Clinical Pharmacology of Recombinant Hirudin

Markwardt, Fritz; Nowak, G; Stürzebecher, Jörg

doi:10.1159/000216274

Cited by 16 publications

(14 citation statements)

References 11 publications

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“…Because hirudin is said to be highly thrombin-specific and pharmacodynamically inert (22,23), it appears that its protective effect is caused by inhibition of endogenous thrombin. This, in turn, strongly suggests that endogenous thrombin participates in ischemia-induced neurodegeneration in the hippocampus.…”

Section: Discussionmentioning

confidence: 99%

The protease thrombin is an endogenous mediator of hippocampal neuroprotection against ischemia at low concentrations but causes degeneration at high concentrations

Striggow

Riek

Breder

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

270

262

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We have considered the extracellular serine protease thrombin and its receptor as endogenous mediators of neuronal protection against brain ischemia. Exposure of gerbils to prior mild ischemic insults, here two relatively short-lasting occlusions (2 min) of both common carotid arteries applied at 1-day intervals 2 days before a severe occlusion (6 min), caused a robust ischemic tolerance of hippocampal CA1 neurons. This resistance was impaired if the specific thrombin inhibitor hirudin was injected intracerebroventricularly before each short-lasting insult. Thus, efficient native neuroprotective mechanisms exist and endogenous thrombin seems to be involved therein. In vitro experiments using organotypic slice cultures of rat hippocampus revealed that thrombin can have protective but also deleterious effects on hippocampal CA1 neurons. Low concentrations of thrombin (50 pM, 0.01 unit͞ml) or of a synthetic thrombin receptor agonist (10 M) induced significant neuroprotection against experimental ischemia. In contrast, 50 nM (10 units͞ml) thrombin decreased further the reduced neuronal survival that follows the deprivation of oxygen and glucose, and 500 nM even caused neuronal cell death by itself. Degenerative thrombin actions also might be relevant in vivo, because hirudin increased the number of surviving neurons when applied before a 6-min occlusion. Among the thrombin concentrations tested, 50 pM induced intracellular Ca 2؉ spikes in fura-2-loaded CA1 neurons whereas higher concentrations caused a sustained Ca 2؉ elevation. Thus, distinct Ca 2؉ signals may define whether or not thrombin initiates protection. Taken together, in vivo and in vitro data suggest that thrombin can determine neuronal cell death or survival after brain ischemia. T he extracellular serine protease thrombin, a well known, key player in blood coagulation and platelet activation, has been found to be expressed in different brain regions (1, 2). Its physiological importance in the central nervous system is emphasized further by the parallel expression of the highly specific thrombin inhibitor protease nexin-1 (3, 4) and PAR-1, the classical thrombin receptor (2, 5-7). Some recent evidence indicates that thrombin and its receptor might be involved in neurodegenerative processes observed after different insults such as stroke, traumatic brain injuries, and heart arrest or as a frequent consequence of bypass surgeries (8-11). Normal brain function depends critically on a permanent supply of glucose and oxygen. Depending on its source, a disruption of the cerebrospinal blood flow leads to global or focal ischemia (hypoxia͞ hypoglycemia) and irreversible neuronal damage. Prothrombin as well as the classical thrombin receptor are expressed in brain regions that are particularly vulnerable to ischemia, e.g., neocortex, cortex, striatum, hypothalamus, hippocampus, and cerebellum (2). Furthermore, studies performed on isolated cells (neurons, astrocytes) have demonstrated that nanomolar concentrations of thrombin exert cytotoxic effects (12-14). How...

