Poster Abstracts 1 to 10
ASENT 5th Annual Meeting
Thursday, March 13 - Saturday, March 15, 2003
Capital Hilton Hotel, Washington, DC
Poster Abstracts
1
SALSOLINOL INDUCES THE SYNTHESIS OF METALLOTHIONIEN ISOFORMS, WHICH PROVIDE NEUROPROTECTIVE ACTIONS IN DOPAMINERGIC NEURONS.
S. Wanpen, S. Shavali and M. Ebadi
Department of Pharmacology, University of North Dakota
School of Medicine and Health Sciences, Grand Forks, ND, 58201
The factors that contribute to the progression of Parkinson’s disease are a defect in mitochondrial respiration and the generation of excess hydrogen peroxide, which in the presence of a deficiency of striatal gluthione becomes converted to highly neurotoxic hydroxyl radicals, which are prevented from forming by selegiline and once formed are scavenged by metallothionein (MT). We have studied the neurotoxic actions of 1-Benzyk-tetrahydroisoquinoline (1-BnTIQ) in dopaminergic SH-SY5Y cells and found that the induction of MT or administration of Coenzyme Q10 attenuated the 1-BnTIQ induced DNA condensation/fragmentation. Furthermore, by employing ESR, we examined the free radicals scavenging effects of zinc MT isoforms I and II on four types of free radicals. Solution of 0.15 mM of MT-1 and 0.3 mM of MT-II were able to scavenge hydroxyl radicals generated in iron-mediated Fenton reactions. Moreover, MT-1 scavenged almost 90% of the superoxide generated by the hypoxanthine and xanthine oxidase system, while MT-II could only scavenge 40%. In addition, while zinc MT protected against damaging effects of hydroxyl radicals, copper MT produced hydroxyl radicals and hence possessed prooxidant properties. The molecular mechanism of salsolinol induced apoptosis has also been studied. Salsolinol induces DNA condensation/fragmentation; and causes activation of JNK/SPAK regulated down stream transcriptional activities via phosphorylation of c-Jun. The induction of metallothionein represents the protective response against the cytotoxic effect of the neurotoxin. The elevated expression of calbindin-D28K, zinc, MT, glutathione peroxidase, and Bc1-2 gene in experimental models of Parkinsonism may reflect the activation of several neuroprotective mechanisms in order to counter balance the excess generation of reactive oxygen species in the striatum of patients with Parkinson’s disease. (Supported in part by a grant from USPHS 2R01 NS 34566-09).
2
A NOVEL TREATMENT FOR PARKINSON’S DISEASE USING AN IMMUNOMODULATOR
B. Sredni*, R. Geffen*, D. Longo^, F. Shalit*, M.P. Mattson^ and Yadid G.*
*Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan, Israel.
^National Institute on Aging, Gerontology Research Center, Baltimore, MD 21224, USA.
Degeneration of the nigrostriatal pathway is characteristic for Parkinson’s disease (PD). Current treatments are aimed at pharmacologically augmenting striatal dopamine but do not prevent continued neuron degeneration. A protective/restorative treatment that would slow, prevent or even reverse the degenerative process is therefore needed. Herein we tested whether a mediator of cytokine production can be sued to effectively treat PD in an animal model. We investigated the effect of the immunomodulator AS101 on an astrocytes cell line (SVG). Our results show that AS101 (0.1-1 µg/ml) stimulates the production and secretion of GDNF and IL-6 in this cell in a dose and a time dependent manner. In order to test the in vivo effect of AS101, the compound (2µg/10µ1/10min) was injected directly into the substantia nigra (SN) or striatum of male Sprague-Dawley rats, resulting in two-fold increase of IL-6, IL-1 and GDNF mRNA levels. Administration of AS101 (15µg for 3 days at 0.2µg/µ1/hr) into the diseased SN of partially 6-OHDA lesion rats, reduced apomorphine-induced rotation by 90% and improved spontaneous behavior. Dopamine (DA) and DA metabolites as well as IL-6 levels were higher in the AS101 treated rats compared to PBS treated rats in both the SN and striatum, as determined by HPLC analysis. These results were supported by increased immunohistochemically staining of tyrosine hydroxylase. Our findings suggest a novel treatment that may provide a dopaminergic supportive environment in the diseased SN.
