Discovery of imidazo[1,2-a]pyridine-based anthelmintic targeting cholinergic receptors of Haemonchus contortus.

We report the synthesis of a series of imidazo[1,2-a]pyridine-based molecules as anthelmintic against the livestock parasite Haemonchus contortus. The molecules were tested by using Larval Paralysis Test (LPT), in order to target ionic channels, as most of the prominent marketed anthelminthics present such mechanism of action. The most active compound (5e) displayed paralysis on H. contortus stage 3 larvae until 31.25 µM. Effect of 5e on H. contortus cholinergic receptors (L-AChR1 and 2) was characterized via electrophysiological measurement and a rare antagonist mode of action was unveiled.

Antibacterial Polyketide Heterodimers from Pyrenacantha kaurabassana Tubers.

Two heterodimers comprising anthraquinone and methylbenzoisocoumarin moieties (1 and 2) were isolated, together with emodin and physcion from the tubers of Pyrenacantha kaurabassana. The structures of 1 and 2 were established by NMR spectroscopy, including the analysis of a 2D INADEQUATE spectrum. On the basis of the data obtained, the structures that were previously proposed in the literature for these compounds were revised. Compounds 1 and 2 showed antibacterial activity against three different strains of Staphylococcus aureus. Compound 2 also showed bactericidal activity against Helicobacter pylori.

Regiocontrolled microwave assisted bifunctionalization of 7,8-dihalogenated imidazo[1,2-a]pyridines: a one pot double-coupling approach.

The reactivity of the 7-chloro-8-iodo- and 8-chloro-7-iodoimidazo[1,2-a]pyridines 1a-e diversely substituted on the 2 position, towards Suzuki-Miyaura, Sonogashira, and Buchwald-Hartwig cross-coupling reactions as well as cyanation was evaluated. Various methodologies are proposed to introduce aryl, heteroaryl, alkyne, amine or cyano groups in the two positions depending on the nature of the substituent present in position 2. In both series, the substitution of the iodine atom was totally regioselective and the difficulty was to substitute the chlorine atom in a second step. Until now, only hetero(aryl) groups could be introduced though Suzuki-Miyaura cross-coupling. We overcame this problem evaluating both regioisomers in parallel. The double coupling approach was also studied allowing the one pot Suzuki/Suzuki, cyanation/Sonogashira and cyanation/Buchwald reactions leading to polyfunctionnalized imidazo[1,2-a]pyridines.

Copper(I)-mediated preparation of new pyrano[3',4':4,5]imidazo[1,2-a]pyridin-1-one compounds under mild palladium-free conditions.

A general and efficient Cu(I)-mediated cross-coupling and heterocyclization reaction of 3-iodoimidazo[1,2-a]pyridine-2-carboxylic acid, and terminal alkynes was developed under very mild conditions. This method allows the introduction in one pot of a third ring fused in positions 2 and 3 of the imidazo[1,2-a]pyridine core with reasonable yields and total regioselectivity. This procedure does not require the use of any expensive supplementary additives, and is palladium-free.

Proerectile effects of dopamine D2-like agonists are mediated by the D3 receptor in rats and mice.

