Personalized treatment of hepatitis C genotype 1a in Norway and Sweden 2014-2016: a study of treatment outcome in patients with or without resistance-based DAA-therapy.

OBJECTIVES: Resistance-associated substitutions (RASs) may impair treatment response to direct-acting antivirals (DAA) in hepatitis C virus (HCV) treatment. We investigated the effects of baseline NS3-RASs (Q80K and R155K) and clinically relevant NS5A-RASs in patients with HCV genotype (GT) 1a infection on treatment outcome, with or without resistance-based DAA-treatment. This multi-center study was carried out between 2014 and 2016. PATIENTS/METHODS: Treatment in the intervention group (n = 92) was tailored to baseline resistance. Detection of NS3-RAS led to an NS5A-inhibitor-based regimen and detection of NS5A-RAS to a protease-inhibitor regimen. Patients without baseline RAS in the intervention group and all patients in the control group (n = 101) received recommended standard DAA-treatment. RESULTS: The sustained virologic response rates (SVR) in the intervention and control groups were 97.8% (90/92) and 93.1% (94/101), respectively (p = .174). A trend toward higher SVR-rate in cirrhotic patients (p = .058) was noticed in the intervention group compared to the control group with SVR-rates 97.5% (39/40) and 83.3% (35/42), respectively. All patients with baseline NS3 (Q80K/R155K) or NS5A-RASs in the intervention group achieved SVR with personalized resistance-based treatment. In the control group, five patients with Q80K or R155K at baseline were treated with simeprevir + sofosbuvir and treatment failed in two of them. Furthermore, one of three patients who failed ledipasvir + sofosbuvir treatment had NS5A-RASs at baseline. CONCLUSIONS: In line with the findings of the OPTIMIST-2 trial for Q80K and the EASL-guidelines 2016 for NS5A-RASs, baseline RASs appeared to have an impact on treatment outcome albeit a statistical significance was not observed in this low-prevalence population.

Prevalence of polymorphisms with significant resistance to NS5A inhibitors in treatment-naive patients with hepatitis C virus genotypes 1a and 3a in Sweden.

BACKGROUND: The future treatment of hepatitis C virus (HCV) infection will be combinations of direct-acting antivirals (DAAs) that not only target multiple viral targets, but are also effective against different HCV genotypes. Of the many drug targets in HCV, one promising target is the non-structural 5A protein (NS5A), against which inhibitors, namely daclatasvir, ledipasvir and ombitasvir, have shown potent efficacy. However, since HCV is known to have very high sequence diversity, development of resistance is a problem against but not limited to NS5A inhibitors (i.e. resistance also found against NS3-protease and NS5B non-nucleoside inhibitors), when used in suboptimal combinations. Furthermore, it has been shown that natural resistance against DAAs is present in treatment-naïve patients and such baseline resistance will potentially complicate future treatment strategies. METHODS: A pan-genotypic population-sequencing method with degenerated primers targeting the NS5A region was developed. We have investigated the prevalence of baseline resistant variants in 127 treatment-naïve patients of HCV genotypes 1a, 1b, 2b and 3a. RESULTS: The method could successfully sequence more than 95% of genotype 1a, 1b and 3a samples. Interpretation of fold resistance data against the NS5A inhibitors was done with the help of earlier published phenotypic data. Baseline resistance variants associated with high resistance (1000-50,000-fold) was found in three patients: Q30H or Y93N in genotype 1a patients and further Y93H in a genotype 3a patient. CONCLUSION: Using this method, baseline resistance can be examined and the data could have a potential role in selecting the optimal and cost-efficient treatment for the patient.

Implications of baseline polymorphisms for potential resistance to NS3 protease inhibitors in Hepatitis C virus genotypes 1a, 2b and 3a.

