Neuroprotective Effect of a CNTF-Expressing Lentiviral Vector in the Quinolinic Acid Rat Model of Huntington's Disease (original) (raw)
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Human Gene Therapy, 2002
The ability to regulate gene expression constitutes a prerequisite for the development of gene therapy strategies aimed at the treatment of neurologic disorders. In the present work, we used tetracycline (Tet)-regulated lentiviral vectors to investigate the dose-dependent neuroprotective effect of human ciliary neurotrophic factor (CNTF) in the quinolinic acid (QA) model of Huntington's disease (HD). The Tet system was split in two lentiviruses, the first one containing the CNTF or green fluorescent protein (GFP) cDNAs under the control of the Tet-response element (TRE) and a second vector encoding the transactivator (tTA). Preliminary coinfection study demonstrated that 63.8% 6 2.0% of infected cells contain at least two viral copies. Adult rats were then injected with CNTF-and GFP-expressing viral vectors followed 3 weeks later by an intrastriatal administration of QA. A significant reduction of apomorphine-induced rotations was observed in the CNTFon group. In contrast, GFP-treated animals or CNTF-off rats displayed an ipsilateral turning behavior in response to apomorphine. A selective sparing of DARPP-32-, choline acetyltransferase (ChAT)-, and NADPHd-positive neurons was observed in the striatum of CNTF-on rats compared to GFP animals and CNTF-off group. Enzyme-linked immunosorbent assay (ELISA) performed on striatal samples of rats sacrificed at the same time point indicated that this neuroprotective effect was associated with the production of 15.5 6 4.7 ng CNTF per milligram of protein whereas the residual CNTF expression in the off state (0.54 6 0.02 ng/mg of protein) was not sufficient to protect against QA toxicity. These results establish the proof of principle of neurotrophic factor dosing for neurodegenerative diseases and demonstrate the feasibility of lentiviral-mediated tetracycline-regulated gene transfer in the brain.
Lentiviral Vectors in Huntington's Disease Research and Therapy
Viral Vector Approaches in Neurobiology and Brain Diseases, 2014
We describe here the potential of viral-mediated gene transfer for the modeling and treatment of Huntington's disease, focusing in particular on strategies for the tissue-specific targeting of various CNS cells. The protocols described here cover the design of lentiviral vectors, strategies for modifying their tropism, including the use of various envelopes and tissue-specific promoters, and the potential of miRNA to regulate transgene expression.
Experimental Neurology, 2004
Ciliary neurotrophic factor (CNTF) has been shown to prevent behavioral deficits and striatal degeneration in neurotoxic models of Huntington's disease (HD), but its effect in a genetic model has not been evaluated. Lentiviral vectors expressing the human CNTF or LacZ reporter gene were therefore injected in the striatum of wild-type (WT) and transgenic mice expressing full-length huntingtin with 72 CAG repeats (YAC72). Behavioral analysis showed increased locomotor activity in 5-to 6-month-old YAC72-LacZ mice compared to WT-LacZ animals. Interestingly, CNTF expression reduced the activity levels of YAC72 mice compared to control animals. In both WT and YAC72 mice, CNTF expression was demonstrated in striatal punches, up to a year after lentiviral injection. Stereological analysis revealed that the number of LacZ and DARPP-32-positive neurons were decreased in YAC72-LacZ mice compared to WT-LacZ animals. Assessment of the benefit of CNTF expression in the YAC72 mice was, however, complicated by a down-regulation of DARPP-32 and to a lesser extent of NeuN in all mice treated with CNTF. The expression of the neuronal marker NADPH-d was unaffected by CNTF, but expression of the astrocytic marker glial fibrillary acidic protein (GFAP) was increased. Finally, a reduction of the number of striatal dark cells was observed in YAC mice treated with CNTF compared to LacZ. These data indicate that sustained striatal expression of CNTF can be achieved with lentiviruses. Further studies are, however, needed to investigate the intracellular signaling pathways mediating the long-term effects of CNTF expression on dopamine signaling, glial cell activation and how these changes may affect HD pathology. D
Proceedings of The National Academy of Sciences, 2006
