Search

The success of antiretroviral treatment (ART) is threatened by the emergence of drug resistance mutations (DRM). Since Brazil presents the largest number of people living with HIV (PLWH) in South America we aimed at understanding the dynamics of DRM in this country. We analyzed a total of 20,226 HIV-1 sequences collected from PLWH undergoing ART between 2008-2017. Results show a mild decline of DRM over the years but an increase of the K65R reverse transcriptase mutation from 2.23% to 12.11%. This increase gradually occurred following alterations in the ART regimens replacing zidovudine (AZT) with tenofovir (TDF). PLWH harboring the K65R had significantly higher viral loads than those without this mutation ( p < 0.001). Among the two most prevalent HIV-1 subtypes (B and C) there was a significant ( p < 0.001) association of K65R with subtype C (11.26%) when compared with subtype B (9.27%). Nonetheless, evidence for K65R transmission in Brazil was found both for C and B subtypes. Additionally, artificial neural network-based immunoinformatic predictions suggest that K65R could enhance viral recognition by HLA-B27 that has relatively low prevalence in the Brazilian population. Overall, the results suggest that tenofovir-based regimens need to be carefully monitored particularly in settings with subtype C and specific HLA profiles.

Despite significant advances in HIV treatment over the past 30 years, critical barriers to an HIV cure persist. The HIV reservoir, defined at both the cellular and anatomical level, constitutes the main barrier to cure. While the mechanisms underlying the reservoir are not yet well understood, one theory to explain persistence at the anatomical level is that subtherapeutic exposure to antiretroviral therapy (ART) within certain tissue compartments permits ongoing replication. Characterizing ART pharmacology throughout the body is important in the context of these potential pharmacologic sanctuaries and for maximizing the probability of success with forthcoming cure strategies that rely on latency reversal and require ART to prevent reseeding the reservoir. In this review, we provide a comprehensive overview of ART and latency reversal agent distribution at the site of action for HIV cure (i.e., anatomical sites commonly associated with HIV persistence, such as lymphoid organs and the central nervous system). We also discuss methodologic approaches that provide insight into HIV cure pharmacology, including experimental design and advances within the computational, pharmaceutical, and analytical chemistry fields. The information discussed in this review will assist in streamlining the development of investigational cure strategies by providing a roadmap to ensure therapeutic exposure within the site of action for HIV cure., (© 2021 The Authors. Clinical Pharmacology & Therapeutics © 2021 American Society for Clinical Pharmacology and Therapeutics.)

Antibody-based strategies have been introduced for a number of disease states, but represent a novel approach in the management of human immunodeficiency virus (HIV). Ibalizumab and leronlimab are monoclonal antibodies with unique mechanisms as a CD4-directed post-attachment inhibitor and a C-C chemokine receptor type 5-directed inhibitor, respectively. These antibody-based strategies are generally well tolerated, have a favourable pharmacokinetic profile allowing for less-frequent dosing, and have a high barrier to resistance. Ibalizumab is currently approved by the US Food and Drug Administration (US FDA) for management of multi-drug-resistant (MDR) HIV infection in patients who are failing their current regimens. Clinical data demonstrated impressive antiretroviral activity with ibalizumab among a complex HIV population in combination with an optimized background regimen, where limited therapeutic options exist. To date, leronlimab has not been granted approval by the US FDA, but has been designated fast-track status. Leronlimab is being studied as a maintenance monotherapy agent in virologically suppressed patients, as well as for treatment of MDR HIV infection in patients who are failing their current regimens. Currently available data in both of these potential areas appear promising for leronlimab. The mechanism of action, pharmacokinetic profile, efficacy and safety of these novel antibody-based strategies represent an advance in the management of HIV. Future studies and post-marketing experience will further determine longer-term clinical efficacy, safety and resistance data for ibalizumab and leronlimab., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Covid-19 infection caused by the novel coronavirus SARS-COV-2 continues to be a major global health challenge. Till date, no drug has been approved for the treatment of this infection. A number of medications have been proposed and there are ongoing clinical trials around the world to find a suitable treatment. A recent randomised control trial compared lopinavir/ritonavir with standard care among 199 patients with severe Covid-19 infection and concluded that there was no significant reduction in mortality rate with lopinavir/ritonavir. However, there are a few important lessons which may be learnt from the study apart from the statistical reduction in mortality rate. There was a numerical reduction in mortality rate, less intensive care unit stay and less complications in the lopinavir-ritonavir group. This article points out some of those important lessons with some suggestions for future clinical trials., Competing Interests: Declaration of Competing Interest None declared., (Copyright © 2020. Published by Elsevier B.V.)

We determined the disinfection efficacy and inactivation mechanisms of peracetic acid (PAA)-based sanitizer using pH values relevant for vegetable sanitation against rotavirus (RV) and Tulane virus (TV; a human norovirus surrogate). TV was significantly more resistant to PAA disinfection than RV: for a 2-log 10 reduction of virus titer, RV required 1 mg/liter PAA for 3.5 min of exposure, while TV required 10 mg/liter PAA for 30 min. The higher resistance of TV can be explained, in part, by significantly more aggregation of TV in PAA solutions. The PAA mechanisms of virus inactivation were explored by quantifying (i) viral genome integrity and replication using reverse transcription-quantitative PCR (RT-qPCR) and (ii) virus-host receptor interactions using a cell-free binding assay with porcine gastric mucin conjugated with magnetic beads (PGM-MBs). We observed that PAA induced damage to both RV and TV genomes and also decreased virus-receptor interactions, with the latter suggesting that PAA damages viral proteins important for binding its host cell receptors. Importantly, the levels of genome-versus-protein damage induced by PAA were different for each virus. PAA inactivation correlated with higher levels of RV genome damage than of RV-receptor interactions. For PAA-treated TV, the opposite trends were observed. Thus, PAA inactivates each of these viruses via different molecular mechanisms. The findings presented here potentially contribute to the design of a robust sanitation strategy for RV and TV using PAA to prevent foodborne disease. IMPORTANCE In this study, we examined the inactivation mechanisms of peracetic acid (PAA), a sanitizer commonly used for postharvest vegetable washing, for two enteric viruses: Tulane virus (TV) as a human norovirus surrogate and rotavirus (RV). PAA disinfection mechanisms for RV were mainly due to genome damage. In contrast, PAA disinfection in TV was due to damage of the proteins important for binding to its host receptor. We also observed that PAA triggered aggregation of TV to a much greater extent than RV. These studies demonstrate that different viruses are inactivated via different PAA mechanisms. This information is important for designing an optimal sanitation practice for postharvest vegetable washing to minimize foodborne viral diseases., (Copyright © 2020 American Society for Microbiology.)

We describe a study leading to the discovery of compound 11 , a pan-genotypic hepatitis C virus (HCV) nonstructural protein 5A (NS5A) inhibitor with excellent potency, metabolic stability, and pharmacokinetics (PK). Compound 11 incorporating a 4-silapiperidine group was discovered by further optimizing our previous lead with a triethylsilyl moiety. It displayed great potency against genotype 1 subtype a (GT1a), -1b, -2a, -3a, -4a, -5a, and -6a with an EC 50 range of 0.33-17 pM and improved potency against the resistance-associated variant GT1a&#95;M28T. Pharmacokinetics (PK) study indicated that compound 11 has reasonable PK exposures with a high liver distribution in preclinical animal species (mouse, rat, and dog). The results of a 14 day repeat-dose toxicity study identified the safety of compound 11 .

