The study enrolled healthy male and female volunteers. Eligible subjects were required to be aged 18–45 years; have a body weight greater than 55 kg and a body mass index (BMI) of > 20 and ≤ 30 kg/m2; have vital signs within the normal range; and to be either non-smokers or smoke fewer than 10 cigarettes per day. Females were also required to have a negative serum pregnancy test within 3 weeks of the trial start and a negative urine pregnancy test at the day before dosing. The following exclusion criteria were applied: a history or presence of cholelithiasis, diabetes, tumours in the pituitary gland or hypothalamus, any serious allergy, positive serological test for hepatitis B or C and HIV, hypertension or other significant cardiovascular abnormality. Subjects were also excluded if they had a significant history or clinical evidence of auto-immune, gastrointestinal, haematological, haematopoietic, hepatic, neurological, pancreatic or renal disease, or had a positive drug or alcohol test or chronic use of medication.
The single-centre trial (NCT01034735), carried out at a clinical pharmacology research centre in Germany, had an open, randomized, three-way crossover design. Treatment with r-hGH (Saizen®; Merck Serono S.A. - Geneva) started within 21 days of screening. Each volunteer received three treatments: reconstituted FD (8.8 mg/1.51 mL) and two liquid formulations (5.83 and 8.0 mg/mL). All three treatments were administered as a single subcutaneous dose of 4 mg, to allow proper determination of the PK parameters. The doses were administered in a randomized sequence with a 1-week wash-out period between each administration. The doses were injected into the anterior abdominal wall, around the umbilicus at the top of a skin fold, using a needle and syringe, and were injected at a distance of at least 10 cm from one another in pre-marked locations.
Additionally, intravenous somatostatin was infused continuously from 1 hour before to 24 hours after each dose, achieving a cumulative dose of 3 mg. This pituitary down-regulation was necessary to suppress endogenous GH secretion in the healthy volunteers to allow reliable calculation of PK parameters.
For each r-hGH administration, the subject attended the clinical unit on the day before the study drug administration (days -1, 7 and 14) and stayed in the unit until 26 hours after drug administration (days 2, 9 and 16). Blood samples for PK analysis were taken hourly up to 10 hours, and then at 12, 18 and 24 hours post-dose. Vital signs, safety and tolerability were assessed before and 24–26 hours after each dose and 14 ± 3 days after the last dose.
The trial was conducted in compliance with the Declaration of Helsinki and the ICH Guideline of Good Clinical Practice, in addition to European, US and German directives for proper conduct of clinical drug trials. The study was approved by the national regulatory agency (the Federal Institute for Drugs and Medicinal Products [Bundesinstitut für Arzneimittel und Medizinprodukte]) and the independent ethics committee of the Bavarian Chamber of Physicians. Participants gave written informed consent at the start of the trial.
The primary objective of the study was to assess the bioequivalence of two concentrations of the liquid formulation of r-hGH in comparison with the FD formulation. The secondary objectives were to evaluate safety and tolerability, and to describe the PK parameters of the liquid formulations of r-hGH in comparison with the FD formulation.
Primary PK endpoints were the area under the serum concentration-time curve from time 0 to the last measurable concentration time point after drug administration (AUC0-t) and the maximum serum concentration (Cmax). Secondary PK endpoints included the total area under the serum concentration-time curve extrapolated to infinity (AUC0-∞), time to reach Cmax (tmax), terminal elimination half life (t1/2), apparent volume of distribution (Vz/f) and apparent clearance (CL/f).
Quantitative determination of r-hGH in human serum samples was performed using a 'two-step' sandwich-type immunoassay. The assay was fully validated according to the FDA and EMA guidelines on bioanalytical method validation and best practice recommendations for the validation and implementation of quantitative bioanalytical methods.[9–11] The lower limit of quantification (LLOQ) of the immunoassay was 0.4 ng/mL. The actual administered dose was determined by gravimetric measurement, subtracting the post-dose weight of the syringe from the pre-dose weight and calculating with a density of 1.02 g/mL for the 5.83 mg/mL liquid formulation and FD formulation and 1.03 g/mL for the 8 mg/mL liquid formulation.
