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Dolan Sondhi,
Linda Johnson,
Keith Purpura,
Sebastien Monette,
Mark M Souweidane, Michael G Kaplitt,
Barry Kosofsky,
Kaleb Yohay,
Douglas Ballon,
Jonathan Dyke,
Stephen M Kaminksy,
Neil R Hackett,
Ronald G Crystal
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ABSTRACT: Abstract Late infantile neuronal ceroid lipofuscinosis (LINCL), a fatal, lysosomal storage disorder caused by mutations in the CLN2 gene, results in a deficiency of tripeptidyl-peptidase I (TPP-I) activity in neurons. Our prior studies showed that delivery of the human CLN2 cDNA directly to the CNS, using an adeno-associated virus serotype 2 (AAV2) vector, is safe in children with LINCL. As a second-generation strategy, we have demonstrated that AAVrh.10hCLN2, a rhesus-derived AAV vector, mediates wide distribution of TPP-I through the CNS in a murine model. This study tests the hypothesis that direct administration of AAVrh.10hCLN2 to the CNS of rats and nonhuman primates at doses scalable to humans has an acceptable safety profile and mediates significant CLN2 expression in the CNS. A dose of 10(11) genome copies (GC) was administered bilaterally to the striatum of Sprague Dawley rats with sacrifice at 7 and 90 days with no significant impact except for mild vector-related histopathological changes at the site of vector administration. A dose of 1.8×10(12) GC of AAVrh.10hCLN2 was administered to the CNS of 8 African green monkeys. The vector-treated monkeys did not differ from controls in any safety parameter except for mild to moderate white matter edema and inflammation localized to the administration sites of the vector. There were no clinical sequelae to these localized findings. TPP-I activity was >2 SD over background in 31.7±8.1% of brain at 90 days. These findings establish the dose and safety profile for human clinical studies for the treatment of LINCL with AAVrh.10hCLN2.
Human gene therapy methods. 11/2012;
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ABSTRACT: A large number of studies have demonstrated that the nucleus accumbens (NAC) is a critical site in the neuronal circuits controlling reward responses, motivation, and mood, but the neuronal cell type(s) underlying these processes are not yet known. Identification of the neuronal cell types that regulate depression-like states will guide us in understanding the biological basis of mood and its regulation by diseases like major depressive disorder. Taking advantage of recent findings demonstrating that the serotonin receptor chaperone, p11, is an important molecular regulator of depression-like states, here we identify cholinergic interneurons (CINs) as a primary site of action for p11 in the NAC. Depression-like behavior is observed in mice after decrease of p11 levels in NAC CINs. This phenotype is recapitulated by silencing neuronal transmission in these cells, demonstrating that accumbal cholinergic neuronal activity regulates depression-like behaviors and suggesting that accumbal CIN activity is crucial for the regulation of mood and motivation.
Proceedings of the National Academy of Sciences 06/2012; 109(28):11360-5. · 9.68 Impact Factor
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ABSTRACT: We used a network approach to study the effects of anti-parkinsonian treatment on motor sequence learning in humans. Eight Parkinson's disease (PD) patients with bilateral subthalamic nucleus (STN) deep brain stimulation underwent H(2)(15)O positron emission tomography (PET) imaging to measure regional cerebral blood flow (rCBF) while they performed kinematically matched sequence learning and movement tasks at baseline and during stimulation. Network analysis revealed a significant learning-related spatial covariance pattern characterized by consistent increases in subject expression during stimulation (p = 0.008, permutation test). The network was associated with increased activity in the lateral cerebellum, dorsal premotor cortex, and parahippocampal gyrus, with covarying reductions in the supplementary motor area (SMA) and orbitofrontal cortex. Stimulation-mediated increases in network activity correlated with concurrent improvement in learning performance (p < 0.02). To determine whether similar changes occurred during dopaminergic pharmacotherapy, we studied the subjects during an intravenous levodopa infusion titrated to achieve a motor response equivalent to stimulation. Despite consistent improvement in motor ratings during infusion, levodopa did not alter learning performance or network activity. Analysis of learning-related rCBF in network regions revealed improvement in baseline abnormalities with STN stimulation but not levodopa. These effects were most pronounced in the SMA. In this region, a consistent rCBF response to stimulation was observed across subjects and trials (p = 0.01), although the levodopa response was not significant. These findings link the cognitive treatment response in PD to changes in the activity of a specific cerebello-premotor cortical network. Selective modulation of overactive SMA-STN projection pathways may underlie the improvement in learning found with stimulation.
