Sustained virologic response after therapy with the HCV protease inhibitor narlaprevir in combination with peginterferon and ribavirin is durable through long-term follow-up.

Achievement of a sustained virologic response (SVR) after peginterferon (PEG-IFN) and ribavirin (RBV) treatment is considered to be a marker for the cure of chronic hepatitis C virus (HCV) infection. Long-term follow-up of patients with SVR after treatment with a direct acting antiviral has not yet been described. We used a randomized placebo-controlled, double-blind, two-period phase 1b trial that was conducted in 40 HCV genotype 1 (treatment-naïve and treatment-experienced)-infected patients. Nineteen patients achieved SVR after treatment with the HCV protease inhibitor narlaprevir followed by PEG-IFN/RBV. In these patients, HCV-RNA tests were scheduled at 3, 6, 12 and 24 months after end of treatment. Patients were followed for a median of 27 months (range 15-32) after end of treatment with a median number of follow-up visits of 4 (range 3-8). All patients remained HCV-RNA negative over time. SVR achieved following narlaprevir and PEG-IFN/RBV-therapy was durable up to 32 months after the end of treatment.

Evolutionary dynamics of hepatitis C virus NS3 protease domain during and following treatment with narlaprevir, a potent NS3 protease inhibitor.

Narlaprevir, a hepatitis C virus (HCV) NS3/4A serine protease inhibitor, has demonstrated robust antiviral activity in a placebo-controlled phase 1 study. To study evolutionary dynamics of resistant variants, the NS3 protease sequence was clonally analysed in thirty-two HCV genotype 1-infected patients following treatment with narlaprevir. Narlaprevir monotherapy was administered for one week (period 1) followed by narlaprevir/pegylated interferon-alpha-2b combination therapy with or without ritonavir (period 2) during two weeks, interrupted by a washout period of one month. Thereafter, all patients initiated pegylated interferon-alpha-2b/ribavirin combination therapy. Longitudinal clonal analysis was performed in those patients with NS3 mutations. After narlaprevir re-exposure, resistance-associated mutations at position V36, T54, R155 and A156 were detected in five patients in >95% of the clones. Narlaprevir retreatment resulted in a 2.58 and 5.06 log10 IU/mL viral load decline in patients with and without mutations, respectively (P=<0.01). After treatment, resistant variants were replaced with wild-type virus within 2-24 weeks in three patients. However, the R155K mutation was still observed 3.1 years after narlaprevir dosing in two patients in 5% and 45% of the viral population. Resistant variants could be detected early during treatment with narlaprevir. A slower viral load decline was observed in those patients with resistance-associated mutations detectable by direct population sequencing. These mutations disappeared within six months following treatment with the exception of R155K mutation, which persisted in two patients.

Intra-gastric balloon with lifestyle modification: a promising therapeutic option for overweight and obese patients with metabolic dysfunction-associated steatotic liver disease.

BACKGROUND: Data on effects of intra-gastric balloon (IGB) on metabolic dysfunction-associated steatotic liver disease (MASLD) are scarce, in part with contradictory results, and mainly obtained in tertiary care patients with diabetes and other comorbidities. We here explore effects of IGB in patients with MASLD referred to a first-line obesity clinic. METHODS: In this prospective cohort study, patients with at least significant fibrosis (≥ F2) and/or severe steatosis (S3) according to screening transient elastography (FibroScan®) were offered a second FibroScan® after 6 months lifestyle modification with or without IGB (based on patient preference). RESULTS: 50 of 100 consecutively screened patients (generally non-diabetic) qualified for repeated evaluation and 29 (58%) of those had a second FibroScan®. At baseline, at least significant fibrosis was present in 28% and severe steatosis in 91%. IGB was placed in 19 patients (59%), whereas 10 patients (41%) preferred only lifestyle modification (no differences in baseline characteristics between both groups). After 6 months, liver stiffness decreased markedly in the IGB group (median: from 6.0 to 4.9 kPa, p = 0.005), but not in the lifestyle modification only group (median: from 5.5 to 6.9 kPa, p = 0.477). Steatosis improved in both groups, (controlled attenuation parameter values; IGB, mean ± SD: from 328 ± 34 to 272 ± 62 dB/m, p = 0.006: lifestyle modification only, mean ± SD: from 344 ± 33 to 305 ± 43 dB/m: p = 0.006). CONCLUSION: Both steatosis and fibrosis improve markedly in overweight/obese patients with MASLD after 6 months IGB combined with lifestyle modification. Our results warrant further research into long-term effect of IGB in these patients.

