5-HT1D receptor (Kis = 6 nM)
The primary target of PNU-142633 is the 5-hydroxytryptamine 1D (5-HT1D) receptor, a G protein-coupled receptor that is a member of the serotonin receptor family. PNU-142633 acts as a high-affinity agonist at the human 5-HT1D receptor, with a Ki of 6 nM. It shows remarkable selectivity for the 5-HT1D receptor over the closely related 5-HT1B receptor, with a Ki of >18,000 nM at the human 5-HT1B receptor. This selectivity profile makes it a valuable tool for studying 5-HT1D-specific functions.

Sumatriptan's intrinsic activity was found to be 84%, while PNU-142633's at the human 5-HT1D receptor was shown to be 70% intrinsic activity in a cell sensor cell-based test [1].

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In vitro, PNU-142633 demonstrates high-affinity binding and agonist activity at the 5-HT1D receptor. It binds to the human 5-HT1D receptor with a Ki of 6 nM. In contrast, its affinity for the human 5-HT1B receptor is significantly lower, with a Ki of >18,000 nM. This high selectivity for 5-HT1D over 5-HT1B is a key feature of the compound's pharmacological profile. The compound's agonist activity at 5-HT1D is expected to inhibit neurotransmitter release and produce vasoconstriction.

Neurogenic plasma protein extravasation was stopped in male Hartley guinea pigs treated intravenously with PNU-142633 (0.03-1 mg/kg). It is not possible for PNU-142633 to alter these vascular beds' resistance [1].

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In this randomized, double-blind, placebo-controlled, parallel-group study, patients received a single 50-mg oral dose of a 5-HT(1D) agonist, PNU-142633 (n = 34), or matching placebo (n = 35) during an acute migraine attack. No statistically significant treatment effects were observed at 1 and 2 h after dosing, even after stratifying by baseline headache intensity. At 1 and 2 h post-dose, 8.8% and 29.4% of the PNU-142633 group, respectively, and 8.6% and 40.0% of the placebo group, respectively, experienced headache relief; 2.9% and 8.8% of the PNU-142633 group and 0% and 5.7% of the placebo group were free of headache pain. Adverse events associated with PNU-142633 treatment included chest pain (two patients) and QTc prolongation (three patients). Results from this study suggest that anti-migraine efficacy is not mediated solely through the 5-HT(1D) receptor subtype, although this receptor may contribute, at least in part, to the adverse cardiovascular effects observed with 5-HT agonist medications[2].

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In vivo, PNU-142633 has demonstrated anti-migraine efficacy. It blocks neurogenic plasma protein extravasation and inhibits sympathetically evoked tachycardia responses in vivo. These effects are consistent with the activation of 5-HT1D receptors, which are known to play a role in migraine pathophysiology. However, the compound was discontinued in Phase 2 clinical trials for the treatment of migraine. Detailed in vivo efficacy data are not extensively reported.

The present study describes the preclinical pharmacology of a highly selective 5-HT1D receptor agonist PNU-142633. PNU-142633 binds with a Ki of 6 nm at the human 5-HT1D receptor and a Ki of> 18 000 nm at the human 5-HT1B receptor. The intrinsic activity of PNU-142633 at the human 5-HT1D receptor was determined to be 70% that of 5-HT in a cytosensor cell-based assay compared with 84% for that of sumatriptan. PNU-142633 was equally effective as sumatriptan and a half-log more potent than sumatriptan in preventing plasma protein extravasation induced by electrical stimulation of the trigeminal ganglion. Like sumatriptan, PNU-142633 reduced the increase in cat nucleus trigeminal caudalis blood flow elicited by electrical stimulation of the trigeminal ganglion compared with the vehicle control. The direct vasoconstrictor potential of PNU-142633 was evaluated in vascular beds. Sumatriptan increased vascular resistance in carotid, meningeal and coronary arteries while PNU-142633 failed to alter resistance in these vascular beds. These data are discussed in relation to the clinical findings of PNU-142633 in a phase II acute migraine study [1].

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In vitro receptor binding assays for PNU-142633 typically involve measuring its affinity for the 5-HT1D and 5-HT1B receptors. Membrane preparations from cells expressing the human receptors are incubated with a radiolabeled ligand (e.g., [³H]-5-HT or a selective 5-HT1D radioligand) in the presence of varying concentrations of PNU-142633. The Ki values are calculated from competition binding curves, with PNU-142633 showing a Ki of 6 nM for 5-HT1D and >18,000 nM for 5-HT1B.

Cellular assays for PNU-142633 are performed in cells expressing the 5-HT1D receptor. The compound's agonist activity is assessed by measuring its ability to activate receptor-mediated signaling pathways, such as inhibition of cAMP production (as 5-HT1D is a Gi-coupled receptor). Cells are treated with PNU-142633, and cAMP levels are measured. The compound's potency and efficacy as a 5-HT1D agonist are determined from concentration-response curves.

Animal/Disease Models: Male Hartley guinea pig (250-300 g) stimulated with bipolar electrodes [1]
Doses: 0.03 mg/kg, 0.1 mg/kg, 0.3 mg/kg, 1 mg/kg
Route of Administration: intravenous (iv) (iv)injection
Experimental Results: External Significant reduction in osmosis was effective at doses of 0.1 and 0.3 mg/kg, but not at doses of 0.03 and 1.0 mg/kg.