Dopamine D(2)-like agonists induce penile erection (PE) and yawning in a variety of species, effects that have been suggested recently to be specifically mediated by the D(4) and D(3) receptors, respectively. The current studies were aimed at characterizing a series of D(2), D(3), and D(4) agonists with respect to their capacity to induce PE and yawning in the rat and the proerectile effects of apomorphine [(R)-(-)-5,6,6a,7-tetrahydro-6-methyl-4H-dibenzo-[de,g]quinoline-10,11-diol hydrochloride] in wild-type and D(4) receptor (R) knockout (KO) mice. All D(3) agonists induced dose-dependent increases in PE and yawning over a similar range of doses, whereas significant increases in PE or yawning were not observed with any of the D(4) agonists. Likewise, D(2), D(3), and D(4) antagonists were assessed for their capacity to alter apomorphine- and pramipexole (N'-propyl-4,5,6,7-tetrahydrobenzothiazole-2,6-diamine dihydrochloride)-induced PE and yawning. The D(3) antagonist, PG01037 [N-{4-[4-(2,3-dichlorophenyl)-piperazin-1-yl]-trans-but-2-enyl}-4-pyridine-2-yl-benzamide hydrochloride], inhibited the induction of PE and yawning, whereas the D(2) antagonist, L-741,626 [3-[4-(4-chlorophenyl)-4-hydroxypiperidin-l-yl]methyl-1H-indole], reversed the inhibition of PE and yawning observed at higher doses. The D(4) antagonist, L-745,870 [3-(4-[4-chlorophenyl]piperazin-1-yl)-methyl-1H-pyrrolo[2,3-b]pyridine trihydrochloride], did not alter apomorphine- or pramipexole-induced PE or yawning. A role for the D(3) receptor was further supported because apomorphine was equipotent at inducing PE in wild-type and D(4)RKO mice, effects that were inhibited by the D(3) antagonist, PG01037, in both wild-type and D(4)R KO mice. Together, these studies provide strong support that D(2)-like agonist-induced PE and yawning are differentially mediated by the D(3) (induction) and D(2) (inhibition) receptors. These studies fail to support a role for the D(4) receptor in the regulation of PE or yawning by D(2)-like agonists.

Influence of 6- or 8-substitution on the antiviral activity of 3-arylalkylthiomethylimidazo[1,2-a]pyridine against human cytomegalovirus (CMV) and varicella-zoster virus (VZV): part II.

The synthesis of original imidazo[1,2-a]pyridines bearing a thioether side chain at the 3 position and diversely substituted on the 6 or 8 position, and their antiviral activities are reported. From the synthesized compounds, the imidazo[1,2-a]pyridines bearing a 5 membered heterocycle (thiophene, furane or pyrrole) in the 6 position or a phenylthio group in the 6 or 8 position were the most potent against human cytomegalovirus (CMV) and varicella-zoster virus (VZV), whereas several other congeners, while less potent, were more selective in their inhibitory activity against VZV and CMV. These compounds showed similar activity against thymidine kinase competent (TK+) and deficient (TK-) VZV strains, demonstrating a mechanism of action independent of the viral thymidine kinase.

Imidazo[1,2-b]pyridazines targeting Toxoplasma gondii calcium-dependent protein kinase 1 decrease the parasite burden in mice with acute toxoplasmosis.

The current therapeutic arsenal for toxoplasmosis is restricted to drugs non-specific to the parasite which cause important side effects. Development of more efficient and specific anti-Toxoplasma compounds is urgently needed. Imidazo[1,2-b]pyridazines designed to inhibit the calcium-dependent protein kinase 1 of Toxoplasma gondii (TgCDPK1) and effective against tachyzoite growth in vitro at submicromolar ranges were modified into hydrochloride salts to be administered in vivo in a mouse model of acute toxoplasmosis. All protonated imidazo[1,2-b]pyridazine salts (SP230, SP231 and SP232) maintained their activity on TgCDPK1 and T. gondii tachyzoites. Rat and mouse liver microsomes were used to predict half-life and intrinsic clearance, and the pharmacokinetic profile of the most rapidly degraded imidazo[1,2b]pyridazine salt (SP230) was determined in serum, brain and lungs of mice after a single administration of 50 mg/kg. Compounds were then tested in vivo in a murine model of acute toxoplasmosis. Mice infected with tachyzoites of the ME49 strain of T. gondii were treated for 4, 7 or 8 days with 25 or 50 mg/kg/day of SP230, SP231 or SP232. The parasite burdens were strongly diminished (>90% reduction under some conditions) in the spleen and the lungs of mice treated with imidazo[1,2-b]pyridazine salts compared with untreated mice, without the need for pre-treatment. Moreover, no increases in the levels of hepatic and renal toxicity markers were observed, demonstrating no significant signs of short-term toxicity. To conclude, imidazo[1,2-b]pyridazine salts have strong efficacy in vivo on acute toxoplasmosis and should be further tested in a model of mouse congenital toxoplasmosis.

Recent progress in the pharmacology of imidazo[1,2-a]pyridines.

Imidazo[1,2-a]pyridine is a bicyclic system with a bridgehead nitrogen atom, of growing interest in medicinal chemistry. The paper deals with the recent progress realised in the comprehension of the pharmacological properties of this scaffold. From the many imidazo[1,2-a]pyridine analogues described in the literature, those discussed herein will be presented in three parts concerning first the enzyme inhibitors, then the receptor ligands and finally the anti-infectious agents.

