Abstract 3494: LOXblock-1 Provides Efficient, Specific, And Long-lasting Neuroprotection Against Transient Focal Ischemia In Mice By Inhibiting 12/15-Lipoxygenase

Abstract

Background and Purpose - Acute phase stroke treatment options are currently very limited. We have previously demonstrated 12/15-lipoxygenase (12/15-LOX) to initiate an apoptotic cell death cascade, contributing to neuronal cell death, but also vascular leakage and edema formation in the ischemic brain. The current study was designed to test if a novel inhibitor LOXBlock-1, can specifically reduce 12/15-LOX activity in vivo, and provide efficient and long-lasting protection against stroke-induced injury. Methods - In vitro protection by LOXBlock-1 was tested in HT22 cells. A cell-based reporter assay determined activation of HIF1a and NRF2. The lipoxygenase metabolite 12-HETE was measured using a commercial EIA kit, and confirmed by HPLC/mass spectrometry. We injected LOXBlock-1 intraperitoneally in mice subjected to middle cerebral artery occlusion (MCAO). Immunohistochemistry was used to detect MDA-modified lysine in the ischemic brain, TTC and H&E stains were used to determine infarct size. Neurological severity score and corner test were used to evaluate behavioral outcome. Intracerebral hemorrhage was induced by collagenase injection. Results - Nanomolar LOXBlock-1 protected HT22 cells against oxidative glutamate toxicity. LOXBlock-1 reduced 12-HETE levels, without activating HIF1a- and NRF2-dependent pathways. One day after MCAO, MDA-modified lysine co-localized with 12/15-LOX in the peri-infarct cortex, and LOXBlock-1 reduced hemispheric swelling, levels of 12-HETE, and infarct size. After 14 days, infarct size remained reduced, and behavioral outcome was improved. LOXBlock-1 did not increase collagenase-induced hemorrhage. Conclusion - 12/15-LOX inhibition by LOXBlock-1 provides effective and long-lasting protection against transient focal ischemia in mice, without worsening hemorrhage. LOXBlock-1 may be a suitable acute phase stroke treatment.