Correlation of pharmacodynamic activity, pharmacokinetics, and anti-product antibody responses to anti-IL-21R antibody therapeutics following IV administration to cynomolgus monkeys.

BACKGROUND: Anti-IL-21R antibodies are potential therapeutics for the treatment of autoimmune diseases. This study evaluated correlations between the pharmacodynamic (PD) activity, pharmacokinetics, and anti-product antibody responses of human anti-IL-21R antibodies Ab-01 and Ab-02 following IV administration to cynomolgus monkeys. METHODS: The PD assay was based on the ability of recombinant human IL-21 (rhuIL-21) to induce expression of the IL-2RA gene in cynomolgus monkey whole blood samples ex vivo. Monkeys screened for responsiveness to rhuIL-21 stimulation using the PD assay, were given a single 10 mg/kg IV dosage of Ab-01, Ab-02, or a control antibody (3/group), and blood samples were evaluated for PD activity (inhibition of IL-2RA expression) for up to 148 days. Anti-IL-21R antibody concentrations and anti-product antibody responses were measured in serum using immunoassays and flow cytometry. RESULTS: Following IV administration of Ab-01 and Ab-02 to cynomolgus monkeys, PD activity was observed as early as 5 minutes (first time point sampled). This PD activity had good correlation with the serum concentrations and anti-product antibody responses throughout the study. The mean terminal half-life (t1/2) was approximately 10.6 and 2.3 days for Ab-01 and Ab-02, respectively. PD activity was lost at approximately 5-13 weeks for Ab-01 and at approximately 2 weeks for Ab-02, when serum concentrations were relatively low. The estimated minimum concentrations needed to maintain PD activity were approximately 4-6 nM for Ab-01 and approximately 2.5 nM for Ab-02, and were consistent with the respective KD values for binding to human IL-21R. For Ab-01, there was noticeable inter-animal variability in t1/2 values (approximately 6-14 days) and the resulting PD profiles, which correlated with the onset of anti-product antibody formation. While all three Ab-01-dosed animals were positive for anti-Ab-01 antibodies, only one monkey (with the shortest t1/2 and the earliest loss of PD activity) had evidence of neutralizing anti-Ab-01 antibodies. All three Ab-02-dosed monkeys developed neutralizing anti-Ab-02 antibodies. CONCLUSIONS: For anti-IL-21R antibodies Ab-01 and Ab-02, there was good correlation between PD activity and PK profiles following IV administration to cynomolgus monkeys. Compared with Ab-01, Ab-02 was eliminated markedly faster from the circulation, which correlated with a shorter duration of PD activity.

In vitro potency, pharmacokinetic profiles, and pharmacological activity of optimized anti-IL-21R antibodies in a mouse model of lupus.

Using phage display, we generated a panel of optimized neutralizing antibodies against the human and mouse receptors for interleukin 21 (IL-21), a cytokine that is implicated in the pathogenesis of many types of autoimmune disease. Two antibodies, Ab-01 and Ab-02, which differed by only four amino acids in V(L) CDR3, showed potent inhibition of human and mouse IL-21R in cell-based assays and were evaluated for their pharmacological and pharmacodynamic properties. Ab-01, but not Ab-02, significantly reduced a biomarker of disease (anti-dsDNA antibodies) and IgG deposits in the kidney in the MRL-Fas(lpr) mouse model of lupus, suggesting that anti-IL-21R antibodies may prove useful in the treatment of lupus. Ab-01 also had a consistently higher exposure (AUC(0-infinity)) than Ab-02 following a single dose in rodents or cynomolgus monkeys (2-3-fold or 4-7-fold, respectively). Our data demonstrate that small differences in CDR3 sequences of optimized antibodies can lead to profound differences in in vitro and in vivo properties, including differences in pharmacological activity and pharmacokinetic profiles. The lack of persistent activity of Ab-02 in the MRL-Fas(lpr) mouse lupus model may have been a consequence of faster elimination, reduced potency in blocking the effects of mouse IL-21R, and more potent/earlier onset of the anti-product response relative to Ab-01.

Biodistribution of [125I]-labeled therapeutic proteins: application in protein drug development beyond oncology.

The majority of biodistribution studies of therapeutic proteins published to date focus on tumor-targeting agents. In this report we present a number of case studies that demonstrate the utility of biodistribution studies during preclinical development of biotherapeutics for non oncology indications, as well as provide a practical perspective on the methodology applied to these studies. For the commonly used classes of biologics (such as human monoclonal antibodies), biodistribution profiles may be compared to those of other therapeutics of the same class and compounds with unexpected off-target mediated uptake may be identified. Temporal biodistribution profiles may be used to address kinetics and reversibility of target- and/or off-target-mediated accumulation. In cases when circulating biotherapeutic is rapidly eliminated from circulation due to the formation of anti-product antibodies, tissue data may provide useful insight on test article exposure at the site of therapeutic action (or at the site of toxicity). Comparison of temporal biodistribution profiles between the genetically engineered and wild-type mouse strains or between the disease models and healthy animals may provide useful insight on sites and kinetics of target-mediated elimination. Finally, biodistribution studies will be a useful tool to study in vivo disposition for a variety of existing and upcoming novel classes of protein compounds.

