Analogues of FAUC 73 revealing new insights into the structural requirements of nonaromatic dopamine D3 receptor agonists

Employing the nonaromatic D3 agonist FAUC 73 as a lead compound, the dopaminergic enynes 1a,b and the diene 2 (FAUC 206) were synthesized via palladium-catalyzed cross-coupling. FAUC 206 showed remarkable D3 affinity and enhanced selectivity over D4 when compared to the lead compound. To learn more about the bioactive structure of the diene moiety, computational studies including DFT-based conformational analysis and calculations of the magnetic shielding properties were performed. The electrostatic properties of the pharmacophoric pi-systems were visualized by diagnostic MEP maps.     

Fancy bioisosteres: synthesis and dopaminergic properties of the endiyne FAUC 88 as a novel non-aromatic D3 agonist

Enlargement of the pi-electronic system of the non-aromatic D3 agonist FAUC 73 led to dopaminergic endiynes of type 1 being synthesized via the bromovinyl triflate 7a as a key intermediate when palladium catalyzed coupling reactions were exploited for the introduction of the (aza)alkyne substituents. As the first neuroreceptor active endiyne, FAUC 88 (1c) displayed high and selective dopamine D3 receptor affinity (K(ihigh) = 3.2 nM) and substantial ligand efficacy (72%, EC(50) = 2.5 nM). Similarities between molecular electrostatic potentials induced by the catechol subunit of the genuine neurotransmitter and those of its non-aromatic endiyne bioisostere are discussed.     

Synthesis and biological investigations of dopaminergic partial agonists preferentially recognizing the D4 receptor subtype

Aminomethyl-substituted biaryls bearing a pyrazole or triazole moiety were synthesized and investigated for dopamine and serotonin receptor binding. The N-arylpyrazoles 3b,f,g and 4 revealed Ki values in the subnanomolar range (0.28-0.70 nM) for the dopamine D4 receptor subtype. Employing both mitogenesis and GTPgammaS assays, ligand efficacy was evaluated indicating partial agonist properties. Interestingly, the tetrahydropyrimidine 4 (FAUC 2020) displayed significant intrinsic selectivity for D2(long) over D2(short).     

Di- and trisubstituted pyrazolo[1,5-a]pyridine derivatives: synthesis, dopamine receptor binding and ligand efficacy

Based on the lead molecule FAUC 113, a series of di- and trisubstituted pyrazolo[1,5-a]pyridine derivatives was synthesized and investigated for their dopamine receptor binding profile. The carbonitrile 11a (FAUC 327) showed excellent pharmacological properties combining high D4 affinity (K(i)=1.5 nM) and selectivity with significant intrinsic activity (31%) in low nanomolar concentrations (EC50=1.5 nM).     

Fused azaindole derivatives: molecular design,synthesis and in vitro pharmacology leading to the preferential dopamine D3 receptor agonist FAUC 725

Computational studies based on the similarity of molecular electrostatic potential maps initiated the synthesis of the tricyclic target compounds 1 (FAUC 725) and 2. Receptor binding studies at the dopamine receptor subtypes D1, D2(long), D2(short), D3 and D4 showed that the azaindole 1 revealed D3 affinity (K(i)=0.54 nM) comparable to the lead pramipexole and enhanced selectivity over D2 and D4. Mitogenesis experiments indicated substantial intrinsic activity for the D3 selective dipropylamine 1. Based on the structure of (S)-3-PPP, bioisosteric replacement and conformational restriction leading to the test compound 2 was not fruitful.     

2,4-Disubstituted pyrroles: synthesis,traceless linking and pharmacological investigations leading to the dopamine D4 receptor partial agonist FAUC 356

Solution-phase synthesis and a solid-phase supported approach to piperazinylmethyl substituted pyrroles are described. Receptor binding studies and the measurement of D4 ligand efficacy led to the ethynylpyrrole 1d (FAUC 356) exerting selective D4 binding and substantial ligand efficacy (66%, EC(50)=1.9nM). This activity profile might be of interest for the treatment of ADHD.     

