Design, synthesis, and pharmacological evaluation of some 2-[4-morpholino]-3-aryl-5-substituted thiophenes as novel anti-inflammatory agents: generation of a novel anti-inflammatory pharmacophore

Compounds incorporating a thiophene moiety, a pi excess five membered heterocycle, have attracted a great deal of research interest owing to the therapeutic utility of the template as useful drug molecular scaffolding. Recently we reported the anti-inflammatory activity profile exhibited by two thiophene analogs, AP84 and AP82 in acute and chronic models of inflammation. The good activity profile exhibited by AP84, a 3-(substituted aryl)-2-(4-morpholino)-5-heteroaryl substituted analog of thiophene, in the formalin induced rat paw edema chronic model as compared to a weak activity in acute carrageenin induced rat paw edema, and the slightly better protection exhibited in the acute model by AP82 (27%), the 5-aroyl analog provided an impetus for a proper exploration of their structural types. In this paper we report the synthesis and pharmacological evaluation of some novel, 2-(4-morpholino)-3-(substituted aryl)-5-substituted thiophenes, as possible anti-inflammatory leads. The 3-(4-chlorophenyl)-2-(4-morpholino) thiophene analogs AP49, AP158, and AP88 provided a protection of 20%, 23%, and 20%, respectively, when screened for anti-inflammatory activity in carrageenin induced rat paw edema, an acute in vivo model, comparable to that of AP82, at a dose level of 100mg/kg body weight p.o. compared to ibuprofen as standard. The replacement of the 3-(4-chlorophenyl) moiety with the 3-phenyl moiety gave rise to AP50 (30%), AP159 (38%), AP27 (0%), and AP92 (38%), with three analogs being more active in the acute model. Alteration of the group para to the phenyl ring at third position, from chloro, to methyl mercapto gave rise to the 3-(4-methylmercapto-phenyl) analogs AP54 (20%), AP160 (0%), and AP73 (52%), with only one analog appearing to be better than AP82. These results indicate that 4-methane sulfonyl aroyl group at 5-position and other substituents of different quadrants of Craig plot on the phenyl moiety at the third position could lead to more potent candidates. However, alteration of aroyl to substituted pyridyl at 5-position with a phenyl group at the third position as in AP26 gave rise to much better protection (66%) again reinforcing the importance of the heteroaryl ring at the fifth position and implying its utility in the composition of a novel pharmacophore for designing better trisubstituted thiophenes as anti-inflammatory agents.     

Design, synthesis, and SAR studies of some 5-aliphatic oximino esters of thiophene as potential anti-inflammatory leads: comparative biological activity profile of aliphatic oximes vs aromatic oximes

A series of novel tetra substituted thiophenes were synthesized, characterized, and evaluated for their anti-inflammatory activity in carrageenin induced rat paw edema model-an acute in vivo model. Compounds V1, V3, V11, V12, V17, and V18 showed good anti-inflammatory activity, indicating the importance of oxime moiety in modulating the activity. The structure-activity relationship studies explore "the aliphatic oxime esters" attached via a ketone bridge to fifth position of the thiophene, and indicate that this feature may enhance the anti-inflammatory activity as compared to aromatic oximes. Since free radicals are implicated in various inflammatory disorders, the free radical scavenging activity of some of the synthesized candidates was assessed using 1,1-diphenyl-2-picryl hydrazyl assay. The oxime containing analogs exhibited weak to moderate activity as free radical scavengers in DPPH assay. A plausible reasoning for its free radical scavenging ability is discussed. All the compounds were also screened in nitro blue tetrazolium model, to assess them as superoxide anion radical scavengers. A direct correlation between anti-inflammatory activity and free radical scavenging activity was not seen. The results disclose a new class of anti-inflammatory agents designed and synthesized for the first time wherein the utility of aliphatic oxime esters in modulating the anti-inflammatory activity profile is apparent. This will give us potential anti-inflammatory leads.     