The future interferon-free treatment of hepatitis C virus (HCV) infection could include NS3 protease inhibitors (PIs) for potent pan-genotypic effect. We studied the prevalence of pre-existing PI resistance associated amino acid variants (RAVs) in 126 treatment-naive patient samples of HCV genotypes 1a, 2b and 3a, the most common genotypes in Sweden. The NS3 genes were each amplified by nested PCR method with degenerated primers to enable a broad genotype analysis. Population sequencing method was used, and the sequences were aligned with the NS3 sequence from HCV genotype 1a H77 strain. Interpretation of fold-change resistance to NS3 candidate drugs were done from already published phenotypic resistance data. The prevalence of known PI RAVs at baseline in genotype 1a was 28% (15/53), either single (V36L or Q80K/R) or combinations (T54A/S and V55A/I) of mutation(s). In genotype 2b, specific mutations like V36L, Q80G and S122R of viral NS3 protease gene were found in 100% (11/11). These may be the natural polymorphisms unique to genotype 2b. Similarly, specific mutations like V36L and D168Q were found uniquely in all 3a samples (30/30). The natural PI RAVs found in genotype 1a, although with relatively weak resistance, could still render up to 10-fold-resistance to the approved (boceprevir and telaprevir) and the 2nd generation PIs (faldaprevir and simeprevir). Moreover, the natural polymorphisms in genotype 2b (i.e. S122R) and 3a (i.e. D168Q), with inherent PI drug resistance of up to 20 and 700 fold respectively, would explain why current PIs are primarily directed against genotype 1.

Bartonella spp. seroprevalence in healthy Swedish blood donors.

Serum samples were collected from healthy blood donors in 5 regions in Sweden in 1999, i.e. from the local Blood Centres (collecting facilities) in Boden, Jönköping, Lund, Skövde, and Uppsala. In total, 498 serum samples (63% males, 37% females) were received and tested by immunofluorescence assay for antibodies against B. elizabethae, B. grahamii, B. henselae (Houston-1), B. henselae (Marseille), B. quintana, and B. vinsonii subsp. vinsonii. An overall Bartonella spp. seroprevalence of 16.1% was found, with a predominance of immunoreactivity to B. elizabethae, at 14.1%; B. grahamii, 2.6%; B. henselae (Houston-1), 1.2%; B. henselae (Marseille), 1.8%; B. quintana, 0.2%; and B. vinsonii subsp. vinsonii, 0.0%. Univariate and multivariate analyses of epidemiological and demographical information revealed an increased rate of B. elizabethae seropositivity in blood donors working outdoors, being out in the wild a minimum of once a week, hunting moose, having cat contact, and travelling to Eastern Europe. Living in the southern region of Sweden (Lund area) was associated with decreased seropositivity to B. elizabethae.

Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on trace elements, inflammation and viral clearance in the myocardium during coxsackievirus B3 infection in mice.

A myocarditic coxsackievirus B3 (CB3) infection in adult male A/J mice was used to investigate the effects of 2,3,7,8-tetrachlorodibenzo-para-dioxin (TCDD) exposure on mortality and on inflammatory lesion, virus and trace element contents of the heart. The mice were injected with four weekly intraperitoneal (i.p.) injections of TCDD (a loading dose of 5 microg/kg followed by three maintenance doses of 1.4 microg/kg). To reach a steady-state body burden of TCDD the mice were allowed a 90-day recovery period before infection with CB3 virus. TCDD increased the infection-induced mortality rate, whereas in TCDD-exposed mice, heart lesions at day 7 after the virus inoculation (median value 0.67% of the tissue section area; interquartile range 0.28; not statistically significant) were one-third of that in non-exposed infected mice (2.07% of the tissue section area; interquartile range 3.06). The size of the inflammatory heart lesion correlated to the amount of virus (r(s) = 0.829, P < 0.01) as well as to the calcium (Ca: r(s) = 0.725, P < 0.01) and the magnesium (Mg: r(s) = -0.615, P < 0.05) contents. In TCDD-exposed mice in situ hybridisation of viral RNA in the myocardium at day 7 showed a tendency to decreased amounts of virus, as well as a less pronounced increase in myocardial Ca content, both supporting a milder myocardial disease after TCDD exposure. No effect of TCDD exposure was seen on the zinc (Zn) or selenium (Se) levels in the myocardium. In conclusion, although TCDD seemed to have a limiting effect on viral replication and the development of the inflammatory lesion in the myocardium, mortality was increased by TCDD in this infection model. However, TCDD had no significant effects on the selected trace elements that could be of importance for the severity of the inflammatory lesion (Ca, Se), for the local host response activation (Zn) or for the development of myocardial disease complications (Mg). Accordingly, the increased mortality may be a result of an infection-induced increase in TCDD toxicity to vital organs other than the heart, and/or a TCDD-induced change in the tissue affinity and virulence of the virus, possibly causing involvement of other target organs in the infectious process and changed pathogenesis.