3
GLOBAL STATISTICAL TESTS FOR PARKINSON’S DISEASE CLINICAL TRIALS
Huang, P., Tilley, B.C., Palesch, Y., Jaskwhich, J.D., Bergmann, K., Guimaraes, P.
Medical University of South Carolina, Charleston, SC 29425
Abstract. Comparing treatment efficacy in Parkinson’s disease studies often involves multiple correlation outcomes to obtain an objective assessment due to the lack of known biomarkers for the disease. Several types of statistical tests have been proposed in the literature for treatment comparisons with multiple outcomes. However, which test is appropriate depends on how we define a beneficial treatment. The advantages and disadvantages of these tests and the situations where these tests are appropriate to use are discussed here. We then focus on the case when treatment efficacy requires that the majority of outcomes have shown improved results. We review the principles of available nonparametric global statistical tests and illustrate their applications using data from previously conducted trials on Parkinson’s disease.
Objective. To compare Parkinson’s disease treatment when treatment efficacy requires evidence of treatment benefit from a majority of correlated outcomes.
Background. Clinical trials for Parkinson’s disease typically have multiple outcomes. However, most trials are designed based on a single primary outcome, such as some weighted linear combination of several outcomes. When no single outcome is accepted as a gold standard for treatment comparison, results based on a single outcome often leave findings from other outcomes unclear. On the other hand, in order to examine several outcomes simultaneously, one often makes sample size unnecessarily large in order to preserve the overall statistical significant level. Global statistics test (GST’s) can consider all outcomes simultaneously with a prespecified significant level and combine evidence from each single outcome to yield an assessment for the overall treatment difference. This provides a more objective conclusion about treatment efficacy than Bonferroni adjustment and Hotelling’s T2 test when treatment efficacy requires improvement over majority of outcomes.
Methods. We define a parameter that measures the overall treatment effect based on multiple correlated outcomes. Subject’s total ranks from different outcomes are used in nonparametric GST. The advantage of using GST is shown through the comparisons of power and length of confidence interval between GST and univariate tests.
Conclusion. GST’s are more powerful tests for comparing treatment effects with multiple correlated outcomes, especially the preference of new treatment requires evidence from several outcome measures. GST’s should be considered for use in Parkinson’s disease clinical trials.
4
DETERMINANTS AND CONSEQUENCES OF DOSE CHANGES IN A FLEXIBLE DOSE STUDY
Lipkovich, I., Ahmed, S., Ahl, J., Kinon, B., Rotelli, M.,
Eli Lilly and Company, Indianapolis, IN 46825
Flexible dose studies allow the does to be adjusted during the course of the treatment and contain useful information on the condition of patients preceding and following dose changes. In the present analysis we identify factors that predict dose changes during a flexible dose study, and determine whether dose changes predict changes in efficacy and side effects. Data from a 12-week, double-blind, flexible-dose study ion patients with acute mania randomized to receive olanzapine (OLZ) (5-20 mg/d, N=234) or haloperidol (HAL) (3-15 mg/d, N=219) were analyzed post hoc. Associations between dose changes and efficacy, extrapyramidal symptoms (EPS), weight changes (as measured by changes in Body Mass Index), and unsolicited EPS related adverse events (EPSAEs) were modeled using repeated measures analyses. For both therapies, baseline illness severity as well as level of efficacy achieved, EPS scales, and severity of EPSAEs experienced during the previous week predicted dose changes. Patients whose dose was decreased showed less improvement on subsequent visits (p=.02 for OLZ and p=0.002 for HAL). The direction of change in EPS scales followed the direction of change in dose of HLA, but these changes were not predicted by dose change. Efficacy and tolerability are predictive of antipsychotic dose adjustments. For both treatments dose changes affect efficacy. For HAL, EPS were influenced by change in dose. For OLZ, overall EPS rates were lower and were not affected by dose changes, which may allow for more aggressive dosing to increase therapeutic benefit.
