Lucy A. McNamara

HIV-1 infects multipotent progenitor cells causing cell death and establishing latent cellular reservoirs

HIV causes a chronic infection characterized by depletion of CD4+ T lymphocytes and the development of opportunistic infections. Despite drugs that inhibit viral spread, HIV infection has been difficult to cure because of uncharacterized reservoirs of infected cells that are resistant to highly active antiretroviral therapy (HAART) and the immune response. Here we used CD34+ cells from infected people as well as in vitro studies of wild-type HIV to show infection and killing of CD34+ multipotent hematopoietic progenitor cells (HPCs). In some HPCs, we detected latent infection that stably persisted in cell culture until viral gene expression was activated by differentiation factors. A unique reporter HIV that directly detects latently infected cells in vitro confirmed the presence of distinct populations of active and latently infected HPCs. These findings have major implications for understanding HIV bone marrow pathology and the mechanisms by which HIV causes persistent infection.

The antiviral factor APOBEC3G enhances the recognition of HIV-infected primary T cells by natural killer cells

APOBEC3G (A3G) is an intrinsic antiviral factor that inhibits the replication of human immunodeficiency virus (HIV) by deaminating cytidine residues to uridine. This causes guanosine-to-adenosine hypermutation in the opposite strand and results in inactivation of the virus. HIV counteracts A3G through the activity of viral infectivity factor (Vif), which promotes degradation of A3G. We report that viral protein R (Vpr), which interacts with a uracil glycosylase, also counteracted A3G by diminishing the incorporation of uridine. However, this process resulted in activation of the DNA-damage–response pathway and the expression of natural killer (NK) cell–activating ligands. Our results show that pathogen-induced deamination of cytidine and the DNA-damage response to virus-mediated repair of the incorporation of uridine enhance the recognition of HIV-infected cells by NK cells.APOBEC3G (A3G) is an intrinsic antiviral factor that inhibits HIV replication by deaminating cytidine residues to uridine. This causes G-to-A hypermutation in the opposite strand and results in viral inactivation. HIV counteracts A3G through the activity of viral infectivity factor (Vif), which promotes A3G degradation. We report that viral protein R (Vpr), which interacts with a uracil glycosylase, also counteracts A3G by reducing uridine incorporation. However, this process results in activation of the DNA damage response pathway and expression of NK cell activating ligands. Our results reveal that pathogen-induced cytidine deamination and the DNA damage response to viral-mediated repair of uridine incorporation enhance recognition of HIV-infected cells by NK cells.

HIV-1 utilizes the CXCR4 chemokine receptor to infect multipotent hematopoietic stem and progenitor cells

HIV infection is characterized by gradual immune system collapse and hematopoietic dysfunction. We recently showed that HIV enters multipotent hematopoietic progenitor cells and establishes both active cytotoxic and latent infections that can be reactivated by myeloid differentiation. However, whether these multipotent progenitors include long-lived hematopoietic stem cells (HSCs) that could establish viral reservoirs for the life of the infected person remains unknown. Here we provide direct evidence that HIV targets long-lived HSCs and show that infected HSCs yield stable, multilineage engraftment in a xenograft model. Furthermore, we establish that the capacity to use the chemokine receptor CXCR4 for entry determines whether a virus will enter multipotent versus differentiated progenitor cells. Because HSCs live for the life span of the infected person and are crucial for hematopoietic health, these data may explain the poor prognosis associated with CXCR4-tropic HIV infection and suggest HSCs as long-lived cellular reservoirs of latent HIV.

Hematopoietic stem/precursor cells as HIV reservoirs.

Purpose of reviewAlthough latent HIV-1 infection in CD4+ T cells contributes to HIV persistence, there is mounting evidence that other viral reservoirs exist. Here, we review recent data suggesting that the infection of hematopoietic progenitor cells creates additional reservoirs for HIV in vivo. Recent findingsNew studies suggest that some types of hematopoietic progenitor cells have the potential to generate reservoirs for HIV. This review focuses on two types that can be infected by HIV in vitro and in vivo: multipotent hematopoietic progenitor cells in the bone marrow and circulating mast cell progenitors. Of these two types, only CD34+ bone marrow cells have been shown to harbor latent provirus in HIV-positive individuals with undetectable viral loads on highly active antiretroviral therapy (HAART). Latent infection of these long-lived cell types may create a significant barrier to HIV eradication; the infection of hematopoietic stem cells in particular could lead to an HIV reservoir that does not appreciably decay over the lifespan of the host. SummaryTo eradicate HIV infection, it will be necessary to purge all viral reservoirs in the host. The findings highlighted here suggest that multipotent hematopoietic progenitor cells and possibly tissue mast cells may constitute significant reservoirs for HIV that must be addressed in order to eliminate HIV infection. Future studies are needed to determine which types of CD34+ cells are infected in vivo and whether infected CD34+ cells contribute to residual viremia in people with undetectable viral loads on HAART.

