Toward safer thrombolytic agents in stroke: molecular requirements for NMDA receptor-mediated neurotoxicity.
Current thrombolytic therapy for acute ischemic stroke with tissue-type plasminogen activator (tPA) has clear global benefits. Nevertheless, evidences argue that in addition to its prohemorrhagic effect, tPA might enhance excitotoxic necrosis. In the brain parenchyma, tPA, by binding to and then cleaving the amino-terminal domain (ATD) of the NR1 subunit of N-methyl-D-aspartate (NMDA) glutamate receptors, increases calcium influx to toxic levels. We show here that tPA binds the ATD of the NR1 subunit by a two-sites system (K(D)=24 nmol/L). Although tenecteplase (TNK) and reteplase also display two-sites binding profiles, the catalytically inactive mutant TNKS478A displays a one-site binding profile and desmoteplase (DSPA), a kringle 2 (K2) domain-free plasminogen activator derived from vampire bat, does not interact with NR1. Moreover, we show that in contrast to tPA, DSPA does not promote excitotoxicity. These findings, together with three-dimensional (3D) modeling, show that a critical step for interaction of tPA with NR1 is the binding of its K2 domain, followed by the binding of its catalytic domain, which in turn cleaves the NR1 subunit at its ATD, leading to a subsequent potentiation of NMDA-induced calcium influx and neurotoxicity. This could help design safer new generation thrombolytic agents for stroke treatment.
Lopez-Atalaya
Jose P
JP
INSERM, INSERM U919 'serine proteases and pathophysiology of the neurovascular unit', GIP Cyceron, Caen Cedex, France.
Roussel
Benoit D
BD
Levrat
Denis
D
Parcq
Jérôme
J
Nicole
Olivier
O
Hommet
Yannick
Y
Benchenane
Karim
K
Castel
Hervé
H
Leprince
Jérôme
J
To Van
Denis
D
Bureau
Ronan
R
Rault
Sylvain
S
Vaudry
Hubert
H
Petersen
Karl-Uwe
KU
Santos
Jana Sopkova-de Oliveira
JS
Ali
Carine
C
Vivien
Denis
D
D000595:Amino Acid Sequence; D000818:Animals; D020134:Catalytic Domain; D002478:Cells, Cultured; D064420:Drug-Related Side Effects and Adverse Reactions; D005343:Fibrinolytic Agents / Q000633:toxicity*; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D010960:Plasminogen Activators / Q000378:metabolism; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D016194:Receptors, N-Methyl-D-Aspartate / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D020521:Stroke / Q000378:metabolism*; D046509:Tissue Culture Techniques; D010959:Tissue Plasminogen Activator / Q000737:chemistry / Q000378:metabolism
D005343:Fibrinolytic Agents; C409466:NR1 NMDA receptor; D016194:Receptors, N-Methyl-D-Aspartate; C069588:salivary plasminogen activator alpha 1, Desmodus rotundus; D010960:Plasminogen Activators; D010959:Tissue Plasminogen Activator