Antiviral Therapy

In haematopoietic stem cell transplant (HSCT) recipients, cytomegalovirus (CMV) infection contributes significantly to morbidity and mortality in both the early and late post-transplant period. Ganciclovir (GCV) is the treatment of choice for CMV, but requires intravenous administration, a fact that complicates its long-term use. Oral valganciclovir (VGCV) and intravenous GCV were recently shown to have similar efficacy for pre-emptive CMV treatment in solid organ transplant recipients, but relatively limited data are available in HSCT recipients. The objectives of this study were to compare the efficacy of VGCV versus intravenous GCV or foscarnet (FOS) for pre-emptive therapy of active CMV infection in allogeneic HSCT and to determine the incidence of adverse effects and relapses.

This was a 2-year prospective, comparative cohort study in which 237 episodes of pre-emptive therapy for active CMV infection were collected in 166 allogeneic HSCT recipients out of 717 included in the Spanish Network for Research on Infection in Transplantation (RESITRA/REIPI) database. Intravenous GCV was the first-line treatment in 112 episodes, intravenous FOS in 38 episodes, and oral VGCV in 87 episodes.

VGCV was used as pre-emptive therapy for active CMV infection in 87 episodes. Excluding episodes considered as relapse, VGCV was as successful (91.4% [74/81]) as GCV (83.7% [87/14]) or FOS (75.8% [25/33]). In the VGCV arm, 7 (8.6%) cases were considered treatment failures: 4 (4.9%) because of adverse events, 1 (1.2%) due to persistent viral activity and 2 (2.5%) based on clinical decision. There were also 6 (7.4%) cases of recurrent infection. No statistically significant differences were found when compared to GCV or FOS.

In allogeneic HSCT recipients, VGCV seemed effective and safe in the pre-emptive therapy of active CMV infection.

Recent studies have indicated that interferon (IFN) or pegylated interferon (PEG-IFN) plus ribavirin therapy can achieve sustained virological response (SVR) against HCV equally in dual HBV–HCV infection and in HCV monoinfection. Whether these therapies can reduce hepatocellular carcinoma (HCC) development in dual HBV–HCV infection remains unclear.

A total of 135 dually-infected patients with active hepatitis C receiving IFN or PEG-IFN plus ribavirin therapy were enrolled. The cumulative incidence of HCC was compared to that in 1,470 HCV-monoinfected patients.

Based on the Cox proportional hazards model, dual infection was an independent factor for HCC development in all 1,605 chronic hepatitis C patients with or without positive hepatitis B surface antigen receiving IFN or PEG-IFN plus ribavirin (hazard ratio (HR)=1.864, 95% CI 1.052–3.303; P=0.033). In dually-infected patients, HCC developed in 4 of 96 with HCV SVR and 11 of 39 without HCV SVR (P<0.001) after a median follow-up of 4.6 years. Age (HR=1.175, 95% CI 1.070–1.291; P=0.001) and non-HCV-SVR (HR=7.874, 95% CI 2.375–26.32; P=0.001) were independent factors for HCC development. Subgroup analysis showed that HCC occurrence was lower in patients with HCV SVR and HBV DNA levels <2,000 IU/ml at the end of treatment or follow-up compared to those with HCV SVR and HBV DNA levels ≥2,000 IU/ml (P=0.034) and those without HCV SVR (P<0.001).

Sustained HCV clearance by IFN or PEG-IFN plus ribavirin therapy may significantly reduce HCC in HBV–HCV dually-infected patients, whereas persistence or reactivation of HBV decreases the benefit of HCV SVR.

There are no convincing data supporting the routine use of pre-emptive therapy against HBV reactivation in various loco-regional therapies for hepatocellular carcinoma (HCC). This study investigated the incidence, severity and risk factors of HBV reactivation during loco-regional therapies.

A total of 205 prospectively enrolled patients were classified in order of increasing intensity of loco-regional therapies: local ablation therapy (LAT; 43 patients), transarterial chemotherapy using adriamycin (TAC-ADR; 93 patients) or combined epirubicin-cisplatin (TAC-EC; 26 patients), and combined chemo-radiotherapy (TAC-EC+RT; 43 patients).

