TZA: a novel assay for measuring the latent HIV-1 reservoir

Gupta, Phalguni; Sanyal, Anwesha; Mailliard, Robbie B.

doi:10.1080/14737159.2017.1384315

Cited by 5 publications

(6 citation statements)

References 34 publications

(47 reference statements)

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“…3f ). TZM-bl cells stably expressing CD4, CCR5, CXCR4, and carrying a β-galactosidase gene under the control of HIV-1 long terminal repeat promoter have been used for more sensitive detection of replication-competent HIV-1 than traditional virus outgrowth assays or quantitative RT-PCR 49 , 50 . By this highly sensitive TZM-bl cell-based galactosidase reporter analysis (TZA) assay, we detected infectious HIV-1 in the spleen and bone marrow cells of mouse 1222 but not mouse 1216 (Fig.…”

Section: Resultsmentioning

confidence: 99%

Clearance of HIV infection by selective elimination of host cells capable of producing HIV

Li¹,

Liu²,

Bauch³

et al. 2020

Nat Commun

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The RNA genome of the human immunodeficiency virus (HIV) is reverse-transcribed into DNA and integrated into the host genome, resulting in latent infections that are difficult to clear. Here we show an approach to eradicate HIV infections by selective elimination of host cells harboring replication-competent HIV (SECH), which includes viral reactivation, induction of cell death, inhibition of autophagy and the blocking of new infections. Viral reactivation triggers cell death specifically in HIV-1-infected T cells, which is promoted by agents that induce apoptosis and inhibit autophagy. SECH treatments can clear HIV-1 in >50% mice reconstituted with a human immune system, as demonstrated by the lack of viral rebound after withdrawal of treatments, and by adoptive transfer of treated lymphocytes into uninfected humanized mice. Moreover, SECH clears HIV-1 in blood samples from HIV-1-infected patients. Our results suggest a strategy to eradicate HIV infections by selectively eliminating host cells capable of producing HIV.

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Section: Resultsmentioning

confidence: 99%

Clearance of HIV infection by selective elimination of host cells capable of producing HIV

Li¹,

Liu²,

Bauch³

et al. 2020

Nat Commun

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“…HIV-1 replication competent reservoir in resting CD4 + T cells. Human splenocytes from human spleen organoids of individual mock or ART-treated chronic HIV-1 infected humanized mice with undetectable viral load (<10 copies per ml) were used in the TZA method (40,72). Human resting CD4 + T cells were stimulated with CD3/28 to reactivate the HIV reservoir, and infectious units per million cells (IUPM) was determined following previously published methods.…”

Section: Methodsmentioning

confidence: 99%

“…Withdrawal of ART in viral load-suppressed chronic HIV-infected BLTS humanized mice resulted in viral load rebound within 2 weeks, with viral load levels reaching set point levels within 8 weeks after ART withdrawal ( Figure 5B). For analysis of replication-competent HIV in the BLTS humanized mouse model, we utilize the TZM-bl cell line, which stably expresses CD4, CCR5, and CXCR4 and carries an integrated copy of the β-galactosidase (β-gal) gene under control of an HIV-1 long terminal repeat (LTR) promoter, to quantify inducible replication competent HIV-1 in an assay, termed TZA (40,41). We demonstrated the presence of replication-competent HIV in human spleen organoid-derived human resting CD4 + T cells (~5% [~2.5 × 10 6 to 4 × 10 6 ] of total splenocytes [~50 × 10 6 to 80 × 10 6 ]) from ART-treated, HIV-infected BLTS humanized mice ( Figure 5C).…”

Section: Figure 2 Lymphoid Tissues Development In the Blts Humanizedmentioning

confidence: 99%

Human immunodeficiency virus infection induces lymphoid fibrosis in the BM-liver-thymus-spleen humanized mouse model