Tetra substituted thiophenes as anti-inflammatory agents: exploitation of analogue-based drug design

A series of 17 novel tetra substituted thiophenes was designed, synthesized, and screened for anti-inflammatory activity in carrageenin induced rat paw edema model, an acute in vivo model. The lead molecule selected was Tenidap, a dual COX/LOX inhibitor. Compounds I (43%), III (60%), IV (60%), and VIII (64%) showed moderate to good anti-inflammatory activity. The best candidate among the whole series was VIII, which gave 64% protection to the inflamed paw. The side chain of candidate VIII had resemblance to that of Romazarit, a DMARD, which was withdrawn due to its toxicity profile. A probable reason for the metabolic stability of the proposed side chain not having the possibility of generating peroxy type radicals or acrylic acid moieties, unlike Romazarit, is discussed. The biological activity exhibited by the three designed series was in the order of methyl amino series > ethyl amino series > carbethoxy amino series. The –(CO)–CH₂–COOR side chain at the fifth position as in candidate VIII, the methyl amino group at the second position, and the ester at the third position of the thiophene can be considered as a three-point pharmacophore for designing better anti-inflammatory agents. The present study is a classical example of the exploitation of an analogue based drug design, which culminated in the development of good anti-inflammatory agents that have the potential of becoming dual inhibitors.     

Design,synthesis and evaluation of some pyrazolo[3,4-d]pyrimidine derivatives bearing thiazolidinone moiety as anti-inflammatory agents

Two new series of pyrazolo[3,4-d]pyrimidine bearing thiazolidinone moiety were designed and synthesized. The newly synthesized compounds were evaluated for their in vitro (COX-1 and COX-2) inhibitory assay. Compounds that showed promising COX-2 selectivity were further subjected to in vivo anti-inflammatory screening applying formalin induced paw edema (acute model) and cotton-pellet induced granuloma (chronic model) assays using celecoxib and diclofenac sodium as reference drugs. The histopathological and ulcerogenic potential were also determined. In vivo anti-inflammatory data showed that compounds 2, 6, 7d displayed anti-inflammatory activity higher than both references in the formalin induced paw edema model. On the other hand, compounds 2, 3d, 3e, 7b and 7d displayed anti-inflammatory activity greater than or nearly equivalent to diclofenac sodium in the cotton pellet-induced granuloma assay. Moreover, most of the tested compounds revealed good gastrointestinal safety profile. Collectively, compounds 2 and 7d were considered as promising candidates in managing both acute and chronic inflammation with safe gastrointestinal margin.     

Design, synthesis and pharmacological evaluation of novel tetrasubstituted thiophene analogues as anti-inflammatory agents

A new series of tetrasubstituted thiophene analogues (4a-4f, 5a-5f and 8a-8i) were designed incorporating the pharmacophoric features of COX-1 (as in fenamates), 5-LOX and the p38 MAP kinase inhibitors. The designed series was synthesized by nucleophilic addition of aryl/aroylisothiocyanate and enamine (2) yielding the addition product l-(alpha-Carbomethoxy-beta-aminothiocrotonoyl)-aryl/aroyl amines (3/7); which on reaction with substituted phenacyl bromides gave the targeted tetrasubstituted thiophene esters (4a-4f / 8a-8i). The tetrasubstituted thiophenes esters (4a-4f ) on hydrolysis with one equivalent of potassium hydroxide solution in methanol at room temperature gave corresponding acids (5a-5f ). All the targeted compounds were evaluated for their anti-inflammatory activity in carrageenin-induced rat hind paw oedema model at the doses of 10, 20 and 40 mg/kg body weight using standard drugs mefanamic acid and ibuprofen. The compounds (4c, 4e, 4f, 5f, 8a- 8i) which gave reasonable protection to the inflamed paw, eliciting good or moderate comparable anti-inflammatory activity were selected for investigating their analgesic activity using acetic acid induced writhing response test in albino mice at 10 mg/kg dose using standard drug ibuprofen and in order to arrive at possible mechanism of their anti-inflammatory activity, in vitro antioxidant nitric oxide radical scavenging assay at the concentrations of 5, 10, 15, 20, 25, 30 and 35 microg/mL were performed using standard drug ascorbic acid.     

Anti-inflammatory and antimicrobial activities of triterpenoids from Strobilanthes callosus nees.

