Given the relatively small number of patients studied, it is possible that our observations are made by chance, and that there is no genotype-dependent effect of ITX5061 on viral kinetics and further clinical studies are required to explore this further. SR-BI is expressed on the endothelial cells lining the sinusoids that play a major role in the clearance Liquiritin of adenoviral particles from the circulation [30]. ITX5061 in patients undergoing liver transplantation at a single centre (Queen Elizabeth Hospital Birmingham, UK). All patients gave informed consent and ethical approval was given by the UK National Research Ethics Service (reference 10/H0301/36). Patients were allocated sequentially to a no treatment control group or to treatment with ITX5061, 150 mg/day via the enteral route for 1 week. Treatment duration was determined with reference to the known safety profile of ITX5061 in patients without liver disease. Although it was intended that 10 subjects would be enrolled into each group, an interim analysis following the enrolment of the first 5 patients suggested that more detailed HCV kinetic monitoring would provide a more robust baseline of viral kinetics in the untreated patients. The control group was therefore increased to 13 subjects. The study was registered at clinicaltrials.gov (“type”:”clinical-trial”,”attrs”:”text”:”NCT01292824″,”term_id”:”NCT01292824″NCT01292824). Population The study enrolled men and women between the ages of 18 and 65 years who were suitable for liver transplantation. Subjects with HCV associated Rabbit Polyclonal to OR5B3 end-stage liver disease or HCC were enrolled regardless of their infecting genotype or previous anti-viral treatment. Subjects co-infected with HBV or HIV were excluded, as were patients receiving a liver from a HCV positive donor. Study drug ITX5061 was formulated as a 25 mL solution for oral or nasogastric use containing 150 mg drug in a vehicle containing 20% (w/w) hydroxypropyl-beta-cyclodextrin in 10 mM aqueous citric acid. A dose of 150 mg was selected following pre-clinical studies predicting a 10-fold excess over the EC90 for inhibiting HCV entry [18]. Dosing at 150 mg was further supported by studies conducted in the initial development of ITX5061 where this dose was sufficient to block uptake of HDL (the physiological ligand of SR-BI) as evidenced by increased serum HDL levels in treated study participants [17]. The first dose was administered orally approximately 1 hour before the induction of anaesthesia. A second dose was given via a nasogastric tube on arrival to the intensive care unit following liver transplantation and then once daily for 7 days thereafter. Pharmacokinetics Plasma ITX5061 concentrations were measured by liquid chromatography/mass spectrometry [20]. Since ITX5061 is primarily metabolised in the liver an interim analysis of ITX5061 plasma concentrations was performed on the first 3 treated subjects. Review of these data by the trial steering group and by the Medicines and Health Regulatory Authority UK, recommended continued enrolment and treatment of the remaining 7 patients. HCV replication kinetics Plasma was collected at screening, before surgery, at the time of transplantation, and during a follow up period of 90 days. HCV RNA levels were measured on admission to hospital, immediately following the induction of anaesthesia, at the time of portal vein clamping (the start of the anhepatic phase), immediately before perfusion of the allograft, and an hour later. Plasma samples were collected every 4 hours during the first post-transplant day, daily for the first week, weekly for the first month, and monthly thereafter up to 90 days. Plasma HCV RNA was measured using the COBAS TaqMan HCV Test v.2.0 in a Health Protection Agency UK accredited laboratory. Viral sequencing HCV RNA was purified from plasma obtained immediately before surgery and 7 days later. Each sample was analysed by UDPS of the viral structural genes (core, E1, E2 and P7) including the hypervariable region (HVR) using genotype specific primers (Suppl. Table 1). Amplicons were ligated to adaptors (Nextera Tagmentation), amplified by emulsion polymerase chain reaction (PCR) and sequenced on a 454 GS Junior (Roche). The raw sequence outputs (reads) were assembled using the Assemble Viral 454 [21] and VICUNA assembler software [22] to form a consensus assembly. The reads were corrected for systematic 454 errors and aligned to the consensus assembly using the ReadClean 454 and V-Phaser algorithms [23]. Average sequence lengths varied from 342 to 405 nucleotides and typically 3900 reads had been generated Liquiritin for every sample, a complete of 15 to 29 106 bases and the average insurance of 350 to 500 reads for every base. Heat-maps from the viral envelope (E2) area had been generated to graphically represent series polymorphisms. Genetic variety within examples, and divergence between examples had been assessed by determining genetic distance quotes. Liquiritin Pairwise evaluations of sequences allowed quotes of genetic variety of viral quasispecies before and after therapy. Figures The principal endpoint of the.