First-in-human study of the pharmacokinetics and antiviral activity of IDX375, a novel nonnucleoside hepatitis C virus polymerase inhibitor.

IDX375 is a potent and selective palm-binding nonnucleoside inhibitor of the hepatitis C virus (HCV) genotype 1 polymerase. This first-in-human study evaluated the safety, tolerability, and pharmacokinetics of IDX375 in healthy volunteers, as well as its antiviral activity in HCV-infected patients. IDX375, as a choline salt, was administered for 1 day to 40 healthy male volunteers (25- to 200-mg IDX375-equivalent single ascending doses and a 200-mg twice-daily [BID] dose) and three patients chronically infected with HCV genotype 1 (200 mg BID only). IDX375 was well absorbed and well tolerated by all of the study participants. A single-day 200-mg BID dose resulted in exposure-related anti-HCV activity with maximal 0.5 to 1.1 log(10) reductions in plasma HCV RNA. These observations support further clinical investigations of IDX375.

New developments in the antiviral treatment of hepatitis C.

Chronic hepatitis C virus (HCV) infection is a major cause of liver cirrhosis and hepatocellular carcinoma. HCV is endemic in most parts of the world, with an estimated 170 million people infected worldwide and 3-4 million new cases each year. HCV-related end-stage liver disease is now the main indication for liver transplantation in the USA and Western Europe. Unfortunately, no vaccine or immunoglobulin is available to prevent HCV infection. Currently, HCV treatment consists of the combined administration of pegylated interferon and ribavirin for a period of 24-48 weeks, resulting in complete viral eradication in 40-80% of patients, depending on genotype, viral load and patient characteristics. This therapy is often accompanied with side-effects that affect compliance and reduce treatment outcomes. Recently, reliable in vitro culture systems have been developed which accelerated antiviral therapy research. Many new specifically targeted antiviral therapies for hepatitis C (STAT-C) and treatment strategies are evaluated in clinical trials. These new antiviral agents are expected to improve treatment significantly with potentially shorter treatment duration. The most promising antiviral agents will be reviewed.

Sofosbuvir Add-on to Ribavirin Treatment for Chronic Hepatitis E Virus Infection in Solid Organ Transplant Recipients Does Not Result in Sustained Virological Response.

Ribavirin is effective for treating immunocompromised patients with chronic hepatitis E virus infection. However, ribavirin treatment is not always successful. We describe 3 solid organ transplant recipients treated with sofosbuvir and ribavirin after failing ribavirin monotherapy. Complete elimination of hepatitis E virus could not be achieved.

Treatment of chronic hepatitis C virus infection - Dutch national guidelines.

