Benjamin Boucherle

Discovery of alpha-aminoazaheterocycle-methylglyoxal adducts as a new class of high-affinity inhibitors of cystic fibrosis transmembrane conductance regulator chloride channels.

The cystic fibrosis transmembrane conductance regulator (CFTR) represents the main Cl– channel in the apical membrane of epithelial cells for cAMP-dependent Cl– secretion. Here we report on the synthesis and screening of a small library of nontoxic α-aminoazaheterocycle-methylglyoxal adducts, inhibitors of wild-type (WT) CFTR and G551D-, G1349D-, and F508del-CFTR Cl– channels. In whole-cell patch-clamp experiments of Chinese hamster ovary (CHO) cells expressing WT-CFTR, we recorded rapid and reversible inhibition of forskolin-activated CFTR currents in the presence of the adducts 5a and 8a,b at 10 pM concentrations. Using iodide efflux experiments, we compared concentration-dependent inhibition of CFTR with glibenclamide (IC50 = 14.7 μM), 3-[(3-trifluoromethyl)phenyl]-5-[(4-carboxyphenyl-)methylene]-2-thioxo-4-thiazolidinone (CFTRinh-172) (IC50 = 1.2 μM), and α-aminoazaheterocycle-methylglyoxal adducts and identified compounds 5a (IC50 = 71 pM), 8a,b (IC50 = 2.5 nM), and 7a,b (IC50 = 3.4 nM) as the most potent inhibitors of WT-CFTR channels. Similar ranges of inhibition were also found when these compounds were evaluated on CFTR channels with the cystic fibrosis mutations F508del (in temperature-corrected human airway epithelial F508del/F508del CF15 cells)-, G551D-, and G1349D-CFTR (expressed in CHO and COS-7 cells). No effect of compound 5a was detected on the volume-regulated or calcium-regulated iodide efflux. Picomolar inhibition of WT-CFTR with adduct 5a was also found using a 6-methoxy-N-(3-sulfopropyl)-quinolinium fluorescent probe applied to the human tracheobronchial epithelial cell line 16HBE14o–. Finally, we found comparable inhibition by 5a or by CFTRinh-172 of forskolin-dependent short-circuit currents in mouse colon. To the best of our knowledge, these new nontoxic α-aminoazaheterocycle-methylglyoxal adducts represent the most potent compounds reported to inhibit CFTR chloride channels.

Identification of PDZ ligands by docking-based virtual screening for the development of novel analgesic agents.

Disrupting the interaction between the PDZ protein, PSD-95, and its target ligands (such as the glutamate NMDA receptor or the serotonin 5-HT2A receptor) was found to reduce hyperalgesia in various models of neuropathic pain. Here, we set out to identify lead molecules which would interact with PSD-95, and hence, would potentially display analgesic activity. We describe the virtual screening of the Asinex and Cambridge databases which together contain almost one million molecules. Using three successive docking filters and visual inspection, we identified three structural classes of molecules and synthesized a potential lead compound from each class. The binding of the molecules with the PDZ domains of PSD-95 was assessed by (1)H-(15)N HSQC NMR experiments. The analgesic activity of the best ligand, quinoline 2, was evaluated in vivo in a model of neuropathic pain and showed promising results.

Structure-Based Design of PDZ Ligands as Inhibitors of 5-HT2A Receptor/PSD-95 PDZ1 Domain Interaction Possessing Anti-hyperalgesic Activity

Disrupting the interaction between the PDZ protein PSD-95 and the C-terminal domain of the 5-HT2A serotonin receptor has been shown to reduce hyperalgesia in a rodent model of neuropathic pain. Here, we designed and synthesized PDZ ligands capable of binding to the first PDZ domain (PDZ1) of the PSD-95 protein and evaluated their biological activity in vitro and in vivo. A series of substituted indoles was identified by docking simulations, and six novel analogues were synthesized. Three analogues displayed strong interactions with the first PDZ domain (PDZ1) of PDZ-95 in (1)H-(15)N heteronuclear single-quantum coherence (HSQC) experiments and two of them were able to inhibit the interaction between PSD-95 and the 5-HT2A receptor in vitro. We identified compound 8b as the analogue able to significantly suppress mechanical hyperalgesia in an experimental model of traumatic neuropathic pain in the rat. This effect was suppressed by the coadministration of the 5-HT2A receptor antagonist M100907, consistent with an inhibitory effect upon 5-HT2A receptor/PSD-95 interaction. Finally, we determined an NMR-restraint driven model structure for the PSD95 PDZ1/8b complex, which confirms that indole 8b binds to the putative PDZ-ligand binding site.

