Investigational agents for treatment of traumatic brain injury.

INTRODUCTION: Traumatic brain injury (TBI) is a major cause of death and disability worldwide. To date, there are no pharmacologic agents proven to improve outcomes from TBI because all the Phase III clinical trials in TBI have failed. Thus, there is a compelling need to develop treatments for TBI. AREAS COVERED: The following article provides an overview of select cell-based and pharmacological therapies under early development for the treatment of TBI. These therapies seek to enhance cognitive and neurological functional recovery through neuroprotective and neurorestorative strategies. EXPERT OPINION: TBI elicits both complex degenerative and regenerative tissue responses in the brain. TBI can lead to cognitive, behavioral, and motor deficits. Although numerous promising neuroprotective treatment options have emerged from preclinical studies that mainly target the lesion, translation of preclinical effective neuroprotective drugs to clinical trials has proven challenging. Accumulating evidence indicates that the mammalian brain has a significant, albeit limited, capacity for both structural and functional plasticity, as well as regeneration essential for spontaneous functional recovery after injury. A new therapeutic approach is to stimulate neurovascular remodeling by enhancing angiogenesis, neurogenesis, oligodendrogenesis, and axonal sprouting, which in concert, may improve neurological functional recovery after TBI.

Targeting nitric oxide in the subacute restorative treatment of ischemic stroke.

INTRODUCTION: Stroke remains the leading cause of adult disability. Thus, it is imperative to develop restorative therapies for ischemic stroke designed specifically to treat the intact brain tissue to stimulate functional benefit. Therapies targeting amplification of brain repair processes with nitric oxide (NO) donors and phosphodiesterase type 5 (PDE5) inhibitors in preclinical studies are emerging and showing improvement of functional recovery after stroke. AREAS COVERED: This review will mainly cover the effect of NO donors, which produce NO, and PDE5 inhibitors, which elevate cyclic guanosine 3',5'-monophosphate (cGMP), on neural restorative events in ischemic brain and highlight mechanisms underlying their restorative therapeutic activity. EXPERT OPINION: During stroke recovery, interwoven restorative events occur in ischemic brain, which include angiogenesis, neurogenesis, oligodendrogenesis, astrogliosis and neurite outgrowth. Emerging preclinical data indicate that restorative therapies targeting multiple parenchymal cells including neural stem cells, cerebral endothelial cells, astrocytes, oligodendrocytes, neurons would be more effective than agents with a single cell target. Preclinical data suggest that elevated cGMP levels induced by NO donors and PDE5 inhibitors act on cerebral endothelial cells, neural stem cells and oligodendrocyte progenitor cells to enhance stroke-induced angiogenesis, neurogenesis and oligodendrogenesis, respectively. These interacting remodeling events collectively improve neurological function after stroke.

Multimodal neuroprotection induced by PACAP38 in oxygen-glucose deprivation and middle cerebral artery occlusion stroke models.

Pituitary adenylate cyclase activating peptide (PACAP), a potent neuropeptide which crosses the blood-brain barrier, is known to provide neuroprotection in rat stroke models of middle cerebral artery occlusion (MCAO) by mechanism(s) which deserve clarification. We confirmed that following i.v. injection of 30 ng/kg of PACAP38 in rats exposed to 2 h of MCAO focal cerebral ischemia and 48 h reoxygenation, 50 % neuroprotection was measured by reduced caspase-3 activity and volume of cerebral infarction. Similar neuroprotective effects were measured upon PACAP38 treatment of oxygen-glucose deprivation and reoxygenation of brain cortical neurons. The neuroprotection was temporally associated with increased expression of brain-derived neurotrophic factor, phosphorylation of its receptor-tropomyosin-related kinase receptor type B (trkB), activation of phosphoinositide 3-kinase and Akt, and reduction of extracellular signal-regulated kinases 1/2 phosphorylation. PACAP38 increased expression of neuronal markers beta-tubulin III, microtubule-associated protein-2, and growth-associated protein-43. PACAP38 induced stimulation of Rac and suppression of Rho GTPase activities. PACAP38 downregulated the nerve growth factor receptor (p75(NTR)) and associated Nogo-(Neurite outgrowth-A) receptor. Collectively, these in vitro and in vivo results propose that PACAP exhibits neuroprotective effects in cerebral ischemia by three mechanisms: a direct one, mediated by PACAP receptors, and two indirect, induced by neurotrophin release, activation of the trkB receptors and attenuation of neuronal growth inhibitory signaling molecules p75(NTR) and Nogo receptor.

