CHI) and primary HIV-1 infection (PHI), since specific recommendations for PHI are also provided in France.Guidelines for initiating HIV treatment are regularly revised. We explored how physicians in France have applied these evolving guidelines for ART initiation over the last decade in two different situations: chronic (Data came from the ANRS PRIMO (1267 patients enrolled during PHI in 1996–2010) and COPANA (800 subjects enrolled at HIV diagnosis in 2004–2008) cohorts. We defined as guidelines-inconsistent during PHI and CHI, patients meeting criteria for ART initiation and not treated in the following month and during the next 6 months, respectively.3 in 2006, only 55% of the patients with CD4≤350 were treated and 66% in 2008. During CHI, ART was more frequently initiated in patients who met the criteria at entry (96%) than during follow-up: 83% when recommendation to treat was 200 and 73% when it was 350 cells/mm3. Independent risk factors for not being treated during CHI despite meeting the criteria were lower viral load, lower educational level, and poorer living conditions.ART initiation during PHI dramatically decreased from 91% of patients in 1996–99 to 22% in 2007 and increased to 60% in 2010, following changes in recommendations. In 2007, however, after the CD4 count threshold was raised to 350 cells/mmHIV ART initiation guidelines are largely followed by practitioners in France. What can still be improved, however, is time to treat when CD4 cell counts reach the threshold to treat. Risk factors for lack of timely treatment highlight the need to understand better how patients’ living conditions and physicians’ perceptions influence the decision to initiate treatment. Combined antiretroviral therapy (ART) has substantially reduced morbidity and mortality in HIV-infected individuals since its introduction in 1996 333 emerged 333.At the beginning of the HAART era in 1996, treatment initiation in chronically HIV-infected patients was largely recommended in France 3. The new European AIDS Clinical Society (EACS) guidelines 3. However, as in France, ART is recommended when the CD4 count is below 500 cells/mm3, or even sooner for pregnant women and people with comorbidities such as HIV-associated kidney disease, neurocognitive impairment, human papillomavirus (HPV)-associated cancers , hepatitis B or C coinfection.The 2010 WHO guidelines 3 in 2006 and since 2010 ART initiation has been recommended at a CD4 count below 500 cells/mm3, as in chronic infection. Moreover, in the early era of cART, because of concerns regarding toxicity, PHI treatment was recommended for 18–24 months only. Treatment interruption has not been recommended since 2008.Most countries do not have systematic recommendations regarding treatment initiation during primary HIV infection (PHI). In the US guidelines, treatment initiation during PHI is obligatory only if the patient is pregnant. In France, initially no specific recommendations were made for PHI before 2002; since then specific recommendations are regularly published, and are based on CD4 count, the presence of more than 3 clinical symptoms, severe neurological abnormalities or pregnancy. What has changed in the last decade is the CD4 cell count for treatment initiation: the CD4 cut-off was raised from 200 to 350 cells/mmprimary HIV infection (PHI) and chronic HIV infection (CHI). Data came from two long-term follow-up studies: the ANRS PRIMO The aim of this study was to explore how physicians in France have applied clinical practice guidelines for antiretroviral therapy initiation during the last decade in two different situations: The ANRS PRIMO cohort is the largest cohort of HIV patients enrolled during PHI. A total of 1267 HIV-1-infected adults were enrolled between June 1996 and December 2010 in 85 hospitals in France. Primary infection was confirmed by an incomplete Western blot, or detectable p24 antigenemia or detectable plasma viral load with a negative or weakly reactive enzyme-linked immunosorbent assay (ELISA), or an interval of less than 6 months between a negative and positive ELISA test. Clinical and biological data were collected at Day 0, months M1, M3, M6 and every 6 months thereafter The ANRS COPANA study enrolled 800 patients newly diagnosed with HIV between 2004 and 2008, in 36 hospitals in France In both studies patients had to be antiretroviral-naïve at enrollment. All the participants gave informed written consent and the two studies were approved by the Paris-Cochin Ethics Committee. No specific recommendations for treatment initiation were given in these 2 cohorts, apart from regularly revised recommendations.primary-HIV infection (PHI); and during chronic-HIV infection (CHI).Guidelines-concordant clinical practice was studied in 2 different situations: ART initiation during 3 in 2004; <350 cells/mm3 in 2006 and less than 500 cells/mm3 in 2010. No specific recommendations were made between 1996 and 2002. We defined as guidelines-inconsistent ART initiation during PHI the cases of patients who met criteria for ART initiation according to contemporary recommendations but were not treated in the month following PHI diagnosis.In order to evaluate guideline-concordant clinical practice of ART initiation during PHI we defined time periods during which major changes in ART initiation recommendations were made in France: treat if CD4 count <200 cells/mm3 in 2002 and ≤350 cells/mm3 in 2008. All patients with clinical AIDS were to be treated. Cases of patients eligible for treatment but not treated in the next 6 months, while still on active follow-up, were considered as guidelines-inconsistent ART initiation during CHI.In chronic HIV infection, ART initiation guidelines evolved as follows: ART started if CD4 count <200 cells/mm3 in July 2008 for chronic patients, and we considered as eligible for treatment a patient whose first CD4 count below 350 cells/mm3 occurred in 2009.In France, HIV guidelines are published in July and we considered as eligible for treatment patients meeting criteria for treatment initiation from the year that followed the change in the recommendations. For example, the CD4 threshold was raised to 350 cells/mmThe characteristics of the patients of the two cohort studies were described. Continuous covariates were categorized according to the median of observed values or using published cut-off values where appropriate.The percentage of patients treated within 1 month following PHI diagnosis according to the calendar period and the CD4 cell count  was illustrated graphically.ART initiation in chronic HIV infection was studied by combining patients from the two cohorts. We defined patients with CHI as those enrolled in the COPANA cohort and patients from the PRIMO cohort still untreated at 6 months following PHI diagnosis. The analysis was conducted separately depending on whether the criteria for ART initiation were met at entry or occurred during follow-up from Month 6.Factors associated with guidelines-inconsistent ART initiation were identified using the Chi2 test and the Wilcoxon rank-sum test for dichotomous and continuous variables, respectively. The odds-ratios (ORs) of factors associated with guidelines-inconsistent ART initiation were estimated using a logistic regression model. Covariates with a P-value ≤0.2 were included in the multivariate analysis.Analysis was performed with SAS version 9.1 . A P-value <0.05 was considered statistically significant.). Patients who were enrolled at PHI were predominantly male, native French, with a higher CD4 cell count and a higher HIV viral load than subjects enrolled during CHI. Patients diagnosed during chronic infection were more frequently native Africans infected through heterosexual intercourse. Low CD4 cell count (<200 cells/mm3) was seen in 4% and 20% of patients at PHI and CHI diagnosis, respectively. Fifty-five percent of the patients during PHI had an HIV viral load higher than 5 log copies/mL versus 25% during CHI. The percentage of HCV and HBV co-infection was overall less than 5%.A total of 2067 patients were included in the ANRS PRIMO and COPANA cohorts, with a median of follow-up of 4.0 years (IQR 1.7–7.5) and 4.4 (IQR 3.1–5.5), respectively. As expected, the two populations differed in several baseline characteristics Table 1. Before 2002, no specific guidelines were available and overall 79% of the patients were treated in the month that followed PHI diagnosis. Of note, the percentage began to decrease in 2000, before official guidelines changes: 91% of the patients were treated in 1996–1999 vs. 67% in 2000–2002 The percentage then gradually declined to 22% in 2007, and afterwards the trend was reversed with ART initiation during PHI gradually increasing over time to 60% overall in 2010 .3; IQR: 49; 165) than those who met these criteria during follow-up , p<0.0001. This was also the case when CD4 cell count was the only criteria for treatment initiation  vs. 177  respectively). No other significant differences between the two groups were observed after adjustment for CD4 cell count at the time criteria for initiating treatment were met.Here we considered chronically infected patients meeting criteria for treatment initiation from the year that followed the change in recommendations (n = 254). The 113 patients who met criteria for ART initiation at study entry initiated ART more often in the next 6 months than the 151 who attained these criteria during follow-up  Table 2.3. The treatment initiation coverage subsequently observed in 2009–2010 was only 73% , p = 0.11), educational level<university , p = 0.02), living at their family or friends’ place  , p = 0.01) and calendar period 2009–2010 vs. 2005–2008 , p = 0.03). Women were not less treated than men in multivariate analysis , p = 0.19). The clinic center effect was no longer significant. No significant interaction was found between calendar period and all the considered factors. Furthermore, adjustment for the cohort (COPANA versus PRIMO) did not modify the results.Here we provide an insight into how practitioners have applied guidelines for ART initiation in HIV clinical practice during the last 15 years in France, in two contrasted situations, PHI and CHI.3 in 2006. There was a degree of inertia in applying these new guidelines, since it took more than 1.5 years after their publication to reach just 66% of treatment initiation coverage for patients with a CD4 cell count <350 cells/mm3.We observed dramatic changes in ART initiation during PHI over time, from 91% in 1996–99 to only 22% in 2007 and then 60% in 2010, following changes in recommendations. The overall trend showed that concordance with guidelines was not optimal after recommendations of ART initiation raised the CD4 cell count to 350 cells/mm3. In 2009–2010, the level of ART initiation in patients with CD4 cell count ≤350 cells/mm3 was still suboptimal (73%); fortunately during this period all patients with clinical AIDS or CD4 cell count ≤200 cells/mm3 had timely initiation of ART. Our study shows that the practitioner’s and the patient’s perception of the need for treatment initiation during CHI depends on whether the CD4 threshold is reached at diagnosis or during follow-up. ART initiation was 96% in patients who had reached the CD4 cell count threshold to treat at entry, but was less timely when the CD4 threshold was reached during active follow-up. This better adherence to recommendations at entry than during follow-up might be partly explained by the lower level of CD4 at entry, and/or by less opportunity for interactions with the patient when criteria for ART initiation are met at the first visit.In CHI the same “inertia effect” as in PHI was observed in 2008, when ART initiation was firmly recommended in asymptomatic patients with CD4 cell count <350 cells/mmIn other medical fields , significant gaps have also been found between clinical practice and guidelines 3) which physicians may have considered less urgent than the treatment of more advanced immunodeficiency (CD4<200 cells/mm3). Patients therefore received less timely treatment in 2009–2010 than in 2005–2008. Discrepancies between national and international recommendations may be implied as well. Currently, ART initiation is recommended for asymptomatic HIV patients with a CD4 cell count <500 cells/mm3 in France and more recently in the USA, while European AIDS Clinical Society (EACS) guidelines 3. A better concordance between national and international guidelines might improve practitioners’ responsiveness to implementation of guidelines.Barriers to adherence to practice guidelines by physicians can be categorized as follows: barriers affecting the physician’s knowledge (lack of awareness or lack of familiarity), attitudes , or behavior  In our study, patients with guidelines-inconsistent ART initiation had a lower HIV viral load. It appears that a viral load <5 log copies/mL led some physicians to delay ART initiation, even when the CD4 threshold was reached. Similar results were found in the Swiss cohort when exploring adherence to recommended combinations of first-line ART regimens according to US guidelines These social parameters may influence the patient’s but also the physician’s perception of the need for treatment. The Swiss cohort study found different results, but investigated not treatment initiation but rather adherence to recommended first-line regimens: more educated patients were more often treated with a regimen at odds with the guidelines 3).It is important to note that systematic reviews and practice guidelines are population-centered and not patient-centered, which makes individual treatment decisions not as easy as might be hoped. Furthermore, before the drawing up of evidence-based recommendations, there is clinical practice experience. In this study, we observed a subset of patients who were treated in anticipation of changes in recommendations or whose treatment even led to such changes  to investigate provider-related factors. Another limitation was the lack of routine recording of the reasons for not initiating ART. A specific study of the reasons for nonadherence to guidelines might shed light on the problem so that more effective interventions can be implemented to overcome barriers that prevent physicians from adhering to guidelines, as in In conclusion, guidelines for initiation of ART in HIV infection are largely followed by practitioners in France. There is, however, room for improvement in the time to treatment when CD4 cell counts reach the threshold to treat. Risk factors for not receiving timely treatment highlight the need for better understanding of how patients’ living conditions and physicians’ perception influence the decision to initiate treatment. In the new area of treatment as prevention (TasP) The ANRS PRIMO and COPANA cohorts are funded by the ANRS. No current external funding sources for this study.