Journal of Neuroscience 02/2012; 32(8):2804-13. · 7.11 Impact Factor
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Peter A LeWitt,
Ali R Rezai,
Maureen A Leehey,
Steven G Ojemann,
Alice W Flaherty,
Emad N Eskandar,
Sandra K Kostyk,
Karen Thomas,
Atom Sarkar,
Mustafa S Siddiqui, [......],
Jason M Schwalb,
Kathleen L Poston,
Jaimie M Henderson,
Roger M Kurlan,
Irene H Richard,
Lori Van Meter,
Christine V Sapan,
Matthew J During, Michael G Kaplitt,
Andrew Feigin
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ABSTRACT: Gene transfer of glutamic acid decarboxylase (GAD) and other methods that modulate production of GABA in the subthalamic nucleus improve basal ganglia function in parkinsonism in animal models. We aimed to assess the effect of bilateral delivery of AAV2-GAD in the subthalamic nucleus compared with sham surgery in patients with advanced Parkinson's disease.
Patients aged 30-75 years who had progressive levodopa-responsive Parkinson's disease and an overnight off-medication unified Parkinson's disease rating scale (UPDRS) motor score of 25 or more were enrolled into this double-blind, phase 2, randomised controlled trial, which took place at seven centres in the USA between Nov 17, 2008, and May 11, 2010. Infusion failure or catheter tip location beyond a predefined target zone led to exclusion of patients before unmasking for the efficacy analysis. The primary outcome measure was the 6-month change from baseline in double-blind assessment of off-medication UPDRS motor scores. This trial is registered with ClinicalTrials.gov, NCT00643890.
Of 66 patients assessed for eligibility, 23 were randomly assigned to sham surgery and 22 to AAV2-GAD infusions; of those, 21 and 16, respectively, were analysed. At the 6-month endpoint, UPDRS score for the AAV2-GAD group decreased by 8·1 points (SD 1·7, 23·1%; p<0·0001) and by 4·7 points in the sham group (1·5, 12·7%; p=0·003). The AAV2-GAD group showed a significantly greater improvement from baseline in UPDRS scores compared with the sham group over the 6-month course of the study (RMANOVA, p=0·04). One serious adverse event occurred within 6 months of surgery; this case of bowel obstruction occurred in the AAV2-GAD group, was not attributed to treatment or the surgical procedure, and fully resolved. Other adverse events were mild or moderate, likely related to surgery and resolved; the most common were headache (seven patients in the AAV2-GAD group vs two in the sham group) and nausea (six vs two).
The efficacy and safety of bilateral infusion of AAV2-GAD in the subthalamic nucleus supports its further development for Parkinson's disease and shows the promise for gene therapy for neurological disorders.
Neurologix.
The Lancet Neurology 03/2011; 10(4):309-19. · 23.46 Impact Factor
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ABSTRACT: The advent of viral gene therapy technology has contributed greatly to the study of a variety of medical conditions, and there is increasing promise for clinical translation of gene therapy into human treatments. Adeno-associated viral (AAV) vectors provide one of the more promising approaches to gene delivery, and have been used extensively over the last 20 years. Derived from nonpathogenic parvoviruses, these vectors allow for stable and robust expression of desired transgenes in vitro and in vivo. AAV vectors efficiently and stably transduce neurons, with some strains targeting neurons exclusively in the brain. Thus, AAV vectors are particularly useful for neurodegenerative diseases, which have led to numerous preclinical studies and several human trials of gene therapy in patients with Parkinson's disease, Alzheimer's disease, and pediatric neurogenetic disorders. Here, we describe an efficient and reliable method for the production and purification of AAV serotype 2 vectors for both in vitro and in vivo applications.
Methods in molecular biology (Clifton, N.J.) 01/2011; 793:443-55.
