Stapled Peptides with Therapeutic Potential for Ischemic Stroke by Blocking the Endocytosis of GluA2 AMPAR.

Marc Fisher MD Kennedy Lees MD Section Editors: The neutral results of the SAINT II study1 dashed the hopes for a second ischemic stroke drug in the near future. Although this is certainly disappointing for stroke neurologists, a careful analysis of the NXY-059 development program reveals several deficiencies, which if properly addressed, might revitalize and strengthen the field of neuroprotection for acute ischemic stroke. This critique points out several potential reasons for the failure of SAINT II to replicate the positive findings of SAINT 1,2 discusses the preclinical problems with NXY-059, and provides a perspective on the development of future neuroprotective agents for acute ischemic stroke. The SAINT II trial1 investigated the efficacy of the free-radical–trapping agent, NXY-059, for the treatment of acute ischemic stroke and failed to confirm the positive results of SAINT I.2 In the latter study with 1699 patients, NXY-059 showed for the first time in a large phase III trial that a drug other than tissue plasminogen activator (t-PA) might lead to significant improvement on the modified Rankin Scale (mRS) using a novel and controversial outcome analysis (Rankin shift analysis). The larger SAINT II trial with 3306 patients not only failed to replicate improvement on the same primary end point but all secondary end points (neurological deficits on the National Institutes of Health Stroke Scale (NIHSS) at 90 days and activities of daily living on the Barthel Index at 90 days) were identical between treatment groups. Besides the obvious possibility that the drug itself was inactive, which may be considered due to the known instability of free-radical–trapping agents, several additional problems were evident. Both trials were designed, in part, as combination studies, testing the addition of NXY-059 to standard therapy compared with t-PA alone in the 3-hour time window. SAINT II gave …

The development of additional effective therapies for acute ischemic stroke remains a challenging but critical endeavor. Intravenous recombinant tissue plasminogen activator (rtPA) initiated within 3 hours of stroke onset remains the only approved and validated therapy for acute ischemic stroke, and regulatory approval has expanded recently. Many other therapies have been evaluated, and these trials have either been inconclusive or negative.1 These acute stroke trials do provide valuable information concerning how to implement future trials and some glimmers of hope about existing data. Some of the lessons learned from prior acute stroke trials that will help to guide future trials are outlined below. The two fundamental approaches to the development of acute stroke therapy remain reperfusion and neuroprotection. This short review will focus on the current status of both approaches and how they might be combined, hopefully in the near future. Negative acute stroke treatment trials may be explained by the following: 1. The agents evaluated in clinical trials may not have been adequately tested in preclinical studies to provide robust confirmation of efficacy in appropriate animal models. 2. Side effects precluded adequate drug assessment or did not allow use of adequate drug concentrations. 3. Because of macro-occlusions and, perhaps, micro-occlusions, the drug did not penetrate into or beyond the penumbral tissue in adequate concentrations. 4. Trials included patients not appropriate for the purported mechanism of action of the drug being tested. 5. Patients were included too late after stoke onset to allow for adequate assessment of the drug’s efficacy, and imaging studies were not done to identify patients with appropriate tissue for treatment. 6. Trials have been inadequately powered to detect modest treatment effects. 7. Trials included too many patients with mild or very severe deficits in whom treatment effects are likely difficult to assess with currently used outcome measures. 8. The single primary outcome measure …

Protecting the brain from stroke: Huoxue Rongluo formula (HXRLF) targets ferroptosis for neuroprotection.