During the follow-up, 62 (30.2%) patients developed HBV reactivation. Multivariate analysis identified HBV DNA levels >10⁴ copies/ml (P=0.041) and treatment option (P=0.001) to be independent predictors of HBV reactivation. There was a significant trend for increasing risk of reactivation with increasing intensity of therapy, with hazard ratios of 1.0 for LAT, 2.45 for TAC-ADR, 4.19 for TAC-EC and 10.17 for TAC-EC+RT. The severity of reactivated disease was also increased with increasing treatment intensity (P-value for trend <0.05). Only one of the patients with low-level viraemia receiving LAT alone developed reactivation, whereas a substantial number of patients with high-level viraemia eventually developed reactivation.

High-level viraemia and high-level treatment intensity are the major risk factors for HBV reactivation during loco-regional therapy. Trends are evident for the increased risk and severity of reactivation with the aggressiveness of treatment. Pre-emptive antiviral therapy should be recommended for all patients with high-level viraemia irrespective of treatment option, or those undergoing any intensive therapy.

Despite a rising incidence of acute HCV in patients infected with HIV, the optimal therapeutic strategy (pegylated interferon-α [PEG-IFN-α] monotherapy or in combination with ribavirin) is still under debate.

A total of 23 HIV-infected patients were prospectively diagnosed with acute HCV and treated with PEG-IFN-α2a monotherapy (180 μg/week) for 24 or 48 weeks. Add-on ribavirin was allowed from week 4 of therapy onwards. There were three patients who were not included for different reasons. Blood samples were routinely drawn for viral load measurement and IL28B polymorphism analysis.

Spontaneous viral clearance occurred in 1 (4%) patient. Nineteen patients (13 genotype 1 and 6 genotype 4) received treatment with PEG-IFN-α monotherapy (3 with add-on ribavirin) resulting in a rapid virological response (HCV RNA<50 IU/ml at week 4) in 7 (37%) patients. A sustained virological response (SVR) was reached in 7 (37%) patients, whereas 9 (47%) patients were null-responders to treatment (that is, <2 log₁₀ drop in HCV RNA at week 12 of therapy). The unfavourable G allele of the IL28B polymorphism rs8099917 was detected in 66% of the non-responders. In case of re-emergence of HCV viraemia after treatment discontinuation, sequence analysis of quasispecies confirmed an HCV relapse in 3 patients while 2 patients were re-infected by their previously non-responding partner.

PEG-IFN-α monotherapy resulted in a low SVR rate and a high percentage of null-response, whereas non-SVR was associated with a polymorphism in the IL28B gene (rs8099917).

The thymidine kinase (TK) and DNA polymerase (pol) genes of herpes simplex virus type-1 (HSV-1) and type-2 (HSV-2) are important targets for genotypic determination of HSV resistance to antiviral drugs. The knowledge of gene polymorphism is an absolute requirement for the correct interpretation of genotypic findings.

In this study, the natural polymorphism of TK and DNA pol genes was examined by DNA sequencing in 56 HSV-1 and 12 HSV-2 strains sensitive to acyclovir.

In 56 HSV-1 strains, 26 different non- synonymous polymorphism-associated mutations were detected in the TK gene. To our knowledge, 8 of them have never been reported in the literature. In the TK gene of 12 HSV-2 strains, 6 polymorphism-related non-synonymous mutations were observed, whereas there was 1 novel mutation in 1 strain. The DNA pol gene of 53 HSV-1 isolates contained 47 distinct polymorphism-associated amino acid substitutions and 11 substitutions were found in the DNA pol gene of 12 HSV-2 strains. Altogether, 31 novel substitutions could be identified in the DNA pol gene of HSV-1 and 3 in HSV-2 strains. In these strains, any resistance to foscarnet was excluded.

The 43 novel non-synonymous mutations enrich the knowledge about the natural genetic polymorphism of TK and DNA pol in clinical HSV strains. The findings have to be considered for genotypic analysis of HSV in case of clinical resistance.