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Brian Alexander,
Jennifer Warner-Schmidt,
Therese Eriksson,
Carol Tamminga,
Margarita Arango-Lievano,
Margarita Arango-Llievano,
Subroto Ghose,
Mary Vernov,
Mihaela Stavarache,
Mihaela Stavarche,
Sergei Musatov,
Marc Flajolet,
Per Svenningsson,
Paul Greengard, Michael G Kaplitt
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ABSTRACT: The etiology of major depression remains unknown, but dysfunction of serotonergic signaling has long been implicated in the pathophysiology of this disorder. p11 is an S100 family member recently identified as a serotonin 1B [5-hydroxytryptamine 1B (5-HT(1B))] and serotonin 4 (5-HT(4)) receptor-binding protein. Mutant mice in which p11 is deleted show depression-like behaviors, suggesting that p11 may be a mediator of affective disorder pathophysiology. Using somatic gene transfer, we have now identified the nucleus accumbens as a key site of p11 action. Reduction of p11 with adeno-associated virus (AAV)-mediated RNA interference in the nucleus accumbens, but not in the anterior cingulate, of normal adult mice resulted in depression-like behaviors nearly identical to those seen in p11 knockout mice. Restoration of p11 expression specifically in the nucleus accumbens of p11 knockout mice normalized depression-like behaviors. Human nucleus accumbens tissue shows a significant reduction of p11 protein in depressed patients when compared to matched healthy controls. These results suggest that p11 loss in rodent and human nucleus accumbens may contribute to the pathophysiology of depression. Normalization of p11 expression within this brain region with AAV-mediated gene therapy may be of therapeutic value.
Science translational medicine 10/2010; 2(54):54ra76. · 7.80 Impact Factor
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Mark M Souweidane,
Justin F Fraser,
Lisa M Arkin,
Dolan Sondhi,
Neil R Hackett,
Stephen M Kaminsky,
Linda Heier,
Barry E Kosofsky,
Stefan Worgall,
Ronald G Crystal, Michael G Kaplitt
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ABSTRACT: The authors conducted a phase I study of late infantile neuronal ceroid lipofuscinosis using an adenoassociated virus serotype 2 (AAV2) vector containing the deficient CLN2 gene (AAV2(CU)hCLN2). The operative technique, radiographic changes, and surgical complications are presented.
Ten patients with late infantile neuronal ceroid lipofuscinosis disease each underwent infusion of AAV2(CU)hCLN2 (3 x 10(12) particle units) into 12 distinct cerebral locations (2 depths/bur hole, 75 minutes/infusion, and 2 microl/minute). Innovative surgical techniques were developed to overcome several obstacles for which little or no established techniques were available. Successful infusion relied on preoperative stereotactic planning to optimize a parenchymal target and diffuse administration. Six entry sites, each having 2 depths of injections, were used to reduce operative time and enhance distribution. A low-profile rigid fixation system with 6 integrated holding arms was utilized to perform simultaneous infusions within a practical time frame. Dural sealant with generous irrigation was used to avoid CSF egress with possible subdural hemorrhage or altered stereotactic registration.
Radiographically demonstrated changes were seen in 39 (65%) of 60 injection sites, confirming localization and infusion. There were no radiographically or clinically defined complications.
The neurosurgical considerations and results of this study are presented to offer guidance and a basis for the design of future gene therapy or other clinical trials in children that utilize direct therapeutic delivery.
Journal of Neurosurgery Pediatrics 08/2010; 6(2):115-22. · 1.53 Impact Factor
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ABSTRACT: Classical estrogen receptor-signaling mechanisms involve estradiol binding to intracellular nuclear receptors [estrogen receptor-alpha (ERalpha) and estrogen receptor-beta (ERbeta)] to promote changes in protein expression. Estradiol can also exert effects within seconds to minutes, however, a timescale incongruent with genomic signaling. In the brain, estradiol rapidly potentiates stimulated dopamine release in the striatum of female rats and enhances spontaneous rotational behavior. Furthermore, estradiol rapidly attenuates the K(+)-evoked increase of GABA in dialysate. We hypothesize that these rapid effects of estradiol in the striatum are mediated by ERalpha located on the membrane of medium spiny GABAergic neurons. This experiment examined whether overexpression of ERalpha in the striatum would enhance the effect of estradiol on rotational behavior and the K(+)-evoked increase in GABA in dialysate. Ovariectomized female rats were tested for rotational behavior or underwent microdialysis experiments after unilateral intrastriatal injections of a recombinant adeno-associated virus (AAV) containing the human ERalpha cDNA (AAV.ERalpha) into the striatum; controls received either the same vector into areas outside the striatum or an AAV containing the human alkaline phosphatase gene into the striatum (AAV.ALP). Animals that received AAV.ERalpha in the striatum exhibited significantly greater estradiol-induced contralateral rotations compared with controls and exhibited behavioral sensitization of contralateral rotations induced by a low-dose of amphetamine. ERalpha overexpression also enhanced the inhibitory effect of estradiol on K(+)-evoked GABA release suggesting that disinhibition of dopamine release from terminals in the striatum resulted in the enhanced rotational behavior.
