scholarly journals Potentiation of drug toxicity through virus latency reversal promotes preferential elimination of HIV infected cells

2022 ◽  
Author(s):  
Tung Truong ◽  
Manuel Hayn ◽  
Camilla Kaas Frich ◽  
Lucy Kate Ladefoged ◽  
Morten Jarlstad Olesen ◽  
...  
Keyword(s):  
Cell Culture ◽  
Drug Toxicity ◽  
Hiv Protease ◽  
Lead Compound ◽  
Mixed Cell ◽  
Virus Latency ◽  
C Terminus ◽  
Mixed Cell Population ◽  
Latently Infected ◽  
Infected Cells

Eliminating latently infected cells is a highly challenging, indispensable step towards the overall cure for HIV/AIDS. We recognized that the unique HIV protease cut site (Phe-Pro) can be reconstructed using a potent toxin, monomethyl auristatin F (MMAF), which features Phe at its C-terminus. We hypothesized that this presents opportunities to design prodrugs that are specifically activated by the HIV protease. To investigate this, a series of MMAF derivatives was synthesized and evaluated in cell culture using latently HIV-infected cells. Cytotoxicity of compounds was enhanced upon latency reversal by up to 11-fold. In a mixed cell population, nanomolar concentrations of the lead compound depleted predominantly the HIV-infected cells and in doing so markedly enriched the pool with the uninfected cells. Despite expectation, mechanism of action of the synthesized toxins was not as HIV protease-specific prodrugs, but likely through the synergy of toxicities between the toxin and the reactivated virus.

scholarly journals Induction of Autophagy to Achieve a Human Immunodeficiency Virus Type 1 Cure

Cells ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1798
Author(s):  
Grant R. Campbell ◽  
Stephen A. Spector
Keyword(s):  
Human Immunodeficiency Virus ◽  
Antiretroviral Therapy ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Functional Cure ◽  
Immunodeficiency Virus ◽  
Latently Infected ◽  
Infected Cells ◽  
Hiv 1

Effective antiretroviral therapy has led to significant human immunodeficiency virus type 1 (HIV-1) suppression and improvement in immune function. However, the persistence of integrated proviral DNA in latently infected reservoir cells, which drive viral rebound post-interruption of antiretroviral therapy, remains the major roadblock to a cure. Therefore, the targeted elimination or permanent silencing of this latently infected reservoir is a major focus of HIV-1 research. The most studied approach in the development of a cure is the activation of HIV-1 expression to expose latently infected cells for immune clearance while inducing HIV-1 cytotoxicity—the “kick and kill” approach. However, the complex and highly heterogeneous nature of the latent reservoir, combined with the failure of clinical trials to reduce the reservoir size casts doubt on the feasibility of this approach. This concern that total elimination of HIV-1 from the body may not be possible has led to increased emphasis on a “functional cure” where the virus remains but is unable to reactivate which presents the challenge of permanently silencing transcription of HIV-1 for prolonged drug-free remission—a “block and lock” approach. In this review, we discuss the interaction of HIV-1 and autophagy, and the exploitation of autophagy to kill selectively HIV-1 latently infected cells as part of a cure strategy. The cure strategy proposed has the advantage of significantly decreasing the size of the HIV-1 reservoir that can contribute to a functional cure and when optimised has the potential to eradicate completely HIV-1.

scholarly journals Targeting the latent human cytomegalovirus reservoir for T-cell-mediated killing with virus-specific nanobodies

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Timo W. M. De Groof ◽  
Elizabeth G. Elder ◽  
Eleanor Y. Lim ◽  
Raimond Heukers ◽  
Nick D. Bergkamp ◽  
...  
Keyword(s):  
Gene Expression ◽  
Human Cytomegalovirus ◽  
Early Gene ◽  
Latent Reservoir ◽  
Solid Organ ◽  
Cell Transplant ◽  
Viral Receptor ◽  
Transplant Patients ◽  
Latently Infected ◽  
Infected Cells