Recent progress in medicinal chemistry of D4 agonists.

In the last decades, the physiological and pharmacological properties of dopamine receptors were controversial principally because of the lack of selective ligand for some receptor subtypes. Since 1997, some specific D4 agonists have been described and have allowed a therapeutic approach. We report here, compounds described as D4 agonist and when available the SAR. The major studies for physiological implications and their potential biological applications are also reported and principally their interest in erectile dysfunction.

2-[(4-phenylpiperazin-1-yl)methyl]imidazo(di)azines as selective D4-ligands. Induction of penile erection by 2-[4-(2-methoxyphenyl)piperazin-1-ylmethyl]imidazo[1,2-a]pyridine (PIP3EA), a potent and selective D4 partial agonist.

A series of novel 2-[(4-phenylpiperazin-1-yl)methyl]imidazoazines and aza-analogues were prepared and screened at selected dopamine, serotonin, and adrenergic receptor subtypes. 2-Substituted imidazopyridines and pyridazines presented high affinities and selectivities for D4 dopamine receptors. Whereas functional experiments indicated neutral antagonists or weak partial agonist effects for most of the target compounds, the 2-methoxyphenyl substituted 2-piperazinylmethylimidazopyridine 3c (PIP3EA) displayed substantial agonist efficacy in mitogenesis experiments and GTPgammaS binding tests, resulting in EC50 values of 3.0 (46%) and 4.5 nM (57%), respectively. Our D4 agonist 3c induced penile erection in vivo when administered to rats. This effect was inhibited by L-745,870 a D4 selective antagonist, confirming the mechanistic pathway.

Two biflavonoids from Ouratea nigroviolacea.

The leaves of Ouratea nigroviolacea (Ochnaceae) afforded two biflavonoids, ouratine A and B together with agathisflavone and stigmasterol. The biflavonoids were characterized as 4'-O-methylated apigeninyl-(I-6, II-8)-4'-O-methylatedapigenin and 4'-O-methylated apigeninyl-(I-6, II-8) apigenin by spectral and chemical transformation studies.

2,3-Diarylimidazo[1,2-a]pyridines as potential inhibitors of UV-induced keratinocytes apoptosis: synthesis, pharmacological properties and interactions with model membranes and oligonucleotides by NMR.

Four 2,3-diarylimidazo[1,2-a]pyridines (I, 1a-c) were synthesized as inhibitors of UV-induced apoptosis and showed quite different properties. First, only the pyridinyl derivative I showed protection in molt cells. From the supposed intracellular target, phospholipid membrane models were studied by (1)H, (2)H and (31)P NMR spectroscopy. All these molecules can incorporate the membrane bilayer of small unilamellar vesicles of lecithin (SUV). However, I is clearly closed to the external polar head of the lipids, and is relatively mobile in the layer. Conversely, the other molecules are strongly immobilized in the deep part of the external layer. (31)P solid-state NMR spectra recorded on phospholipid dispersions (multilayers vesicles (MLV)) completely excluded any detergent effect or any modification of temperature transition. The only structural or dynamic effect observed was a homogeneous, but limited, reduction in the chemical shift anisotropy in the presence of I, in agreement with its superficial location. (2)H NMR experiment performed on the same model using perdeuterated phospholipids showed no significant fluidity reduction at the level of terminal CD(3) groups in the presence of 1a-c, according to their deep location. Finally, their interactions with synthetic oligonucleotide, d(CGATCG)(2) was studied showing non specific interactions of 1a on the external GC pair, while no interaction was observed with the other derivatives.

Development of new highly potent imidazo[1,2-b]pyridazines targeting Toxoplasma gondii calcium-dependent protein kinase 1.

