iconstar paper   HIV Articles  
Back grey arrow rt.gif
 
 
Impact of monotherapy on HIV-1 reservoir, immune activation, and co-infection with Epstein-Barr virus
 
 
  Download the PDF here
 
September 19, 2017
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0185128
 
Thirty-two patients >18 year old with documented history of HIV-1 infection, stable cART, undetectable plasmaviremia (HIV-1 RNA <50 copies/ml) and good immunological status (CD4 >350 cell/μl and nadir CD4 >100 cell/μl) for at least 6 months were switched from cART to mART with one PI (Lopinavir/ritonavir).
 
In conclusion, the results of the study outlined here show that mART has a lower control on HIV-1 reservoir and immune activation than cART. The higher risk of intermittent viremia or virological failure episodes in mART strategy may be due to a persistent perturbation of the immune system in response to a persistent replication of HIV-1 reservoir and vice versa. These findings underline the fact that patients treated with mART should be studied not only for classical parameters, i.e. HIV-1 plasmaviremia and CD4 cell count, but also for HIV-1 reservoirs and immune activation. Moreover, the long-term monitoring of B-cell activation is important to avoid the risk of EBV-related malignancies.
 
Conclusions
 
Long-term mART is associated with higher levels of T- and B-cell activation and, conversely to cART, does not reduce the size of HIV-1 reservoir and EBV co-infection. Notably, levels of mtDNA remained stable in the cART group, but significantly rose in the mART one (p<0.001). [from Jules: so what is Dual ART affect on HIV-DNA, we don’t know?]
 
During follow-up, no cART patients had virological failure, while one mART-treated patient had one HIV-1 RNA blip (defined as a transitory episode of HIV-1 RNA >50 copies/ml preceded and followed by plasmaviremia <50 copies/ml), one mART-treated patient had two HIV-1 RNA blips and 3 mART-treated patients had virological failure (defined as two consecutive HIV-1 RNA >50 copies/ml), at week 12 (1 patient) and at week 36 (2 patients) of follow-up. The patients regained virological control by adding the previous NRTI-backbone and were excluded from the mART group when they were returned to cART. No significant adverse effects and no deaths were observed in the two groups of patients during the study period.
 
HIV-1 reservoir
 
At baseline, the median (interquartile range-[IQR]) levels of HIV-1 DNA and intracellular HIV-1 RNA were similar in the two groups (Fig 1A and 1B). Overall, beyond individual patients’ variability, HIV-1 DNA levels remained stable in the mART group throughout the study period (24 [10-112], 55 [16-169], and 33 [10-186] copies/106 PBMC at baseline (T0), week 48 (T48), and week 96 (T96), respectively; p = 0.778), but slightly decreased in the cART group (16 [10-137], 31 [10-59], and 10 [10-10] copies/106 PBMC at T0, T48, and T96, respectively; p = 0.050) (Fig 1A). The intracellular HIV-1 RNA levels did not significantly vary in the mART group (680 [161-1318], 441 [102-1374], and 1035 [552-1836] copies/106 PBMC at T0, T48, and T96, respectively; p = 0.733), while they decreased in the cART group (835 [385-1274], 595 [184-1261], and 358 [204-753] copies/106 PBMC at T0, T48, and T96, respectively; p = 0.037) (Fig 1B). At the end of follow-up, the mART-treated patients had significantly higher levels of HIV-1 DNA and intracellular HIV-1 RNA with respect to the cART-treated patients (Fig 1A and 1B).
 
At the end of follow-up, the mART-treated patients had significantly higher levels of HIV-1 DNA and intracellular HIV-1 RNA with respect to the cART-treated patients (Fig 1A and 1B).
 
Immune activation
 
At baseline, percentages of activated T cells (CD3+CD8+CD38+) [21,23] were similar in the two groups (Fig 2A). The activated T cells significantly increased in the mART group during the follow-up (3.58 [2.31-4.60], 3.73 [2.36-5.33], and 4.46 [3.23-6.79] %CD3+CD8+CD38+ at T0, T48, and T96, respectively; p = 0.014), but they did not vary significantly in the cART-treated patients (3.72 [1.80-4.38], 2.81 [2.03-4.74], and 3.41 [1.54-4.85] %CD3+CD8+CD38+ at T0, T48, and T96, respectively; p = 0.870) (Fig 2A). At the end of follow-up, percentages of activated T cells were significantly higher in the mART-treated than in cART-treated patients (Fig 2A).
 
Microbial translocation

 
At baseline, there were no differences in any of the microbial translocation markers studied (mtDNA, 16S rDNA, sCD14 and LPS) (Fig 3A, 3B, 3C and 3D). During follow-up, the levels of mtDNA significantly increase in the mART-treated patients (1254 [271-2546], 1221 [425-3153], and 4482 [2170-5213] copies/μl at T0, T48, and T96, respectively; p<0.001), while they did not significantly vary in the cART-treated patients (2030 [377-4424], 2213 [637-8104], and 2858 [983-4352] copies/μl at T0, T48, and T96, respectively; p = 0.940) (Fig 3A). No significant changes were observed in the other markers during the follow-up (Fig 3B, 3C and 3D). At the end of follow-up, mART-treated patients had significantly higher levels of mtDNA than cART-treated patients (Fig 3A).
 