Pharmacokinetic and pharmacodynamic modeling of a humanized anti-IL-13 antibody in naive and Ascaris-challenged cynomolgus monkeys.

PURPOSE: Neutralization of IL-13 is an attractive approach for treatment of asthma. In this report, we developed a novel PK-PD model that described the relationship between the circulating concentrations of total IL-13 and a neutralizing anti-IL-13 antibody (Ab-02) in the model of acute airway inflammation induced by Ascaris challenge to cynomolgus monkeys, as well as in naive monkeys. METHODS: Cynomolgus monkeys were administered a single intravenous or subcutaneous dose of Ab-02. Total IL-13 and Ab-02 concentrations were measured by immunoassays. RESULTS: Modeling and simulations indicated that: (1) Ascaris challenge induced approximately three-fold increase in circulating IL-13 concentrations, when compared to naive animals, consistent with the notion that Ascaris-induced airway inflammation was IL-13-mediated; (2) the transient increase in total IL-13 concentrations observed in both naive and Ascaris-challenged monkeys following Ab-02 administration was due to the increase in Ab-02-bound IL-13, while free IL-13 was decreased; and (3) the extent and duration of neutralization of circulating IL-13 were different in naive and Ascaris-challenged monkeys for the same Ab-02 dose regimen. CONCLUSIONS: The PK-PD model presented in this report may be applied to study drug-ligand interactions when a free ligand cannot be directly assayed but total ligand concentrations are modulated by the drug administration.

Preclinical pharmacokinetics, interspecies scaling, and tissue distribution of humanized monoclonal anti-IL-13 antibodies with different IL-13 neutralization mechanisms.

Numerous animal and in vitro studies suggest that neutralization of IL-13 is an attractive approach for therapeutic intervention in asthma. In this paper we describe preclinical pharmacokinetics (PK), interspecies scaling, and biodistribution of two humanized anti-IL-13 IgG1 monoclonal antibodies, Ab-01 and Ab-02, with different IL-13 neutralization mechanisms. PK parameters of Ab-01 and Ab-02 following IV or SC dosage to mouse, rat, cynomolgus monkey, and sheep, were similar. After IV administration, the elimination of anti-IL-13 antibodies was slow in all species tested and the serum clearance ranged from 0.13 mL/h/kg in monkeys to 0.81 mL/h/kg in mice. Both anti-IL-13 antibodies appeared to be confined primarily to the vascular space, as volume of distribution was relatively small (<120 mL/kg) in all species and tissue-to-serum concentration ratios (in mice and rats) were low (<0.5) in the tissues examined. The elimination half-life ranged from 3-6 days in mice to 14-17 days in monkey and sheep. In monkeys, PK parameters appeared to be approximately linear in the 1-100 mg/kg dose range. Following SC administration, the bioavailability of anti-IL-13 antibodies was 60-90% in all species tested. PK profile of Ab-02 in the model of acute airway inflammation (induced by Ascaris challenge) was, in general, similar to that in unchallenged monkeys; however, volume of distribution and clearance tended to decrease in Ascaris-challenged animals. Allometric scaling suggested that anti-IL-13 antibodies would likely to have a favorable PK profile, such as slow clearance and long terminal half-life, following IV or SC administration to humans.

Effect of von Willebrand factor on the pharmacokinetics of recombinant human platelet glycoprotein Ibalpha-immunoglobulin G1 chimeric proteins.

PURPOSE: Recombinant human platelet glycoprotein Ibalpha-immunoglobulin G1 chimeric proteins (GPIbalpha-Ig) have varying levels of anti-thrombotic activities based on their ability to compete for platelet mediated adhesion to von Willebrand Factor (vWF). Valine substituted GPIbalpha-Ig chimeras, at certain position, increase the binding affinity to vWF over its "wild-type" GPIbalpha-Ig analog. The purpose of this study was to determine the pharmacokinetics of two valine substituted GPIbalpha-Ig chimeras, GPIbalpha-Ig/1V (valine substitution at 239 position) and GPIbalpha-Ig/2V (double valine substitution at 233 and 239 position), in mice, rats and dogs. METHODS: Head-to-head comparisons of pharmacokinetics of GPIbalpha-Ig/1V and GPIbalpha-Ig/2V were investigated in rats and dogs after intravenous administration. Since vWF precipitates in the serum but not in plasma preparation, the concentration-time profiles of GPIbalpha-Ig/2V in rats were examined from the same blood samples for determination of matrix effect. The disposition of GPIbalpha-Ig/2V was also compared in vWF-deficient versus wild-type mice. RESULTS: For GPIbalpha-Ig/2V, the serum clearances were 2.62+/-0.27 ml/hr/kg in rats and 1.97+/-0.24 ml/hr/kg in dogs. The serum clearances of less potent GPIbalpha-Ig/1V were 1.08+/-0.08 and 0.97+/-0.19 ml/hr/kg in rats and dogs, respectively. In addition, the serum clearance of GPlbalpha-Ig/2V of 1.53 ml/hr/kg in vWF-deficient mice was lower than that in wild-type mice of 2.79 ml/hr/kg. CONCLUSION: The difference in disposition for valine substituted forms of GPIbalpha-Ig in laboratory animals are likely affected by their enhanced binding affinity for circulating vWF.