Youhei Sohma

Novel and efficient synthesis of difficult sequence-containing peptides through O–N intramolecular acyl migration reaction of O-acyl isopeptides

A novel and efficient method for the synthesis of difficult sequence-containing peptides has been developed based on the synthesis of O-acyl isopeptides followed by an O-N intramolecular acyl migration reaction, resulting in a remarkable improvement of the yields.

“Click Peptides”—Chemical Biology‐Oriented Synthesis of Alzheimer's Disease‐Related Amyloid β Peptide (Aβ) Analogues Based on the “O‐Acyl Isopeptide Method”

A clear understanding of the pathological mechanism of amyloid β peptide (Aβ) 1–42, a currently unexplained process, would be of great significance for the discovery of novel drug targets for Alzheimers disease (AD) therapy. To date, though, the elucidation of these Aβ1–42 dynamic events has been a difficult issue because of uncontrolled polymerization, which also poses a significant obstacle in establishing experimental systems with which to clarify the pathological function of Aβ1–42. We have recently developed chemical biology‐oriented pH‐ or phototriggered “click peptide” isoform precursors of Aβ1–42, based on the “O‐acyl isopeptide method”, in which a native amide bond at a hydroxyamino acid residue, such as Ser, is isomerized to an ester bond, the target peptide subsequently being generated by an O–N intramolecular acyl migration reaction. These click peptide precursors did not exhibit any self‐assembling character under physiological conditions, thanks to the presence of the one single ester bond, and were able to undergo migration to give the target Aβ1–42 in a quick and easy, one‐way (so‐called “click”)conversion reaction. The use of click peptides could be a useful strategy to investigate the biological functions of Aβ1–42 in AD through inducible activation of Aβ1–42 self‐assembly.

O-N intramolecular acyl migration reaction in the development of prodrugs and the synthesis of difficult sequence-containing bioactive peptides

NOintramolecular acyl migration in Ser‐ or Thr‐containing peptides is a well‐known side reaction in peptide chemistry. It results in the mutual conversion of ester and amide bonds. Our medicinal chemistry study focused on the fact that the O‐acyl product can be readily converted to the original N‐acyl form under neutral or slightly basic conditions in an aqueous buffer and the liberated ionized amino group enhances the water solubility of O‐acyl products. Because of this, we have developed a novel class of “ON intramolecular acyl migration”‐type water‐soluble prodrugs of HIV‐1 protease inhibitors. These prodrugs released the parent drugs via a simple chemical mechanism with no side reaction. In this study, we applied this strategy to important cancer chemotherapeutic agents, paclitaxel and its derivatives, to develop water‐soluble taxoid prodrugs, and found that these prodrugs, 2′‐O‐isoform of taxoids, showed promising results with higher water solubility and proper kinetics in their parent drug formation by a simple pH‐dependent chemical mechanism with ON intramolecular acyl migration. These results suggest that this strategy would be useful in toxicology and medical economics.

Design and Folding of [GluA4(OβThrB30)]Insulin (“Ester Insulin”): A Minimal Proinsulin Surrogate that Can Be Chemically Converted into Human Insulin

Insulin biosynthesis involves the efficient folding of a single polypeptide-chain precursor, with concomitant formation of three disulfides, to give proinsulin and the subsequent enzymatic removal of the C-peptide to give mature insulin.[1,2] A proinsulin- or mini-proinsulin-based approach is currently used in the recombinant production of human insulin.[3,4] However, recombinant production of insulin analogues is effectively limited to the creation of mutants from the twenty genetically encoded amino acids. In contrast to this, total chemical synthesis of insulin would in principle enable the incorporation of a wide range of non-natural amino acids and other chemical modifications into the molecule,[5] and would thus enable the full exploration of the medicinal chemistry of this important therapeutic molecule. Until now, however, we have lacked an efficient approach to the chemical synthesis of human insulin.[5] This has impeded development of next-generation insulin analogues containing non-standard side chains, D-amino acids[6,7] or other novel chemical structural features.

“Click Peptide“: pH‐Triggered in Situ Production and Aggregation of Monomer Aβ1–42

Into neutral: We demonstrate the unique features of a pH click peptide based on an O‐acyl isopeptide method. Under acidic conditions, the click peptide remains in a monomeric form. Upon increase of the pH to 7.4, the click peptide is quickly able to convert into Aβ1–42 through an O‐to‐N intramolecular acyl migration. Further study using this pH click peptide would elucidate the pathological role of Aβ1–42 in Alzheimers disease.

