Pyrazoles as Anti-inflammatory and Analgesic Agents: In-vivo and In-silico Studies.

BACKGROUND: Pyrazole is a well-known nucleus in the pharmacy field with a wide range of other activities in addition to anti-inflammatory and analgesic, i.e., anticonvulsant, antiviral, and anticancer activities. There are well-known marketed drugs having pyrazole moiety as celecoxib, and lonazolac as COX-II inhibitors. AIMS: We aim to synthesize better anti-inflammatory than existing ones. Thiophene is also known for its analgesic and anti-inflammatory action. Thus, the fusion of both gives better anti-inflammatory agents. In the present studies, derivatives from two series of pyrazole were prepared by reacting substituted chalcone (3a-3f) derivatives prepared from 2-acetyl thiophene. They substituted aromatic aldehydes with phenyl hydrazine to form (5a-5f) and with 2, 4-dinitro phenyl hydrazine giving compounds (6a-6f) separately. METHODS: Purified and characterized pyrazoles have been analyzed for in-vivo analgesic and anti-inflammatory activities by using standard methods. Compounds 5e, 5f, and 6d were proved to be potent analgesics and series (5a-5f) was found to have anti-inflammatory action, which was further validated using docking and ADME studies. RESULTS: The ADME profile of synthesized compounds was found to be satisfactory. CONCLUSION: The synthesized compounds can serve as lead for further drug designing.

Transient receptor potential vanilloid 1 involved in the analgesic effects of total flavonoids extracted from Longxuejie ().

OBJECTIVE: To evaluate the analgesic effects of total flavonoids of Longxuejie (Resina Dracaenae Cochinchinensis) (TFDB) and explore the possible analgesic mechanism associated with transient receptor potential vanilloid 1 (TRPV1). METHODS: Whole-cell patch clamp technique was used to observe the effects of TFDB on capsaicin-induced TRPV1 currents. Rat experiments in vivo were used to observe the analgesic effects of TFDB. Western blot and immunofluorescence experiments were used to test the change of TRPV1 expression in DRG neurons induced by TFDB. RESULTS: Results showed that TFDB inhibited capsaicin-induced TRPV1 receptor currents in acutely isolated dorsal root ganglion (DRG) neurons of rats and the half inhibitory concentration was (16.7 ± 1.6) mg/L. TFDB (2-20 mg/kg) showed analgesic activity in the phase Ⅱ of formalin test and (0.02-2 mg per paw) reduced capsaicin-induced licking times of rats. TFDB (20 mg/kg) was fully efficacious on complete Freund's adjuvant (CFA)-induced inflammatory thermal hyperalgesia and capsaicin could weaken the analgesic effects. The level of TRPV1 expressions of DRG neurons was also decreased in TFDB-treated CFA-inflammatory pain rats. CONCLUSION: All these results indicated that the analgesic effect of TFDB may contribute to their modulations on both function and expression of TRPV1 channels in DRG neurons.

Efficacy assessment of novel methanimine derivatives in chronic constriction injury-induced neuropathic model: An in-vivo, ex-vivo and In-Silico approach.

The multicomponent etiology, complex clinical implications, dose-based side effect and degree of pain mitigation associated with the current pharmacological therapy is incapable in complete resolution of chronic neuropathic pain patients which necessitates the perpetual requirement of novel medication therapy. Therefore, this study explored the ameliorative aptitude of two novel methanimine imitative like (E)-N-(4-nitrobenzylidene)-4­chloro-2-iodobenzamine (KB 09) and (E)-N-(4-methylbenzylidene)-4­chloro-2-iodobenzamine (KB 10) in chronic constriction injury (CCI) of sciatic nerve induced neuropathic pain in rat model. Standard behavioral tests like dynamic and static allodynia, cold, thermal and mechanical hyperalgesia along with rotarod activity were performed at various experimental days like 0, 3, 7, 14 and 21. Enzyme linked immunosorbent assay (ELISA) on spinal tissue and antioxidant assays on sciatic nerve were executed accompanied by molecular docking and simulation studies. Prolonged ligation of sciatic nerve expressively induced hyperalgesia as well as allodynia in rats. KB 09 and KB 10 substantially attenuated the CCI elicited hyperalgesia and allodynia. They significantly reduced the biomarkers of pain and inflammation like Interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) in ELISA and while enhanced the GSH, SOD and CAT and diminished the MDA levels during antioxidant assays. KB 09 displayed -9.62 kcal/mol with TNF-α and -7.68 kcal/mol binding energy with IL-6 whereas KB 10 exhibited binding energy of -8.20 kcal/mol with IL-6 while -11.68 kcal/mol with TNF-α and hence both trial compounds ensured stable interaction with IL-6 and TNF-α during computational analysis. The results advocated that both methanimine derivatives might be novel candidates for attenuation of CCI-induced neuropathic pain prospects via anti-nociceptive, anti-inflammatory and antioxidant mechanisms.