Synthesis and radioiodination of selective ligands for the dopamine D3 receptor subtype

Starting from FAUC 365, a series of iodine substituted heteroaryl carboxamides has been synthesized revealing high affinity and selectivity for the dopamine D3 receptor. Binding data showed a 15-560-fold selectivity for the dopamine D3 over D2. A 2,3-dichloro substitution pattern on the phenylpiperazine moiety led to the highest subtype selectivity, whereas the 2-methoxy substituted compounds showed superior D3 affinity. Suitable precursors were radioiodinated with high radiochemical yields (53-85%) leading to potential imaging agents for the D3 receptor by SPET.     

Dopaminergic 7-aminotetrahydroindolizines: ex-chiral pool synthesis and preferential D3 receptor binding

Starting from both isomers of enantiopure asparagine, heterocyclic bioisosteres of the preferential dopamine D3 receptor agonist (R)-7-OH-DPAT were investigated when SAR studies led to the 3-formyl substituted aminoindolizine (S)-1e (FAUC 54) displaying a K(i) value of 6.0 nM for the high affinity D3 binding site. In contrast, D3 affinity of the enantiomer (R)-1e was 300 fold lower.     

A novel structural framework for α(1A/D)-adrenoceptor selective antagonists identified using subtype selective pharmacophores

In this study four and five-feature pharmacophores for selective antagonists at each of the three α(1)-adrenoceptor (AR) subtypes were used to identify novel α(1)-AR subtype selective compounds in the National Cancer Institute and Tripos LeadQuest databases. 12 compounds were selected, based on diversity of structure, predicted high affinity and selectivity at the α(1D)- subtype compared to α(1A)- and α(1B)-ARs. 9 out of 12 of the tested compounds displayed affinity at the α(1A) and α(1D) -AR subtypes and 6 displayed affinity at all three α(1)-AR subtypes, no α(1B)-AR selective compounds were identified. 8 of the 9 compounds with α(1)-AR affinity were antagonists and one compound displayed partial agonist characteristics. This virtual screening has successfully identified an α(1A/D)-AR selective antagonist, with low μM affinity with a novel structural scaffold of a an isoquinoline fused three-ring system and good lead-like qualities ideal for further drug development.     

Microwave-assisted Phospha-Michael addition reactions in the 13α-oestrone series and in vitro antiproliferative properties

Microwave-assisted phospha-Michael addition reactions were carried out in the 13α-oestrone series. The exocyclic 16-methylene-17-ketones as α,β-unsaturated ketones were reacted with secondary phosphine oxides as nucleophilic partners. The addition reactions furnished the two tertiary phosphine oxide diastereomers in high yields. The main product was the 16α-isomer. The antiproliferative activities of the newly synthesised organophosphorus compounds against a panel of nine human cancer cell lines were investigated by means of MTT assays. The most potent compound, the diphenylphosphine oxide derivative in the 3-O-methyl-13α-oestrone series (9), exerted selective cell growth-inhibitory activity against UPCI-SCC-131 and T47D cell lines with low micromolar IC₅₀ values. Moreover, it displayed good tumour selectivity property determined against non-cancerous mouse fibroblast cells.     

Synthesis and structure elucidation of potential 6-oxygenated metabolites of (22R)-6alpha,9alpha-difluoro-11beta,21-dihydroxy-1 6alpha,17alpha-propyl methylenedioxypregn-4-ene-3,20-dione, and related glucocorticosteroids

(22R)-6alpha,9alpha-Difluoro-11beta,21-dihydroxy-16 alpha,17alpha-propylmethylenedioxypregn-4-ene-3,20-dione (rofleponide) is a synthetic glucocorticosteroid with high affinity for the rat thymus glucocorticoid receptor and a very high biotransformation rate demonstrated through incubation with a human liver S9 subcellular fraction. Because oxidation in the 6-position is an important metabolic pathway of glucocorticosteroids, the potential 6beta-hydroxy and 6-oxo metabolites of rofleponide were synthesized to be used as reference compounds. Three alternative routes were used to reach the 6-hydroxy compound: (a) a one-step procedure involving allylic oxidation of rofleponide by selenium dioxide, (b) selenium dioxide oxidation of the corresponding 1,4-diene followed by selective 1,2-hydrogenation using Wilkinson's catalyst, and (c) autoxidation of a 3-methoxypregna-3,5-diene derivative. All three routes proceeded stereospecifically. Routes (a) and (c) gave approximately the same overall yield of the 6beta-hydroxy epimer, whereas the overall yield from route (b) was much lower, primarily because of incomplete 1,2-hydrogenation. The 6-oxo compound was prepared through Pfitzner/Moffat oxidation of the 6-hydroxy compound. The stereochemistry of the 6-hydroxy substituent is discussed on the basis of 1H-NMR spectroscopy and supplementary 2D NOESY experiments.     