Treatment is switched before the overgrowth and archiving of mutant strains that carry resistance against the induction regimen and would limit its future re-use. In silico modelling shows that an optimal trade-off is achieved by switching treatment at The human immunodeficiency virus (HIV) can be suppressed by highly active anti-retroviral therapy (HAART) in the majority of infected patients. Nevertheless, treatment interruptions inevitably result in viral rebounds from persistent, latently infected cells, necessitating lifelong treatment. Virological failure due to resistance development is a frequent event and the major threat to treatment success. Currently, it is recommended to change treatment after the confirmation of virological failure. However, at the moment virological failure is detected, drug resistant mutants already replicate in great numbers. They infect numerous cells, many of which will turn into latently infected cells. This pool of cells represents an archive of resistance, which has the potential of limiting future treatment options. The objective of this study was to design a treatment strategy for treatment-naive patients that decreases the likelihood of early treatment failure and preserves future treatment options. We propose to apply a single, pro-active treatment switch, following a period of treatment with an induction regimen. The main goal of the  In 1996, the tremendous clinical success of highly active antiretroviral therapy had led many researchers to believe that the eradication of HIV would be feasible. However, it was soon realized that inducible pro-virus persists in latently infected cells despite ongoing therapy and that the latent reservoir prevents HIV eradication within the patients lifetime Latent infection is established when Despite the impressive improvement of antiviral therapy, many patients still experience virological failure caused by the selection of drug resistant virus populations. Current guidelines recommend changing treatment after the confirmation of virological failure. However, in the face of the rapid viral turnover this approach could be sub-optimal Induction-maintenance (IM) approaches are used for the treatment of a growing number of infectious- and neoplastic diseases Another approach to overcome the development of resistance is to alternate antiretroviral therapy We propose an approach that combines the advantages of conventional IM- and treatment alternation strategies, but minimizes their inherent disadvantages. We suggest a single, pro-active treatment switch from an inducer drug combination to a maintenance combination. The inducer drug combination should rapidly lower the viral population size and eliminate resistant mutants. Subsequently, it will be replaced by a maintenance drug regimen with a completely different resistance profile, before drug resistant strains are archived.We have previously introduced a novel model of virus dynamics and adaptation We have extended the existing viral dynamics model, described in prior to proviral genomic integration , respectively. A histogram of the derived  switching times from a total of 6000 simulations is shown in next drug combination (maintenance combination). We therefore evaluate In The derived switch-times are displayed in In order to reflect the clinical practice of HIV care, we have implemented the following routine for assessing the efficacy of the applied treatment combinations.Our virtual patients are monitored every month for efficacy assessment until virus levels fall below the limit of detection (50 HIV RNA/mL plasma). Thereafter, they are monitored every other month. Virological failure has been defined according to treatment guidelines We implemented conventional HAART in the following way: The virtual patients are initially treated with a drug combination consisting of two nucleoside reverse transcriptase inhibitors (NRTIs) and one non-nucleoside reverse transcriptase inhibitor (NNRTI) (e.g. tenofovir (TDF) + emtricitabine (FTC) + efavirenz (EFV)), until virological failure is detected, in which case treatment is changed to a second line regimen consisting of a protease inhibitor (PI), an integrase inhibitor (InI) and an entry inhibitor (EI) (e.g. ritonavir (RTV) -boosted PI + raltegravir (RLV) + maraviroc (MVR)).In the proposed induction-maintenance-strategy, patients are initially treated with a combination consisting of a PI, an InI and an EI, until In the following, we performed 1000 hybrid stochastic-deterministic simulations for each relevant parameter set  a very simple treatment strategy that can lead to significant reductions in virological failure in comparison to conventional HAART treatment. A unique drug combination (inducer combination) is used for a short time (80 days) and pro-actively switched to a maintenance combination. The purpose of the inducer combination is to decrease viral population size and thereby increase the likelihood that the subsequent therapy (maintenance) will achieve durable suppression. Clinical implementation of this novel treatment strategy requires only one additional clinical visit at 80 days in comparison with the conventional HAART therapy. The important finding of our study is, that although two drug combinations are always utilized during the proposed induction-maintenance strategy, less archiving of drug resistance occurs in comparison with a conventional treatment strategy, where a second treatment line would be applied only in the case of virological failure or toxicity. Less drug resistance archiving implies that more treatment options will be available for the follow-up and long-term management of HIV-infected patients when the proposed induction-maintenance treatment strategy is used (see current drug combination and would limit the further use of this drug combination (risk of the strategy), (ii) while at the same time, it has to be applied long enough to possibly eradicate viral mutants, which are resistant to the next drug combination (the benefit of the strategy).The time for switching between combinations de novo developed). As discussed above, we determine the abundance of mutants at the time of therapy initiation by utilizing the deterministic fix-point as starting condition for our simulations. We have shown the non-inferiority of our approach in in vivo for drugs with a large genetic barrier in vivo the inducer combination could possibly be applied for a longer time frame than the 80 days utilized in our model, if the genetic distance between wildtype and fully drug resistant mutant was greater than considered here (greater than one point mutation). However, our results demonstrate that even a very short time (80 days) in which the inducer combination is applied, can improve the clinical outcome significantly  and also on their genetic distance to the wildtype  are cleared in vivo data, Chun et al. One limitation of the proposed induction-maintenance strategy is the potential inability to eliminate viral strains, that carry resistance to the maintenance therapy. This is particularly the case, if viral mutants, which carry resistance against all (or at least the majority of) drugs in the maintenance combination, are archived in the latent reservoir prior to treatment initiation. In Infection with drug resistant strains, mainly against established drug classes, is a major, growing health concern For our strategy, we have chosen drugs from novel classes  for the inducer-combination, while we selected drugs from well-established classes for the maintenance combination . This has the following rationale: The inducer combination will only be applied for a short time (80 days), while the maintenance combination could possibly be applied for much longer periods of time . Second or third generation drugs within the established drug-classes are often more convenient to apply (e.g. once daily dosing) and are less toxic, which has important implications for the long-term management of HIV Some drug classes can cause a distinct viral load decline. In particular, the only approved InI raltegravir causes a more rapid viral load decay, compared with other HIV inhibitors Intuitively, it might be more advantageous to use drug resistance tests to guide treatment switches, instead of using a fixed time for a pro-active switch from inducer- to maintenance combination in silico study, we considered time-invariant, as well as anatomically homogeneous average drug efficacy In our Based on a recent, successful pre-exposure prophylaxis (PrEP) trial, where emtricitabine (FTC) + tenofovir (TDF) were given to high-risk individuals prior to proviral genomic integration  setting for numerical simulation. This approach (i) takes into account stochastic fluctuations in the slow reaction processes; and (ii) reduces the computational costs for the simulation of the fast (deterministic) system dynamics. We used the direct hybrid method proposed in In brief, the hybrid method comprises the following algorithmic workflow:Set initial time Generate two uniformly distributed pseudo-random variables Set Take the integer Update Set atlab−6 and an absolute error tolerance of 10−9. Our simulation code is provided in The above algorithmic scheme requires the use of numerical integrators that allow to stop integration in step (3) when a stochastic reaction event is detected at a time For the model above (eq. (3)), the reproductive numbers, which indicate the expected number of offspring in the next generation, are defined as follows: the reproductive number Since infected cells are also pathogens, which can lead to a rebound of the disease even in the absence of any virus, we also determined their basic reproductive numbers under a given treatment We have previously introduced the reproductive capacity Figure S1Time and probability of virological failure depends on pool-size of archived drug-resistant virus. A: The median time until virological failure, in relation to the number of fully-resistant archived virus . B: Probability that virological failure occurs within two years after initiation of HAART therapy as a function of the number of fully-resistant archived virus. 500 stochastic-deterministic runs were performed for each pool size of the latently infected drug-resistant reservoir. Parameter values used: (PDF)Click here for additional data file.Table S1Determination of relevant parameter space for further investigation. We assessed virological failure rates after one year of triple drug therapy for varying values of efficacy (PDF)Click here for additional data file.Source Code S1The File ‘HAART.m’ can be used to simulate the kinetics of HIV after application of conventional HAART treatment in MATLAB.(M)Click here for additional data file.Source Code S2The File ‘HIVmodel.m’ builds the original HIV model used throughout the manuscript for use in MATLAB.(M)Click here for additional data file.Source Code S3The File ‘InductionMaintenance.m’ can be used to simulate the kinetics of HIV after under the proposed induction-maintenance therapy in MATLAB.(M)Click here for additional data file.Source Code S4The File ‘PatientMonitoring.m’ contains the MATLAB implementation of routine patient monitoring.(M)Click here for additional data file.Source Code S5The File ‘ReadMeFirst.txt’ Contains a description of all supplied source code files, contact details, information on runtime and execution and a copy of the GNU public license.(TXT)Click here for additional data file.Source Code S6The File ‘SpeciesLevelsIndices.pdf’ contains an interpretation of the output generated by executing the provided MATLAB Source Code Files –S4.(PDF)Click here for additional data file.