Funding provided by Eli Lilly and Company.
5
TREATMENT-EMERGENT EXTRAPYRAMIDAL SYMPTOMS DURING TREATMENT WITH OLANZAPINE OR RISPERIDONE
Lipkovich, I., Kollack-Walker, S., Ahmed, S.
Eli Lilly and Company, Indianapolis, IN 46285.
This study examines the dynamic expression of extrapyramidal symptoms (EPS) in patients treated with olanzapine (OLZ) or risperidone (RIS). Data from a double-blind, parallel group, controlled 28-week study in psychotic patients comparing OLZ (10-20 mg/d; n=172) and RIS (4-12 mg/d, n=167) were analyzed posthoc using longitudinal repeated measures analysis. EPS were evaluated at baseline and during 28-week treatment period using standard measurement scales and spontaneously reported adverse events. Data were analyzed for all patients, and subgroups of patients with and without EPS at baseline. RIS patients were divided into > of < 6 mg/d modal dose subgroups. Longitudinal analysis revealed greater reduction in Barnes Global (p=.023) and Simpson-Angus (p<.001) scores following treatment with OLZ compared to RIS. Patients with preexisting EPS at baseline (>0 score) showed less improvement in Barnes Global (p=.026) and Simpson-Angus (p=.007) scores with RIS. Patients with no EPS at baseline showed greater increases in Simpson-Angus score (p=.020) during treatment with RIS. Improvement in Simpson-Angus score was greater with OLZ compared to RIS at either low or high modal dose. Olanzapine treatment led to significantly greater improvement in EPS compared to risperidone. Furthermore, olanzapine had a lower liability for treatment-induced parkinsonian symptoms.
Funding provided by Eli Lilly and Company.
6
PHASE III TRIAL OF EBSELEN
Oeda, J., Kidoguchi, E., Masayasu, H., Yamaguchi, T.,
Department of Medical Planning & Coordination, Daiichi Pharmaceutical Co., Ltd., Tokyo, 134-8630, Japan.
National Cardiovascular Center, Osaka, 565-0873, Japan.
a) Ebselen is a seleno-organic compound which exhibits antioxidative activity through intracellular redox regulation. Ebselen is not merely a free radical scavenger as it is a mimic of enzyme glutathione peroxidase, and has recently been found to be an excellent substrate for thioredoxin reductase, and acted as thioredoxin reductase peroxidase (Zhao R et al, PNAS 2002:99:8579-8584). A previous clinical study in 300 acute patients (Yamaguchi et al. Stroke 1998:29:12-17) treated within 48 hours of stroke demonstrated that overall ebselen treatment achieved a significantly better outcome (Glasgow Outcome Scale, GOS) than placebo at 1 month (P=.023), but not at 3 months (P=.056), although the improvement of the outcome was observed in patients with sever or moderate impairment. Neurological status was assessed by the Modified Mathew scale; a score from 0 to 34, 35 to 74, and 75 to 100 was considered to indicate severe, moderate and mild impairment, respectively. A score of 100 indicates the absence of impairment. The outcome of patients who received early ebselen treatment within 24 hours of onset was significantly superior to that of patients treated after 24 hours. In addition, the effect of ebselen on outcome was more prominent in patients with lesions involving the cerebral cortex than in those with deep-brain lesions. In order to confirm the reproducibility of the neuroprotective effect of this compound, a new study was designed in patients with acute ischemic stroke with cortical involvement. b) Those patients with disturbed consciousness, demonstrable cortical signs, CT/MRI lesions in the cortical branch territory or cortical border zone, or severe stenosis/occlusion of main cerebral arteries were entered into the study. Patients with lacunar, infarction, transient ischemic attack, or intracranial hemorrhage were excluded. Treatment was started within 24 hours of onset. The concomitant use of drugs was not prohibited except thrombolytics. The percentage of stroke patients with “good recovery” according to the GOS assessment (primary endpoint) will be compared between the Ebselen and placebo groups. c) The study, started in March 2001, is currently ongoing. Patient enrollment completed in September 2002 with 394 patients. d) The blind will be broken Q2 of 2003.