The National Research Council has recommended development of polio antiviral drugs to assist in management of outbreaks and to mitigate adverse consequences of vaccination. V-073 is a small molecule poliovirus capsid inhibitor that is being developed for these purposes. Antiviral use raises the potential of treatment-emergent resistance. Understanding virological consequences of resistance is important.

Six independent laboratory-derived V-073-resistant poliovirus variants were characterized for their ability to be neutralized by conventional vaccine-induced immune sera, to elicit serum neutralizing antibodies upon CD-1 mouse immunization, and to replicate in and to cause paralysis of TgPVR21 mice.

V-073-resistant variants were effectively neutralized by oral poliovirus vaccine and inactivated poliovirus vaccine human immune sera. All variants elicited virus neutralizing antibody titres in CD-1 mice that were comparable to drug-susceptible parental and Sabin vaccine strain viruses. Infection efficiency of TgPVR21 mice by variants was comparable to (1 of 6 variants) or considerably lower than (5 of 6 variants) parental viruses. Drug-resistant variants replicated to levels comparable to (1 of 6 variants) or substantially less than (5 of 6 variants) their drug-susceptible parental viruses and were on average 1.4 log₁₀ (range 0.3 to >2.8 log₁₀) less neurovirulent.

Laboratory-derived V-073-resistant variants exhibit clear attenuation of pathogenic properties while maintaining immunological features of drug-susceptible viruses.

Data on the use of oseltamivir in infants is limited. We documented the clinical presentations of infants aged <12 months hospitalized with laboratory-confirmed influenza A in the southern hemisphere winter of 2009 and compared outcomes in relation to oseltamivir therapy.

Data were extracted from prospectively collected and collated influenza case reports (June– September 2009) ascertained through Paediatric Active Enhanced Disease Surveillance, an in-patient surveillance system operating at the Children's Hospital at Westmead (CHW), Sydney, NSW, Australia. Nosocomial cases were excluded.

Of 56 infants with definite influenza (35 pandemic H1N1 2009) admitted to the CHW, 20 were treated with oseltamivir. Overall, 12 (60%) of those treated with oseltamivir were aged <6 months. Cough, fever and coryza were the most common clinical features (≥70%). Vomiting was present on admission in 31.4%. All 7 cases presenting with vomiting then given antiviral treatment had reduction of vomiting and the other 13 did not develop vomiting on treatment. There were three infants with hypoxaemia (oxygen saturation ≤93%) on presentation in the treatment group compared with none in the control group (P=0.04). The median duration of hospital stay was the same in both groups (2.5 days).

The use of oseltamivir was well tolerated in hospitalized infants. Vomiting, one of the widely reported side effects of oseltamivir, was found to be more a presenting symptom of influenza than a side effect of oseltamivir. Based on pulse oximetry, oseltamivir-treated cases may have been more severe on presentation, but there was no significant difference in length of hospital stay.

The aim of this study was to investigate long-term renal function in HIV-infected adults initiating antiretroviral therapy (ART) with a CD4⁺ T-cell count <200 cells/mm³ in Africa.

This was an observational analysis within the DART trial randomizing 3,316 adults to routine laboratory and clinical monitoring (LCM) or clinically driven monitoring (CDM). Serum creatinine was measured pre-ART (all ≤360 μmol/l), at weeks 4 and 12, then every 12 weeks for 4-5 years; estimated glomerular filtration rate (eGFR) was determined using the Cockcroft-Gault formula. We analysed eGFR changes, and cumulative incidences of eGFR<30 ml/min/1.73 m² and chronic kidney disease (CKD; <60 ml/min/1.73 m² or 25% decrease if <60 ml/min/1.73 m² pre-ART; confirmed >3 months).