Modern antiretroviral therapy (ART) is assumed to allow a certain degree of nonadherence while still maintaining complete suppression of viral replication ("forgiveness"), as virological suppression, measured by the commercial plasma viral load assays, is common at adherence levels >55-70% . Yet, it+ count, 620 cells/mm3). For every patient, three longitudinal samples, taken with 3-4 month intervals, were analyzed. One-week mean adherence to ART (percentage of prescribed doses taken) prior to the sampling moments was measured electronically.Levels of HIV-1 unspliced RNA (usRNA) and viral DNA were quantified by seminested real-time PCR -4 in PBMP=0.006). Remarkably, the change in adherence patterns from optimal through improving to poor was paralleled by a gradual increase in the corresponding viral RNA trends. Significant differences between the poor adherers and the remaining patients were observed in time-weighted changes from baseline (P=0.0006) and regression slopes (P=0.009) of usRNA, but not of viral DNA. These effects were independent of the therapy regimen or the time of virological suppression. Same effects were observed in a subset of patients whose plasma viremia was constantly undetectable (n=30).Adherence never fell below7O% in any patient, and concurrent plasma viral loads of 109/120 (91%) PBMC samples were undetectable (<50 cop/ ml); for 10/11 remaining samples they were <100 cop/ml. Longitudinally, 23 patients were constantly 100% adherent, eight demonstrated improving adherence in time, and nine ("poor adherers") showed decreasing, variable, or constantly <100% adherence. Notwithstanding the lack of virological rebound in any of the patients, poor adherence, but not optimal or improving adherence, caused a significant longitudinal increase in usRNA levels  on HIV RNA levels in PBMC strongly suggests new replication cycles despite ART, and not simply enhanced HIV-1 transcription in cells infected prior to therapy initiation. Our results represent the first evidence indicating that constant optimal adherence to modern ART may be necessary to stop all HIV replication.


Antiviral treatment with nucleoside analogs has been used for chronic hepatitis B (CHB). Each kind of nucleoside analog has its own characteristics and suitability for patients. Telbivudine  is the newest nucleoside analog, with strong and rapid viral suppression. However, its resistance rate is relatively high during long-term application, due to low genetic barriers to resistance. So, it is necessary to increase the effect and reduce resistance with effective management, according to baseline factors and early on-treatment responses.To reveal possible predictive factors of the effect of telbivudine (LdT) treatment on naïve HBeAg-positive chronic hepatitis B (CHB) patients to optimize treatment.A total 71 naïve chronic hepatitis B (CHB) patients who met the inclusion criteria were enrolled. All patients were treated with LdT 600 mg Qd for at least 52 weeks. Multiple logistic regression analyses were done to investigate the predictive values of baseline factors and responses at Week 24.The reduction in hepatitis virus B (HBV) DNA level was 6.44 ± 2.38 lg copies/mL at Week 52 compared with baseline. The complete virus response (CVR), biochemical response (BR), serological response (SR), and drug resistance (DR) were 61.99%, 77.46%, 35.21%, and 8.45% respectively. By multiple regression analysis, baseline alanine aminotransferase (ALT) levels significantly affected CVR , and baseline ALT and baseline HBV DNA levels were independent compact factors of SR . The differences in CVR, SR, and DR in patients with ALT > 120 Iu/mL compared with patients with ALT ≤ 120 Iu/mL were statistically significant. The differences in SR in patients with HBV DNA > 107 copies/mL compared with patients with HBV DNA ≤ 107 copies/mL were statistically significant. Additionally, CVR, BR, and SR were differed significantly between patients with HBV DNA lower than 300 copies/mL at Week 24 and patients with HBV DNA higher than 300 copies/mL .There were more responders among naïve HBeAg-positive chronic hepatitis B patients with lower HBV DNA levels  and higher ALT values  to LdT treatment. Adjustments for treatment strategy should be considered if HBV DNA > 300 copies/mL at Week 24 is observed. In recent years, much evidence has confirmed that antiviral treatment with nucleoside analogs of chronic hepatitis B (CHB) can inhibit viral replication, improve liver function, and reduce the incidence of decompensated liver diseases, including cirrhosis and liver cancer, and thus improve the prognosis of diseases. Each kind of nucleoside analog has its own characteristics and suitability for patients. So, it is necessary to find out the characteristics and impact factor of each medicine to improve its clinical application. Of the four available nucleoside analogs in China , telbivuIn this study, a total of 71 naïve HBeAg-positive CHB patients who received LdT monotherapy for 52 weeks were enrolled, and impact factors on effect and resistance were analyzed. We wished to identify the possible predictive factors of the effect of LdT treatment on naïve HBeAg-positive CHB patients to optimized the application of LdT.Patients who visited the outpatient clinic of West China Hospital of Sichuan University from March 2008 to August 2009 and met the inclusion criteria were enrolled in this study. The inclusion criteria were as follows: positivity for hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HbeAg) for at least 6 months and without history of antiviral therapy with nucleoside analogs; alanine aminotransferase (ALT) levels between 2 to 10 times the upper normal levels (UNL); and hepatitis virus B (HBV) DNA levels ≥ 105 copies/mL. Patients were excluded if they were coinfected with human immunodeficiency virus (HIV) or other hepatitis viruses  or had nonalcoholic fatty liver disease (NAFLD), alcoholic liver disease, posthepatitic cirrhosis, or hepatocellular carcinoma. The diagnosis of fatty liver depended on the characteristic rise in liver enzymes, including γ-glutamyltransferase; change in ultrasound Doppler; and other underlying diseases with metabolic disorder. The diagnosis of alcoholic liver disease depended on alcohol consumption, history of alcohol use, and a special imaging examination, including ultrasound Doppler and computed tomography. Liver cirrhosis was confirmed through invasive and noninvasive techniques, including liver stiffness measurement, special imaging examination, and liver biopsy. Hepatocellular carcinoma was diagnosed by elevated A fetal protein (AFP) and characteristic imaging manifestation, including computed tomography (CT) and magnetic resonance imaging (MRI). After patients signed an informed consent form, they were given LdT 600 mg daily as antiviral treatment for at least 52 weeks. During the treatment period, patients were not given any other antiviral drugs or medicines to protect liver function.Analyses of liver and renal functions, which included serum levels of total bilirubin (TB), ALT, albumin (ALB), blood urea nitrogen (BUN), blood creatinine (Cr), amylase, and lipase, were performed at baseline and at Weeks 12, 24, 36, and 52 of LdT treatment using an automatic biochemistry analyzer . The status of HBsAg, HBeAg, and antibodies to HBeAg (anti-HBe) was measured by microparticle enzyme-linked immunosorbent assay (ELISA) at baseline and Weeks 24 and 52. Serum HBV DNA was quantified by Cobas TaqMan Real-Time Polymerase Chain Reaction (PCR) Assay (Roche Diagnostics)  at baseline and Weeks 12, 24, 36, and 52, with a linear range between 300 and 109 copies/mL. LdT-associated mutations were assessed via direct sequencing if virological breakthrough occurred during the treatment. All assays were performed in the microbiological laboratories of West China Hospital, Sichuan University.Virological response (VR) was defined as the level of decline at the end of treatment. A complete virological response (CVR) was defined as a reduction in HBV DNA levels to less than the level of detection (< 300 copies/mL). Biochemical response (BR) was defined as a normalization of ALT levels. Serological response (SR) was defined as disappearance of HBeAg with or without the appearance of hepatitis B e antibody (HbeAb). Virological breakthrough was defined as an increase in serum HBV DNA > 1 log copy/mL compared to the on-treatment nadir