LMD and LC-MS-based chemical constituents and pharmacological effects assessment for two different processing methods of the root of Paeonia lactiflora Pall.

The plant of Paeonia lactiflora Pall. belongs to Ranunculaceae, and its root can be divided into two categories according to different processing methods, which included that one was directly dried without peeling the root of the P. lactiflora (PR), and the other was peeled the root of the P. lactiflora (PPR) after boiled and dried. To evaluate the difference of chemical components, UPLC-ESI-Q-Exactive Focus-MS/MS and UPLC-QQQ-MS were applied. The distribution of chemical components in different tissues was located by laser microdissection (LMD), especially the different ingredients. A total of 86 compounds were identified from PR and PPR. Four kind of tissues were isolated from the fresh root of the P. lactiflora (FPR), and 54 compounds were identified. Especially the content of gallic acid, albiflorin, and paeoniflorin with high biological activities were the highest in the cork, but they were lower in PR than that in PPR, which probably related to the process. To illustrate the difference in pharmacological effects of PR and PPR, the tonifying blood and analgesic effects on mice were investigated, and it was found that the tonifying blood and analgesic effects of PPR was superior to that of PR, even though PR had more constituents. The material basis for tonifying blood and analgesic effect of the root of P. lactiflora is likely to be associated with an increase in constituents such as paeoniflorin and paeoniflorin lactone after boiled and peeled. The study was likely to provide some theoretical support for the standard and clinical application.

Bioassay-guided isolation of anti-inflammatory and antinociceptive metabolites among three Moroccan Juniperus leaves extract supported with in vitro enzyme inhibitory assays.

ETHNOPHARMACOLOGICAL RELEVANCE: Herbs of the genus Juniperus (family Cupressaceae) have been commonly used in ancestral folk medicine known as "Al'Araar" for treatment of rheumatism, diabetes, inflammation, pain, and fever. Bioassay-guided isolation of bioactives from medicinal plants is recognized as a potential approach for the discovery of novel drug candidates. In particular, non-addictive painkillers are of special interest among herbal phytochemicals. AIM OF THE STUDY: The current study aimed to assess the safety of J. thurifera, J. phoenicea, and J. oxycedrus aqueous extracts in oral treatments; validating the traditionally reported anti-inflammatory and analgesic effects. Further phytochemical investigations, especially for the most bioactive species, may lead to isolation of bioactive metabolites responsible for such bioactivities supported with in vitro enzyme inhibition assays. MATERIALS AND METHODS: Firstly, the acute toxicity study was investigated following the OECD Guidelines. Then, the antinociceptive, and anti-inflammatory bioactivities were evaluated based on chemical and mechanical trauma assays and investigated their underlying mechanisms. The most active J. thurifera n-butanol fraction was subjected to chromatographic studies for isolating the major anti-inflammatory metabolites. Moreover, several enzymatic inhibition assays (e.g., 5-lipoxygenase, protease, elastase, collagenase, and tyrosinase) were assessed for the crude extracts and isolated compounds. RESULTS: The results showed that acute oral administration of the extracts (300-500 mg/kg, p. o.) inhibited both mechanically and chemically triggered inflammatory edema in mice (up to 70% in case of J. thurifera) with a dose-dependent antinociceptive (tail flick) and anti-inflammatory pain (formalin assay) activities. This effect was partially mediated by naloxone inhibition of the opioid receptor (2 mg/kg, i. p.). In addition, 3-methoxy gallic acid (1), quercetin (2), kaempferol (3), and ellagic acid (4) were successfully identified being involved most likely in J. thurifera extract bioactivities. Nevertheless, quercetin was found to be the most potent against 5-LOX, tyrosinase, and protease with IC50 of 1.52 ± 0.01, 192.90 ± 6.20, and 399 ± 9.05 µM, respectively. CONCLUSION: J. thurifera extract with its major metabolites are prospective drug candidates for inflammatory pain supported with inhibition of inflammatory enzymes. Interestingly, antagonism of opioid and non-opioid receptors is potentially involved.