Journal of Neuroscience 03/2009; 29(6):1897-903. · 7.11 Impact Factor
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ABSTRACT: The authors investigated whether the insertion of deep brain stimulation electrodes into the subthalamic nucleus can alter regional brain metabolism in the absence of stimulation.
Six patients with Parkinson disease (PD) underwent preoperative FDG PET scanning, and again after STN electrode implantation with stimulation turned off.
Compared with baseline values, glucose utilization was reduced in the postoperative off-stimulation scans in the putamen/globus pallidus and in the ventral thalamus (p < 0.01), and there was increased metabolism in the sensorimotor cortex and cerebellum (p < 0.005). The expression of a specific PD-related spatial covariance pattern measured in the FDG PET data did not change after electrode implantation (p = 0.36), nor was there a significant change in clinical motor ratings (p = 0.44). Differences in PD-related spatial covariance pattern expression among the patients after electrode implantation did, however, correlate with the number of microelectrode recording trajectories placed during surgery (r = -0.82, p < 0.05).
These findings suggest that electrode implantation can impart a microlesion effect on regional brain function. Nonetheless, these local changes did not cross the threshold of network modulation needed to achieve clinical benefit.
Journal of Neurosurgery 03/2009; 110(6):1278-82. · 2.96 Impact Factor
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Wencheng Liu,
Cristofol Vives-Bauza,
Rebeca Acín-Peréz,
Ai Yamamoto,
Yingcai Tan,
Yanping Li,
Jordi Magrané,
Mihaela A Stavarache,
Sebastian Shaffer,
Simon Chang, Michael G Kaplitt,
Xin-Yun Huang,
M Flint Beal,
Giovanni Manfredi,
Chenjian Li
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ABSTRACT: Mutations in PTEN induced kinase 1 (PINK1), a mitochondrial Ser/Thr kinase, cause an autosomal recessive form of Parkinson's disease (PD), PARK6. Here, we report that PINK1 exists as a dimer in mitochondrial protein complexes that co-migrate with respiratory chain complexes in sucrose gradients. PARK6 related mutations do not affect this dimerization and its associated complexes. Using in vitro cell culture systems, we found that mutant PINK1 or PINK1 knock-down caused deficits in mitochondrial respiration and ATP synthesis. Furthermore, proteasome function is impaired with a loss of PINK1. Importantly, these deficits are accompanied by increased alpha-synclein aggregation. Our results indicate that it will be important to delineate the relationship between mitochondrial functional deficits, proteasome dysfunction and alpha-synclein aggregation.
PLoS ONE 02/2009; 4(2):e4597. · 4.09 Impact Factor
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Frontiers of neurology and neuroscience 02/2009; 25:174-9.
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Michael G Kaplitt
Frontiers of neurology and neuroscience 02/2009; 25:180-8.
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ABSTRACT: The mitochondrial protease OMI (also known as HtrA2) has been implicated in Parkinson's Disease (PD) and deletion or protease domain point mutations have shown profound neuropathologies in mice. A beneficial role by OMI, in preserving cell viability, is assumed to occur via the avoidance of dysfunctional protein turnover. However relatively few substrates for mitochondrial Omi are known. Here we report our identification of three novel mitochondrial substrates that impact metabolism and ATP production. Using a dual proteomic approach we have identified three interactors based upon ability to bind to OMI, and/or to persist in the proteome after OMI activity has been selectively inhibited. One candidate, the chaperone HSPA8, was common to each independent study. Two others (PDHB subunit and IDH3A subunit) did not appear to bind to OMI, however persisted in the mito-proteome when OMI was inhibited. Pyruvate dehydrogenase (PDH) and isocitrate dehydrogenase (IDH) are two key Kreb's cycle enzymes that catalyse oxidative decarboxylation control points in mitochondrial respiration. We verified both PDHB and IDH3A co-immunoprecipitate with HSPA8 and after elution, were degraded by recombinant HtrA2 in vitro. Additionally our gene expression studies, using rotenone (an inhibitor of Complex I) showed Omi expression was silenced when pdhb and idh3a were increased when a sub-lethal dose was applied. However higher dose treatment caused increased Omi expression and decreased levels of pdhb and idh3a transcripts. This implicates mitochondrial OMI in a novel mechanism relating to metabolism.