Background: The effect of baseline resistance-associated substitutions (RAS) on the sustained virologic response at 12 weeks (SVR12) among chronic hepatitis C (CHC) patients receiving the second generation, pan-genotypic glecaprevir/pibrentasvir (G/P) regimen is unclear., Aim: To assess the effect of RAS on the SVR12 in CHC patients treated with G/P regimen., Methods: The EMBASE, MEDLINE and Cochrane central register of controlled trials databases were searched for relevant studies published before 1 March 2019. The principal outcome was to compare the SVR12 in CHC patients with and without baseline RAS, particularly in genotype-1, genotype-3 and direct-acting anti-virals (DAAs) failure patients. The outcomes were pooled using a random-effects model and odds ratio (OR) was calculated. The risk of bias was assessed using the Cochrane risk of bias tools for randomised and nonrandomised interventional studies., Results: After initially identifying 410 studies, 3302 patients from 17 studies were included. Among 50 cases of virologic failures, 48% had genotype-3 infection, 44% genotype-1 infection and 36% DAA-failure patients. Baseline RAS were present in 44(88%) patients. The most common NS5a and NS3 mutations were Y93H and A166S respectively. The odds of SVR12 were significantly reduced in patients with any baseline RAS (NS3 and/or NS5a) (OR 0.32, 95%C I[0.15, 0.65], I 2  = 0%) and NS5a substitutions (OR 0.36, 95%CI [0.18,0.73]). The impact of RAS on SVR12 was significant among genotype-3 patients, but not among genotype-1 or DAA-failure cases. The presence of Y93H and A30K mutations significantly impacted SVR12 rates in genotype-3 patients., Conclusion: Baseline NS3 or NS5a RAS, especially the NS5a substitutions-A30K, Y93H, decrease the odds of achieving SVR12 in genotype-3 CHC patients., (© 2020 John Wiley & Sons Ltd.)

HIV diversification facilitates immune escape and complicates antiretroviral therapy. In this study, we take advantage of a humanized-mouse model to probe the contribution of APOBEC3 mutagenesis to viral evolution. Humanized mice were infected with isogenic HIV molecular clones (HIV-WT, HIV-45G, and HIV-ΔSLQ) that differ in their abilities to counteract APOBEC3G (A3G). Infected mice remained naive or were treated with the reverse transcriptase (RT) inhibitor lamivudine (3TC). Viremia, emergence of drug-resistant variants, and quasispecies diversification in the plasma compartment were determined throughout infection. While both HIV-WT and HIV-45G achieved robust infection, over time, HIV-45G replication was significantly reduced compared to that of HIV-WT in the absence of 3TC treatment. In contrast, treatment responses differed significantly between HIV-45G- and HIV-WT-infected mice. Antiretroviral treatment failed in 91% of HIV-45G-infected mice, while only 36% of HIV-WT-infected mice displayed a similar negative outcome. Emergence of 3TC-resistant variants and nucleotide diversity were determined by analyzing 155,462 single HIV reverse transcriptase gene ( RT ) and 6,985 vif sequences from 33 mice. Prior to treatment, variants with genotypic 3TC resistance (RT-M184I/V) were detected at low levels in over a third of all the animals. Upon treatment, the composition of the plasma quasispecies rapidly changed, leading to a majority of circulating viral variants encoding RT-184I. Interestingly, increased viral diversity prior to treatment initiation correlated with higher plasma viremia in HIV-45G-infected animals, but not in HIV-WT-infected animals. Taken together, HIV variants with suboptimal anti-A3G activity were attenuated in the absence of selection but displayed a fitness advantage in the presence of antiretroviral treatment. IMPORTANCE Both viral (e.g., RT) and host (e.g., A3G) factors can contribute to HIV sequence diversity. This study shows that suboptimal anti-A3G activity shapes viral fitness and drives viral evolution in the plasma compartment in humanized mice., (Copyright © 2020 American Society for Microbiology.)

Background: HIV drug resistance and suboptimal adherence are the main reasons for treatment failure among HIV-infected individuals. As genotypic resistance testing is not routinely available in resource-limited settings such as Uganda, data on transmitted and acquired resistance are sparse., Methods: This observational follow-up study assessed the virological outcomes of patients diagnosed with virological failure or transmitted HIV drug resistance in 2015 at the adults' outpatient clinic of the Infectious Diseases Institute in Kampala, Uganda. Initially, 2430 patients on antiretroviral therapy (ART) underwent virological monitoring, of which 190 had virological failure and were subsequently eligible for this follow-up study. Nine patients diagnosed with transmitted drug resistance were eligible. In patients with a viral load > 1000 copies/mL, genotypic resistance testing was performed., Results: Of 190 eligible patients, 30 (15.8%) had either died or were lost to follow-up. A total of 148 (77.9%) were included, of which 98 had had a change of ART regimen, and 50 had received adherence counseling only. The majority was now on second-line ART (N = 130, 87.8%). The median age was 39 years (interquartile range: 32-46), and 109 (73.6%) were women. Virological failure was diagnosed in 29 (19.6%) patients, of which 24 (82.8%) were on second-line ART. Relevant drug resistance was found in 25 (86.2%) cases, of which 12 (41.3%) carried dual and 7 (24.1%) triple drug resistance., Conclusion: Two years after initial virological failure, most patients followed up by this study had a successful virological outcome. However, a significant proportion either continued to fail or died or was lost to follow-up.

Background: Contemporary antiretroviral therapies (ART) and management strategies have diminished both human immunodeficiency virus (HIV) treatment failure and the acquired resistance to drugs in resource-rich regions, but transmission of drug-resistant viruses has not similarly decreased. In low- and middle-income regions, ART roll-out has improved outcomes, but has resulted in increasing acquired and transmitted resistances. Our objective was to review resistance to ART drugs and methods to detect it, and to provide updated recommendations for testing and monitoring for drug resistance in HIV-infected individuals., Methods: A volunteer panel of experts appointed by the International Antiviral (formerly AIDS) Society-USA reviewed relevant peer-reviewed data that were published or presented at scientific conferences. Recommendations were rated according to the strength of the recommendation and quality of the evidence, and reached by full panel consensus., Results: Resistance testing remains a cornerstone of ART. It is recommended in newly-diagnosed individuals and in patients in whom ART has failed. Testing for transmitted integrase strand-transfer inhibitor resistance is currently not recommended, but this may change as more resistance emerges with widespread use. Sanger-based and next-generation sequencing approaches are each suited for genotypic testing. Testing for minority variants harboring drug resistance may only be considered if treatments depend on a first-generation nonnucleoside analogue reverse transcriptase inhibitor. Different HIV-1 subtypes do not need special considerations regarding resistance testing., Conclusions: Testing for HIV drug resistance in drug-naive individuals and in patients in whom antiretroviral drugs are failing, and the appreciation of the role of testing, are crucial to the prevention and management of failure of ART.

Safe and effective lifelong treatment to control human immunodeficiency virus (HIV) infection is one of the greatest scientific and public health achievements of the past century. The majority of infected individuals able to maintain a daily oral regimen now have a normal or near-normal life expectancy. More than 30 approved drugs and dozens of formulations have produced thousands of possible drug combinations used clinically in the past, but today most patients receive only a handful of high priority and rigorously tested regimens. Unique features of antiretroviral therapy include the need for lifelong treatment to control virus replication and the possibility of rapid emergence of permanent drug resistance if these agents are not properly used. Although three-drug combination oral regimens have radically altered the course of this epidemic, the future will include long-acting injectable and implantable drugs and devices to treat and prevent infection., (© 2018 American Society for Clinical Pharmacology and Therapeutics.)