Latent HIV-1 Infection Occurs in Multiple Subsets of Hematopoietic Progenitor Cells and Is Reversed by NF-κB Activation

ABSTRACT The ability of HIV-1 to establish a latent infection presents a barrier to curing HIV. The best-studied reservoir of latent virus in vivo is resting memory CD4+ T cells, but it has recently been shown that CD34+ hematopoietic progenitor cells (HPCs) can also become latently infected by HIV-1 in vitro and in vivo. CD34+ cells are not homogenous, however, and it is not yet known which types of CD34+ cells support a latent infection. Furthermore, the mechanisms through which latency is established in this cell type are not yet known. Here we report the development of a primary cell model for latent HIV-1 infection in HPCs. We demonstrate that in this model, latent infection can be established in all subsets of HPCs examined, including HPCs with cell surface markers consistent with immature hematopoietic stem and progenitor cells. We further show that the establishment of latent infection in these cells can be reversed by tumor necrosis factor alpha (TNF-α) through an NF-κB-dependent mechanism. In contrast, we do not find evidence for a role of positive transcription elongation factor b (P-TEFb) in the establishment of latent infection in HPCs. Finally, we demonstrate that prostratin and suberoylanilide hydroxamic acid (SAHA), but not hexamethylene bisacetamide (HMBA) or 5-aza-2′-deoxycytidine (Aza-CdR), reactivate latent HIV-1 in HPCs. These findings illuminate the mechanisms through which latent infection can be established in HPCs and suggest common pathways through which latent virus could be reactivated in both HPCs and resting memory T cells to eliminate latent reservoirs of HIV-1.

CD133+ Hematopoietic Progenitor Cells Harbor HIV Genomes in a Subset of Optimally Treated People With Long-Term Viral Suppression

BACKGROUND Hematopoietic progenitor cells (HPCs) in the bone marrow of human immunodeficiency virus (HIV)-infected individuals have been proposed as a persistent reservoir of virus. However, some studies have suggested that HIV genomes detected in HPCs arise from T-cell contamination. METHODS CD133-sorted HPCs and CD133-depleted bone marrow cells were purified from bone marrow specimens obtained from 11 antiretroviral-treated donors in whom the HIV load had been <48 copies/mL for at least 6 months. CD133 and CD3 expression on the cells was assessed by flow cytometry. HIV DNA was quantified by real-time polymerase chain reaction analysis. RESULTS HIV genomes were detected in CD133-sorted samples from 6 donors, including 2 in whom viral loads were undetectable for >8 years. CD3(+) T cells represented <1% of cells in all CD133-sorted samples. For 5 of 6 CD133-sorted samples with detectable HIV DNA, the HIV genomes could not be explained by contaminating CD3(+) T cells. Donors with detectable HIV DNA in HPCs received their diagnosis significantly more recently than the remaining donors but had had undetectable viral loads for similar periods. CONCLUSIONS HIV genomes can be detected in CD133-sorted cells from a subset of donors with long-term viral suppression and, in most cases, cannot be explained by contamination with CD3(+) T cells.

Using epitope predictions to evaluate efficacy and population coverage of the Mtb72f vaccine for tuberculosis

BackgroundThe Mtb72f subunit vaccine for tuberculosis, currently in clinical trials, is hoped to provide improved protection compared to the current BCG vaccine. It is not clear, however, whether Mtb72f would be equally protective in the different human populations suffering from a high burden of tuberculosis. Previous work by Hebert and colleagues demonstrated that the PPE18 protein of Mtb72f had significant variability in a sample of clinical M. tuberculosis isolates. However, whether this variation might impact the efficacy of Mtb72f in the context of the microbial and host immune system interactions remained to be determined. The present study assesses Mtb72fs predicted efficacy in people with different DRB1 genotypes to predict whether the vaccine will protect against diverse clinical strains of M. tuberculosis in a diverse host population.ResultsWe evaluated the binding of epitopes in the vaccine to different alleles of the human DRB1 Class II MHC protein using freely available epitope prediction programs and compared protein sequences from clinical isolates to the sequences included in the Mtb72f vaccine. This analysis predicted that the Mtb72f vaccine would be less effective for several DRB1 genotypes, due either to limited vaccine epitope binding to the DRB1 proteins or to binding primarily by unconserved PPE18 epitopes. Furthermore, we found that these less-protective DRB1 alleles are found at a very high frequency in several populations with a high burden of tuberculosis.ConclusionAlthough the Mtb72f vaccine candidate has shown promise in animal and clinical trials thus far, it may not be optimally effective in some genotypic backgrounds. Due to variation in both M. tuberculosis protein sequences and epitope-binding capabilities of different HLA alleles, certain human populations with a high burden of tuberculosis may not be optimally protected by the Mtb72f vaccine. The efficacy of the Mtb72f vaccine should be further examined in these particular populations to determine whether additional protective measures might be necessary for these regions.