At ART initiation, median CD4⁺ T-cell count was 86 cells/mm³; 1,492 (45%) participants had mild (60–<90 ml/min/1.73 m²), 237 (7%) moderate (30–<60 ml/min/1.73 m²) and 7 (0.2%) severe (15–<30 ml/min/1.73 m²) decreases in eGFR. First-line ART was zidovudine/lamivudine plus tenofovir (74%), abacavir (9%) or nevirapine (17%). By 4 years, cumulative incidence of eGFR<30 ml/min/1.73 m² was 2.8% (n=90) and CKD was 5.0% (n=162). Adjusted eGFR increases to 4 years were 1, 9 and 6 ml/min/1.73 m² with tenofovir, abacavir and nevirapine, respectively (P<0.001), and 4 and 2 ml/min/1.73 m² for LCM and CDM, respectively (P=0.005; 2 and 3 ml/min/1.73 m² to 5 years; P=0.81).

On all regimens and monitoring strategies, severe eGFR impairment was infrequent; differences in eGFR changes were small, suggesting that first-line ART, including tenofovir, can be given safely without routine renal function monitoring.

Antiviral activity of TMC435, an oral, once-daily, HCV NS3/4A protease inhibitor, was evaluated with pegylated interferon-α2a/ribavirin (P/R) in HCV genotype-1 patients.

Optimal Protease inhibitor Enhancement of Response to TherApy (OPERA-1; TMC435-C201; NCT00561353) is a Phase IIa, randomized, placebo-controlled study. Treatment-naive patients (n=74) received 25, 75 or 200 mg TMC435 once daily, or placebo for 7 days followed by 21 days of triple therapy with P/R, or triple therapy for 28 days. Treatment-experienced patients (n=37; 56.8% with cirrhosis) received 75, 150 or 200 mg TMC435 once daily, or placebo with P/R for 28 days. Patients continued P/R up to week 48.

Treatment-naive patients who received initial monotherapy had a rapid decline in HCV RNA by day 3. At day 7, HCV RNA reductions were greatest for the 75 and 200 mg doses (0.02, -2.63, -3.43 and -4.13 log₁₀ IU/ml for placebo, and TMC435 25, 75 and 200 mg, respectively). At day 28, all patients who received triple therapy with TMC435 75 or 200 mg had HCV RNA<25 IU/ml versus 4/9 for placebo. In total, 18/28 treatment- experienced patients (9/9 prior relapsers, 9/19 non-responders) who received TMC435 had HCV RNA<25 IU/ml at day 28 versus 0/9 for placebo; similar results were observed for the 150 and 200 mg doses. Most adverse events were grade 1/2. No relevant changes in laboratory parameters occurred, except mild and reversible bilirubin elevations, mostly at the 200 mg dose.

Once-daily TMC435 with P/R showed potent, dose-dependent antiviral activity over 28 days, and had a favourable tolerability profile.

The interaction between HIV-1 gp120 and CCR5 N terminus is critical for R5-virus entry and affects CCR5 antagonists’ activity. Knowledge of how different genetic signatures of gp120 V3 domain effect the strength of this interaction is limited.

HIV-1 coreceptor usage was assessed in 251 patients using enhanced-sensitivity Trofile assay and V3 sequencing plus tropism prediction by Geno2pheno algorithm. Bayesian partitional model and recursive model selection have been used to define V3 genetic determinants correlated with different coreceptor usage. Gp120 interaction with CCR5 N terminus was evaluated by docking-analysis/molecular–dynamic simulations starting from the model described previously.

Selected V3 genetic determinants (beyond known aminoacidic positions) significantly correlate with CCR5-or CXCR4-usage, and modulate gp120 affinity for CCR5 N terminus. This is the case for N5Y and N7K, absent in CCR5-using viruses and present in 4.5% and 6% of CXCR4-using viruses, respectively, and A19V, occurring in 2.6% of CCR5-using viruses and 22.0% of CXCR4-using viruses (P=10^-2 to 10^-7). Their presence determines a decreased affinity for CCR5 N terminus even stronger than that observed in the presence of the well-known mutation S11R (N5Y: -6.60 Kcal/mol; N7K: -5.40 Kcal/mol; A19V: -5.60 Kcal/mol; S11R: -6.70 Kcal/mol; WT: -6.90 Kcal/mol). N7K significantly increases the distance between V3 position 7 and sulphotyrosine at CCR5 position 14 (crucial for binding to gp120; from 4.22 Å to 8.30 Å), thus abrogating the interaction between these two important residues.

