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HIV Reservoir Size
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Written by Alain Lafeuillade
http://www.hiv-reservoir.net/
Friday, 03 June 2011 13:37
http://www.hiv-reservoir.net/index.php/Latest-News-on-HIV-Reservoirs-Eradication/hiv-reservoir-size.html
Size of HIV Reservoir
In a study published in the Journal of Infectious Diseases, Tae-Wook Chun et al. have analyzed the correlations between the size of HIV proviral DNA reservoirs, residual plasma viremia and immune activation in 127 individuals on effective ART.
This study (1) included 127 individuals who had received ART (various regimens) for a median of 6.5 years (range, 1.3-15.8 years) and who had achieved suppression of plasma viremia. The median CD4+ and CD8+ T-cell were 580 cells/mm3 of blood and 760 cells/mm3, respectively. All participants included in this study maintained undetectable levels of plasma viremia (<50 copies/mL) and had fewer than 3 viral ''blips'' (defined as <100 HIV RNA copies/mL) after initiation of ART, as determined by frequent blood sampling (at least 3 times per year).
The plasma from the majority of the study participants (63.0%) contained detectable plasma viremia, whereas there was no measurable HIV RNA in the plasma of 37.0% (median HIV RNA level: 2.6 copies/ml). The median copy number of HIV proviral DNA for all study participants examined was 775.1 copies (range, <2.6-6890.6 copies) per 106 CD4+ T cells.
The frequency of CD4+ T cells carrying HIV proviral DNA in infected individuals with undetectable plasma viremia (median, 448.5 copies per 106 CD4+ T cells) as a group was lower than that of individuals with detectable plasma viremia (median, 1027.2 copies per 106 CD4+ T cells).
However, 38% of the study participants with undetectable plasma viremia carried >775 copies per 106 CD4+ T cells (median), suggesting that, at least in some individuals, the size of the HIV reservoir in the CD4+ T-cell compartment in the peripheral blood may not necessarily equate to the level of residual plasma HIV.
No correlation was found between markers of immune activation (C-reactive protein, D-dimer, IL-6, soluble TNF receptor I, CD4+ CD38+, and CD8+ CD38+) and residual plasma viremia.
This study suggests that reactivation of the latent viral reservoir may not be the sole source of residual plasma viremia. Considering the relatively high frequency of HIV infection in the CD4+ T-cell compartment of nearly 40% of the study participants exhibiting undetectable plasma viremia (0 copy) and the lack of any correlation between residual plasma viremia and various markers of immune activation in blood, there is a possibility that residual viral replication, with or without the detection of plasma viremia, may also originate from productively infected CD4+ T cells in various lymphoid tissues.
Reference
Chun TW, Murray D, Justement JS, Hallahan CW, Moir S, Kovacs C, Fauci AS. Relationship Between Residual Plasma Viremia and the Size of HIV Proviral DNA Reservoirs in Infected Individuals Receiving Effective Antiretroviral Therapy. J Infect Dis. 2011; 204(1):135-8.
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Relationship Between Residual Plasma Viremia and the Size of HIV Proviral DNA Reservoirs in Infected Individuals Receiving Effective Antiretroviral Therapy
The Journal of Infectious Diseases July 1 2011
Tae-Wook Chun,1 Danielle Murray,1 J. Shawn Justement,1 Claire W. Hallahan,2 Susan Moir,1 Colin Kovacs,3 and Anthony S. Fauci1
1Laboratory of Immunoregulation, and 2Biostatistics Research Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland; and 3Department of Medicine, University of Toronto, Ontario, Canada
"Here, we demonstrate that residual plasma viremia correlates with the size of the CD4+ T cell viral reservoir, but not with markers of immune activation, suggesting that reactivation of the latent viral reservoir may not be the sole source of residual plasma viremia. Novel therapeutic strategies aimed at targeting the source of residual viremia may be necessary to achieve viral eradication.....We have demonstrated detectable levels of residual plasma viremia (1-49 copies/mL) in the majority of study participants receiving ART in whom plasma viremia had been suppressed for extended periods of time, as measured by a standard clinical assay (with a limit of detection of 50 copies/mL). Furthermore, we found a correlation between the level of residual plasma viremia and the frequency of CD4+ T cells carrying HIV proviral DNA."
"considering the relatively high frequency of HIV infection in the CD4+ T-cell compartment of nearly 40% of the study participants exhibiting undetectable plasma viremia (0 copy) and the lack of any correlation between residual plasma viremia and various markers of immune activation in blood, there is a possibility that residual viral replication, with or without the detection of plasma viremia, may also originate from productively infected CD4+ T cells in various lymphoid tissues. In this regard, recent studies investigating the effect of intensification of conventional ART have demonstrated evidence for the existence of ongoing viral replication, as measured by levels of cell-associated viral DNA in blood [12] and RNA in the GALT [13]; however, in other studies with similar settings, but in which plasma viremia was the sole virological indicator, there was no effect of drug intensification [14, 15]. With regard to these apparent discrepancies, it is important to point out that it is possible to have low levels of viral replication and cell-to-cell spread of virus, particularly in lymphoid organs, without such replication being reflected in the levels of plasma viremia, even as measured by the most sensitive assays."
