New HIV-1 circulating recombinant form 94: from phylogenetic detection of a large transmission cluster to prevention in the age of geosocial-networking apps in France, 2013 to 2017.

BackgroundEnding the HIV pandemic must involve new tools to rapidly identify and control local outbreaks and prevent the emergence of recombinant strains with epidemiological advantages.AimThis observational study aimed to investigate in France a cluster of HIV-1 cases related to a new circulating recombinant form (CRF). The confirmation this CRF's novelty as well as measures to control its spread are presented.MethodsPhylogenetic analyses of HIV sequences routinely generated for drug resistance genotyping before 2018 in French laboratories were employed to detect the transmission chain. The CRF involved was characterised by almost full-length viral sequencing for six cases. Cases' clinical data were reviewed. Where possible, epidemiological information was collected with a questionnaire.ResultsThe transmission cluster comprised 49 cases, mostly diagnosed in 2016-2017 (n = 37). All were infected with a new CRF, CRF94_cpx. The molecular proximity of this CRF to X4 strains and the high median viraemia, exceeding 5.0 log10 copies/mL, at diagnosis, even in chronic infection, raise concerns of enhanced virulence. Overall, 41 cases were diagnosed in the Ile-de-France region and 45 were men who have sex with men. Among 24 cases with available information, 20 reported finding partners through a geosocial networking app. Prevention activities in the area and population affected were undertaken.ConclusionWe advocate the systematic use of routinely generated HIV molecular data by a dedicated reactive network, to improve and accelerate targeted prevention interventions. Geosocial networking apps can play a role in the spread of outbreaks, but could also deliver local targeted preventive alerts.

Phenotypic analysis of HIV-1 E157Q integrase polymorphism and impact on virological outcome in patients initiating an integrase inhibitor-based regimen.

Objectives: To assess the phenotypic susceptibility of the E157Q polymorphism in HIV-1 integrase (IN) and the virological outcome of patients infected with E157Q-mutated virus initiating an IN inhibitor (INI)-based regimen. Methods: This was a multicentre study assessing IN sequences from INI-naive patients among 17 French HIV clinical centres. E157Q site-directed mutants in pNL4.3 and pCRF02_AG contexts were assessed in a recombinant phenotypic assay. Results: Prevalence of the E157Q polymorphism was 2.7% among 8528 IN sequences from INI-naive patients and its distribution was 1.7%, 5.6% and 2.2% in B, CRF02_AG and various non-B subtypes, respectively. Thirty-nine INI-naive patients with E157Q-mutated virus initiated an INI-based regimen. Among them, 15 had a viral load (VL) <50 copies/mL at initiation and virological suppression was maintained during the first year of follow-up in all but two exhibiting a viral blip. Twenty-four patients had a VL > 50 copies/mL at the time of INI-based regimen initiation. Among them eight were receiving a first-line regimen and the only two patients who did not reach VL < 50 copies/mL at week 24 were receiving elvitegravir. The 16 remaining patients were ART experienced in virological failure with drug-resistant viruses displaying several virological outcomes independently of the genotypic susceptibility score. Phenotypic analyses showed a fold change in EC50 of 0.6, 0.9 and 1.9 for raltegravir, dolutegravir and elvitegravir, respectively, in a subtype B context, and 1.1, 1.9 and 2.4 for raltegravir, dolutegravir and elvitegravir, respectively, in a CRF02_AG context. Conclusions: Assessment of virological response in 39 patients initiating an INI-based regimen with E157Q-mutated virus, in combination with phenotypic analysis, suggests that particular attention should be paid to antiretroviral-naive patients and dolutegravir should be preferentially used in these patients.

Efficacy and tolerance of dolutegravir-based combined ART in perinatally HIV-1-infected adolescents: a French multicentre retrospective study.

