Simple Method for Screening of HIV Drug Resistance in Resource-Limited Settings

在资源有限的环境中筛查 HIV 耐药性的简单方法

基本信息

批准号：
10384759
负责人：
Mario Stevenson
金额：
$ 30万
依托单位：
DISCIDIUM BIOSCIENCES, LLC
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-08-17 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10384759
关键词：
Address Alleles Anti-Retroviral Agents Antiretroviral drug resistance Antiretroviral resistance Biological Assay Blood Caring Clinical Cold Chains Complementary DNA Data Detection Development Encapsulated False Negative Reactions Fluorescence Fluorescent Probes Freeze Drying Genes Genetic Polymorphism Genome Genotype Goals Gold HIV HIV Infections HIV drug resistance HIV resistance HIV therapy HIV-1 HIV-1 drug resistance HIV-1 protease Health Personnel Human immunodeficiency virus test Individual Infection Infrastructure Interruption Label Laboratories Lamivudine Light Methods Modification Mutation Nucleosides Peptide Hydrolases Performance Persons Pharmaceutical Preparations Phase Plasma Point Mutation Polymerase Polymers RNA RNA purification RNA-Directed DNA Polymerase Reaction Reagent Recommendation Regimen Research Priority Resistance Resource-limited setting Reverse Transcription Sampling Screening procedure Source Testing Treatment outcome Tube United States National Institutes of Health Viral Visualization Vulnerable Populations Whole Blood base commercialization cost cost effective design efavirenz improved inhibitor low and middle-income countries mutant non-nucleoside reverse transcriptase inhibitors novel strategies particle reconstitution resistance mutation resistance to lamivudine screening tool treatment optimization viral RNA

项目摘要

Abstract Drug resistance to antiretroviral therapy for HIV infection is a serious clinical problem without cost-effective solutions in low and middle-income countries (LMICs) where standard genotypic resistance assays are unaffordable. Discidium's goal is to develop a single tube assay that uses lyophilized reactants, standard PCR and in-tube readout, that can be used as a screening resistance test and in laboratories with minimal infrastructure. We have exploited a Taq polymerase that absolutely requires a terminal 3´ base match that, with appropriate primers matched to resistance-associated polymorphisms, forms the basis for a low-cost allele- specific PCR resistance assay. This assay reveals the presence of first line resistance mutations to the most common NRTI that underscore resistance in the vast majority of individuals failing therapy in LMICs. We have also developed a fluorescence approach that allows direct visualization of PCR products under a blue light source. As such, the assay is single-step in that amplification and readout occur in the same tube. Our preliminary data supports the use of this assay to target the most common mutations that confer resistance to ARTs (K103N for NNRTIs and M184V for NRTIs), and we propose to optimize the use of this assay in unprocessed blood, and rigorously validate the assay performance using HIV-1 harboring the K103N and M184V mutations. This assay has the potential to be developed into a complete kit that health care workers can use to perform HIV-1 resistance testing in LMICs to guide optimal management of HIV-1-infected individuals.

抽象的 HIV感染的抗逆转录病毒治疗的耐药性是一个严重的临床问题，没有成本效益低收入和中等收入国家 (LMIC) 的解决方案，这些国家采用标准基因型耐药性测定 Discidium 的目标是开发一种使用冻干反应物、标准 PCR 的单管检测方法。和管内读数，可用作筛选电阻测试，并在实验室中使用我们开发了一种 Taq 聚合酶，它绝对需要一个末端 3' 碱基匹配，与抗性相关多态性相匹配的适当引物，构成了低成本等位基因的基础特异性 PCR 抗性检测该检测揭示了大多数一线抗性突变的存在。常见的 NRTI 强调了中低收入国家绝大多数治疗失败的个体的抵抗力。还开发了一种荧光方法，可以在蓝光下直接观察 PCR 产物因此，该测定是单步的，扩增和读出发生在同一管中。初步数据支持使用该测定来针对最常见的耐药突变 ART（NNRTI 为 K103N，NRTI 为 M184V），我们建议优化该测定在未经处理的血液，并使用含有 K103N 和的 HIV-1 严格验证检测性能 M184V 突变有可能被开发成供医护人员使用的完整试剂盒。可用于在中低收入国家进行 HIV-1 耐药性检测，以指导 HIV-1 感染者的最佳管理个人。