Who's in favor of translational cell therapy for stroke: STEPS forward please?

A consortium of translational stem cell and stroke experts from multiple academic institutes and biotechnology companies, under the guidance of the government (FDA/NIH), is missing. Here, we build a case for the establishment of this consortium if cell therapy for stroke is to advance from the laboratory to the clinic.

Atorvastatin extends the therapeutic window for tPA to 6 h after the onset of embolic stroke in rats.

We investigated the neuroprotective effect of atorvastatin in combination with delayed thrombolytic therapy in a rat model of embolic stroke. Rats subjected to embolic middle cerebral artery (MCA) occlusion were treated with atorvastatin at 4 h, followed by tissue plasminogen activator (tPA) at 6 or 8 h after stroke. The combination of atorvastatin at 4 h and tPA at 6 h significantly decreased the size of the embolus at the origin of the MCA, improved microvascular patency, and reduced infarct volume, but did not increase the incidence of hemorrhagic transformation compared with vehicle-treated control animals. However, monotherapy with tPA at 6 h increased the incidence of hemorrhagic transformation and failed to reduce infarct volume compared with the control group. In addition, adjuvant treatment with atorvastatin at 4 h and with tPA at 6 h reduced tPA-induced upregulation of protease-activated receptor-1, intercellular adhesion molecule-1, and matrix metalloproteinase-9, and concomitantly reduced cerebral microvascular platelet, neutrophil, and fibrin deposition compared with rats treated with tPA alone at 6 h. In conclusion, a combination of atorvastatin and tPA extended the therapeutic window for stroke to 6 h without increasing the incidence of hemorrhagic transformation. Atorvastatin blocked delayed tPA-potentiated adverse cerebral vascular events, which likely contributes to the neuroprotective effect of the combination therapy.

Emerging treatments for traumatic brain injury.

BACKGROUND: This review summarizes promising approaches for the treatment of traumatic brain injury (TBI) that are in either preclinical or clinical trials. OBJECTIVE: The pathophysiology underlying neurological deficits after TBI is described. An overview of select therapies for TBI with neuroprotective and neurorestorative effects is presented. METHODS: A literature review of preclinical TBI studies and clinical TBI trials related to neuroprotective and neurorestorative therapeutic approaches is provided. RESULTS/CONCLUSION: Nearly all Phase II/III clinical trials in neuroprotection have failed to show any consistent improvement in outcome for TBI patients. The next decade will witness an increasing number of clinical trials that seek to translate preclinical research discoveries to the clinic. Promising drug- or cell-based therapeutic approaches include erythropoietin and its carbamylated form, statins, bone marrow stromal cells, stem cells singularly or in combination or with biomaterials to reduce brain injury via neuroprotection and promote brain remodeling via angiogenesis, neurogenesis, and synaptogenesis with a final goal to improve functional outcome of TBI patients. In addition, enriched environment and voluntary physical exercise show promise in promoting functional outcome after TBI, and should be evaluated alone or in combination with other treatments as therapeutic approaches for TBI.

Intracellular free calcium mediates glioma cell detachment and cytotoxicity after photodynamic therapy.

Photofrin photodynamic therapy (PDT) caused a dose-dependent decrease of enzymatic cell detachment by trypsin/ethylenediamine tetra-acetic acid (EDTA) in human glioma U251n and U87 cells. This happened coincidently with the increase of intracellular free calcium ([Ca(2+)](i)). Thapsigargin, which increased [Ca(2+)](i), induced further decrease in enzymatic cell detachment and increased cytotoxicity. Opposite effects were observed when 1,2-bis(2-aminophenoxy) ethane-N,N,N',N'-tetra-acetic acid tetrakis, an intracellular Ca(2+) chelator, was used. PDT-induced changes in [Ca(2+)](i) and cell detachment were not blocked by calcium channel antagonists nickel (Ni(2+)) or nimodipine, nor were they altered when cells were irradiated in a buffer free from Ca(2+) and magnesium (Mg(2+)), suggesting that [Ca(2+)](i) is derived from the internal calcium stores. Decreased cell migration was observed after PDT, as assessed by chemotactic and wound-healing assays. Our findings indicated that internal calcium store-derived [Ca(2+)](i) plays an important role in PDT-induced enzymatic cell detachment decrease and cytotoxicity. Cell migration may be affected by these changes.

Contralesional axonal remodeling of the corticospinal system in adult rats after stroke and bone marrow stromal cell treatment.