AbstractLatent human cytomegalovirus (HCMV) infection is characterized by limited gene expression, making latent HCMV infections refractory to current treatments targeting viral replication. However, reactivation of latent HCMV in immunosuppressed solid organ and stem cell transplant patients often results in morbidity. Here, we report the killing of latently infected cells via a virus-specific nanobody (VUN100bv) that partially inhibits signaling of the viral receptor US28. VUN100bv reactivates immediate early gene expression in latently infected cells without inducing virus production. This allows recognition and killing of latently infected monocytes by autologous cytotoxic T lymphocytes from HCMV-seropositive individuals, which could serve as a therapy to reduce the HCMV latent reservoir of transplant patients.

scholarly journals The Paramyxovirus Simian Virus 5 V Protein Slows Progression of the Cell Cycle

2000 ◽  
Vol 74 (19) ◽  
pp. 9152-9166 ◽  
Author(s):  
Grace Y. Lin ◽  
Robert A. Lamb
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Simian Virus ◽  
S Phase ◽  
V Protein ◽  
Simian Virus 5 ◽  
C Terminus ◽  
M Phase ◽  
Infected Cells ◽  
Affect Cell Cycle Progression

ABSTRACT Infection of cells by many viruses affects the cell division cycle of the host cell to favor viral replication. We examined the ability of the paramyxovirus simian parainfluenza virus 5 (SV5) to affect cell cycle progression, and we found that SV5 slows the rate of proliferation of HeLa T4 cells. The SV5-infected cells had a delayed transition from G1 to S phase and prolonged progression through S phase, and some of the infected cells were arrested in G2 or M phase. The levels of p53 and p21CIP1were not increased in SV5-infected cells compared to mock-infected cells, suggesting that the changes in the cell cycle occur through a p53-independent mechanism. However, the phosphorylation of the retinoblastoma protein (pRB) was delayed and prolonged in SV5-infected cells. The changes in the cell cycle were also observed in cells expressing the SV5 V protein but not in the cells expressing the SV5 P protein or the V protein lacking its unique C terminus (VΔC). The unique C terminus of the V protein of SV5 was shown previously to interact with DDB1, which is the 127-kDa subunit of the multifunctional damage-specific DNA-binding protein (DDB) heterodimer. The coexpression of DDB1 with V can partially restore the changes in the cell cycle caused by expression of the V protein.

Galectin-3 is involved in HIV-1 expression through NF-κB activation and associated with Tat in latently infected cells

2019 ◽  
Vol 260 ◽  
pp. 86-93 ◽  
Author(s):  
Mika Okamoto ◽  
Akemi Hidaka ◽  
Masaaki Toyama ◽  
Masanori Baba
Keyword(s):  
Latently Infected ◽  
Galectin 3 ◽  
Infected Cells ◽  
Hiv 1

scholarly journals The Early Gene hhi1 Reactivates Heliothis zea Nudivirus 1 in Latently Infected Cells

2009 ◽  
Vol 84 (2) ◽  
pp. 1057-1065 ◽  
Author(s):  
Yueh-Lung Wu ◽  
Carol P. Wu ◽  
Song-Tay Lee ◽  
Han Tang ◽  
Chi-Hua Chang ◽  
...  
Keyword(s):  
Viral Infection ◽  
Critical Role ◽  
Heliothis Zea ◽  
Virus Titer ◽  
Viral Reactivation ◽  
Early Gene ◽  
Mechanistic Study ◽  
Early Genes ◽  
Latently Infected ◽  
Infected Cells

