Curcumin Derivative GT863 Inhibits Amyloid-Beta Production via Inhibition of Protein N-Glycosylation.

Amyloid-ß (Aß) peptides play a crucial role in the pathogenesis of Alzheimer's disease (AD). Aß production, aggregation, and clearance are thought to be important therapeutic targets for AD. Curcumin has been known to have an anti-amyloidogenic effect on AD. In the present study, we performed screening analysis using a curcumin derivative library with the aim of finding derivatives effective in suppressing Aß production with improved bioavailability of curcumin using CHO cells that stably express human amyloid-ß precursor protein and using human neuroblastoma SH-SY5Y cells. We found that the curcumin derivative GT863/PE859, which has been shown to have an inhibitory effect on Aß and tau aggregation in vivo, was more effective than curcumin itself in reducing Aß secretion. We further found that GT863 inhibited neither ß- nor γ-secretase activity, but did suppress γ-secretase-mediated cleavage in a substrate-dependent manner. We further found that GT863 suppressed N-linked glycosylation, including that of the γ-secretase subunit nicastrin. We also found that mannosidase inhibitors that block the mannose trimming step of N-glycosylation suppressed Aß production in a similar fashion, as was observed as a result of treatment with GT863. Collectively, these results suggest that GT863 downregulates N-glycosylation, resulting in suppression of Aß production without affecting secretase activity.

Specific mutations in presenilin 1 cause conformational changes in γ-secretase to modulate amyloid ß trimming.

γ-Secretase generates amyloid beta peptides (Aß) from amyloid precursor protein through multistep cleavages, such as endoproteolysis (ε-cleavage) and trimming (γ-cleavage). Familial Alzheimer's disease (FAD) mutations within the catalytic subunit protein of presenilin 1 (PS1) decrease γ-cleavage, resulting in the generation of toxic, long Aßs. Reducing long Aß levels has been proposed as an AD therapeutic strategy. Previously, we identified PS1 mutations that are active in the absence of nicastrin (NCT) using a yeast γ-secretase assay. Here, we analysed these PS1 mutations in the presence of NCT, and found that they were constitutively active in yeast. One triple, 13 double, and 5 single mutants enhanced ε-cleavage activity up to 2.7-fold. Furthermore, L241I, F411Y, S438P and F441L mutations modulated trimming activities to produce more short-Aß in yeast microsomes. When introduced in mouse embryonic fibroblasts, these mutations possessed similar or reduced ε-cleavage activity. However, two mutations, L241I and S438P, modulated trimming activities and changed the conformation of transmembrane domain 1, the substrate recognition site. These mutants had the opposite modulatory effects of FAD mutations and produced more short Aßs and fewer long Aßs. Our results provide insights into the relationship between PS1 conformational changes and γ-secretase activities.

In vivo analysis of supersaturation/precipitation/absorption behavior after oral administration of pioglitazone hydrochloride salt; determinant site of oral absorption.

The purpose of this study was to evaluate in vivo supersaturation/precipitation/absorption behavior in the gastrointestinal (GI) tract based on the luminal concentration-time profiles after oral administration of pioglitazone (PG, a highly permeable lipophilic base) and its hydrochloride salt (PG-HCl) to rats. In the in vitro precipitation experiment in the classic closed system, while the supersaturation was stable in the simulated gastric condition, PG drastically precipitated in the simulated intestinal condition, particularly at a higher initial degree of supersaturation. Nonetheless, a drastic and moderate improvement in absorption was observed in vivo at a low and high dose of PG-HCl, respectively. Analysis based on the luminal concentration of PG after oral administration of PG-HCl at a low dose revealed that most of the dissolved PG emptied from the stomach was rapidly absorbed before its precipitation in the duodenum. At a high dose of PG-HCl, PG partly precipitated in the duodenum but was absorbed to some extent. Therefore, the extent of the absorption was mainly dependent on the duodenal precipitation behavior. Furthermore, a higher-than expected absorption after oral administration of PG-HCl from in vitro precipitation study may be due to the absorption process in the small intestine, which suppresses the precipitation by removal of the drug. This study successfully clarify the impact of the absorption process on the supersaturation/precipitation/absorption behavior and key absorption site for a salt formulation of a highly permeable lipophilic base based on the direct observation of in vivo luminal concentration. Our findings may be beneficial in developing an ideal physiologically based pharmacokinetic model and in vitro predictive dissolution tools and/or translating the in silico and in vitro data to the in vivo outcome.

Microsatellite polymorphism in the heme oxygenase-1 gene promoter is associated with susceptibility to cerebral malaria in Myanmar.

Cerebral malaria (CM) is a serious complication of Plasmodium falciparum malaria, and its pathogenesis leading to coma remains unknown. Heme oxygenase-1 (HO-1) catalyzes heme breakdown, eventually generating bilirubin, iron and carbon monoxide. The HO-1 gene promoter contains a polymorphic (GT)n repeat which may influence the expression level of HO-1. To explore the correlation between this (GT)n polymorphism and susceptibility to CM, we analyzed the frequencies of the (GT)n alleles in 120 Myanmarese patients with uncomplicated malaria (UM) and 30 patients with CM. The frequency of homozygotes for the short (GT)n alleles (<28 repeats) in CM patients was significantly higher than those in UM patients (P < 0.008, OR = 3.14). Thus, short (GT)n alleles represent a genetic risk factor for CM.