The novel calpain inhibitor SJA6017 improves functional outcome after delayed administration in a mouse model of diffuse brain injury.
A principal mechanism of calcium-mediated neuronal injury is the activation of neutral proteases known as calpains. Proteolytic substrates for calpain include receptor and cytoskeletal proteins, signal transduction enzymes and transcription factors. Recently, calpain inhibitors have been shown to provide benefit in rat models of focal head injury and focal cerebral ischemia. The present study sought to investigate, in experiment 1, the time course of calpain-mediated cytoskeletal injury in a mouse model of diffuse head injury by measuring the 150- and 145-kDa alpha-spectrin breakdown products (SBDP). Secondly, in experiment 2, we examined the effect of early (20 min postinjury) administration of the novel calpain inhibitor SJA6017 on functional outcome measured 24 h following injury and its effect on posttraumatic alpha-spectrin degradation. Lastly, in experiment 3, we examined the effect of delayed (4 or 6 h postinjury) administration of SJA6017 on 24-h postinjury functional outcome. In experiment 1, isoflurane-anesthetized male CF-1 mice (18-22 g) were subjected to a 750 g-cm weight drop-induced injury and were sacrificed for SBDP analysis at postinjury times of 30 min, and 1, 2, 6, 24 and 48 h (plus sham). In experiments 2 and 3, mice were injured as described, and delivered a single tail vein injection of either SJA6017 (0.3, 1, or 3 mg/kg) or vehicle (administered immediately, 4 or 6 h postinjury [3 mg/kg]). Functional outcome was evaluated in both studies, and, in experiment 2, 24-h postinjury assessment of SBDPs was determined. Following injury, the level of SBDP 145 was significantly different from sham at 24 and 48 h in cortical and at 24 h in the hippocampal tissues and at 48 h in the striatum. Immediate postinjury administration of SJA6017 resulted in a dose-related improvement in 24-h functional outcome (p < 0.05 at 3 mg/kg). Significance was maintained after a 4-h delay of the 3 mg/kg, but was lost after a 6-h delay. Despite improvement in functional outcome at 24 h, SJA6017 did not reduce spectrin breakdown in cortical or hippocampal tissues. These results support a role for calpain-mediated neuronal injury and the potential for a practical therapeutic window for calpain inhibition following traumatic brain injury. However, measurements of regional spectrin degradation may not be the most sensitive marker for determining the effects of calpain inhibition.
Kupina
N C
NC
Neuroscience Therapeutics, Pfizer Global Research and Development-Ann Arbor Laboratories, Michigan 48015, USA.
Nath
R
R
Bernath
E E
EE
Inoue
J
J
Mitsuyoshi
A
A
Yuen
P W
PW
Wang
K K
KK
Hall
E D
ED
D000818:Animals; D001522:Behavior, Animal / Q000187:drug effects; D015153:Blotting, Western; D001930:Brain Injuries / Q000188:drug therapy* / Q000473:pathology / Q000523:psychology; D002154:Calpain / Q000037:antagonists & inhibitors*; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism; D003599:Cytoskeleton / Q000473:pathology; D004151:Dipeptides / Q000627:therapeutic use*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000627:therapeutic use*; D008297:Male; D051379:Mice; D013049:Spectrin; D013997:Time Factors
D004151:Dipeptides; D004791:Enzyme Inhibitors; C109145:N-(4-fluorophenylsulfonyl)-L-valyl-L-leucinal; D013049:Spectrin; D002154:Calpain