BACKGROUND AND PURPOSE: Motor recovery after stroke is associated with neuronal reorganization in bilateral hemispheres. We investigated contralesional corticospinal tract remodeling in the brain and spinal cord in rats after stroke and treatment of bone marrow stromal cells. METHODS: Adult male Wistar rats were subjected to permanent right middle cerebral artery occlusion. Phosphate-buffered saline or bone marrow stromal cells were injected into a tail vein 1 day postischemia. An adhesive removal test was performed weekly to monitor functional recovery. Threshold currents of intracortical microstimulation on the left motor cortex for evoking bilateral forelimb movements were measured 6 weeks after stroke. When intracortical microstimulation was completed, biotinylated dextran amine was injected into the left motor cortex to anterogradely label the corticospinal tract. At 4 days before euthanization, pseudorabies virus-152-EGFP and 614-mRFP were injected into left or right forelimb extensor muscles, respectively. All animals were euthanized 8 weeks after stroke. RESULTS: In normal rats (n=5), the corticospinal tract showed a unilateral innervation pattern. In middle cerebral artery occlusion rats (n=8), our data demonstrated that: 1) stroke reduced the stimulation threshold evoking ipsilateral forelimb movement; 2) EGFP-positive pyramidal neurons were increased in the left intact cortex, which were labeled from the left stroke-impaired forelimb; and 3) biotinylated dextran amine-labeled contralesional axons sprouted into the denervated spinal cord. Bone marrow stromal cells significantly enhanced all 3 responses (n=8, P<0.05). CONCLUSIONS: Our data demonstrated that corticospinal tract fibers originating from the contralesional motor cortex sprout into the denervated spinal cord after stroke and bone marrow stromal cells treatment, which may contribute to functional recovery.

Low-level laser therapy applied transcranially to rats after induction of stroke significantly reduces long-term neurological deficits.

BACKGROUND AND PURPOSE: Low-level laser therapy (LLLT) modulates various biological processes. In the present study, we assessed the hypothesis that LLLT after induction of stroke may have a beneficial effect on ischemic brain tissue. METHODS: Two sets of experiments were performed. Stroke was induced in rats by (1) permanent occlusion of the middle cerebral artery through a craniotomy or (2) insertion of a filament. After induction of stroke, a battery of neurological and functional tests (neurological score, adhesive removal) was performed. Four and 24 hours poststroke, a Ga-As diode laser was used transcranially to illuminate the hemisphere contralateral to the stroke at a power density of 7.5 mW/cm2. RESULTS: In both models of stroke, LLLT significantly reduced neurological deficits when applied 24 hours poststroke. Application of the laser at 4 hours poststroke did not affect the neurological outcome of the stroke-induced rats as compared with controls. There was no statistically significant difference in the stroke lesion area between control and laser-irradiated rats. The number of newly formed neuronal cells, assessed by double immunoreactivity to bromodeoxyuridine and tubulin isotype III as well as migrating cells (doublecortin immunoactivity), was significantly elevated in the subventricular zone of the hemisphere ipsilateral to the induction of stroke when treated by LLLT. CONCLUSIONS: Our data suggest that a noninvasive intervention of LLLT issued 24 hours after acute stroke may provide a significant functional benefit with an underlying mechanism possibly being induction of neurogenesis.

Recovery in a patient with locked-in syndrome.

BACKGROUND AND PURPOSE: Sildenafil citrate has been shown to enhance neurogenesis, angiogenesis, synaptogenesis, and neurological outcome by augmentation of cyclic guanosine monophosphate (cGMP) levels in animal models of ischemic stroke. Whether sildenafil citrate may be helpful for recovery in human stroke is unknown at this time. METHODS: A 41-year-old woman with locked-in syndrome due to pontine infarction began receiving 150 mg of oral sildenafil citrate daily on a compassionate use basis in August 2003 and continues treatment at this time. Magneto-encephalography (MEG) was performed at 12 and 17 months after stroke. RESULTS: No serious adverse events have occurred. Significant milestone recoveries including standing, use of both arms, talking, and full return of swallowing have occurred, particularly after nine months of treatment. The MEG showed a significantly increased amplitude in the somatosensory cortex. CONCLUSION: Daily use of high dose sildenafil citrate appears to be safe in this patient with stroke resulting in locked-in syndrome. Further studies will be required to establish safety and efficacy.

Local fluorouracil chemotherapy interferes with neural and behavioral recovery after brain tumor-like mass compression.