The anti-inflammatory and antimicrobial activities of the 95% ethanol extract, benzene fraction and isolated triterpenoids of Strobilanthes callosus were investigated. In the carrageenan-induced paw edema inflammation model, the taraxerol showed a high reduction of edema, but the antimicrobial effect observed was lower at the two doses employed. These results confirm the use of this plant in folk medicine as an anti-inflammatory and antimicrobial herbal drug.     

Absence of effect of chronic angiotensin II type 1 receptor blockade on endogenous kinin concentrations-induced paw edema model in the rat.

The effects of chronic treatment with losartan. an AT1 receptor antagonist, on the tissue content of bradykinin (BK) and des-Arg9-BK and on their pharmacological effects were examined in the carrageenan-induced paw edema model (0.5% solution, 50 microl/paw) in the rat. These effects were compared with those of angiotensin-converting enzyme inhibitors (ACEi). For this purpose, rats were chronically treated with losartan (3, 10 and 30 mg/kg/day) and enalapril or quinapril (1 mg/kg/day). Endogenous BK and des-Arg9-BK tissue contents at the site of local inflammation were measured by highly sensitive and specific enzyme immunoassays. Losartan 3 mg/kg/day for 7, 14 and 28 days had no significant effect on carrageenan-induced paw edema, but both losartan 10 and 30 mg/kg/day for 14 days significantly increased the hindpaw volume by 50% at 3 h and by 59% at 5 h. These effects, similar to those measured for ACEi, were inhibited by icatibant, a B2 kinin receptor antagonist (32.5 nmol/paw), that reduced carrageenan-induced paw edema to the level seen in vehicle-treated rats. In the same model, and contrary to ACEi, losartan 3, 10 and 30 mg/kg/day for 14 days had no significant effect on endogenous BK and des-Arg9-BK levels in the local inflammatory site or on circulating and tissue ACE activities. These results show, at least in that model, that the potentiating effects of losartan on carrageenan-induced paw edema are independent of the concentrations of endogenous kinins.     

Structure-guided approach on the role of substitution on amide-linked bipyrazoles and its effect on their anti-inflammatory activity

A structure-guided modelling approach using COX-2 as a template was used to investigate the effect of replacing the chloro atom located at the chlorophenyl ring of amide-linked bipyrazole moieties, aiming at attaining better anti-inflammatory effect with a good safety profile. Bromo, fluoro, nitro, and methyl groups were revealed to be ideal candidates. Consequently, new bipyrazole derivatives were synthesised. The in vitro inhibitory COX-1/COX-2 activity of the synthesised compounds exhibited promising selectivity. The fluoro and methyl derivatives were the most active candidates. The in vivo formalin-induced paw edoema model confirmed the anti-inflammatory activity of the synthesised compounds. All the tested derivatives had a good ulcerogenic safety profile except for the methyl substituted compound. In silico molecular dynamics simulations of the fluoro and methyl poses complexed with COX-2 for 50 ns indicated stable binding to COX-2. Generally, our approach delivers a fruitful matrix for the development of further amide-linked bipyrazole anti-inflammatory candidates.     

LIG抗炎,解热活性的研究

本文研究了当归提取物LIG的抗炎,解热活性.研究发现LIG(2.5,5,10 mg/kg)对角叉菜和右旋糖苷诱导的足肿胀的抑制率分别为22.2%,49.4%,76.5%和20.8%,44.2%,75.3%(P＜0.001);对棉球肉芽肿的抑制率为29.2%,44.9%,58.8%(P＜O.001);对小鼠白细胞迁移的抑制率为20.7%,35.6%,48.2%(P＜0.001).此外,LIG可降低啤酒酵母导致的高热(P＜0.001).并且,LIG的抗炎,解热作用均成剂量依赖性.以上结果显示LIG具有非常强的抗炎,解热活性.     