that was confirmed in two consecutive tests. Drug resistance (DR) was defined as the emergence of virological breakthrough and the presence of drug-resistant mutations. The curative effect of LdT was assessed at Weeks 12, 24, and 52 during treatment. Rates of CVR, BR, SR, and DR were assessed at the time points above respectively. Baseline factors, including age, gender, ALT, and DNA levels, were analyzed for their effects on LdT treatment. HBV DNA levels at Week 24 were analyzed as impact factors during the treatmentQuantitative data were presented as the mean ± standard deviation (SD), categorical data were presented as counts and percentages, and HBV DNA levels were presented as log-transformation. Data were analyzed using SPSS, version 13.0 . ANOVA was used to analyze the reduction in HBV DNA levels at Weeks 12, 24, and 52 respectively. Pearson chi-square was used to compare CVR, BR, SR, and DR at Week 52 between patients with different baseline factors. Logistic regression analysis was used to investigate the baseline predictors for the response to treatment. Age, baseline DNA, and baseline ALT were quantitative variables. Gender was the qualitative variable. In all cases, tests of significance were 2-tailed, and statistical significance was defined as P < 0.05.A total of 71 patients were included, comprising 60 (84.5%) males and 11 (15.5%) females, with ages ranging from 17 to 46 years (mean 28 ± 8.337). Baseline data were as follows: the median level of HBV DNA was 1.25 ×108 copies/mL, and the mean ALT level was 183.99 ± 131.87 IU/mL.The reduction in HBV DNA levels was 4.71 ± 1.88, 5.84 ± 2.04, and 6.44 ± 2.38 lg copies/mL at Weeks 12, 24, and 52 respectively, compared with baseline . The difALT normalization was achieved in 39.44% (28/71), 64.79% (46/71), and 77.46% (55/71) of patients at Weeks 12, 24, and 52, respectively.The disappearance of HBeAg was observed in 21.13% (15/71), 28.17% (20/71), and 35.21% (25/71) of patients at Weeks 12, 24, and 52, respectively. HBeAg/Anti-HBe seroconversion was achieved in 9.86% (7/71), 16.90% (12/71) and 23.94% (17/71) of patients at Weeks 12, 24, and 52, respectively.By Week 52, viral breakthrough occurred in 7 (9.86%) patients, 6 of which (8.45%) had confirmed genotypic resistance, comprising 5 cases with rtM204I and 1 case with rtLl80M + rtM204V.Overall, LdT demonstrated a good safety profile. All levels of serum electrolytes were normal. Mild adverse reactions were observed in 4 (5.63%) patients, 3 of whom had a mild to moderate increase in blood creatine kinase without myalgia or muscle weakness at Week 24, 36, and 36, respectively. In the follow-up period, increased creatine kinase all reduced in the next 3 to 6 months without special treatment or drug withdrawal. Another patient suffered from insomnia, which disappeared after symptomatic treatment. No renal failure or pancreatitis occurred in any patient.As shown in All patients were divided into two groups: baseline ALT > 120 Iu/mL and ≤ 120 Iu/mL. At Week 52, the CVR, BR, SR, and DR of 41 patients with baseline ALT > 120 Iu/mL were 73.2% (30/41), 85.4% (35/41), 46.3% (19/41), and 2.4% (1/41), respectively. In the 30 patients with baseline ALT ≤ 120 Iu/mL, the CVR, BR, SR, and DR were 46.7% (14/30), 66.7% (20/30), 20.0% (6/30), and 16.7% (5/30), respectively. The differences in CVR, SR, and DR between the two groups were statistically significant . No significant difference was found in BR between the two groups  .All patients were divided into two groups: baseline HBV DNA > 107 copies/mL and ≤ 107 copies/mL. At Week 52, the CVR, BR, SR, and DR of 54 patients with baseline HBV DNA > 107 copies/mL were 59.3% (32/54), 77.8% (42/54), 27.8% (15/54), and 5.6% (3/54), respectively. In the 17 patients with HBV DNA ≤ 107 copies/ml, the CVR, BR, SR, and DR were 70.6% (12/17), 76.5% (13/17), 58.8% (10/17), and 17.6% (3/17), respectively. The differences in SR between the two groups were statistically significant . No significant difference was found in CVR, BR, or DR between the two groups  .For 32 patients with HBV DNA < 300 copies/mL at Week 24, the CVR, BR, SR, and DR at week 52 were 90.63% (29/32), 90.63% (29/32), 62.50% (20/32), and 6.25% (2/32), respectively. But, for the other 39 patients with HBV DNA ≥ 300 copies/mL at Week 24, the CVR, BR, SR, and DR at Week 52 were 38.46% (15/39), 66.67% (26/39), 12.82% (5/39), and 10.26% (4/39), respectively. The differences in CVR, BR, and SR between the two groups were statistically significant . But, the difference in DR between the two groups was not statistically significant  .The importance of optimizing therapy with nucleoside analogs lies in their long-term treatment course and inevitable resistance . How to This study investigated several baseline host and viral factors that may influence antiviral efficacy, including age, gender, and baseline HBV DNA and ALT levels. The multiple regression statistical analyses showed that patient age and gender had no impact on CVR, BR, SR, or DR. Baseline ALT level significantly affected CVR, suggesting that higher ALT levels lead to a higher CVR, similar to results of the GLOBE clinical trial  and otheThe optimization strategy also includes medication adjustments according to early response during treatment, except according to baseline factor . For difOf the 71 cases in this study, drug resistance occurred in 6 patients (8.45%). However, the observation period lasted only 52 weeks. With an extension of the treatment course, the occurrence of drug resistance may increase gradually. Therefore, it is necessary to monitor the occurrence of drug resistance with ongoing treatment. Usually, Ldt is regarded as a safe medicine, even for pregnant woman . In thisIn summary, our results suggest the following conclusions: LdT has good efficacy in naïve HBeAg-positive chronic hepatitis B patients, with a good safety profile. In particular, there were more responders among patients with lower HBV DNA levels, especially less than 107 copies/ml, and higher ALT values, especially higher than 120 Iu/ml at baseline, to LdT therapy. At the same time, it is crucial to focus on the response at Week 24 during the course of treatment. If HBV DNA level is still > 300 copies/mL at Week 24, alterations or adjustments to the treatment strategy should be considered. The primary limitations of this study are the relatively short observation time period and the small number of patients. However, the results of this study suggest some clues and indicators of clinical value in the 52-week observation period. It is worth performing further studies by continuing treatment for longer terms.


Nephrotoxicity of tenofovir (TDF)-containing HAART may be associated to several factors related to treatment or to patient's characteristics. We tried to assess Glomerular Filtration Rate (GFR) changes and their relationship with multiple clinical parameters in an Italian cohort receiving a TDF-containing HAART.OSMA-1 , a multicenter Italian study, was designed to evaluate since February 2008 the efficacy and the safety of TDF-based regimen in a real-life clinical setting. HIV infected, therapy naïve subjects were enrolled. GFR was estimated using Cockcroft-Gault (CG), Modification of Diet in Renal Disease (MDRD) and Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equations. Statistical analyses used a parametric test and a mixed model was used to analysis changes from baseline to 6, 12, and 24 months, with the different variables as fixed effect plus visit  and baseline value (continuous as covariate).We consecutively enrolled 172 patients . At baseline median CD4+ cell count was 225 cells/µL (range 2 - 701), HIVRNA > 100000 copies/mL in 43.6%. Median Body Mass Index (BMI) was 22.6 for males and 21.9 for females. A boosted PI was given in 60.5% of cases. At 6-mos evaluation, Women had greater declines in GFR vs. Men, independently from the GFR equation used . No further decline was documented after 6 months. Factors associated with demography or treatment are shown in Table Our data suggest that long-term use of TDF is associated with a modest decline in GFR that occurs predominantly at the beginning (in the first 6 months) of therapy in HAART-naive women. However, the decline is small, does not seem to worsen over time, and may not have a relevant clinical effect. A decreasing GFR was also found in elder patients, in smokers and when a boosted PI is used.


