Megumi Kawai

The structure and conformation of HC-toxin

Abstract Difference nuclear magnetic resonance studies and amino acid oxidase experiments establish the sequence and configuration of amino acids in HC-toxin as cyclo(L-Aoe-D-Pro-L-Ala-D-Ala). HC-toxin adopts the bis-γ-turn conformation in solution previously found for the cytostatic cyclic tetrapeptide chlamydocin.

Structure-activity profiles of macrolactam immunosuppressant FK-506 analogues

The immunosuppressive agent FK‐506 has received much attention due to its efficacy and potency in the areas of transplant rejection and autoimmune disease. Calcineurin, a Ca2+‐calmodulin activated phosphatase, was recently implicated in the immunosuppressive mechanism of FK‐506. In our ongoing search for superior immunosuppressive agents, we have synthesized several analogues of FK‐506 and tested their mechanistic and immunosuppressive actions. It was found that C‐18 hydroxyl analogues of ascomycin, an analogue of FK‐506 also called FR900520, bound tightly to immunophilin FKBP‐12, but do not show any immunosuppressive activity in vitro or in vivo despite good bioavail‐ability. Further, they reverse the inhibition of calcineurin caused by FK‐506/FKBP‐12 complex.

Conformation of two non-immunosuppressive FK506 analogs when bound to FKBP by isotope-filtered NMR

The 3D structure of two unlabeled FK506 analogs, (R)‐ and (S)‐[18‐OH]ascomycin, when bound to [U‐13C,14N]FKBP were determined by isotope‐filtered 2D NMR experiments. The structures for the R and S isomers that bind tightly to FKBP but lack immunosuppresive activity are compared to each other and to the conformation of the potent immunosuppressant, ascomycin, when bound to FKBP. The results are interpreted in terms of calcineurin binding to the FKBP/ascomycin complex.

Total Synthesis of the Cyclic Tetrapeptide, HC-Toxin

Abstract The amino acid sequence of the host-specific phytotoxin, HC-toxin, cyclo(L-Ala-D-Ala-L-Aoe-D-Pro) has been confirmed by total synthesis.

Stereoselective synthesis and absolute configuration of epoxyketones in chlamydocin and related cyclic tetrapeptides

Abstract The epoxyketone group in the amino acid, 2S-amino-8-oxo-9,10-epoxydecanoic acid (Aoe) has been synthesized by chiral epoxidation of the corresponding allylic alcohol cyclic tetrapeptide precursor to form chlamydocin and epichlamydocin. These compounds have been used as standards to assign by circular dichroism spectroscopy the Aoe epoxyketone configurations in HC-toxin and WF-3161.

Synthesis of VS-105: A novel and potent vitamin D receptor agonist with reduced hypercalcemic effects.

We have synthesized a novel vitamin D receptor agonist VS-105 ((1R,3R)-5-((E)-2-((3αS,7αS)-1-((R)-1-((S)-3-hydroxy-2,3-dimethylbutoxy)ethyl)-7α-methyldihydro-1H-inden-4(2H,5H,6H,7H,7αH)-ylidene)ethylidene)-2-methylenecyclohexane-1,3-diol). Preparation of a-ring phenylphosphine oxide 11, followed by Wittig-Horner coupling of 11 with the protected 25-hydroxy Grundmanns ketone 22 generated the precursor 12. Deprotection of the TBDMS groups of 12 produced the target compound VS-105. The biological profiles of VS-105 were evaluated using in vitro assays (VDR receptor binding, VDR reporter gene and HL-60 differentiation) in comparison to calcitriol (the endogenous hormone) or paricalcitol. Furthermore, the PTH suppressing potency and hypercalcemic side effects of VS-105 were evaluated in the 5/6 nephrectomized uremic rats in comparison to paricalcitol. Combining various changes at 20-epi, 22-oxa, 24-methyl, and 2-methylene yielded VS-105 that not only is highly potent in inducing functional responses in vitro, but also effectively suppresses PTH in a dose range that does not affect serum calcium in the 5/6 nephrectomized uremic rats.

Vitamin K-dependent carboxylase: Synthesis of an inhibitor of the glutamyl binding site

Liver microsomes contain a vitamin K and O2‐dependent carboxylase that converts peptide‐bound glutamyl residues to γ‐carboxyglutamyl residues. The peptide Boc‐O‐phospho—Ser‐O‐phospho—Ser—Leu‐OMe has now been synthesized. This peptide inhibits the carboxylation of endogenous protein precursors by a detergent‐solubilized preparation of the carboxylase and is an apparent competitive inhibitor of the carboxylation of Phe—Leu—Glu—Glu—Leu.

Studies on an immunosuppressive macrolactam, ascomycin: Synthesis of a C-33 hydroxyl derivative

Ascomycin 2, a close analogue of the immunosuppressant FK506 1, was modified to incorporate a hydroxyl group at the C-33 position. This increased the aqueous solubility of ascomycin by a hundred-fold at pH 7.4 and by approximately 300-fold at pH 6.5. Ascomycin 3 also exhibited an excellent immunosuppressive activity in vitro, as tested in a human mixed lymphocyte proliferation (HuMLR) assay, and in vivo using a rat popliteal lymph node (rPLN) hyperplasia assay.