ABSTRACT Heliothis zea nudivirus 1 (HzNV-1), previously known as Hz-1 virus, is an insect virus able to establish both productive and latent infections in several lepidopteran insect cells. Here, we have cloned and characterized one of the HzNV-1 early genes, hhi1, which maps to the HindIII-I fragment of the viral genome. During the productive viral infection, a 6.2-kb hhi1 transcript was detectable as early as 0.5 h postinfection (hpi). The level of transcript reached a maximum at 2 hpi and gradually decreased after 4 hpi. The transcript was not detectable during the latent phase of viral infection. Upon cycloheximide treatment, much higher levels of hhi1 transcript were detected throughout the productive viral infection cycle, suggesting that newly synthesized proteins are not needed for the expression of hhi1. Nevertheless, viral coinfection can further stimulate the expression of transfected hhi1 promoter in a plasmid. Transient hhi1 expression in latently infected cells resulted in a significant increase in virus titer and viral DNA propagation, suggesting that hhi1 plays a critical role in viral reactivation. Additional experiments showed that six early genes, which possibly function in transcription or DNA replication, were activated in the latent cells upon hhi1 transfection. Among these six genes, orf90 and orf121 expression could be induced by hhi1 alone without the need for other viral genes. Our discovery should be useful for future mechanistic study of the switches of latent/productive HzNV-1 viral infections.

scholarly journals Quantitation of mast cell heparin by flow cytofluorometry.

1976 ◽  
Vol 24 (12) ◽  
pp. 1231-1238 ◽  
Author(s):  
L Enerbäck ◽  
G Berlin ◽  
I Svensson ◽  
I Rundquist
Keyword(s):  
Mast Cell ◽  
Mast Cells ◽  
Mixed Cell ◽  
Flow Cytofluorometry ◽  
Frequency Distributions ◽  
Cell Counts ◽  
Mixed Cell Population ◽  
Number Of Cells ◽  
Excellent Reproducibility

Mast cells can be automatically identified in a mixed cell population by flow cytofluorometry after Berberine sulphate staining. Volume specific counts of the total number of cells and number of mast cells, as well as frequency distributions of fluorescence intensities of mast cells, based on a large number of cells, can be rapidly obtained. Results obtained by microscope fluorometry of cells identified by phase contrast microscopy showviously published results it may be inferred that the fluorescence intensity of individual mast cells is proportional to mast cell heparin content. The automated cell counts correlated very well with manual hemocytometer counts. Both cell counts and the determination of mean mast cell fluorescence showed excellent reproducibility.

scholarly journals Probabilistic control of HIV latency and transactivation by the Tat gene circuit

2018 ◽  
Vol 115 (49) ◽  
pp. 12453-12458 ◽  
Author(s):  
Youfang Cao ◽  
Xue Lei ◽  
Ruy M. Ribeiro ◽  
Alan S. Perelson ◽  
Jie Liang
Keyword(s):  
Cell Phenotype ◽  
Control Mechanisms ◽  
Viral Production ◽  
Gene Circuit ◽  
Hiv Tat ◽  
Latently Infected ◽  
Shock And Kill ◽  
Infected Cells ◽  
Cell Phenotypes ◽  
Druggable Targets

The reservoir of HIV latently infected cells is the major obstacle for eradication of HIV infection. The “shock-and-kill” strategy proposed earlier aims to reduce the reservoir by activating cells out of latency. While the intracellular HIV Tat gene circuit is known to play important roles in controlling latency and its transactivation in HIV-infected cells, the detailed control mechanisms are not well understood. Here we study the mechanism of probabilistic control of the latent and the transactivated cell phenotypes of HIV-infected cells. We reconstructed the probability landscape, which is the probability distribution of the Tat gene circuit states, by directly computing the exact solution of the underlying chemical master equation. Results show that the Tat circuit exhibits a clear bimodal probability landscape (i.e., there are two distinct probability peaks, one associated with the latent cell phenotype and the other with the transactivated cell phenotype). We explore potential modifications to reactions in the Tat gene circuit for more effective transactivation of latent cells (i.e., the shock-and-kill strategy). Our results suggest that enhancing Tat acetylation can dramatically increase Tat and viral production, while increasing the Tat–transactivation response binding affinity can transactivate latent cells more rapidly than other manipulations. Our results further explored the “block and lock” strategy toward a functional cure for HIV. Overall, our study demonstrates a general approach toward discovery of effective therapeutic strategies and druggable targets by examining control mechanisms of cell phenotype switching via exactly computed probability landscapes of reaction networks.

scholarly journals Latently KSHV-Infected Cells Promote Further Establishment of Latency upon Superinfection with KSHV