Introduction: Transplantation of neural stem cells (NSCs) provides a promising therapy for stroke. Its efficacy, however, might be limited because of massive grafted-cell death following transplantation, as well as insufficient capability for tissue repair. Chemical preconditioning primes the cells to the ‘‘state of readiness’’ to withstand the rigors of lethal ischemia. In this study, we investigated whether cellular reprogramming of NSCs with minocycline or interleukin-6 (IL-6), both of which are known to possess cytoprotective properties, facilitates the effectiveness of stem cell therapy in ischemic stroke. Methods: NSCs harvested from the subventricular zone of fetal mice were preconditioned with minocycline (10 μM) or IL-6 (20 ng/ml) for 24 hours in vitro . For an in vitro study, the NSCs were subjected to oxygen-glucose deprivation and reoxygenation (n=4). For an in vivo study, the NSCs were transplanted into mouse brains 6 hours or 7 days after transient middle cerebral artery occlusion (n=8). Histological and behavioral tests were examined from days 0 to 28 after stroke. Results: Minocycline preconditioning up-regulated the expression of Nrf2 (2.7-fold), as well as Nrf2-regulated antioxidant genes (HO-1: 4.0-fold/NQO1: 20-fold), and induced the NSCs to release paracrine factors, including BDNF, NGF, GDNF, and VEGF. IL-6 preconditioning activated STAT3-mediated up-regulation of SOD2 (2.3-fold), a primary mitochondrial antioxidant enzyme, and induced VEGF secretion (2.2-fold). Transplantation of minocycline- or IL-6-preconditioned NSCs significantly reduced grafted cell death, attenuated infarct size, and improved neurological performance compared with non-preconditioned NSCs. This minocycline- or IL-6-induced amelioration of ischemic insults was abolished by transfecting the NSCs with Nrf2- or STAT3-small interfering RNA before transplantation. Conclusions: Chemical preconditioning, which reprograms NSCs to tolerate oxidative stress after ischemic reperfusion injury and to induce trophic effects, is a simple and beneficial approach for enhancing the effectiveness of cell transplantation therapy in ischemic stroke.

Section Editors: Marc Fisher MD Antoni Davalos MD The Food and Drug Administration (FDA) evaluates applications for new human drugs, biologics, and complex medical devices. Companies must obtain FDA approval to legally market these products. In August, the FDA gave Concentric Medical clearance to market its Merci Retriever system to “remove blood clots from the brain in patients experiencing an ischemic stroke.” Given that the FDA is charged with “protecting the public health by assuring the safety, efficacy, and security of… biological products and medical devices…, ” “advancing public health by helping to speed innovations that make medicines … more effective, safer, and more affordable,” and “helping the public get the accurate, science-based information they need to use medicines … to improve their health,”1 the FDA’s decision to approve the Merci Retriever system is of concern. The pathways to approval are reviewed by Felten et al in the accompanying article and are outlined in Figure 1. Figure 1. Potential pathways for device approval. The decision to approve the Merci Retriever was based on data from the MERCI (Mechanical Embolus Removal in Cerebral Ischemia) Trial; the approval was granted through the 510(k) process. The Merci Retriever system includes a flexible nickel titanium (nitinol) wire that obtains a helical shape once it is passed through the tip of the guidance catheter. In practice, the catheter/wire is passed distal to the thrombus, the catheter is removed, and the helical configuration assumed by the wire; the clot is then trapped in the helix and withdrawn from the vasculature (Figure 2). The 510(k) clearance means that the Merci Retriever was felt to be substantially equivalent to a predicate device. In this case, the predicate device was the Concentric Retriever, which itself received 510(k) clearance by the FDA in May 2001 for “use in …

A novel transient middle cerebral artery (MCA) occlusion model in rats was used to evaluate the effects of nimodipine on 45Ca accumulation, brain oedema and mortality. Nimodipine (1 microgram/kg/min, IV, 30 min) administered immediately after 3 hr of transient unilateral MCA occlusion significantly attenuated the post-ischaemic increase of tissue water content, partly 45Ca accumulation in the parietotemporal and frontal cortices ipsilateral to the left MCA occlusion 3 hr after reperfusion. Nimodipine decreased the mortality rate at 6 and 9 hr time points after recirculation, although the survival rate at the 24 hr point after recirculation was not different from the control group. These results suggest that nimodipine has beneficial effects in the early phase of reperfusion period.

