N-Difluoromethyl-triazole as a constrained scaffold in peptidomimetics.

The N-difluoromethyl triazolo-ß-aza-ε-amino acid present in the core of peptides led to constrained conformations due to CH-F and NH-F interactions. Pseudotetrapeptides were obtained in excellent yields directly by click chemistry between azidodifluoroacetamides and alkynes, both linked to an amino acid. This work demonstrates that the N-difluoromethyltriazole scaffold can induce extended structures to ß-strand mimics.

ß-Hairpin mimics containing a piperidine-pyrrolidine scaffold modulate the ß-amyloid aggregation process preserving the monomer species.

Alzheimer's disease is a neurodegenerative disorder linked to oligomerization and fibrillization of amyloid ß peptides, with Aß1-42 being the most aggregative and neurotoxic one. We report herein the synthesis and conformational analysis of Aß1-42-amyloid related ß-hairpin peptidomimetics, built on a piperidine-pyrrolidine semi rigid ß-turn inducer and bearing two small recognition peptide sequences, designed on oligomeric and fibril structures of Aß1-42. According to these peptide sequences, a stable ß-hairpin or a dynamic equilibrium between two possible architectures was observed. These original constructs are able to greatly delay the kinetics of Aß1-42 aggregation process as demonstrated by thioflavin-T fluorescence, and transmission electron microscopy. Capillary electrophoresis indicates their ability to preserve the monomer species, inhibiting the formation of toxic oligomers. Furthermore, compounds protect against toxic effects of Aß on neuroblastoma cells even at substoichiometric concentrations. This study is the first example of acyclic small ß-hairpin mimics possessing such a highly efficient anti-aggregation activity. The protective effect is more pronounced than that observed with molecules which have undergone clinical trials. The structural elements made in this study provide valuable insights in the understanding of the aggregation process and insights to explore the design of novel acyclic ß-hairpin targeting other types of amyloid-forming proteins.

Access to novel functionalized trifluoromethyl ß-lactams by ring expansion of aziridines.

From carboxylic acid trifluoromethyl aziridines, halogeno ß-lactams were obtained stereoselectively by ring expansion. Different conditions such as radical, organometallic reactions allowed easy and selective access to CF3-ß-lactams substituted at the C-3 position.

(19)F NMR monitoring of the eukaryotic 20S proteasome chymotrypsin-like activity: an investigative tool for studying allosteric regulation.

The proteasome displays three distinct proteolytic activities. Currently, proteasome inhibitors are evaluated using specific fluorescent substrates for each of the individual active sites. However, the photophysical properties of the commonly used fluorophores are similar and thus, the simultaneous monitoring of the three proteolytic activities is not possible. We have developed a bimodal fluorescent fluorinated substrate as a novel tool to study the chymotrypsin-like (ChT-L) proteolytic activity and its regulation by inhibitors and by substrates of trypsin-like (T-L) and caspase-like sites (PA). We demonstrate that this substrate is reliable to evaluate the ChT-L inhibitory activity of new molecules either by fluorescence or (19)F NMR spectroscopy. We have found that the ChT-L activity is dramatically reduced in the presence of T-L and PA substrates. This work provides a proof of concept that the fluorinated substrate enables investigation of the allosteric regulation of the ChT-L activity.

Discovery of a new efficient chiral ligand for copper-catalyzed enantioselective Michael additions by high-throughput screening of a parallel library.

A combinatorial library of 125 chiral Schiff base ligands 5 was synthesized with the use of solution-phase parallel synthesis and solid-phase extraction (SPE) techniques to scavenge excess reagents and reaction by-products and avoid chromatography. The synthetic methodology coupled five N-Boc-protected beta-amino sulfonyl chlorides 1a-e with five different amines 2f-j to give 25 N-Boc sulfonamides 3, which were in turn deprotected and coupled with five salicylaldehydes 4p-t to give 125 ligands 5 in good yields and of sufficient purity to be used in ligand-catalyzed reactions. These ligands were tested in the copper-catalyzed conjugate addition of dialkyl zinc to cyclic and acyclic enones. A multisubstrate high-throughput screening of the library was performed with an equimolar mixture of 2-cyclohexenone and 2-cycloheptenone (9 and 10, respectively, 0.2 mmol total), with 5.5 mol% ligand 5 (0.011 mmol) and 5 mol% Cu(OTf)2 (OTf= OSO2CF3) (0.010 mmol) in 1:1 toluene/ hexane at - 20 degrees C. From the screening of the library, 5bhr was identified as the best ligand, which yielded 3-ethylcyclohexanone (12) and 3-ethylcycloheptanone (13) in 82% and 81% ee, respectively, and complete conversions. Under optimized conditions (2.75 mol% 5bhr, 2.5 mol% copper(i) triflate, toluene as reaction solvent), improved results were obtained for 12 (90% ee, 93% yield) and for 13 (91% ee, 95% yield). Selected ligands 5 were also tested in the addition of Me2Zn to 2-cyclohexenone (9, ee up to 79%), of Et2Zn to 2-cyclopentenone (11, ee up to 80%) and to acyclic enones 16 and 17 (ee up to 50%).

Synthesis, modelling and NK1 antagonist evaluation of a non-rigid cyclopropane-containing analogue of CP-99,994.

A non-rigid cyclopropane-containing diamine analogue of CP-99,994 was synthesised and was found to have only moderate NK1 receptor binding affinity. Molecular dynamics calculations of the conformational space of the former compound gave good correlation between observed activity and a recently published pharmacophore model, lending predictive value to the latter.

Ascites in patients undergoing intermittent haemodialysis at Kenyatta National Hospital.

In a two-year-period (August 1984 to August 1986), 77 patients were admitted into the maintenance haemodialysis programme at Kenyatta National Hospital. 24 (31.5%) of these had ascites during haemodialysis. Nine (37.5%) of the patients who had ascites had prior peritoneal dialysis, while 15 (62.5%) had congestive cardiac failure at the time of development of the ascites. In 21 (87.5%), the ascites responded to therapy with diuretics, salt and fluid restriction, antibiotics when indicated and to ultrafiltration during dialysis. In 3 (12.5%) of the patients, the ascites developed in the absence of any predisposing cause. The ascites progressively increased in amount and was associated with marked wasting. These patients were considered to have refractory ascites of haemodialysis.

Problems with a renal replacement programme in a developing country.

Since August 1984 patients with end-stage renal disease in Kenya have been started on haemodialysis with a view to renal transplantation. In a two year period (August 1984-August 1986) 77 patients mean age 29.6 years (49 males), have been dialysed. The mean duration on dialysis prior to death or transplantation was 2.9 months (range 1 day to 11 months). Fifty patients (65%) died while on dialysis, including 2 who had had unsuccessful transplantation. Fourteen patients were still on dialysis, 11 had discharged themselves to peripheral hospitals for conservative management, and 2 had had successful renal transplantation. The possible causes of this abysmal experience include admission of critically ill patients, shortage of trained staff, over-dependence on arteriovenous shunts for vascular access, lack of centralization of patient management, recurrent shortage of essential equipment and reagents and a slow pace of transplantation.