Huntington's disease (HD) is a fatal, genetic, neurological disorder resulting from a trinucleotide repeat expansion in the gene that encodes for the protein huntingtin. These excessive repeats confer a toxic gain of function on huntingtin, which leads to the degeneration of striatal and cortical neurons and a devastating motor, cognitive, and psychological disorder. Trophic factor administration has emerged as a compelling potential therapy for a variety of neurodegenerative disorders, including HD. We previously demonstrated that viral delivery of glial cell line-derived neurotrophic factor (GDNF) provides structural and functional neuroprotection in a rat neurotoxin model of HD. In this report we demonstrate that viral delivery of GDNF into the striatum of presymptomatic mice ameliorates behavioral deficits on the accelerating rotorod and hind limb clasping tests in transgenic HD mice. Behavioral neuroprotection was associated with anatomical preservation of the number and size of striatal neurons from cell death and cell atrophy. Additionally, GDNF-treated mice had a lower percentage of neurons containing mutant huntingtin-stained inclusion bodies, a hallmark of HD pathology. These data further support the concept that viral vector delivery of GDNF may be a viable treatment for patients suffering from HD. gene therapy | neurodegeneration | neuroprotection | polyglutamine | adenoassociated virus
Experimental Neurology, 2003
Huntington's disease (HD) is an autosomal dominant disorder caused by an expanded polyglutamine (CAG) tract at the IT15 locus on chromosome 4. These excessive repeats lead to the degeneration of striatal and cortical neurons resulting in a devastating cognitive, psychiatric, and motor disorder for which no treatments are available. Neurotrophic factors support the viability of striatal neurons suggesting that they might prevent the inevitable neural degeneration and its accompanying functional decline associated with HD. The present study investigated whether glial cell line-derived neurotrophic factor (GDNF) delivered by an adeno associated virus could provide structural and functional neuroprotection in a rat model of HD. Lewis rats received bilateral injections of either AAV-GDNF (n = 12) or AAV-green fluorescence protein (AAV-GFP, n = 12) into the striatum followed 2 weeks later by chronic subcutaneous infusions of the mitochondrial toxin, 3-nitropropionic acid (3-NP, 38 mg/kg). All rats underwent 4 weeks of behavioral testing and were then sacrificed. Following 3-NP, the performance by AAV-GFP-treated rats on a raised platform motor task deteriorated while the performance by AAV-GDNF-treated rats was near normal (P < 0.001). AAV-GDNF-treated rats also received better scores on a blinded semi-quantitative neurological scale compared to rats receiving AAV-GFP (P < 0.001). Histological analyses supported our behavioral findings. 3-NP-treated rats receiving AAV-GDNF displayed 70% more NeuN-immunoreactive neurons compared to 3-NP-treated rats receiving AAV-GFP (P = 0.002). Similar findings were seen with dopamine-and-adenosine-3'5'-monophosphate-regulated phosphoprotein (DARPP-32) staining. These data indicate that the viral-mediated gene transfer of GDNF into the striatum provides neuroanatomical and behavioral protection in a rodent model of HD.
Human Gene Therapy, 1999
Huntington's disease (HD) is a genetic disorder leading to the degeneration of striatal GABAergic output neurons. No treatment is currently available for this devastating disorder, although several neurotrophic factors, including brain-derived neurotrophic factor (BDNF), have been shown to be beneficial for striatal neuron survival. We analyzed the effect of adenovirusmediated transfer of the BDNF gene in a model of HD. Using a stereological procedure, three groups of rats were given an intrastriatal injection of adenovirus encoding BDNF, ß-galactosidase or sham surgery. Two weeks after treatment, the animals were lesioned with quinolinic acid (QUIN), a toxin which induces striatal neuron death by an excitotoxic process. One month after the lesion, histological study revealed that striatal neurons were protected only in rats treated with the BDNF adenovirus. Volume measurements showed that the QUIN-induced lesions were 55% smaller in the BDNF adenovirus-treated group than in the ßgalactosidase adenovirus-treated group (p<0.05), and the sham-treated group (p<0.05). To determine the survival of striatal GABAergic output neurons after the QUINinduced lesion, we immunostained brain sections with DARPP-32, an antibody specific for striatal output neurons. Prior treatment with the BDNF adenovirus resulted in a cell survival of 64%, whereas that after ßgalactosidase treament was 46% (p<0.05), showing that the BDNF adenovirus protected the striatal neurons. These results indicate that transfer of the BDNF gene is of therapeutic value for Huntington's disease.
