Drug News & Perspectives 2010, 23(9): 549-556 THOMSON REUTERS LOOKING AHEAD Targeting both NMDA receptors and free NEU2000, AN NR2B-SELECTIVE, radicals may provide MODERATE NMDA RECEPTOR enhanced ANTAGONIST AND POTENT SPIN neuroprotection against TRAPPING MOLECULE FOR hypoxic-ischemic injury. STROKE confer substantial neuroprotection in ani- by Sung Ig Cho, Ui Jin Park, mal models of stroke have failed to show SUMMARY Jun-Mo Chung and Byoung Joo Gwag beneficial effects in clinical trials for stroke. Excess activation of ionotropic gluta- Free radicals mediate an additional route of mate receptors, primarily N-methyl-D- Stroke is a cerebrovascular injury caused by neuronal cell death after ischemia and aspartate (NMDA) receptors and free the interruption of blood flow to the brain reperfusion. Several antioxidants have radicals, evoke nerve cell death follow- due to thrombosis, embolic particles or advanced to clinical trials including edar- ing hypoxic-ischemic brain injury in var- blood vessel bursts. Stroke is the leading avone, a hydroxyl radical scavenger that has ious animal models. However, clinical cause of serious, long-term disability in shown beneficial effects in patients with trials in stroke patients using NMDA adults and the second leading cause of transient ischemia and which was approved receptor antagonists have failed to death in the U.S. and Europe (1). Rates of as a neuroprotective drug in Japan and show efficacy primarily due to the limit- stroke mortality and burden are more China. ed therapeutic time window for neuro- affected in low-income countries including protection and a narrow therapeutic NMDA receptor antagonists and antioxi- eastern Europe, northern Asia and central index. In comparison, antioxidants pro- dants are expected to confer neuroprotec- Africa (2). Ischemic stroke, a major type of longed the time window for neuropro- tion in human stroke if drug safety and the stroke caused by thrombosis and arterial tection in animal models of ischemic therapeutic time window are justified. As embolism, can cause neuronal death rapid- stroke and showed greater therapeutic NMDA receptor activation and free radicals ly in core areas, which is accompanied by potential in clinical trials for ischemic are expected to contribute to neuronal secondary death in the ischemic penumbra stroke. Excess activation of NMDA death through distinct signaling pathways subsequent to activation of multiple death receptors and free radicals mediate the at different points of time after hypoxic pathways. Tissue plasminogen activator, a two separate pathways of nerve cell ischemic injury, it is conceivable to reason thrombolytic agent approved in 1996 by the death in stroke and a safe and multi- that targeting both NMDA receptors and U.S. Food & Drug Administration, is used to functional drug that can block both free radicals may provide enhanced neuro- dissolve the clot and to improve neurologi- routes in the brain will likely provide a protection against hypoxic-ischemic injury cal outcome for acute ischemic stroke with- better therapeutic outcome in patients and increase the therapeutic time window, if in 3 hours after the onset of symptoms (3). with stroke. Derivatives of the lead safety is compromised. Mechanisms and interventional therapies structures of sulfasalazine and aspirin for ischemic neuronal death have been have led to the discovery of a new mol- THE ROLE OF NMDA RECEPTORS IN extensively investigated to establish a novel ecule, Neu2000, that has demonstrat- ISCHEMIC NEURONAL DEATH neuroprotection therapy with a greater ther- ed excellent neuroprotection against apeutic time window than that of throm- The excitatory neurotransmitter glutamate NMDA- and free radical-induced cell bolytic agents (4). Preclinical studies have is released and accumulates in ischemic death. Neu2000 is an NR2B-selective, provided extensive evidence that excess brain areas deprived of glucose and oxygen moderate NMDA receptor antagonist (Fig. 1). Abnormal accumulation of gluta- activation of N-methyl-D-aspartate (NMDA) with potent cell-permeable, spin trap- 2+ receptors mediates rapidly evolving neu- mate triggers influx and overload of Ca ping antioxidant action even at ronal death following ischemic insults. primarily through excess activation of the nanomolar concentrations. Nonclinical However, NMDA receptor antagonists that NMDA receptors. This increase in intracellu- and human phase I studies demonstrat- 2+ 2+ lar Ca ([Ca ]i) in neurons then triggers ed that Neu2000 can be translated to neuronal injury by activating cytotoxic pro- treat patients with stroke with better Correspondence: B.J. Gwag, [email protected] or teins such as nitric oxide synthase, calpain efficacy and therapeutic time window. [email