The development of this guideline was initiated and coordinated by the Netherlands Association of Gastroenterologists and Hepatologists (Nederlandse Vereniging van Maag-Darm-Leverartsen). The aim is the establishment of practical guidelines in the evaluation and antiviral treatment of patients with chronic hepatitis C virus (HCV) infection. This includes recommendations for the initial evaluation of patients, the choice and duration of antiviral therapy and the follow-up after antiviral therapy. Hepatitis C is a slowly progressive disease. The initial evaluation of chronically HCV-infected patients should include liver biochemistry testing, virological testing and abdominal ultrasound imaging. Liver biopsy is no longer a routine procedure. Antiviral treatment should be considered for all HCV-infected patients. Current antiviral treatment is a long-term process and is associated with substantial side effects. When deciding whether to start treatment or not, the chance of successful treatment (80% with hepatitis C genotype 2 and 3 and 50% with hepatitis C genotype 1 and 4), the fibrosis stage, the expected side effects and the compliance of the patient should be taken into consideration. In the absence of significant fibrosis and necroinflammation in liver biopsy, postponing treatment is an option. Current antiviral treatment is contraindicated in patients with Child-Pugh-class B or C cirrhosis. The possibility of a liver transplantation should be investigated in these patients. Significant comorbidity with a limited life expectancy is an absolute contraindication for antiviral treatment Treatment of chronic hepatitis C consists of administration of peginterferon and ribavirin for 24 or 48 weeks. Patients with hepatitis C genotype 1 or 4 are treated for 48 weeks. Patients with hepatitis C genotype 2 or 3 are treated for 24 weeks. In patients with undetectable HCV RNA after four weeks (28 days) of treatment, a shorter treatment is equally effective (12 to 16 weeks for hepatitis C genotype 2 or 3; 24 weeks for hepatitis C genotype 1 or 4). Outpatient clinic visits are recommended at the start and after 2, 4, 8, and 12 weeks of treatment, and thereafter every four to six weeks until the end of treatment. It is recommended to stop treatment if the HCV RNA level has not decreased by at least 2 log10 IU/ml (c/ml) after 12 weeks of treatment or when HCV RNA is still detectable after 24 weeks of treatment. The recommended frequency of outpatient clinic visits for patients who are not being treated is once every six months in patients with cirrhosis, otherwise every 12 months. It is expected that new anti-HCV-medication (STAT-C, specifically targeted antiviral therapy for HCV) will become available in the near future. Therefore treatment of chronic HCV infection will probably be more effective in the future.

Miravirsen dosing in chronic hepatitis C patients results in decreased microRNA-122 levels without affecting other microRNAs in plasma.

BACKGROUND: MicroRNA-122 (miR-122) is an important host factor for hepatitis C virus replication. Administration of miravirsen, an anti-miR-122 oligonucleotide, resulted in a dose dependent and prolonged decrease in HCV RNA levels in chronic hepatitis C patients. AIM: To assess the plasma level of various miRNAs in patients dosed with miravirsen. METHODS: We included 16 of 36 chronic hepatitis C patients who received five injections of either 3 mg/kg (n = 4), 5 mg/kg (n = 4), 7 mg/kg (n = 4) miravirsen or placebo (n = 4) over a 4-week period in a double-blind, randomised phase 2a study. Plasma levels of 179 miRNAs were determined by qPCR and compared between patients dosed with miravirsen or placebo. RESULTS: Median plasma miR-122 level at baseline in patients receiving miravirsen was 3.9 × 10(3) compared to 1.3 × 10(4) copies/4 µL in placebo-dosed patients (P = 0.68). At week 1, 4, 6 and 10/12, patients dosed with miravirsen had respectively a median 72-fold, 174-fold, 1109-fold and 552-fold lower expression of miR-122 than at baseline (P = 0.001, as compared to patients receiving placebo). At week 4 of dosing, miRNA-profiling demonstrated a significant lower expression of miR-210 and miR-532-5p compared to baseline (3.0 and 4.7-fold lower respectively). However, subsequent longitudinal analysis showed no significant differences in miR-210 and miR-532-5p plasma levels throughout the study period. CONCLUSIONS: We demonstrated a substantial and prolonged decrease in plasma miR-122 levels in patients dosed with miravirsen. Plasma levels of other miRNAs were not significantly affected by antagonising miR-122.

Treatment of young rats with cholestyramine or a hypercholesterolemic diet does not influence the response of serum cholesterol to dietary cholesterol in later life.

Groups of 10 female Wistar rats (aged 4 weeks) were fed for 29 days either a low-cholesterol commercial diet, a commercial diet containing 2% (w/w) cholesterol, 0.5% cholate and 5% olive oil or a diet containing 2% cholestyramine. The rats were then fed the low-cholesterol commercial diet for the next 91 days and the high-cholesterol diet for another 29 days. There was no significant difference between the groups in the increase of cholesterol in serum and liver during the last period of cholesterol feeding. A fourth group of 10 animals was fed the diet containing cholesterol and cholate during the entire experimental period of 149 days. By the end of the experiment serum cholesterol in these animals was lower and liver cholesterol was higher than in the 3 groups fed the high-cholesterol diet during days 120-149 of the experiment. This study does not present evidence for imprinting effects of early diet manipulation on the later cholesterolemic response to a high cholesterol diet.

