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Type 1-programmed dendritic cells drive antigen-specific latency reversal and immune elimination of persistent HIV-1.

ABSTRACT: BACKGROUND:Despite the success of antiretroviral therapy (ART), latent HIV-1 continues to persist in a long-lived population of resting memory CD4+ T cells within those who are infected. Finding a safe and effective means to induce latency reversal (LR) during ART to specifically expose this latent HIV-1 cellular reservoir for immune elimination has been a major barrier to a functional cure. METHODS:In this study, we test the use of antigen-presenting type 1-polarized, monocyte-derived dendritic cells (MDC1) generated from chronic HIV-1-infected individuals on ART as a means to induce HIV-1 latency reversal in autologous CD4+ T cells harboring replication-competent provirus. We use the same MDC1 for ex-vivo generation of autologous HIV-1 antigen-specific CD8+ cytotoxic T cells (CTL) and test their effector responses against the MDC1-exposed HIV-1- infected CD4+ T cell targets. FINDINGS:MDC1 presentation of either HIV-1 or cytomegalovirus (CMV) antigens to CD4+ T cells facilitated HIV-1 LR. This antigen-driven MDC1-mediated LR was sharply diminished with blockade of the CD40L/CD40 'helper' signaling pathway. Importantly, these antigen-presenting MDC1 also activated the expansion of CTL capable of killing the exposed HIV-1-infected targets. INTERPRETATION:Inclusion of virus-associated MHC class II 'helper' antigens in MDC1-based HIV-1 immunotherapies could serve both as a targeted means to safely unmask antigen-specific CD4+ T cells harboring HIV-1, and to support CTL responses that can effectively target the MDC1-exposed HIV-1 cellular reservoir as a functional cure strategy. FUND: This study was supported by the NIH-NAID grants R21-AI131763, U01-AI35041, UM1-AI126603, and T32-AI065380.

SUBMITTER: Kristoff J

PROVIDER: S-EPMC6557749 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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Type 1-programmed dendritic cells drive antigen-specific latency reversal and immune elimination of persistent HIV-1.

Kristoff Jan J Palma Mariana L ML Garcia-Bates Tatiana M TM Shen Chengli C Sluis-Cremer Nicolas N Gupta Phalguni P Rinaldo Charles R CR Mailliard Robbie B RB

EBioMedicine 20190402

<h4>Background</h4>Despite the success of antiretroviral therapy (ART), latent HIV-1 continues to persist in a long-lived population of resting memory CD4<sup>+</sup> T cells within those who are infected. Finding a safe and effective means to induce latency reversal (LR) during ART to specifically expose this latent HIV-1 cellular reservoir for immune elimination has been a major barrier to a functional cure.<h4>Methods</h4>In this study, we test the use of antigen-presenting type 1-polarized, ...[more]

PMID: 30952614

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Project description:Quantification of cell associated HIV RNA (ca-RNA) is one of the most important and commonly used methods to evaluate the performance of latency-reversing agents (LRAs). Copies of HIV RNA measured by qPCR, are often normalized to the input RNA or cell number. However, these could be affected by biological variability and/or technical errors, which can be avoided by using an internal reference gene. To obtain reliable data, it is essential to select stable reference genes (RGs) of which the expression is not influenced by biological variability, the type of cells, or the LRAs used. However, to date, no study has carefully evaluated RG stability following LRA exposure. We analyzed the stability of six widely used RGs (GAPDH, TBP, YWHAZ, UBE2D2, HPRT1 and RPL27A) in human PBMC and CD4+ T cells. LRA exposure significantly influenced the stability of these RGs. Overall, TBP, UBE2D2, and RPL27A were the most stable RGs in all tested conditions. TBP was generally the most stable RG whereas GAPDH varied the most. Finally, we evaluated the impact of applying different RG normalizers to host genes and HIV ca-RNA data. Altered results were observed both in host and HIV gene expression when unstable RGs were used. Our data underline the importance of testing the stability of RGs utilized to evaluate LRA-induced HIV ca-RNA expression. To our knowledge, this is the first careful evaluation of the stability of RGs after LRA exposure and will significantly contribute to the quality of data analysis in regard to gene expression.IMPORTANCELatency-reversing agents (LRAs) are ubiquitously used in the "shock-and-kill" HIV cure strategy and their performance is often evaluated by ex-vivo quantification of cell associated HIV RNA. HIV RNA, measured by qPCR, is often normalized to internal reference genes, but the expression of these genes should not be influenced by the experimental settings. We found that treatment of human PBMC and CD4+ T cells with LRAs significantly altered the expression of several commonly used reference genes, such as GAPDH. Finally, we evaluate the impact of different reference genes on normalization of host genes and HIV cell associated RNA expression and demonstrated that using unstable reference genes dramatically altered experimental outcome. Our data highlight the importance of using reference genes that are unaffected by LRAs under study to correctly evaluate host gene and cell associated HIV RNA expression induced by latency-reversing agents.

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Type 1-programmed dendritic cells drive antigen-specific latency reversal and immune elimination of persistent HIV-1.

Publications

Type 1-programmed dendritic cells drive antigen-specific latency reversal and immune elimination of persistent HIV-1.

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