Samal¹,

Kelly²,

Na-Shatal³

et al. 2018

JCI Insight

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A major pathogenic feature associated with HIV infection is lymphoid fibrosis, which persists during antiretroviral therapy (ART). Lymphoid tissues play critical roles in the generation of antigen-specific immune response, and fibrosis disrupts the stromal network of lymphoid tissues, resulting in impaired immune cell trafficking and function, as well as immunodeficiency. Developing an animal model for investigating the impact of HIV infection-induced lymphoid tissue fibrosis on immunodeficiency and immune cell impairment is critical for therapeutics development and clinical translation. Said model will enable in vivo mechanistic studies, thus complementing the well-established surrogate model of SIV infection-induced lymphoid tissue fibrosis in macaques. We developed a potentially novel human immune system-humanized mouse model by coengrafting autologous fetal thymus, spleen, and liver organoids under the kidney capsule, along with i.v. injection of autologous fetal liver-derived hematopoietic stem cells, thus termed the BM-liver-thymus-spleen (BLTS) humanized mouse model. BLTS humanized mouse model supports development of human immune cells and human lymphoid organoids (human thymus and spleen organoids). HIV infection in BLTS humanized mice results in progressive fibrosis in human lymphoid tissues, which was associated with immunodeficiency in the lymphoid tissues, and lymphoid tissue fibrosis persists during ART, thus recapitulating clinical outcomes.

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“…Using a TZM‐bl cells‐based virus outgrowth assay (VOA), 59,60 we observed that the mice with no HIV‐1 rebound after SECH withdrawal in the blood were also negative for HIV‐1 in the spleen and bone marrow cells 32 . We also used an in vivo humanized mouse‐based VOA (hmVOA)—an extremely rigorous assay for measuring the clearance of the virus 16,61,62 .…”

Section: Elimination Of Hiv‐1‐infected T Cells In Vitro By Viral Reac...mentioning

confidence: 99%

Clearance of HIV-1 or SIV reservoirs by promotion of apoptosis and inhibition of autophagy: Targeting intracellular molecules in cure-directed strategies

Chen

Budai

et al. 2022

Journal of Leukocyte Biology

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The reservoirs of the HIV display cellular properties resembling long‐lived immune memory cells that could be exploited for viral clearance. Our interest in developing a cure for HIV stems from the studies of immunologic memory against infections. We and others have found that long‐lived immune memory cells employ prosurvival autophagy and antiapoptotic mechanisms to protect their longevity. Here, we describe the rationale for the development of an approach to clear HIV‐1 by selective elimination of host cells harboring replication‐competent HIV (SECH). While reactivation of HIV‐1 in the host cells with latency reversing agents (LRAs) induces viral gene expression leading to cell death, LRAs also simultaneously up‐regulate prosurvival antiapoptotic molecules and autophagy. Mechanistically, transcription factors that promote HIV‐1 LTR‐directed gene expression, such as NF‐κB, AP‐1, and Hif‐1α, can also enhance the expression of cellular genes essential for cell survival and metabolic regulation, including Bcl‐xL, Mcl‐1, and autophagy genes. In the SECH approach, we inhibit the prosurvival antiapoptotic molecules and autophagy induced by LRAs, thereby allowing maximum killing of host cells by the induced HIV‐1 proteins. SECH treatments cleared HIV‐1 infections in humanized mice in vivo and in HIV‐1 patient PBMCs ex vivo. SECH also cleared infections by the SIV in rhesus macaque PBMCs ex vivo. Research efforts are underway to improve the efficacy and safety of SECH and to facilitate the development of SECH as a therapeutic approach for treating people with HIV.

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TZA: a novel assay for measuring the latent HIV-1 reservoir

Cited by 5 publications

References 34 publications

Clearance of HIV infection by selective elimination of host cells capable of producing HIV

Clearance of HIV infection by selective elimination of host cells capable of producing HIV

Human immunodeficiency virus infection induces lymphoid fibrosis in the BM-liver-thymus-spleen humanized mouse model

Clearance of HIV-1 or SIV reservoirs by promotion of apoptosis and inhibition of autophagy: Targeting intracellular molecules in cure-directed strategies

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