Esterases in inbred strains of mice with differential cholesterolemic responses to a high-cholesterol diet.

Specific esterase isoenzyme patterns in plasma may be associated with responsiveness of serum cholesterol to dietary cholesterol. In rabbits and rats the presence and absence of a high-mobility, anodal esterase band on electrophoresis have been shown to be associated with hypo- and hyperresponsiveness, respectively. We fed for 28 days male mice of 7 inbred strains either a low-cholesterol, commercial diet or a diet containing 2% (w/w) cholesterol, 0.5% cholic acid and 5% olive oil. Feeding the high-cholesterol diet revealed marked inter-strain differences in the responses of plasma and liver cholesterol; the increases ranged from 21 to 129% and from 10 to 80-fold, respectively. There was no association between esterase isoenzyme patterns in plasma and the sensitivity to the high-cholesterol diet. The mean baseline plasma total esterase activity tended to be positively associated with the absolute response of plasma cholesterol to the high-cholesterol diet (r = 0.56; n = 7), but the positive relationship between the baseline concentration of the ES-1 component in plasma and the cholesterolemic response was stronger (r = 0.84; n = 7; P less than 0.05). The high-cholesterol diet caused a significant increase in plasma total esterase activities in 6 out of the 7 strains. Evidence is presented that the increase in plasma total esterase activity, which was associated with an increase in the activity and concentration of the so-called ES-2 isoenzyme, is the result of an enhanced release of esterases from the intestine, rather than from the liver. A significant, positive correlation was found between the baseline intestinal esterase activity and the cholesterolemic response after cholesterol feeding (r = 0.83; n = 7; P less than 0.05).

Biochemical characterization of pituitary-dependent hyperadrenocorticism in the dog.

The biochemical characterization of 22 cases of pituitary-dependent hyperadrenocorticism in the dog, is reported. The principal characteristics of the disease include excessive and non-rhythmic production of cortisol, decreased sensitivity of the hypothalamic-pituitary system to the suppressive effects of dexamethasone, decreased responsiveness of the pituitary-adrenocortical system to the stimulus of insulin-induced hypoglycaemia and increased responsiveness of the system to stimulation with lysine-vasopressin. From these observations it is concluded that pituitary-dependent hyperadrenocorticism in the dog is a valid model for study of the pathogenesis of the disease in man. For the diagnosis of hyperadrenocorticism itself, the measurement of the concentration of corticosteroids in a single sample of plasma obtained 8 h after intravenous injection of 0.01 mg dexamethasone/kg was sufficient. The level of 11-hydroxycorticosteroids was less than 140 nmol/1 plasma in normal dogs, whereas higher values were found in dogs with hyperadrenocorticism. For purposes of differential diagnosis, measurement of the level of corticosteroids in the plasma both before and 4 h after intravenous injection of 0.05 mg dexamethasone/kg is adequage: suppression is obtained only in cases of pituitary-dependent hyperadrenocorticism.

A common deletion in two gamma ray induced rat pulmonary tumor cell lines.

Subtraction hybridization was performed on normal WAG/Rij rat DNA with DNA from a syngeneic Ir-192 induced pulmonary tumor cell line L37. The residual DNA was amplified by means of sequence-independent PCR. This procedure yielded a sequence, of which multiple copies are present in normal rat DNA. In the tumor line L37 two restriction fragments hybridizing with this repeat sequence are lacking. In another Ir-192 induced pulmonary tumor line, L33, one of these fragments was also lacking. This indicates a common deletion in the two tumor lines.

Biodegradation pathway of 2-chlorodibenzo-p-dioxin and 2-chlorodibenzofuran in the biphenyl-utilising strain JB1.

The biphenyl-utilising Burkholderia (previously Alcaligenes) strain JB1 is also able to degrade a number of chlorinated dibenzo-p-dioxins and dibenzofurans. In this study, 4-chlorocatechol and a chlorotrihydroxydiphenyl ether were identified as metabolites of 2-chlorodibenzo-p-dioxin. 5-Chlorosalicylic acid and a chlorotrihydroxybiphenyl were metabolites of 2-chlorodibenzofuran. These results show that degradation of these compounds follows pathways in which the initial reaction is angular dioxygenation, followed by cleavage of an ether bridge. This pathway is similar to that used by dibenzofuran-degrading strains such as Sphingomonas sp. strain RW1.