Effect of Nimodipine on Ischemia-Induced Brain Edema and Mortality in a Novel Transient Middle Cerebral Artery Occlusion Model

To the Editor: Brey et al1 reported what is to their knowledge the first study to demonstrate a prospective association between sera cofactor-dependent anticardiolipin antibodies and stroke independent of other risk factors as well as myocardial infarction (MI). In addition to lending support to basic research that has shown the pathogenicity of antiphospholipid-protein antibodies (aPL) in thrombosis,2 this well-conducted epidemiological study of Japanese-American men enrolled in the Honolulu Heart Program and followed for up to 20 years provides evidence for the role of aPL as potentially important markers and/or causes of increased vascular risk associated with ischemic stroke and MI. It is known that stroke is a multisystemic disorder involving mechanisms of thrombotic and neurotoxic coupling.3 Biochemical markers including glutamate, homocysteine (a sulfinic analog of aspartate4), and N -methyl-d-aspartate (NMDA) receptor autoantibodies (aAb) are independently associated with neurotoxicity and can be measured in blood.5 The aPLs are a part of the structural components of …

Ischemic stroke (IS) is a major cause of mortality and disability worldwide. However, the pathogenesis of IS remains unknown, and methods for early prediction and diagnosis of IS are lacking. Metabolomics can be applied to biomarker discovery and mechanism exploration of IS by exploring metabolic alterations. In this review, 62 IS metabolomics studies in the murine model published from January 2006 to December 2020 in the PubMed and Web of Science databases were systematically reviewed. Twenty metabolites (e.g., lysine, phenylalanine, methionine, tryptophan, leucine, lactate, serine, N-acetyl-aspartic acid, and glutathione) were reported consistently in more than two-third murine studies. The disturbance of metabolic pathways, such as arginine biosynthesis; alanine, aspartate and glutamate metabolism; aminoacyl-tRNA biosynthesis; and citrate cycle, may be implicated in the development of IS by influencing the biological processes such as energy failure, oxidative stress, apoptosis, and glutamate toxicity. The transient middle cerebral artery occlusion model and permanent middle cerebral artery occlusion model exhibit both common and distinct metabolic patterns. Furthermore, five metabolites (proline, serine, LysoPC (16:0), uric acid, glutamate) in the blood sample and 7 metabolic pathways (e.g., alanine, aspartate, and glutamate metabolism) are shared in animal and clinical studies. The potential biomarkers and related pathways of IS in the murine model may facilitate the biomarker discovery for early diagnosis of IS and the development of novel therapeutic targets.

Intracranial bleeding is the most feared complication of thrombolytic therapy in acute stroke. The risk of brain hemorrhage is the main argument of the European authorities not to approve recombinant tissue plasminogen activator (rtPA), and the fear of hurting patients with rtPA explains its limited use in North America. The common argument is, “Treatment with rt-PA may have some beneficial effect, but that is traded off by a considerable risk of symptomatic hemorrhage.” This argument is false and based on misunderstanding and misconception. The misunderstanding: There is no such trade-off. The National Institute of Neurological Disorders and Stroke (NINDS) rtPA Stroke Study Group observed 2 patients (0.6%) with symptomatic and 1 patient (0.3%) with fatal hemorrhages in the placebo group (n=312) and 20 patients (6.4%) with symptomatic and 9 patients (2.9%) with fatal hemorrhages in the rtPA group (n=312).1 Despite this supposed excess in risks caused by rtPA treatment (odds ratios [OR], 10.6 and 9.2), rtPA treatment significantly reduced the risk for disability and death (modified Rankin Scale >1 at 12 months after stroke) from 73% to 59% (reduction for death alone: 28% to 24%).2 In both European Cooperative Acute Stroke Studies (ECASS) 1 and 2, rtPA increased the risk for parenchymal hematomas (OR, 3.0 and 4.2), but reduced the overall risk for disability and death by 6% and 8% (NS).3,4⇓ A similar observation—an overall risk reduction for disability and death despite an increased risk for intracranial hemorrhages—was made in the Multicenter Acute Stroke Trials (MAST) -Europe and -Italy. Why …

