Glucocerebrosidase (GCase) activity modulation by 2-alkyl trihydroxypiperidines: Inhibition and pharmacological chaperoning.

The enzyme glucocerebrosidase (GCase) has become an important therapeutic target due to its involvement in pathological disorders consequent to enzyme deficiency, such as the lysosomal storage Gaucher disease (GD) and the neurological Parkinson disease (PD). Pharmacological chaperones (PCs) are small compounds able to stabilize enzymes when used at sub-inhibitory concentrations, thus rescuing enzyme activity. We report the stereodivergent synthesis of trihydroxypiperidines alkylated at C-2 with both configurations, by means of the stereoselective addition of Grignard reagents to a carbohydrate-derived nitrone in the presence or absence of Lewis acids. All the target compounds behave as good GCase inhibitors, with IC50 in the micromolar range. Moreover, compound 11a behaves as a PC in fibroblasts derived from Gaucher patients bearing the N370/RecNcil mutation and the homozygous L444P mutation, rescuing the activity of the deficient enzyme by up to 1.9- and 1.8-fold, respectively. Rescues of 1.2-1.4-fold were also observed in wild-type fibroblasts, which is important for targeting sporadic forms of PD.

Multimerization of DAB-1 onto Au GNPs affords new potent and selective N-acetylgalactosamine-6-sulfatase (GALNS) inhibitors.

Gold glyconanoparticles (Au GNPs) decorated with the natural iminosugar DAB-1 at different densities are reported. These new multivalent iminosugar architectures strongly and selectively inhibit N-acetylgalactosamine-6-sulfatase (GALNS), whose deficiency is connected to the lysosomal storage disease Morquio A. The combination of the dendrimeric technique with the synthetic strategy employed for Au GNP preparation allowed the enhancement of the multivalent presentation of the iminosugar onto the surface of gold nanoparticles, which resulted in the best GALNS inhibitor reported to date (IC50 = 520 nM).

Accessing 2-substituted piperidine iminosugars by organometallic addition/intramolecular reductive amination: aldehyde vs. nitrone route.

A dual synthetic strategy to afford 2-substituted trihydroxypiperidines is disclosed. The procedure involved Grignard addition either to a carbohydrate-derived aldehyde or to a nitrone derived thereof, and took advantage of an efficient ring-closure reductive amination strategy in the final cyclization step. An opposite diastereofacial preference was demonstrated in the nucleophilic attack to the two electrophiles, which would finally produce the same piperidine diastereoisomer as the major product. However, use of a suitable Lewis acid in the Grignard addition to the nitrone allowed reversing the selectivity, giving access to 2-substituted piperidines with the opposite configuration at C-2.

From glycals to aminosugars: a challenging test for new stereoselective aminohydroxylation and related methodologies.

The introduction of amino functionalities in a regio- and stereoselective manner onto sugar scaffolds represents a great challenge in carbohydrate synthesis. The most relevant methods to access 1-, 2-, 3-amino or 1,2-diaminosugars starting from glycals and 2,3-hexenopyranosides derived from them are concisely reviewed. The main synthetic strategies for accessing this class of compounds are classified in intermolecular and intramolecular approaches and the key features of each class are discussed. This review highlights how carbohydrate derivatives always pose great challenges representing a benchmark for assessing the efficiency of stereoselective strategies, and aims to give the readers inspiration for the development of new procedures.

Stereoselective approach to hydroxyindolizidines: protection/deprotection of the nitrone functionality via cycloaddition/retrocycloaddition.

The enantiomerically pure indolizidine (-)-21 has been synthesized starting from L-malic acid. The key intermediate 20 has been assembled through an intramolecular 1,3-dipolar cycloaddition of a nitrone generated in situ by retrocycloaddition from isoxazolidine 17 or 18. The configuration of the new three stereocenters was set up with complete control in the cycloaddition step. The presented synthetic route provides a general and highly selective methodology toward indolizidines having the [1,8a]-cis configuration.

Total synthesis of (-)-rosmarinecine by intramolecular cycloaddition of (S)-malic acid derived pyrroline N-oxide.

[reaction in text] Straightforward total syntheses of (-)-rosmarinecine have been achieved from L-malic acid derived pyrroline N-oxides by two novel useful cascade processes, which join the family of domino reactions. Both strategies, which furnished the target alkaloid in enantioenriched and enantiopure forms, respectively, allow complete control of configuration at all the three newly created contiguous stereogenic centers.

The effect of pH on the thermal resistance of Clostridium sporogenes (PA 3679) in asparagus purée acidified with citric acid and glucono-delta-lactone.

The influence of type of acid, pH and temperature on heat resistance of Clostridium sporogenes (PA 3679) spores were investigated in white asparagus pureé acidified with citric acid and glucono-delta-lactone (GDL). The pH values studied were: 4.5, 4.8, 5.1 and 5.4 at temperatures of 110, 115, 118 and 121 degrees C. The addition of citric acid and GDL to reduce pH significantly diminished heat resistance of the spores. The two acids investigated differed in their effect on heat resistance at the various pH-levels. The most pronounced effect was observed at the lower heat treatment temperatures investigated. The z values ranged from 10.24 to 13.09 degrees C in the asparagus purée with acids added and with significant differences between the two acids and the various pH levels.

One-pot synthesis of cyclic nitrones and their conversion to pyrrolizidines: 7a-epi-crotanecine inhibits alpha-mannosidases.

[reaction: see text] A new straightforward and inexpensive one-pot procedure is described for the preparation of enantiopure five-membered cyclic nitrones starting from the corresponding lactols. Its efficiency relies on the condensation of unprotected hydroxylamine with readily available lactols and on the chemoselectivity of the subsequent esterification with methanesulfonyl chloride. The targeted enantiomerically pure pyrroline N-oxides are versatile synthetic intermediates: one of the nitrones so-obtained has been converted into new polyhydroxypyrrolizidines, analogues of the alkaloids rosmarinecine and crotanecine, which were assayed for their inhibitory activities toward 22 commercially available glycosidase enzymes. One of them ((-)-7a-epi-crotanecine) is a potent and selective inhibitor of alpha-mannosidases from jack beans and almonds.

Measurement of testosterone and its 5-alfa-reduced metabolites in human prostatic tissue using isotope dilution mass spectrometry.

5 alpha-Dihydrotestosterone (DHT) has been widely measured in human prostatic tissue using RIA since it is found to be involved in pathogenesis of human prostatic hyperplasia (BHP) and to be the best index for the follow-up of patients affected by prostatic cancer under endocrine treatment. A GC/MS method for the simultaneous determination of testosterone (T), dihydrotestosterone (DHT), 5 alpha-androstan-3 alpha,17 beta-diol (3 alpha-diol) and 5 alpha-androstan-3 beta-17 beta-diol (3 beta-diol) in prostatic tissue was developed based on the isotopic dilution technique. Trideuterated internal standards of each compound were previously synthesized in our laboratory. After previous extraction and purification on Sep-Pak C-18 cartridges and Lipidex DEAP columns, T and its metabolites were measured as heptafluorobutyric esters (HFB). Quantitative analysis was performed on a VG 7070 EQ mass spectrometer equipped with a fused silica capillary column using the Selected Ion Monitoring technique. Steroid values (means +/- SD; ng/g tissue) found in 9 human hypertrophic prostates were: T, 0.71 +/- 0.43; DHT, 4.46 +/- 1.41; 3 alpha-diol, 0.34 +/- 0.23; 3 beta-diol, 0.14 +/- 0.32.