Cyclic amidines as benzamide bioisosteres: EPC synthesis and SAR studies leading to the selective dopamine D4 receptor agonist FAUC 312

Investigation of conformationally restricted benzamide bioisosteres led to the chiral phenyltetrahydropyrimidine derivative ent2a (FAUC 312) displaying strong and highly selective dopamine D4 receptor binding (K(i(high))=1.5 nM). Mitogenesis experiments indicated 83% ligand efficacy when compared to the unselective agonist quinpirole. The target compounds of type 2 and 3 were synthesized in enantiopure form starting from asparagine.     

Gastrointestinal Replacement Solution

The mean sodium concentration of 61 gastrointestinal aspirations in peritonitis was 103 mEq/l. (S.D. 16·66), of 16 aspirations in vagotomy 88 mEq/l. (S.D. 19·49), of 12 aspirations in perforated duodenal ulcer 81 mEq/l. (S.D. 19·49), and of 15 aspirations in intestinal obstruction 89 mEq/l. (S.D. 19·49). The mean potassium concentrations were 9, 9·9, 13, and 8·8 mEq/l. respectively, and the mean chloride concentrations 122, 131, 125, and 112 mEq/l. respectively. It is suggested that a gastrointestinal replacement solution should contain 100 mEq of sodium, 12 mEq of potassium, and 122 mEq of chloride per litre; 50 g. of dextrose or 100 g. of fructose may be added to provide energy.     

MyD88-dependent activation of dendritic cells and CD4(+) T lymphocytes mediates symptoms, but is not required for the immunological control of parasites during rodent malaria

We investigated the role of different TLRs and MyD88 in host resistance to infection and malaria pathogenesis. TLR2(-/-), TLR4(-/-), TLR6(-/-), TLR9(-/-) or CD14(-/-) mice showed no change in phenotypes (parasitemia, body weight and temperature) when infected with Plasmodium chabaudi chabaudi (AS). MyD88(-/-) mice displayed comparable ability to wild type animals in controlling and clearing parasitemia. Importantly, MyD88(-/-) mice exhibited impaired production of TNF-alpha and IFN-gamma as well as attenuated symptoms, as indicated by changes in body weight and temperature during parasitemia. Consistently, CD11b(+) monocytes and CD11c(+) dendritic cells from infected MyD88(-/-) mice were shown impaired for production of pro-inflammatory cytokines, and in initiating CD4(+) T cell responses. Importantly, the inhibition of T cell activation with anti-CD134L, mostly inhibited IFN-gamma, partially inhibited TNF-alpha production, and protected the animals from malaria symptoms. Our findings suggest that MyD88 and possibly its associated TLRs expressed by dendritic cells play an important role in pro-inflammatory responses, T cell activation, and pathogenesis of malaria, but are not critical for the immunological control of the erythrocytic stage of P. chabaudi.     

Benzamide bioisosteres incorporating dihydroheteroazole substructures: EPC synthesis and SAR leading to a selective dopamine D4 receptor partial agonist (FAUC 179).

Conformationally restricted benzamide bioisosteres were investigated when the chiral phenyldihydroimidazole derivative 4e (FAUC 179) showed strong and highly selective dopamine D4 receptor binding (K(i)high=0.95nM). Mitogenesis experiments indicated partial agonist properties (42%). EPC syntheses of the target compounds of type 4 were performed starting from alpha-amino acids.     