High Risk for Invasive Meningococcal Disease Among Patients Receiving Eculizumab (Soliris) Despite Receipt of Meningococcal Vaccine

Use of eculizumab (Soliris, Alexion Pharmaceuticals), a terminal complement inhibitor, is associated with a 1,000-fold to 2,000-fold increased incidence of meningococcal disease (1). Administration of meningococcal vaccines is recommended for patients receiving eculizumab before beginning treatment (2,3). Sixteen cases of meningococcal disease were identified in eculizumab recipients in the United States during 2008-2016; among these, 11 were caused by nongroupable Neisseria meningitidis. Fourteen patients had documentation of receipt of at least 1 dose of meningococcal vaccine before disease onset. Because eculizumab recipients remain at risk for meningococcal disease even after receipt of meningococcal vaccines, some health care providers in the United States as well as public health agencies in other countries recommend antimicrobial prophylaxis for the duration of eculizumab treatment; a lifelong course of treatment is expected for many patients. Heightened awareness, early care seeking, and rapid treatment of any symptoms consistent with meningococcal disease are essential for all patients receiving eculizumab treatment, regardless of meningococcal vaccination or antimicrobial prophylaxis status.

Comparison of the Predicted Population Coverage of Tuberculosis Vaccine Candidates Ag85B-ESAT-6, Ag85B-TB10.4, and Mtb72f via a Bioinformatics Approach

The Bacille-Calmette Guérin (BCG) vaccine does not provide consistent protection against adult pulmonary tuberculosis (TB) worldwide. As novel TB vaccine candidates advance in studies and clinical trials, it will be critically important to evaluate their global coverage by assessing the impact of host and pathogen variability on vaccine efficacy. In this study, we focus on the impact that host genetic variability may have on the protective effect of TB vaccine candidates Ag85B-ESAT-6, Ag85B-TB10.4, and Mtb72f. We use open-source epitope binding prediction programs to evaluate the binding of vaccine epitopes to Class I HLA (A, B, and C) and Class II HLA (DRB1) alleles. Our findings suggest that Mtb72f may be less consistently protective than either Ag85B-ESAT-6 or Ag85B-TB10.4 in populations with a high TB burden, while Ag85B-TB10.4 may provide the most consistent protection. The findings of this study highlight the utility of bioinformatics as a tool for evaluating vaccine candidates before the costly stages of clinical trials and informing the development of new vaccines with the broadest possible population coverage.

Interferon Alfa Therapy: Toward an Improved Treatment for HIV Infection

Highly active antiretroviral therapy(HAART) is an expensive, lifelong treat-ment for human immunodeficiencyvirus (HIV) infection that is associatedwith significant toxicity. Despite advanc-es in treatment, there remains a need fornovel therapies for HIV infection. Onestrategy is to achieve a functional cure,in which treatment can be safely stoppeddespite the presence of residual virus, byfinding approaches that enhance the im-mune response to HIV. Such a therapywould require boosting a patient’sim-mune system to suppress viral replicationbelow clinically relevant levels, therebyeliminating the need for HAART. A ster-ilizing cure, by contrast, is one in whichall functional HIV genomes within thebody are eliminated. Latent HIV genomesrepresent a major barrier to a sterilizingcure because these genomes do notproduce viral proteins and thus cannotbe eliminated by the immune system orby current therapies [1].In this issue, Azzoni and colleaguesassessed the potential of pegylated (Peg)interferon alfa-2a to suppress HIV repli-cation [2]. In what could be a first steptowards a functional cure, they foundthat Peg–interferon alfa-2a permitted asubset of subjects to maintain low viralloads for 12–24 weeks in the absence ofHAART. Furthermore, they found thatin patients with a favorable virologic re-sponse to Peg interferon alfa-2a, levelsof HIV provirus in peripheral bloodmononuclear cells (PBMCs) werereduced. This finding suggests that Peg–interferon alfa-2a may eliminate latentHIV genomes, a possibility that wouldshed light on the search for a sterilizingas well as a functional cure for HIV.Interferon alfa is used clinically forthe treatment of hepatitis B and C virusinfections, and several studies have as-sessed the capacity of this cytokine tolimit HIV replication in vivo [3–9].Mul-tiple studies confirm that interferon alfacan reduce viral load and delay diseaseprogression in viremic patients [3,5–11].However, interferon alfa’s potential tosuppress viremia in patients with well-controlled viral loads is less clear.Recently, one study subjected HAART-treated patients with a viral load of<400 copies/mL forat least 6 months anda CD4