The neuropeptidase glutamate carboxypeptidase II (GCPII) hydrolyzes N-acetyl-L-aspartyl-L-glutamate (NAAG) to liberate N-acetylaspartate and glutamate. GCPII was originally cloned as PSMA, an M(r) 100,000 type II transmembrane glycoprotein highly expressed in prostate tissues. PSMA/GCPII is located on the short arm of chromosome 11 and functions as both a folate hydrolase and a neuropeptidase. Inhibition of brain GCPII may have therapeutic potential in the treatment of certain disease states arising from pathologically overactivated glutamate receptors. Recently, we reported that certain urea-based structures act as potent inhibitors of GCPII (J. Med. Chem. 2001, 44, 298). However, many of the potent GCPII inhibitors prepared to date are highly polar compounds and therefore do not readily penetrate the blood-brain barrier. Herein, we elaborate on the synthesis of a series of potent, urea-based GCPII inhibitors from the lead compound 3 and provide assay data for these ligands against human GCPII. Moreover, we provide data revealing the ability of one of these compounds, namely, 8d, to reduce the perception of inflammatory pain. Within the present series, the gamma-tetrazole bearing glutamate isostere 7d is the most potent inhibitor with a K(i) of 0.9 nM. The biological evaluation of these compounds revealed that the active site of GCPII likely comprises two regions, namely, the pharmacophore subpocket and the nonpharmacophore subpocket. The pharmacophore subpocket is very sensitive to structural changes, and thus, it appears important to keep one of the glutamic acid moieties intact to maintain the potency of the GCPII inhibitors. The site encompassing the nonpharmacophore subpocket that binds to glutamate's alpha-carboxyl group is sensitive to structural change, as shown by compounds 6b and 7b. However, the other region of the nonpharmacophore subpocket can accommodate both hydrophobic and hydrophilic groups. Thus, an aromatic ring can be introduced to the inhibitor, as in 8b and 8d, thereby increasing its hydrophobicity and thus potentially its ability to cross the blood-brain barrier. Intrathecally administered 8d significantly reduced pain perception in the formalin model of rat sensory nerve injury. A maximal dose of morphine (10 mg) applied in the same experimental paradigm provided no significant increase in analgesia in comparison to 8d during phase 1 of this pain study and modestly greater analgesia than 8d in phase 2. These urea-based inhibitors of GCPII thus offer a novel approach to pain management.

The peptide neurotransmitter N-acetylaspartylglutamate (NAAG) acts as an agonist at group II metabotropic glutamate receptors (mGluRs). NAAG is inactivated by extracellular peptidase activity yielding glutamate and N-acetylaspartate. We recently developed a series of potent NAAG peptidase inhibitors, including ZJ-11, ZJ-17 and ZJ-43. In the present study, we examined the effects of intrathecally administered ZJ-11 and ZJ-17 and intravenously administered ZJ-11 and ZJ-43 in the rat formalin test (an inflammatory pain model) and in the rat partial sciatic nerve ligation model (a neuropathic pain model). Intrathecal injection of ZJ-11 or ZJ-17 or intravenous injection of ZJ-11 or ZJ-43 suppressed both phases of the agitation behaviour induced by paw formalin injection. Intrathecal and intravenous injection of ZJ-11 suppressed the expression of Fos-like immunoreactivity, induced by paw formalin injection, in laminae I-II in segments L4-L5 of the spinal cord, suggesting an action on sensory spinal transmission. Partial sciatic nerve ligation induced significant mechanical allodynia 7 days after the nerve injury. Intrathecal injection of ZJ-11 or ZJ-17 or intravenous administration of ZJ-11 or ZJ-43 attenuated the level of mechanical allodynia induced by this nerve ligation. These effects of intrathecally or intravenously administered ZJ compounds in both the formalin test and the partial sciatic nerve ligation model were completely antagonized by pretreatment with LY-341495, a highly selective group II mGluR antagonist. Thus, elevation of extracellular NAAG, induced by the inhibition of NAAG peptidase, activates group II mGluRs and produces an analgesic effect in neuropathic and inflammatory and pain models. In contrast, peptidase inhibition did not affect the threshold for withdrawal from a noxious mechanical stimulus or from an acute thermal stimulus in the hotplate test.

Recent genetic and pharmacological studies have suggested that the metabotropic glutamate receptor subtype 5 (mGluR5) may represent a druggable target in identifying new therapeutics for the treatment of various central nervous system disorders including drug abuse. In particular, considerable attention in the mGluR5 field has been devoted to identifying ligands that bind to the allosteric modulatory site, distinct from the site for the primary agonist glutamate. Both 2-methyl-6-(phenylethynyl)pyridine (MPEP) and its analogue 3-[(2-methyl-4-thiazolyl)ethynyl]pyridine (MTEP) have been shown to be selective and potent noncompetitive antagonists of mGluR5. Because of results presented in this study showing that MTEP prevents the reinstatement of cocaine self-administration caused by the presentation of environmental cues previously associated with cocaine availability, we have prepared a series of analogues of MTEP with the aim of gaining a better understanding of the structural features relevant to its antagonist potency and with the ultimate aim of investigating the effects of such compounds in blunting the self-administration of cocaine. These efforts have led to the identification of compounds showing higher potency as mGluR5 antagonists than either MPEP or MTEP. Two compounds 19 and 59 exhibited functional activity as mGluR5 antagonists that are 490 and 230 times, respectively, better than that of MTEP.