Co-infection with EBV-DNA
 
At baseline, the levels of intracellular EBV-DNA were similar in the mART and cART groups (Fig 4). During follow-up, the levels of EBV-DNA did not vary within the mART group (589 [285-1558], 433 [10-1190], and 550 [12-1459] copies/106 PBMC at T0, T48, and T96, respectively; p = 0.725), while they decreased significantly in the cART-treated patients (398 [156-1470], 200 [140-565], and 24 [10-270] copies/106 PBMC at T0, T48, and T96, respectively; p = 0.006) (Fig 4). At the end of follow-up, levels of EBV-DNA was higher in the mART than in cART group (Fig 4).
 
Discussion

 
A body of evidence indicate that mART in HIV-1-infected patients is associated to a higher rate of intermittent viremia and viral rebound with respect to that observed in cART-treated patients [2, 4-7]. Although this would suggest that mART has a lower impact on HIV-1 reservoir, the data produced using HIV-1 DNA load on PBMC as a marker of HIV-1 reservoir size are controversial [11-13].
 
In the present study, the HIV-1 reservoir size was estimated by measuring both HIV-1 DNA and intracellular HIV-1 RNA, a marker of ongoing viral activation/replication that may continuously refill the viral reservoir. During the study period, beyond individual patients’ variability, we found an overall slight decline of both markers in the cART group, whereas they remained fairly stable in the mART-patients. Thus, while cART treatment seemed to continuously reduce the size of the HIV-1 reservoir, confirming previous findings [25], a subliminal HIV-1 replication may contribute to refilling the HIV-1 reservoir in mART-treated patients. Chronic immune activation/inflammation is a hallmark of HIV-1 pathogenesis [10]. The findings that the percentage of both activated T and B cells rises in mART-treated patients, supports the hypothesis that mART has a lower impact on the HIV-1 reservoir. Since memory T cells are the preferential target for HIV-1 their activation may contribute to maintaining the viral reservoir by both activating HIV-1 replication and expanding latently infected cells [26]. A potential limitation of this study was the sole use of the CD38 marker on CD8 T cells to estimate their activation, and, unfortunately, the limited number of cells did not allow us to analyse the status of CD4 T cell activation/proliferation, even with other markers.
 
The activation of T and B cells is mainly triggered by microbial products. Indeed, pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) induce a potent innate immune response through the engagement of several Toll-like receptors (TLRs), which lead to T and B activation [27]. Notably, we did not find significant changes in PAMPs levels neither in the cART, as expected, nor in the mART-treated patients. Thus, the control of HIV-1 replication induced by mART may be sufficient to prevent the massive HIV-1-induced T-cell depletion that causes damage to the intestinal mucosa, promoting translocation of microbial products into circulation [27,28].
 
Interestingly, we found that in mART-treated patients there was an increase in levels of mtDNA. It has been suggested that NRTI-based cART is associated to diminished mitochondrial functions [29], as these drugs may interfere with mtDNA polymerase γ and interrupt mtDNA replication, leading to its depletion [30]. The presence of NRTIs may thus explain why cART-treated patients have lower mtDNA levels than patients treated only with PI. Once released from cells, mtDNA levels, significantly higher in mART-treated than in cART-treated patients, may substantially contribute to stimulating T and B cells. To our knowledge, this is the first time that activation of B cells has been investigated in mART-treated patients. Chronic B-cell stimulation and expansion of EBV-infected B-cells [31-34] may increase the risk of EBV-related malignancies [16, 35, 36]. The hyperactivation of B cells by HIV-1 is characterized by a higher expression of activation markers (CD80 and CD86) [31] and of activated memory B cells (CD21lowCD27+ B cells) which have undergone HIV-1-induced activation and differentiation to plasmablasts [24]. We found that levels of activated B-cell significantly increased over time in the mART-treated patients, while no differences were noted in the cART-treated ones. Moreover, while levels of EBV-DNA decreased in the cART group, they remained stable in the mART group. Thus, the control of immune activation status seems to play a role not only in reducing HIV-1 reservoir, but also in lowering EBV-DNA levels. By contrast, the mART, which is not as efficient as cART in controlling DAMPs levels and B-cell hyperactivation, may promote EBV reactivation and/or polyclonal expansion of EBV-infected B cells, favoring the onset of EBV-related malignancies.
 
In conclusion, the results of the study outlined here show that mART has a lower control on HIV-1 reservoir and immune activation than cART. The higher risk of intermittent viremia or virological failure episodes in mART strategy may be due to a persistent perturbation of the immune system in response to a persistent replication of HIV-1 reservoir and vice versa. These findings underline the fact that patients treated with mART should be studied not only for classical parameters, i.e. HIV-1 plasmaviremia and CD4 cell count, but also for HIV-1 reservoirs and immune activation. Moreover, the long-term monitoring of B-cell activation is important to avoid the risk of EBV-related malignancies. We are aware that limitation of this study is the low number of patients studied; a larger clinical studies are warranted to confirm these results.
 
Fig 1. HIV-1 reservoir levels in mART-treated and cART-treated patients. HIV-1 DNA (A) and intracellular HIV-1 RNA (B) levels in mART and cART patients at baseline, at 48 and 96 weeks of follow-up. Each symbol represents one patient. The lines indicate the median and the 25±75th percentiles.

graph

 
 
 
 
  iconpaperstack View Older Articles   Back to Top   www.natap.org