Drug Transport at the Blood-Testis Barrier

血睾屏障的药物转运

基本信息

批准号：
8490705
负责人：
Nathan J Cherrington
金额：
$ 35.25万
依托单位：
UNIVERSITY OF ARIZONA
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-06-17 至 2015-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8490705
关键词：
Acquired Immunodeficiency Syndrome Activities of Daily Living Anti-Retroviral Agents Basement membrane Biological Blood - brain barrier anatomy Blood-Testis Barrier Carrier Proteins Cessation of life Characteristics Chemicals Development Didanosine Diffusion Disease Drug Delivery Systems Drug Exposure Drug Transport Ensure Exclusion Germ Cells HIV Highly Active Antiretroviral Therapy Human Immunohistochemistry Indinavir Lopinavir Measures Methods Molecular Movement Mus Organ P-Glycoprotein Patients Penetration Permeability Persons Pharmaceutical Preparations Phenotype Physiological Property Protease Inhibitor Regulation Relative (related person)Reproduction Resistance Rest Risk Role Seminal fluid Seminiferous tubule structure Site Specificity Surface System Tenofovir Testing Testis Therapeutic Tight Junctions United States Viral Load result Virus Wild Type Mouse Zidovudine abacavir antiretroviral therapy base basolateral membrane carrier mediated transport design emtricitabine inhibitor/antagonist luminal membrane men non-nucleoside reverse transcriptase inhibitors novel nucleoside analog public health relevance research study response sertoli cell small hairpin RNA transmission process uptake

项目摘要

DESCRIPTION (provided by applicant): Since its introduction in 1996, highly active antiretroviral therapy (HAART) has been credited with a marked and sustained reduction in AIDS-related death and disease in the United States. As with any pharmacotherapeutic agent, drug delivery to the site of action is absolutely required to elicit a favorable response. A significant obstacle to complete eradication of the HIV virus from the patient's body is the existence of biological barriers creating sanctuaries where the virus remains free from drug exposure. Two such barriers exist, ensuring the virus' survival and possible development of a resistant phenotype: the blood- brain barrier (BBB) and the blood-testis barrier (BTB). While there are no current methods that fully recapitulate the BBB to study the transport of drugs, we have developed a novel means of studying the intact BTB. The BTB protects the developing germ cells from the effects of potentially disruptive chemical interactions as well as immunological influences. Although this normal, protective function of the BTB is physiologically beneficial for reproduction, it can also provide a sanctuary for the HIV virus during antiretroviral therapy. This sanctuary site for HIV is particularly significant because the presence of HIV virus in the ejaculate of infected men is the major means of transmission to uninfected persons. Therefore, even while a patient is being effectively treated with a cocktail of antiretroviral drugs, the selective exclusion of the drugs at the BTB increases the viral load of the ejaculate and potential for transmission. Indeed, protease inhibitors and non-nucleoside reverse transcriptase inhibitors are actively excluded at the BTB and do not reach therapeutic concentrations in the testis. Alternatively, nucleoside analog drugs (NSAs) offer a unique opportunity to examine the characteristics of the BTB as well as the possibility of utilizing endogenous transporters as a means of circumventing the BTB since a number of NSAs are capable of accumulating in the semen of treated men. As the major component of the blood-testis barrier, Sertoli cells separate the adluminal compartment of the seminiferous tubules (STs) from the rest of the body by the formation of tight junctions below developing germ cells. In order to reach the adluminal space, drugs must first avoid barrier-function transporters before subsequently passing through Sertoli cells either by passive diffusion or carrier-mediated transport. Inward-facing transporter proteins, located at the basolateral and luminal membranes, allow specific compounds that cannot pass by passive diffusion to cross the BTB via transepithelial secretion. Selective movement of compounds across Sertoli cells therefore comprises the physiologic or functional aspect of the BTB. Therefore, we hypothesize that specific transporters are present at the basolateral and luminal membranes of Sertoli cells that allow specific NSA drugs to penetrate the BTB. The following aims have been designed to test this hypothesis: 1. Determine the constitutive expression and cellular localization of the major drug transporter proteins at the blood-testis barrier, as well as penetration of NSAs and PIs into the testis. 2. Determine the ability of selected inward-facing transporters (particularly Ent1, Ent2, Mate1 and Mate2) to support transepithelial secretion of NSAs and PIs. 3. Determine the functional capacity and molecular specificity of isolated Sertoli cells and intact STs to transport NSAs (didanosine, azidothymidine, abacavir, emtricitabine, tenofovir) and PIs (lopinavir and indinavir).The major emphasis of the current proposal is to focus on mechanisms by which therapeutics can successfully utilize endogenous transepithelial transporters to accumulate in the adluminal compartment, even within the context of the functional physiologic barrier.