Changes in prostaglandin E2 (PGE2) levels in paw exudate, stomach and kidney of arthritic rats: effects of flosulide

To further examine the organ-specific toxic effects of selective and non-selective COX-2 inhibitors in adjuvant arthritis (CAA), we assessed the PGE2 concentration in various organs. AA was induced by intradermal injection of Mycobacterium butyricum. Fourteen days after inoculation, AA rats were selected and treated orally every day for two weeks with the selective COX-2 inhibitor, flosulide, or the COX-1-COX-2 inhibitor, indomethacin. The time-course of paw swelling was determined. At the end of treatments, PGE2 was extracted from paw, stomach (wall and mucosa) and kidney and its concentration was determined by ELISA. Paw edema increase was accompanied by a rise in PGE2 concentration. PGE2 also increased in stomach (mucosa and wall) and kidney. The anti-inflammatory treatment with flosulide (5 mg/kg x day), and indomethacin (1 mg/kg x day), reduced plantar edema by 98.0% and 74.4% respectively. Both drugs greatly decreased PGE2 levels in paw (73.7-53.2%), stomach wall (84.5-80.3%), stomach mucosa (109.9-110.9%) and kidney (92.9-97.5% respectively). However, PGE2 reductions in AA rats did not fall significantly below control values.     

5-Arylidene-2-imino-4-thiazolidinones: design and synthesis of novel anti-inflammatory agents

The synthesis and pharmacological activity of 5-arylidene-2-imino-4-thiazolidinones (3a-8a) are described. All derivatives exhibited significant activity levels in models of acute inflammation such as carrageenan-induced paw and pleurisy edema in rats. In particular, 5-(3-methoxyphenylidene)-2-phenylimino-3-propyl-4-thiazolidinone (3a) displayed high levels of carrageenan-induced paw edema inhibition comparable to those of indomethacin. In addition the ability of such a new class of anti-inflammatory agents to inhibit COX isoforms was assessed in murine monocyte/macrophage J774 cell line assay. 5-(4-Methoxyphenylidene)-2-phenylimino-3-propyl-4-thiazolidinone (6a), the most interesting compound in such an experiment, was docked in the known active site of COX-2 protein and showed that its 4-methoxyarylidene moiety can easily occupy the COX-2 secondary pocket considered as the critical interaction for COX-2 selectivity.     

Investigation of the nature of MIP recognition: the development and characterisation of a MIP for Ibuprofen

This paper describes the rational design, generation and testing of a molecularly imprinted polymer specific for Ibuprofen. Ibuprofen is a member of the class of drugs termed non-steroidal anti-inflammatory drugs (NSAIDS). In the present study, Ibuprofen was used as a template molecule for the preparation of molecularly imprinted polymers. A MIP has been produced which is capable of recognising Ibuprofen in aqueous media. Furthermore, Ibuprofen can be selectively extracted from aqueous conditions by molecularly imprinted solid phase extraction (MISPE). Recoveries were typically high (>80%) and good selectivity for Ibuprofen over structurally related analogues was seen. Moreover, the nature of the recognition between MIP and template has been investigated by NMR and molecular modelling to analyse whether or not it is possible to predict how well a given MIP will perform under set conditions. In addition, the physical characteristics of the MIP have been investigated including the particle size distribution on exposure of the MIP to different solvents. This has been related to the ability of the MIP to rebind Ibuprofen under the same conditions. The data from the characterisation of the MIP has been used to further enhance the understanding of the nature of MIP recognition.     

Synthesis and anti-inflammatory activity of a series of N-substituted naproxen glycolamides: nitric oxide-donor naproxen prodrugs

A series of glycolamide naproxen prodrugs containing a nitrate group as a nitric oxide (NO) donor moiety has been synthesized. These compounds were evaluated for their anti-inflammatory activity, naproxen release, and gastric tolerance. Compounds 4a, 4b, 5a, 5b, 7b, and 7c exhibited anti-inflammatory activity equivalent to that of the parent NSAID, naproxen-Na, in the rat carrageenan paw edema model. At equimolar doses relative to naproxen-Na, the NO-donor glycolamide derivatives 4a, 4b, 5a, 5b, 7b, and 7c were gastro-sparing in the rat. Naproxen formation from these NO-donor glycolamides varied among the structures examined, with the N-substituent on the amide group having a particular influence, and demonstrated their prodrug nature. Compound 7b was selected for exemplary demonstration that the glycolamide nitrates can be bioactivated to release NO. These data open the possibility that naproxen glycolamide nitrates may represent a safer alternative to naproxen as anti-inflammatory medicines.     