Background and Purpose: We previously reported the neuroprotective effects of trans-arterial regional hypothermia in rat transient middle cerebral artery occlusion (tMCAO) model (ISC 2014, ISC 2015). The tMCAO model is representative of the clinical setting of successful recanalization in acute ischemic stroke. However, it is not always possible to achieve successful recanalization. Herein, we investigated the neuroprotective effects of trans-arterial regional hypothermia in permanent MCAO (pMCAO) model as well. Methods: Three groups were provided to investigate the neuroprotective effects. Regional hypothermia group had a single infusion of 20ml/kg 10°C cold saline via the ipsilateral internal carotid artery for 15 min at 0 hr or 1 hr after the onset of pMCAO. Vehicle group had a 37°C warm saline infusion in the same manner. Control group had no infusion after pMCAO. The rats were tested for neurological score after 48-hour pMCAO and then sacrificed to evaluate infarction size and pathological condition. Immunohistochemical analysis was performed to investigate the neuronal apoptosis (NeuN, cleaved caspase 3), glial reaction (GFAP), and microglial activation (Iba-1). Results: The regional hypothermia group showed significantly better neurological score compared with the other groups (p<0.01). Infarct volume was significantly smaller in the regional hypothermia group (8.1±4.7% at 0 hr and 15.7±5.9% at 1 hr) than in the vehicle group (35.8±5.6% at 0 hr and 34.0±3.6% at 1 hr) and also in the control group (35.1±8.9%) (p<0.01). Neuronal apoptosis (p<0.01), reactive gliosis (p<0.01), and microglial activation (p<0.05) in the peri-infarct area were significantly inhibited in the regional hypothermia group compared with the other groups. The degree of the inhibition was somewhat more evident at 0 hr than at 1 hr. Conclusions: Our result demonstrated that trans-arterial regional hypothermia provided robust neuroprotection in pMCAO model as well as tMCAO model, suggesting that this therapy may have a wide clinical application in ischemic stroke treatment.

Transplantation of neural stem cells provides a promising therapy for stroke. Its efficacy, however, might be limited because of massive grafted-cell death after transplantation, and its insufficient capability for tissue repair. Interleukin 6 is a pro-inflammatory cytokine involved in the pathogenesis of various neurological disorders. Paradoxically, interleukin 6 promotes a pro-survival signalling pathway through activation of signal transducer and activator of transcription 3. In this study, we investigated whether cellular reprogramming of neural stem cells with interleukin 6 facilitates the effectiveness of cell transplantation therapy in ischaemic stroke. Neural stem cells harvested from the subventricular zone of foetal mice were preconditioned with interleukin 6 in vitro and transplanted into mouse brains 6 h or 7 days after transient middle cerebral artery occlusion. Interleukin 6 preconditioning protected the grafted neural stem cells from ischaemic reperfusion injury through signal transducer and activator of transcription 3-mediated upregulation of manganese superoxide dismutase, a primary mitochondrial antioxidant enzyme. In addition, interleukin 6 preconditioning induced secretion of vascular endothelial growth factor from the neural stem cells through activation of signal transducer and activator of transcription 3, resulting in promotion of angiogenesis in the ischaemic brain. Furthermore, transplantation of interleukin 6-preconditioned neural stem cells significantly attenuated infarct size and improved neurological performance compared with non-preconditioned neural stem cells. This interleukin 6-induced amelioration of ischaemic insults was abolished by transfecting the neural stem cells with signal transducer and activator of transcription 3 small interfering RNA before transplantation. These results indicate that preconditioning with interleukin 6, which reprograms neural stem cells to tolerate oxidative stress after ischaemic reperfusion injury and to induce angiogenesis through activation of signal transducer and activator of transcription 3, is a simple and beneficial approach for enhancing the effectiveness of cell transplantation therapy in ischaemic stroke.

Knockdown of Smox protects the integrity of the blood-brain barrier through antioxidant effect and Nrf2 pathway activation in stroke

The Common Racially Diverse Human PAR4 Variant Thr120 Causes Worse Outcomes in a Mouse Stroke Model Via Platelet-Mediated NET Formation: Prevention By P-Selectin Blockade

MP-124, a novel poly(ADP-ribose) polymerase-1 (PARP-1) inhibitor, ameliorates ischemic brain damage in a non-human primate model