Objectives: To assess the safety and efficacy of a dolutegravir-based regimen in perinatally HIV-1-infected adolescents. Patients and methods: We conducted a retrospective multicentre study of 50 adolescents beginning dolutegravir-based treatment regimens between January 2014 and December 2015. Clinical and biological data collected before and after dolutegravir initiation were analysed. The primary endpoint was the proportion of patients achieving a plasma viral load (PVL) <50 copies/mL within 3 months of dolutegravir initiation (for patients with detectable viraemia at baseline) and maintaining virological suppression (PVL <50 copies/mL) until the last follow-up visit (for all patients). Results: Virological suppression was noted for 17/50 adolescents at baseline. Dolutegravir-based regimens maintained virological success in 14/17 patients (82%). The other three patients experienced a transient viral rebound, before PVL fell to < 50 copies/mL again, with no need to change the antiretroviral regimen. Thirty-three viraemic adolescents were enrolled. All but one had already received antiretroviral drugs. Virological success was achieved and maintained in 19/33 subjects (58%). Another three adolescents with initial virological failure had an undetectable PVL at the end of follow-up, with reinforced measures to improve compliance. Overall, sustained virological success was observed in 66% of patients and 78% of patients had an undetectable PVL at the last visit. Dolutegravir was well tolerated. Only one patient stopped treatment for severe drug-related adverse effects (dizziness and sleep disturbance). No emergence of resistance mutations was observed in patients with virological failure. Conclusions: Dolutegravir was safe and virologically effective in these patients, for whom multiple interventions were required to improve compliance.

Antiretroviral-naive and -treated HIV-1 patients can harbour more resistant viruses in CSF than in plasma.

OBJECTIVES: The neurological disorders in HIV-1-infected patients remain prevalent. The HIV-1 resistance in plasma and CSF was compared in patients with neurological disorders in a multicentre study. METHODS: Blood and CSF samples were collected at time of neurological disorders for 244 patients. The viral loads were >50 copies/mL in both compartments and bulk genotypic tests were realized. RESULTS: On 244 patients, 89 and 155 were antiretroviral (ARV) naive and ARV treated, respectively. In ARV-naive patients, detection of mutations in CSF and not in plasma were reported for the reverse transcriptase (RT) gene in 2/89 patients (2.2%) and for the protease gene in 1/89 patients (1.1%). In ARV-treated patients, 19/152 (12.5%) patients had HIV-1 mutations only in the CSF for the RT gene and 30/151 (19.8%) for the protease gene. Two mutations appeared statistically more prevalent in the CSF than in plasma: M41L (P=0.0455) and T215Y (P=0.0455). CONCLUSIONS: In most cases, resistance mutations were present and similar in both studied compartments. However, in 3.4% of ARV-naive and 8.8% of ARV-treated patients, the virus was more resistant in CSF than in plasma. These results support the need for genotypic resistance testing when lumbar puncture is performed.

Initiation of rilpivirine, tenofovir and emtricitabine (RPV/TDF/FTC) regimen in 363 patients with virological vigilance assessment in 'real life'.

OBJECTIVES: To study the single-tablet regimen (STR) combination rilpivirine/tenofovir/emtricitabine (RPV/TDF/FTC) as soon as it became available. We describe a 14 month follow-up in a real clinical setting with a focus on resistance to RPV/TDF/FTC and polymorphisms associated with these drugs. METHODS: We estimated drug resistance at STR baseline by combining all available resistance tests, resulting in a cumulative virtual genotype. Physicians were advised of current or archived resistance mutations for the three drugs. Virological response was analysed according to resistance genotype at baseline. RESULTS: Three hundred and sixty-three patients received RPV/TDF/FTC; 79% had received previous treatment and RPV/TDF/FTC was the result of a switch of one drug to rilpivirine in two-thirds of cases. The cumulative genotype showed 4% of rilpivirine resistance mutations at baseline and 16% of polymorphisms concerning non-nucleoside reverse transcriptase inhibitors (NNRTIs). With a median duration of STR of 8 months, 78% of patients with these polymorphisms were virologically suppressed compared with 96% with wild-type genotypes. Five genotypes were determined during the follow-up, revealing three rilpivirine resistance-associated mutations: E138Q/Y181I, M230L and K101P (potentially with a K101Q intermediate). CONCLUSIONS: This observational study reflects routine clinical practice and the relevance of virological advice. It also confirms the efficacy of this STR (RPV/TDF/FTC) for naive and virologically suppressed pretreated patients with a low prevalence of virological failure and resistance if the cumulative baseline genotype is free of resistance to NNRTIs and/or polymorphisms associated with NNRTIs.