Mechanisms Involved in the Anti-Inflammatory Action of a Polysulfated Fraction from Gracilaria cornea in Rats

The anti-inflammatory mechanisms of the sulfated polysaccharidic fraction obtained from red marine alga Gracilaria cornea (Gc-FI) were investigated using a paw edema model induced in rats by different inflammatory agents (carrageenan, dextran, serotonin, bradykinin, compound 48/80 or L-arginine). Gc-FI at the doses of 3, 9 or 27 mg/kg, subcutaneously - s.c., significantly inhibited rat paw edema induced by carrageenan and dextran, as confirmed by myeloperoxidase and Evans’ blue assessments, respectively. Gc-FI (9 mg/kg, s.c.) inhibited rat paw edema induced by histamine, compound 48/80 and L-arginine. Additionally, Gc-FI (9 mg/kg, s.c.) inhibited Cg-induced edema in animals with intact mast cells but did not inhibit that with degranulated mast cells by compound 48/80, revealing a protective role on mast cell membranes. Gc-FI down-regulated the IL-1β, TNF-α and COX-2 mRNA and protein levels compared with those of the carrageenan group, based on qRT-PCR and immunohistochemistry analyses. After inhibition with ZnPP IX, a specific heme oxygenase-1 (HO-1) inhibitor, the anti-inflammatory effect of Gc-FI was not observed in Cg-induced paw edema, suggesting that the anti-inflammatory effect of Gc-FI is, in part, dependent on the integrity of the HO-1 pathway. Gc-FI can target a combination of multiple points involved in inflammatory phenomena.     

pH-sensitive niosomes: Effects on cytotoxicity and on inflammation and pain in murine models

pH-sensitive nonionic surfactant vesicles (niosomes) by polysorbate-20 (Tween-20) or polysorbate-20 derivatized by glycine (added as pH sensitive agent), were developed to deliver Ibuprofen (IBU) and Lidocaine (LID). For the physical-chemical characterization of vesicles (mean size, size distribution, zeta potential, vesicle morphology, bilayer properties and stability) dynamic light scattering (DLS), small angle X-ray scattering and fluorescence studies were performed. Potential cytotoxicity was evaluated on immortalized human keratinocyte cells (HaCaT) and on immortalized mouse fibroblasts Balb/3T3. In vivo antinociceptive activity (formalin test) and anti-inflammatory activity tests (paw edema induced by zymosan) in murine models were performed on drug-loaded niosomes. pH-sensitive niosomes were stable in the presence of 0 and 10% fetal bovine serum, non-cytotoxic and able to modify IBU or LID pharmacological activity in vivo. The synthesis of stimuli responsive surfactant, as an alternative to add pH-sensitive molecules to niosomes, could represent a promising delivery strategy for anesthetic and anti-inflammatory drugs.     

Design,synthesis and pharmacological evaluation of novel tetrasubstituted thiophene analogues as anti-inflammatory agents

A new series of tetrasubstituted thiophene analogues (4a-4f, 5a-5f and 8a-8i) were designed incorporating the pharmacophoric features of COX-1 (as in fenamates), 5-LOX and the p38 MAP kinase inhibitors. The designed series was synthesized by nucleophilic addition of aryl/aroylisothiocyanate and enamine (2) yielding the addition product l-(α-Carbomethoxy-β-aminothiocrotonoyl)-aryl/aroyl amines (3/7); which on reaction with substituted phenacyl bromides gave the targeted tetrasubstituted thiophene esters (4a-4f / 8a-8i). The tetrasubstituted thiophenes esters (4a-4f ) on hydrolysis with one equivalent of potassium hydroxide solution in methanol at room temperature gave corresponding acids (5a-5f ). All the targeted compounds were evaluated for their anti-inflammatory activity in carrageenin-induced rat hind paw oedema model at the doses of 10, 20 and 40 mg/kg body weight using standard drugs mefanamic acid and ibuprofen. The compounds (4c, 4e, 4f, 5f, 8a- 8i) which gave reasonable protection to the inflamed paw, eliciting good or moderate comparable anti-inflammatory activity were selected for investigating their analgesic activity using acetic acid induced writhing response test in albino mice at 10 mg/kg dose using standard drug ibuprofen and in order to arrive at possible mechanism of their anti-inflammatory activity, in vitro antioxidant nitric oxide radical scavenging assay at the concentrations of 5, 10, 15, 20, 25, 30 and 35 μg/mL were performed using standard drug ascorbic acid.     