Tolerability was assessed by adverse events (AEs) queried on an ongoing basis from the day prior to dosing to at least 14 ± 3 days after the last dose administration. Additionally, tolerability was evaluated immediately pre-dose, at the time of administration (injection-site reactions [ISRs] only), 5 minutes, and 2, 4, 6, 12 and 24 hours after each injection. ISRs were assessed by the investigator, whereas itching at the injection site and pain were assessed by the subject; a 100 mm visual analogue scale (VAS) was used to assess pain.
Safety assessments comprised physical examination, vital signs (blood pressure, pulse rate, body temperature), supine 12-lead ECG recordings and laboratory tests. The physical examination was made at screening and at 14 ± 3 days after each administration. Vital signs were taken immediately pre-dose, 1, 6, 12 and 24 hours after treatment administration, and at the post-study visit. ECG measurements were taken before each dose.
A total of 24 evaluable subjects (four subjects for each treatment sequence) were required to provide approximately 90% power for demonstrating bioequivalence. This calculation assumed intra-subject coefficients of variation of approximately 12% for AUC and 17% for Cmax, based on earlier studies with the FD formulation and values between 0.95 and 1.05 for the true treatment ratio test/reference. The 95% confidence level (CI) was used to adjust for multiplicity. To account for potential drop-outs, 30 subjects were enrolled.
Subjects were randomly assigned in chronological order to a treatment sequence, according to the randomization list, with an equal number of subjects in each of the six treatment sequences. Randomization of each subject to a treatment sequence occurred immediately before dosing on study day 1.
PK Analysis and Statistical Methods
PK analyses were performed for the PK population and the safety analyses were performed for the safety population. The PK population included all subjects who had been treated according to the protocol in all trial periods who fulfilled the following criteria: all inclusion/exclusion criteria satisfied; adequate trial medication compliance; absence of relevant protocol violations and availability for the primary target variables for at least one treatment. The safety population included all subjects who received at least one dose of trial medication and had follow-up safety data.
The PK parameters of r-hGH were calculated according to non-compartmental standard methods using the validated computer program KINETICA (Version 4.4.1, Thermo, Philadelphia, USA). Statistical analysis was performed using the computer program package SAS System for Windows (TM version 8.2; SAS Institute, Cary NC, USA).
A mixed model was fitted to each of the log-transformed PK endpoints, Cmax, AUC0-t, Cmax adjusted to target dose (Cmax adj) and AUC0-t adjusted to target dose (AUC0-t adj) with fixed effect terms for treatment, period and sequence, and subject as a random effect. In compliance with the relevant FDA and EMA guidelines,[9–11] for each of the two test liquid formulations, bioequivalence was concluded if the 95% CIs for the estimated test/reference ratios of geometric means of AUC0-t and Cmax were within the standard pre-specified acceptance range for bioequivalence (0.80–1.25). In addition, a further correction for the measured active content (as determined by the certificate of analysis of each formulation) was applied to the ratio estimates and confidence limits in order to further assess bioequivalence (Table 1).
All PK parameters were presented in a descriptive way per treatment group (number of subjects [n], mean, standard deviation [SD], median, geometric mean, standard error of mean [SEM], coefficient of variation, minimum [Min] and maximum [Max] values). Mean serum concentrations were also described per time point and treatment. Values below the LLOQ were taken as zero for descriptive statistics of concentrations.
The numerical values for tolerability and safety variables were summarized descriptively for the safety population.
BMC Clin Pharmacol. 2010;10(14):1-7. © 2010
BioMed Central Ltd.
Cite this: Comparison of the Pharmacokinetics, Safety and Tolerability of Two Concentrations of a New Liquid Recombinant Human Growth Hormone Formulation versus the Freeze-dried Formulation - Medscape - Oct 20, 2010.