Shuichiro Kakimoto

Amelioration of Neuropathic Pain by Novel Transient Receptor Potential Vanilloid 1 Antagonist AS1928370 in Rats without Hyperthermic Effect

Transient receptor potential vanilloid 1 (TRPV1) is activated by a variety of stimulations, such as endogenous ligands and low pH, and is believed to play a role in pain transmission. TRPV1 antagonists have been reported to be effective in several animal pain models; however, some compounds induce hyperthermia in animals and humans. We discovered the novel TRPV1 antagonist (R)-N-(1-methyl-2-oxo-1,2,3,4-tetrahydro-7-quinolyl)-2-[(2-methylpyrrolidin-1-yl)methyl]biphenyl-4-carboxamide (AS1928370) in our laboratory. AS1928370 bound to the resiniferatoxin-binding site on TRPV1 and inhibited capsaicin-mediated inward currents with an IC50 value of 32.5 nM. Although AS1928370 inhibited the capsaicin-induced Ca2+ flux in human and rat TRPV1-expressing cells, the inhibitory effect on proton-induced Ca2+ flux was extremely small. In addition, AS1928370 showed no inhibitory effects on transient receptor potential vanilloid 4, transient receptor potential ankyrin 1, and transient receptor potential melastatin 8 in concentrations up to 10 μM. AS1928370 improved capsaicin-induced secondary hyperalgesia and mechanical allodynia in an L5/L6 spinal nerve ligation model in rats with respective ED50 values of 0.17 and 0.26 mg/kg p.o. Furthermore, AS1928370 alleviated inflammatory pain in a complete Freunds adjuvant model at 10 mg/kg p.o. AS1928370 had no effect on rectal body temperature up to 10 mg/kg p.o., although a significant hypothermic effect was noted at 30 mg/kg p.o. In addition, AS1928370 showed no significant effect on motor coordination. These results suggest that blockage of the TRPV1 receptor without affecting the proton-mediated TRPV1 activation is a promising approach to treating neuropathic pain because of the potential wide safety margin against hyperthermic effects. As such, compounds such as ASP1928370 may have potential as new analgesic agents for treating neuropathic pain.

Minodronic acid, a third-generation bisphosphonate, antagonizes purinergic P2X2/3 receptor function and exerts an analgesic effect in pain models

The P2X(2/3) receptor has an important role in the nociceptive transmission. Minodronic acid is a third third-generation bisphosphonate and a potent inhibitor of bone resorption. We found that minodronic acid inhibited alpha,beta-methylene ATP-induced cation uptake with the potency higher than that of suramin in the P2X(2/3) receptor receptor-expressing cells. Other bisphosphonates did not show such activity. Subcutaneously administered (10-50 mg/kg) minodronic acid significantly inhibited the alpha,beta-methylene ATP-, acetic acid- and formalin-induced nociceptive behaviors in mice. These unique effects of minodronic acid would be beneficial for the treatment of accelerated bone turnover diseases accompanied by bone pain, including bone metastases.

AS1069562, the (+)-Isomer of Indeloxazine, Exerts Analgesic Effects in a Rat Model of Neuropathic Pain with Unique Characteristics in Spinal Monoamine Turnover

