Ichiro Umemura

A potent and specific agonist, Suc-[Glu9, Ala11,15]-endothelin-1(8-21), IRL 1620, for the ETB receptor

A series of C-terminal linear peptides of endothelin (ET)-1 and their N alpha-succinyl (Suc) analogs were synthesized and their binding affinities for the two subtypes of ET receptor, ETA and ETB, in porcine lung membranes were examined. Among the synthetic analogs, Suc-[Glu9,Ala11,15]-ET-1(8-21), IRL 1620, was the most potent and specific ligand for the ETB receptor (KiETA/KiETB approximately equal to 120,000) as judged by the Ki values for ETA (1.9 microM) and ETB (16 pM) receptors. IRL 1620 was 60 times more selective for the ETB receptor than ET-3 (KiETA/KiETB approximately equal to 1,900). IRL 1620 (10(-9)-10(-7) M) induced contractions of the guinea pig trachea with a comparable potency to those of ET-1 or ET-3, suggesting that IRL 1620 is a potent ETB receptor agonist.

An endothelin B receptor-selective antagonist: IRL 1038, [Cys11-Cys15]-endothelin-1(11–21)

In the inhibition of specific binding or [125]endothelins (ETs) to membranes from various tissues of rats, guinea pigs, pigs and humans, [Cys11‐Cys15]‐ET‐1(11–21), IRL 1038, has a much higher affinity for ETB receptors (K i = 6–11 nM) than for ETA receptors (K i = 0.4–0.7 μM). In contraction assays, with ET‐3 as a stimulant, 3 μM IRL 1038 antagonized the ETB receptor‐mediated contraction of guinea pig ileal and tracheal smooth muscle without any significant agonistic activity, but did not effect the ETA receptor‐mediated contraction of rat aortic smooth muscle. IRL 1038 is, therefore, considered to be the first antagonist selective to the ETB receptor.

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.

A reversible radioligand specific for the ETB receptor: [125I]Tyr13-Suc-[Glu9,Ala11,15]-endothelin-1(8- 21), [125I]IRL 1620.

Suc-[Glu9,Ala11,15]-endothelin(ET)-1(8-21), IRL 1620, is a linear ET-analog specific for the ET-isopeptide-nonselective ETB receptor. The radio-iodinated analog, [125I]IRL 1620, showed a single class of saturable binding to the ETB receptors in porcine lung membranes with a Kd of 18 pM and a Bmax of 930 fmol/mg protein, which are almost comparable to the values obtained with [125I]ET-3 (6 pM and 900 fmol/mg protein). In competitive binding assays with [125I]IRL 1620, unlabeled ET-1, ET-3, IRL 1620 and [monoiodo-Tyr13]-IRL 1620 showed almost identical displacement curves with Ki of 8 to 16 pM. However, [125I]IRL 1620 was dissociated from the binding sites by addition of an excess amount (100 nM) of any of these unlabeled peptides, each with the same t1/2 of 100 min. This was in marked contrast to [125I]ET-3 which was hardly dissociated from the binding sites.

Retraction concerning an endothelin B receptor-selective antagonist: Urade, Y., Fujitani, Y., Oda, K., Watakabe, T., Umemura, L., Takai, M., Okada, T., Sakata, K. and Karaki, H. (1992) FEBS Lett. 311, 12-16 : An endothelin B receptor-selective antagonist: IRL 1038, [Cys11-Cys15]-endothelin-1(11-21)