Molecular design, construction and analgesic mechanism insights into the novel transdermal fusion peptide ANTP-BgNPB.

Toad venom, a traditional Chinese medicine, exhibits remarkable medicinal properties of significant therapeutic value. The peptides present within toad venom possess a wide range of biological functions, yet the neuropeptide B (NPB) and it modification requires further exploration to comprehensively understand its mechanisms of action and potential applications. In this study, a fusion peptide, ANTP-BgNPB, was designed to possess better analgesic properties through the transdermal modification of BgNPB. After optimizing the conditions, the expression of ANTP-BgNPB was successfully induced. The molecular dynamics simulations suggested that the modified protein exhibited improved stability and receptor binding affinity compared to its unmodified form. The analysis of the active site of ANTP-BgNPB and the verification of mutants revealed that GLN3, SER38, and ARG42 were crucial for the protein's recognition and binding with G protein-coupled receptor 7 (GPR7). Moreover, experiments conducted on mice using the hot plate and acetic acid twist body models demonstrated that ANTP-BgNPB was effective in transdermal analgesia. These findings represent significant progress in the development of transdermal delivery medications and could have a significant impact on pain management.

Discovery of first-in-class highly selective TRPV1 antagonists with dual analgesic and hypoglycemic effects.

Analgesia and blood sugar control are considered as two main unmet clinical needs for diabetes related neuropathic pain patients. Transient receptor potential vanilloid type-1 (TRPV1) channel is a highly validated target for pain perception, while no TRPV1 antagonists have been approved due to hyperthermia side effects. Herein, two series of new TRPV1 antagonists with flavonoid skeleton were designed by the structure-based drug design (SBDD) strategy. After comprehensive evaluation, compound CX-3 was identified as a promising TRPV1 antagonist. CX-3 exhibited equivalent TRPV1 antagonistic activity with classical TRPV1 antagonist BCTC in vitro, and exerted better analgesic activity in vivo than that of BCTC in the formalin induced inflammatory pain model without hyperthermia risk. Moreover, CX-3 exhibited robust glucose-lowering effects and showed high selectivity over other ion channels. Overall, these findings identified a first-in-class highly selective TRPV1 antagonist CX-3, which is a promising candidate to target the pathogenesis of diabetes related neuropathic pain.

The Potential Antinociceptive Effect and Mechanism of Cannabis sativa L. Extract on Paclitaxel-Induced Neuropathic Pain in Rats Uncovered by Multi-Omics Analysis.

Cannabis sativa L. (hemp) is a herbaceous plant rich in cannabinoids with a long history of use in pain treatment. The most well-characterized cannabinoids, cannabidiol (CBD) and Δ9-tetrahydrocannabinol (Δ9-THC), garnered much attention in chemotherapy-induced peripheral neuropathy (CIPN) treatment. However, few studies have investigated the biological benefits and mechanism of hemp extract on CIPN. In the present study, hemp extract (JG) rich in cannabinoids was extracted by supercritical fluid carbon dioxide extraction (SFCE). The antinociceptive efficacy was evaluated using a paclitaxel-induced peripheral neuropathy (PIPN) rat model based on behavioral tests. Further omics-based approaches were applied to explore the potential mechanisms. The results showed that JG decreased mechanical allodynia, thermal hyperalgesia, and inflammatory cytokines in PIPN rats significantly. Transcriptome analysis identified seven key genes significantly regulated by JG in PIPN model rats, mainly related to the neuroactive ligand-receptor interaction pathway, PPAR signaling pathway, and cAMP signaling pathway. In metabolomic analysis, a total of 39 significantly altered metabolites were identified, mainly correlated with pentose and glucuronate interconversions and the glycerophospholipid metabolism pathway. Gut microbiota analysis suggested that increased community Lachnoclostridium and Lachnospiraceae_UCG-006 in PIPN rats can be reversed significantly by JG. In conclusion, hemp extract exhibited antinociceptive effects on PIPN. The analgesic mechanism was probably related to the regulation of inflammation, neuroactive ligand-receptor interaction pathway, sphingolipid metabolism, etc. This study provides novel insights into the functional interactions of Cannabis sativa L. extract on PIPN.