7
A NEW MOLECULAR TRIGGER FOR NERVE REGENERATION
Palerme, J.S., Rongy-Mimouni, M., Landau, A.M., Desbarats, J.
Department of Physiology, McGill University, Montreal, QC, H3G 1Y6, Canada.
Fas (CD95) is a member of the tumor necrosis factor receptor (TNF-R) superfamily, best known for its role as a “death receptor.” Fas can trigger neuron apoptosis in animal models of stroke in vivo, and in motoneurons in vitro. However, we have previously shown the Fas can also promote growth, proliferation, and differentiation in a variety of cell types. Here we show that Fas engagement on sensory neurons can induce a dramatic regenerative response, resulting in prolific axon growth in vitro and accelerated recovery after nerve damage in vivo (Nature Cell Biol. 2003, 5:118-115). We demonstrate that Fas-mediated axon regeneration is independent of the Fas apoptotic pathway, but requires activation of the Extracellular-signal Regulated Kinase (ERK) pathway. Furthermore, our data indicate that Fas engagement can promote neuron survival and preserve neuritogenic potential, as well as stimulate new axon growth. Fas engagement triggered axon regeneration that was morphologically and kinetically similar to that induced by the neurotrophin nerve growth factor (NGF). However, DNA microarray analysis revealed a family of genes uniquely upregulated during Fas-stimulated neural regeneration, that was not induced during NGF-stimulate regeneration. Our data show that Fas is a trigger for axon regrowth, and suggest that Fas may engage an as yet unstudied intracellular program of neural regeneration. This new mechanism for promoting neuron survival and axon regrowth may lead to a novel approach in the treatment of neurodegenerative diseases such as Parkinson’s Disease.
This work was funded by NIH grant NS43773 to J.D.
8
ASSESSMENT OF NOVEL NEUROPROTECTIVE THERAPIES FOR ACUTE SPINAL CORD INJURY IN THE “POST-METHYLPREDNISOLONE ERA”
Schwartz, G.1,2, Liu, Y.2, Fehlings, M.G.1,2,
1Institute of Medical Science, University of Toronto, Toronto, ON, Canada
2Krembil Neuroscience Centre, University Health Network, Toronto, ON, Canada.
Methylprednisolone (MPS) remains the only pharmacotherapeutic agent used clinically after spinal cord injury (SCI). Given the modest benefits and potential toxicity of MPS, other neuroprotective approaches are required. Previously, we showed that acute administration of riluzole (Ril), a sodium channel blocker, is neuroprotective in rodents after SCI. Further experimentation to determine Ril’s and the novel sodium channel blocker topiramate’s (TP) optimal dose, time window for administration, and potential synergy with the MPSS was done in an in vitro organotypic slice model of SCI. Weight-drop injured slices were treated at several time points pre and post SCI with 10 µM of Ril, TP, MPS, or Ril+MPS. Fluorescent, propidium iodide (PI)-labeled cells were imaged by confocal microscopy at designated times post treatment. Mean PI cell counts from each treatment group were quantified to generate an index of cell death. Analysis by ANOVA indicated a significant effect of treatment (p=3.7x10-9 and p=9.29x10-13) with the neuroprotective drug treated slices consistently having less cell death than control slices. These findings suggest that the sodium channel blockers, Ril and TP, and MPS are effective neuroprotective strategies for SCI. Further preclinical in vivo evaluation is necessary prior to translation to clinical trials.