Raltegravir is the first integrase inhibitor to get into the clinic and the MK-0518 Expanded Access Program (EAP) started in Italy in 2007, so that most patients now have an observation period of two years. Up to date a few studies have described the durability of raltegravir-containing regimen with a follow up of 96 weeks [Patients from the MK0518 EAP were followed-up prospectively since enrolment in the study. Clinical and laboratory data were collected every 2 - 4 months after commercial availability.Out of 250 patients enrolled in the MK0518 EAP in our centres, 229 are still on the same regimen. The mean follow up was 80 weeks. At the time of present analysis (96 weeks) 133 patients were evaluated. Eighty-nine% of these had HIV-RNA < 50 Cp/mmc. Two patients temporarily stopped therapy, one developing the integrase 72I mutation plus 2 additional RT mutations, and both stably resuppressed the virus with the same regimen. Seven patients discontinued therapy due to virological failure: primary mutations for integrases were detected in all the samples  in addition to other mutations . Four patients with non-primary mutations detected continued the same therapy maintaining viral replication below 4000 copies/ml, with 1.9 log decrease in HIV-RNA. One patient was lost to follow up, four discontinued due gastrointestinal AEs and one for CK elevation, while eight died during the observation period, 4 of non-Hodgkin’s lymphoma (NHL), 1 of acute myocardial infarction, 2 of end stage liver disease and 1 post-transplantation for HCV-related cirrhosis. The immunological gain of the patients who remained on therapy is good in a salvage setting: +230 CD4/mmc.The MK0518 EAP seems to have contributed in a relevant measure to obtain full viral suppression for two years in most of our salvage patients, with an important immunologic gain and very few adverse events, in a situation were active companion drugs were really difficult to find out . The number of deaths (including NHL that represented the cause of death for 50% of the cases) has not been related to the drug regimen, but more likely to the advanced stage of HIV infection.


Adolescents have been identified as a high-risk group for poor adherence to and defaulting from combination antiretroviral therapy (cART) care. However, data on outcomes for adolescents on cART in resource-limited settings remain scarce.We developed an observational study of patients who started cART at The AIDS Service Organization (TASO) in Uganda between 2004 and 2009. Age was stratified into three groups: children (≤10 years), adolescents (11–19 years), and adults (≥20 years). Kaplan-Meier survival curves were generated to describe time to mortality and loss to follow-up, and Cox regression used to model associations between age and mortality and loss to follow-up. To address loss to follow up, we applied a weighted analysis that assumes 50% of lost patients had died.p<0.001), patients with a low initial CD4 cell count (p<0.001), patients with advanced WHO clinical disease stage (p<0.001), and shorter duration of time receiving cART (p<0.001). The crude mortality rate was lower for children , than adolescents  and adults .A total of 23,367 patients were included in this analysis, including 810 (3.5%) children, 575 (2.5%) adolescents, and 21 982 (94.0%) adults. A lower percentage of children (5.4%) died during their cART treatment compared to adolescents (8.5%) and adults (10%). After adjusting for confounding, other features predicted mortality than age alone. Mortality was higher among males (This study is the largest assessment of adolescents receiving cART in Africa. Adolescents did not have cART mortality outcomes different from adults or children. Programmatic evaluations of HIV/AIDS in resource-limited settings have historically focused on adult and child populations 13 In this relatively small sample, (n = 154), the authors reported significantly worse virological suppression in adolescents versus adults in a cohort of patients from nine southern African countries receiving privately purchased cart Adolescence can be a confusing time for youth, especially those living with a chronic and often stigmatized disease. A number of challenges have been identified that may compromise positive outcomes of care for adolescents. They may be particularly rebellious, may not have caregivers unlike younger children, and there many be challenges associated with puberty and disease Not all settings in Africa are alike, neither can we expect that outcomes in a privately purchased cART program will be similar to a publicly funded one. This study compares survival and loss to follow-up of adolescents to children and adults using a large dataset from a nationally representative cohort of HIV patients receiving free cART in Uganda.This study received ethical approval from TASO Administrative Research Board, a Uganda National Science and Technology Council approved board, and from University of British Columbia. Informed consent was not required as this was routinely collected operational data and the institutional review boards waived the need for consent.3The AIDS Support Organization (TASO) is the largest and oldest national non-governmental AIDS service organization in Africa. Founded in 1987, TASO currently provides psychosocial support, clinical care, and cART free of charge to people with HIV in Uganda. TASO started prescribing cART in 2004 and now provides treatment to approximately 24 000 patients throughout 11 clinical sites in Uganda. Criteria used for initiation of cART include World Health Organization (WHO) clinical disease stage III or IV or a CD4 cell count below 250 cells/mmThis study is based on data collected routinely for clinical monitoring and evaluation purposes at TASO. Clinicians complete standardized patient forms detailing patient demographics, clinical, psychosocial, and drug utilization data at each patient visit. These data are then entered into the TASO data collection database at each site by trained data capturers. All data are anonymized using a unique, confidential identification number.Clinic staff, nurses and clinical officers or physicians offer adherence monitoring and clinical support. For community-based recipients of care, a field monitoring team equipped with motorcycles is responsible for patient adherence, social support, and follow-up. This team, which includes medical attendants who conduct HIV testing, adherence counselling, clinical observation, and provide cART to patients, visits patients who fail to show for any appointment for three months or longer and patients who have requested home-based care.The present study includes all patients who initiated cART since the start of the programme (2004), irrespective of age. For each patient, the following baseline information was recorded: age at start of cART (years), gender , exact baseline CD4 cell count, WHO clinical disease stage, loss to follow-up status, date last seen for care, and where applicable, date of death.p-value of <0.05 considered significant. All analyses were conducted using SAS version 8 .The cohort was stratified into three age groups: children (age ≤10 years), adolescents (age 11–19 years), and adults (age ≥20 years). These age definitions have been used before by Nachega et al. p<0.001).A total of 23,367 patients were included in this analysis. There were 810 (3.5%) children, 575 (2.5%) adolescents, and 21,982 (94.0%) adults at baseline. Follow-up time differed between the age groups. Adults were followed for a median of 32 months , and children and adolescents were followed for 28 months  and 28 months , respectively (p<0.001). Baseline CD4 at the time of cART initiation was significantly different across the age groups (p<0.001): child patients started cART at a median CD4 count of 238 cells/µl , adolescent patients started at a median of 131 cells/µl,  and adult patients started at a median of 142 cells/µl . A lower percentage of children (5.4%) died during their cART treatment compared to adolescents (8.5%) and adults (10%). There were also no significant differences in WHO clinical disease stage and loss to follow-up between the 3 age groups.p<0.001), patients with a CD4 cell count lower than 100 cells/mm3 at cART initiation (p<0.001), patients with a WHO clinical disease stage of III or IV at cART initiation (p<0.001), and patients who started cART at an earlier time point (p<0.001). The crude mortality rate was lower for children , than adolescents  and adults .Overall, mortality was higher among males (p-value of the log-rank test was <0.001, suggesting an initial difference in the survival distributions between the age groups.p-value of the log-rank test was 0.134, suggesting there is no difference in the distributions of age for loss to follow-up.This study is the largest assessment of clinical outcomes among adolescents receiving cART in Africa. In this study, crude adolescent mortality was significantly different from child patients, yet, loss to follow-up was not. After adjusting for explanatory variables, we did not demonstrate a significance difference in either mortality or loss to follow up across groups.The recent report from southern Africa by Nachega et al. found that adolescents have worse outcomes compared to their adult counterparts in terms of virologic suppression and adherence As most adolescents would be infected at birth, many individuals would have died before achieving adolescence. As the number of adolescents enrolled in treatment grows and patient live longer, the question of adherence and retention will become increasingly important for health care practitioners. Often adolescents fall through the cracks between pediatric care and adult care. Specialized adolescent health care clinics providing counseling, testing, and treatment have been developed in countries like South Africa to meet the needs of HIV-positive and at-risk youth As with almost all assessments of mortality in African cohorts, we found that male gender was independently predictive of mortality. This finding is inconsistent with advocacy for increased attention to female issues and we hope that the medical and advocacy communities can promote an evidence-based strategy to responding to local epidemics. In our experience, and others, male patients are typically late to receive cART, have more advanced illness, and have worse clinical outcomes As with any study of this nature, there were several limitations. Loss to follow-up may have led to a misclassification of mortality as loss to follow up. TASO uses active retention strategies to locate patients who do not attend their scheduled appointments, thus reducing degree of lost to follow-up. We also attempted to overcome the issue of loss to follow-up in the present study with the use of our assumption that 50% of those lost had died 50% It should also be noted that CD4 cell count data at cART initiation, was not complete. The lack of complete CD4 cell counts is a reflection of the diverse settings in which TASO works in Uganda. This problem is also common in other resource-constrained settings Strengths of the study include the large sample size and long-term follow-up. The cohort includes patients receiving care throughout Uganda, and thus captures a wide range of differing patient experiences based on regional variation. Furthermore, the use of active retention to reduce loss to follow-up has resulted in higher patient retention rates than similar cohort studies.In conclusion, our study confirms earlier assertions that providing cART to adolescent patients is a complex undertaking. Adolescents have been overlooked in the literature and also in programming. They have unique needs that require tailored services and targeted research. As this population becomes increasingly important in the epidemic, further investigation into the causes of loss to follow up and mortality are needed for this subgroup of patients in order to design and evaluate supportive strategies for this vulnerable population.