Using a structure-based design approach, we have developed a new series of imidazo[1,2-b]pyridazines, targeting the calcium-dependent protein kinase-1 (CDPK1) from Toxoplasma gondii. Twenty derivatives were thus synthesized. Structure-activity relationships and docking studies confirmed the binding mode of these inhibitors within the ATP binding pocket of TgCDPK1. Two lead compounds (16a and 16f) were then identified, which were able to block TgCDPK1 enzymatic activity at low nanomolar concentrations, with a good selectivity profile against a panel of mammalian kinases. The potential of these inhibitors was confirmed in vitro on T. gondii growth, with EC50 values of 100 nM and 70 nM, respectively. These best candidates also displayed low toxicity to mammalian cells and were selected for further in vivo investigations on murine model of acute toxoplasmosis.

A small-molecule cell-based screen led to the identification of biphenylimidazoazines with highly potent and broad-spectrum anti-apicomplexan activity.

An in vitro screening of the anti-apicomplexan activity of 51 compounds, stemming from our chemical library and from chemical synthesis, was performed. As a study model, we used Toxoplasma gondii (T. gondii), expressing ß-galactosidase for the colorimetric assessment of drug activity on parasites cultivated in vitro. This approach allowed the validation of a new series of molecules with a biphenylimidazoazine scaffold as inhibitors of T. gondii growth in vitro. Hence, 8 molecules significantly inhibited intracellular replication of T. gondii in vitro, with EC50 < 1 µM, while being non-toxic for human fibroblasts at these concentrations. Most attractive candidates were then selected for further biological investigations on other apicomplexan parasites (Neospora caninum, Besnoitia besnoiti, Eimeria tenella and Plasmodium falciparum). Finally, two compounds were able to inhibit growth of four different apicomplexans with EC50 in the submicromolar to nanomolar range, for each parasite. These data, including the broad anti-parasite spectrum of these inhibitors, define a new generation of potential anti-parasite compounds of wide interest, including for veterinary application. Studies realized on E. tenella suggest that these molecules act during the intracellular development steps of the parasite. Further experiments should be done to identify the molecular target(s) of these compounds.

Synthesis and antiviral activities of 3-aralkylthiomethylimidazo[1,2-b]pyridazine derivatives.

The synthesis of novel substituted 3-aralkylthiomethylimidazo[1,2-b]pyridazines is reported. All of the synthesized compounds are devoid of antiviral activity against the replication of human immunodeficiency virus. However, compounds 6-chloro-8-methyl-3-phenethylthioimidazo[1,2-b]pyridazine and 6-chloro-2-methyl-3-phenethylthioimidazo[1,2-b]pyridazine are potent inhibitors of the replication of human cytomegalovirus in vitro, while compounds 6-chloro-2-methyl-3-benzylthiomethylimidazo[1,2-b]pyridazine and 6-chloro-2-methyl-3-phenethyl-thioimidazo[1,2-b]pyridazineare inhibitors of the replication of varicella-zoster virus. The results presented here suggest that compound 10 should be considered as a new lead in the development of antiviral agents.

New flavonoids C-glycosides from Rhabdophyllum arnoldianum.

Two new flavone glycosides, 3″-O-acetyl-7-O-methylvitexin (1) and 6″-α-rhamnopyranosyl-7-O-methylvitexin (2), along with nine known compounds (3-11) were isolated from the leaves of Rhabdophyllum arnoldianum (Ochnaceae). The structures of the new compounds were established by detailed spectroscopic studies and mass spectrometry, while known compounds were characterised by direct comparison of their reported NMR data with those found in the literature. All these compounds were the first reported from Rhabdophyllum genus. The biological assays on crude extracts and compounds of this plant demonstrated that the crude extracts possess significant antimicrobial activity against Gram-positive bacteria.

High performance thin layer chromatography-densitometry: a step further for quality control of cranberry extracts.

A method for selective quantitation of catechin, proanthocyanidin (PAC) A2 and PAC-B1 in American cranberry (Vaccinium macrocarpon) extracts using high performance thin layer chromatography (HPTLC)-densitometry is presented. Methylene chloride/ethyl acetate/formic acid (6:10:1, v/v) as the mobile phase and 1% vanillin hydrochloric solution as staining reagent were used. In these conditions, three standards considered as quality markers, catechin, PAC-A2 and PAC-B1, were well resolved allowing simultaneous quantitation on one plate. All standards were quantified in the range of 0.7-5 µg with RSD of repeatability and intermediate precision not exceeding 5%. Catechin, PAC-A2 and PAC-B1 profiles of cranberry extracts were analysed regarding global PAC amounts obtained by BL-DMAC assay. It appears clearly that HPTLC-densitometry process provides additional information which, combined with BL-DMAC results, allows qualitative and quantitative control of cranberry extracts. Particularly, densitometric assay highlighted degradation of PACs in a 7 day-extract, leading to high overestimation with BL-DMAC protocol.