Topiramate was generously provided by Johnson & Johnson R & D.
Supported by the Dennis and Lucy Wing Scholarship Fund and the Ontario Neurotrauma Foundation.
9
4-METHYLCATECHOL PREVENTS EXPERIMENTAL ANTIMITOTIC-INDUCED NEUROPATHY IN THE RAT
Baillet-Mignard, C., Andriambeloson, E., Romanet, V., Callizot, N.
Neurofit SA, Rue Jean Sapidus, F-67400 Illkirch, France.
Cis-diamine-dichloro-platinium (cisplatin) and vincristine are drugs of choice for treatment of various types of cancers. The use of these anti-tumoral drugs is limited by serious adverse events, especially peripheral neuropathy. The neuropathy induced by cisplatin is predominantly sensory with initial complaints of distal paresthesiae while that induced by vincristine is a sensory and motoneuropathy. The present study was conducted in order to explore in rat models the value of 4-methylcatechol (4-MC), an inducer of nerve growth factor (NGF), as therapeutic approach to treat these two types of antimitotic-induced neuropathies in 2 rat models. In these models, sensory and motor impairments were evaluated, in parallel, by behavioral tests (hot plate and walking test), electrophysiological recordings (motor and reflex waves) and morphological analyses. Results showed that behavioral defects along with nerve dysfunction induced by antimitotic intoxication were improved by 4-MC (10 µg/kg/day; p.o.) treatment. These observations were consistent with morphological alteration of myelinated fibers and with marked reduction of degenerated fibers within the sciatic nerve from 4-MC to treat sensory disorders most probably via NGF-dependent mechanism. In addition, as 4-MC treatment hampered also the progression of motor dysfunction, a NGF-independent mechanism(s) could be suspected. Altogether, these observations suggest that 4-MC may represent an attractive approach for prophylaxis and therapy of chemotherapy-induced neuropathy.
10
MULTIMODAL NEUROPROTECTION: THE CAFFEINOL EXPERIENCE
Labiche, L.A., Aronowski, J., Grotta, J.C.
The University of Texas Health Science Center, Houston, Texas 77030.
The purpose of these studies was to identify the most effective multi-modal neuroprotectant strategy that can be safely coupled with reperfusion. The novel combinant of caffeine and ethanol (caffeinol) has demonstrated robust neuroprotection in our rodent model of focal transient ischemia. A significant 83% reduction in infarct volume was observed after the intravenous administration of caffeine 10 mg/kg and 5% ethanol 0.325 g/kg over 120 minutes. Significant benefit was seen even with treatment delayed up to 3 hours post-stroke. The target therapeutic plasma ranges were found to be 5-10 mcg/ml caffeine and 30-50 mg/dl ethanol. No increased rates of intracerebral hemorrhage were observed in vivo with combination of caffeinol and rt-PA. Furthermore, the neuroprotective effect of the combination of caffeinol and mild systemic hypothermia (35oC) was more robust than any other treatment alone. Therefore, an open-label, dose-escalation study of caffeinol was performed in 23 acute ischemic stroke patients to determine the optimal dose of caffeinol with regard to safety and achievement of therapeutic serum levels. Four dosage regimens were tested with doses ranging from caffeine 6-8 mg/kg and ethanol 0.2-0.4 g/kg administered within 6 hours of stroke onset. Concurrent administration of rt-PA was allowed, but not required. A does was successfully identified that achieved “therapeutic” plasma levels based on animal studies. This dosing schedule, caffeine 8 mg/kg and 10% ethanol 0.4 g/kg, was clinically well tolerated and did not produce any significant adverse effects. The next stage of our clinical research will evaluate the safety and feasibility of administering both caffeinol (within 4 hours) and mild systemic hypothermia (within 5 hours) in acute ischemic stroke patients treated with rt-PA.