Pentagastrin stimulated gastric secretion in the dog (orogastric aspiration technique).

An orogastric aspiration technique is presented for the estimation of pentagastrin stimulated gastric secretion in the anaesthetised dog. Reference values for volume, pH and hydrogen ion output, 'acidity' and hydrochloric acid output, chloride, sodium, potassium and pepsin outputs were measured and calculated in 23 clinically healthy dogs. It was concluded that 30 and 45 minute hydrogen ion, hydrochloric acid and chloride peak outputs were the most suitable parameters for the acid secreting capacity of the parietal cells. Reference values for 30 and 45 minute peak outputs were respectively 1.63 to 3.61 and 1.50 to 3.35 mmol per hour per kg0.75 (hydrogen ion), 1.86 to 3.64 and 1.65 to 3.42 mmol per hour per kg0.75 (hydrochloric acid) and 2.35 to 4.13 and 1.98 to 3.91 mmol per hour per kg0.75 (chloride). Reproducibility of pepsin outputs was so poor that they cannot be used as parameters for gastric secretory capacity.

Dietary cholesterol-fat type combinations and carbohydrate and lipid metabolism in rats and mice.

In rats fed semipurified cholesterol-free diets, dietary corn oil induced higher levels of liver cholesterol, but lower concentrations of plasma cholesterol, plasma triglycerides and blood D-3-hydroxybutyric acid, than did coconut fat. Addition of cholesterol to the diets (1%, w/w) greatly increased liver cholesterol levels but did not affect the corn oil effect upon liver and plasma cholesterol. In contrast, dietary cholesterol prevented the corn oil effects upon plasma triglycerides and blood D-3-hydroxybutyric acid. Liver glycogen levels in rats increased significantly after cholesterol feeding, the effect being independent of dietary fat type. In mice, dietary corn oil, compared to coconut fat, elevated liver cholesterol only when the diet contained cholesterol. Corn oil caused an increase of liver glycogen in mice in the presence, but not in the absence of dietary cholesterol. This study shows that dietary cholesterol-fat type combinations influence various aspects of lipid and carbohydrate metabolism in rats and mice.

Capillary and venous blood compared with arterial blood in the measurement of acid-base and blood gas status of dogs.

Acid-base and blood gas status of capillary blood from the ear was compared with acid-base and blood gas status of arterial and venous blood in dogs with induced hypoxemia, hypercapnea, and hypovolemia. It was concluded that capillary blood samples can replace arterial blood samples for the measurement of acid-base and blood gas status of dogs with severe impairment of lung function. When circulation is impaired capillary blood samples cannot replace arterial blood samples for the measurement of acid-base and blood gas status of dogs. With impaired capillary circulation, the capillary-arterial differences become unacceptably large. It is, therefore, advisable to submit samples of capillary blood for analysis only if circulation is not impaired and microtubes can be filled quickly and easily.

Gamma camera imaging of bilateral adrenocortical hyperplasia and adrenal tumors in the dog.

Gamma camera imaging of the adrenal glands was done in 8 dogs with hyperadrenocorticism and 4 normal dogs given intravenous injections of 131I-19-iodocholesterol. In normal dogs, both adrenal glands could be visualized separately, and there was no difficulty in distinguishing among the images of normal glands, hyperplastic glands, and functional adrenal tumors. In addition, gamma camera imaging enabled the correct surgical site to be selected for removal of adrenal tumors. Hyperadrenocorticism was diagnosed in 8 dogs by evaluation of urinary 17-hydroxycorticosteroid (OHCS) excretion rates, urinary 17-OHCS and plasma 11beta-OHCS RESPONSES TO DEXAMETHASONE SUPPRESSION OF ENDOGENOUS ADRENOCORTICOTROPIN (ACTH) secretion, and plasma 11beta-OHCS response to intravenous administration of ACTH. Base line 17-OHCS excretion increased in 5 of the 8 dogs. Plasma 11beta-OHCS concentrations were not decreased by dexamethasone administration in the 4 dogs subsequently found to have adrenal tumors; however, there was an exaggerated increase in plasma 11beta-OHCS concentration after administration of ACTH in 3 of the 4 dogs which had bilateral adrenocortical hyperplasia.