AS1069562 [(R)-2-[(1H-inden-7-yloxy)methyl]morpholine monobenzenesulfonate] is the (+)-isomer of indeloxazine, which had been used clinically for the treatment of cerebrovascular diseases with multiple pharmacological actions, including serotonin (5-HT) and norepinephrine (NE) reuptake inhibition. Here we investigated the analgesic effects of AS1069562 in a rat model of chronic constriction injury (CCI)–induced neuropathic pain and the spinal monoamine turnover. These effects were compared with those of the antidepressants duloxetine and amitriptyline. AS1069562 significantly elevated extracellular 5-HT and NE levels in the rat spinal dorsal horn, although its 5-HT and NE reuptake inhibition was much weaker than that of duloxetine in vitro. In addition, AS1069562 increased the ratio of the contents of both 5-HT and NE to their metabolites in rat spinal cord, whereas duloxetine slightly increased only the ratio of the content of 5-HT to its metabolite. In CCI rats, AS1069562 and duloxetine significantly ameliorated mechanical allodynia, whereas amitriptyline did not. AS1069562 and amitriptyline significantly ameliorated thermal hyperalgesia, and duloxetine tended to ameliorate it. Furthermore, AS1069562, duloxetine, and amitriptyline significantly improved spontaneous pain–associated behavior. In a gastric emptying study, AS1069562 affected gastric emptying at the same dose that exerted analgesia in CCI rats. On the other hand, duloxetine and amitriptyline significantly reduced gastric emptying at lower doses than those that exerted analgesic effects. These results indicate that AS1069562 broadly improved various types of neuropathic pain–related behavior in CCI rats with unique characteristics in spinal monoamine turnover, suggesting that AS1069562 may have potential as a treatment option for patients with neuropathic pain, with a different profile from currently available antidepressants.

Spontaneous and evoked pain-associated behaviors in a rat model of neuropathic pain respond differently to drugs with different mechanisms of action

Given that patients with neuropathic pain suffer a mixture of spontaneous and evoked pain symptoms, we assessed the effects of drugs with different mechanism of action on spontaneous and evoked pain-associated behaviors in a rat model of neuropathic pain induced by chronic constriction injury (CCI) of the sciatic nerve. Frequent aberrant limb movement on the operated side was measured to assess spontaneous pain-associated behavior, and mechanical allodynia and thermal hyperalgesia were evaluated to assess evoked pain-associated behaviors. These three types of behavior were assessed after administration of the following drugs: pregabalin (α2δ-subunit ligand), morphine (μ-opioid receptor agonist), perampanel (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid [AMPA] receptor antagonist), clonidine, dexmedetomidine (α2-adrenoceptor agonists), and diclofenac (non-steroidal anti-inflammatory drug [NSAID]). Pregabalin at an oral dose of 10 or 30mg/kg significantly alleviated frequent aberrant limb movement and mechanical allodynia, but not thermal hyperalgesia. Morphine at a subcutaneous dose of 1 or 3mg/kg significantly improved all three types of behavior. Perampanel at an oral dose of 1mg/kg attenuated only frequent aberrant limb movement. Intraperitoneal administration of clonidine (0.01 or 0.03mg/kg) and dexmedetomidine (0.03mg/kg) significantly improved all three types of behavior, while diclofenac did not relieve any of the behaviors. Pregabalin, clonidine, and dexmedetomidine significantly decreased motor performance at doses close to analgesic doses in the rotarod test. The present study demonstrates that responses to spontaneous and evoked pain symptoms in neuropathic pain condition differ depending on a drugs mechanism of action. The selection and application of drugs according to the specific symptoms would be considered for the medication of patients with neuropathic pain.

Discovery of an 8-methoxytetrahydroisoquinoline derivative as an orally active N-type calcium channel blocker for neuropathic pain without CYP inhibition liability

In lead optimization efforts starting from the tetrahydroisoquinoline (S)-1, we identified 2-{[(2R)-2-hydroxypropyl]amino}-1-[(1S)-8-methoxy-1-phenyl-3,4-dihydroisoquinolin-2(1H)-yl]ethanone ((1S)-8t) as a novel orally active small-molecule N-type calcium channel blocker without CYP inhibition liability. CYP3A4 inhibition profile was improved by reducing the lipophilicity of compound (S)-1. Moreover, introduction of a methoxy group to the C-8 position of tetrahydroisoquinoline led to identification of (1S)-8t, which eliminated CYP2D6 inhibition liability. Oral administration of (1S)-8t exerted efficacy in a rat spinal nerve ligation (SNL) model of neuropathic pain with an ED50 value of 2.8 mg/kg.