protected] and endonuclease and by impairing mito- Copyright © 2010 Prous Science, S.A.U. or its licensors. All rights reserved. CCC: 0214-0934/2010. DOI: 10.1358/dnp.2010.23.9.1513493 549 NEU2000 FOR STROKE S.I. Cho et al. ROLE OF FREE RADICALS IN ISCHEMIC NEURONAL DEATH Ischemic injury produces free radicals through multiple pathways. Activation of 2 NMDA receptors causes the accumulation ↓ 2+ of [Ca ]i that triggers production of super- oxide and nitric oxide via the activation of xanthine oxidase, nitric oxide synthase and 2+ NADPH (nicotinamide adenine dinucleotide phosphate) oxidase (14-16). Reoxygenation ↑ or reperfusion can give rise to superoxide and H2O2 by the oxidation of accumulated hypoxanthine and the uncoupling of elec- · ↑ ↑ 2 2 tron flow from mitochondria electron trans- port chain (Fig. 1) (4). During reperfusion, 2+ transient metal ions such as ferrous and fer- 2+ ↑ ric ions are uptaken and accumulate in vul- nerable cells, which results in •OH produc- · tion by the Fenton and the Haber–Weiss reactions (17-20). The therapeutic potential of antioxidants has been well documented in animal models of stroke with a longer therapeutic time window than NMDA recep- tor antagonists (21). Four antioxidants have advanced to clinical trials for acute Figure 1. Role of NMDA receptors and free radicals in ischemic injury. Interrupted blood flow to the brain ischemia. First, tirilazad, a lipid peroxidation causes deprivation of oxygen and glucose () that results in loss of ATP (), leading to membrane inhibitor, failed to show significant beneficial 2+ depolarization, Ca entry into the presynaptic terminal () and glutamate release from the synaptic effects in six randomized, controlled trials in vesicles into the synaptic cleft (). Accumulated glutamate causes influx and accumulation of Ca2+ patients with acute ischemic stroke. An through excess activation of postsynaptic NMDA receptors (NMDAR) ( ). Sustained Ca2+ overload for a few minutes can induce neuronal death. Following reperfusion of reoxygenation, cells are exposed to additional phase III study using a higher .– dosage of tirilazad was stopped with ques- excess oxygen molecules that can be converted to superoxide (O2 ) and hydrogen peroxide (H2O2) in .– mitochondria. O2 and H2O2 can be further converted to hydroxyl radicals (•OH) through the Fenton tions emerging in Europe regarding its safe- reaction and the Haber–Weiss reaction catalyzed by ferrous and ferric ions. Such toxic free radicals ty (22). Secondly, the administration of mediate slowly evolving secondary nerve cell death following ischemic injury. ebselen, a glutathione peroxidase mimic, to patients with acute ischemic stroke within 24 hours after stroke onset, improved out- come for 1 month but not after 3 months, in chondria and endoplasmic reticulum. receptor subtype 2B (NR2B)-selective man- Japan (23, 24). The modified Mathew Scale 2+ Neurons overloaded with Ca for even a few ners, respectively, showed no efficacy in and modified Barthel Index scores were also minutes can undergo neuronal death, sug- phase III trials for acute stroke therapy (9). improved in stroke patients subjected to gesting that NMDA receptors mediate rapid The lack of beneficial effects of NMDA early treatment with ebselen. In Japan, a and fulminant neuronal death following receptor antagonists in clinical trials is placebo-controlled, double-blind, random- ischemic injury (5, 6). Furthermore, NMDA largely attributable to the narrow therapeu- ized, multicenter trial of ebselen has been receptor antagonists prevent Ca2+ entry and tic time window and unwanted side effects initiated for patients with acute cortical overload in cultured neurons deprived of such as neuronal vacuolization and necrosis infarction but further clinical outcomes have glucose and oxygen and markedly attenuate primarily in the posterior cingulate and ret- not yet been reported. Thirdly, the phase III neuronal death induced by oxygen-glucose rosplenial cortex (10-12). Moreover, adminis- study of edaravone, a free radical scavenger, deprivation and in animals subjected to tration of potent NMDA receptor antago- was performed with 250 patients with acute ischemic stroke in Japan, which showed a transient cerebral ischemia (7, 8). nists can produce psychiatric adverse effects significant improvement in functional out- in humans as shown with selfotel, which In contrast to the neuroprotective effects come within 3 months, as evaluated by the verified in animal models of stroke, NMDA caused marked psychiatric and neurological modified Rankin scale (25). Edaravone was receptor antagonists have not improved side effects in human stroke patients at a approved for the treatment of acute stroke neurological outcomes in stroke patients subtherapeutic plasma