Consistent high concentration of the viral microRNA BART17 in plasma samples from nasopharyngeal carcinoma patients--evidence of non-exosomal transport.

BACKGROUND: Because latent Epstein Barr (EBV)-infection is a specific characteristic of malignant nasopharyngeal carcinoma (NPC), various molecules of viral origin are obvious candidate biomarkers in this disease. In a previous study, we could show in a few clinical samples that it was possible to detect a category of EBV microRNAs called miR-BARTs in the plasma of at least a fraction of NPC patients. The first aim of the present study was to investigate the status of circulating miR-BART17-5p (one of the miR-BARTs hereafter called miR-BART17) and EBV DNA in a larger series of NPC plasma samples. The second aim was to determine whether or not circulating miR-BART17 was carried by plasma exosomes. PATIENTS AND METHODS: Plasma samples were collected from 26 NPC patients and 10 control donors, including 9 patients with non-NPC Head and Neck squamous cell carcinoma and one healthy EBV carrier. Concentrations of miR-BART17 and two cellular microRNAs (hsa-miR-16 and -146a) were assessed by real-time quantitative PCR with spike-in normalization and absolute quantification. In addition, for 2 patients, exosome distributions of miR-BART17 and miR-16 were investigated following plasma lipoprotein fractionation by isopycnic density gradient ultrcentrifugation. RESULTS: The miR-BART17 was significantly more abundant in plasma samples from NPC patients compared to non-NPC donors. Above a threshold of 506 copies/mL, detection of miR-BART17 was highly specific for NPC patients (ROC curve analysis: AUC=0.87 with true positive rate = 0.77, false positive rate = 0.10). In this relatively small series, the concentration of plasma miR-BART17 and the plasma EBV DNA load were not correlated. When plasma samples were fractionated, miR-BART17 co-purified with a protein-rich fraction but not with exosomes. CONCLUSIONS: Detection of high concentrations of plasma miR-BART17 is consistent in NPC patients. This parameter is, at least in part, independent of the viral DNA load. Circulating miR-BART17 does not co-purify with exosomes.

Changes in visual acuity in patients with wet age-related macular degeneration treated with intravitreal ranibizumab in daily clinical practice: the LUMIERE study.

PURPOSE: To survey compliance with recommended intravitreal ranibizumab treatment protocols in daily clinical practice in France, with reference to outcomes. METHODS: A retrospective, descriptive, observational study in patients with subfoveal wet age-related macular degeneration treated with ranibizumab. All historical data for the study period, including demographic, treatment, and disease details and visual acuity measurements (baseline, Month 3, and Month 12), were recorded retrospectively at least 12 months after the beginning of treatment. RESULTS: In 551 patients followed by 16 ophthalmologists, 12 months of intravitreal ranibizumab treatment induced a mean visual acuity gain of 3.2 ± 14.8 Early Treatment Diabetic Retinopathy Study-equivalent letters. Fewer than 40% of patients received the recommended treatment of initial 3 monthly injections. More than 50% had to wait >8 days between diagnosis and treatment. At Month 3, visual acuity gain was greater in patients who had received recommended induction and in whom treatment was initiated quickly. At Month 12, the induction-related effect had largely disappeared but the time-to-treatment effect persisted. Patients had an average of 5.1 injections (2.6 during induction period). No patients were monitored monthly as stipulated in the guidelines. CONCLUSION: Although poor compliance with recommendations has been reflected in mediocre outcomes, there is evidence that practice is improving.

Evaluation of the genotypic prediction of HIV-1 coreceptor use versus a phenotypic assay and correlation with the virological response to maraviroc: the ANRS GenoTropism study.