Direct-acting antivirals (DAAs) targeting NS5A are broadly effective against hepatitis C virus (HCV) infections, but sustained virological response rates are generally lower in patients infected with genotype (gt)-1a than gt-1b viruses. The explanation for this remains uncertain. Here, we adopted a highly accurate, ultra-deep primer ID sequencing approach to intensively study serial changes in the NS5A-coding region of HCV in gt-1a- and gt-1b-infected subjects receiving a short course of monotherapy with the NS5A inhibitor, elbasvir. Low or undetectable levels of viremia precluded on-treatment analysis in gt-1b-infected subjects, but variants with the resistance-associated substitution (RAS) Y93H in NS5A dominated rebounding virus populations following cessation of treatment. These variants persisted until the end of the study, two months later. In contrast, while Y93H emerged in multiple lineages and became dominant in subjects with gt-1a virus, these haplotypes rapidly decreased in frequency off therapy. Substitutions at Q30 and L31 emerged in distinctly independent lineages at later time points, ultimately coming to dominate the virus population off therapy. Consistent with this, cell culture studies with gt-1a and gt-1b reporter viruses and replicons demonstrated that Y93H confers a much greater loss of replicative fitness in gt-1a than gt-1b virus, and that L31M/V both compensates for the loss of fitness associated with Q30R (but not Y93H) and also boosts drug resistance. These observations show how differences in the impact of RASs on drug resistance and replicative fitness influence the evolution of gt-1a and gt-1b viruses during monotherapy with an antiviral targeting NS5A., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Drug resistance is a major obstacle faced by therapist in treating HIV infected patients. The reason behind these phenomena is either protein mutation or the changes in gene expression level that induces resistance to drug treatments. These mutations affect the drug binding activity, hence resulting in failure of treatment. Therefore, it is necessary to conduct resistance testing in order to carry out HIV effective therapy. This study combines both sequence and structural features for predicting HIV resistance by applying SVM and Random Forests classifiers. The model was tested on the mutants of HIV-1 protease and reverse transcriptase. Taken together the features we have used in our method, total contact energies among multiple mutations have a strong impact in predicting resistance as they are crucial in understanding the interactions of HIV mutants. The combination of sequence-structure features offers high accuracy with support vector machines as compared to Random Forests classifier. Both single and acquisition of multiple mutations are important in predicting HIV resistance to certain drug treatments. We have discovered the practicality of these features; hence, these can be used in the future to predict resistance for other complex diseases.

The product limit or Kaplan-Meier (KM) estimator is commonly used to estimate the survival function in the presence of incomplete time to event. Application of this method assumes inherently that the occurrence of an event is known with certainty. However, the clinical diagnosis of an event is often subject to misclassification due to assay error or adjudication error, by which the event is assessed with some uncertainty. In the presence of such errors, the true distribution of the time to first event would not be estimated accurately using the KM method. We develop a method to estimate the true survival distribution by incorporating negative predictive values and positive predictive values, into a KM-like method of estimation. This allows us to quantify the bias in the KM survival estimates due to the presence of misclassified events in the observed data. We present an unbiased estimator of the true survival function and its variance. Asymptotic properties of the proposed estimators are provided, and these properties are examined through simulations. We demonstrate our methods using data from the Viral Resistance to Antiviral Therapy of Hepatitis C study., (Copyright © 2017 John Wiley & Sons, Ltd.)

Treatment of HIV has reduced HIV/AIDS-related mortality. Sustaining >90% virologic suppression in sub-Saharan Africa requires decentralized care and prevention services to rural communities. In Zimbabwe, the number of people receiving antiretroviral treatment (ART) has increased rapidly. However, access to treatment monitoring tools such as viral load and drug resistance testing is limited. We assessed virologic treatment outcomes among ART recipients in Nyamutora, a rural community receiving bimonthly ART and prevention services. We enrolled all ART recipients (143) at 6-monthly visits in the Nyamutora community in 2014 and 2015. Whole blood samples were collected in K-EDTA tubes, transported to Harare for CD4 counts and viral load testing, and genotype was obtained in participants with viral loads >1,000 copies/ml. Ages ranged from 2 to 75 years (median 43 years) with a median 42 months on ART at follow-up. Eight of 143 (6%) had viral loads >1,000 copies/ml at one of the 3 visits, 7 on first-line nevirapine (NVP)-based ART and 1 on second-line LPV/r-based ART. Seven participants had sequence data available, and five had drug resistance mutations, K65R, T69N, K101E, K103N, Y181C/I, M184V, and G190A. Virologic failure (p = .001) and drug resistance mutations (p = .01) on first-line NVP-based ART were associated with younger age by univariate exact logistic regression. The participants had high viral suppression (94%) despite less than optimal (NVP based) ART regimens without laboratory monitoring. Virologic failure and drug resistance were higher among children and adolescents. Effective ART delivery to the community achieved high rates of virologic suppression and minimal drug resistance.

Purpose: The chemistry, pharmacology, pharmacodynamics, pharmacokinetics, efficacy, safety, dosage, administration, and role of elbasvir-grazoprevir in the treatment of hepatitis C virus (HCV) infection are reviewed., Summary: Elbasvir-grazoprevir was recently approved by the Food and Drug Administration for the treatment of chronic HCV genotype 1 or 4 infections with or without ribavirin in patients with or without compensated cirrhosis. Elbasvir exhibits antiviral activity against HCV genotypes 1a, 1b, 2a, 3a, and 4a. Elbasvir-grazoprevir undergoes fecal excretion, does not require dosage adjustment in patients with renal impairment, and is contraindicated in moderate and severe hepatic impairment. In Phase II and III clinical trials, elbasvir-grazoprevir administered orally for 12 weeks was shown to achieve a high sustained virological response 12 weeks after the end of treatment. Elbasvir-grazoprevir is a once-daily, fixed-dose combination tablet that can be taken without regard to food. The adverse drug reactions most commonly reported include fatigue, headache, and nausea. Elbasvir-grazoprevir is indicated with ribavirin for treatment-naive and treatment-experienced patients with genotype 1a with baseline NS5A polymorphisms, for treatment-experienced patients with genotype 1b, and for treatment-experienced patients with genotype 4., Conclusion: Elbasvir-grazoprevir achieves a high cure rate in the treatment of patients with chronic HCV with a once-daily oral regimen and without serious adverse effects; however, it requires close monitoring of liver function values. It is an effective option for patients with HCV genotype 1a, 1b, or 4 with or without compensated cirrhosis and is a particularly attractive option in patients with chronic kidney disease receiving hemodialysis and in patients with HIV coinfection., (Copyright © 2017 by the American Society of Health-System Pharmacists, Inc. All rights reserved.)

Influenza A virus envelop protein hemagglutinin (HA) plays important roles in viral entry. We previously have reported that MBX2546, a novel influenza A virus inhibitor, binds to HA and inhibits HA-mediated membrane fusion. In this report, we show that (i) both binding and stabilization of HA by MBX2546 are required for the inhibition of viral infection and (ii) the binding of HA by MBX2546 represses the low-pH-induced conformational change in the HA, which is a prerequisite for membrane fusion. Mutations in MBX2546-resistant influenza A/PR/8/34 (H1N1) viruses are mapped in the HA stem region near the amino terminus of HA2. Finally, we have modeled the binding site of MBX2546 using molecular dynamics and find that the resulting structure is in good agreement with our results. Together, these studies underscore the importance of the HA stem loop region as a potential target for therapeutic intervention.