A novel isoquinoline alkaloid HJ-69 isolated from Zanthoxylum bungeanum attenuates inflammatory pain by inhibiting voltage-gated sodium and potassium channels.

ETHNOPHARMACOLOGY RELEVANCE: Zanthoxylum bungeanum Maxim. (Z. bungeanum), a member of the Rutaceae family, has a rich history of traditional use in Asia for treating arthritis and toothache conditions. As characteristic chemical components, numerous kinds of alkaloids have been extracted from plants and their diverse biological activities have been reported. However, research on the isoquinoline alkaloid, a specific type of alkaloids, in Z. bungeanum was scarce. AIM OF THE STUDY: The study aimed to isolate a novel isoquinoline alkaloid from Z. bungeanum and explore its pharmacological activity in vitro and analgesic activity in vivo. MATERIALS AND METHODS: Isoquinoline alkaloid isolation and identification from Z. bungeanum were conducted using chromatographic and spectroscopic methods. The whole-cell patch-clamp technique was applied to assess its impact on neuronal excitability, and endogenous voltage-gated potassium (Kv) and sodium (Nav) currents in acutely isolated mouse small-diameter dorsal root ganglion (DRG) neurons. Its inhibitory impacts on channels were further validated with HEK293 cells stably expressing Nav1.7 and Nav1.8, and Chinese hamster ovary (CHO) cells transiently expressing Kv2.1. The formalin inflammatory pain model was utilized to evaluate the potential analgesic activity in vivo. RESULTS: A novel isoquinoline alkaloid named HJ-69 (N-13-(3-methoxyprop-1-yl)rutaecarpine) was isolated and identified from Z. bungeanum for the first time. HJ-69 significantly suppressed the firing frequency and amplitudes of action potentials in DRG neurons. Consistently, it state-dependently inhibited endogenous Nav currents of DRG neurons, with half maximal inhibitory concentration (IC50) values of 13.06 ± 2.06 µM and 30.19 ± 2.07 µM for the inactivated and resting states, respectively. HJ-69 significantly suppressed potassium currents in DRG neurons, which notably inhibited the delayed rectifier potassium (IK) currents (IC50 = 6.95 ± 1.29 µM) and slightly affected the transient outward potassium (IA) currents (IC50 = 523.50 ± 39.16 µM). Furtherly, HJ-69 exhibited similar potencies on heterologously expressed Nav1.7, Nav1.8, and Kv2.1 channels, which correspondingly represent the main components in neurons. Notably, intraperitoneal administration of 30 mg/kg and 100 mg/kg HJ-69 significantly alleviated pain behaviors in the mouse inflammatory pain model induced by formalin. CONCLUSION: The study concluded that HJ-69 is a novel and active isoquinoline alkaloid, and the inhibition of Nav and Kv channels contributes to its analgesic activity. HJ-69 may be a promising prototype for future analgesic drug discovery based on the isoquinoline alkaloid.

Discovery of an Ortho-Substituted N-Cyclopropylmethyl-7α-phenyl-6,14-endoethano-tetrahydronorthebaine Derivative as a Selective and Potent Kappa Opioid Receptor Agonist with Subsided Sedative Effect.

Research into kappa opioid receptor (KOR) agonists with attenuated central-nervous-system side effects is a critical focus for developing productive and safe analgesics. Herein, a series of ortho-substituted N-cyclopropylmethyl-7α-phenyl-6,14-endoethano-tetrahydronorthebaines were designed, synthesized, and subjected to bioassays. Compound 7a exhibited high subtype selectivity and potent agonistic activity toward KOR (KOR, Ki = 3.9 nM, MOR/KOR = 270, DOR/KOR = 1075; [35S]GTPγS binding, EC50 = 3.4 nM). Additionally, this compound exhibited robust and persistent antinociceptive effects in rodent models with different animal strains (hot plate test, ED50 = 0.20-0.30 mg/kg, i.p.; abdominal constriction test, ED50 = 0.20-0.60 mg/kg, i.p.), with its KOR-mediated mechanism for antinociception firmly established. Notably, compound 7a, unlike conventional KOR agonists, displayed minimal sedation and aversion at the antinociceptive ED50 dose. This feature addresses a crucial limitation in existing KOR agonists, positioning compound 7a as a promising novel therapeutic agent.

