The persistence of latently infected cells in patients under combinatory antiretroviral therapy (cART) is a major hurdle to HIV-1 eradication. Strategies to purge these reservoirs are needed and ...activation of viral gene expression in latently infected cells is one promising strategy. Bromodomain and Extraterminal (BET) bromodomain inhibitors (BETi) are compounds able to reactivate latent proviruses in a positive transcription elongation factor b (P-TEFb)-dependent manner. In this study, we tested the reactivation potential of protein kinase C (PKC) agonists (prostratin, bryostatin-1 and ingenol-B), which are known to activate NF-κB signaling pathway as well as P-TEFb, used alone or in combination with P-TEFb-releasing agents (HMBA and BETi (JQ1, I-BET, I-BET151)). Using in vitro HIV-1 post-integration latency model cell lines of T-lymphoid and myeloid lineages, we demonstrated that PKC agonists and P-TEFb-releasing agents alone acted as potent latency-reversing agents (LRAs) and that their combinations led to synergistic activation of HIV-1 expression at the viral mRNA and protein levels. Mechanistically, combined treatments led to higher activations of P-TEFb and NF-κB than the corresponding individual drug treatments. Importantly, we observed in ex vivo cultures of CD8+-depleted PBMCs from 35 cART-treated HIV-1+ aviremic patients that the percentage of reactivated cultures following combinatory bryostatin-1+JQ1 treatment was identical to the percentage observed with anti-CD3+anti-CD28 antibodies positive control stimulation. Remarkably, in ex vivo cultures of resting CD4+ T cells isolated from 15 HIV-1+ cART-treated aviremic patients, the combinations bryostatin-1+JQ1 and ingenol-B+JQ1 released infectious viruses to levels similar to that obtained with the positive control stimulation. The potent effects of these two combination treatments were already detected 24 hours post-stimulation. These results constitute the first demonstration of LRA combinations exhibiting such a potent effect and represent a proof-of-concept for the co-administration of two different types of LRAs as a potential strategy to reduce the size of the latent HIV-1 reservoirs.
The main aim of this study was to determine whether HIV replication can be controlled following interruption of treatment started early in the course of infection (CD4 >350 cells/μl and viral load ...<50 000 copies/ml), but not during the primary infection.
Patients enrolled in a multicenter trial of treatment interruption (ANRS 116 SALTO) with CD4 above 450 cells/μl and viral load below 400 copies/ml at treatment interruption were selected for this second analysis. We determined the proportion of patients whose plasma HIV-RNA load remained below 400 copies/ml during the first 12 months of treatment interruption, and baseline factors predictive of time to loss of viral control. Viral load rebound was defined as two successive values above 400 copies/ml, or as one value above 400 copies/ml, followed by treatment resumption.
We studied 95 patients with a median CD4 nadir of 382 cells/μl (340-492). At treatment interruption, the median CD4 cell count and HIV-DNA load were 813/μl (695-988) and 206 copies/10 peripheral blood mononuclear cells (PBMCs) (53-556). Twelve months after treatment interruption, seven patients still had viral load below 400 copies/ml (Kaplan-Meier estimate 7.5%, 95% confidence interval 3.7-14.6), and four of them still had viral load below 400 copies/ml at 36 months. A multivariable Cox proportional-hazards model showed that time to loss of viral control was more shorter in patients with HIV-DNA at least 150 copies/10 PBMCs at treatment interruption (hazard ratio 2.1, 95% confidence interval 1.3-3.3, P = 0.002) than in those with HIV-DNA below 150 copies/10 PBMCs.
Patients who have low HIV-DNA levels at antiretroviral treatment interruption are more likely to maintain viral control for long periods.
To characterize viro-immunological outcomes following long-term combined antiretroviral therapy (cART) initiated during primary HIV infection (PHI) or chronic HIV infection (CHI) and to identify ...factors predictive of optimal viro-immunological responder (OVIR) status.
This was a prospective, single-centre cohort study of HIV-1-infected patients on effective cART. Total cell-associated HIV DNA levels and T cell counts before and during treatment were used to identify factors predictive of OVIR status {i.e. low HIV DNA level <2.3 log10 copies/10(6) peripheral blood mononuclear cells (PBMCs), together with normalization of the absolute/relative CD4+ T cell counts and CD4+/CD8+ ratio}.
A total of 307 patients were enrolled, of whom 35 started cART during PHI (<4 months post-infection) and 272 during CHI. HIV DNA decay was modelled with a non-linear mixed-effects model that showed two phases of HIV DNA decay, both of which were significantly more pronounced in the PHI group. At the end of follow-up, after a median of 4 years of viral suppression (<50 copies/mL), HIV DNA levels were lower in the PHI group than in the CHI group (median = 2.15 versus 2.84 log10 copies/10(6) PBMCs; P< 0.0001). Immune reconstitution was more rapid and sustained in the PHI group (median = 883 versus 619 CD4+ cells/mm(3); 41% versus 31% CD4+; CD4+/CD8+ 1.31 versus 0.77; all P< 0.0001). Finally, OVIR status was obtained in 19/35 (54%) and 7/272 (3%) patients in the PHI and CHI groups (P< 0.0001), respectively. In a logistic regression analysis, cART initiation during PHI (OR = 16, 95% CI = 3.5-72.3) and HIV DNA level <3.3 log10 before treatment (OR = 4.8, 95% CI = 1.2-19.3) were independently predictive of OVIR status.