描述（由申请人提供）：自1996年引入以来，高度活跃的抗逆转录病毒疗法（HAART）已被认为是美国与艾滋病相关的死亡和疾病的明显且持续减少的。与任何药物治疗剂一样，绝对需要向行动现场提供药物以引起有利的反应。从患者体内完全消除HIV病毒的一个重要障碍是存在生物屏障的存在，从而产生了避难所，使该病毒仍然免于药物暴露。存在两个这样的障碍，可确保病毒的生存和抗性表型的可能发展：血液脑屏障（BBB）和血液潮湿屏障（BTB）。尽管目前没有完全概括BBB来研究药物运输的方法，但我们已经开发了一种研究完整BTB的新方法。 BTB保护发育中的生殖细胞免受潜在破坏性化学相互作用以及免疫学影响的影响。尽管BTB的这种正常的保护功能在生理上对繁殖有益，但在抗逆转录病毒疗法期间，它也可以为HIV病毒提供庇护所。这种艾滋病毒的保护区尤其重要，因为在感染男性的射精中存在艾滋病毒病毒是向未感染的人传播的主要手段。因此，即使患者正在用抗逆转录病毒药物的鸡尾酒有效治疗，在BTB处的药物的选择性排除也会增加射精的病毒载荷和传播潜力。实际上，蛋白酶抑制剂和非核苷逆转录酶抑制剂在BTB处被积极排除，并且在睾丸中未达到治疗浓度。另外，核苷类模拟药物（NSAS）为检查BTB的特征以及利用内源性转运蛋白作为围绕BTB的一种手段而提供了独特的机会，因为许多NSA能够在处理过男性精液中积聚。作为血液尾屏障的主要组成部分，Sertoli细胞通过形成在发育中的生殖细胞下方的紧密连接处的形成，将精神小管（STS）的腹腔隔室与身体的其余部分分开。为了到达adluminal空间，在随后通过被动扩散或载体介导的转运穿过Sertoli细胞之前，必须首先避免使用屏障功能转运蛋白。位于基底外侧和腔内膜上的朝向转运蛋白蛋白允许无法通过被动扩散通过的特定化合物通过transepithelial分泌通过BTB。因此，化合物跨Sertoli细胞的选择性运动包括BTB的生理或功能方面。因此，我们假设特定的转运蛋白存在于Sertoli细胞的基底外侧和腔膜上，这些转运蛋白允许特定的NSA药物穿透BTB。以下目的是为了检验这一假设的设计：1。确定主要药物转运蛋白在血液测testis屏障上的组成型表达和细胞定位，以及NSA和PIS和PIS渗透到睾丸中。 2。确定选定的朝内转运蛋白（尤其是ENT1，ENT2，MATE1和MATE2）支持NSA和PIS的跨层分泌的能力。 3.确定孤立的塞托利细胞和完整的STS的功能能力和分子特异性运输NSA（Didanosine，Azidothymidine，abacavir，Emtricitabine，Tenofovir）和PIS（Lopinavir和Indinavir）。即使在功能生理屏障的背景下，转运蛋白也会积聚在辅助室中。