The significance of bile pigment deconjugation by beta-glucuronidase in canine hyperbilirubinemia.

In dogs, the differentiation between haemolytic and cholestatic hepatobiliary diseases cannot be achieved by measuring of the unconjugated:conjugated bilirubin ratio, which is in contrast with generally held clinical concepts. The overlap of the bilirubin ratios between the two groups of icterus-generating diseases might in part be explained by deconjugation of conjugated bilirubin. Enzymatic cleavage by hepatic beta-glucuronidase might result in higher unconjugated bilirubin (UCB) fractions in cholestatic disease. The influence of deconjugation of bilirubins by beta-glucuronidase was investigated in 25 healthy dogs and 35 dogs with spontaneous hyperbilirubinemia due to either hepatobiliary or haemolytic disease. UCB and its mono- and diconjugates were measured by alkaline methanolysis and HPLC in plasma and liver tissue. The activity of beta-glucuronidase was also measured in both liver and plasma. In addition, semiquantitative histochemical quantitation of bilirubins in liver tissue was performed. The concentration and the fraction of UCB in plasma of dogs with hepatobiliary disease were not significantly different from those of dogs with autoimmune haemolytic anaemia. There was a correlation between the fraction of UCB in liver and plasma of jaundiced dogs (r = 0.42, P less than 0.01) and between the histochemically estimated and the biochemically measured total bilirubin concentration in liver tissue. There was no correlation between the beta-glucuronidase activity and either unconjugated or monoconjugated bilirubin in plasma or liver of diseased animals. The fraction and the concentration of UCB in the liver of dogs with hepatic and with haemolytic disease were identical. It is concluded that beta-glucuronidase activity is not the significant factor in explaining the similar levels and fractions of UCB in dogs with hyperbilirubinemia due to either hepatobiliary or haemolytic disease.

Hyperlipoproteinaemia associated with atherosclerosis and cutaneous xanthomatosis in a cat.

Bilateral uveitis and multiple xanthomas (fat deposits in the skin) are described in a 3.5-year-old ovariohysterectomized female Persian cat. The cat had been treated for 2 years with corticosteroids. Examinations included a routine blood chemistry profile, radiographic examination of the thorax and abdomen, histopathological examination of multiple skin punch biopsies, and analysis of blood lipid components by cellulose-acetate electrophoresis and by preparative ultracentrifugation studies. Total lipid values were 23 g/l. Ultracentrifugation studies indicated strongly elevated VLDL and LDL fractions and a decreased concentration of the HDL fraction. Because of sudden blindness the cat was euthanized at the request of the owner. Autopsy revealed massive atherosclerotic changes in the large abdominal vessels, the wall of the aorta, and the coronary vessels. Although the exact pathogenesis remains uncertain, these unusual findings might be explained by a primary hyperlipoproteinaemia, complicated by long-term use of corticosteroids.

Corticosteroid-induced alkaline phosphatase isoenzyme in the diagnosis of canine hypercorticism.

Corticosteroid-induced isoenzyme of alkaline phosphatase (AP) can easily be demonstrated in canine plasma as a routine procedure because of its greater heat stability at 65 degrees C in comparison with that of other AP-isoenzymes. In this study the accuracy of this test for the diagnosis of hypercorticism was investigated. The AP-65 degrees C test had its highest efficiency when applied to plasma AP levels exceeding 150 units/litre. In a group of 146 dogs, clinically suspected of having hyperadrenocorticism, the test had a sensitivity of 0.92 and a positive predictive value for a positive test result of 0.89. Its lack of specificity (0.44) makes it unsuitable as a diagnostic test. The main application of AP-65 degrees C is in detecting hypercorticism in dogs by routine laboratory measurements, as was demonstrated in 711 dogs, in which a positive predictive value for the presence of hypercorticism of 0.89 was found.