Initiating cART during PHI represents a major opportunity to reduce HIV reservoirs and achieve optimal immune reconstitution.
Background. The stable immune control of human immunodeficiency virus (HIV) in long-term nonprogressors (LTNPs) with protective human leukocyte antigen (HLA) alleles raises the question of whether ...and how these alleles influence the immune distribution of the HIV reservoirs. Methods. Cell-associated HIV-DNA levels were quantified in blood sorted resting CD4 T-cell subsets from 8 LTNPs with and 10 without HLA-B*27 or HLA-B*57 alleles (HLA-B27/B57). Results. A remarkably lower infection level of central memory CD4 T cells (T CM ) was an exclusive feature that distinguished the HLA-B27/B57 HIV reservoirs from the other ones. In LTNPs, T CM protection was correlated with preservation of T CM counts, which correlated positively with the magnitude of HIV Gag-specific CD8 T cells. In HLA-B27/B57 LTNPs, a lower activation level of their memory CD4 T cells was associated with lower amounts of cell HIV-DNA in each resting memory CD4 subset and were also associated with higher ratios of HIV Gag-specific CD8 T cells per infected resting CD4 T cell (effector/target E/T). As a result, HLA-B27/B57 E/T ratios were negatively correlated with the contribution of memory CD4 T-cell subsets to the total HIV reservoirs. Conclusions. The potent antiviral immunity governed by the protective HLA-B27/B57 alleles, by limiting T CM infection and pool exhaustion, are associated with a reduced T CM contribution to the HIV reservoir.
How HIV controllers (HICs) maintain undetectable viremia without therapy is unknown. The strong CD8+ T-cell HIV suppressive capacity found in many, but not all, HICs may contribute to long-lasting ...viral control. However, other earlier defense mechanisms may be involved. Here, we examined intrinsic HIC cell resistance to HIV-1 infection. After in vitro challenge, monocyte-derived macrophages and anti–CD3-activated CD4+ T cells from HICs showed low HIV-1 susceptibility. CD4 T-cell resistance was independent of HIV-1 coreceptors and affected also SIVmac infection. CD4+ T cells from HICs expressed ex vivo higher levels of p21Waf1/Cip1, which has been involved in the control of HIV-1 replication, than cells from control subjects. However, HIV restriction in anti–CD3-activated CD4+ T cells and macrophages was not associated with p21 expression. Restriction inhibited accumulation of reverse transcripts, leading to reduction of HIV-1 integrated proviruses. The block could be overcome by high viral inocula, suggesting the action of a saturable mechanism. Importantly, cell-associated HIV-1 DNA load was extremely low in HICs and correlated with CD4+ T-cell permissiveness to infection. These results point to a contribution of intrinsic cell resistance to the control of infection and the containment of viral reservoir in HICs.
Reactivation of HIV gene expression in latently infected cells together with an efficient cART has been proposed as an adjuvant therapy aimed at eliminating/decreasing the reservoir size. Results ...from HIV clinical trials using deacetylase inhibitors (HDACIs) question the efficiency of these latency‐reversing agents (LRAs) used alone and underline the need to evaluate other LRAs in combination with HDACIs. Here, we evaluated the therapeutic potential of a demethylating agent (5‐AzadC) in combination with clinically tolerable HDACIs in reactivating HIV‐1 from latency first in vitro and next ex vivo. We showed that a sequential treatment with 5‐AzadC and HDACIs was more effective than the corresponding simultaneous treatment both in vitro and ex vivo. Interestingly, only two of the sequential LRA combinatory treatments tested induced HIV‐1 particle recovery in a higher manner than the drugs alone ex vivo and at concentrations lower than the human tolerable plasmatic concentrations. Taken together, our data reveal the benefit of using combinations of 5‐AzadC with an HDACI and, for the first time, the importance of treatment time schedule for LRA combinations in order to reactivate HIV.
Synopsis
Sequential administration of latency‐reversing agents (LRAs), namely deacetylase inhibitors (HDACIs) and demethylating agents, together with an efficient cART, could represent an adjuvant anti‐HIV‐1 therapy to induce viral production and possibly reduce the size of latent HIV‐1 reservoirs.
A sequential treatment with 5‐AzadC and HDACIs is more effective both in vitro and ex vivo at inducing HIV gene expression than the corresponding simultaneous treatments, highlighting for the first time the importance of treatment schedule for LRAs combinations.
Combining 5‐AzadC + panobinostat and 5‐AzadC + romidepsin show reactivation potentials at concentrations lower than the human tolerable plasmatic concentrations.
These findings are valuable for designing future anti‐latency therapeutic strategies and thereby constitute a step forward in achieving HIV remission.
Sequential administration of latency‐reversing agents (LRAs), namely deacetylase inhibitors (HDACIs) and demethylating agents, together with an efficient cART, could represent an adjuvant anti‐HIV‐1 therapy to induce viral production and possibly reduce the size of latent HIV‐1 reservoirs.
