Anne Plum

The mechanism of protraction of insulin detemir, a long-acting, acylated analog of human insulin.

AbstractPurpose. Insulin detemir has been found in clinical trials to be absorbed with very low variability. A series of experiments were performed to elucidate the underlying mechanisms. Methods. The disappearance from an injected subcutaneous depot and elimination studies in plasma were carried out in pigs. Size-exclusion chromatography was used to assess the self-association and albumin binding states of insulin detemir and analogs. Results. Disappearance T50% from the injection depot was 10.2 ± 1.2 h for insulin detemir and 2.0 ± 0.1 h for a monomeric acylated insulin analog. Self-association of acylated insulin analogs with same albumin affinity in saline correlated with disappearance rate and addition of albumin to saline showed a combination of insulin detemir self association and albumin binding. Intravenous kinetic studies showed that the clearance and volume of distribution decreased with increasing albumin binding affinity of different acylated insulin analogs. Conclusions. The protracted action of detemir is primarily achieved through slow absorption into blood. Dihexamerization and albumin binding of hexameric and dimeric detemir prolongs residence time at the injection depot. Some further retention of detemir occurs in the circulation where albumin binding causes buffering of insulin concentration. Insulin detemir provides a novel principle of protraction, enabling increased predictability of basal insulin.

In vitro protein binding of liraglutide in human plasma determined by reiterated stepwise equilibrium dialysis.

Liraglutide is a human glucagon-like peptide-1 (GLP-1) analogue approved for the treatment of type 2 diabetes. It is based on human GLP-1 with the addition of a 16-carbon fatty acid, which facilitates binding to plasma proteins, thus prolonging the elimination half-life and allowing once-daily administration. It has not been possible to quantify liraglutide protein binding by ultrafiltration (the usual method of choice), as the lipophilic molecule becomes trapped in the filter membrane. Therefore, the aim of this study was to develop a methodology that could determine the extent of liraglutide binding to plasma proteins in vitro. We report here the details of a novel reiterated stepwise equilibrium dialysis assay that has successfully been used to quantify liraglutide plasma protein binding. The assay allowed quantification of liraglutide binding to proteins in purified plasma protein solutions and human plasma samples and was effective at plasma dilutions as low as 5%. At a clinically relevant liraglutide concentration (104 pM), greater than 98.9% of liraglutide was bound to protein. Specific binding to human serum albumin and α1-acid glycoprotein was 99.4% and 99.3%, respectively. The novel methodology described herein could have an application in the quantification of plasma protein binding of other highly lipophilic drug molecules.

Pharmacokinetic and pharmacodynamic properties of insulin aspart and human insulin

The preferred approach to determine the pharmacokinetic (PK) and pharmacodynamic (PD) properties of insulin analogues is the euglycemic glucose clamp. Currently, non-compartmental data analytical approaches are used to analyze data. The purpose of the present study is to propose a novel compartmental-model for analysis of data from glucose clamp studies. Data used in this trial only involved 18 of the 20 originally treated subjects. Data was obtained from a crossover trial where 18 healthy subjects each received a single subcutaneous (sc) dose of 1.2 nmol/kg (body weight) insulin aspart (IAsp) or 1.2 nmol/kg human insulin (HI) during a euglycemic glucose clamp after overnight fast. Serum insulin and glucose concentrations were measured and the glucose infusion rate (GIR) was adjusted after dosing, to maintain blood glucose near basal levels. Individual model parameters were estimated for IAsp, HI, and the corresponding glucose and GIR data. We found statistically significant differences between most of the HI and IAsp pharmacokinetic parameters, including the sigmoidicity of the time course of absorption (1.5 for HI vs. 2.1 for IAsp (unit less), P=0.0005, Wilcoxon Signed-rank test), elimination rate constant (0.010 min−1 for HI vs. 0.016 min−1 for IAsp (P=0.002)). The PD model parameters were mostly not different, except for the rate of insulin action (0.012 min−1 for HI vs. 0.017 min−1 for IAsp (P=0.03)). The model may provide a framework to account for different PK properties when estimating the PD properties of insulin and insulin analogues in glucose clamp experiments.

Kernel Function Smoothing of Insulin Absorption Kinetics

Drug absorption kinetics are usually described by simple descriptive statistics such as the time until 50% is absorbed or by parametric analysis, e.g., compartmental modelling. This paper describes how the absorption alternatively can be estimated nonparametrically by means of kernel function smoothing. This avoids the need for modelling the lag phase and it is still possible to estimate the absorption rate. The methods are applied to analyse the disappearance of radioactively labelled insulin from a subcutaneous depot. The size of the depot is measured by external counting. One application is the absorption of the fast-acting human insulin Actrapid, where the amount of insulin in blood is also predicted. Another application is the study of variation in absorption of a long-acting insulin analogue, Insulin 174, which was found to have significantly less variation than a comparable crystalline insulin suspension preparation.

Validity and use of a non-parallel insulin assay for pharmacokinetic studies of the rapid-acting insulin analogue, insulin aspart.

A radioimmunoassay (RIA) for insulin was validated for reliable measurement of the human insulin analogue, insulin aspart, by correction of non-linear measurements. Specificity was equivalent for several species of insulin, except insulin aspart. A non-linear hyperbolic model fitted insulin aspart with a correction formula for non-linearity of: z = 1503y/(1398 - y), where y denotes measured concentration and z denotes true concentration. Matrix-effects were insignificant for human, porcine, and canine heparin-plasma and for human and porcine serum. The coefficient of variation was below 15% for 80–800 pmol/L human and porcine insulin and for 80–600 pmol/L insulin aspart. The limit of detection for insulin aspart was 11.5 pmol/L with a lower limit of quantification of 17.5 pmol/L. Dilution of serum with Pharmacia dilution media introduced no significant error. In conclusion, this paper demonstrates that a non-parallel radioimmunoassay can be used to estimate accurate concentrations of insulin aspart.