Immune-escape mutations and stop-codons in HBsAg develop in a large proportion of patients with chronic HBV infection exposed to anti-HBV drugs in Europe

Background: HBsAg immune-escape mutations can favor HBV-transmission also in vaccinated individuals, promote immunosuppression-driven HBV-reactivation, and increase fitness of drug-resistant strains. Stop-codons can enhance HBV oncogenic-properties. Furthermore, as a consequence of the overlapping structure of HBV genome, some immune-escape mutations or stop-codons in HBsAg can derive from drug-resistance mutations in RT. This study is aimed at gaining insight in prevalence and characteristics of immune-associated escape mutations, and stop-codons in HBsAg in chronically HBV-infected patients experiencing nucleos(t)ide analogues (NA) in Europe. Methods: This study analyzed 828 chronically HBV-infected European patients exposed to ≥ 1 NA, with detectable HBV-DNA and with an available HBsAg-sequence. The immune-associated escape mutations and the NA-induced immune-escape mutations sI195M, sI196S, and sE164D (resulting from drug-resistance mutation rtM204 V, rtM204I, and rtV173L) were retrieved from literature and examined. Mutations were defined as an aminoacid substitution with respect to a genotype A or D reference sequence. Results: At least one immune-associated escape mutation was detected in 22.1% of patients with rising temporal-trend. By multivariable-analysis, genotype-D correlated with higher selection of ≥ 1 immune-associated escape mutation (OR[95%CI]:2.20[1.32-3.67], P = 0.002). In genotype-D, the presence of ≥ 1 immune-associated escape mutations was significantly higher in drug-exposed patients with drug-resistant strains than with wild-type virus (29.5% vs 20.3% P = 0.012). Result confirmed by ana

Construction de promoteurs chimères dirigeant l'expression d'un gène rapporteur dépendante de la protéine EBNA1 de l'EBV dans les cellules épithéliales

Le virus d'Epstein-Barr (EBV) est associé à de nombreuses tumeurs épithéliales, notamment aux carcinomes du nasopharynx et carcinomes mammaires. Etant donné l'importance du nombre de patients touchés par les carcinomes mammaires et l'association de l'EBV aux tumeurs de plus mauvais pronostic, il est essentiel de proposer de nouvelles approches thérapeutiques dont la cible serait les cellules épithéliales tumorales EBV positives. L'expression de gènes viraux dans les cellules tumorales de carcinomes à l'EBV est une voie hautement sélective pour cibler l'expression de vecteurs thérapeutiques dans les cellules tumorales. La présence de la protéine virale EBNA1 dans les cellules tumorales infectées par l'EBV peut être utilisée pour activer dans ces cellules l'expression de vecteurs contenant un couple promoteur cis-activateur. La fixation de la protéine EBNA1 sir des séquences répétées de la région de la famille de répétion FR active dans les lymphocytes B de nombreux promoteurs EBV spécifiques et ubiquitaires. Nous avons étudié la combinaison du couple FR/ EBNAI avec différents promoteurs, EBV spécifiques (LMP2A actif dans les cellules épithéliales de carcinome du nasopharynx) et ubiquitaires (RSV et SV40) sur l'état d'activation et d'expression d'un gène rapporteur, le gène de la luciférase dans les cellules épithéliales. L'état d'activation et d'expression des différents promoteurs dans les cellules épithéliales mammaires infectées ou non par l'EBV a montré que l'activité des promoteurs est augmentée dans ces cellules quand FR et EBNAI1 sont présents. Cependant, cette sensibilité au couple FR/EBNA1 varie en fonction des promoteurs et du type cellulaire...PARIS7-Villemin (751102101) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

Analytical validation of hepatitis B core‐related antigen (HBcrAg) using dried blood spots (DBS)

International audienceLimited access to nucleic acid testing (NAT) to quantify HBV DNA levels, an essential tool to determine anti-HBV treatment eligibility, represents a significant barrier to scale up HBV diagnostic services in resource-limited countries. Hepatitis B core-related antigen (HBcrAg) has the potential to become an affordable alternative because of its low cost (US$ <15/assay) and strong correlation with HBV DNA levels in treatment-naïve patients. However, the current assay requires plasma or serum. To further facilitate its application to decentralized settings, we developed and evaluated a standardized procedure to quantify HBcrAg using dried blood spots as a tool to diagnose HBV-infected people with high viraemia. We evaluated the following elution method optimized to quantify HBcrAg: suspension of a punched blood-soaked disc (11 mm) of Whatman 903 Protein Saver Card in 450 µL of PBS 0.05% Tween 20, followed by an incubation for 4 h at room temperature and a centrifugation at 10,000 g for 10 minutes. 150 µL of DBS eluate was used to quantify HBcrAg using chemiluminescent enzyme immunoassay (LUMIPULSE® G600II, Fujirebio). The limit of detection of dried blood spot HBcrAg in relation with HBV DNA levels was 19,115 IU/mL across the five major HBV genotypes (A/B/C/D/E). A strong linear correlation was confirmed between dried blood spot HBcrAg and HBV DNA levels (r = 0.94, p < 0.0001) in samples with high viral loads (range: 3.7–7.0 log IU/mL). The coefficient of variation ranged between 4.0–11.2% for repeatability and 3.9–12.2% for reproducibility. Analytical specificity was 100% (95% CI: 83.9–100%) in HBV-negative samples. Using our elution method, it may be possible to identify HBV-infected patients with high viraemia who need antiviral therapy using dried blood spot and HBcrAg. A large-scale clinical validation is warranted in resource-limited countries

