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596. AAV Gene Delivery of the Anti-Tau Antibody PHF1 Reduces Brain Tau Pathology in P301L Mice
Neurologic Diseases (Including Ophthalmic and Auditory Diseases) III Neurologic Diseases (Including Ophthalmic and Auditory Diseases) III 596. AAV Gene Delivery of the Anti-Tau Antibody PHF1 Reduces Brain Tau Pathology in P301L Mice
Wencheng Liu,1 Lingzhi Zhao,1 Maria J. Chiuchiolo,2 Fangmin Yu,1 Thomas Woo,1 Man-Ying Wong,1 Brittany Black,1 Peter Davies,3 Clarisse Jose,2 Jonathan Rosenberg,2 Stephen M. Kaminsky,2 Dolan Sondhi,2 Ronald G. Crystal,2 Steven M. Paul.1 1 Appel Alzheimer’s Disease Research Institute, Feil Family Brain and Mind Research Institute, Weill Cornell Medical College, New York, NY; 2Department of Genetic Medicine, Weill Cornell Medical College, New York, NY; 3Feinstein Institute for Medical Research, North Shore LIJ, Manhassett, NY. Anti-tau immunotherapy has been proposed as a promising therapy for various tauopathies including Alzheimer’s disease (AD), frontotemporal dementia (FTD) and progressive supranuclear palsy (PSP). However, there are limitations to passive immunization including the need for repeated administration and the very low levels of antibody that get into brain from the circulation. To circumvent the disadvantages of passive immunization, we have used serotype rh.10 adeno-associated virus vectors to deliver an anti-tau monoclonal antibody PHF1, known to reduce tau pathology following passive immunization in several tauopathy mouse models directly to the CNS. To accomplish this, we stereotactically administered 1010 viral genomes (vg) of AAVrh.10-PHF1 or AAVrh.10-mCherry as a control, bilaterally into the hippocampus of young (3.5-month old) homozygous P301L mice, a model that develops robust tau pathology in an age- and brain region-dependent manner. Six months after injection, mice were sacrificed to measure the presence of anti-tau antibody as well as the effects of treatment on tau pathology in several brain regions using specific ELISAs and immunohistochemistry (IHC) for quantifying pathological and normal tau. In pilot experiments using wild-type C57BL/6 and P301L mice, we observed much higher levels of PHF1 antibody in the hippocampus when delivered via the AAVrh.10 vector compared to passive immunization (³100-fold higher). We observed that treatment with AAVrh.10-PHF1 resulted in a marked antibody-dependent reduction in tau pathology in P301L mice, including a highly significant reduction in tau pathology in the hippocampus (≥80–90% p=0.001) compared to the control vector. There was also no reduction in total tau measured by ELISA in any brain region examined. Based on these observations, AAVrh.10 gene delivery of anti-tau monoclonal antibodies to the CNS may represent a novel therapeutic strategy for treating various tauopathies including FTD, PSP and AD.
597. AAV.NT-3 Treatment Attenuates Spontaneous Autoimmune Peripheral Polyneuropathy
Mehmet E. Yalvac,1 William D. Arnold,2,3 Cilwyn Braganza,1 Lei Chen,1 Jerry R. Mendell,1,3 Zarife Sahenk.1,3 1 Center for Gene Therapy, Nationwide Children’s Hospital, Columbus, OH; 2Department of Physical Medicine and Rehabilitation, The Ohio State University, Columbus, OH; 3 Department of Neurology, The Ohio State University, Columbus, OH. The spontaneous autoimmune peripheral polyneuropathy (SAPP) model in B7-2 knockout non-obese diabetic (NOD) mice demonstrates overlapping clinical features with chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). Secondary axonal loss is prominent in the progressive phase of this neuropathy. Neurotrophin 3 (NT-3) is an important autocrine factor supporting Molecular Therapy Volume 23, Supplement 1, May 2015 Copyright © The American Society of Gene & Cell Therapy
Schwann cell (SC) survival and differentiation and stimulates neurite outgrowth and myelination. Recently, NT-3 delivered in bone marrowderived neural stem cells was shown to exert therapeutic effect, by reducing the inflammation and accelerating remyelination in experimental encephalomyelitis (EAE), an animal model of multiple sclerosis. These findings led to considerations of AAV.NT-3 gene therapy via intramuscular delivery into SAPP mice to attenuate the disease. The results show AAV.NT-3 injection into gastrocnemius muscle of 25 week old SAPP mice produced measurable NT-3 levels in the serum at 7 weeks post gene delivery, sufficient to protect sciatic nerve motor function and compound muscle action potentials (CMAPs). Sciatic nerves of mice, receiving gene transfer had a significantly greater number of remyelinated fibers and decreased infiltration of CD3+ T cells compared to controls. These data correlated with reduced expression of the pro-inflammatory cytokines, interleukin-1β (IL-1β) and tumor necrosis factor (Tnf-α) and increased expression of anti-inflammatory markers, interleukin-10 and forkhead box-P3 (Foxp3) in the sciatic nerve. The treatment also increased the percentage of CD4+CD25+Foxp3+ regulatory T cells (Tregs) in the spleen. This proof-of-principle study established a therapeutic benefit by neuroprotective and immunomodulatory effects of AAV-NT-3 treatment in SAPP. This study demonstrates the potential clinical translational path for AAV delivered NT-3 for refractory cases of CIDP..