We analyzed the latent reservoir, the residual viremia and episomal 2LTR DNA to examine the relationship between these measures and the HIV-1 latent reservoir, immune activation, lymphocyte subsets , and markers associated with bacterial translocation.We performed an open-label pilot phase II clinical trial involving chronically HIV-1-infected patients receiving stable antiretroviral therapy whose regimen was intensified with 48 weeks of maraviroc therapy+ or CD8+ counts, although a significant decrease was found in the proportion of HLA-DR+/CD38+ CD4+ and CD8+ T-cells. LPS and sCD14 levels increased.Overall a non significant reduction in the size of the latent reservoir was found (p = 0.068). A mean reduction of 1.82 IUPM was observed in 4 patients with detectable latent reservoir at baseline after 48 weeks of intensification. No effect on plasma residual viremia was observed. Unexpectedly, all the patients had detectable 2LTR DNA circles at week 24, while none of them showed those circles at the end of the study. No changes were detected in CD4Intensification with MVC was associated with a trend to a decrease in the size of the latent HIV-1 reservoir in memory T cells. No impact on residual viremia was detected. Additional studies with larger samples are needed to confirm the results.NCT00795444ClinicalTrials.gov  Antiretroviral therapy (ART) can reduce plasma HIV-1 RNA levels to <50 copies/ml + T cells in chronically HIV-1-infected patients with controversial results. Ramratnam et al. found an accelerated decay of the HIV-1 latent reservoir after intensification therapy with abacavir with or without efavirenz + T cell reservoir in patients receiving a four-drug combination as initial antiretroviral therapy provided discordant results, namely, a reduction in the reservoir in patients treated during acute infection Although intensification of current ART with potent drugs could potentially decrease residual viremia and prevent replenishment of viral reservoirs, prior intensification studies have not demonstrated any impact on residual viremia + T cells are rapidly depleted and T-cell activation is high. Since monitoring intestinal cells is difficult, expression of surrogate markers such as the mucosal homing receptor α4β7 integrin on circulating T cells has been proposed to correlate with loss or restoration of intestinal CD4+ T cells and could prove helpful in monitoring the success of therapeutic strategies + T cells Bacterial translocation and a low level of ongoing viral replication have been associated with an increase in immune activation in HIV-1-infected patients Maraviroc (MVC) is a potent new antiretroviral agent approved for the treatment of HIV-1 infection that blocks interaction between the virus and the CCR5 co-receptor, a crucial step in the HIV-1 life cycle + T cells in chronically HIV-1-infected patients receiving antiretroviral therapy. We also analyzed residual viremia and episomal 2LTR DNA to examine the relationship between these measures and the HIV-1 latent reservoir, immune activation, lymphocyte subsets , and markers associated with bacterial translocation. The hypothesis was that if these factors are causally linked, altering one of them with the intervention should result in alteration of the others.We performed a prospective open-label pilot phase II clinical trial to assess the effect of MVC intensification on latently infected CD4The protocol for this trial and supporting CONSORT checklist are available as supporting information; see We performed an open-label pilot phase II clinical trial to evaluate the effect of MVC on the cellular HIV-1 reservoir in patients receiving ART. The study was conducted at Hospital Universitario Ramón y Cajal in Madrid, Spain between 2008 and 2010. This independent clinical trial (NCT00795444) had a follow up of 48 weeks of intensification with MVC .ceic.hrc@salud.madrid.org). All patients provided their written informed consent for participation, which included sample collection and laboratory determinations.The study was carried out according to the recommendations of the Declaration of Helsinki and current Spanish legislation on clinical trials. It was approved by the AEMPS  and our Independent Ethics Committee , and no previous treatment with MVC. Patients were recruited from two hospitals , both in Madrid, Spain.Nine patients met all the following inclusion criteria: undetectable plasma viral load (pVL) by standard commercial assays (<50 copies HIV-1 RNA/ml) for at least two years; ART with three or more drugs for at least two years; CD4Patient's enrollment, allocation and follow up are described in Samples were collected at baseline and after 12, 24, 36, and 48 weeks of intensification with MVC. A total of 300 ml of heparinized whole blood was drawn to quantify the latent HIV-1 reservoir; 50 ml of whole blood with EDTA was drawn to obtain plasma and isolate peripheral blood mononuclear cells (PBMC). Samples were cryopreserved in the HIV Biobank of the Spanish AIDS Research Network (RIS) following current procedures In addition, two control groups  were also analyzed for some measurements.Two baseline determinations (three months apart) were performed to investigate the accuracy of the method and the stability of the reservoir.+ T cells (CD4+/HLA-DR-/CD25-) were isolated and purified using magnetic beads according to the manufacturer's recommendations . CD4+ T-cell purity greater than 99% with less than 0.4% activation was confirmed by flow cytometry. The isolated resting CD4+ T cells were plated in replicate limiting dilutions and co-cultured with allogenic irradiated PBMC from seronegative donors and phytohemagglutinin. The activated cell culture was fed once a week with PBMC from healthy donors after depletion of CD8+ T cells. On days 15 and 21, culture supernatants were tested for HIV-1 replication using the HIV-1 p24 antigen assay . Infected cell frequencies were determined by the maximum likelihood method and expressed as infectious units per million (IUPM) resting CD4+ T cells + T cells obtained from the total PBMCs conditioned the number of replicates of each dilution. The assay was performed to each point as duplicate fivefold dilution series.We carried out a previously described co-culture assay + T cells. Thereafter, the limit of detection decreased to 0.023 IUPM, since the total blood extraction volume was increased after approval of the amended protocol by the IEC.From baseline to week 12 of follow-up, the limit of detection of the assay was 0.12 IUPM resting CD4Residual viremia was measured with internally controlled an ultrasensitive quantitative real-time RT-PCR (single copy assay), as reported elsewhere The presence of HIV-1 episomal 2LTR DNA circles was detected by qualitative PCR. To maximize the recovery of 2LTR DNA circles and overcome the lack of sensitivity of this technique 5′-TAAGATGGGTGGCAAGTGGTCA; and reverse, 5′-TCTACTTGTCCATGCATGGCTT.A nested PCR flanking the episomal 2LTR DNA circle junction was designed. In the first round, 5–20 µL of episomal DNA were amplified in a 50-µL reaction with the following primers: forward, 5′-AATCTCTAGCAGTACTGGAAG; reverse, 5′-GCGCTTCAGCAAGCCGAGTCCT). PCR products were analyzed on a 1% agarose gel stained with GelRed .The second PCR round was performed using 1 to 2 µL of the first reaction product as the template and primers spanning the unique junction formed by ligation of 5′ and 3′ LTR sequences -Cy7, CD4-peridinin chlorophyll protein complex (PerCP), CD8-phycoerythrin (PE)-Cy7, CD38-phycoerythrin (PE), and HLA-DR-allophycocyanin (APC). Lymphocyte subpopulations were defined as naïve (CD45RA+CCR7+), T central memory , T effector memory , and T effector memory RA+ . To analyze lymphocyte subsets this antibody combination was used: CD3-allophycocyanin (APC)-Cy7, CD4-peridinin chlorophyll protein complex (PerCP), CD8-phycoerythrin (PE)-Cy7, CD45RA- phycoerythrin (PE), and CCR7-allophycocyanin (APC), β7-allophycocyanin (APC). Antibodies were from Becton Dickinson , and an unstained control was performed for all samples. Briefly, 100 µL of blood were lysed with 200 µl of FACS Lysing solution (Becton Dickinson) for 30 min at room temperature, incubated with the antibodies during 20 min at 4°C, washed and resuspended in PBS containing 1% azida. Cells were analyzed in a Gallios flow cytometer . At least 15000 CD3+ cells were collected for each sample and analyzed with Kaluza software (Beckman-Coulter) initially gating lymphocytes according to morphological parameters. The gating was always the same between different time points.Fresh EDTA anticoagulated whole blood was used to analyze CD4Two commercial assays were used to evaluate bacterial translocation from plasma samples. Plasma bacterial lipopolysaccharide (LPS) was measured using QCL-1000 Limulus Amebocyte Lysate  according to the manufacturer's protocol. This test quantifies endotoxins from the cell wall of gram-negative bacteria. Plasma sCD14 was quantified using the Quantikine® Human sCD14 Immunoassay , according to the manufacturer's instructions. Samples were run in duplicate in all cases.This is an exploratory pilot study that is trying to prove a concept. A treatment group of 10 patients has been considered adequate, based on data from previous studies published by other authors on the same subject.t test for independent samples was used to compare normally distributed continuous variables and the Mann-Whitney test to compare non-normally distributed continuous variables. Categorical variables were described as proportions. The association between categorical variables was evaluated using the chi-square test. A Spearman correlation was used. Statistical analysis was performed using SPSS software 16.0 .Continuous variables were expressed as the median and interquartile range and discrete variables as percentages. The + and CD8+ T-cell counts were 711 and 784 cells/mm3, respectively. All patients were receiving nucleoside reverse transcriptase inhibitor (NRTI)–containing regimens combined with non-nucleoside reverse transcriptase inhibitors (NNRTIs) in two cases (22%), with protease inhibitors (PIs) in five cases (56%), and with a third nucleoside in two cases (22%). The median duration of ART before study entry was 75 months.The baseline characteristics of the patients are summarized in MVC was well tolerated. Two patients interrupted the study early. Data from these patients were included only until the study interruption. None of them was included in the final analysis at week 48.One patient, who was coinfected with hepatitis C virus, had a significant increase in liver enzymes after the visit at week 12; the other discontinued all antiretroviral drugs due to poor adherence after the visit at week 12. The remaining patients maintained an HIV viral load <50 copies/ml until the end of the study.+ T cells decreased after 48 weeks of MVC intensification, although the difference was not significant (p = 0.068) (The number of latently HIV-1-infected memory CD4= 0.068) . Three p= 0.068) , and thi= 0.068)  or afterThe impact on the cellular reservoir was not associated with significant changes in residual viremia. At baseline, 8 of the 9 patients had undetectable residual viremia using the single copy assay. A significant increase in residual viremia was observed after 12 weeks of MVC intensification compared to baseline (p = 0.027). Afterwards, residual viremia decreased progressively until the end of the study, with no significant differences compared to baseline  and at week 24 in all 9 patients . At week 48, episomal 2LTR DNA circles were again undetectable in all patients .