Pharmacological characterization of a novel centrally permeable P2X7 receptor antagonist: JNJ‐47965567

Abstract

Background and Purpose An increasing body of evidence suggests that the purinergic receptor P2X, ligand‐gated ion channel, 7 (P2X7) in the CNS may play a key role in neuropsychiatry, neurodegeneration and chronic pain. In this study, we characterized JNJ‐47965567, a centrally permeable, high‐affinity, selective P2X7 antagonist. Experimental Approach We have used a combination of in vitro assays (calcium flux, radioligand binding, electrophysiology, IL‐1β release) in both recombinant and native systems. Target engagement of JNJ‐47965567 was demonstrated by ex vivo receptor binding autoradiography and in vivo blockade of Bz‐ATP induced IL‐1β release in the rat brain. Finally, the efficacy of JNJ‐47965567 was tested in standard models of depression, mania and neuropathic pain. Key Results JNJ‐47965567 is potent high affinity (pKi 7.9 ± 0.07), selective human P2X7 antagonist, with no significant observed speciation. In native systems, the potency of the compound to attenuate IL‐1β release was 6.7 ± 0.07 (human blood), 7.5 ± 0.07 (human monocytes) and 7.1 ± 0.1 (rat microglia). JNJ‐47965567 exhibited target engagement in rat brain, with a brain EC50 of 78 ± 19 ng·mL−1 (P2X7 receptor autoradiography) and functional block of Bz‐ATP induced IL‐1β release. JNJ‐47965567 (30 mg·kg−1) attenuated amphetamine‐induced hyperactivity and exhibited modest, yet significant efficacy in the rat model of neuropathic pain. No efficacy was observed in forced swim test. Conclusion and Implications JNJ‐47965567 is centrally permeable, high affinity P2X7 antagonist that can be used to probe the role of central P2X7 in rodent models of CNS pathophysiology.

Purinergic mechanisms in neuroinflammation: An update from molecules to behavior

“…To study the manic pole of bipolar disorder, genetic deficiency and pharmacological blockade of P2X7 receptors attenuated hyperactivity induced by amphetamine in all studies examined (Bhattacharya et al, 2013;Csolle et al, 2013a;Gubert et al, 2014;Lord et al, 2014). These findings indicate that the presence and endogenous activation of P2X7 receptors contributes to behavioral changes induced by either negative or positive challenges, and the blockade of the receptor alleviates these fluctuations.…”

Pharmacology of a Novel Central Nervous System–Penetrant P2X7 Antagonist JNJ-42253432

“…1321N1 cells expressing the rat P2X7 were grown in the same media supplemented with 10% fetal bovine serum and 100 mg/ml Zeocin (Invivogen, San Diego, CA). For isolation of primary astrocytes, rat brains were dissected from 3-day-old neonatal rats as described by Bhattacharya et al (2013).…”

“…Twenty-micron-thick tissue sections at the level of the hippocampus were prepared for autoradiography. P2X7 radioligand binding autoradiography was determined at room temperature with 30 nM [ 3 H]A-804598 in 50 mM Tris-HCl incubation buffer containing 0.1% bovine serum albumin as previously described (Bhattacharya et al, 2013). Nonspecific binding was measured with 100 mM A-740003 (N-[1-[[(cyanoamino)(5-quinolinylamino)methylene]amino]-2,2-dimethylpropyl]-3,4-dimethoxybenzeneacetamide).…”

“…Specifically, the P2X7 ion channel in the brain has attracted interest due to the genetic association with depression and bipolar disorder, coupled with emerging science on the role of the P2X7-IL-1b pathway in central neuroinflammation (Bennett, 2007;Sperlagh et al, 2012;Bhattacharya et al, 2013;Iwata et al, 2013). There is growing evidence that strengthens the role of P2X7 in mood disorders.…”

Preclinical Evaluation of a P2X7 Receptor–Selective Radiotracer: PET Studies in a Rat Model with Local Overexpression of the Human P2X7 Receptor and in Nonhuman Primates

“…In addition, several animal models for neurodegenerative disorders such as Alzheimer's disease and Huntington's disease (9)(10)(11) showed that proinflammatory cytokines correlated with upregulated P2X7R expression and increased adenosine triphosphate sensitivity, putting forward P2X7R as a valuable alternative marker for microglial activation and inflammation (8,12). In a recent study, 3 H-A-804598 was used to determine the ex vivo brain P2X7R occupancy of different P2X7R antagonists, indicating that this class of molecules crosses the blood-brain barrier and supporting the feasibility of in vivo visualization of P2X7R with PET in the brain (13)(14)(15). An additional hurdle, however, is the P2X7R interspecies difference reported for several molecules, giving rise to different binding affinities for mouse, rat, and human P2X7R (hP2X7R) (16)(17)(18).…”