Genotypic algorithms for prediction of HIV-1 coreceptor usage need to be evaluated in a clinical setting. We aimed at studying (i) the correlation of genotypic prediction of coreceptor use in comparison with a phenotypic assay and (ii) the relationship between genotypic prediction of coreceptor use at baseline and the virological response (VR) to a therapy including maraviroc (MVC). Antiretroviral-experienced patients were included in the MVC Expanded Access Program if they had an R5 screening result with Trofile (Monogram Biosciences). V3 loop sequences were determined at screening, and coreceptor use was predicted using 13 genotypic algorithms or combinations of algorithms. Genotypic predictions were compared to Trofile; dual or mixed (D/M) variants were considered as X4 variants. Both genotypic and phenotypic results were obtained for 189 patients at screening, with 54 isolates scored as X4 or D/M and 135 scored as R5 with Trofile. The highest sensitivity (59.3%) for detection of X4 was obtained with the Geno2pheno algorithm, with a false-positive rate set up at 10% (Geno2pheno10). In the 112 patients receiving MVC, a plasma viral RNA load of <50 copies/ml was obtained in 68% of cases at month 6. In multivariate analysis, the prediction of the X4 genotype at baseline with the Geno2pheno10 algorithm including baseline viral load and CD4 nadir was independently associated with a worse VR at months 1 and 3. The baseline weighted genotypic sensitivity score was associated with VR at month 6. There were strong arguments in favor of using genotypic coreceptor use assays for determining which patients would respond to CCR5 antagonist.

Extra-cellular release and blood diffusion of BART viral micro-RNAs produced by EBV-infected nasopharyngeal carcinoma cells.

BACKGROUND: Nasopharyngeal carcinoma (NPC) is a human epithelial malignancy consistently associated with the Epstein-Barr virus. The viral genome is contained in the nuclei of all malignant cells with abundant transcription of a family of viral microRNAs called BART miRNAs. MicroRNAs are well known intra-cellular regulatory elements of gene expression. In addition, they are often exported in the extra-cellular space and sometimes transferred in recipient cells distinct from the producer cells. Extra-cellular transport of the microRNAs is facilitated by various processes including association with protective proteins and packaging in secreted nanovesicles called exosomes. Presence of microRNAS produced by malignant cells has been reported in the blood and saliva of tumor-bearing patients, especially patients diagnosed with glioblastoma or ovarian carcinoma. In this context, it was decided to investigate extra-cellular release of BART miRNAs by NPC cells and their possible detection in the blood of NPC patients. To address this question, we investigated by quantitative RT-PCR the status of 5 microRNAs from the BART family in exosomes released by NPC cells in vitro as well as in plasma samples from NPC xenografted nude mice and NPC patients. RESULTS: We report that the BART miRNAs are released in the extra-cellular space by NPC cells being associated, at least to a large extent, with secreted exosomes. They are detected with a good selectivity in plasma samples from NPC xenografted nude mice as well as NPC patients. CONCLUSIONS: Viral BART miRNAs are secreted by NPC cells in vitro and in vivo. They have enough stability to diffuse from the tumor site to the peripheral blood. This study provides a basis to explore their potential as a source of novel tumor biomarkers and their possible role in communications between malignant and non-malignant cells.

Perinatal acquisition of drug-resistant HIV-1 infection: mechanisms and long-term outcome.

