A mouse viral outgrowth assay for the detection of residual HIV-1 reservoirs

用于检测残留 HIV-1 病毒库的小鼠病毒生长测定

• 批准号: 9298573
• 负责人: JOEL N BLANKSON
• 金额: $ 40.5万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-08 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9298573
• 关键词: Adoptive Transfer; Allogeneic Bone Marrow Transplantation; Allogenic; Alpha Cell; Antigens; B-Lymphocytes; Berlin; Biological Assay; Bone Marrow; Bone Marrow Transplantation; Boston; CCR5 gene; CD28 gene; CD3 Antigens; CD4 Positive T Lymphocytes; Case Study; Cell Death; Cells; DNA; Data; Defective Viruses; Detection; Development; Flushing; Frequencies; Future; Gut associated lymphoid tissue; HIV; HIV-1; Hematologic Neoplasms; Hematopoietic; Highly Active Antiretroviral Therapy; Human; Immune response; Immunodeficient Mouse; Individual; Injectable; Interruption; Latent Virus; Lead; Malignant Neoplasms; Measures; Mediating; Methods; Mus; Mutation; Natural Killer Cells; Outcome; Patients; Peripheral; Peripheral Blood Mononuclear Cell; Primary Infection; Procedures; RNA; Regimen; Reporting; Residual state; Rest; Shock; Stem cell transplant; Study Subject; T-Lymphocyte; Testing; Therapeutic Intervention; Viral; Viral Antigens; Viral Cytopathogenic Effect; Viremia; Virus; Virus Latency; Work; antiretroviral therapy; base; experience; immune activation; in vivo; killings; novel; prevent; public health relevance; response; screening

 DESCRIPTION (provided by applicant): Latently infected CD4+ T cells represent the major barrier to HIV-1 eradication. Many strategies are being developed to eliminate these cells, but the field lacks a simple sensitive assay that can screen for these cells. PCR based assays cannot distinguish between replication-competent versus defective virus and recent studies have shown that the current state of the art culture assay measures only a small fraction of the replication-competent virus present. In this proposal we aim to develop a sensitive assay that is capable of selectively amplifying replication-competent virus from latently infected CD4+ T cells. Such an assay could inform the decision of whether or not to discontinue cART in patients after a therapeutic intervention is made. The need for such an assay is highlighted by the recent report of the 2 "Boston patients" who were treated with allogeneic stem cell transplantation for malignancies. No HIV DNA was detected by PCR in either patient from as many as 200 million PBMCs (<0.07 copies/106 PBMCs), but after cART was discontinued there was a rapid rebound in viremia in both patients. This outcome is undesirable is it negates the theoretical advantages to having smaller HIV-1 reservoirs. The development of a simple but sensitive assay that can screen a very large number of CD4+ T cells may help prevent the occurrence of a similar scenario from occurring in the future. This work will be important for the development of strategies for HIV-1 eradication since the assay will help inform the decision of whether or not to discontinue antiretroviral therapy.

 描述（由申请方提供）：潜伏感染的CD 4 + T细胞是HIV-1根除的主要障碍。目前正在开发许多策略来消除这些细胞，但该领域缺乏一种简单敏感的检测方法来筛选这些细胞。基于PCR的测定不能区分有复制能力的病毒与有缺陷的病毒，并且最近的研究表明，现有技术的培养测定仅测量存在的有复制能力的病毒的一小部分。在这个建议中，我们的目标是开发一种灵敏的检测，能够选择性扩增复制能力的病毒从潜伏感染的CD 4 + T细胞。这种测定可以告知在进行治疗干预后是否停止患者的cART的决定。最近报道的2例“波士顿患者”因恶性肿瘤接受异基因干细胞移植治疗，突出了对这种测定的需要。在两名患者中，通过PCR从多达2亿个PBMC中均未检测到HIV DNA（<0.07拷贝/106个PBMC），但在cART停止后，两名患者的病毒血症迅速反弹。这种结果是不可取的，因为它否定了具有较小HIV-1储库的理论优势。开发一种简单但灵敏的检测方法，可以筛选大量的CD 4 + T细胞，这可能有助于防止将来发生类似情况。 这项工作对于制定HIV-1根除战略非常重要，因为该检测将有助于决定是否 停止抗逆转录病毒治疗。

项目成果

The Effect of Latency Reversal Agents on Primary CD8+ T Cells: Implications for Shock and Kill Strategies for Human Immunodeficiency Virus Eradication.

• DOI: 10.1016/j.ebiom.2016.04.019
• 发表时间: 2016-06
• 期刊: EBioMedicine
• 影响因子: 11.1
• 作者: Walker-Sperling VE;Pohlmeyer CW;Tarwater PM;Blankson JN
• 通讯作者: Blankson JN

A CD3/CD28 microbead-based HIV-1 viral outgrowth assay.

基于 CD3/CD28 微珠的 HIV-1 病毒生长测定。

• 发表时间: 2017
• 期刊: Journal of virus eradication
• 影响因子: 5.5
• 作者: Kuzmichev,YuryV;Veenhuis,RebeccaT;Pohlmeyer,ChristopherW;Garliss,CarolineC;Walker-Sperling,VictoriaEk;Blankson,JoelN
• 通讯作者: Blankson,JoelN