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Marked prevention of ischemic brain injury by Neu2000, an NMDA antagonist and antioxidant derived from aspirin and sulfasalazine

Neurotech Pharmaceuticals Co., Ajou University School of Medicine, Suwon, Gyeonggido, South Korea.
Journal of Cerebral Blood Flow & Metabolism (Impact Factor: 5.34). 07/2007; 27(6):1142-51. DOI: 10.1038/sj.jcbfm.9600418
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ABSTRACT Excitotoxicity and oxidative stress mediate neuronal death after hypoxic-ischemic brain injury. We examined the possibility that targeting both N-methyl-D-aspartate (NMDA) receptor-mediated excitotoxicity and oxidative stress would result in enhanced neuroprotection against hypoxic-ischemia. 2-Hydroxy-5-(2,3,5,6-tetrafluoro-4-trifluoromethyl-benzylamino)-benzoic acid (Neu2000) was derived from aspirin and sulfasalazine to prevent both NMDA neurotoxicity and oxidative stress. In cortical cell cultures, Neu2000 was shown to be an uncompetitive NMDA receptor antagonist and completely blocked free radical toxicity at doses as low as 0.3 micromol/L. Neu2000 showed marked neuroprotection in a masked fashion using histology and behavioral testing in two rodent models of focal cerebral ischemia without causing neurotoxic side effects. Neu2000 protected against the effects of middle cerebral artery occlusion, even when delivered 8 h after reperfusion. Single bolus administration of the drug prevented gray and white matter degeneration and spared neurologic function for over 28 days after MACO. Neu2000 may be a novel therapy for combating both NMDA receptor-mediated excitotoxicity and oxidative stress, the two major routes of neuronal death in ischemia, offering profound neuroprotection and an extended therapeutic window.

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    • "Neu2000 [2-hydroxy-5-(2,3,5,6-tetrafluoro-4 trifluoromethyl benzylamino)-benzoic acid] is a derivative of acetylsalicylic acid and sulfasalazine, a conjugate of 5-aminosalicylic acid and sulfapyridine (Gwag et al., 2007). Neu2000 has been shown to be neuroprotective against N-methyl-D-aspartate (NMDA)-and ironmediated neurotoxicity in cortical neuron cultures (Gwag et al., 2007; Cho et al., 2010). Neu2000 also limited the activation of proapoptotic proteins in a mouse model of amyotrophic lateral "
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    ABSTRACT: Neu2000 [2-hydroxy-5-(2,3,5,6-tetrafluoro-4 trifluoromethylbenzylamino) benzoic acid] is a dual-acting neuroprotective agent that functions both as a noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonist and a free radical scavenger. In the present study, we investigated the scavenging activity of Neu2000 on various classes of reactive oxygen species and reactive nitrogen species (ROS/RNS) as well as its efficacy for reducing free radicals and oxidative stress/damage induced in spinal cord mitochondrial preparations. Neu2000 exerted scavenging activity against superoxide, nitric oxide, and hydroxyl radicals, and efficiently scavenged peroxynitrite. In the mitochondrial studies, Neu2000 markedly inhibited ROS/RNS and hydrogen peroxide levels following antimycin treatment. In addition, Neu2000 effectively scavenged hydroxyl radicals generated by iron(III)-ascorbate, reduced protein carbonyl formation mediated by hydroxyl radicals and peroxynitrite, and prevented glutathione oxidation caused by tert-butyl hydroperoxide in isolated mitochondria. Interestingly, incubation of isolated mitochondria with Neu2000 followed by centrifugation and removal of the supernatant also resulted in a concentration-dependent decrease in lipid peroxidation. This observation suggests that Neu2000 enters mitochondria to target free radicals or indirectly affects mitochondrial function in a manner that promotes antioxidant activity. The results of the present study demonstrate that Neu2000 possesses potent in vitro antioxidant activity due, most likely, to its active phenoxy group.