BACKGROUND: Primary-HIV-1-infection in newborns that occurs under antiretroviral prophylaxis that is a high risk of drug-resistance acquisition. We examine the frequency and the mechanisms of resistance acquisition at the time of infection in newborns. PATIENTS AND METHODS: We studied HIV-1-infected infants born between 01 January 1997 and 31 December 2004 and enrolled in the ANRS-EPF cohort. HIV-1-RNA and HIV-1-DNA samples obtained perinatally from the newborn and mother were subjected to population-based and clonal analyses of drug resistance. If positive, serial samples were obtained from the child for resistance testing. RESULTS: Ninety-two HIV-1-infected infants were born during the study period. Samples were obtained from 32 mother-child pairs and from another 28 newborns. Drug resistance was detected in 12 newborns (20%): drug resistance to nucleoside reverse transcriptase inhibitors was seen in 10 cases, non-nucleoside reverse transcriptase inhibitors in two cases, and protease inhibitors in one case. For 9 children, the detection of the same resistance mutations in mothers' samples (6 among 10 available) and in newborn lymphocytes (6/8) suggests that the newborn was initially infected by a drug-resistant strain. Resistance variants were either transmitted from mother-to-child or selected during subsequent temporal exposure under suboptimal perinatal prophylaxis. Follow-up studies of the infants showed that the resistance pattern remained stable over time, regardless of antiretroviral therapy, suggesting the early cellular archiving of resistant viruses. The absence of resistance in the mother of the other three children (3/10) and neonatal lymphocytes (2/8) suggests that the newborns were infected by a wild-type strain without long-term persistence of resistance when suboptimal prophylaxis was stopped. CONCLUSION: This study confirms the importance of early resistance genotyping of HIV-1-infected newborns. In most cases (75%), drug resistance was archived in the cellular reservoir and persisted during infancy, with or without antiretroviral treatment. This finding stresses the need for effective antiretroviral treatment of pregnant women.

High clustering of acute HCV infections and high rate of associated STIs among Parisian HIV-positive male patients.

BACKGROUND: Increasing incidence of hepatitis C virus (HCV) infection in human immunodeficiency virus (HIV)-positive men having sex with men (MSM) has been described in recent years. Phylogenetic analyses of acute HCV infections were undertaken to characterize the dynamics during the epidemic in Paris, and associated sexually transmitted infections (STIs) were evaluated. METHODS: Sanger sequencing of polymerase gene was performed. Maximum likelihood phylogenies were reconstructed using FastTree 2.1 under a GTR+CAT model. Transmission chains were defined as clades with a branch probability ≥0.80 and intraclade genetic distances <0.02 nucleotide substitutions per sites. STIs detected ≤1 month before HCV diagnosis were considered. RESULTS: Among the 85 studied patients, at least 81.2% were MSM. Respectively, 47.6%, 39.0%, 11.0% and 2.4% were infected with genotypes 1a, 4d, 3a and 2k. At least 91.8% were co-infected with HIV. HCV re-infection was evidenced for 24.7% of patients and STIs for 20.0% of patients. Twenty-two transmission chains were identified, including 52 acute hepatitis C (11 pairs and 11 clusters from three to seven patients). CONCLUSIONS: These results revealed strong clustering of acute HCV infections. Thus, rapid treatment of both chronic and acute infections is needed among this population to decrease the prevalence of HCV, in combination with preventive behavioural interventions.

HIV-1-infected patients from the French National Observatory experiencing virological failure while receiving enfuvirtide.

OBJECTIVES: We studied gp41 mutations associated with failing enfuvirtide salvage therapy. METHODS: This multicentre study involved patients with HIV-1 plasma viral load (pVL) > 5000 copies/mL after at least 3 months of uninterrupted enfuvirtide therapy and with plasma samples available at inclusion (T0), at initial enfuvirtide failure (T1) and at last follow-up visit during continued failing enfuvirtide therapy (T2). The HR-1 and HR-2 domains of the gp41 gene were sequenced at T0, T1 and T2. RESULTS: Ninety-nine patients were enrolled. At baseline, the median pVL and CD4 cell count were 5.1 log copies/mL and 72 cells/mm(3), respectively. Based on the ANRS Resistance Group algorithm, the proportion of patients harbouring viruses with enfuvirtide resistance mutations increased significantly between T0 and T1. In the HR-1 domain, the V38A/M, Q40H, N42T, N43D and L45M mutations wereselected (P < 0.02). In the HR-2 domain, no mutations were significantly selected during the follow-up. None of the mutations was associated with a CD4 cell count increment. CONCLUSIONS: Mutations selected during failing enfuvirtide salvage therapy are mainly located in the HR-1 domain of the gp41 gene, between codons 38 and 45. No mutations were associated with an increase in the CD4 cell count.

Detection of numerous HIV-1/MO recombinants in France.