In the inhibition of specific binding of [125I]endothelins (ETs) to membrane from various tissues of rats, guinea pigs, pigs and humans, [Cys11-Cys15]-ET-1(11-21), IRL 1038, has a much higher affinity for ETB receptors (Ki = 6-11 nM) than for ETA receptors (Ki = 0.4-0.7 microM). In contraction assays, with ET-3 as a stimulant, 3 microM IRL 1038 antagonized the ETB receptor-mediated contraction of guinea pig ileal and tracheal smooth muscle without any significant agonistic activity, but did not effect the ETA receptor-mediated contraction of rat aortic smooth muscle. IRL 1038 is therefore, considered to be the first antagonist selective to the ETB receptor.The article describes the synthetic peptide IRL 1038 as having a much higher affinity to ETB receptors (Ki = 6 11 nM) than to ET, receptors (Ki = 4OC700 nM) in various tissue membranes. It was also shown that this peptide antagonized the ET,-mediated contraction of guinea pig ileal and tracheal smooth muscles. Since the publication of the data, we have synthesized more than 20 different batches of IRL 1038, some of which showed potencies and selectivities almost equivalent to the published data. However, the other batches, including batches synthesized by external suppliers, exhibited significantly higher Ki of 5&400 nM for the binding to ET, receptors in porcine lung membranes. We have very carefully investigated the physico-chemical properties of IRL 1038 in aqueous solutions at different pH (pH 7-10) and peptide concentrations, using fluorescence and proton NMR spectroscopy. At high pH (pH lo), the peptide does not aggregate and two kinds of non-aggregated states were observed by NMR spectroscopy. There was, however, no evident correlation between the two non-aggregated states and binding af-

Effect of Cathepsin K Inhibitors on Bone Resorption

On the basis of the pyrrolopyrimidine core structure that was previously discovered, cathepsin K inhibitors having a spiro amine at the P3 have been explored to enhance the target, bone marrow, tissue distribution. Several spiro structures were identified with improved distribution toward bone marrow. The representative inhibitor 7 of this series revealed in vivo reduction in C-terminal telopeptide of type I collagen in rats and monkeys.

IRL 2500: A potent ETB selective endothelin antagonist

Abstract Combination of a glycine substitution scan on the C-terminal dodecapeptide analog of ET-1 and a substance P antagonist screen on the basis of a homology study of the rhodopsin superfamily of seven-transmembrane receptors yielded in the development of IRL 2500, a potent ETB selective endothelin antagonist.

Structural study on silkworm eclosion hormone fragment (1–34) in solution by proton nuclear magnetic resonance spectroscopy

Eclosion hormone (EH) is a neuropeptide hormone which controls the ecdysis behavior in insect. The three dimensional structure of the N-terminal fragment (1-34) of the eclosion hormone which was predicted to contain a compact region crucial for the EH activity was studied in 50% d3-trifluoroethanol(TFE)/50% H2O at pH 3 and 298 K by 1H NMR spectroscopy with the combined use of distance geometry and molecular dynamics calculations. NMR results indicated that the fragment actually assumes an alpha-helix between Ala10 and Gln20, but no rigid structure is present from Cys21 through the C-terminus and for the N-terminal region (Ser1-Asp9). The elucidated structure was compared with the predicted structure of the native EH for the further development of the design of the insecticide.

Discovery of a Novel Piperidine-Based Inhibitor of Cholesteryl Ester Transfer Protein (CETP) That Retains Activity in Hypertriglyceridemic Plasma

Herein we describe the discovery and characterization of a novel, piperidine-based inhibitor of cholesteryl ester transfer protein (CETP) with a core structure distinct from other reported CETP inhibitors. A versatile synthesis starting from 4-methoxypyridine enabled an efficient exploration of the SAR, giving a lead molecule with potent CETP inhibition in human plasma. The subsequent optimization focused on improvement of pharmacokinetics and mitigation of off-target liabilities, such as CYP inhibition, whose improvement correlated with increased lipophilic efficiency. The effort led to the identification of an achiral, carboxylic acid-bearing compound 16 (TAP311) with excellent pharmacokinetics in rats and robust efficacy in hamsters. Compared to anacetrapib, the compound showed substantially reduced lipophilicity, had only modest distribution into adipose tissue, and retained potency in hypertriglyceridemic plasma in vitro and in vivo. Furthermore, in contrast to torcetrapib, the compound did not increase aldosterone secretion in human adrenocortical carcinoma cells nor in chronically cannulated rats. On the basis of its preclinical efficacy and safety profile, the compound was advanced into clinical trials.