In vivo effects of a selected thiourea derivative 1-(2-chlorobenzoyl)-3-(2,3-dichlorophenyl) against nociception, inflammation and gastric ulcerogenicity: Biochemical, histopathological and in silico approaches.

The current study was designed to investigate the potential of a synthetic therapeutic agent for better management of pain and inflammation, exhibiting minimal to non-existent ulcerogenic effects. The effect of 1-(2-chlorobenzoyl)-3-(2,3-dichlorophenyl) thiourea was assessed through model systems of nociception and anti-inflammatory activities in mice. In addition, the ulcerogenic potential was evaluated in rats using the NSAID-induced pyloric ligation model, followed by histopathological and biochemical analysis. The test was conducted on eight groups of albino rats, comprising of group I (normal saline), groups II and III (aspirin® at doses of 100 mg/kg and 150 mg/kg, respectively), groups IV and V (indomethacin at doses of 100 mg/kg and 150 mg/kg, respectively), and groups VI, VII, and VIII (lead-compound at 15 mg/kg, 30 mg/kg and 45 mg/kg doses, respectively). Furthermore, molecular docking analyses were performed to predict potential molecular target site interactions. The results showed that the lead-compound, administered at doses of 15, 30, and 45 mg/kg, yielded significant reductions in chemically and thermally induced nociceptive pain, aligning with the levels observed for aspirin® and tramadol. The compound also effectively suppressed inflammatory response in the carrageenan-induced paw edema model. As for the ulcerogenic effects, the compound groups displayed no considerable alterations compared to the aspirin® and indomethacin groups, which displayed substantial increases in ulcer scores, total acidity, free acidity, and gastric juice volume, and a decrease in gastric juice pH. In conclusion, these findings suggest that our test compound exhibits potent antinociceptive, anti-inflammatory properties and is devoid of ulcerogenic effects.

Ligustilide covalently binds to Cys703 in the pre-S1 helix of TRPA1, blocking the opening of channel and relieving pain in rats with acute soft tissue injury.

ETHNOPHARMACOLOGICAL RELEVANCE: The natural anodyne Ligustilide (Lig), derived from Angelica sinensis (Oliv.) Diels and Ligusticum chuanxiong Hort., has been traditionally employed for its analgesic properties in the treatment of dysmenorrhea and migraine, and rheumatoid arthritis pain. Despite the existing reports on the correlation between TRP channels and the analgesic effects of Lig, a comprehensive understanding of their underlying mechanisms of action remains elusive. AIM OF THE STUDY: The objective of this study is to elucidate the mechanism of action of Lig on the analgesic target TRPA1 channel. METHODS: The therapeutic effect of Lig was evaluated in a rat acute soft tissue injury model. The analgesic target was identified through competitive inhibition of TRP channel agonists at the animal level, followed by Fluo-4/Ca2+ imaging on live cells overexpressing TRP proteins. The potential target was verified through in-gel imaging, colocalization using a Lig-derived molecular probe, and a drug affinity response target stability assay. The binding site of Lig was identified through protein spectrometry and further analyzed using molecular docking, site-specific mutation, and multidisciplinary approaches. RESULTS: The administration of Lig effectively ameliorated pain and attenuated oxidative stress and inflammatory responses in rats with soft tissue injuries. Moreover, the analgesic effects of Lig were specifically attributed to TRPA1. Mechanistic studies have revealed that Lig directly activates TRPA1 by interacting with the linker domain in the pre-S1 region of TRPA1. Through metabolic transformation, 6,7-epoxyligustilide (EM-Lig) forms a covalent bond with Cys703 of TRPA1 at high concentrations and prolonged exposure time. This irreversible binding prevents endogenous electrophilic products from entering the cysteine active center of ligand-binding pocket of TRPA1, thereby inhibiting Ca2+ influx through the channel opening and ultimately relieving pain. CONCLUSIONS: Lig selectively modulates the TRPA1 channel in a bimodal manner via non-electrophilic/electrophilic metabolic conversion. The epoxidized metabolic intermediate EM-Lig exerts analgesic effects by irreversibly inhibiting the activation of TRPA1 on sensory neurons. These findings not only highlight the analgesic mechanism of Lig but also offer a novel nucleophilic attack site for the development of TRPA1 antagonists in the pre-S1 region.