BACKGROUND: The broad genetic divergence of HIV-1/O relative to HIV-1/M has important implications for diagnosis, monitoring and treatment. Despite this divergence, some HIV-1/M+O dual infections and HIV-1/MO recombinant forms have been reported, mostly in Cameroon, where both groups are prevalent. Here, we describe the characteristics of such infections detected in France in 10 new patients, and discuss their implications for biological and clinical practice, owing to the presence of group O species. METHODS: The French National Reference Centre for HIV received samples within the framework of mandatory notification of HIV infections, and for expert analysis. A strategy combining serotyping, viral quantification, group-specific molecular amplification and whole-genome sequencing was used for strain characterization and complementary investigations. RESULTS: We identified one patient with M+O infection, three patients with M+O infection associated with an MO recombinant, and six patients with only an MO recombinant. These atypical infections were detected upon strain characterization (n = 4) or because of anomalies during patient monitoring (n = 6). We identified eight new URF_MO, all but one originating from Cameroon. Interestingly, two distinct recombinant strains were found in two unrelated patients, representing possible precursors of a CRF_MO. CONCLUSION: Our work highlights the fact that the continuous evolution of HIV can hinder diagnosis and complicate clinical practice. We stress that unexpected results during diagnosis or monitoring necessitate further serological and molecular exploration, these atypical infections influence biological and therapeutic management and necessitate appropriate tools, and specific surveillance is necessary, especially as the frequency of such infections may be underestimated.

Response to HAART in French patients with resistant HIV-1 treated at primary infection: ANRS Resistance Network.

OBJECTIVE: The aim of the study was to analyse the response to highly active antiretroviral therapy (HAART) initiated at the time of primary HIV infection (PHI) in patients infected with a virus resistant to > or = 1 drug of their treatment compared with patients infected with a wild-type virus. METHODS: We analysed data from 350 patients who were enrolled from 1996-2004 in the French ANRS PRIMO Cohort or in the ANRS Resistance Group and treated with HAART during PHI. During the study period, HAART was initiated before the result of the genotypic resistance test was available. We compared patients infected with a virus resistant to > or = 1 drug of their regimen (GR group, n = 46) with patients harbouring a wild-type virus (WT group, n = 304). Virological and immunological response to treatment according to drug-resistance profile was analysed 3 months and 6 months after HAART initiation. RESULTS: In GR and WT groups, HIV RNA level was < 400 copies/ml in 68% and 83% (P = 0.02) and < 50 copies/ml in 23% and 40% (P = 0.08) 3 months after HAART initiation. In multivariable logistic regression taking into account gender, age, boosted PI regimen, plasma HIV RNA and CD4+ T-cell count at HAART initiation, patients with virus resistant to > or = 1 drug of their regimen were significantly less likely to achieve undetectable viral load at month 3 (odds ratio 0.32, 95% confidence interval 0.15-0.72) than the others. This difference was sustained up to month 6. CONCLUSION: In this large cohort of HAART-treated PHI-patients, the presence of drug resistance mutations led to suboptimal response to early therapy.

A new insertion in the HIV-1 reverse transcriptase gene inducing major resistance to non-nucleoside reverse transcriptase inhibitors.

We identified an HIV-1 isolate with a 3 base pairs insertion in the 100-105 region of the reverse transcriptase gene (RT) along with a G190E and a V75A mutation. Virus carrying the insertion alone or in association with G190A was not infectious. The association of G190E and the 100-105 insertion displayed a high level of resistance to non-nucleoside reverse transcriptase inhibitors; the addition of the insertion to G190E may increase the activity of RT.

Stable prevalence of genotypic drug resistance mutations but increase in non-B virus among patients with primary HIV-1 infection in France.