Asthma and chronic obstructive pulmonary disease (COPD) exacerbations are commonly associated with respiratory syncytial virus (RSV), rhinovirus (RV) and influenza A virus (IAV) infection. The ensuing airway inflammation is resistant to the anti-inflammatory actions of glucocorticoids (GCs). Viral infection elicits transforming growth factor-β (TGF-β) activity, a growth factor we have previously shown to impair GC action in human airway epithelial cells through the activation of activin-like kinase 5 (ALK5), the type 1 receptor of TGF-β. In the current study, we examine the contribution of TGF-β activity to the GC-resistance caused by viral infection. We demonstrate that viral infection of human bronchial epithelial cells with RSV, RV or IAV impairs GC anti-inflammatory action. Poly(I:C), a synthetic analog of double-stranded RNA, also impairs GC activity. Both viral infection and poly(I:C) increase TGF-β expression and activity. Importantly, the GC impairment was attenuated by the selective ALK5 (TGFβRI) inhibitor, SB431542 and prevented by the therapeutic agent, tranilast, which reduced TGF-β activity associated with viral infection. This study shows for the first time that viral-induced glucocorticoid-insensitivity is partially mediated by activation of endogenous TGF-β., Competing Interests: The authors have declared that no competing interests exist.

Background: In resource-limited settings, where resistance testing is unavailable, confirmatory testing for patients with high viral loads (VL) delays antiretroviral therapy (ART) switches for persons with resistance. We developed a risk score algorithm to predict need for ART change by identifying resistance among persons with persistently elevated VL., Methods: We analyzed data from a Phase IV open-label trial. Using logistic regression, we identified demographic and clinical characteristics predictive of need for ART change among participants with VLs ≥1000 copies/ml, and assigned model-derived scores to predictors. We designed three models, including only variables accessible in resource-limited settings., Results: Among 290 participants with at least one VL ≥1000 copies/ml, 51 % (148/290) resuppressed and did not have resistance testing; among those who did not resuppress and had resistance testing, 47 % (67/142) did not have resistance and 53 % (75/142) had resistance (ART change needed for 25.9 % (75/290)). Need for ART change was directly associated with higher baseline VL and higher VL at time of elevated measure, and inversely associated with treatment duration. Other predictors included body mass index and adherence. Area under receiver operating characteristic curves ranged from 0.794 to 0.817. At a risk score ≥9, sensitivity was 14.7-28.0 % and specificity was 96.7-98.6 %., Conclusions: Our model performed reasonably well and may be a tool to quickly transition persons in need of ART change to more effective regimens when resistance testing is unavailable. Use of this algorithm may result in public health benefits and health system savings through reduced transmissions of resistant virus and costs on laboratory investigations.

Background: The assessment of adherence to antiretroviral therapy (ART) is important in order to predict treatment outcomes. Lamivudine (3TC) is one of the most widely used NRTIs in China, but its concentrations in hair and association with virologic failure and drug resistance have not been studied., Methods: We conducted a cross-sectional survey to investigate 3TC concentrations in hair as a predictor of virologic failure and drug resistance among HIV patients receiving free ART. We also compared the capacity of hair 3TC concentrations with self-reported adherence in predicting virologic responses. Hair 3TC concentrations were detected through the LC-MS/MS system., Results: In patients without HIV drug resistance (HIVDR), with a threshold hair 3TC concentration of 260 ng/g, the sensitivity and specificity in predicting virologic suppression were 76.9% and 89.9%, respectively. Some factors, including CD4+ cell counts, initial treatment regimens with 3TC, and current regimens with second-line drugs, influenced the association between hair 3TC concentrations and virologic suppression. In patients who experienced virologic failure with HIVDR, with a threshold of 180 ng/g, the sensitivity and specificity were 70.0% and 74.4%, respectively. Hair 3TC concentrations had higher sensitivity and specificity in predicting virologic failure and drug resistance than self-reported adherence., Conclusions: The hair 3TC concentration was a stronger indicator than self-reported adherence in predicting virologic failure and drug resistance in HIV patients receiving free ART.

Purpose: The use of a three-drug regimen for the prevention of perinatal transmission of zidovudine- and nevirapine-resistant HIV is described., Summary: A 17-year-old Hispanic woman infected with HIV arrived at our clinic for the management of her first pregnancy. The patient was in her second trimester of her pregnancy, had not previously been treated with antiretroviral therapy, and was only taking daily prenatal vitamins at the time of her first clinic visit. Her HIV RNA viral load was 240 copies/mL, and the virus was resistant to both zidovudine and nevirapine. Nelfinavir (compounded suspension), lamivudine, and zidovudine were prescribed for the mother, though she was generally nonadherent to therapy. Nelfinavir, lamivudine, and zidovudine were initiated for the newborn within eight hours of delivery. Six months later, the patient returned to the clinic in the first trimester of her second pregnancy. At this visit, her HIV RNA viral load was 120 copies/mL. After the birth of her second child, the infant received the same regimen received by her firstborn: zidovudine 4 mg/kg orally twice daily for six weeks, lamivudine 2 mg/kg orally twice daily for two weeks, and nelfinavir 55 mg/kg orally twice daily for two weeks. At four months of age, each infant was found to be HIV-negative., Conclusion: A prophylactic regimen that included nelfinavir, lamivudine, and zidovudine was used to prevent perinatal transmission of HIV in two neonates. The regimen was well tolerated, and both infants were determined to be HIV-negative at four months of age., (Copyright © 2016 by the American Society of Health-System Pharmacists, Inc. All rights reserved.)

HIV-1 protease variants resist drugs by active and non-active-site mutations. The active-site mutations, which are the primary or first set of mutations, hamper the stability of the enzyme and resist the drugs minimally. As a result, secondary mutations that not only increase protein stability for unhindered catalytic activity but also resist drugs very effectively arise. While the mechanism of drug resistance of the active-site mutations is through modulating the active-site pocket volume, the mechanism of drug resistance of the non-active-site mutations is unclear. Moreover, how these allosteric mutations, which are 8-21 Å distant, communicate to the active site for drug efflux is completely unexplored. Results from molecular dynamics simulations suggest that the primary mechanism of drug resistance of the secondary mutations involves opening of the flexible protease flaps. Results from both residue- and community-based network analyses reveal that this precise action of protease is accomplished by the presence of robust communication paths between the mutational sites and the functionally relevant regions: active site and flaps. While the communication is more direct in the wild type, it traverses across multiple intermediate residues in mutants, leading to weak signaling and unregulated motions of flaps. The global integrity of the protease network is, however, maintained through the neighboring residues, which exhibit high degrees of conservation, consistent with clinical data and mutagenesis studies.

Influenza is a common disease affecting many children each year. In a number of cases, particularly in children <2 years old and in those with severe chronic underlying disease, influenza can be complicated by lower respiratory tract infections, acute otitis media, rhinosinusitis, febrile seizures, dehydration or encephalopathy. Oseltamivir is the influenza virus drug that is most commonly studied in children for both the treatment and prevention of influenza. To avoid the risk that children with mild influenza or patients suffering from different viral infections receive oseltamivir, oseltamivir treatment should be recommended only in severe influenza cases, especially if confirmed by reliable laboratory tests. However, therapy must be initiated considering the risk of complications and the presence of severe clinical manifestations at age- and weight-appropriate doses. Because the vaccine remains the best option for preventing influenza and its complications, prophylaxis using oseltamivir should only be considered in select patients.