Chemical Investigation and Antinociceptive Activity Evaluation of Marrubium Vulgare L. Aqueous Extract.

Marrubium vulgare L. (Lamiaceae) has a long history of use in traditional herbal medicine for the treatment of respiratory tract infections, inflammatory conditions, and pain. This study aimed to investigate the chemical composition, acute toxicity, and antinociceptive effects of the aqueous extract from M. vulgare leaves (AEMV). Antioxidant activity was evaluated using DPPH and reducing power assays. The chemical composition of AEMV was determined through LC-MS/MS, and the levels of total phenolics, flavonoids, and condensed tannins were quantified. Acute oral toxicity was assessed in male Swiss mice with a single oral dose of AEMV (1, 2, 5 g/kg). The analgesic impact was examined through writhing, hot plate, and formalin tests. Our findings not only confirmed the safety of the extract in animal models but also revealed significant antioxidant activity in AEMV. High-performance liquid chromatography (HPLC) analysis identified important bioactive compounds, with marrubiin being a major component. Furthermore, AEMV demonstrated robust antinociceptive properties in all conducted tests, highlighting its potential as a valuable natural source of bioactive compounds suitable for a wide range of therapeutic applications.

Evaluation of Rhododendri Mollis Flos and its representative component as a potential analgesic.

Rhododendri Mollis Flos (R. mole Flos), the dried flowers of Rhododendron mole G. Don, have the ability to relieve pain, dispel wind and dampness, and dissolve blood stasis, but they are highly poisonous. The significance of this study is to explore the analgesic application potential of R. mole Flos and its representative component. According to the selected processing methods recorded in ancient literature, the analgesic activities of wine- and vinegar-processed R. mole Flos, as well as the raw product, were evaluated in a writhing test with acetic acid and a formalin-induced pain test. Subsequently, the HPLC-TOP-MS technique was utilized to investigate the changes in active components before and after processing once the variations in activities were confirmed. Based on the results, rhodojaponin VI (RJ-Vl) was chosen for further study. After processing, especially in vinegar, R. mole Flos did not only maintain the anti-nociception but also showed reduced toxicity, and the chemical composition corresponding to these effects also changed significantly. Further investigation of its representative components revealed that RJ-VI has considerable anti-nociceptive activity, particularly in inflammatory pain (0.3 mg/kg) and peripheral neuropathic pain (0.6 mg/kg). Its toxicity was about three times lower than that of rhodojaponin III, which is another representative component of R. mole Flos. Additionally, RJ-VI mildly inhibits several subtypes of voltage-gated sodium channels (IC50 > 200 µM) that are associated with pain or cardiotoxicity. In conclusion, the chemical substances and biological effects of R. mole Flos changed significantly before and after processing, and the representative component RJ-VI has the potential to be developed into an effective analgesic.

Discovery of new 2-(3-(naphthalen-2-yl)-4,5-dihydro-1H-pyrazol-1-yl)thiazole derivatives with potential analgesic and anti-inflammatory activities: In vitro, in vivo and in silico investigations.

