In Vivo Receptor Occupancy

Overview:  Measures the degree to which the test drug actually occupies its receptor in vivo.  Receptor occupancy is determined by measuring the ability of a systemic dose of the test drug (non-radioactive) to compete with binding of a radiolabeled drug to the receptor.  The test is useful to determine the dose or plasma concentration of the test drug needed to reach a therapeutically effective occupancy level.

Protocol summary:  Administer (unlabeled) test drug to the animal (p.o./i.v./s.c.).  After a period of time for drug absorption (depending on administration route and pharmacokinetics) inject a standard radiotracer for the targeted receptor intravenously.  30 min later sacrifice animal.  Collect blood and dissect out brain and/or other tissues.  Weigh and digest tissues and count radioactivity.

Data analysis:  Radioactivity levels are determined in both a receptor-rich region and a reference, receptor-deficient, region and both converted to DPM/mg tissue.   Receptor occupancy of the test (non-radioactive) drug is determined from its ability to reduce the radioactivity in the receptor rich region relative to that in the reference (receptor deficient region).  Complete occupancy (100 %) would typically reduce this ratio to unity, i.e.  no difference in counts between the receptor-rich region and reference region. 

Suggested testing:  5 doses of the test drug plus a vehicle control and one or more doses of a positive control.  n = 6 - 8 animals per dose.

Examples of radiotracers for in vivo receptor occupancy measurements:

Custom Services:  The receptor-radiotracer combinations listed in the table above were selected on the basis of having been determined in the laboratories of the co-founders as giving receptor-specific binding signal sufficient to enable in vivo occupancy measurements by competing drugs in mice.  We will be happy to evaluate other radiotracer-receptor combinations upon request.

Ex vivo receptor occupancy determinations:  For drugs with high affinity to their targeted receptor, an estimate of receptor occupancy can instead be made in some cases using an ex vivo approach.  In this technique the unlabeled test drug is administered to the animal.  However, receptor occupancy is subsequently measured in vitro immediately following sacrifice of the animal using either a homogenate radioligand binding assay or with in vitro autoradiography.  A short non-equilibrium incubation time in the radiotracer is used to avoid appreciable dissociation of the unlabeled test drug before the termination of the assay.