Profiles of liver fibrosis evolution during long-term tenofovir treatment in HIV-positive patients coinfected with hepatitis B Running title: Liver fibrosis evolution in HIV/HBV coinfection

International audienceBackground & AimsData on liver fibrosis evolution and its involvement in liver-related morbidity are scarce in individuals with human immunodeficiency virus (HIV) and hepatitis B virus (HBV) co-infection during treatment. We identified profiles of liver fibrosis evolution in coinfected patients undergoing tenofovir (TDF).MethodsWe included 169 HIV-HBV-coinfected patients on TDF-based antiretroviral therapy. Virological and clinical data were obtained at TDF-initiation and every 6-12 months. From data on non-invasive liver fibrosis assessments collected yearly (FibroTest®), we established clusters of individuals with similar liver fibrosis evolution using group-based trajectory models.ResultsFour profiles of liver fibrosis evolution were established from a median follow-up of 7.6 years (IQR = 3.1-13.1): low fibrosis with no progression (29.6%, profile A), low fibrosis with progression (22.5%, profile B), moderate fibrosis with high fluctuation (39.6%, profile C), and cirrhosis with no regression (8.3%, profile D). When compared to profile A, baseline HBeAg-positive status was associated with profiles B (P = .007) and C (P = .004), older age with profiles C (P < .001) and D (P = .001), exposure to second-generation protease inhibitors with profile C (P = .004), and CD4+ <500/mm3 at the last visit with profiles C (P = .02) and D (P = .002). Incident liver-related events occurred in profiles other than A (B, n = 1/38; C, n = 6/67; D, n = 3/14) and all five cases of hepatocellular carcinoma occurred in profiles C (n = 2) and D (n = 3).ConclusionsTDF-treated HIV-HBV coinfected individuals do not seem to benefit from comparable levels of liver fibrosis regression as in HBV mono-infection. Liver-related morbidity occurs mainly in those with fluctuating or consistently high fibrosis levels

Persistent HBV replication and serological response during up to 15 years of tenofovir-based antiretroviral therapy in HIV/HBV-coinfected patients: a multicentre prospective cohort study

International audienceObjectivesTo determine the extent of hepatitis B virus (HBV) suppression and its association with seroclearance of hepatitis ‘e’ antigen (HBeAg) and hepatitis B surface antigen (HBsAg) in HIV/HBV-coinfected patients undergoing long-term tenofovir-based antiretroviral therapy (ART).MethodsWe prospectively followed 165 HIV/HBV-coinfected patients undergoing tenofovir-based ART. Serum HBV-DNA viral loads and HBeAg and HBsAg status were obtained at tenofovir initiation and every 6–12 months. We calculated the proportion achieving virological response (VR, <60 IU/mL) during follow-up. We also calculated rates of HBeAg- and HBsAg-seroclearance, which were compared between those who achieved versus never achieved VR during follow-up using an Exact binomial test.ResultsDuring a median 8.1 years (IQR = 4.0–13.2) of tenofovir treatment, 152 (92.1%) patients were able to achieve VR and 13 (7.9%) never achieved VR (median HBV-DNA at the end of follow-up = 608 IU/mL, range = 67–52 400 000). The prevalence of individuals with detectable HBV-DNA (≥60 IU/mL) decreased during tenofovir treatment: 15.1% (n = 14/93) at 5 years, 3.2% (n = 2/62) at 10 years and, 3.2% (n = 1/31) at 15 years. 44/96 HBeAg-positive patients (6.15/100 person-years) had HBeAg-seroclearance and 13/165 patients overall (0.87/100 person-years) had HBsAg-seroclearance. No difference in HBeAg-seroclearance was observed between those who achieved versus never achieved VR (7.4 versus 3.7/100 person-years, P = 0.33), while HBsAg-seroclearance was only observed in those with VR (1.0 versus 0/100 person-years, P = 0.49; respectively). Individuals with VR also had a higher frequency of undetectable HIV-RNA during treatment (P < 0.001).ConclusionsDuring long-term tenofovir-based ART for HIV/HBV coinfection, persistent HBV viraemia is apparent, but becomes less frequent over time. HBsAg-seroclearance only occurred in those with full HBV and relatively high HIV suppression