    Toxicology in Vitro 12/2012; 27(2). DOI:10.1016/j.tiv.2012.12.011 · 3.21 Impact Factor
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    • "Focal cerebral ischemia Male Sprague–Dawley rats (260–300 g) were anesthetized with chloral hydrate (400 mg/kg, i.p.) and subjected to focal cerebral ischemia by transient occlusion of the right middle cerebral artery (rMCA) and both common carotid arteries (CCAs), as previously described (Tamura et al. 1981; Gwag et al. 2007). In brief, the CCAs were exposed through a vertical midline neck incision. "
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    ABSTRACT: Alternative splicing of tau exon 10 influences microtubule assembly and stability during development and in pathological processes of the central nervous system. However, the cellular events that underlie this pre-mRNA splicing remain to be delineated. In this study, we examined the possibility that ischemic injury, known to change the cellular distribution and expression of several RNA splicing factors, alters the splicing of tau exon 10. Transient occlusion of the middle cerebral artery reduced tau exon 10 inclusion in the ischemic cortical area within 12 h, resulting in the induction of three-repeat (3R) tau in cortical neurons. Ubiquitinated protein aggregates and reduced proteasome activity were also observed. Administration of proteasome inhibitors such as MG132, proteasome inhibitor I and lactacystin reduced tau exon 10 splicing in cortical cell cultures. Decreased levels of Tra2beta, an RNA splicing factor responsible for tau exon 10 inclusion, were detected both in cortical cell cultures exposed to MG132 and in cerebral cortex after ischemic injury. Taken together, these findings suggest that transient focal cerebral ischemia reduces tau exon 10 splicing through a mechanism involving proteasome-ubiquitin dysfunction and down-regulation of Tra2beta.
    Journal of Neurochemistry 04/2010; 114(1):160-70. DOI:10.1111/j.1471-4159.2010.06732.x · 4.24 Impact Factor
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    • "Based upon the pharmacological actions of sulfasalazine, the novel neuroprotectant 2-hydroxy-5- (2,3,5,6-tetrafluoro-4-trifluoromethyl-benzylamino)-benzoic acid was designed and named 'Neu2000/NEU'. NEU is much more potent than sulfasalazine in protecting neurons against NMDAR-mediated excitotoxicity and free radical injury (Gwag et al. 2007). NEU dramatically reduces glutamate toxicity and prevents ischemic cell death with a fairly long therapeutic window. "
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    ABSTRACT: Neu2000 [NEU, 2-hydroxy-5-(2,3,5,6-tetrafluoro-4-trifluoromethyl-benzylamino)-benzoic acid], a derivative of sulfasalazine, attenuates NMDA-induced neuronal toxicity. Here we investigated the effects of NEU on the NMDA receptor (NMDAR) using whole-cell patch clamp technique to determine the molecular mechanisms underlying its neuroprotective role. NEU reversibly suppressed NMDA responses in an uncompetitive manner with fast binding kinetics. Its inhibition of NMDAR activity depended on both the concentration and the use of agonist but not on the membrane potential. NEU accelerated NMDA desensitization without affecting the binding affinity of NMDAR for its agonists and stabilized the closed state of NMDAR. Therefore, NEU should effectively alleviate disorders that are a result of glutamate excitoxicity with fewer side effects because it is a low-affinity gating modifier that antagonizes NMDAR in an uncompetitive manner. Moreover, in the presence of ifenprodil (an NR2B antagonist) but not NVP-AAM077 [(R)-[(S)-1-(4-bromo-phenyl)-ethylamino]-(2,3-dioxo-1,2,3,4-tetrahydro-quinoxalin-5-yl)-methyl]-phosphonic acid, an NR2A antagonist], the extent of NEU block was decreased, suggesting that NEU is an NR2B-specific antagonist.
    Journal of Neurochemistry 04/2009; 109(5):1261-71. DOI:10.1111/j.1471-4159.2009.06044.x · 4.24 Impact Factor
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