Joining the global demand for the discovery of potent NSAIDs with minimized ulcerogenic effect, new pyrazole clubbed thiazole derivatives 5a-o were designed and synthesized. The new derivatives were initially evaluated for their analgesic activity. Eight compounds 5a, 5c, 5d, 5e, 5f, 5h, 5m, and 5o showed higher activity than Indomethacin (potency = 105-130 % vs. 100 %). Subsequently, they were picked for further evaluation of their anti-inflammatory activity, ulcerogenic liability as well as toxicological studies. Derivatives 5h and 5m showed a potential % edema inhibition after 3 h (79.39 % and 72.12 %, respectively), with a promising safety profile and low ulcer indices (3.80 and 3.20, respectively). The two compounds 5h and 5m were subjected to in vitro COX-1 and COX-2 inhibition assay. The candidate 5h showed nearly equipotent COX-1 inhibition (IC50 = 38.76 nM) compared to the non-selective reference drug Indomethacin (IC50 = 35.72 nM). Compound 5m expressed significant inhibitory activities and a higher COX-2 selectivity index (IC50 = 87.74 nM, SI = 2.05) in comparison with Indomethacin (SI = 0.52), with less selectivity than Celecoxib (SI = 8.31). Simulation docking studies were carried out to gain insights into the binding interaction of compounds 5h and 5m in the vicinity of COX-1 and COX-2 enzymes that illustrated the importance of pyrazole clubbed thiazole core in hydrogen bonding interactions. The thiazole motif of compounds 5h and 5m exhibited a well orientation toward COX-1 Arg120 key residue by hydrogen bonding interactions. Compound 5h revealed an additional arene-cation interaction with Arg120 that could rationalize its superior COX-1 inhibitory activity. Compounds 5h and 5m overlaid the co-crystallized ligand Celecoxib I differently in the active site of COX-2. Compound 5m showed an enhanced accommodation with binding energy of - 6.13 vs. - 1.70 kcal/mol of compounds 5h. The naphthalene ring of compound 5m adopted the Celecoxib I benzene sulfonamide region that is stabilized by hydrogen-arene interactions with the hydrophobic sidechains of the key residues Ser339 and Phe504. Further, the core structure of compound 5m, pyrazole clubbed thiazole, revealed deeper hydrophobic interactions with Ala513, Leu517 and Val509 residues. Finally, a sensitive and accurate UPLC-MS/MS method was developed for the simultaneous estimation of some selected promising pyrazole derivatives in rat plasma. Accordingly, compounds 5h and 5m were suggested to be promising potent analgesic and anti-inflammatory agents with improved safety profiles and a novel COX isozyme modulation activity.

Phellintremulins A-C, antinociceptive sesquiterpenoids from the medicinal fungus Phellinus tremulae.

Phellintremulin A (1), a rearranged sesquiterpenoid with an unprecedented bicyclic backbone, and two previously unreported illudane-type sesquiterpenoids, namely phellintremulin B (2) and phellintremulin C (3), together with two known analogues (±)‒4 and (±)‒5, were isolated from cultures of the medicinal fungus Phellinus tremulae. Their structures and absolute configurations were established by means of spectroscopic data and HRESIMS analyses, as well as ECD and NMR calculations. A plausible biogenesis for 1 was discussed. The electrophysiological experiments showed that phellintremulins (A‒C) can inhibit Nav current in DRG neuron cells at 10 µM, with percentage inhibitions of 23.2%, 49.3%, and 31.7%, respectively. The antinociceptive activities of phellintremulins (A‒C) were evaluated via the acetic acid-induced writhing test in mice at a dose of 3 mg/kg. They showed significant antinociceptive effects with percentages of inhibition of 43.8%, 54.4%, and 50.6%, respectively, and phellintremulin B and C expressed more potent analgesic effect than lidocaine.

Novel drug-drug salt crystals of metformin with ibuprofen or naproxen: Improved solubility, dissolution rate, and synergistic antinociceptive effects.

As the monotherapy of available analgesics is usually accompanied by serious side effects or limited efficacy in the management of chronic pain, multimodal analgesia is widely used to achieve improved benefit-to-risk ratios in clinic. Drug-drug salts are extensively researched to optimize the physicochemical properties of active pharmaceutical ingredients (APIs) and achieve clinical benefits compared with individual APIs or their combination. New drug-drug salt crystals metformin-ibuprofen (MET-IBU) and metformin-naproxen (MET-NAP) were prepared from metformin (MET) and two poorly water-soluble anti-inflammatory drugs (IBU and NAP) by the solvent evaporation method. The structures of these crystals were confirmed by single crystal and powder X-ray diffraction, Hirshfeld surface, Fourier transform infrared spectroscopy and thermal analysis. Both MET-IBU and MET-NAP showed significantly improved solubility and intrinsic dissolution rate than the pure IBU or NAP. The stability test indicated that MET-IBU and MET-NAP have excellent physical stability under stressing test (10 days) and accelerated conditions (3 months). Moreover, isobolographic analysis suggested that MET-IBU and MET-NAP exerted potent and synergistic antinociceptive effects in λ-Carrageenan-induced inflammatory pain in mice, and both of them had an advantage in rapid pain relief. These results demonstrated the potential of MET-IBU and MET-NAP to achieve synergistic antinociceptive effects by developing drug-drug salt crystals.

