Direct thrombin inhibitors built on the azaphenylalanine scaffold provoke degranulation of mast cells

Luka Peternel; Mateja Štempelj; Manica Cerne; Anamarija Zega; Aleš Obreza; Marko Oblak; Gorazd Drevenšek; Metka V. Budihna; Lovro Stanovnik; Uroš Urleb

doi:10.1160/TH05-10-0683

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2006; 95(02): 294-300
DOI: 10.1160/TH05-10-0683

Wound Healing and Inflammation/Infection

Schattauer GmbH

Direct thrombin inhibitors built on the azaphenylalanine scaffold provoke degranulation of mast cells

Authors

Luka Peternel

¹Drug Discovery, Lek Pharmaceuticals d.d., Ljubljana, Slovenia

²Institute of Pharmacology and Experimental Toxicology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia
Mateja Štempelj

²Institute of Pharmacology and Experimental Toxicology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia
Manica Cerne

¹Drug Discovery, Lek Pharmaceuticals d.d., Ljubljana, Slovenia
Anamarija Zega

³Faculty of Pharmacy, University of Ljubljana, Ljubljana, Slovenia
Aleš Obreza

³Faculty of Pharmacy, University of Ljubljana, Ljubljana, Slovenia
Marko Oblak

¹Drug Discovery, Lek Pharmaceuticals d.d., Ljubljana, Slovenia
Gorazd Drevenšek

²Institute of Pharmacology and Experimental Toxicology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia
Metka V. Budihna

²Institute of Pharmacology and Experimental Toxicology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia
Lovro Stanovnik

²Institute of Pharmacology and Experimental Toxicology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia
Uroš Urleb

¹Drug Discovery, Lek Pharmaceuticals d.d., Ljubljana, Slovenia

³Faculty of Pharmacy, University of Ljubljana, Ljubljana, Slovenia

Abstract

Summary

The main structural feature of direct thrombin inhibitor LK-732 responsible for the appropriate interaction at the thrombin active site is a strong basic group. A possibility that a strong basic group of LK-732 might contribute to the mast cell degranulation effect and consequent reduction of tracheal air flow (TAF) and fall of mean arterial blood pressure (MAP) in rats was investigated in the present study. At doses up to5 mg/kg (i. v.), LK-732 did not cause significant changes of TAF and MAP. At 7 mg/kg (i. v.),a sudden reduction of TAF and a fall of MAP was observed within 5 min after LK-732 administration (75% mortality, p=0. 007). A less basic direct thrombin inhibitor LK-658 (21 mg/ kg, i. v.) did not significantly disturb TAF and MAP. A reduction of TAF and a fall of MAP caused by LK-732 (7 mg/kg, i. v.) was almost completely abolished in rats with degranulated mast cells (0% mortality, p=0. 008). LK-732 concentration-dependently degranulated rat peritoneal mast cells in vitro (pEC₅₀=1. 92±0. 05 µM). A structure-activity relationship (SAR) study revealed that the terminal basic groups attached to the aromatic ring are responsible for the mast cell degranulation effect. A good correlation was observed between mast cell degranulation and pK_b of analogues of LK-732 (R²=0. 49), but not between mast cell degranulation and thrombin K_i (R²=0. 23). LK-732-induced reduction of TAF, the fall of MAP and high mortality originate from LK732-induced mast cell degranulation. As judged by the SAR study, this effect could be overcome by reducing the basicity of LK-732.

Keywords

Thrombin inhibitors - azaphenylalanine scaffold - mast cell degranulation - basicity - rat

Full Text

References

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