"Achieving eradication of HIV in infected individuals receiving ART remains a daunting challenge for the scientific community. To achieve a functional cure, as defined by the absence of detectable HIV for extended periods of time in the absence of ART, therapeutic strategies aimed at eliminating cellular reservoirs in various tissue compartments must be accompanied by comprehensive virological assays that monitor infected CD4+ T cells that may or may not contribute to residual plasma viremia."
Abstract
Residual plasma viremia (<50 copies/mL) persists in certain human immunodeficiency virus (HIV)-infected individuals receiving antiretroviral therapy (ART); however, the relationship between the degree of residual plasma viremia, the size of HIV reservoirs, and the level of immune activation has not been delineated. Here, we demonstrate that residual plasma viremia correlates with the size of the CD4+ T cell viral reservoir, but not with markers of immune activation, suggesting that reactivation of the latent viral reservoir may not be the sole source of residual plasma viremia. Novel therapeutic strategies aimed at targeting the source of residual viremia may be necessary to achieve viral eradication.
Prolonged suppression of plasma viremia is now achievable in most human immunodeficiency virus (HIV)-infected individuals receiving antiretroviral therapy (ART) [1]. Nonetheless, it has not been possible to eradicate HIV by ART alone, likely due in part to the persistence of various viral reservoirs [2-6]. A number of previous studies have demonstrated that HIV persists in latently and productively infected CD4+ T cells in peripheral blood [2-4] as well as in gut-associated lymphoid tissues (GALT) [6, 7] of infected individuals receiving ART who have maintained undetectable plasma viremia for prolonged periods of time, as measured by clinically relevant assays (with a typical limit of detection of 50 HIV RNA copies/mL of plasma). With the advent of a laboratory-based real-time polymerase chain reaction (PCR) assay capable of detecting single copies of HIV RNA in plasma [8], several studies have recently demonstrated the presence of residual plasma viremia ranging from 1 to 49 copies/mL in some infected individuals receiving ART [8-10]. One such study observed multiphasic decay of residual plasma viremia and speculated that latently infected, resting CD4+ T cells and/or unidentified viral reservoirs, which are capable of producing low levels of genetically identical virions for prolonged periods of time without cellular turn-over, may be responsible for the persistence of residual plasma viremia in infected individuals receiving ART for extended periods of time [10, 11]. However, the relationship between residual plasma viremia and the frequency of CD4+ T cells carrying HIV proviral DNA and/or markers of immune activation has not been fully delineated. We conducted the present study to address this issue.
DISCUSSION
The persistence of HIV proviral DNA and infectious virus in CD4+ T cells has long been considered as one of the major impediments to eradicating virus in infected individuals receiving ART for extended periods of time and in whom plasma viremia was suppressed below the level of detection, as measured by a number of US Food and Drug Administration-approved clinical assays (which have a limit of detection of 50 copies/mL) [2-4]. In recent years, studies using a laboratory-based real-time PCR assay that is capable of detecting single HIV RNA copies in plasma have shown that residual plasma viremia 50 copies/mL can be observed in certain infected individuals who are receiving effective ART [9, 10]. Given that delineating the mechanism by which residual plasma viremia persists and identifying the source of the <50 HIV RNA copies/mL in plasma may have profound implications for eradicating HIV in infected individuals receiving ART, it is important to understand the relationship between residual plasma viremia and various immunological and virological parameters, such as the level of cell-associated HIV proviral DNA and markers of immune activation.