Multitarget µ-Opioid Receptor Agonists─Neuropeptide FF Receptor Antagonists Induce Potent Antinociception with Reduced Adverse Side Effects.

The design of bifunctional compounds is a promising approach toward the development of strong analgesics with reduced side effects. We here report the optimization of the previously published lead peptide KGFF09, which contains opioid receptor agonist and neuropeptide FF receptor antagonist pharmacophores and is shown to induce potent antinociception and reduced side effects. We evaluated the novel hybrid peptides for their in vitro activity at MOP, NPFFR1, and NPFFR2 and selected four of them (DP08/14/32/50) for assessment of their acute antinociceptive activity in mice. We further selected DP32 and DP50 and observed that their antinociceptive activity is mostly peripherally mediated; they produced no respiratory depression, no hyperalgesia, significantly less tolerance, and strongly attenuated withdrawal syndrome, as compared to morphine and the recently FDA-approved TRV130. Overall, these data suggest that MOP agonist/NPFF receptor antagonist hybrids might represent an interesting strategy to develop novel analgesics with reduced side effects.

Phytochemical, Pharmacological, and Molecular Docking Study of Dry Extracts of Matricaria discoidea DC. with Analgesic and Soporific Activities.

Pineapple weed (Matricaria discoidea DC.) is a widespread plant in Europe and North America. In ethnomedicine, it is well-known for its anti-inflammatory and spasmolytic activities. The aim of this research was to develop novel methods of M. discoidea processing to obtain essential oil and dry extracts and to investigate their phytochemical compositions. Moreover, the molecular docking of the main substances and the in vivo studies on their soporific and analgesic activities were conducted. The essential oil and two dry extracts from M. discoidea were prepared. A total of 16 phenolic compounds (seven flavonoids, seven hydroxycinnamic acids, and two phenolic acids) in the dry extracts were identified by means of UPLC-MS/MS. In the essential oil, nine main terpenoids were identified by gas chromatography (GC). It was shown that phenolic extraction from the herb was successful when using 70% ethanol in a triple extraction method and at a ratio of 1:14-1:16. The in vivo studies with rodents demonstrated the analgesic activity of the M. discoidea extracts and improvements in the sleep of animals. The dry extracts of M. discoidea did not show any toxicity. The molecular docking analysis showed a high probability of COX-1,2 inhibition and NMDA receptor antagonism by the extracts.

Asymmetric Carbene-Alkyne Metathesis-Mediated Cascade: Synthesis of Benzoxazine Polychiral Polyheterocycles and Discovery of a Novel Pain Blocker.

This study introduces a novel approach for synthesizing Benzoxazine-centered Polychiral Polyheterocycles (BPCPHCs) via an innovative asymmetric carbene-alkyne metathesis-triggered cascade. Overcoming challenges associated with intricate stereochemistry and multiple chiral centers, the catalytic asymmetric Carbene Alkyne Metathesis-mediated Cascade (CAMC) is employed using dirhodium catalyst/Brønsted acid co-catalysis, ensuring precise stereo control as validated by X-ray crystallography. Systematic substrate scope evaluation establishes exceptional diastereo- and enantioselectivities, creating a unique library of BPCPHCs. Pharmacological exploration identifies twelve BPCPHCs as potent Nav ion channel blockers, notably compound 8 g. In vivo studies demonstrate that intrathecal injection of 8 g effectively reverses mechanical hyperalgesia associated with chemotherapy-induced peripheral neuropathy (CIPN), suggesting a promising therapeutic avenue. Electrophysiological investigations unveil the inhibitory effects of 8 g on Nav1.7 currents. Molecular docking, dynamics simulations and surface plasmon resonance (SPR) assay provide insights into the stable complex formation and favorable binding free energy of 8 g with C5aR1. This research represents a significant advancement in asymmetric CAMC for BPCPHCs and unveils BPCPHC 8 g as a promising, uniquely acting pain blocker, establishing a C5aR1-Nav1.7 connection in the context of CIPN.