A key target in the treatment of HIV-1/AIDS has been the viral protease. Here we first studied in silico the evolution of protease resistance. Primary active site resistance mutations were found to weaken interactions between protease and both inhibitor and substrate P4-P4' residues. We next studied the effects of secondary resistance mutations, often distant from the active site, on protease binding to inhibitors and substrates. Those secondary mutations contributed to the rise of multi-drug resistance while also enhancing viral replicative capacity. Here many secondary resistance mutations were found in the HIV-1 protease substrate-grooves, one on each face of the symmetrical protease dimer. The protease active site binds substrate P4-P4' residues, while the substrate-groove allows the protease to bind residues P12-P5/P5'-P12', for a total of twenty-four residues. The substrate-groove secondary resistance mutations were found to compensate for the loss of interactions between the inhibitor resistant protease active site and substrate P4-P4' residues, due to primary resistance mutations, by increasing interactions with substrate P12-P5/P5'-P12' residues. In vitro experiments demonstrated that a multi-drug resistant protease with substrate-groove resistance mutations was slower than wild-type protease in cleaving a peptide substrate, which did not allow for substrate-groove interactions, while it had similar activity as wild-type protease when using a Gag polyprotein in which cleavage-site P12-P5/P5'-P12' residues could be bound by the protease substrate-grooves. When the Gag MA/CA cleavage site P12-P5/P5'-P12' residues were mutated the multi-drug resistant protease cleaved the mutant Gag significantly slower, indicating the importance of the protease S-grooves in binding to substrate., (Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)

Unlabelled: During HIV-1 infection of cells, the viral capsid plays critical roles in reverse transcription and nuclear entry of the virus. The capsid-targeting small molecule PF74 inhibits HIV-1 at early stages of infection. HIV-1 resistance to PF74 is complex, requiring multiple amino acid substitutions in the viral CA protein. Here we report the identification and analysis of a novel PF74-resistant mutant encoding amino acid changes in both domains of CA, three of which are near the pocket where PF74 binds. Interestingly, the mutant virus retained partial PF74 binding, and its replication was stimulated by the compound. The mutant capsid structure was not significantly perturbed by binding of PF74; rather, the mutations inhibited capsid interactions with CPSF6 and Nup153 and altered HIV-1 dependence on these host factors and on TNPO3. Moreover, the replication of the mutant virus was markedly impaired in activated primary CD4(+) T cells and macrophages. Our results suggest that HIV-1 escapes a capsid-targeting small molecule inhibitor by altering the virus's dependence on host factors normally required for entry into the nucleus. They further imply that clinical resistance to inhibitors targeting the PF74 binding pocket is likely to be strongly limited by functional constraints on HIV-1 evolution., Importance: The HIV-1 capsid plays critical roles in early steps of infection and is an attractive target for therapy. Here we show that selection for resistance to a capsid-targeting small molecule inhibitor can result in viral dependence on the compound. The mutant virus was debilitated in primary T cells and macrophages--cellular targets of infection in vivo. The mutations also altered the virus's dependence on cellular factors that are normally required for HIV-1 entry into the nucleus. This work provides new information regarding mechanisms of HIV-1 resistance that should be useful in efforts to develop clinically useful drugs targeting the HIV-1 capsid., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Background: Preexposure prophylaxis (PrEP) with emtricitabine plus tenofovir disoproxil fumarate (FTC/TDF) or TDF alone reduces the risk of human immunodeficiency virus (HIV) acquisition. Understanding the risk of antiretroviral resistance selected by PrEP during breakthrough infections is important because of the risk of treatment failure during subsequent antiretroviral use., Methods: Within the largest randomized trial of FTC/TDF versus TDF as PrEP, plasma samples were tested for HIV with resistance mutations associated with FTC (K65R and M184IV) and TDF (K65R and K70E), using 454 sequencing., Results: Of 121 HIV seroconverters, 25 received FTC/TDF, 38 received TDF, and 58 received placebo. Plasma drug levels in 26 individuals indicated PrEP use during or after HIV acquisition, of which 5 had virus with resistance mutations associated with their PrEP regimen. Among those with PrEP drug detected during infection, resistance was more frequent in the FTC/TDF arm (4 of 7 [57%]), compared with the TDF arm (1 of 19 [5.3%]; P = .01), owing to the FTC-associated mutation M184IV. Of these cases, 3 had unrecognized acute infection at PrEP randomization, and 2 were HIV negative at enrollment., Conclusions: These results suggest that resistance selected by PrEP is rare but can occur both with PrEP initiation during acute seronegative HIV infection and in PrEP breakthrough infections and that FTC is associated with a greater frequency of resistance mutations than TDF., (© The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Background & Aims: We evaluated the antiviral response of Asian or Pacific Islander (API) patients with chronic hepatitis B (CHB) who had baseline high viral load (HVL), defined as pre-treatment hepatitis B virus (HBV) DNA ≥9 log10 copies/ml, following up to 288 weeks of tenofovir disoproxil fumarate (TDF) treatment., Methods: A total of 205 HBeAg-negative and HBeAg-positive self-described API patients received 48 weeks of TDF 300 mg (HVL n = 18) or adefovir dipivoxil 10 mg (HVL n = 15) in a blinded fashion, followed by open-label TDF for an additional 240 weeks. The proportions of HVL vs. non-HVL patients with HBV DNA <400 copies/ml were compared. Mean declines in HBV DNA were evaluated in API vs. non-API patients., Results: Throughout the first 72 weeks of treatment, a smaller proportion of HVL API patients reached HBV DNA <400 copies/ml than non-HVL API patients. However, after this timepoint similar proportions of HVL and non-HVL API patients achieved HBV DNA <400 copies/ml (100% vs. 97%, respectively), which was maintained through week 288, where 92% of HVL patients and 99% of non-HVL API patients on treatment had HBV DNA <400 copies/ml. During the 288 weeks of treatment, API patients had similar mean HBV DNA declines as non-API patients, regardless of whether patients were HVL or non-HVL. No API HVL patient had persistent viremia at week 288. No resistance was detected among HVL or non-HVL patients., Conclusions: API patients with HVL CHB achieve HBV DNA <400 copies/ml with long-term TDF treatment; however, achieving viral suppression may take longer for HVL patients relative to non-HVL API patients., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

After the World Health Organization officially declared the end of the first pandemic of the XXI century in August 2010, the influenza A(H1N1)pdm09 virus has been disseminated in the human population. In spite of its sustained circulation, very little on phylogenetic data or oseltamivir (OST) resistance is available for the virus in equatorial regions of South America. In order to shed more light on this topic, we analysed the haemagglutinin (HA) and neuraminidase (NA) genes of influenza A(H1N1)pdm09 positive samples collected during the pandemic period in the Pernambuco (PE), a northeastern Brazilian state. Complete HA sequences were compared and amino acid changes were related to clinical outcome. In addition, the H275Y substitution in NA, associated with OST resistance, was investigated by pyrosequencing. Samples from PE were grouped in phylogenetic clades 6 and 7, being clustered together with sequences from South and Southeast Brazil. The D222N/G HA gene mutation, associated with severity, was found in one deceased patient that was pregnant. Additionally, the HA mutation K308E, which appeared in Brazil in 2010 and was only detected worldwide the following year, was identified in samples from hospitalised cases. The resistance marker H275Y was not identified in samples tested. However, broader studies are needed to establish the real frequency of resistance in this Brazilian region.