In the present study, we set out to determine the degrees of residual plasma viremia in a large number of infected individuals receiving ART and to determine immunological or virological parameters that may correlate with residual plasma viremia. Residual plasma viremia was determined in quadruplicate using an automated system to minimize quantitative errors often associated with the detection of extremely low levels of viral RNA. We have demonstrated detectable levels of residual plasma viremia (1-49 copies/mL) in the majority of study participants receiving ART in whom plasma viremia had been suppressed for extended periods of time, as measured by a standard clinical assay (with a limit of detection of 50 copies/mL). Furthermore, we found a correlation between the level of residual plasma viremia and the frequency of CD4+ T cells carrying HIV proviral DNA. Of note, due to the limited amounts of blood obtained from the study subjects, we could not conduct the quantitative coculture assays that are required for determining the frequency of cells carrying replication-competent HIV. It has been speculated that residual plasma viremia originates from the cellular activation of latently infected, resting CD4+ T cells [10]. In this regard, we have previously shown that activated CD4+ T cells in the peripheral blood of HIV-infected individuals receiving ART spontaneously released virions in the absence of activating stimuli [5]. In addition, we have reported higher frequencies of HIV infection in CD4+ T cells in the GALT and evidence for cross-infection between the blood and tissue compartments of infected individuals receiving ART [6]. The data presented in the present study do not rule out the possibility of reactivation of latently infected, resting CD4+ T cells contributing to residual plasma viremia. However, considering the relatively high frequency of HIV infection in the CD4+ T-cell compartment of nearly 40% of the study participants exhibiting undetectable plasma viremia (0 copy) and the lack of any correlation between residual plasma viremia and various markers of immune activation in blood, there is a possibility that residual viral replication, with or without the detection of plasma viremia, may also originate from productively infected CD4+ T cells in various lymphoid tissues. In this regard, recent studies investigating the effect of intensification of conventional ART have demonstrated evidence for the existence of ongoing viral replication, as measured by levels of cell-associated viral DNA in blood [12] and RNA in the GALT [13]; however, in other studies with similar settings, but in which plasma viremia was the sole virological indicator, there was no effect of drug intensification [14, 15]. With regard to these apparent discrepancies, it is important to point out that it is possible to have low levels of viral replication and cell-to-cell spread of virus, particularly in lymphoid organs, without such replication being reflected in the levels of plasma viremia, even as measured by the most sensitive assays.
Achieving eradication of HIV in infected individuals receiving ART remains a daunting challenge for the scientific community. To achieve a functional cure, as defined by the absence of detectable HIV for extended periods of time in the absence of ART, therapeutic strategies aimed at eliminating cellular reservoirs in various tissue compartments must be accompanied by comprehensive virological assays that monitor infected CD4+ T cells that may or may not contribute to residual plasma viremia.
RESULTS
To determine the level of residual plasma viremia <50 HIV RNA copies/mL in 127 infected individuals receiving ART, we performed an automated viral load assay (Cobas Ampliprep/Cobas Taqman HIV-1 Test, version 2.0; Roche Diagnostics) in quadruplicate for each study participant. As shown in Figure 1A, residual plasma viremia ranged from 0 to 49 HIV RNA copies/mL (median HIV RNA level, 2.6 copies/mL). The plasma from the majority of the study participants (63.0%) contained detectable plasma viremia, whereas there was no measurable HIV RNA in the plasma of 37.0% of the individuals in our study.
To examine the correlation between residual plasma viremia (at the level of <50 HIV RNA copies/mL) and the frequency of CD4+ T cells carrying HIV proviral DNA, genomic DNA was prepared from highly purified CD4+ T cells from peripheral blood of study participants and subjected to real-time PCR specific for HIV proviral DNA. The median copy number of HIV proviral DNA for all study participants examined was 775.1 copies (range, <2.6-6890.6 copies) per 106 CD4+ T cells. There was a direct correlation between the level of residual plasma viremia and the frequency of CD4+ T cells carrying HIV proviral DNA (r = .294; P = .001) (Figure 1B). When the data on HIV proviral DNA burden were stratified on the basis of residual plasma viremia, there was a statistically significant difference between the study subjects with undetectable (0 copies) versus detectable (1-49 copies) plasma viremia (P = .01) (Figure 1C), indicating that the frequency of CD4+ T cells carrying HIV proviral DNA in infected individuals with undetectable plasma viremia (median, 448.5 copies per 106 CD4+ T cells; interquartile range, 166.7-1467.2 copies per 106 CD4+ T cells) as a group is lower than that of individuals with detectable plasma viremia (median, 1027.2 copies per 106 CD4+ T cells; interquartile range, 292.2-3040.7 copies per 106 CD4+ T cells). Of note, 38% of the study participants with undetectable plasma viremia carried ≥775 copies per 106 CD4+ T cells (median value of all data points), suggesting that, at least in some individuals, the size of the HIV reservoir in the CD4+ T-cell compartment in the peripheral blood may not necessarily equate to the level of residual plasma HIV.
To determine the effect of immune activation on residual plasma viremia and the size of the viral reservoir carrying HIV proviral DNA in infected individuals receiving clinically effective ART, we sought to evaluate possible correlations among immune markers (C-reactive protein, D-dimer, IL-6, soluble TNF receptor I, and CD38 expression on CD4+ and CD8+ T cells), residual plasma viremia, and the frequency of CD4+ T cells carrying HIV proviral DNA. There was no correlation among the above parameters (P > .5), with the exception of a direct correlation between CD4+/CD8+ T-cell count ratio and the level of CD4+ T cells carrying HIV proviral DNA (P = .005). However, there was no correlation between markers of immune activation and residual plasma viremia (P > .5).
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