Pediatric Latent HIV Reservoir Characterization and Quantification Assay

项目摘要 公共卫生问题 现代抗逆转录病毒疗法（ART）组合大大提高了艾滋病毒感染者的生存率， 艾滋病毒传播。然而，全世界仍有3800万人感染艾滋病毒，其中1800万人感染了艾滋病毒。 2000万人是15岁以下的儿童，这突出表明迫切需要治疗性疗法。在有限 资源领域，其中儿科艾滋病毒是最常见的，使用新的分析，需要更少的血液体积， 低成本和低劳动强度的方法是必要的，以促进临床试验，旨在减少规模 和/或实现治愈或无ART缓解。 SBIR第一阶段项目的最终价值将证明我们的技术能够表征和 量化儿科HIV-1储库，这仍然是HIV治疗领域未满足的关键需求。 当前解决方案的问题以及产品如何满足未满足的需求 RNAamp允许直接检测RNA，这提供了对HIV复制的最准确测量。 合格储层。该检测方法被证明比HIV更快，更可靠，更定量 基于QVOA、RT-qPCR、RT-ddPCR、iSCA和IPDA的储库测定。RNA靶扩增和 检测是通过使用短的，序列特异性的， profluorophore和催化探针。重现性显著（重复测定之间的差异<4%）和限度 检测限（LoD）为1个HIV感染细胞/1 x 106个PBMC或1 - 10个RNA拷贝/mL，使用标准血样 （10 - 30 mL）。 Jan Biotech的RNAamp快速，直接RNA测试，使用少量血液样本，易于样品制备， 可靠性和灵敏度，定量能力和短时间的结果，非常适合表征和 定量的反弹能力的儿科艾滋病毒水库。 方法概述 Jan Biotech开发了一种检测方法，其治疗后反弹时间的预测准确度为0.92 早期/急性治疗成人受试者的中断。该测定法RNAamp利用非酶的化学方法， 扩增细胞相关的HIV RNA靶点，以提供快速、灵敏和定量的检测， 艾滋病病毒的储存库没有其他检测方法（包括QVOA、IPDA、iSCA、RT-qPCR或RT-ddPCR）提供预测 对于相同的成年参与者，准确度大于0.2。该建议是开发用于交叉的RNAamp- 子类型（即，非B亚型）HIV储库检测，专门用于儿科检测，为此使用了小 RNAamp易于测试的样品至关重要。儿童艾滋病毒感染者通常在早期接受治疗， 艾滋病毒感染，往往在出生时治疗，医生监测或自发中断治疗， 在这个人群中很常见儿科治疗有效性和治疗中断临床试验通常包括 非B亚型HIV感染者。 合作伙伴和独特资源 Jan Biotech，Inc.，具有分子诊断开发方面的专业知识，将咨询Jonathan Li，MD，MMSC 医学助理教授，哈佛医学院，在艾滋病临床试验组（ACTG）的研究和 Harris Gelbard，医学博士，哲学博士，罗切斯特大学医学院。 第一阶段具体目标 具体目标1：开发HIV-1交叉亚型（CS）RNAamp多重检测试剂盒 具体目的2：使用非B亚型HIV感染细胞将CS-RNAamp与Pol RT-qPCR相关联 具体目标3：使用CS-RNAamp HIV储库表征进行儿科人体样本检测 二期建成后的市场 吉利德和默克都对这项技术感兴趣。将寻求监管批准， 商业化

PROJECT SUMMARY ABSTRACT Public Health Problem Modern antiretroviral therapy (ART) combinations greatly improve survival of HIV infected individuals and reduce HIV transmission. However, there are still 38 million people living with HIV (PWH) worldwide and of those 1.8 million are children under the age of 15, highlighting the imperative need for curative therapies. In limited resource areas, where pediatric HIV is most common, the use of new assays that require less blood volumes, lower cost and less labor-intensive methods are necessary to facilitate clinical trials aimed at reducing the size of the latent reservoir and/or to achieve a cure or ART-free remission. The ultimate value of the SBIR Phase I project will demonstrate the ability of our technology to characterize and quantify the pediatric HIV-1 reservoir, which remains a critical unmet need in the HIV cure field. Issues with Current Solutions & How Product Meets Unmet Needs RNAamp allows direct detection of RNA, which provides the most accurate measure of the HIV replication competent reservoir. The assay is proven to be faster and have better reliability and quantification than HIV reservoir assays based on QVOA, RT-qPCR, RT-ddPCR, iSCA and IPDA. RNA target amplification and detection is accomplished by oligonucleotide-templated photoreduction using short, sequence-specific profluorophore and catalytic probes. Reproducibility is outstanding (4% variance between replicates) and Limit of Detection (LoD) is 1 HIV-infected cells/1 x 106 PBMCs or 1-10 RNA copies/mL, using a standard blood draw (10-30 mL). Jan Biotech’s RNAamp quick, direct RNA test, with its use of small blood samples, ease of sample preparation, reliability and sensitivity, quantitative ability and short time to result, is ideally suited for characterization and quantification of the rebound-competent pediatric HIV reservoir. Summary of Approach Jan Biotech has developed an assay with a prediction accuracy of 0.92 for time to rebound after treatment interruption for early/acute-treated adult participants. The assay, RNAamp, utilizes nonezymatic, chemical amplification of cell-associated HIV RNA targets to provide quick, sensitive, and quantitative measurement of the HIV reservoir. No other assay, including QVOA, IPDA, iSCA, RT-qPCR, or RT-ddPCR, provided a prediction accuracy of greater than 0.2 for the same adult participants. This proposal is to develop RNAamp for cross- subtype (i.e., non-B subtype) HIV reservoir detection, specifically for pediatric testing, for which the use of small samples easily tested by RNAamp is crucial. Pediatric HIV-infected individuals are typically treated during early HIV infection, often treated at birth, and physician-monitored or spontaneous treatment interruption is more frequent in this population. Pediatric treatment efficacy and treatment interruption clinical trials typically include non-B subtype HIV-infected participants. Collaborators and Unique Resources Jan Biotech, Inc., with expertise in molecular diagnostic development, will consult with Jonathan Li, MD, MMSc Assistant Professor of Medicine, Harvard Medical School, in a AIDS Clinical Trials Group (ACTG) study and Harris Gelbard, MD, PhD, University of Rochester Medical School. Phase I Specific Aims Specific Aim 1: Develop HIV-1 cross-subtype (CS) RNAamp multiplexed assay Specific Aim 2: Correlate CS-RNAamp to Pol RT-qPCR using non-B subtype HIV-infected cells Specific Aim 3: Pediatric human sample testing using CS-RNAamp HIV reservoir characterization Market after Phase II Completion Both Gilead and Merck are commercially interested in the technology. Regulatory approval will be sought for commercialization.