Key determinants for tropism within the V3 sequence, confirmed by structure- and by phenotypictropism, have been identified. This information can be used for a finer tuning of potential efficacy of CCR5-antagonists in clinical practice, and to provide molecular implications for design of new entry inhibitors.

Recently, we identified increased rates of CD4⁺ T-cell apoptosis in HCV-infected HIV-positive patients as a potential mechanism for enhanced mortality in patients with HIV/HCV coinfection. Since this effect might be attributed to changes in receptor-induced apoptosis, we studied expression and function of Fas ligand (FasL) and its death receptor Fas on CD4⁺ T-cells in HIV/HCV coinfection.

In this cross-sectional study, we simultaneously analysed surface expression of Fas and FasL on CD4⁺ T-cells and serum levels of soluble FasL in HCV/HIV-coinfected, HIV-monoinfected and HCV-monoinfected patients. Susceptibility to FasL-induced apoptosis was analysed by incubating isolated peripheral blood mono-nuclear cells with rhFasL followed by measuring CD4⁺ T-cell apoptosis.

HIV and HCV monoinfection were associated with significantly enhanced surface expression of Fas. Highest Fas expression was detected in HIV/HCV-coinfected patients and correlated with low CD4⁺ T-cell counts. By contrast, elevated levels of soluble and cellular FasL were found only in patients with HIV infection, but not in patients with HCV infection. Importantly, enhanced Fas expression in HCV/HIV coinfection rendered CD4⁺ T-cells more susceptible towards FasL-induced apoptosis. While effective HAART normalized expression and secretion of FasL in HIV-infected and HIV/HCV-coinfected patients, expression of Fas decreased only slightly and still remained significantly elevated as compared with healthy controls.

Our findings suggest a synergistic mechanism in HIV/HCV coinfection between up-regulation of Fas expression on CD4⁺ T-cells and HIV-induced elevated levels of cellular and soluble FasL. Together, both effects contribute to enhanced apoptosis of CD4⁺ T-cells in HIV/HCV coinfection.

Sex differences in the natural history of HIV infection may vary between resource-rich and resource-limited settings.

Baseline characteristics from a randomized clinical trial of treatment-naive subjects conducted at sites in Africa, Asia, the Caribbean, and North and South America were analysed to determine if there were significant differences by sex.

Of the 1,571 participants, 740 (47.1%) were women. Women had higher mean screening CD4⁺ T-cell counts (mean 15 cells higher; P<0.001), lower mean haemoglobin and creatinine clearance, a lower mean baseline HIV-1 viral load (4.85 log₁₀ versus 5.05 log₁₀ copies/ml; P<0.001) and were less likely to have a prior AIDS diagnosis than men. The sex difference in viral load was related to CD4⁺ T-cell count; however, it was independent of country and persisted within the strata with CD4⁺ T-cell count <200 cells/mm³.

Women in resource-limited settings have lower levels of plasma HIV-1 RNA and appear to present for enrolment into clinical trials at an earlier stage of disease than men. The biological basis for lower viral load in women compared to men remains unexplained. It will be important to determine if the sex differences observed at baseline impact clinical outcomes once the PEARLS clinical trial is completed.

Bone mineral density decreases with anti-retroviral therapy (ART) initiation, although the pathogenesis, including the role of tenofovir (TDF), is unclear. This study assessed changes in bone-turnover markers, osteoprotegerin (OPG), soluble receptor activator for nuclear factor-κβ ligand (sRANKL), and inflammation in subjects initiating TDF- versus non-TDF- containing regimens, and determined the relationship between bone turnover, OPG/sRANKL and inflammation.