+ T cells decreased significantly after 12, 24, and 36 weeks of MVC intensification , only to increase after 48 weeks, although the difference with baseline was not significant (p = 0.6) (+ T-cell activation at baseline was significantly higher (p<0.001), with no significant differences at weeks 12 and 24 .The level of activation of CD4p = 0.6) . Compare+ T cells also decreased significantly after 12 weeks and until the end of the study  (The level of activation of CD8ctively) . The lev+ or CD8+ T cells were found during follow-up. A significant increase in CD8+ effector memory and TemRA T cell count was found during the first 24 and 36 weeks after MVC intensification, respectively. Thereafter, the differences compared to baseline were not significant . The level of CD8+ naive T cells was similar during follow-up . No significant differences were observed in any CD4+ T cell subsets during follow-up (data not shown).No differences in the absolute number or percentage of CD4nificant . On the + naïve and memory T cells expressing a β7 receptor significantly increased after 12 weeks of intensification  (+ T cells at week 12 compared to baseline (p = 0.046), and a trend was observed during the rest of the follow up  . These l+ T cells expressing a β7 receptor and the level of naive CD4+ T cells expressing β7 receptor at week 24 . Similarly, the level of LPS at week 36 correlated directly with that of CD8+ effector memory T cells .A significant direct correlation was found between sCD14 levels and LPS levels at baseline  while a similar trend was observed during the rest of the follow up. Furthermore, the LPS level correlated directly with both the proportion of CD4+ T cells. No impact on residual viremia was observed.In this pilot study, intensification of treatment with MVC in patients chronically infected with HIV-1 receiving successful long-term ART was associated with a trend to an accelerated decay in the pool of latently infected CD4+ T-cell counts, or, more recently, to reduce immune activation. With the exception of one study that showed a decrease in HIV RNA levels after abacavir intensification, none of the studies has been able to show significant improvements with intensified therapy compared to the standard three-drug regimens Intensification of successful antiretroviral therapy has been explored in two different ways. Some studies have evaluated its role for clinical purposes, that is, to achieve better virological control or a greater increase in CD4+ and CD4+ T cells strongly argues against this possibility.We observed a reduction in the size of the latent reservoir after intensification of treatment with MVC, although this reduction was not significant, probably due to the small sample size. This finding is consistent with other studies that included patients with different baseline characteristics + T cells and, therefore, could not induce viral replication + T cells and, through intracellular signalling, increase transcriptional activation of the latent virus. As an alternative hypothesis, CCR5 blockade by MVC could lead natural ligands or chemokines to bind to other receptors and induce latent HIV transcriptional activation through distinct signalling pathways. Both hypotheses would be consistent with the reduced number of latently infected memory T cells found in this study, the transient increase in the number of copies of HIV RNA seen in some patients, and the increased bacterial translocation plus migration of α4β7 cells to the gut. This possibility is further supported by the increased immune activation in intestinal tissue biopsies that was observed in another MVC intensification study Other findings in our study may help explain the effect on the latent reservoir. Unexpectedly, 2LTR DNA circles were transiently detectable during MVC intensification. Due to its mechanism of action, MVC is not expected to increase 2LTR DNA circles, even when inhibiting persistent viral replication. The increase in levels of this marker suggests that treatment with MVC in patients with undetectable viral load may induce viral replication. Preclinical studies showed that MVC had no agonistic activity on activated CD4Neither of the above hypotheses can account for the diminished immune activation that we detected. MVC has been shown to decrease immune activation in clinical trials, and it could be assumed that a different mechanism drives the impact on immune activation + T cell counts parallels the increase in bacterial translocation. These increments are balanced by a transient decrease in central memory CD8+ T cell counts, while naive CD8+ T cell levels remain stable + T cells and especially on activated CD8 T cells support the idea of migration of these cells to the intestine, where bacterial translocation takes place. This migration of activated cells to the gut could account for the reduced immune activation observed in the periphery. In this sense, immune events detected in peripheral blood may not reflect what takes place in the intestine, as is the case for the huge CD4+ T-cell destruction that occurs in the intestine of the SIV rhesus macaque during acute infection, which is not reflected in lymph nodes or peripheral blood [50].The transient increase in TEM and TemRA CD8Our work has a series of limitations. First, it is a pilot study with a small sample size, thus limiting the strength of the conclusions. Second, no control group was included. The study was designed as a proof of concept with no guarantee of any spontaneous effect in the sample. Finally, the methodological tools used for the different measurements and their interpretations are still imperfect.+ T cells. These effects are not reflected in reduced residual plasma viremia and may be associated with the activity of the drug in an anatomical reservoir, most likely the intestine. The mechanism by which MVC decrease the HIV-1 reservoir is under investigation. The decrease we observed, however, is by no means large enough to eradicate HIV. New therapeutic strategies, probably based on anti-latency drugs, are necessary to eradicate viral reservoirs. In this sense, complete inhibition of viral replication at all sites can be presumed to be necessary before these new treatment strategies can be applied. The exact role of treatment intensification in eradication remains undefined, and a large, comparative trial with adequate methodology is needed to provide a definitive answer to this relevant question.In summary, our study shows positive effects of MVC intensification on latently HIV-1-infected CD4Protocol S1Trial Protocol.(DOC)Click here for additional data file.Checklist S1CONSORT Checklist.(DOC)Click here for additional data file.


Guidance is needed on best medical management for advanced HIV disease with multidrug resistance (MDR) and limited retreatment options. We assessed two novel antiretroviral (ARV) treatment approaches in this setting.10 copies/ml, and 59% had a prior AIDS diagnosis. Median follow-up was 4.0 years in 1249 person-years of observation. There were no statistically significant differences in the primary composite outcome of ADE or death between re-treatment options of standard versus intensive ART , or between immediate retreatment initiation versus interruption before re-treatment , or in the rate of non-HIV associated serious adverse events between re-treatment options.We conducted a 2×2 factorial randomized open label controlled trial in patients with a CD4 count ≤300 cells/µl who had ARV treatment (ART) failure requiring retreatment, to two options (a) re-treatment with either standard (≤4 ARVs) or intensive (≥5 ARVs) ART and b) either treatment starting immediately or after a 12-week monitored ART interruption. Primary outcome was time to developing a first AIDS-defining event (ADE) or death from any cause. Analysis was by intention to treat. From 2001 to 2006, 368 patients were randomized. At baseline, mean age was 48 years, 2% were women, median CD4 count was 106/µl, mean viral load was 4.74 logWe did not observe clinical benefit or harm assessed by the primary outcome in this largest and longest trial exploring both ART interruption and intensification in advanced MDR HIV infection with poor retreatment options.NCT00050089Clinicaltrials.gov  Despite the effectiveness of current antiretroviral (ARV) treatment (ART) Optimal medical management remains unclear for MDR HIV with limited re-treatment options. Clinical management strategies include either continuing current or alternative ARVs in an ART regimen of up to four ARVs, intensifying ART with at least five and up to nine ARVs Several studies have addressed treatment interruption in patients with MDR HIV and are reviewed elsewhere In addition, health outcomes other than AIDS related disease have emerged as very important indicators of burden and impact of health in advanced HIV disease Our primary hypothesis was that ART intensification  would result in increased clinical benefit in terms of prolonging life and delaying the occurrence of new or recurrent AIDS events compared to continued standard treatment. A 2×2 factorial study design allowed us to test a second hypothesis regarding interruption, along with standard treatment or intensification. At the time of study development and protocol approval, conflicting data existed as to whether there was any benefit of interruption, for any duration; and only uncontrolled data existed as to the possible benefit of intensification. Our initial hypothesis regarding interruption was that it would result in a clinical benefit, and that when combined with intensification, any clinical benefit would be further increased. Our objective therefore was to investigate these clinical management strategies for MDR HIV infection in advanced disease with limited ARV retreatment options.The protocols for this trial and supporting CONSORT checklist are available as supporting information; see http://clinicaltrials.gov, number NCT00050089. Primary outcome was time to first new or recurrent AIDS-defining illness or death from any cause. Secondary outcome was time to first non-HIV related SAE. Outcomes were reviewed by an Endpoints Review Committee (ERC) blinded to randomization, classified either as ADEs according to the U.S. CDC revised AIDS case definition http://www.hivnet.ubc.ca/e/home/optima/protocol/), or as adverse events. Deaths (by ERC) were reviewed for attribution , and all SAEs (by STB and DWC) were reviewed for attribution .OPTIMA was a randomized 2×2 factorial clinical trial conducted in three countries  at over 70 clinical centers. The trial was registered at The protocol was approved by independent Research Ethics Boards at each site. The trial was performed in accordance with the principles of Good Clinical Practice and the Declaration of Helsinki. All volunteers signed written informed consent before any trial related procedure.HIV-1 infected patients were recruited between June 2001 and June 2006, and followed until study closure on December 31, 2007. They were eligible if the clinician was considering a change of ART because they