2021 ◽  
Vol 22 (21) ◽  
pp. 11994
Author(s):  
Chen Gam ze Letova ◽  
Inna Kalt ◽  
Meir Shamay ◽  
Ronit Sarid
Keyword(s):  
Latent Infection ◽  
Fluorescent Proteins ◽  
Cell Types ◽  
Lytic Cycle ◽  
Viral Reservoir ◽  
Virus Spread ◽  
Viral Genomes ◽  
Latently Infected ◽  
Infected Cells

Kaposi’s sarcoma-associated herpesvirus (KSHV) is a cancer-related virus which engages in two forms of infection: latent and lytic. Latent infection allows the virus to establish long-term persistent infection, whereas the lytic cycle is needed for the maintenance of the viral reservoir and for virus spread. By using recombinant KSHV viruses encoding mNeonGreen and mCherry fluorescent proteins, we show that various cell types that are latently-infected with KSHV can be superinfected, and that the new incoming viruses establish latent infection. Moreover, we show that latency establishment is enhanced in superinfected cells compared to primary infected ones. Further analysis revealed that cells that ectopically express the major latency protein of KSHV, LANA-1, prior to and during infection exhibit enhanced establishment of latency, but not cells expressing LANA-1 fragments. This observation supports the notion that the expression level of LANA-1 following infection determines the efficiency of latency establishment and avoids loss of viral genomes. These findings imply that a host can be infected with more than a single viral genome and that superinfection may support the maintenance of long-term latency.

scholarly journals The gammaherpesvirus 68 viral cyclin facilitates reactivation by promoting latent gene expression

2019 ◽  
Author(s):  
Brian F Niemeyer ◽  
Joy E Gibson ◽  
Jennifer N Berger ◽  
Lauren M Oko ◽  
Eva Medina ◽  
...  
Keyword(s):  
Gene Expression ◽  
Critical Role ◽  
Productive Infection ◽  
Viral Gene ◽  
Cellular Distribution ◽  
Viral Cyclin ◽  
Latently Infected ◽  
Infected Cells ◽  
Virally Encoded ◽  
The Impact

AbstractGammaherpesviruses establish life-long infections within their host and have been shown to be the causative agents of devastating malignancies. Chronic infection within the host is mediated through cycles of transcriptionally quiescent stages of latency with periods of reactivation into more active lytic and productive infection. The mechanisms that regulate reactivation from latency remain poorly understood. Previously, we defined a critical role for the viral cyclin in promoting reactivation from latency. Disruption of the viral cyclin had no impact on the frequency of cells containing viral genome during latency, yet it remains unclear whether the viral cyclin influences latently infected cells in a qualitative manner. To define the impact of the viral cyclin on latent gene expression, we utilized a viral cyclin deficient variant expressing a LANA-beta-lactamase fusion protein (LANA::βla), to enumerate both the cellular distribution and frequency of latent gene expression. Disruption of the viral cyclin did not affect the cellular distribution of latently infected cells, but did result in a significant decrease in the frequency of cells that expressed LANA::βla across multiple tissues and in both immunocompetent and immunodeficient hosts. Strikingly, whereas the cyclin-deficient virus had a reactivation defect in bulk culture, sort purified cyclin-deficient LANA::βla expressing cells were fully capable of reactivation. These data emphasize that the γHV68 latent reservoir is comprised of at least two distinct stages of infection characterized by differential latent gene expression, and that a primary function of the viral cyclin is to promote latent gene expression within infected cells in vivo.AUTHOR SUMMARYGammaherpesviruses are ubiquitous viruses with oncogenic potential that establish latency for the life of the host. These viruses can emerge from latency through reactivation, a process that is controlled by the immune system. Control of viral latency and reactivation is thought to be critical to prevent γHV-associated disease. This study focuses on a virally-encoded cyclin that is required for reactivation from latency. By characterizing how the viral cyclin influences latent infection in pure cell populations, we find that the viral cyclin has a vital role in promoting viral gene expression during latency. This work provides new insight into the function of a virally encoded cyclin in promoting reactivation from latency.

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