This was a longitudinal observational study comparing levels of bone turnover markers (C- terminal telopeptide of type I collagen [CTX] and osteocalcin [OC]), OPG, sRANKL and inflammatory cytokines (soluble tumour necrosis factor [TNF]-α receptor [sTNFR]-I, sTNFR-II and interleukin-6) prior to ART and 6-12 months after ART initiation with a TDF- versus non-TDF-containing regimen in HIV-infected subjects 18-50 years old.

A total of 87 subjects were enrolled (TDF n=44 and non-TDF n=43). Groups were similar except subjects on TDF had a lower CD4⁺ T-cell nadir (P<0.01) and were more likely to receive a protease inhibitor (PI; P=0.03). At pre-ART, 35% and 1% of subjects had CTX and OC above the normal range, respectively. Both increased with ART initiation, whereas OPG, sRANKL and inflammatory markers significantly decreased. In multivariate models, increases in OC were associated with TDF use, PI use and pre-ART levels of sTNFR-I, whereas increases in CTX were associated with CD4⁺ T-cell nadir <50 cell/mm³. Increases in bone markers were unrelated to pre-ART levels of OPG/sRANKL and changes in OPG/sRANKL after ART initiation.

TDF use, PI use, TNF-α activity and advanced HIV disease are associated with changes in bone turnover markers, underscoring the complicated interaction between ART, bone turnover, inflammation and immune status, which extend beyond the OPG/RANKL system.

Both IL28B gene polymorphisms and serum levels of interleukin (IL)-10, IL-12p40 and IL-18 have been reported to affect the outcome of natural and pegylated interferon and ribavirin-treated HCV infection.

To clarify their association and predictive value in treatment outcome of genotype 1 HCV- infected patients, we measured pretreatment serum IL-10, IL-12p40 and IL-18 levels using multiplex assays and determined IL28B gene polymorphisms (rs 8099917) in 52 cases with chronic hepatitis C.

High baseline levels of IL-10 (P<0.001) and low levels of IL-12p40 (P<0.001) were significantly associated with a non-virological response (NVR) in our cohort. The IL28B polymorphism was tested and TT, TG or GG genotypes were found in 60%, 38% and 2% of patients, respectively, with corresponding NVR rates of 10%, 60% and 100% (P<0.001). Serum cytokine levels were significantly correlated with IL28B gene polymorphisms. When serum IL-10 levels were stratified at 5.0 pg/ml, NVR rates were 50% versus 0% (P=0.004) for the TT genotype and 87% versus 0% (P=0.001) for the TG or GG genotypes. Similarly, low IL-12p40 levels were associated with an NVR in patients with TG or GG genotypes (P=0.006). In multivariate analysis, high IL-10, low IL-12p40 and IL28B TG or GG genotypes were independently associated with an NVR.

Serum IL-10 and IL-12p40 levels in combination with IL28B genotype, especially G-allele carriage, are strong predictive markers of an NVR to HCV treatment with pegylated interferon and ribavirin.

Interleukin (IL)-6, a pleiotropic cytokine, is increased in various types of chronic liver disease, including chronic hepatitis C (CHC). It was reported recently that IL-6 is associated with insulin resistance, iron metabolism and interferon resistance, which may affect the outcome of antiviral treatment. In this study, we investigated the association of serum IL-6 levels with outcomes of pegylated interferon (PEG-IFN) plus ribavirin (RBV) combination therapy.

We included 149 CHC patients and measured serum IL-6 levels at baseline and at 4, 8 and 12 weeks, and the end of treatment in 49 patients. We performed univariate and multivariate regression analyses for the association of IL-6 levels and clinical and laboratory parameters and treatment responses.

Serum IL-6 levels were significantly higher in CHC patients than healthy subjects. Pretreatment IL-6 levels of male patients were inversely correlated with sustained virological response (SVR) in univariate analysis (P=0.012). In male patients with SVR, serum IL-6 levels decreased significantly at 4 weeks of treatment (P=0.029) and remained significantly lower than those of non-SVR patients after 4, 8 and 12 weeks of PEG-IFN plus RBV therapy.

Our results suggest that baseline levels of IL-6, as well as their decrease during treatment, are correlated to outcomes of PEG-IFN plus RBV therapy in male patients. Further analyses of IL-6 may provide new strategies for difficult-to-treat CHC patients and prevention of hepatocarcinogenesis.