Effect of viral replication and liver fibrosis on all-cause mortality in HIV/HBV coinfected individuals: a retrospective analysis of a 15-year longitudinal cohort

International audienceBackground: In individuals co-infected with HIV and hepatitis B virus (HBV), widespread tenofovir (TDF)-containing antiretroviral therapy (ART) has led to substantial decreases in HBV-DNA and HIV-RNA detection. However, the link between viral replication, liver fibrosis, and mortality remains unclear.Methods: 300 HIV-HBV co-infected individuals undergoing ART were prospectively followed. Virological and clinical data were obtained at baseline and every 6-12 months. We quantified the association between HBV-DNA, HIV-RNA, and liver fibrosis with risk of all-cause mortality using a joint longitudinal-survival model. Viral detection, viral loads, and time-averaged cumulative viral loads of HIV and HBV were modeled as three separate exposures.Results:During a median 10.5 years (IQR=4.0-14.6), the proportion undergoing TDF-containing ART (baseline=18.7%, end of follow-up=79.1%) and with undetectable HBV-DNA (baseline=36.7%, end of follow-up=94.8%) substantially increased. HIV-RNA was mostly undetectable during follow-up (76.6%). 42 participants died (incidence rate=1.30/100person-years, 95%CI=0.96-1.76). The leading causes of death were non-AIDS/non-liver-related malignancies (28.6%), followed by liver-related (16.7%), AIDS-related (16.7%), and other (16.7%). All-cause mortality was associated with HBV-DNA viral load (adjusted-HR per log10IU/mL=1.41, 95%CI=1.04-1.93, p=0.03) or time-averaged cumulative HBV-DNA (adjusted-HR per log10IU-years=1.37, 95%CI=1.03-1.83, p=0.03), but not undetectable HBV-DNA (adjusted-HR=0.30, 95%CI=0.08-1.09, p=0.08). Advanced liver fibrosis at baseline was also associated with increased mortality rates (adjusted-HR=2.35, 95%CI=1.16-4.76, p=0.02). No significant association between HIV-RNA replication and mortality was observed.Conclusions: Concurrent and historical HBV replication and liver fibrosis are important drivers of all-cause mortality in largely TDF-treated HIV-HBV co-infected individuals, despite one-fifth of deaths being liver-related. HBV-DNA and liver fibrosis remain important prognostic indicators for this patient population

Kinetics of Hepatitis B Core–Related Antigen and Anti–Hepatitis B Core Antibody and Their Association With Serological Response in Human Immunodeficiency Virus–Hepatitis B Coinfection

International audienceBackground: The aim of the current study was to describe the kinetics of quantified hepatitis B core-related antigen (qHBcrAg) and quantified anti-hepatitis B core antibody (qAnti-HBc) during tenofovir (TDF) treatment and assess their ability to predict hepatitis B e antigen (HBeAg) seroclearance in patients coinfected with human immunodeficiency virus (HIV) and hepatitis B virus.Methods: Serum qHBcrAg, qAnti-HBc, and hepatitis B virus DNA were obtained at TDF initiation and every 6-12 months. The on-treatment kinetics of qHBcrAg (ΔqHBcrAg) and qAnti-HBc (ΔqAnti-HBc) were estimated using mixed-effect linear regression. Hazard ratios (HRs) assessing the association between markers and HBeAg seroclearance were calculated using proportional hazards regression, and the sensitivity (Se) and specificity (Sp) of marker levels in predicting HBeAg seroclearance were assessed using time-dependent receiving operating characteristic curves.Results: During a median of 4.6 years, the cumulative incidences of hepatitis B surface antigen and HBeAg seroclearance were 3.2% (n = 5 of 158) and 27.4% (n = 26 of 95), respectively. ΔqHBcrAg was biphasic in HBeAg-positive patients (-0.051 and -0.011 log10 U/mL/mo during ≤18 and >18 months, respectively) and monophasic in HBeAg-negative patients. ΔqAnti-HBc was monophasic regardless of HBeAg status. In HBeAg-positive patients, baseline qHBcrAg and qAnti-HBc levels were associated with HBeAg seroclearance (adjusted HR, 0.48/log10 U/mL [95% confidence interval, .33-.70] and unadjusted HR, 1.49/log10 Paul Ehrlich Institute units/mL [1.08-2.07], respectively). Cutoffs with the highest accuracy in predicting HBeAg seroclearance at 36 months were qHBcrAg <6.5 log10 U/mL at month 24 (Se, 1; Sp, 0.58) and baseline qAnti-HBc ≥4.1 log10 Paul Ehrlich Institute units/mL (Se, 0.42; Sp, 0.81).Conclusions: In coinfected patients undergoing TDF, qHBcrAg/qAnti-HBc could be of use in monitoring HBeAg seroclearance