In this study, we investigated the impact of intracerebral delivery of chemotherapy on functional recovery from focal cortical tissue displacement, characteristic of brain tumors. Unilateral focal brain compression was induced by epidural implantation of an inverted hemisphere-shaped bead over the sensorimotor cortex. Microinjections of a total of 1mg chemoagent fluorouracil or the same volume of saline were made into the compressed cortex. Behavioral tests of forelimb sensorimotor function were conducted during 4 weeks' observation. Rats subjected to any of the three types of lesions, saline microinjection plus cortical compression, chemoagent microinjection alone, or chemoagent microinjection combined with cortical compression, demonstrated significant behavioral deficits in several sensorimotor tasks, compared with saline-microinjected control animals. In placing tests, behavioral deficits elicited by each single treatment were worsened by combined treatment with chemoagent microinjection and focal cortical compression. Concurrently, local delivery of chemoagent into the compressed cortex induced increased cortical tissue loss, necrosis and apoptosis. These data indicate that local chemotherapy exacerbates compression-induced neurological impairment, and a model of controlled focal cortical compression may provide a valuable means to improve anti-cancer therapeutic designs with reduced deterioration of brain function.

Interruption of functional recovery by the NMDA glutamate antagonist MK801 after compression of the sensorimotor cortex: implications for treatment of tumors or other mass-related brain injuries.

Glutamate antagonists have recently been shown to limit tumor growth, providing potential new therapeutic targets and strategies against brain tumors. Here, we demonstrate that the glutamate NMDA receptor antagonist MK801, after a delay, adversely reverses functional recovery in rats with compressive mass lesions of the sensorimotor cortex. Our data suggest that the controlled focal cortical compression model may be a valuable pre-clinical tool to screen compounds for the treatment of brain tumors. It may be possible to use this model to develop interventions that maintain anti-cancer effects but with diminished harm to bystander tissue and brain plasticity.

Assessing combination treatments in acute stroke: preclinical experiences.

BACKGROUND: Acute ischemic stroke is a complex disease. Treatment success may require combining different therapeutic approaches. An obvious treatment combination in acute ischemic stroke is a thrombolytic therapy, adjuvant with a neuroprotective agent to have better stroke recovery. SUMMARY OF REVIEW: Complete factorial designs can assess the synergy of combination treatments and distinguish them from supper-additive, additive or sub-additive effects. A factorial design, a two-way analysis of variance and a novel graphic technique can detect and illustrate interactions of two treatments, which were used to evaluate combination treatments to extend the therapeutic window for thrombolytic therapy in an embolic stroke model on rats. We hypothesized synergy or additive effects on stroke recovery when combining thrombolytic therapy with either an antagonist to the integrin CD11b/CD18 (UK-279,276) or a glycoprotein IIb/IIIa receptor inhibitor. CONCLUSIONS: Factorial designs offer an efficient approach to study synergistic effects of two treatments. Our graphical technique provides a powerful, intuitive and quantitative explanation of joint treatment effects. Combining thrombolytic therapy with a neuroprotectant yielded a super-additive or additive treatment effect for two preclinical experiments designed to extend the thrombolytic therapeutic window for stroke.

Inhibition of factor Xa reduces ischemic brain damage after thromboembolic stroke in rats.

BACKGROUND AND PURPOSE: Factor Xa (FXa) is a key coagulation protease and target for novel antithrombotic agents for prevention and treatment of diverse thromboembolic disorders. In the present study we describe the effect of a novel, potent, and selective FXa inhibitor, DPC602, on brain damage and neurobehavioral consequence in a rat thromboembolic model of stroke. METHODS: Thromboembolic stroke was induced in rats by placement of an autologous clot into the middle cerebral artery. RESULTS: Laser-Doppler monitoring of cerebral blood flow demonstrated that DPC602 (8 mg/kg, single IV/IP bolus pretreatment) markedly improved cerebral blood flow after thromboembolic stroke by 25% to 160% (n=6; P<0.001) at 1 to 6 hours. DPC602 demonstrated concentration- and time-dependent reductions in infarct size, with maximal effect (89% reduction; n=14; P<0.001) at the highest dose over controls. Neurological function was also significantly improved in DPC602-treated rats at days 1, 3, and 7 (n=13; P<0.01). DPC602 treatment did not cause cerebral hemorrhage, assessed by free hemoglobin in the ischemic brain tissues. CONCLUSIONS: These data suggest that anticoagulation with a selective FXa inhibitor might ameliorate the extent of ischemic brain damage and neurological deficits after a thromboembolic event. Enhanced clot dissolution and early reperfusion may account for the cerebrovascular-protective effect of the drug.