In 2012, the complete genomic sequence of a new and potentially harmful influenza A-like virus from bats (H17N10) was identified. However, infectious influenza virus was neither isolated from infected bats nor reconstituted, impeding further characterization of this virus. Here we show the generation of an infectious chimeric virus containing six out of the eight bat virus genes, with the remaining two genes encoding the haemagglutinin and neuraminidase proteins of a prototypic influenza A virus. This engineered virus replicates well in a broad range of mammalian cell cultures, human primary airway epithelial cells and mice, but poorly in avian cells and chicken embryos without further adaptation. Importantly, the bat chimeric virus is unable to reassort with other influenza A viruses. Although our data do not exclude the possibility of zoonotic transmission of bat influenza viruses into the human population, they indicate that multiple barriers exist that makes this an unlikely event.

Using hepatitis C virus (HCV) and interferon (IFN) resistance as a proof of concept, we have devised a new method for calculating the effect of a drug on a viral population, as well as the resistance of the population's individual intrahost variants. By means of next-generation sequencing, HCV variants were obtained from sera collected at nine time points from 16 patients during the first 48 h after injection of IFN-α. IFN-resistance coefficients were calculated for individual variants using changes in their relative frequencies, and for the entire intrahost viral population using changes in viral titer. Population-wide resistance and presence of IFN-resistant variants were highly associated with pegylated IFN-α2a/ribavirin treatment outcome at week 12 (P = 3.78 × 10(-5) and 0.0114, respectively). This new method allows an accurate measurement of resistance based solely on changes in viral titer or the relative frequency of intrahost viral variants during a short observation time.

NS5B, a hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp) that plays a key role in viral replication, is an important target in the discovery of antiviral agents. PF-00868554 is a potent non-nucleoside inhibitor (NNI) that binds to the Thumb II allosteric pocket of NS5B polymerase and has shown significant promise in phase II clinical trials. Unfortunately, several PF-00868554 resistant mutants have been identified. M423 variants were the most common NS5B mutations that occurred after PF-00868554 monotherapy. In this study, we used molecular dynamics (MD) simulations, binding free energy calculations and free energy decomposition to explore the drug resistance mechanism of HCV to PF-00868554 resulting from three representative mutations (M423T/V/I) in NS5B polymerase. Free energy decomposition analysis reveals that the loss of binding affinity mainly comes from the reduction of both van der Waals (ΔE(vdw)) and electrostatic interaction contributions in the gas phase (ΔE(ele)). Further structural analysis indicates that the location of PF-00868554 and the binding mode changed due to mutation of the residue at the 423 site of NS5B polymerase from methionine to threonine, isoleucine or valine, which further resulted in the loss of binding ability of PF-00868554 to NS5B polymerase. The obtained computational results will have important value for the rational design of novel non-nucleoside inhibitors targeting HCV NS5B polymerase.

Cytomegalovirus (CMV) is an important pathogen often encountered after solid organ transplantation and is associated with increased morbidity and mortality. Resistance of CMV to antiviral agents is becoming more common but with few treatment strategies. Two specific mutations in the CMV genome--the UL97 and UL54 genes--correlate with antiviral drug resistance. We describe a 49-year-old, CMV-seronegative woman who received a CMV-seropositive donor kidney transplant and appropriate CMV prophylaxis. Approximately 1 month after transplantation, the patient developed CMV viremia that responded to valganciclovir. She was later diagnosed with recurrent CMV infection, CMV resistance, and both the UL97 and UL54 gene mutations. The patient responded to foscarnet and significant reduction of immunosuppression; she was negative for CMV viremia for the next 12 months. This case illustrates the importance of having heightened awareness for the possibility of panresistant CMV early and decreasing immunosuppression as the cornerstone of treatment., (© 2013 American College of Clinical Pharmacy.)

The activity of the phosphite of acycloguanosine (P-ACG) and six antivirals was tested individually and in double and triple combinations on two strains of the herpes simplex virus (HSV) type 1 (sensitive to acyclovir and resistant to acyclovir) using the CPE inhibition method in the Vero E6 cell microcultures. These are: phosphite of acycloguanosine (P-ACG), Ara-A, cidofovir (CDV), ribavirin (Rib), phosphonoformate (PFA), glycyrrhizic acid (GLN) and alpha-interferon (alpha-IFN). All studied double combinations and triple combinations including P-ACV inhibited replication of both HSV strains more effectively than any drug alone. Various types of interactions depending on the virus type were observed in both viral models: synergistic (double combinations P-ACG with PFA, CDV, Rib, alpha-IFN and triple combinations P-ACG with alpha-IFN +PFA, alpha-IFN +AraA, alpha-IFN +CDV, PFA+CDV, PFA+Rib, CDV+AraA, CDV+Rib, CDV+GLN,PFA+AraA) and additive (P-ACG with AraA and PFA+GLN). Neither antagonism nor interference was noted for any combinations. Adduced results suggest that these combinations might be clinically useful for the treatment of certain herpes simplex virus type 1 infections.

The emergence of influenza drug resistance is a major public health concern. The molecular basis of resistance to oseltamivir (Tamiflu) is investigated using a computational assay involving multiple 500 ns unrestrained molecular dynamics (MD) simulations of oseltamivir complexed with mutants of H1N1-2009 influenza neuraminidase. The simulations, accelerated using graphics processors (GPUs), and using a fully explicit model of water, are of sufficient length to observe multiple drug unbinding and rebinding events. Drug unbinding occurs during simulations of known oseltamivir-resistant mutants of neuraminidase. Molecular-level rationalizations of drug resistance are revealed by analysis of these unbinding trajectories, with particular emphasis on the dynamics of the mutant residues. The results indicate that MD simulations can predict weakening of binding associated with drug resistance. In addition, visualization and analysis of binding site water molecules reveal their importance in stabilizing the binding mode of the drug. Drug unbinding is accompanied by conformational changes, driven by the mutant residues, which results in flooding of a key pocket containing tightly bound water molecules. This displaces oseltamivir, allowing the tightly bound water molecules to be released into bulk. In addition to the role of water, analysis of the trajectories reveals novel behavior of the structurally important 150-loop. Motion of the loop, which can move between an open and closed conformation, is intimately associated with drug unbinding and rebinding. Opening of the loop occurs coincidentally with drug unbinding, and interactions between oseltamivir and the loop seem to aid in the repositioning of the drug back into an approximation of its original binding mode on rebinding. The similarity of oseltamivir to a transition state analogue for neuraminidase suggests that the dynamics of the loop could play an important functional role in the enzyme, with loop closing aiding in binding of the substrate and loop opening aiding the release of the product.