Resistant HCV populations may pre-exist in patients before NS3 protease inhibitor therapy and would likely be selected under specific antiviral pressure. The higher prevalence and lower rate of response to treatment associated with HCV genotype 1 infections has led to drug discovery efforts being focused primarily on enzymes produced by this genotype. Protease inhibitors may also be useful for non-genotype-1-infected patients, notably for non-responders.

We investigated the prevalence of dominant resistance mutations and polymorphism in 298 HCV pro-tease-inhibitor-naive patients infected with HCV genotypes 1, 2, 3, 4 or 5. Genotype-specific NS3 primers were designed to amplify and sequence the NS3 protease gene.

None of the 233 analysed sequences contained major telaprevir (TVR) or boceprevir (BOC) resistance mutations (R155K/T/M, A156S/V/T and V170A). Some substitutions (V36L, T54S, Q80K/R, D168Q and V170T) linked to low or moderate decreases in HCV sensitivity to protease inhibitors were prevalent according to genotype (between 2% and 100%). Other than genotype signature mutations at positions 36, 80 and 168, the most frequent substitution was T54S (4 genotype 1 and 2 genotype 4 sequences). All genotype 2–5 sequences had the non-genotype-1 signature V36L mutation known to confer low-level resistance to both TVR and BOC.

We have developed an HCV protease NS3 inhibitor resistance genotyping tool suitable for use with HCV genotypes 1–5. Polymorphism data is valuable for interpreting genotypic resistance profiles in cases of failure of anti-HCV NS3 protease treatment.

To study the prevalence, predictors and outcome of late HIV diagnosis in the Icona cohort, according to the new European consensus definition of late diagnosis.

In this observational cohort study we investigated patients diagnosed with HIV over 3 months preceding enrolment who were defined as diagnosed late if they presented with AIDS or a CD4⁺ T-cell count ≤350/mm³ (European consensus definition). We estimated the prevalence of late diagnosis, identified factors associated with being diagnosed late and looked at the prognostic value of the European consensus definition of late presentation to predict subsequent clinical progression (new AIDS events or death).

In total, 1,438/2,276 patients (63%) were defined as diagnosed late using the new European Consensus definition. Of these, 387 (16%) were AIDS-presenters. Predictors of being diagnosed late were older age, non-Italian origin, high HIV RNA and unemployment (versus retirement). A total of 293 patients showed clinical progression (3 events/100 person-years of follow-up, 95% CI: 2.7–3.4). Presenting late was strongly associated with a >5-fold increased risk of disease progression.

In our observational setting with free access to care, more than 60% of new HIV diagnoses occurred below the recommended threshold for initiating anti-retroviral treatment. Presenting late for care was associated with a high risk of clinical progression.

We studied prevalence of etravirine (ETR) and rilpivirine (RPV) resistance in HIV-1 subtype CRF01_AE infection with first-line non-nucleoside reverse transcriptase inhibitor (NNRTI) failure.

A total of 225 adults failing two nucleoside reverse transcriptase inhibitors (NRTIs) plus 1 NNRTI in Thailand with HIV RNA>1,000 copies/ml were included. Genotypic resistance results and HIV-1 subtype were interpreted by Stanford DR database. ETR resistance was calculated by the new Monogram weighted score (Monogram WS; ≥4 indicating high-level ETR resistance) and by DUET weighted score (DUET WS; 2.5–3.5 and ≥4 resulted in intermediate and reduce ETR response, respectively). RPV resistance interpretation was based on previous reports.