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Section: Discussionmentioning

confidence: 99%

The protease thrombin is an endogenous mediator of hippocampal neuroprotection against ischemia at low concentrations but causes degeneration at high concentrations

Striggow

Riek

Breder

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

270

262

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“…The analogous thrombin inhibitor, hirudin, was isolated from the leech Hirudo medicinalis. Hirudin is also a potent (K i 20 fM) and competitive thrombin inhibitor and a purified recombinant version has entered clinical use [38][39][40][41]. Comparative studies with both proteins have concluded that TAP yields an equivalent anticoagulant benefit with a reduced incidence of bleeding side effects [42,43].…”

Section: Possible Points Of Interventionmentioning

confidence: 99%

Perspectives on Factor Xa Inhibition

Rai¹,

Sprengeler²,

Elrod³

et al. 2001

CMC

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Blood coagulation involves a complex cascade of enzymatic reactions, ultimately generating fibrin, the basis of all blood clots. This cascade is comprised of two arms, the intrinsic and extrinsic pathways which converge at factor Xa to form the common pathway. Factor Xa activates prothrombin to thrombin, which in turn catalyzes the conversion of fibrinogen to fibrin. Recently, both natural and synthetic factor Xa inhibitors have shown promising pharmacological effects in animal models of thrombosis. Accordingly, factor Xa has emerged as a compelling target for pharmacological intervention and much recent effort has focused on selective and potent inhibition of this key enzyme. Factor Xa and other enzymes in the coagulation cascade belong to the trypsin-like serine protease family, the various members of which are involved in numerous physiological functions in the body. Hence, to avoid toxicity and adverse side effects, it is important to selectively inhibit the target enzyme. Achieving the needed selectivity has proved challenging due to the high degree of structural homology around the active site of this class of enzymes. This article provides a brief review of the strategies currently being employed to develop oral anticoagulants and, more specifically, the structural features of protein-ligand binding that have been utilized to achieve potency and selectivity toward factor Xa. Additionally, selected lead molecules will be discussed to highlight binding motifs used to attain both potency and selectivity in drug candidates.

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“…After pharmacotoxicological profiling of rH, we began preclinical and clinicopharmacological investigations in 1988 (82,85, 87,90,92,105). These studies and subsequent ones from other researchers showed that the pharmacological properties of rH are very similar to those of nH.…”

Section: Preclinical Studiesmentioning

confidence: 99%

Hirudin: The Promising Antithrombotic

Markwardt

1992

Cardiovascular Drug Reviews

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It has long been known that medicinal leeches (Hirudo medicinalis) contain a substance with anticoagulant properties. In early studies, aqueous extracts and dry preparations from leeches containing minute amounts of the active substance were used to prevent blood from clotting. Only after the development of protein chemistry and the elucidation of the biochemistry of blood coagulation was the anticoagulant agent, hirudin, isolated and its chemical nature and mode of action clarified.The historical background of hirudin isolation as well as the individual steps of its biochemical and pharmacological characterization have been described in several reviews (36,68,70,72,74,79). BIOCHEMISTRYHirudin is extracted from the homogenized heads of medicinal leeches and enriched by precipitation procedures followed by ion-exchange chromatography and gel filtration (2,65,98,137). Affinity chromatography on matrix-bound thrombin can be used to obtain highly purified hirudin preparations (139,140). The pure anticoagulant agent obtained is a carboxyhydrate-free single chain miniprotein containing 65 amino acids with a molecular weight of about 7 kDa. The amino acid composition is characterized by a remarkably high content of acidic amino acids at the C-terminal, the absence of arginine, methionine, and tryptophan, and the presence of a sulfated tyrosine residue and three intramolecular disulfide bridges (62,65,70).Considerable progress in the isolation, purification, and analysis of hirudin has shown the medicinal leech generates several variants of the miniprotein in trace amounts. The isoinhibitor forms share 85-90% amino acid sequence homology. Their main structural elements, which include N-terminal hydrophobic amino acids, followed by a core region linked by three disulfide bridges and the unique clustering of acidic amino acid residues in the C-terminal portion remain constant (4,17,18,113). The N-terminal amino acids Val-Val can be replaced by Ile-Thr (Fig. 1).

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Clinical Pharmacology of Recombinant Hirudin

Cited by 16 publications

References 11 publications

The protease thrombin is an endogenous mediator of hippocampal neuroprotection against ischemia at low concentrations but causes degeneration at high concentrations

The protease thrombin is an endogenous mediator of hippocampal neuroprotection against ischemia at low concentrations but causes degeneration at high concentrations

Perspectives on Factor Xa Inhibition

Hirudin: The Promising Antithrombotic

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