Api88 is a novel antibacterial designer peptide to treat systemic infections with multidrug-resistant gram-negative pathogens

The emergence of multiple-drug-resistant (MDR) bacterial pathogens in hospitals (nosocomial infections) presents a global threat of growing importance, especially for Gram-negative bacteria with extended spectrum β-lactamase (ESBL) or the novel New Delhi metallo-β-lactamase 1 (NDM-1) resistance. Starting from the antibacterial peptide apidaecin 1b, we have optimized the sequence to treat systemic infections with the most threatening human pathogens, such as Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii. The lead compound Api88 enters bacteria without lytic effects at the membrane and inhibits chaperone DnaK at the substrate binding domain with a K(D) of 5 μmol/L. The Api88-DnaK crystal structure revealed that Api88 binds with a seven residue long sequence (PVYIPRP), in two different modes. Mice did not show any sign of toxicity when Api88 was injected four times intraperitoneally at a dose of 40 mg/kg body weight (BW) within 24 h, whereas three injections of 1.25 mg/kg BW and 5 mg/kg BW were sufficient to rescue all animals in lethal sepsis models using pathogenic E. coli strains ATCC 25922 and Neumann, respectively. Radioactive labeling showed that Api88 enters all organs investigated including the brain and is cleared through both the liver and kidneys at similar rates. In conclusion, Api88 is a novel, highly promising, 18-residue peptide lead compound with favorable in vitro and in vivo properties including a promising safety margin.     

Chemical synthesis,microbial transformation and biological evaluation of tetrahydroprotoberberines as dopamine D1/D2 receptor ligands

Dopamine D1/D2 receptors are important targets for drug discovery in the treatment of central nervous system diseases. To discover new and potential D1/D2 ligands, 17 derivatives of tetrahydroprotoberberine (THPB) with various substituents were prepared by chemical synthesis or microbial transformation using Streptomyces griseus ATCC 13273. Their functional activities on D1 and D2 receptors were determined by cAMP assay and calcium flux assay. Seven compounds showed high activity on D1/D2 receptor with low IC₅₀ values less than 1?µM. Especially, top compound 5 showed strong antagonistic activity on both D1 and D2 receptor with an IC₅₀ of 0.391 and 0.0757?µM, respectively. Five compounds displayed selective antagonistic activity on D1 and D2 receptor. The SAR studies revealed that (1) the hydroxyl group at C-9 position plays an important role in keeping a good activity and small or fewer substituents on ring D of THPBs may also stimulate their effects, (2) the absence of substituents at C-9 position tends to be more selective for D2 receptor, and (3) hydroxyl substitution at C-2 position and the substitution at C-9 position may facilitate the conversion of D1 receptor from antagonist to agonist. Molecular docking simulations found that Asp 103/Asp 114, Ser 107/Cys 118, and Trp 285/ Trp 386 of D1/ D2 receptors are the key residues, which have strong interactions with the active D1/D2 compounds and may influence their functional profiles.     

Optimal salt bridge for Trp-cage stabilization

Gai and co-workers [Bunagan, M. R., et al. (2006) J. Phys. Chem. B 110, 3759-3763] reported computational design studies suggesting that a D9E mutation would stabilize the Trp-cage. Experimental studies for this mutation were reported in 2008 [Hudaky, P., et al. (2008) Biochemistry 47, 1007-1016]; the authors suggested that [D9E]-TC5b presented a more compact and melting resistant structure because of the "optimal distance between the two sides of the molecule". Nonetheless, the authors reported essentially the same circular dichroism (CD) melting temperature, 38 ± 0.3 °C, for TC5b and its [D9E] mutant. In this study, a more stable Trp-cage, DAYAQ WLKDG GPSSG RPPPS, was examined by nuclear magnetic resonance and CD with the following mutations: [D9E], [D9R,R16E], [R16O], [D9E,R16O], [R16K], and [D9E,R16K]. Of these, the [D9E] mutant displayed the smallest acidification-induced change in the apparent T(m). In analogy to the prior study, the CD melts of TC10b and its [D9E] mutant were, however, very similar; all of the other mutations were significantly fold destabilizing by all measures. A detailed analysis indicates that the original D9-R16 salt bridge is optimal with regard to fold cooperativity and fold stabilization. Evidence of salt bridge formation is also provided for a swapped pair, the [D9R,R16E] mutant. Model systems reveal that an ionized aspartate at the C-terminus of a helix significantly decreases intrinsic helicity, a requirement for Trp-cage fold stability. The CD evidence that was cited as supporting increased fold stability for [D9E]-TC5b at higher temperatures appears to be a reflection of increased helix stability in both the folded and unfolded states rather than a more favorable salt bridge. Our study also provides evidence of other Trp-cage stabilizing roles of the R16 side chain.