Purinergic P2X receptor activation induces emetic responses in ferrets and Suncus murinus (house musk shrews)

Despite the rapid progress made in understanding the significant role played by signalling via extracellular ATP in physiology and pathology, there has been no clear information generated on its involvement in the emetic response.

Discovery of a 1-isopropyltetrahydroisoquinoline derivative as an orally active N-type calcium channel blocker for neuropathic pain.

N-type calcium channel blockade is a promising therapeutic approach for the treatment of neuropathic pain. Starting from lead compound (S)-1, we focused our optimization efforts on potency for N-type calcium channel inhibition and improvement of CYP inhibition profile. 2-{[(1-Hydroxycyclohexyl)methyl]amino}-(1R)-(1-isopropyl-6-methoxy-3,4-dihydroisoquinolin-2(1H)-yl)ethanone oxalate ((R)-5r) was identified as a novel orally active small-molecule N-type calcium channel inhibitor with reduced CYP inhibition liability. Oral administration of (R)-5r improved mechanical allodynia in a spinal nerve ligation model of neuropathic pain in rats with an ED50 value of 2.5 mg/kg.

Discovery of novel tetrahydroisoquinoline derivatives as orally active N-type calcium channel blockers with high selectivity for hERG potassium channels

N-type calcium channels represent a promising target for the treatment of neuropathic pain. The selective N-type calcium channel blocker ziconotide ameliorates severe chronic pain but has a narrow therapeutic window and requires intrathecal administration. We identified tetrahydroisoquinoline derivative 1a as a novel potent N-type calcium channel blocker. However, this compound also exhibited potent inhibitory activity against hERG channels. Structural optimizations led to identification of (1S)-(1-cyclohexyl-3,4-dihydroisoquinolin-2(1H)-yl)-2-{[(1-hydroxycyclohexyl)methyl]amino}ethanone ((S)-1h), which exhibited high selectivity for hERG channels while retaining potency for N-type calcium channel inhibition. (S)-1h went on to demonstrate in vivo efficacy as an orally available N-type calcium channel blocker in a rat spinal nerve ligation model of neuropathic pain.

Assessment of canine sensory function by using sine-wave electrical stimuli paradigm

The paradigm of sine-wave electrical stimuli has been used for sensory neurological assessment in humans. In the present study, we applied the paradigm to the dog for the quantitative assessment of sensory function. Sine-wave electrical current stimuli at frequencies of 2000, 250, and 5Hz were delivered to bipolar electrodes attached to the skin surface of the hind paws. The stimulation intensity was gradually increased, and the minimum intensity required to elicit the lifting behavior in the stimulated paw was determined as current threshold (CT) for each of the three frequencies. Dogs consistently showed the lifting behavior at CTs without showing aversive behaviors such as vocalization and wriggling. The baseline CTs (mean+/-SEM, n=12) were 4430+/-110microA for CT2000, 2215+/-173microA for CT250, and 2305+/-152microA for CT5. The CTs immediately increased after bolus intravenous injection of fentanyl at 10microg/kg, although the significant increase disappeared within 1h. The time course for the CTs was parallel to that of plasma fentanyl concentration. In conclusion, the present study applied the paradigm of transcutaneous sine-wave electrical stimuli to the dog, and used the hind paw lifting as endpoint behavior. This paradigm is simple, non-invasive, useful in the assessment of sensory function, and can be adapted to investigate the pharmacokinetics/pharmacodynamics relation of drugs. Further studies are needed to give the conclusive interpretation of the endpoint behavior.

Analgesic effects of ASP3662, a novel 11β‐hydroxysteroid dehydrogenase 1 inhibitor, in rat models of neuropathic and dysfunctional pain

Glucocorticoids are a major class of stress hormones known to participate in stress‐induced hyperalgesia. Although 11β‐hydroxysteroid dehydrogenase 1 (11β‐HSD1) is a key enzyme in the intracellular regeneration of glucocorticoids in the CNS, its role in pain perception has not been assessed. Here, we examined the effects of ASP3662, a novel 11β‐HSD1 inhibitor, on neuropathic and dysfunctional pain.