A Novel In Vivo Receptor Occupancy Methodology for the Glucocorticoid Receptor: Toward An Improved Understanding of Lung Pharmacokinetic/Pharmacodynamic Relationships

Abstract

Investigation of pharmacokinetic/pharmacodynamic (PK/PD) relationships for inhaled drugs is challenging because of the limited possibilities of measuring tissue exposure and target engagement in the lung. The aim of this study was to develop a methodology for measuring receptor occupancy in vivo in the rat for the glucocorticoid receptor (GR) to allow more informative inhalation PK/PD studies. From AstraZeneca's chemical library ofpropoxy]indazol-1-yl]-N-methyl-benzamide] was identified to have properties that are useful as a tracer for GR in vitro. When given at an appropriate dose (30 nmol/kg) to rats, compound 1 functioned as a tracer in the lung and spleen in vivo using liquid chromatography-tandem mass spectrometry bioanalysis. The methodology was successfully used to show the dose-receptor occupancy relationship measured at 1.5 hours after intravenous administration of fluticasone propionate (20, 150, and 750 nmol/kg) as well as to characterize the time profile for receptor occupancy after a dose of 90 nmol/kg i.v. The dose giving 50% occupancy was estimated as 47 nmol/kg. The methodology is novel in terms of measuring occupancy strictly in vivo and by using an unlabeled tracer. This feature confers key advantages, including occupancy estimation not being influenced by drug particle dissolution or binding/dissociation taking place postmortem. In addition, the tracer may be labeled for use in positron emission tomography imaging, thus enabling occupancy estimation in humans as a translatable biomarker of target engagement.

Revealing the Regional Localization and Differential Lung Retention of Inhaled Compounds by Mass Spectrometry Imaging

“…Rats were anesthetized at the time of sampling [time points, 2 and 30 minutes, were chosen based on a previously published study (5) ] with isoflurane (FORENE Ò ; Abbott Scandinavia AB, Solna, Sweden), and a terminal blood sample was collected as previously described (23) to isolate plasma and stored at -20°C until analysis. The heart and lungs were removed from the thorax en bloc.…”

Physiologically Based Pharmacokinetic/Pharmacodynamic Modeling Accurately Predicts the Better Bronchodilatory Effect of Inhaled Versus Oral Salbutamol Dosage Forms

“…In fact, there are no experimental methodologies available to do this, although indirect approaches such as target occupancy have been employed. (3,5) The inability to define the target site concentration(s) of inhaled drug even in preclinical species clearly poses a huge challenge for developing inhaled drug treatments with the objective of optimizing drug potency and targeting of relevant lung structures. Currently used methodologies to evaluate inhaled drug pharmacology and predicting therapeutic dosing in man seem to still largely depend on doseresponse studies in preclinical species with functional readouts at the level of the lung as a whole.…”

Glucocorticoid-independent modulation of GR activity: Implications for immunotherapy

“…Information about serum concentrations of GCs and receptor occupancy in relevant target cells for different doses should be useful in predicting the lowest doses required for efficacy with minimal side-effects, but such information is not readily available in the literature. However, a recent study in mice showed no correlation between receptor occupancy and serum GC concentrations (Boger et al, 2015). This study also established that a GC dose of 47nmol/kg was required to occupy 50% of receptor and that more of the receptor was occupied at higher doses.…”