OBJECTIVE: To evaluate the frequency of drug-resistant HIV-1 viral strains from patients presenting with primary infection in 1999-2000 and to survey the molecular epidemiology of these viruses circulating in France. METHODS: Resistance mutations were detected by sequencing the reverse transcriptase and the protease genes in plasma samples from 249 individuals. Phylogenetic analysis was performed on the reverse transcriptase genes. RESULTS: Ten per cent of patients [26/249; 95% confidence interval (CI) 7-15%] presented with virus mutations associated with resistance to at least one antiretroviral drug. The distribution of the resistance mutations was as follows: to nucleoside reverse transcriptase inhibitors in 19 (8%; 95% CI 5-12%) and to non-nucleoside reverse transcriptase inhibitors in 10 (4%; 95% CI 2-7%). Primary resistance mutations to protease inhibitors were detected in 14 (6%; 95% CI 3-9%). Twelve patients (5%; 95% CI 3-8%) presented with virus harbouring mutations associated with resistance to two or three classes of antiretroviral drugs. The median HIV RNA in plasma at enrollment was lower in patients with one or more drug resistance mutations than in patients with no mutations (5.05 log versus 5.47 log, P = 0.05). Phylogenetic analysis revealed that 19% (14-24%) of patients harboured HIV-1 non-B subtype strains; this proportion remained high when Caucasian patients only were considered (14%). CONCLUSION: This study, performed within the French network on HIV-1 primary infection survey, revealed no change in the frequency of resistant viral strains over time, but showed an increasing prevalence of non-B subtypes overall and among Caucasian individuals.

Genotypic determinants of the virological response to tenofovir disoproxil fumarate in nucleoside reverse transcriptase inhibitor-experienced patients.

OBJECTIVE: To assess the genotypic determinants of the virological response to tenofovir disoproxil fumarate (TDF) in a multicentre cohort of antiretroviral (ARV)-experienced patients receiving TDF as a part of a salvage therapy. METHODS: HIV-1 genotype was assessed at baseline in a subgroup of 161 patients of the French expanded access program receiving a stable TDF-including regimen for 3 months or more. Reverse transcriptase mutations associated with the viral load decrease at month 3 with a P-value <0.15 were retained for the construction of a mutation score. The score was then validated using a multivariate analysis and bootstrap resampling method. RESULTS: The strongest association with decrease in viral load was observed with a set of seven mutations (TDF mutation score) that consisted of M41L, E44D, D67N, T69D/N/S, L74V, L210W and T215Y/F RT mutations. The RT K65R mutation and the insertions at codon 69 were not included in the analysis due to low prevalences. A TDF mutation score of < or = 2 predicted the absence of resistance to TDF and > or = 6 mutations predicted resistance to TDF with corresponding reductions in viral load of -1.3 +/- 1.1, and +0.1 +/- 0.7 log10 copies/ml, respectively. In patients with a TDF mutation score of 3-5, the decrease in viral load was -0.8 +/- 1.0 log10 copies/ml and was considered possibly resistant. In the multivariate analysis, a TDF mutation score > or = 6, previous use of amprenavir, indinavir and lopinavir, and co-prescription of didanosine were associated with a worse virological response. The bootstrap analysis showed the robustness of the TDF mutation score. CONCLUSION: In ARV-experienced patients receiving TDF-containing regimens, a score derived from seven reverse transcriptase mutations was shown to be independently predictive of the virological response.

Persistence of multidrug-resistant HIV-1 without antiretroviral treatment 2 years after sexual transmission.

OBJECTIVES: To understand the virological mechanisms of 2-year persistence of multidrug-resistant virus without selective antiretroviral pressure in HIV-1-infected patients. PATIENTS AND METHODS: Two patients were contaminated recently by their HIV-1-infected partners, who had received, before the transmission, all available antiretroviral drugs and who exhibited a severe therapeutic failure. The resistance mutations analysis was performed by clonal sequencing of 1.2 kb of pol gene in plasma of index and sources patients. Sequencing of HIV-1 DNA was performed in PBMCs of index patients. RESULTS: Genotypic testing performed in index patients at time of seroconversion showed resistance mutations to three classes of drugs. All mutations were linked on the same viral genome and all quasispecies carried all mutations. No wild-type virus was detected. The same results were found in source patients and showed that all mutations were transmitted. In the index patients, all mutations persisted over 2 years without antiretroviral treatment. Moreover, the resistance mutations were all archived in the cellular reservoir. Viral load and CD4 count of index patients remained unchanged during 2 years of follow-up. DISCUSSION: Only multidrug-resistant viruses were detected in the source patients and could be transmitted in index patients. In the latter, an expansion of predominant multidrug-resistant quasispecies and the 'archival' of all mutations were observed. These results explain the persistence of mutations and suggest that it is highly difficult to return to a wild-type viral population, sensitive to an antiretroviral treatment. The treatment of index patients is limited and the major risk is the transmission of these multidrug-resistant viruses. This work was presented in part in the XII International HIV Drug Resistance Workshop, Los Cabos, Mexico, June 2003; and in the 2nd IAS Conference on HIV Pathogenesis & Treatment, Paris, France, July 2003.