HIV-2 infection is characterized by a very low replication rate in most cases and low progression. This necessitates an approach to patient monitoring that differs from that for HIV-1 infection. ...Here, a new highly specific and sensitive method for HIV-2 DNA quantification was developed. The new test is based on quantitative real-time PCR targeting the long terminal repeat (LTR) and
regions and using an internal control. Analytical performance was determined in three laboratories, and clinical performance was determined on blood samples from 63 patients infected with HIV-2 group A (
= 35) or group B (
= 28). The specificity was 100%. The 95% limit of detection was three copies/PCR and the limit of quantification was six copies/PCR. The within-run coefficients of variation were between 1.03% at 3.78 log
copies/PCR and 27.02% at 0.78 log
copies/PCR. The between-run coefficient of variation was 5.10%. Both manual and automated nucleic acid extraction methods were validated. HIV-2 DNA loads were detectable in blood cells from all 63 patients. When HIV-2 DNA was quantifiable, median loads were significantly higher in antiretroviral-treated than in naive patients and were similar for groups A and B. HIV-2 DNA load was correlated with HIV-2 RNA load (
= 0.68; 95% confidence interval CI, 0.4 to 0.8;
< 0.0001). Our data show that this new assay is highly sensitive and quantifies the two main HIV-2 groups, making it useful for the diagnosis of HIV-2 infection and for pathogenesis studies on HIV-2 reservoirs.
Therapeutic control of HIV replication reduces the size of the viral reservoir, particularly among central memory CD4+ T cells, and this effect might be accentuated by early treatment.
We examined ...the effect of ART initiated at the time of the primary HIV infection (early ART), lasting 2 and 6 years in 11 and 10 patients, respectively, on the HIV reservoir in peripheral resting CD4+ T cells, sorted into naive (TN), central memory (TCM), transitional memory (TTM) and effector memory (TEM) cells, by comparison with 11 post-treatment controllers (PTCs).
Between baseline and 2 years, CD4+ T cell subset numbers increased markedly (P < 0.004) and HIV DNA levels decreased in all subsets (P < 0.009). TTM cells represented the majority of reservoir cells at both timepoints, T cell activation status normalized and viral diversity remained stable over time. The HIV reservoir was smaller after 6 years of early ART than after 2 years (P < 0.019), and did not differ between PTCs and patients treated for 6 years. One patient, who had low reservoir levels in all T cell subsets after 2 years of treatment similar to the levels in PTCs, spontaneously controlled viral replication during 18 months off treatment.
Early prolonged ART thus limits the size of the HIV reservoir, protects long-lived cells from persistent infection and may enhance post-treatment control.
Combination antiretroviral therapy (cART) reduces HIV-associated morbidities and mortalities but cannot cure the infection. Given the difficulty of eradicating HIV-1, a functional cure for ...HIV-infected patients appears to be a more reachable short-term goal. We identified 14 HIV patients (post-treatment controllers PTCs) whose viremia remained controlled for several years after the interruption of prolonged cART initiated during the primary infection. Most PTCs lacked the protective HLA B alleles that are overrepresented in spontaneous HIV controllers (HICs); instead, they carried risk-associated HLA alleles that were largely absent among the HICs. Accordingly, the PTCs had poorer CD8+ T cell responses and more severe primary infections than the HICs did. Moreover, the incidence of viral control after the interruption of early antiretroviral therapy was higher among the PTCs than has been reported for spontaneous control. Off therapy, the PTCs were able to maintain and, in some cases, further reduce an extremely low viral reservoir. We found that long-lived HIV-infected CD4+ T cells contributed poorly to the total resting HIV reservoir in the PTCs because of a low rate of infection of naïve T cells and a skewed distribution of resting memory CD4+ T cell subsets. Our results show that early and prolonged cART may allow some individuals with a rather unfavorable background to achieve long-term infection control and may have important implications in the search for a functional HIV cure.
Understanding factors affecting the size and the evolution of the HIV reservoir is essential for the development of curative strategies. This study aimed to assess the impact of antiretroviral ...therapy (ART) initiated during primary infection (PHI)
chronic infection (CHI) on the levels and dynamics of integrated HIV-1 DNA, a biomarker of viral persistence.
Integrated and total HIV-1-DNA were measured in the blood of 92 patients treated during PHI (early group) and 41 during CHI (deferred group), at diagnosis, ART initiation, and 12-24 months on treatment.
On ART, detectable (>1.78 log
copies/10
PBMCs) integrated HIV-1 DNA levels were significantly lower in the early
deferred group (2.99 log
3.29 log
= 0.005). The proportion of undetectable integrated HIV-1 DNA tended to be higher in the early group
deferred group (61 %
46 %; p = 0.133).
Treatment initiated at PHI limits the levels of integrated HIV-1 DNA in blood. However, initiating treatment at CHI does not allow reaching such low levels in most patients, probably because the stable proviruses at that stage are present in the less prone to elimination long-lived cells. Thus, early ART could provide an opportunity to preparing for functional cure and eradication strategies.
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