryonic stem cells (hESCs), and induced pluripotent stem cells (iPSCs) in vitro to produce haploid cells. The hSertCs have been available to researchers worldwide through a partnership with Lonza (Walkersville, MD) since May 2009, and have been used to create a model of the blood-testis barrier (BTB) in coated inserts of transwell plates. By silencing and overexpressing genes that encode germ-cell-specific cytoplasmic RNA-binding proteins, we have been able to modulate human germ-cell formation and developmental progression, to promote later stages of meiosis and development of haploid gametes. In Phase I, we aim to establish a model using primary human Sertoli, Leydig, and peritubular myoid cells to form a functional (BTB) in a closed hollow fiber system mimicking physiologic shear stress conditions, and to perform analytical and toxicological studies of the cells. There are three Specific Aims for Phase I. 1. Develop an in vitro human Sertoli cell-based, 3-dimensional (3-D) model of the testis. 2. Develop methods to detect and quantify the cells within the testis model. 3. Demonstrate the effect of reproductive toxins on the viability of human testicular cells. In Phase II, we intend to drive the differentiation of induced pluripotent stem cells (iPSCs) and spermatogonial stem cells (SSCs) to the haploid state within the model, and test the effect of toxins on this process. From this effort, we expect to identify reducible, reliable, and relevant endpoints for reproductive toxicant testing, and then to establish outcome markers and assays enabling for F.D.A. approval of the model in conjunction with other toxicology testing required for all New Drug Applications. The need for more relevant and improved reproductive toxicology testing represents a significant commercial opportunity, especially in the pharmaceutical industry. The bioassay product would be provided to industry on a fee-for-service basis initially. This effort potentially could lead to the capability to replicate normal human spermatogenesis in vitro and generate de novo mature sperm for severely infertile, azoospermic men, including cancer survivors. PUBLIC HEALTH RELEVANCE: More than 87,000 chemicals have been developed and distributed discarded over the past 50 years. The vast majority of these have not been tested for potential toxic effects in humans or animals. There is an unmet need for more cost-effective bioassay systems that will mimic and assess human exposures and their reproductive effects for systematic, prospective investigation into potentially toxic substances. The overall goal of the research is to develop a functional in vitro human cell-based testis model of spermatogenesis as a clinically relevant product for reproductive toxicology. Better identification of reproductive toxins could help protect developing fetuses from unwanted toxic exposures, reduce the incidence of male infertility, potentially lower rates of testis cancer, and create safer drugs for the marketplace. Also, part of our interest in developing a testis model is develop the capability to replicate normal human spermatogenesis in vitro to potentially generate de novo mature sperm for severely infertile, azoospermic men, including cancer survivors.

描述（由申请人提供）：在过去的50年中，已经开发和分发了87,000多种化学品。其中绝大多数尚未对人类或动物的潜在毒性作用进行测试。大多数系统的毒理学研究都是在动物中进行的，非常昂贵。尽管动物研究贡献了重要的信息，但它们可能不会紧密近似人类的暴露或暴露效应。对具有成本效益的生物测定系统的需求未满足，该系统模仿和评估人类暴露及其对系统的潜在有毒物质的前瞻性研究的生殖作用。该研究的总体目标是开发一种在体外人类细胞基于人类细胞的睾丸生成模型，作为生殖毒理学临床相关的产品。第一代产品的开发是通过我们从成年人类供体（HSERTC）和操纵原始生殖细胞（PGC），人类胚胎干细胞（HESC）和诱导多能干细胞（IPSC）中的原始生殖细胞（PGC）分离和传播原发性sertoli细胞并传播原代细胞（IPSC）中产生hapero haploid细胞的。自2009年5月以来，通过与Lonza（马里兰州Walkersville）的合作伙伴关系，HSERTC可以为全世界的研究人员提供，并被用来在Transwell Plates的涂层插入物中创建一个血式障碍物（BTB）模型。通过沉默和表达过表达的基因，这些基因编码细菌特异性的细胞质结合蛋白，我们能够调节人类种系的形成和发育进展，以促进减数分裂和单倍体配子的发育的后期阶段。在第一阶段，我们旨在使用原代人Sertoli，Leydig和周围肌动物细胞建立模型，以在模仿生理剪切应力条件的封闭空心纤维系统中形成功能（BTB），并对细胞进行分析和毒理学研究。第1期。1。开发睾丸的基于体外的人类Sertoli细胞的3维（3-D）模型。 2。开发检测和量化睾丸模型中细胞的方法。 3。证明生殖毒素对人睾丸细胞的生存能力的影响。在第二阶段，我们打算将诱导的多能干细胞（IPSC）和精子干细胞（SSC）驱动到模型中的单倍体状态，并测试毒素对此过程的影响。通过这项努力，我们希望确定可降低，可靠和相关的终点，以进行生殖有毒物质测试，然后确定为F.D.A.带来的结果标记和测定。该模型与所有新药应用所需的其他毒理学测试结合使用。需要更相关和改善的生殖毒理学测试是一个重要的商业机会，尤其是在制药行业。最初，生物测定产品将以付费服务提供给行业。这项努力可能会导致能力在体外复制正常的人类精子发生，并为包括癌症幸存者在内的严重不育，具有无菌的偶氮化男性产生从头成熟的精子。 公共卫生相关性：在过去50年中，已经开发和分发了87,000多种化学品。其中绝大多数尚未对人类或动物的潜在毒性作用进行测试。需要更具成本效益的生物测定系统的需求，这些系统将模仿和评估人类暴露及其生殖作用，以实施对潜在有毒物质的系统，前瞻性研究。该研究的总体目标是开发一种在体外人类细胞基于人类细胞的睾丸生成模型，作为生殖毒理学临床相关的产品。更好地识别生殖毒素可以帮助保护发育中的胎儿免受不必要的毒性暴露，降低男性不育症的发生，潜在地降低睾丸癌的发生率，并为市场创造更安全的药物。同样，我们对开发睾丸模型的兴趣的一部分是发展能够在体外复制正常的人类精子发生，以产生从头开始成熟的精子，以实现严重不育，无敏感的男性，包括癌症幸存者。