Peptide insertions in reverse transcriptase pol gene of human immunodeficiency virus type 1 as a rare cause of persistent antiretroviral therapeutic failure.

Peptide insertions in codons 67-71 of the reverse transcriptase (RT) pol gene were detected in 11 (2.7%) of 414 genotypic analyses performed in a hospital cohort of 2900 outpatients with human immunodeficiency virus type 1 (HIV-1) infection. The duration of antiretroviral treatment (bi- or tri-therapy) before the detection of insertions ranged from 12 to 60 months. Dipeptide insertions were detected in ten patients, of which the most frequent was serine-serine. A monopeptide insertion was diagnosed once. The amino-acid composition patterns of insertions varied with time in five of the 11 patients. Peptide insertions were always associated with various patterns of pre-existing or appearing resistance mutations in the RT pol gene to different antiretroviral drugs. Genotypic-guided treatment resulted in virological and immunological improvement in two patients. In contrast, the remaining patients did not respond to any of the various antiretroviral regimens prescribed. Furthermore, various patterns of resistance mutations developed to the prescribed antiretroviral drugs, with AIDS-related conditions leading to death in two patients. It was concluded that peptide insertion in this region of the HIV-1 RT pol gene constitutes a rare cause of persistent therapeutic failure, and that management of such patients remains challenging despite successive genotypic analyses aimed at detecting mutations conferring antiretroviral drug resistance.

A cohort study of treatment-experienced HIV-1-infected patients treated with raltegravir: factors associated with virological response and mutations selected at failure.

This study aimed to identify factors associated with virological response (VR) to raltegravir (RAL)-containing regimens in 468 treatment-experienced but integrase inhibitor-naive HIV-1 patients receiving a RAL-containing regimen. VR was defined at Month 6 (M6) as HIV-1 RNA viral load (VL) <50 copies/mL. The impacts on VR of baseline integrase mutations, VL, CD4 count, genotypic sensitivity score for nucleoside reverse transcriptase inhibitors, non-nucleoside reverse transcriptase inhibitors and protease inhibitors, and the number of new antiretrovirals used for the first time associated with RAL were investigated. For patients with VL >50 copies/mL at M6, integrase mutations selected were characterised. Median baseline VL was 4.2 log(10)copies/mL (IQR 3.3-4.9 log(10) copies/mL) and CD4 count was 219 cells/mm(3) (IQR 96-368 cells/mm(3)). At M6, 71% of patients were responders. In multivariate analysis, baseline VL and CD4 count and ≥ 2 new antiretrovirals among darunavir, etravirine, maraviroc and enfuvirtide were associated with VR to RAL. Neither HIV-1 subtype nor baseline integrase polymorphisms were associated with VR to RAL. Among 63 failing patients at M6, selection of ≥ 1 change in the integrase gene was observed in 49 (77.8%), and 27/63 (42.9%) were considered as RAL-associated resistance mutations. Factors independently associated with the occurrence of ≥ 1 RAL-associated resistance mutation were VL at failure >3 log(10) and having no new drugs associated with RAL. RAL showed great potency in treatment-experienced patients. The number of new drugs associated with RAL was an important factor associated with VR. HIV-1 subtype and baseline integrase polymorphisms do not influence the RAL VR.