Human Gene Therapy, 2004
Huntington's disease (HD) is an autosom al dominant genetic disease with devastating clinical effects on cognitive, psychologic al, and motor functions. These clinical symptoms primarily relate to the progressive loss of medium-spiny GABA-ergic neurons of the striatum. There is no known treatment to date. Several neurotrophic factors have, however, demonstrated the capacity to protect striatal neurons in various experimental models of HD. This includes the ciliary neurotrophic factor (CNTF), the substance examined in this protocol. An ex vivo gene therapy approach based on encapsulated genetically modified BHK cells will be used for the continuous and long-term intracerebral delivery of CNTF. A device, containing up to 10 6 human CNTF-producing BHK cells surrounded by a semipermeable membrane, will be implanted into the right lateral ventricle of 6 patients. Capsules releasing 0.15-0.5 m g CNTF/day will be used. In this phase I study, the principal goal will be the evaluation of the safety and tolerability of the procedure. As a secondary goal, HD symptoms will be analyzed using a large battery of neuropsychological, motor, neurological, and neurophysiological tests and the striatal pathology monitored using MRI and PET-scan imaging. It is expected that the gene therapy approach described in this protocol will mitigate the side effects associated with the peripheral administration of recombinant hCNTF and allow a well-tolerated, continuous intracerebroventricular delivery of the neuroprotective factor. 1723 BACKGROUND H U N TIN G TO N 'S D IS EA SE (HD) is a genetic, autosom al dominant disease, with com plete penetrance, that specifically affects the brain. It inexorably provokes a profound cognitive impairment, motor abnormalities and severe psychiatric disorders. The disease can start clinically at any time in life but begins, in general, during the fourth decade. This means that it affects young adults who are working and, in many cases, have young children. They rapidly lose their autonomy and need constant care, first by the fam ily then in a hospital. They die after ten to twenty years (17 years as a mean) after the initiation of symptoms, showing profound dementia and abnormal motor behavior (see Ref. 10 for review). The total prevalence of HD is around 3 in 10,000 in Western countries, including 1 in 10,000 clinically identified HD and 2 in 10,000 gene carriers who will
Human Gene Therapy, 2000
Huntington's disease (HD) is an autosomal dominant genetic disease with devastating clinical effects on cognitive, psychological, and motor functions. These clinical symptoms primarily relate to the progressive loss of medium-spiny GABA-ergic neurons of the striatum. There is no known treatment to date. Several neurotrophic factors have, however, demonstrated the capacity to protect striatal neurons in various experimental models of HD. This includes the ciliary neurotrophic factor (CNTF), the substance examined in this protocol. An ex vivo gene therapy approach based on encapsulated genetically modified BHK cells will be used for the continuous and long-term intracerebral delivery of CNTF. A device, containing up to 106 human CNTF-producing BHK cells surrounded by a semipermeable membrane, will be implanted into the right lateral ventricle of 6 patients. Capsules releasing 0.15-0.5 microg CNTF/day will be used. In this phase I study, the principal goal will be the evaluation of the safety and tolerability of the procedure. As a secondary goal, HD symptoms will be analyzed using a large battery of neuropsychological, motor, neurological, and neurophysiological tests and the striatal pathology monitored using MRI and PET-scan imaging. It is expected that the gene therapy approach described in this protocol will mitigate the side effects associated with the peripheral administration of recombinant hCNTF and allow a well-tolerated, continuous intracerebroventricular delivery of the neuroprotective factor.
Experimental Neurology, 1999
The elucidation of the genetic defect in patients with Huntington's disease (HD) has allowed for the detection of individuals at risk for HD prior to the onset of symptoms. Thus ''neuroprotection strategies'' aimed at preventing the neuropathological and behavioral sequelae of this disease might be powerful therapeutically since they could be introduced to healthy patients before the initiation of a massive degenerative cascade principally localized to the striatum. A variety of trophic factors potently protect vulnerable striatal neurons in animal models of HD. A number of experimental variables are critical in determining the success of trophic factors in animal models. In this regard, the method of trophic factor delivery may be crucial, as delivery via genetically modified cells often produces greater and more widespread effects on striatal neurons than infusions of that same factor. The mechanisms by which cellularly delivered trophic factors forestall degeneration and prevent behavioral deficits are complex and often appear to be unrelated to the trophic factor binding to its cognate receptor. In this regard, cells genetically modified to secrete nerve growth factor (NGF) or ciliary neurotrophic factor (CNTF) protect degenerating striatal neurons which do not express either NGF or CNTF receptors. This review will discuss some of the non-receptor-based events that might underlie these effects and present the hypothesis that cellular delivery of certain trophic factors using genetically modified cells may be ready for clinical testing in HD patients. 1999 Academic Press