Background: Non-adherence is one of the strongest predictors of therapeutic failure in HIV-positive patients. Virologic failure with subsequent emergence of resistance reduces future treatment options and long-term clinical success., Methods: Prospective observational cohort study including patients starting new class of antiretroviral therapy (ART) between 2003 and 2010. Participants were naïve to ART class and completed ≥1 adherence questionnaire prior to resistance testing. Outcomes were development of any IAS-USA, class-specific, or M184V mutations. Associations between adherence and resistance were estimated using logistic regression models stratified by ART class., Results: Of 314 included individuals, 162 started NNRTI and 152 a PI/r regimen. Adherence was similar between groups with 85% reporting adherence ≥95%. Number of new mutations increased with increasing non-adherence. In NNRTI group, multivariable models indicated a significant linear association in odds of developing IAS-USA (odds ratio (OR) 1.66, 95% confidence interval (CI): 1.04-2.67) or class-specific (OR 1.65, 95% CI: 1.00-2.70) mutations. Levels of drug resistance were considerably lower in PI/r group and adherence was only significantly associated with M184V mutations (OR 8.38, 95% CI: 1.26-55.70). Adherence was significantly associated with HIV RNA in PI/r but not NNRTI regimens., Conclusion: Therapies containing PI/r appear more forgiving to incomplete adherence compared with NNRTI regimens, which allow higher levels of resistance, even with adherence above 95%. However, in failing PI/r regimens good adherence may prevent accumulation of further resistance mutations and therefore help to preserve future drug options. In contrast, adherence levels have little impact on NNRTI treatments once the first mutations have emerged.

Hepatitis C virus (HCV) is one of the foremost causes of chronic liver disease affecting over 300 million globally. HCV contains a positive-stranded RNA of ~9600 nt and is surrounded by the 5' and 3'untranslated regions (UTR). The only successful treatment regimen includes interferon (IFN) and ribavirin. Like many other viruses, HCV has also evolved various mechanisms to circumvent the IFN response by blocking (1) downstream signaling actions via STAT1, STAT2, IRF9 and JAK-STAT pathways and (2) repertoire of IFN Stimulatory Genes (ISGs). Several studies have identified complex host demographic and genetic factors as well as viral genetic heterogeneity associated with outcomes of IFN therapy. The genetic predispositions of over 2000 ISGS may render the patients to become resistant, thus identification of such parameters within a subset of population are necessary for management corollary. The ability of various HCV genotypes to diminish IFN antiviral responses plays critical role in the establishment of chronic infection at the acute stage of infection, thus highlighting importance of the resistance in HCV treated groups. The recently defined role of viral protein such as C, E2, NS3/NS4 and NS5A proteins in inducing the IFN resistance are discussed in this article. How the viral and host genetic composition and epistatic connectivity among polymorphic genomic sites synchronizes the evolutionary IFN resistance trend remains under investigation. However, these signals may have the potential to be employed for accurate prediction of therapeutic outcomes. In this review article, we accentuate the significance of host and viral components in IFN resistance with the aim to determine the successful outcome in patients., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Background: The emergence and transmission of HIV-1 drug resistance (HIVDR) has raised concerns after rapid global antiretroviral therapy (ART) scale-up. There are limited data on the epidemiology of primary HIVDR in resource-limited settings in Asia. We aimed to determine the prevalence and compare the distribution of HIVDR in a cohort of ART-naïve Asian patients with recent and chronic HIV-1 infection., Methods: Multicenter prospective study was conducted in ART-naïve patients between 2007 and 2010. Resistance-associated mutations (RAMs) were assessed using the World Health Organization 2009 list for surveillance of primary HIVDR., Results: A total of 458 patients with recent and 1,340 patients with chronic HIV-1 infection were included in the analysis. The overall prevalence of primary HIVDR was 4.6%. Recently infected patients had a higher prevalence of primary HIVDR (6.1% vs. 4.0%, p = 0.065) and frequencies of RAMs to protease inhibitors (PIs; 3.9% vs. 1.0%, p<0.001). Among those with recent infection, the most common RAMs to nucleoside reverse transcriptase inhibitors (NRTIs) were M184I/V and T215D/E/F/I/S/Y (1.1%), to non-NRTIs was Y181C (1.3%), and to PIs was M46I (1.5%). Of patients with chronic infection, T215D/E/F/I/S/Y (0.8%; NRTI), Y181C (0.5%; non-NRTI), and M46I (0.4%; PI) were the most common RAMs. K70R (p = 0.016) and M46I (p = 0.026) were found more frequently among recently infected patients. In multivariate logistic regression analysis in patients with chronic infection, heterosexual contact as a risk factor for HIV-1 infection was less likely to be associated with primary HIVDR compared to other risk categories (odds ratio 0.34, 95% confidence interval 0.20-0.59, p<0.001)., Conclusions: The prevalence of primary HIVDR was higher among patients with recent than chronic HIV-1 infection in our cohort, but of borderline statistical significance. Chronically infected patients with non-heterosexual risks for HIV were more likely to have primary HIVDR.

Objectives: To explore the durability of three first-line tenofovir/emtricitabine-based regimens in combination with atazanavir/ritonavir, efavirenz or lopinavir/ritonavir in HIV-1-infected patients., Patients and Methods: A retrospective, longitudinal, multicentre analysis of adult patients enrolled in the Antiretroviral Resistance Cohort Analysis (ARCA), a national prospective observational cohort of HIV-1-infected patients followed up at more than 100 clinical and laboratory units in Italy. Patients eligible were those starting first-line antiretroviral therapy between 1 June 2004 and 15 April 2011 and who were followed up for at least 6 months. The primary endpoint was durability, defined as the time from antiretroviral therapy initiation to first treatment modification. Time-dependent events were analysed by the Kaplan-Meier approach and the Cox proportional hazard model., Results: There are 26,000 HIV-infected patients in the ARCA database, of whom 1654 met study inclusion criteria. Six hundred and thirty-nine (38.6%) received efavirenz, 321 (19.4%) received atazanavir/ritonavir and 694 (41.9%) received lopinavir/ritonavir as a first-line regimen. Over a total observation period of 88 months, equivalent to more than 2805 person-years of follow-up, 618 patients underwent treatment modification. Lopinavir/ritonavir, given twice daily, was associated with a higher discontinuation rate than efavirenz- and atazanavir-based regimens [hazard ratio (HR) 1.83, 95% confidence interval (CI) 1.56-2.15, P = 0.001]. Comparing the once-daily regimens, the rate of discontinuation of efavirenz was higher than that of atazanavir/ritonavir (HR 1.39, 95% CI 1.06-1.83, P = 0.016)., Conclusions: Significant differences in treatment duration were observed among the three studied regimens. Once-daily regimens exhibited greater durability than the twice-daily regimen. Among the specific regimens examined, tenofovir/emtricitabine plus atazanavir/ritonavir showed the greatest durability.

New treatment options for HCV infection with Direct-Acting Antivirals (DAAs) increased SVR rate in treated patients but on the other hand drew attention to the problem of HCV drug resistance. Drug-resistant HCV mutants are present in all infected patients even before the treatment initiation and their number grows significantly over the first few days of DAAs therapy. But HCV has no known genetic form of intra-cellular persistence (does not integrate with host's genome and cannot produce episomal forms) which enables its total eradication. It is likely that the effective interferon-free, based on all-oral DAAs drug combination will be available within the next few years. This paper reviews HCV resistance mechanisms and their significance in treatment. I also presents results of recent DAAs trials.