PLoS ONE 01/2009; 4(9):e7100. · 4.09 Impact Factor
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Stefan Worgall,
Dolan Sondhi,
Neil R Hackett,
Barry Kosofsky,
Minal V Kekatpure,
Nurunisa Neyzi,
Jonathan P Dyke,
Douglas Ballon,
Linda Heier,
Bruce M Greenwald,
Paul Christos,
Madhu Mazumdar,
Mark M Souweidane, Michael G Kaplitt,
Ronald G Crystal
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ABSTRACT: Late infantile neuronal ceroid lipofuscinosis (LINCL) is an autosomal recessive, neurodegenerative lysosomal storage disease affecting the CNS and is fatal by age 8 to 12 years. A total average dose of 2.5 10(12) particle units of an adeno-associated virus (AAV) serotype 2 vector expressing the human CLN2 cDNA (AAV2 CU h-CLN2) was administered to 12 locations in the CNS of 10 children with LINCL. In addition to safety parameters, a neurological rating scale (primary variable) and three quantitative magnetic resonance imaging (MRI) parameters (secondary variables) were used to compare the rate of neurological decline for 18 months in treated subjects compared with untreated subjects. Although there were no unexpected serious adverse events that were unequivocally attributable to the AAV2 CU hCLN2 vector, there were serious adverse effects, the etiology of which could not be determined under the conditions of the experiment. One subject died 49 days postsurgery after developing status epilepticus on day 14, but with no evidence of CNS inflammation. Four of the 10 subjects developed a mild, mostly transient, humoral response to the vector. Compared with control subjects, the measured rates of decline of all MRI parameters were slower, albeit the numbers were too small for statistical significance. Importantly, assessment of the neurologic rating scale, which was the primary outcome variable, demonstrated a significantly reduced rate of decline compared with control subjects. Although the trial is not matched, randomized, or blinded and lacked a contemporaneous placebo/sham control group, assessment of the primary outcome variable suggests a slowing of progression of LINCL in the treated children. On this basis, we propose that additional studies to assess the safety and efficacy of AAV-mediated gene therapy for LINCL are warranted.
Human gene therapy 06/2008; 19(5):463-74. · 4.20 Impact Factor
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ABSTRACT: Parkinson's disease (PD) is characterized by elevated expression of an abnormal metabolic brain network that is reduced by clinically effective treatment. We used fluorodeoxyglucose (FDG) positron emission tomography (PET) to determine the basis for motor improvement in 12 PD patients receiving unilateral subthalamic nucleus (STN) infusion of an adenoassociated virus vector expressing glutamic acid decarboxylase (AAV-GAD). After gene therapy, we observed significant reductions in thalamic metabolism on the operated side as well as concurrent metabolic increases in ipsilateral motor and premotor cortical regions. Abnormal elevations in the activity of metabolic networks associated with motor and cognitive functioning in PD patients were evident at baseline. The activity of the motor-related network declined after surgery and persisted at 1 year. These network changes correlated with improved clinical disability ratings. By contrast, the activity of the cognition-related network did not change after gene transfer. This suggests that modulation of abnormal network activity underlies the clinical outcome observed after unilateral STN AAV-GAD gene therapy. Network biomarkers may be used as physiological assays in early-phase trials of experimental therapies for PD and other neurodegenerative disease.
Proceedings of the National Academy of Sciences 01/2008; 104(49):19559-64. · 9.68 Impact Factor
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Michael G Kaplitt,
Andrew Feigin,
Chengke Tang,
Helen L Fitzsimons,
Paul Mattis,
Patricia A Lawlor,
Ross J Bland,
Deborah Young,
Kristin Strybing,
David Eidelberg,
Matthew J During
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ABSTRACT: Dopaminergic neuronal loss in Parkinson's disease leads to changes in the circuitry of the basal ganglia, such as decreased inhibitory GABAergic input to the subthalamic nucleus. We aimed to measure the safety, tolerability, and potential efficacy of transfer of glutamic acid decarboxylase (GAD) gene with adeno-associated virus (AAV) into the subthalamic nucleus of patients with Parkinson's disease.