New water-soluble prodrugs of HIV protease inhibitors based on O→N intramolecular acyl migration

To improve the low water-solubility of HIV protease inhibitors, we synthesized water-soluble prodrugs of KNI-272 and KNI-279 which are potent HIV-1 protease inhibitors consisting of an Apns-Thz core structure (Apns; allophenylnorstatine, Thz; thiazolidine-4-carboxylic acid) as an inhibitory machinery. The prodrugs, which contained an O-acyl peptidomimetic structure with an ionized amino group leading to the increase of water-solubility, were designed to regenerate the corresponding parent drugs based on the O-->N intramolecular acyl migration reaction at the alpha-hydroxy-beta-amino acid residue, that is allophenylnorstatine. The synthetic prodrugs 3, 4, 6, and 7 improved the water-solubility (>300mg/mL) more than 4000-fold in comparison with the parent compounds, which is the practically acceptable value as water-soluble drugs. These prodrugs were stable as an HCl salt and in a strongly acidic solution corresponding to gastric juice (pH 2.0), and could be converted to the parent compounds promptly in the aqueous condition from slightly acidic to basic pH at 37 degrees C, with the suitable migration rate, via a five-membered ring intermediate. Using a similar method, we synthesized a prodrug (12) of ritonavir, a clinically useful HIV-1 protease inhibitor as an anti-AIDS drug. In contrast to the prodrugs 3, 4, 6, and 7, the prodrug 12 was very slowly converted to ritonavir probably through a six-membered ring intermediate, with the t(1/2) value of 32h that may not be suitable for practical use.

Attenuation of the Aggregation and Neurotoxicity of Amyloid‐β Peptides by Catalytic Photooxygenation

Alzheimers disease (AD), a progressive severe neurodegenerative disorder, is currently incurable, despite intensive efforts worldwide. Herein, we demonstrate that catalytic oxygenation of amyloid-β peptides (Aβ) might be an effective approach to treat AD. Aβ1-42 was oxygenated under physiologically-relevant conditions (pH 7.4, 37 °C) using a riboflavin catalyst and visible light irradiation, with modifications at the Tyr(10) , His(13) , His(14) , and Met(35) residues. The oxygenated Aβ1-42 exhibited considerably lower aggregation potency and neurotoxicity compared with native Aβ. Photooxygenation of Aβ can be performed even in the presence of cells, by using a selective flavin catalyst attached to an Aβ-binding peptide; the Aβ cytotoxicity was attenuated in this case as well. Furthermore, oxygenated Aβ1-42 inhibited the aggregation and cytotoxicity of native Aβ.

Controlled Production of Amyloid β Peptide from a Photo-Triggered, Water-Soluble Precursor “Click Peptide“

In biological experiments, poor solubility and uncontrolled assembly of amyloid β peptide (Aβ) 1–42 pose significant obstacles to establish an experiment system that clarifies the function of Aβ1–42 in Alzheimers disease (AD). Herein, as an experimental tool to overcome these problems, we developed a water‐soluble photo‐“click peptide” with a coumarin‐derived photocleavable protective group that is based on an O‐acyl isopeptide method. The click peptide had nearly 100‐fold higher water solubility than Aβ1–42 and did not self‐assemble, as the isomerized structure in its peptide backbone drastically changed the conformation that was derived from Aβ1–42. Moreover, the click peptide afforded Aβ1–42 quickly under physiological conditions (pH 7.4, 37 °C) by photoirradiation followed by an O–N intramolecular acyl migration. Because the in situ production of intact Aβ1–42 from the click peptide could improve the difficulties in handling Aβ1–42 caused by its poor solubility and highly aggregative nature, this click peptide strategy would provide a reliable experiment system for investigating the pathological function of Aβ1–42 in AD.

Controlled drug release: new water-soluble prodrugs of an HIV protease inhibitor

We designed and synthesized a series of highly water-soluble prodrugs of an HIV protease inhibitor, KNI-727 (1), containing tandem-linked two auxiliary units, a solubilizing moiety and a self-cleavable spacer. Prodrugs with an ionized amino group at the solubilizing moiety exhibited a remarkable increase of water-solubility (>10(4) fold) compared to the parent drug 1. These prodrugs released I not enzymatically, but chemically via an intramolecular cyclization-elimination reaction through an imide formation in physiological conditions. Diversified rates of parent drug release were observed when the chemical structure of both the solubilizing and the spacer moieties were modified. This new approach for water-soluble prodrugs will enable to control chemically the release of parent drug as well as to maintain high water-solubility.

Rational Design and Identification of a Non‐Peptidic Aggregation Inhibitor of Amyloid‐β Based on a Pharmacophore Motif Obtained from cyclo[‐Lys‐Leu‐Val‐Phe‐Phe‐]

Inhibition of pathogenic protein aggregation may be an important and straightforward therapeutic strategy for curing amyloid diseases. Small-molecule aggregation inhibitors of Alzheimers amyloid-β (Aβ) are extremely scarce, however, and are mainly restricted to dye- and polyphenol-type compounds that lack drug-likeness. Based on the structure-activity relationship of cyclic Aβ16-20 (cyclo-[KLVFF]), we identified unique pharmacophore motifs comprising side-chains of Leu(2), Val(3), Phe(4), and Phe(5) residues without involvement of the backbone amide bonds to inhibit Aβ aggregation. This finding allowed us to design non-peptidic, small-molecule aggregation inhibitors that possess potent activity. These molecules are the first successful non-peptidic, small-molecule aggregation inhibitors of amyloids based on rational molecular design.