Median (IQR) age was 38 (34–42) years, 41% were female and CDC A:B:C were 22%:21%:57%. HIV subtypes were 96% CRF01_AE and 4% B. Antiretrovirals at failure were lamivudine (100%), stavudine (93%), nevirapine (90%) and efavirenz (10%) with a median (IQR) duration of 3.4 (1.8–4.5) years. Median (IQR) CD4⁺ T-cell count and HIV RNA were 194 (121–280) cells/mm³ and 4.1 (3.6–4.6) log₁₀ copies/ml, respectively. The common NNRTI mutations were Y181C (41%), G190A (22%) and K103N (19%). The proportion of patients with Monogram WS score ≥4 was 61.3%. By DUET WS, 49.8% and 7.5% of patients were scored 2.5–3.5 and ≥4, respectively. Only HIV RNA≥4 log₁₀ copies/ml at failure was associated with both Monogram WS≥4 (OR 2.3, 95% CI 1.3–3.9; P=0.003) and DUET WS≥2.5 (OR 1.9, 95% CI 1.1–3.3; P=0.02). The RVP resistance-associated mutations (RAMs) detected were K101P (1.8%), Y181I (2.7%) and Y181V (3.6%). All patients with RPV mutation had ETR resistance. No E138R/E138K mutations were detected.

Approximately 60% of patients had high-level ETR resistance. The role of ETR in second-line therapy is limited in late NNRTI failure settings. RVP RAMs were uncommon, but cross-resistance between ETR and RVP was high.

HIV–HBV-coinfected individuals who need to be treated only for their HBV infection have limited therapeutic options, since most approved anti-HBV agents have a risk of selecting for drug-resistant HIV mutants. In vivo data are inconclusive as to whether telbivudine (LdT) may exert antiviral effects against HIV. Thus, we investigated in further detail the antiviral activity and the biochemical properties of LdT against HIV-1.

To investigate the activity of LdT against HIV-1 in humans we analysed viral dynamics and genotypic and phenotypic resistance development in two HIV–HBV-coinfected individuals with no prior antiviral exposure. To investigate the activity of LdT against HIV-1 in vitro, LdT susceptibility for HIV-1 wild-type strains as well as drug-resistant strains was determined. Furthermore, we studied whether the 5′-triphosphate form of LdT (LdT-TP) can act as a substrate for wild-type HIV-1 RT.

In the two patients studied, LdT treatment did not result in a significant decline of HIV-1 RNA load nor in selection of genotypic or phenotypic resistance in HIV-1 RT. In vitro virological analyses demonstrated that LdT had no activity (50% effective concentration >100 μM) against wild type HIV and drug-resistant variants. Biochemical analyses demonstrated that LdT-TP is not incorporated by wild-type HIV-1 RT.

Based on the in vivo and in vitro evidence obtained in this study, we conclude that LdT has no anti-HIV-1 activity and is currently the only selective anti-HBV agent among the five FDA-approved nucleoside/nucleotide analogues for treatment of HBV infections in HIV-infected individuals.

Stavudine, a nucleoside reverse transcriptase inhibitor, is used commonly to treat HIV-infected children in the developing world. The paediatric liquid formulation presents major logistical difficulties in rural and resource-limited areas, and prescribers are frequently forced to employ off-label ‘opened capsule’ dosing methods using the adult capsule. The South African Department of Health (DoH) has advised that caregivers should be instructed to disperse the contents of an adult capsule in 5 ml water and then withdraw the required dose using a syringe. The bioavailability of stavudine using the opened capsule dosing method has not previously been validated.

This was a randomized crossover pharmacokinetic study with each subject serving as his/her own control. A total of 28 healthy HIV-negative adult volunteers were randomized on a 1:1 basis to receive one of the two generic preparations typically available in state hospitals. They were then further randomized to receive either intact or opened 30 mg capsules. After 1 week, those who initially received intact capsules, were given opened capsules and vice versa. The capsule dispersion technique used was identical to that prescribed by the DoH. Serial blood samples were collected and plasma stavudine concentrations were assayed by liquid chromatography tandem mass spectrometry. Stavudine pharmacokinetics were analysed using non-compartmental methods and formulations were compared using ANOVA.

Plasma drug exposure after stavudine administration as a solution using an opened capsule dosing method was found to be bioequivalent to intact capsule administration. This was true for both generics tested.

The opened capsule dosing technique is bio-equivalent to intact capsule dosing for stavudine in HIV-seronegative adults.