We did an open label, safety and tolerability trial of unilateral subthalamic viral vector (AAV-GAD) injection in 11 men and 1 woman with Parkinson's disease (mean age 58.2, SD=5.7 years). Four patients received low-dose, four medium-dose, and four high-dose AAV-GAD at New York Presbyterian Hospital. Inclusion criteria consisted of Hoehn and Yahr stage 3 or greater, motor fluctuations with substantial off time, and age 70 years or less. Patients were assessed clinically both off and on medication at baseline and after 1, 3, 6, and 12 months at North Shore Hospital. Efficacy measures included the Unified Parkinson's Disease Rating Scale (UPDRS), scales of activities of daily living (ADL), neuropsychological testing, and PET imaging with 18F-fluorodeoxyglucose. The trial is registered with the ClinicalTrials.gov registry, number NCT00195143.
All patients who enrolled had surgery, and there were no dropouts or patients lost to follow-up. There were no adverse events related to gene therapy. Significant improvements in motor UPDRS scores (p=0.0015), predominantly on the side of the body that was contralateral to surgery, were seen 3 months after gene therapy and persisted up to 12 months. PET scans revealed a substantial reduction in thalamic metabolism that was restricted to the treated hemisphere, and a correlation between clinical motor scores and brain metabolism in the supplementary motor area.
AAV-GAD gene therapy of the subthalamic nucleus is safe and well tolerated by patients with advanced Parkinson's disease, suggesting that in-vivo gene therapy in the adult brain might be safe for various neurodegenerative diseases.
The Lancet 07/2007; 369(9579):2097-105. · 38.28 Impact Factor
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ABSTRACT: Gene therapy with an adeno-associated viral (AAV) vector encoding the X-linked inhibitor of apoptosis protein (XIAP) in an animal model of cisplatin-induced ototoxicity can elucidate apoptotic pathways in the inner ear.
Cisplatin is limited clinically by ototoxicity associated with apoptosis in the inner ear. The relevant intracellular apoptotic pathways, however, are unknown. XIAP is an antiapoptotic protein that both inhibits caspases and reciprocally regulates the proapoptotic Smac/Omi proteins. AAV-mediated delivery of various XIAP mutants could distinguish between these antiapoptotic pathways in the ear and further the development of specific reagents for gene therapy- mediated prevention of cisplatin-induced ototoxicity.
We administered unilaterally through the round-window AAV-harboring genes encoding wild-type dXIAP, yellow fluorescent protein, or either of two dXIAP point mutants-one deficient in caspase inhibition (dXIAP-d) and the other additionally deficient in the binding of Smac/Omi (dXIAP-t). All rats received a 3-day systemic course of cisplatin. Functional hearing loss was measured by shifts in auditory brainstem response (ABR) thresholds after cisplatin treatment, and hair-cell loss was assessed by whole-mount phalloidin staining of cochlear turns.
Uninjected ears universally displayed high-frequency-specific hair-cell loss and ABR threshold shifts upon cisplatin treatment. Although yellow fluorescent protein had no effect, ears injected with dXIAP exhibited 68% less ABR threshold shift at 32 kHz and 50% less basal-turn outer-hair-cell loss compared with contralateral untreated ears. This protection was maintained in ears injected with dXIAP-d but was abolished in those expressing dXIAP-t, which is incapable of blocking Smac/Omi. CONCLUSIO: Hair-cell apoptosis induced by cisplatin involves the Smac/Omi pathway. Thus, gene therapy with either wild-type dXIAP or Smac/Omi-selective dXIAP-d may be effective to protect against cisplatin-mediated ototoxicity.