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In vivo animal studies with PNU-142633 have been performed to evaluate its anti-migraine potential. In these studies, the compound's ability to block neurogenic plasma protein extravasation and inhibit sympathetically evoked tachycardia responses is assessed. These endpoints are commonly used to evaluate the efficacy of anti-migraine compounds in animal models. However, detailed protocols are not extensively reported in the available literature.

PNU-142633 has a molecular weight of 422.53. It is typically dissolved in DMSO for in vitro studies. The compound is stable when stored as a powder at -20°C. However, detailed pharmacokinetic parameters such as absorption, distribution, metabolism, excretion, half-life, and bioavailability are not extensively reported in the available literature. As a compound that reached Phase 2 clinical trials, it would have undergone pharmacokinetic characterization, but specific data are not detailed.

Comprehensive toxicology data for PNU-142633 are not extensively reported in the available literature. As a compound that was investigated in clinical trials, it would have undergone toxicological evaluation, but specific data are not detailed. The compound is classified as a research-use-only chemical and is not intended for human consumption. The compound was discontinued in Phase 2 for undisclosed reasons.

[1]. McCall RB, et al. Preclinical studies characterizing the anti-migraine and cardiovascular effects of the selective 5-HT1D receptor agonist PNU-142633. Cephalalgia. 2002 Dec;22(10):799-806.
[2]. Gomez-Mancilla B, et al. Safety and efficacy of PNU-142633, a selective 5-HT1D agonist, in patients with acute migraine. Cephalalgia. 2001 Sep;21(7):727-32.

These studies investigated the pharmacological mechanisms of neurogenic dural vasodilation in anesthetized guinea pigs. After establishing a closed cranial window, meningeal (dural) vessels were observed using a live microscope, and their diameters were continuously measured using a video sizing analyzer. Before dilating dural vessels with calcitonin gene-related peptide (CGRP; 1 μg kg⁻¹, intravenously) or local electrical stimulation of the dura mater (up to 300 μA), endothelin-1 (3 μg kg⁻¹, intravenously) was administered to constrict the dural vessels. In guinea pigs pretreated with the CGRP receptor antagonist CGRP((8-37)) (0.3 mg kg⁻¹, intravenously), the vasodilatory response induced by electrical stimulation was inhibited by 85%, indicating that CGRP plays an important role in neurogenic dural vasodilation in this species. Neurogenic dural vasodilation can also be blocked by the 5-HT(1B/1D) receptor agonist rizatriptan (100 μg/kg), with plasma concentrations estimated to be comparable to those required for efficacy in treating migraines. Rizatriptan does not reverse CGRP-induced dural vasodilation, suggesting it acts on presynaptic receptors on the sensory fibers of the trigeminal nerve innervating the dural vessels. Furthermore, the selective 5-HT(1D) receptor agonist PNU-142633 (100 μg kg⁻¹) also blocks neurogenic dural vasodilation, while the 5-HT(1F) receptor agonist LY334370 (3 mg kg⁻¹) does not, suggesting that rizatriptan may block neurogenic vasodilation by inhibiting CGRP release through action on 5-HT(1D) receptors located on the perivascular trigeminal nerve. This mechanism may be one of the mechanisms of action of triptan drugs such as rizatriptan in treating migraines, and suggests that guinea pigs are a suitable animal model for studying the pharmacology of neurogenic dural vasodilatory disorders. Br J Pharmacol. 2001 Aug;133(7):1029-34.

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PNU-142633 is a research-grade compound that was previously investigated as a potential anti-migraine agent but was discontinued in Phase 2 clinical trials. Its primary application is as a pharmacological tool for studying the 5-HT1D receptor. Its high selectivity for 5-HT1D over 5-HT1B makes it a valuable reagent for dissecting the specific functions of the 5-HT1D receptor in migraine pathophysiology and serotonergic signaling.

C24H30N4O3

422.53

422.232

C, 68.22; H, 7.16; N, 13.26; O, 11.36

187665-65-2

9845148

White to off-white solid powder

3.065

2

5

6

31

614

1

CNC(C1=CC2CCO[C@](CCN3CCN(C4=CC=C(C(=O)N)C=C4)CC3)([H])C=2C=C1)=O

PNTVCCRNJOGKGA-QFIPXVFZSA-N

InChI=1S/C24H30N4O3/c1-26-24(30)19-4-7-21-18(16-19)9-15-31-22(21)8-10-27-11-13-28(14-12-27)20-5-2-17(3-6-20)23(25)29/h2-7,16,22H,8-15H2,1H3,(H2,25,29)(H,26,30)/t22-/m0/s1

1H-2-Benzopyran-6-carboxamide, 1-(2-(4-(4-(aminocarbonyl)phenyl)-1-piperazinyl)ethyl)-3,4-dihydro-N-methyl-, (1S)-

PNU 142633; PNU142633; PNU-142633; 187665-65-2; PNU 142633; 1H-2-Benzopyran-6-carboxamide, 1-(2-(4-(4-(aminocarbonyl)phenyl)-1-piperazinyl)ethyl)-3,4-dihydro-N-methyl-, (1S)-; B54P1BQ73L; CHEMBL441095; (1S)-1-[2-[4-(4-carbamoylphenyl)piperazin-1-yl]ethyl]-N-methyl-3,4-dihydro-1H-isochromene-6-carboxamide; UNII-B54P1BQ73L; PNU-142633

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)