Synthesis and anti-inflammatory activity of fructigenine a derivatives

Several derivatives were synthesized from fructigenine A, which was isolated fromPenicillium fructigenum. The anti-inflammatory properties of fructigenine A was evaluatedin vivo with a 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced mouse ear edema model and a carrageenan-induced rat paw edema model. Results showed that the anti-in flammatory activity was significantly higher with fructigenine derivatives than with indomethacin, which was used as a standard. We concluded that fructigenine derivatives could exert an anti-inflammatory effect.     

Anti-inflammatory and antinociceptive potential of major phenolics from Verbascum salviifolium Boiss

The potential effects of flavonoids, phenylethanoid and neolignan glycosides from the aerial parts of Verbascum salviifolium Boiss. were studied in the p-benzoquinone-induced writhing reflex, for the assessment of the antinociceptive activity, and in carrageenan- and PGE1-induced hind paw edema and 12-O-tetradecanoyl-13-acetate (TPA)-induced ear edema models in mice, for the assessment of the anti-inflammatory activity. Through bioassay-guided fractionation and isolation procedures ten compounds from the aqueous extract of the plant, luteolin 7-O-glucoside (1), luteolin 3'-O-glucoside (2), apigenin 7-O-glucoside (3), chrysoeriol 7-O-glucoside (4), beta-hydroxyacteoside (5), martynoside (6), forsythoside B (7), angoroside A (8), dehydrodiconiferyl alcohol-9'-O-beta-D-glucopyranoside (9) and dehydrodiconiferyl alcohol-9-O-beta-D-glucopyranoside (10), were isolated and their structures were elucidated by spectral techniques. Results have shown that 1, 2, 3 and 5 significantly inhibited carrageenan-induced paw edema at a 200 mg/kg dose, while 1, 2 and 5 also displayed anti-inflammatory activity against the PGE1-induced hind paw edema model. However, all the compounds showed no effect in the TPA-induced ear edema model. The compounds 1 and 2 also exhibited significant antinociceptive activity.     

Pimaradienoic Acid Inhibits Carrageenan-Induced Inflammatory Leukocyte Recruitment and Edema in Mice: Inhibition of Oxidative Stress,Nitric Oxide and Cytokine Production

Pimaradienoic acid (PA; ent-pimara-8(14),15-dien-19-oic acid) is a pimarane diterpene found in plants such as Vigueira arenaria Baker (Asteraceae) in the Brazilian savannas. Although there is evidence on the analgesic and in vitro inhibition of inflammatory signaling pathways, and paw edema by PA, its anti-inflammatory effect deserves further investigation. Thus, the objective of present study was to investigate the anti-inflammatory effect of PA in carrageenan-induced peritoneal and paw inflammation in mice. Firstly, we assessed the effect of PA in carrageenan-induced leukocyte recruitment in the peritoneal cavity and paw edema and myeloperoxidase activity. Next, we investigated the mechanisms involved in the anti-inflammatory effect of PA. The effect of PA on carrageenan-induced oxidative stress in the paw skin and peritoneal cavity was assessed. We also tested the effect of PA on nitric oxide, superoxide anion, and inflammatory cytokine production in the peritoneal cavity. PA inhibited carrageenan-induced recruitment of total leukocytes and neutrophils to the peritoneal cavity in a dose-dependent manner. PA also inhibited carrageenan-induced paw edema and myeloperoxidase activity in the paw skin. The anti-inflammatory mechanism of PA depended on maintaining paw skin antioxidant activity as observed by the levels of reduced glutathione, ability to scavenge the ABTS cation and reduce iron as well as by the inhibition of superoxide anion and nitric oxide production in the peritoneal cavity. Furthermore, PA inhibited carrageenan-induced peritoneal production of inflammatory cytokines TNF-α and IL-1β. PA presents prominent anti-inflammatory effect in carrageenan-induced inflammation by reducing oxidative stress, nitric oxide, and cytokine production. Therefore, it seems to be a promising anti-inflammatory molecule that merits further investigation.