NMR evaluation of interactions between substituted-indole and PDZ1 domain of PSD-95

We synthesized small organic molecules designed as PDZ ligands. These indole-based compounds were evaluated for their interaction with the PDZ1 domain of the post-synaptic density 95 (PSD-95) protein. Three molecules were found to interact with the targeted PDZ protein by NMR. One of them showed chemical shift perturbations closely related to the natural ligands.

Synthesis and evaluation of bidentate ligands designed to interact with PDZ domains

We designed bidentate ligands to target PDZ domains through two binding sites: site S0, delimited by the GLGF loop, and site S1, a zone situated around loop β(B)/β(C). A molecular docking study allowed us to design a generic S0 binder, to which was attached a variable size linker, itself linked to an amino acid aimed to interact with the S1 site of PDZ domains. A series of 15 novel bidentate ligands was prepared in 6-11 steps in good overall yield (24-43%). Some of these ligands showed an inhibitory activity against serotonin 5-HT2A receptor/PSD-95 interaction. This was assessed by pull-down assay using a synthetic decapeptide corresponding to the C-terminal residues of the receptor as a bait.

Traditional uses, phytochemistry and pharmacological properties of African Nauclea species: A review

ETHNOPHARMACOALOGICAL RELEVANCE The genus Nauclea in Africa comprises seven species. Among them, N. latifolia, N. diderrichii and N. pobeguinii are widely used by the local population in traditional remedies. Preparation from various parts of plants (e.g. roots, bark, leaves) are indicated by traditional healers for a wide range of diseases including malaria, pain, digestive ailments or metabolic diseases. MATERIALS AND METHODS A literature search was conducted on African species of the genus Nauclea using scientific databases such as Google Scholar, Pubmed or SciFinder. Every document of ethnopharmacological, phytochemical or pharmacological relevance and written in English or French were analyzed. RESULTS AND DISCUSSION The Nauclea genus is used as ethnomedicine all along sub-Saharan Africa. Several local populations consider Nauclea species as a major source of remedies for malaria. In this regard, two improved traditional medicines are currently under development using extracts from N. latifolia and N. pobeguinii. Concerning the chemical composition of the Nauclea genus, indoloquinolizidines alkaloids could be considered as the major class of compounds as they are reported in every analyzed Nauclea species, with numerous structures identified. Based on traditional indications a considerable amount of pharmacological studies were conducted to ensure activity and attempt to link them to the presence of particular compounds in plant extracts. CONCLUSION Many experimental studies using plant extracts of the African species of the genus Nauclea validate traditional indications (e.g. malaria and pain). However, bioactive compounds are rarely identified and therefore, there is a clear need for further evaluations as well as for toxicity experiments. The sustainability of these plants, especially of N. diderrichii, a threatened species, should be kept in mind to adapt local uses and preparation modes of traditional remedies.

A One-Pot Synthesis of Highly Functionalized Purines

Highly substituted purines were synthesized in good to high yields through a one-pot straightforward metal-free scalable method, using the Traube synthesis adapted to Vilsmeier-type reagents. From 5-amino-4-chloropyrimidines, new 9-aryl-substituted chloropurines and intermediates for peptide nucleic acid synthesis were prepared. Variant procedures allowing a rapid synthesis of ribonucleosides and 7-benzylpurine from 5-amidino-6-aminopyrimidines are also reported to illustrate the high potential of this versatile toolbox. This route appears to be particularly interesting in the field of nucleic acids for a direct and rapid access to various new 8-alkylpurine nucleosides.