3-Biphenylimidazo[1,2-a]pyridines or [1,2-b]pyridazines and analogues, novel Flaviviridae inhibitors.

Using Ttou 84 as starting point, a novel class of biphenyl derivatives of imidazo[1,2-a]pyridine and imidazo[1,2-b]pyridazine was designed to optimize the inhibitory properties on the replication of the bovine viral diarrhoea virus (BVDV) and hepatitis C virus (HCV). Three sites of pharmacomodulation were chosen i.e. positions 2, 3 and 6 on the central heterocyclic core structure. From the 49 analogues tested, only compound 18j (3-(2'-hydroxybiphen-3-yl)-2-(2-methoxyphenyl)-6-(thien-3-yl)imidazo[1,2-b]pyridazine) showed antiviral activity in the HCV replicon system reminiscent of selective inhibition (60-70% inhibition). Compound 4f (3-(biphen-3-yl)-2-(4-fluorophenyl)-6-phenylthioimidazo[1,2-a]pyridine) proved to be the most selective inhibitor of BVDV replication and showed no or only marginal cross-resistance with known inhibitors of pestivirus replication. The cross-resistance profile of 4f might indicate that 4f does not interact with the same binding site as BPIP, VP32947, AG110 or LZ37. From 42 analogues tested against both viruses, QSAR studies were discussed in regard to BVDV antiviral activity.

Synthesis and antiviral activity of an imidazo[1,2-a]pyrrolo[2,3-c]pyridine series against the bovine viral diarrhea virus.

A series of imidazo[1,2-a]pyrrolo[2,3-c]pyridines has been prepared and evaluated for their anti-BVDV activities in MDBK cells. From the synthesized analogues bearing modifications of the substituents at positions 2, 3, 7 and 8, compounds 10a, b, 16, 24, 25 and 26 exhibited significant anti-BVDV activities.

Physicochemical properties and membrane interactions of anti-apoptotic derivatives 2-(4-fluorophenyl)-3-(pyridin-4-yl)imidazo[1,2-a]pyridine depending on the hydroxyalkylamino side chain length and conformation: an NMR and ESR study.

Three imidazo[1,2-a]pyridine derivatives 3a-c have been synthesized from p38 kinase inhibitor structures and evaluated as anti-apoptosis agents. These drugs were designed to interact with nucleic acids and membrane interactions by varying the chain length in position 6, from hydroxyethylamino (3a), to hydroxybutylamino (3b) and hydroxyhexylamino (3c). First experiments showed that 3a and 3b were insoluble in water while 3c could be solubilized in water despite its partition coefficient (logP=3.2). This latter feature was explained by the formation of a fifth intramolecular cycle thus allowing supramolecular structure formation (NMR and MD calculations). The interactions with membranes have been studied using (1)H, (2)H, (31)P Nuclear Magnetic Resonance (NMR), Electron Spin Resonance (ESR) and High Resolution-Magic Angle Spinning (HR-MAS). Despite the insolubility of 3a and 3b in water, these derivatives could be partially solubilized by synthetic phospholipidic model membranes (small unilamellar vesicles, SUV). (1)H NMR paramagnetic broadening experiments performed on the same models showed that 3a was located in the external layer, probably close to the surface while 3b only formed external superficial adducts. Supplementary (31)P, (2)H NMR and ESR experiments on phospholipid dispersions confirmed the location of 3a close to the polar headgroup of the external layer of the membrane, this resulting in a 2K lowering of the transition temperature. Moreover, no significant interaction was detected on the deep part of the layer ((2)H NMR and 16NS ESR experiments). This binding was also found in the presence of cell cultures, as revealed by HR-MAS NMR experiments. Conversely, no significant interaction with membranes was found with 3b or 3c. From both the unexpected solubility of 3c and 3a interactions with membranes, further chemical modifications were finally proposed.