had twice failed NRTI based ART including an NNRTI or a PI, or had genotypic or phenotypic evidence of three-class  ARV resistance, and they were currently receiving ART for at least three months. Other inclusion criteria included a CD4 count ≤300/µl, HIV-1 plasma viremia ≥5000 copies/ml , or ≥2500 . Exclusions were age < eighteen years, pregnancy, nursing mothers, AIDS-defining illness within fourteen days of screening, or likelihood of imminent death.Randomization was stratified by CD4 count at screening (above or below 100/µl) and by study site, according to a computer-generated randomization list, in permuted blocks of randomly varying size by the coordinating center to re-treatment with (a) either initial treatment interruption for an intended period of 12 weeks followed by a new ART regimen, or a change of ART regimen without interruption, and (b) change in ART by intensification (to ≥5 ARV) or standard ART (of ≤4 ARV). For patients randomized to ART interruption before re-treatment, the assigned standard or intensified ART option was not communicated to the physician or the patient until the end of the planned interruption. The ARVs in the retreatment regimens were chosen by the treating physician according to treatment history and expected tolerance, and by recent or screening susceptibility testing. Low-dose ritonavir (≤400 mg/day) was not counted as an ARV. Patients were recruited, consented and followed-up in the clinics where they received care for HIV infection and followed-up at 6, 12, 24 and then every 12 weeks after randomization with additional visits at 2 and 6 weeks after treatment interruption and after re-treatment initiation. Blood samples were collected at screening for genotypic resistance testing and virtual phenotype  and at screening and follow-up visits for HIV-1 viral load (VL), CD4 and CD8 T lymphocyte count and routine hematological and biochemical blood tests. Follow-up ARV resistance testing was permitted at the discretion of the treating physician. Incremental phenotypic susceptibility score (PSS) from the baseline vP was calculated as previously described A patient was considered compliant to protocol if after assignment to a treatment interruption it was continued for at least 6 weeks, if they started retreatment within two weeks of their assigned time, and they initiated retreatment with the number of ARVs within the assigned standard or intensive ART option. Periods up to 30 days off ART or off the assigned number of drugs in the standard or intensive ART regimens were allowed. Patients were considered protocol compliant if the reason for change was stated to be high VL, low CD4, severe adverse event or intercurrent illness. Medication compliance was determined at each follow-up visit using an assessment tool adopted from the AIDS Clinical Trials Group adherence questionnaire alpha)  = 0.05; 3) Loss to follow-up at 3.5 years of 10%; 4) cross-over from standard to intensive ART of 5% in year 1, increasing 10% every year thereafter; 5) cross-over from intensive to standard ART of 20% in year 1, decreasing by 50% every year thereafter. With 652 outcomes in 1700 subjects, a 22% relative reduction in hazard would be detected with 93% power. During the study sample size was revised based on observed accrual, outcome and crossover rates, with extended accrual and follow-up periods but no change in other assumptions, such that 261 outcomes in 390 patients would provide a study power of 75%. Also, a change was implemented in the UK due to lack of equipoise among treating physicians regarding ART interruption, to permit a choice of only one of the two randomizations.The original study design Analyses were performed by intention to treat, according to randomly allocated management strategy. Analyses of the primary outcome and survival comparisons were performed using time-to-event methods including Kaplan-Meier plots and the stratified log-rank tests. Analyses also included interaction of treatment assignments, length of follow-up, or calendar time of enrollment on the primary outcome. Treatment differences were estimated by hazard ratios using Cox proportional hazards regression. Other comparisons including outcomes stratified by baseline CD4 count  were made with standard parametric and non-parametric statistical tests. An independent Data and Safety Monitoring Board monitored results on an ongoing basis.The study funders  reviewed and approved the design and conduct of the trial, with external expert reviewers who were represented on the Trial Steering Committee. The OPTIMA Writing Committee and the OPTIMA team had full access to the statistical report and to the study data by request and the Writing Committee had final responsibility for the submitted manuscript.10 copies/ml.Of the 457 patients screened, 368 patients were enrolled , of whom 339 were randomized to the 2×2 factorial and 29 were randomized to standard vs. intensification-ART only . The bas3) with 83%, 53%, 59% taking anti-PCP, other anti-fungal, and antibacterial medications at the last follow-up visit respectively.The study population was heavily ARV-experienced: 96% had taken over three NRTI , 97% at least one NNRTI , 63% at least three PI  and 2.5% enfuvirtide. In the retreatment regimens, 171 (99.4%) of 172 standard-ART and 139 (91.5%) of 152 intensification-ART patients were protocol-compliant in the number of ARVs used with a median number of ARVs of 3 in standard and 5 in intensive ART regimens. The mean number of active ARVs as determined by PSS was 1.3 in the standard and 1.7 in the intensive groups (p<0.03), and 1.4 in the interruption and 1.5 in the no interruption groups (P = NS). The median time to protocol non-compliance resulting in a change in retreatment strategy was 187  weeks for standard, versus 59  weeks for intensification-ART . There was no significant difference in time to changing treatment strategy between the ART interruption before retreatment versus immediate ART strategy. For those assigned ART interruption, the median duration of the interruption was 12 weeks (IQR 12–14 weeks). Use of primary and secondary opportunistic infection prophylaxis was high at baseline, with over 80% of patients overall taking anti-PCP, 25% taking other anti-fungal, and 45% taking other antibacterial medications. This level of prophylaxis was maintained during the study (in patients with CD4<200 cells/mmPneumocystis jirovecii pneumonia (PCP) (8.5%), cytomegalovirus disease (4.8%), HIV wasting syndrome (4.3%), Mycobacterium avium complex infection (3.7%), Kaposi's sarcoma (2.4%), and cryptococcosis (2.4%). There was no significant difference in the number or type ADE between re-treatment options. The ERC adjudicated 52% of deaths to be HIV-related, 2% due to ART medication, 15% unrelated to HIV or ART, and 31% as unattributable.A total of 165 (44.8%) of 368 subjects experienced a primary outcome. This included 67 deaths without a preceding ADE, and 98 ADE of whom 61 died subsequently. Of the 165 primary outcomes, the most common were death (40.9%), esophageal candidiasis (18.3%), Of 368 randomized to standard versus intensive retreatment, there were respectively 83 versus 82 ADE or death outcomes , p = 0.03), and more had ADE or death (39 (81.3%) versus 26 (55.3%), p = 0.007). Finally, tests of main effect interactions demonstrated no statistical significance in AIDS events or death when treatment assignment, length of follow-up, or calendar time of enrollment were considered . Significantly more patients with baseline CD4 ≤100/µl had a new or recurrent ADE with continuation (38 (46.3%)) compared to with interruption (23 (30.7%), p = 0.04). In the lowest baseline CD4 quartile (< 36/µl) more deaths occurred with intensification than standard ART retreatment (34 (70.8%) nsidered , 4, 5, 6185 of 368 (50%) patients developed 481 non HIV-related SAEs. There was no significant difference in number or time to first non HIV-related SAE between standard vs. intensive ART retreatment , or between interruption versus continuation  .Expected changes in CD4 lymphocyte counts and plasma HIV viral load were observed ; viremiaThere are randomized control trials in retreatment of treatment-failing or drug-resistant HIV infection with unpromising retreatment options, which evaluate new ARVs Although no significant advantage or disadvantage to any of the retreatment options was seen in over 1000 person-years of observation, confidence limits here may include a clinically significant effect that is not reflected in the simple outcome rates. For example, the data are consistent with a possible 14% decrease or 59% increase in death or ADE with treatment intensification; and approximately a 32% decline or 30% increase in death or ADE with treatment interruption. Unlike other reports, there was no excess risk of ADE with ART interruption in patients with the highest expected risk defined by lowest baseline CD4 counts. For example, Lawrence et al. reported on a controlled trial of 16-week treatment interruption, which showed an excess of treatment-responsive esophageal candidiasis, but no difference in HRQoL or in survival No other trials of retreatment intensification evaluate clinical outcomes, and there is a valid concern for toxicity, tolerance and cost as well as efficacy of intensified ART. OPTIMA identified no significant overall benefit in health outcomes or harm in excess non-HIV SAEs or drug intolerance. However, maintenance of intensification was much less than a standard ART retreatment option so that no difference in cumulative additive or antagonistic effects from additional medications in MEGA-ART were seen.In an ART era of new and very promising retreatment options of new class ARVs and combinations, what instruction may be taken from these trials of retreatment management options for HIV treatment failure? In the developing world, the greatest burden of HIV and the greatest healthcare resource constraints co-exist. Conventional ARV roll-outs for the many untreated are still expanding and leave little room to allocate added resources for the inevitable adherence- and intolerance-driven failures of ART and treatment-emergent MDR-HIV. In clinical practice, treatment interruption is a common occurrence. A recently completed trial in Uganda exploring short, intermittent HIV treatment interruption in a healthier population found that 7 days on and 7 days off ARVs resulted in more virologic failures compared to 7 day or 5 day continuous treatment cycles which yielded comparable results OPTIMA suggests that in some settings of ART failure with limited prospects for re-treatment, individualization of retreatment for interruption or intensification with conventional ARVs may be considered, particularly with good clinical follow-up and management of opportunistic infection risk. Such decisions should carefully weigh both potential benefits and harms. As trials of this nature in these types of settings are unlikely in the future, a pooling of existing trial datasets may offer the best opportunity to identify principles and particular populations which may gain or lose most in ARV retreatment choices after failure of ART, MDR-HIV and limited retreatment options.Checklist S1CONSORT Checklist.(0.16 MB RTF)Click here for additional data file.Protocol S1Trial Protocol.(1.06 MB PDF)Click here for additional data file.