Determining pharmacological selectivity of the kappa opioid receptor antagonist LY2456302 using pupillometry as a translational biomarker in rat and human

Abstract

Background: Selective kappa opioid receptor antagonism is a promising experimental strategy for the treatment of depression. The kappa opioid receptor antagonist, LY2456302, exhibits ~30-fold higher affinity for kappa opioid receptors over mu opioid receptors, which is the next closest identified pharmacology.

Methods: Here, we determined kappa opioid receptor pharmacological selectivity of LY2456302 by assessing mu opioid receptor antagonism using translational pupillometry in rats and humans.

Results: In rats, morphine-induced mydriasis was completely blocked by the nonselective opioid receptor antagonist naloxone (3mg/kg, which produced 90% mu opioid receptor occupancy), while 100 and 300 mg/kg LY2456302 (which produced 56% and 87% mu opioid receptor occupancy, respectively) only partially blocked morphine-induced mydriasis. In humans, fentanyl-induced miosis was completely blocked by 50mg naltrexone, and LY2456302 dose-dependently blocked miosis at 25 and 60 mg (minimal-to-no blockade at 4-10mg).

Conclusions: We demonstrate, for the first time, the use of translational pupillometry in the context of receptor occupancy to identify a clinical dose of LY2456302 achieving maximal kappa opioid receptor occupancy without evidence of significant mu receptor antagonism.

Keywords: LY2456302; dynorphin; kappa opioid receptor; pupillometry; translational biomarker.

Figure 1.

Dose-dependent in vivo receptor occupancy at putative mu, kappa, and delta opioid receptors (MOR, KOR, and DOR, respectively) by naloxone and LY2456302 60 minutes after administration. a, Naloxone showed dose- and concentration-dependent occupancy of putative opioid receptors in a manner consistent with its in vitro binding affinity, with MOR and KOR fully saturated at higher doses (3–10mg/kg). The doses at which 50% of receptors were occupied (ED₅₀) at MOR, KOR, and DOR were 0.49, 0.75, and 3.45mg/kg, respectively. b, LY2456302 saturated putative KOR at all doses tested (3–300mg/kg). At higher doses, putative MOR and DOR occupancies were observed (ED₅₀ values = 84.4 and 214.6mg/kg, respectively). LY2456302 selectively occupies KOR in the rat at doses <100mg/kg PO. Some engagement of MOR and DOR is evident at 100 and 300mg/kg (corresponding to brain exposures >473ng/g; see supplemental Table S1). Chemical structures of naloxone and LY2456302 are shown in insets.

Figure 2.

Effect of naloxone (NLX) and LY2456302 (LY) on morphine-induced mydriasis in the rat and naltrexone (NTX) and LY2456302 on fentanyl-induced miosis in healthy human subjects. a, Naloxone (3mg/kg subcutaneously) completely blocked morphine-induced mydriasis in the rat, indicating full blockade of mu opioid receptor (MOR). b, LY2456302 produced modest, but significant, blockade of morphine-induced mydriasis in the rat at the highest doses tested (100 and 300mg/kg orally), consistent with 56% and 87% MOR occupancy, respectively. At lower doses, LY2456302 showed no evidence of MOR blockade. c, Naltrexone (50mg) completely blocked fentanyl-induced miosis in healthy human subjects. d, LY2456302 produced dose- and concentration-dependent blockade of fentanyl-induced miosis in healthy subjects, with statistically significant effects at 25 and 60mg. Doses of 4 and 10mg LY2456302 did not significantly affect fentanyl-induced miosis. Data in each panel represent the mean (±SEM) pupil diameter, expressed as a percent of baseline pupil diameter. See supplementary Table S4 for raw pupil diameter values (in millimeters).

Figure 3.

Statistical analysis of the least squares geometric means of the area under (or above) the effect curve (AUEC) of pupil diameter in rats and healthy subjects. Mean estimates of AUEC ratio of drug: placebo across dose levels showed a dose-response relationship in rats (a) and healthy subjects (b). Dotted line indicates a ratio of 1, or no effect on fentanyl-induced mydriasis or miosis, respectively.

Figure 4.

Maximum change in pupil diameter ratio. a, Maximum change in pupil diameter (MaxCPD) ratio in rats (drug:vehicle) is inversely correlated with LY2456302 brain exposure, indicating that, at higher exposures, LY2456302 reduces the magnitude of morphine-induced mydriasis observed. b, MaxCPD ratio in humans (treatment:placebo) is inversely related to LY2456302 exposure and described by the E_max (the estimated maximal effect on MaxCPD ratio) model (E₀ [the baseline ratio]=1.00 [fixed]; E_max=−0.918 [36.1%]; EC₅₀ [the AUC that produces 50% of the E_max]=1270 [72.8%]).