Activity-differential search for amyloid-ß aggregation inhibitors using LC-MS combined with principal component analysis.

Aggregation of amyloid ß42 (Aß42) is one of the hallmarks of Alzheimer's disease (AD). Inhibition of Aß42 aggregation is thus a promising approach for AD therapy. Kampo medicine has been widely used to combat dementias such as AD. Crude drug known as Shoyaku is an ingredient of Kampo that could have potential as a natural source of novel drugs. However, given that a mixture of compounds, rather than singular compounds, could contribute to the biological functions of crude drug, there are very limited studies on the structure and mechanism of each constituent in crude drug which may have anti-Aß42 aggregation properties. Herein we provide an efficient method, using LC-MS combined with principal component analysis (PCA), to search for activity-dependent compounds that inhibit Aß42 aggregation from 46 crude drug extracts originating from 18 plants. Only 5 extracts (Kakou, Kayou, Gusetsu, Rensu, and Renbou) from lotus demonstrated differentially inhibitory activities depending on the part of the plant from which they are derived (e.g. petiole, leaf, root node, stamen, and receptacle, respectively). To compare the anti-aggregative properties of compounds of active crude drug with those of inactive crude drug, these extracts were subjected to LC-MS measurement, followed by PCA. From 12 candidate compounds identified from the analysis, glucuronized and glucosidized quercetin, as well as 6 flavonoids (datiscetin, kaempferol, morin, robinetin, quercetin, and myricitrin), including catechol or flatness moiety suppressed Aß42 aggregation, whereas curcumol, a sesquiterpene, did not. In conclusion, this study offers a new activity-differential methodology to identify bioactive natural products in crude drugs that inhibit Aß42 aggregation and that could be applied to future AD therapies.

Role of the carboxy groups of triterpenoids in their inhibition of the nucleation of amyloid ß42 required for forming toxic oligomers.

Herein we report that a preferable inhibition of the nucleation phase of Aß42, related to the formation of toxic oligomers, by triterpenoids from medicinal herbs originates from a salt bridge of their carboxy groups with Lys16 and 28 in Aß42. Such a direct interaction targeting the monomer, dimer, and trimer suppressed further oligomerization. In contrast, the corresponding congeners without carboxy groups failed to do so.

Semisynthesis and Structure-Activity Studies of Uncarinic Acid C Isolated from Uncaria rhynchophylla as a Specific Inhibitor of the Nucleation Phase in Amyloid ß42 Aggregation.

Oligomers of the 42-mer amyloid-ß protein (Aß42), rather than fibrils, cause synaptic dysfunction in the pathology of Alzheimer's disease (AD). The nucleation phase in a nucleation-dependent aggregation model of Aß42 is related to the formation of oligomers. Uncaria rhynchophylla is one component of "Yokukansan", a Kampo medicine, which is widely used for treating AD symptoms. Previously, an extract of U. rhynchophylla was found to reduce the aggregation of Aß42, but its active principles have yet to be identified. In the present work, uncarinic acid C (3) was identified as an inhibitor of Aß42 aggregation that is present in U. rhynchophylla. Moreover, compound 3 acted as a specific inhibitor of the nucleation phase of Aß42 aggregation. Compound 3 was synthesized from saponin A (10), an abundant byproduct of rutin purified from Uncaria elliptica. Comprehensive structure-activity studies on 3 suggest that both a C-27 ferulate and a C-28 carboxylic acid group are required for its inhibitory activity. These findings may aid the development of oligomer-specific inhibitors for AD therapy.

Structure-based design of substituted piperidines as a new class of highly efficacious oral direct Renin inhibitors.

A cis-configured 3,5-disubstituted piperidine direct renin inhibitor, (syn,rac)-1, was discovered as a high-throughput screening hit from a target-family tailored library. Optimization of both the prime and the nonprime site residues flanking the central piperidine transition-state surrogate resulted in analogues with improved potency and pharmacokinetic (PK) properties, culminating in the identification of the 4-hydroxy-3,5-substituted piperidine 31. This compound showed high in vitro potency toward human renin with excellent off-target selectivity, 60% oral bioavailability in rat, and dose-dependent blood pressure lowering effects in the double-transgenic rat model.

Overcoming hERG issues for brain-penetrating cathepsin S inhibitors: 2-cyanopyrimidines. Part 2.

We describe here orally active and brain-penetrant cathepsin S selective inhibitors, which are virtually devoid of hERG K(+) channel affinity, yet exhibit nanomolar potency against cathepsin S and over 100-fold selectivity to cathepsin L. The new non-peptidic inhibitors are based on a 2-cyanopyrimidine scaffold bearing a spiro[3.5]non-6-yl-methyl amine at the 4-position. The brain-penetrating cathepsin S inhibitors demonstrate potential clinical utility for the treatment of multiple sclerosis and neuropathic pain.

4-Amino-2-cyanopyrimidines: novel scaffold for nonpeptidic cathepsin S inhibitors.

We describe here a novel 4-amino-2-cyanopyrimidine scaffold for nonpeptidomimetic cathepsin S selective inhibitors. Some of the synthesized compounds have sub-nanomolar potency and high selectivity toward cathepsin S along with promising pharmacokinetic and physicochemical properties. The key structural features of the inhibitors consist of a combination of a spiro[2.5]oct-6-ylmethylamine P2 group at the 4-position, a small or polar P3 group at the 5-position and/or a polar group at the 6-position of the pyrimidine.

Discovery of selective and nonpeptidic cathepsin S inhibitors.

Nonpeptidic, selective, and potent cathepsin S inhibitors were derived from an in-house pyrrolopyrimidine cathepsin K inhibitor by modification of the P2 and P3 moieties. The pyrrolopyrimidine-based inhibitors show nanomolar inhibition of cathepsin S with over 100-fold selectivity against other cysteine proteases, including cathepsin K and L. Some of the inhibitors showed cellular activities in mouse splenocytes as well as oral bioavailabilities in rats.

Novel scaffold for cathepsin K inhibitors.

Pyrrolopyrimidine, a novel scaffold, allows to adjust interactions within the S3 subsite of cathepsin K. The core intermediate 10 facilitated the P3 optimization and identified highly potent and selective cathepsin K inhibitors 11-20.

Synthesis and structure-activity relationship of novel RXR antagonists: orally active anti-diabetic and anti-obesity agents.

A series of diazepinylbenzoic acid derivatives were synthesized and tested in the inhibition assay of the transactivation of RXR. Oral treatment of cyano derivatives (16f) was found to show anti-diabetic and anti-obesity effects in KK-A(y) mice.

Synthesis and structure-activity relationship of RXR antagonists based on the diazepinylbenzoic acid structure.

Synthesis and structure-activity relationship of RXR antagonists employing a diazepinylbenzoic acid scaffold are described. Of those antagonists, sulfonamide derivatives (6v and 6w) reveal a high antagonistic activity and good pharmacokinetic properties.