Background: ITMN-191 (RG7227, Danoprevir), as a potential inhibitor of the NS3/4A protease of hepatitis C virus, has been in phase 2 clinical trial. Unfortunately, several ITMN-191 resistance mutants including R155K, A156V, and D168A/E have been identified., Methods: Molecular dynamics simulation, binding free energy calculation and per-residue energy decomposition were employed to explore the binding and resistance mechanism of hepatitis C virus NS3/4A protease to ITMN-191., Results: Based on molecular dynamics simulation and per-residue energy decomposition, the nonpolar energy term was found to be the driving force for ITMN-191 binding. For the studied R155K, A156V, D168A/E mutants, the origin of resistance is mainly from the conformational changes of the S4 and extended S2 binding pocket induced by the studied mutants and further leading to the reduced binding ability to the extended P2 and P4 moieties of ITMN-191., Conclusions: Further structural analysis indicates that the destruction of conservative salt bridges between residues 168 and 155 should be responsible for the large conformation changes of the binding pocket in R155K and D168A/E mutants. For A156V mutation, the occurrence of drug resistance is mainly from the changed binding pocket by a replacement of one bulky residue Val., General Significance: The obtained drug resistance mechanism of this study will provide useful guidance for the development of new and effective HCV NS3/4A inhibitors with low resistance., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The recent development of small molecule compounds that directly inhibit the viral life cycle represents a major milestone for the treatment of chronic hepatitis C virus (HCV) infection. These new drugs that are collectively termed direct-acting antivirals (DAA) include a range of inhibitors of the non-structural (NS) 3/4A protease, NS5B polymerase and NS5A protein. Two NS3/4A protease inhibitors (boceprevir and telaprevir) in combination with pegylated interferon and ribavirin have now been approved for the treatment of chronic HCV genotype 1 infection and cure rates could be increased by 20-30%. However, the majority of DAAs is still in early clinical development. The rapid replication rate of HCV, along with the error-prone polymerase activity leads to a high genetic diversity among HCV virions that includes mutants with reduced susceptibility to DAA-therapy. These resistance-associated variants often occur at very low frequencies. However, during DAA-based treatment, rapid selection of resistance mutations may occur, eventually leading to viral break-through. A number of variants with different levels of resistance have been described in vitro and in vivo for virtually all DAAs. We review the parameters that determine DAA resistance as well as the clinical implications of resistance testing. In addition, the most recent literature and conference data on resistance profiles of DAAs in clinical development and future strategies to avoid the emergence of viral resistance are also discussed., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Background & Aims: Hepatitis C virus (HCV) is a common cause of chronic liver disease. Many patients do not clear the viral infection; little is known about the mechanisms of HCV persistence or the frequent failure of interferon (IFN) to eliminate it. Better culture systems are needed to study viral replication in quiescent liver cells., Methods: We used human hepatoma (Huh7.5) cells and those that had undergone proliferation arrest and differentiation (Huh7.5(dif)) to study the persistence of HCV infection following exposure of the cells to IFN-α and to compare the antiviral effects of IFN-α and IFN-λ. We validated these results with primary human hepatocytes and Huh7 cells that expressed an IFN-inducible fluorophore., Results: Following infection of Huh7.5(dif) cells, HCV replicated persistently and released infectious particles. Long-term exposure of the cells to IFN-α reduced HCV replication ∼1000-fold but did not eliminate the virus; viral replication rebounded after withdrawal of IFN, as it does in patients with chronic HCV infection. HCV replicated at higher levels, but not exclusively, in cells that had a low level of response to IFN-α. Following incubation of cells with equipotent concentrations of IFN-α or IFN-λ, Huh7.5(dif) cells expressed a wider pattern of IFN-stimulated genes than undifferentiated Huh7.5 cells or primary human hepatocytes, indicating that the antiviral response depends on the differentiation status of the cells., Conclusions: We developed a cell culture system using hepatoma cells to study persistent HCV infection during the type I or type III IFN-induced antiviral response. The level and range of the antiviral responses were associated with the differentiation status of the cells. We propose that HCV exploits the stochastic nature of the response of hepatocytes to IFN to sustain persistence., (Copyright © 2012 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Context: Cecropia glaziovii Snethl. (Cecropiaceae), commonly known as "embaúba-vermelha", is widely distributed throughout Latin America and has been reported in Brazilian folk medicine to treat cough, asthma, high blood pressure and inflammation., Objective: Investigate the hepatoprotective properties of crude hydroethanolic extract of C. glaziovii as well as its in vitro antioxidant and antiviral (HSV-1 acyclovir resistant strain) activities., Materials and Methods: The hepatoprotective effect, the antioxidant properties and antiviral activity of crude hydroethanol extract (RCE40) from C. glaziovii leaves were evaluated by carbon-tetrachloride (CCl(4))-induced hepatotoxicity, by TBARS (thiobarbituric acid reactive species) and MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assays, respectively., Results: The RCE40 extract (20 mg/kg) inhibited lipid peroxidation on liver in post injury treatment and decreased serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST). In addition, in this protocol the RCE40 (20 mg/kg) enhanced the activity of hepatic enzymes (SOD/CAT) which are involved in combating reactive oxygen species (ROS), suggesting that it possesses the capacity to attenuate the CCl(4)-induced liver damage. Moreover the RCE40 (20 mg/kg) inhibited TBARS formation induced by several different inductors of oxidative stress showing significant antioxidant activity, including physiologically relevant concentration, as low as 2 µg/mL. Concerning antiviral activity, the RCE40 was effective against herpes simplex virus type 1 replication (29R acyclovir resistant strain) with EC(50) = 40 µg/mL and selective index (SI) = 50., Discussion and Conclusion: These results indicate that C. glaziovii could be a good source of antioxidant and anti-HSV-1 lead compounds.

Background & Aims: Hepatitis C virus (HCV) interacts with apolipoproteins B (apoB) and E (apoE) to form infectious lipoviral particles (LVP). Response to peginterferon is influenced by interferon-stimulated genes (ISGs) and IL28B genotype. LDL cholesterol (LDL-C) also predicts interferon response, therefore we hypothesised that LVP may also be associated with interferon sensitivity., Methods: LVP (HCV RNA density ≤1.07 g/ml) and 'non-LVP' (d >1.07 g/ml) were measured in 72 fasted HCV-G1 patients by iodixanol density gradient ultracentrifugation and the LVP ratio (LVP/LVP+non-LVP) was calculated. Fasting lipid profiles and apolipoproteins B and E were measured. Interferon-gamma-inducible protein 10 kDa (IP10), a marker of ISGs, was measured by ELISA., Results: Complete early virological response (EVR) was associated with lower apoE (23.9±7.7 vs. 36.1±15.3 mg/L, p=0.013), higher LDL-C (p=0.039) and lower LVP ratios (p=0.022) compared to null responders. In multivariate linear regression analysis, apoE was independently associated with LVP (R(2) 19.5%, p=0.003) and LVP ratio (p=0.042), and negatively with LDL-C (p<0.001). IP10 was significantly associated with ApoB (p=0.001) and liver stiffness (p=0.032). IL28B rs12979860 CC was associated with complete EVR (p=0.044), low apoE (CC 28±11 vs. CT/TT 35±13 mg/L, p=0.048) and higher non-LVP (p=0.008). Logistic regression analysis indicated that patients with high LVP ratios were less likely to have EVR (odds ratio 0.01, p=0.018)., Conclusions: In HCV-G1, interferon sensitivity is characterised by low LVP ratios and low apoE levels in addition to higher LDL-C and IL28B rs12979860 CC. Null-response is associated with increased LVP ratio. The association of apoE and LVP with peginterferon treatment response suggests that lipid modulation is a potential target to modify interferon sensitivity., (Copyright © 2012 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)