Ontology & Neurotology 05/2007; 28(3):417-25. · 1.90 Impact Factor
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Marina E Emborg,
Maren Carbon,
James E Holden,
Matthew J During,
Yilong Ma,
Chengke Tang,
Jeffrey Moirano,
Helen Fitzsimons,
Ben Z Roitberg,
Eray Tuccar,
Andrew Roberts, Michael G Kaplitt,
David Eidelberg
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ABSTRACT: Parkinson's disease (PD) is associated with increased excitatory activity within the subthalamic nucleus (STN). We sought to inhibit STN output in hemiparkinsonian macaques by transfection with adeno-associated virus (AAV) containing the gene for glutamic acid decarboxylase (GAD). In total, 13 macaques were rendered hemiparkinsonian by right intracarotid 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine injection. Seven animals were injected with AAV-GAD into the right STN, and six received an AAV gene for green fluorescent protein (GFP). Videotaped motor ratings were performed in a masked fashion on a weekly basis over a 55-week period. At 56 weeks, the animals were scanned with (18)F-fluorodeoxyglucose (FDG) positron emission tomography (PET). Histological examination was performed at the end of the study. No adverse events were observed after STN gene therapy. We found that the clinical rating scores for the two treatment groups had different patterns of change over time (group x time interaction, P<0.001). On FDG PET, the GAD animals exhibited an increase in glucose utilization in the right motor cortex relative to GFP controls (P<0.001). Metabolism in this region correlated with clinical ratings at end point (P<0.01). Histology confirmed GAD expression in treated animals. These findings suggest that STN AAV-GAD is well tolerated and potentially effective in a primate model of PD. The changes in motor cortical glucose utilization observed after gene therapy are consistent with the modulation of metabolic brain networks associated with this disorder.
Journal of Cerebral Blood Flow & Metabolism 03/2007; 27(3):501-9. · 5.01 Impact Factor
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ABSTRACT: Estrogen receptor alpha (ERalpha) plays a pivotal role in the regulation of food intake and energy expenditure by estrogens. Although it is well documented that a disruption of ERalpha signaling in ERalpha knockout (ERKO) mice leads to an obese phenotype, the sites of estrogen action and mechanisms underlying this phenomenon are still largely unknown. In the present study, we exploited RNA interference mediated by adeno-associated viral vectors to achieve focused silencing of ERalpha in the ventromedial nucleus of the hypothalamus, a key center of energy homeostasis. After suppression of ERalpha expression in this nucleus, female mice and rats developed a phenotype characteristic for metabolic syndrome and marked by obesity, hyperphagia, impaired tolerance to glucose, and reduced energy expenditure. This phenotype persisted despite normal ERalpha levels elsewhere in the brain. Although an increase in food intake preceded weight gain, our data suggest that a leading factor of obesity in this model is likely a decline in energy expenditure with all three major constituents being affected, including voluntary activity, basal metabolic rate, and diet-induced thermogenesis. Together, these findings indicate that ERalpha in the ventromedial nucleus of the hypothalamus neurons plays an essential role in the control of energy balance and the maintenance of normal body weight.
Proceedings of the National Academy of Sciences 03/2007; 104(7):2501-6. · 9.68 Impact Factor
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ABSTRACT: Adrenomedullin is a vasoactive peptide that is upregulated in higher-grade gliomas and promotes tumor cell proliferation. Since reduced activity of the anti-oncogene PTEN seems to also correlate with higher tumor grade, this suggests an inverse association between PTEN activity and adrenomedullin expression. PC12 pheochromocytoma and human U251 glioma cell lines were stably transfected with human PTEN or control plasmid. Adrenomedullin expression was analyzed using quantitative PCR and Western blotting. A cell proliferation assay was used to assess adrenomedullin effects on U251 cells overexpressing PTEN. PC12 and U251 cells overexpressing PTEN had 17- and 8-fold decreases in adrenomedullin mRNA levels, respectively, compared to control cells. Cellular and secreted adrenomedullin peptide was similarly reduced. Addition of adrenomedullin to medium of controlled cells induced proliferation, as described previously, but U251 cells overexpressing PTEN did not respond to exogenous adrenomedullin. Further exploration revealed that PTEN also inhibits expression of the gliomas receptor for adrenomedullin, which accounts for this effect. These data were all replicated with an inducible PTEN construct confirming that these effects are not exclusively secondary to chronic overexpression. Given the profound effects of adrenomedullin on tumor cells, this is a novel and previously unidentified mechanism by which alterations in PTEN levels or function may influence tumor growth.
Journal of Neuro-Oncology 10/2006; 79(2):117-23. · 3.21 Impact Factor
