Quick and Accurate Measurements of HIV Broadly Neutralizing Antibody Susceptibility

快速准确地测量 HIV 广泛中和抗体敏感性

基本信息

批准号：
10676422
负责人：
Zizhang Sheng
金额：
$ 100.24万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-04-01 至 2026-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10676422
关键词：
AIDS clinical trial group Address Affect Algorithms Antibodies Area Under Curve Bar Codes Biological Assay Clinical Clinical Trials Cloning Computer Models Consumption DNA DNA analysis Data Detection Development Dideoxy Chain Termination DNA Sequencing Enrollment Exposure to Genetic Genome Genomic DNA Genotype Goals HIV HIV Envelope Protein gp120 HIV Infections HIV Vaccine Trials Network HIV envelope protein Human IgG1 Individual Infection Knowledge Laboratories Length Libraries Link Literature Luciferases Measurement Measures Medical center Minor Minority Neutralization Tests Participant Patients Performance Peripheral Blood Mononuclear Cell Persons Phenotype Predisposition Procedures Property Provirus Integration Research Personnel Resistance Running Sampling Site Societies System Technology Testing Third Generation Sequencing Time Training Universities Validation Variant Viral Viral reservoir Virion Virus antiretroviral therapy assay development biobank clinical development clinical trial enrollment early phase clinical trial env Genes experimental study improved machine learning model multiplex assay nanopore neutralizing antibody novel participant enrollment point of care portability prediction algorithm prevent public health relevance research clinical testing viral RNA

项目摘要

PROJECT SUMMARY HIV-infected individuals harbor viral quasispecies that differ in genetic sequences and susceptibility to broadly neutralizing antibodies (bNAbs) targeting the HIV envelope (Env). Due to difficulties with current assays in confirming viral sensitivity to bNAbs, most clinical trials enrolled participants without knowledge of their viral susceptibility. Thus, an accurate and sensitive assay with quick turnaround (1-2 weeks) is needed. We propose to approach this aim with two prongs: Aim 1, a phenotypic (Env function) assay for bulk Env neutralization assessment and Aim 2, a genotypic (Env sequence) algorithm for high-throughput prediction of neutralization susceptibility. We propose 10 HIV bNAbs targeting 5 distinct gp120 sites for assay development, with 2 non-HIV human IgG1 antibodies as negative controls. Current assays are difficult because of two labor-intensive and time-consuming procedures: Env single-genome amplification (SGA) and individual Env cloning. Aim 1 will address the latter procedure required because the current TZM-bl neutralization assay cannot tease out bulk Env clones that differ in bNAb susceptibility. Aim 1 will apply DNA barcoding to simultaneously track bNAb sensitivity of hundreds of Env variants in a high-throughput manner. This will be achieved by a novel Env- expressing lentiviral system in which each Env is linked to a unique barcode sequence. Aim 2 will address Env sequencing, for which SGA is necessary because Sanger sequencing cannot sequence bulk templates. We propose in Aim 2 to apply a third-generation sequencing technology, Nanopore sequencing, for effective bulk template sequencing and develop machine learning models for a fast and reliable prediction algorithm. Combining with the ample bNAb neutralization data from literature and Aim 1 of this project, trained computational models can be fast, accurate, and high-throughput to predict bNAb susceptibility. Since the detection of a low yield of functional Env sequences and minority variants from participant’s HIV DNA reservoir is essential, we propose in Aim 3 to investigate whether a virion capture assay could be applied to enrich functional Envs. We will also apply a DNA analysis to enrich minor variants from the viral reservoir. We have biospecimens stored onsite and access to a wide distribution of HIV subtypes from the AIDS Clinical Trials Group (ACTG) and HIV Vaccine Trials Network (HVTN) biorepositories for assay development and validation. All proposed R61 Aims have clear milestones defined and the R61 assays will be developed with streamlined and standard operating procedure (SOP) for adaptation to clinical labs. The Columbia Clinical Trials Support Laboratory is fully equipped and staffed to perform the SOPs and provide input and validation on assay performance. Successful development of an accurate and sensitive bNAb susceptibility assay with quick turnaround will alter the enrollment for bNAb trials and greatly improve clinical development of bNAb products.

项目概要 HIV 感染者具有病毒准种，其基因序列和对广泛病毒的易感性不同。由于目前的检测方法存在困难，因此针对 HIV 包膜 (Env) 的中和抗体 (bNAb)。为了确认病毒对 bNAb 的敏感性，大多数临床试验在不了解其病毒情况的情况下招募了参与者因此，我们建议进行快速周转（1-2 周）的准确、灵敏的检测。通过两个方面来实现这一目标：目标 1，用于批量 Env 中和的表型（Env 功能）测定评估和目标 2，一种用于高通量中和预测的基因型（Env 序列）算法我们建议针对 5 个不同 gp120 位点的 10 种 HIV bNAb 进行检测开发，其中 2 种为非 HIV 人类 IgG1 抗体作为阴性对照，目前的测定很困难，因为两个劳动密集型和耗时的程序：Env 单基因组扩增 (SGA) 和单独的 Env 克隆 Aim 1 将。解决后一个所需的程序，因为当前的 TZM-bl 中和测定无法梳理出批量 bNAb 敏感性不同的 Env 克隆将应用 DNA 条形码来同时追踪 bNAb。以高通量方式检测数百种 Env 变体的敏感性这将通过一种新颖的 Env- 来实现。表达慢病毒系统，其中每个 Env 都链接到一个独特的条形码序列 Aim 2 将寻址 Env。测序，为此 SGA 是必需的，因为桑格测序无法对批量模板进行测序。在目标 2 中建议应用第三代测序技术，纳米孔测序，以实现有效批量模板排序并开发机器学习模型，以实现快速可靠的预测算法。结合文献中充足的 bNAb 中和数据和本项目的目标 1，训练计算模型可以快速、准确、高通量地预测 bNAb 敏感性。从参与者的 HIV DNA 库中检测到低产量的功能性 Env 序列和少数变异至关重要，我们在目标 3 中建议研究是否可以应用病毒颗粒捕获测定来富集我们还将应用 DNA 分析来丰富我们拥有的病毒库中的微小变异。现场储存的生物样本，并可从艾滋病临床试验组获得广泛分布的艾滋病毒亚型 (ACTG) 和 HIV 疫苗试验网络 (HVTN) 生物样本库用于检测开发和验证。拟议的 R61 目标具有明确的里程碑，并且 R61 检测将通过简化和简化来开发用于适应临床实验室的标准操作程序 (SOP)。实验室设备齐全，人员配备齐全，可以执行 SOP 并提供分析输入和验证成功开发出准确、灵敏、快速的 bNAb 药敏测定方法。转变将改变 bNAb 试验的招募情况，并极大地改善 bNAb 产品的临床开发。