In vivo & ex vivo receptor occupancy (RO) offers a simple yet powerful way to measure compound/target engagement using a selective radioligand and receptor autoradiography. Beta – imaging technology, developed by Biospace Lab, France, significantly shortens the time needed for assessment of specific binding in the brain (hours versus weeks for tritiated ligands).
Figure 1: (A) Percent receptor occupancy dose response curve for CTP – 354, a subtype – selective GABAA receptor modulator. Rats (n=5) were dosed PO at 30, 10, 3, 1, 0.3, and 0.1 mg/kg with the selective benzodiazepine receptor (GABAA) modulator CTP – 354 in 0.5% methylcellulose. One hour after CTP – 354 administration, animals received a tail vein injection of the selective benzodiazepine receptor (GABAA) antagonist 3H-Ro15-1788 (Flumazenil, 87 Ci/mmol) diluted 1:100 in saline. The amount injected was 1μl/g followed by decapitation 3 minutes post injection. Six (6) 40 μM coronal sections spanning Bregma – 2.2/- 6.4 were mounted on poly-lysine coated slides. Slides were loaded into the cassette of a Beta – imager 2000 (BioSpace Lab, France), and the images of the brain sections were acquired for 40 hours. Surface activity (cpm/mm2) was measured for all six sections using Beta vision + software (BioSpace Lab, France). (B) Percent receptor occupancy dose response curve for CTP – 354 with calculated ED50 of 2.8 mg/kg. Percent receptor occupancy was calculated as: [average vehicle (cpm/mm2)– average drug treatment (cpm/mm2))]/ average vehicle (cpm/mm2) X 100%.
Figure 2: Ex vivo receptor occupancy allows to establish correlation between occupancy at the receptor with both in vivo behavioral efficacy and drug exposure. High correspondence between behaviorally active doses of sazetidine in the forced swim test (2A) and α4β2 nicotinic acetylcholine receptor (nAChR) occupancy in the brain (2B). Specifically, sazetidine was inactive in the forced swim test at 0.3 mg/kg (i.p.) and showed very low receptor occupancy at this dose (<10%). In contrast, doses of sazetidine (1 and 3 mg/kg, i.p.) that produced robust behavioral activity in the forced swim test, corresponded with high levels of occupancy at the α4β2 nAChRs (~40% and ~90% respectively).
Data published in: Psychopharmacology (Berl). (2011) 217(2): 199-210.
Bioanalysis & Microdialysis