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A series of arylglycine-based analogs was synthesized and tested for TRPM8 antagonism in a cell-based functional assay. Following structure-activity relationship studies in vitro, a number of compounds were identified as potent TRPM8 antagonists and were subsequently evaluated in an in vivo pharmacodynamic assay of icilin-induced 'wet-dog' shaking in which compound 12 was fully effective. TRPM8 antagonists of the type described here may be useful in treating pain conditions wherein cold hypersensitivity is a dominant feature.

Intracerebroventricular (i.c.v.) administration to mice of delta9-tetrahydrocannabinol (delta9-THC), WIN 55,212-2 or the endogenous cannabinoid anandamide induced dose-related antinociception in the 55 degrees C warm-water tail-flick test. Pretreatment (24 h, i.c.v.) with pertussis toxin dose-dependently reduced the antinociceptive effect of delta9-THC (955 nmol), WIN 55,212-2 (30 nmol) and anandamide (135 nmol) (IC50 = 0.13, 5.5, and 0.32 nmol, respectively). In contrast, pretreatment (24 h, i.c.v.) with cholera toxin (0.1-3.0 mg) reduced the antinociception of WIN 55,212-2, had minimal effect on delta9-THC, and dose-dependently increased the antinociception of anandamide (ED50 = 0.50 nmol). These data suggest differences in the receptor-effector coupling of delta9-THC, WIN 55,212-2 and anandamide in supraspinal-induced antinociception in mice.

In vitro studies have demonstrated that FMRFamide-related peptide receptors can be coupled to different G-proteins, mediating opposite stimulatory and inhibitory effects. The present study tested whether this duality might extend to effects in vivo. Antinociception in mice of ICV [ d -Met 2 ]FMRFamide, which produced agonist [ED 50 = 36.3 μg (61.6 nmol)] and antagonist [ID 50 = 0.72 μg (1.22 nmol)] actions, was attenuated by 24-h pretreatment with ICV pertussis toxin (ID 50 = 0.55 μg) or cholera toxin (ID 50 = 0.09 μg), suggesting that [ d -Met 2 ]FMRFamide in vivo effects might also be explained by dual G i G s coupling.

A series of benzothiophene-based phosphonates was synthesized and many analogs within the series were shown to be potent antagonists of the TRPM8 channel. The compounds were obtained as a racemic mixture in 5 synthetic steps, and were tested for TRPM8 antagonist activity in a recombinant, canine TRPM8-expressing cell line using a fluorometric imaging plate reader (FLIPR) assay. Structure–activity relationships were developed initially by modification of the core structure and subsequently by variation of the aromatic substituents and the phosphonate ester. Compound 9l was administered intraperitoneally to rats and demonstrated engagement of the TRPM8 target in both prevention and reversal-modes in an icilin-induced ‘wet-dog’ shake model.

Discovered as part of an effort to identify delta opioid (DOPr or DOR) agonist analgesics, JNJ-20788560 and JNJ-39204880 exhibited high DOR affinity, with K(i) values of 1.7 and 2.0nM, respectively, and were selective for DOR over the mu opioid receptor (MOPr or MOR), with 596- and 122-fold selectivity, respectively. Both compounds stimulated DOR but not MOR induced GTPgammaS binding and were effective antihyperalgesic agents in the complete Freund's adjuvant model of thermal hyperalgesia in the rat, with oral ED(50) values of 13.5 and 35mg/kg, corresponding to plasma levels of 1 and 9microM, respectively. Autoradiographic analysis of DOR and MOR occupancy in sections of brain (striatum) and lumbar spinal cord (L4-L6) was determined ex vivo, using radiolabeled naltrindole or DAMGO. Quantitative image analysis resulted in striatal DOR ED(50) values of 6.9 and 10.7mg/kg, for JNJ-20788560 and JNJ-39204880 respectively, and spinal cord values of 6.4 and 3.2mg/kg, respectively. Neither compound dose-dependently occupied MOR within the dose range studied. Thus, this study confirmed the DOR selectively over MOR of both compounds following their oral administration, and further demonstrated dose-dependent DOR occupancy by each compound across its antihyperalgesic dose range. Importantly, these in vitro, in vivo, and ex vivo data revealed that the greater in vitro potency of JNJ-20788560 was paralleled by its greater in vivo potency, although JNJ-39204880 achieved higher plasma levels following its oral administration. The receptor occupancy levels observed at the pharmacologic ED(50) doses of these compounds suggest the need for greater target engagement by JNJ-39204880 than by JNJ-20788560 to elicit a similar therapeutic response.

Two parallel synthetic methods were developed to explore the structure-activity relationships (SAR) of a series of potent opioid agonists. This series of tropanylidene benzamides proved extremely tolerant of structural variation while maintaining excellent opioid activity. Evaluation of several representative compounds from this series in the mouse hot plate test revealed potent antinociceptive effects upon oral administration.

The design and synthesis of novel pyrrolidine-containing bradykinin antagonists, II, are described. Conformational analysis suggested that a pyrrolidine moiety could substitute for the N-methyl cis-amide moiety of FR 173657. The in vitro binding data showed that the (S)-isomer of II was potent in the bradykinin B2 receptor-binding assay with a Ki of 33 nM. The opposite isomer, (R)-II, had a Ki of 46 nM. The in vitro binding data confirmed our conformational hypothesis.

The synthesis and structure–activity relationships of a novel series of aroylpyrrole alkylamides as potent selective bradykinin B2 receptor antagonists are described. Several members of this series display nanomolar affinity at the B2 receptor and show activity in an animal model of antinociception.

FMRFamide (Phe-Met-Arg-Phe-NH 2 ) and several analogs produce centrally-mediated, naloxone-reversible antinociception, but have minimal affinity for opioid receptor (sub)types. In the present study, the antinociception in mice (55°C tail-flick test) produced by supraspinal (intracerebroventricular; ICV) administration of [ d -Met 2 ]-FMRFamide (a stable analog of FMRFamide) was attenuated by pretreatment with ICV oligodeoxyribonucleotide antisense to the opioid μ receptor or by antisense to the G i 2α G-protein subunit. These data suggest that [ d -Met 2 ]-FMRFamide produces its antinociception via an opioid interneuron.