An Ultrasensitive In Vivo Latent HIV Viral Outgrowth Assay Using Humanized Mice

使用人源化小鼠进行超灵敏体内潜伏 HIV 病毒生长测定

• 批准号: 9277371
• 负责人: Ramesh Akkina
• 金额: $ 68.32万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-30 至 2020-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9277371
• 关键词: Adoptive Transfer; Aftercare; Allogenic; Animals; Anti-HIV Therapy; Area; Berlin; Biological Assay; Birth; Boston; CD4 Positive T Lymphocytes; Cells; Child; Coculture Techniques; Colorado; Development; Goals; Growth; HIV; HIV Infections; HIV-1; In Vitro; Infection; Interruption; Latent Virus; Macaca; Methods; Mississippi; Mus; Names; Patients; Peripheral; Pharmaceutical Preparations; Plasma Cells; Residual state; Rest; SIV; Sampling; System; Testing; Tissues; Universities; Viral; Viral reservoir; Virus; Virus Latency; Work; antiretroviral therapy; base; experience; experimental study; humanized mouse; improved; in vivo; innovation; novel; optimism; predicting response; public health relevance; viral rebound

 DESCRIPTION (provided by applicant): ABSTRACT: The fully cured "Berlin patient" example instilled a lot of optimism in the HIV field and the current thrust is to forge ahead with developing novel HIV-1 eradication strategies. In this context, it is essential that reliable tests are developed that can verify significant reductins in inducible viral reservoirs and predict responses to curative strategies after antiretroviral therapy (ART) interruptions (ATI). Other than ATI, viral out growth assays (VOA) that screen for latently infected cells by co-culturing methods have been critical to quantifying inducible or replication competent residual virus that may persist after a curative strategy. However, in some patients with prolonged undetectable plasma and cell-associated HIV-1 such as the two "Boston patients" who underwent allogeneic HSCT and the "Mississippi child" who had undergone early ART right after birth, traditional VOAs could not detect residual virus in peripheral CD4 T cells although virus rebounded after treatment interruption. Thus, it has become important that more sensitive VOAs that employ novel and innovative systems such as HIV susceptible humanized mice (hu-mice) need to be developed and validated. In a recent development, latent virus was successfully recovered from fully virus suppressed SIV infected macaques (as determined by all standard tests) undergoing intensive ART by adoptive transfer of their CD4 T cells to naive animals. This showed that ultralow levels of otherwise undetectable latently infected cells could be captured and induced using an in vivo system. Our current proposal is based on these recent findings and the hypothesis that an in vivo humanized mouse viral outgrowth assay (hmVOA) will be more sensitive than traditional in vitro VOAs at detecting low levels of persistent infectin in HIV patients. Our specific aims are to: 1. Develop and optimize an in vivo humanized-mouse viral outgrowth assay (hmVOA) for latent HIV using patient derived resting CD4 T cells, 2. Compare hmVOA with traditional VOA using patient derived resting CD4 T cells and determine if hmVOA is more sensitive, 3. Determine whether hmVOA can detect persistent HIV infection in patients in which virus is undetectable by any other method, including VOA, 4. Evaluate if hmVOA can more readily detect virus-latent cells from tissue reservoirs/ sanctuaries of long-term fully suppressed patients' compared to standard VOA, 5. Determine if sensitivity of hmVOA can be further improved by in vivo treatment of cell infused mice with HIV latency-reversing agents such as romidepsin and panobinostat.

 描述（由申请人提供）： 摘要：完全治愈的“柏林病人”的例子给艾滋病毒领域注入了很多乐观情绪，目前的重点是继续开发新的艾滋病毒1根除战略。在这种情况下，至关重要的是，开发可靠的测试，可以验证诱导型病毒库的显着减少，并预测抗逆转录病毒治疗（ART）中断（ATI）后对治疗策略的反应。除ATI外，通过共培养方法筛选潜伏感染细胞的病毒外生长测定（VOA）对于定量治疗策略后可能持续存在的可诱导或可复制的残留病毒至关重要。然而，在一些长期检测不到血浆和细胞相关HIV-1的患者中，例如接受同种异体HSCT的两名“波士顿患者”和出生后立即接受早期ART的“密西西比儿童”，传统VOA无法检测到外周CD 4 T细胞中的残留病毒，尽管病毒在治疗中断后反弹。因此，需要开发和验证采用新颖和创新系统（如HIV易感人源化小鼠（hu-mice））的更灵敏的VOA，这一点变得非常重要。在最近的发展中，通过将其CD 4 T细胞过继转移至幼稚动物，成功地从经历强化ART的完全病毒抑制的SIV感染的猕猴（如通过所有标准测试所确定的）中回收潜伏病毒。这表明可以使用体内系统捕获和诱导超低水平的原本不可检测的潜伏感染细胞。我们目前的建议是基于这些最新的研究结果和假设，在体内人源化小鼠病毒生长试验（hmVOA）将比传统的体外VOA检测低水平的持续感染HIV患者更敏感。我们的具体目标是：1.使用患者来源的静息CD 4 T细胞开发并优化潜伏HIV的体内人源化小鼠病毒生长测定（hmVOA），2.使用患者来源的静息CD 4 T细胞将hmVOA与传统VOA进行比较，并确定hmVOA是否更敏感。确定hmVOA是否可以检测到持续的HIV感染的患者中的病毒是无法检测到的任何其他方法，包括VOA，4。评价与标准VOA相比，hmVOA是否可以更容易地从长期完全抑制患者的组织储库/庇护所中检测病毒潜伏细胞，5。确定hmVOA的敏感性是否可以通过用HIV潜伏期逆转剂如罗米地辛和帕比司他体内治疗细胞输注小鼠来进一步提高。