598. Fusion of Mesenchymal Stem Cells Exclusively with “Degenerating” Cerebellar Neurons in Spinocerebellar Ataxia Type 1 Model Mice
Fathul Huda,2 Yiping Fan,1 Ayumu Konno,2 Yasunori Matsuzaki,2 Nobutaka Takahashi,2 Jerry Chan,1 Hirokazu Hirai.2 1 Department of Reproductive Medicine, KK Women’s and Children’s Hospital, Singapore, Singapore; 2Department of Neurophysiology, Gunma University, Gunma, Japan. Mesenchymal stem cells (MSCs) have the potential to migrate to damaged tissues where they participate in tissue repair. Previous studies suggest that intravenous or intracortical injections of MSCs results in the emergence of binucleated cerebellar Purkinje cells (PCs) containing a MSC-derived marker protein in mice suggestive of a fusion event. However, transdifferentiation of MSCs into PCs or transfer of a marker protein from a MSC to a PC cannot be completely ruled-out. Here we unequivocally showed a fusion of human fetal MSCs (hMSCs) with mouse PCs by combination of tetracyclineregulated system with Cre-dependent genetic inversion switch (flip-excision; FLEx). We performed intra-cerebellar injection of adeno-associated viral vectors expressing tetracycline transactivator (tTA) and Cre recombinase into either non-symptomatic (4 weekold) or clearly symptomatic (6 – 8 month-old) spinocerebellar ataxia type 1 (SCA1) mice. Two weeks after the treatment, the mice received injection of 50,000 geneticially-engineered hMSCs that expressed GFP only in the presence of Cre recombinase and tTA, which restricted GFP expression exclusively to cerebellar cells fused with hMSCs. We observed GFP-expressing PCs and interneurons in symptomatic, but not non-symptomatic, SCA1 mice as early as 2 weeks after the MSC injection. The frequency of GFP-expressing PCs did not increase when analysed at longer time points (~23 weeks post hMSC grafting), suggesting that most hMSC fusion events occurred within 2 weeks of delivery. Using this model, we show conclusively that hMSC display a tendency to fuse with degenerating PC and interneurons, and not with normal neurons.
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Wencheng Liu,1 Lingzhi Zhao,1 Maria J. Chiuchiolo,2 Fangmin Yu,1 Thomas Woo,1 Man-Ying Wong,1 Brittany Black,1 Peter Davies,3 Clarisse Jose,2 Jonathan Rosenberg,2 Stephen M. Kaminsky,2 Dolan Sondhi,2 Ronald G. Crystal,2 Steven M. Paul.1 1 Appel Alzheimer’s Disease Research Institute, Feil Family Brain and Mind Research Institute, Weill Cornell Medical College, New York, NY; 2Department of Genetic Medicine, Weill Cornell Medical College, New York, NY; 3Feinstein Institute for Medical Research, North Shore LIJ, Manhassett, NY. Anti-tau immunotherapy has been proposed as a promising therapy for various tauopathies including Alzheimer’s disease (AD), frontotemporal dementia (FTD) and progressive supranuclear palsy (PSP). However, there are limitations to passive immunization including the need for repeated administration and the very low levels of antibody that get into brain from the circulation. To circumvent the disadvantages of passive immunization, we have used serotype rh.10 adeno-associated virus vectors to deliver an anti-tau monoclonal antibody PHF1, known to reduce tau pathology following passive immunization in several tauopathy mouse models directly to the CNS. To accomplish this, we stereotactically administered 1010 viral genomes (vg) of AAVrh.10-PHF1 or AAVrh.10-mCherry as a control, bilaterally into the hippocampus of young (3.5-month old) homozygous P301L mice, a model that develops robust tau pathology in an age- and brain region-dependent manner. Six months after injection, mice were sacrificed to measure the presence of anti-tau antibody as well as the effects of treatment on tau pathology in several brain regions using specific ELISAs and immunohistochemistry (IHC) for quantifying pathological and normal tau. In pilot experiments using wild-type C57BL/6 and P301L mice, we observed much higher levels of PHF1 antibody in the hippocampus when delivered via the AAVrh.10 vector compared to passive immunization (³100-fold higher). We observed that treatment with AAVrh.10-PHF1 resulted in a marked antibody-dependent reduction in tau pathology in P301L mice, including a highly significant reduction in tau pathology in the hippocampus (≥80–90% p=0.001) compared to the control vector. There was also no reduction in total tau measured by ELISA in any brain region examined. Based on these observations, AAVrh.10 gene delivery of anti-tau monoclonal antibodies to the CNS may represent a novel therapeutic strategy for treating various tauopathies including FTD, PSP and AD.
597. AAV.NT-3 Treatment Attenuates Spontaneous Autoimmune Peripheral Polyneuropathy
Mehmet E. Yalvac,1 William D. Arnold,2,3 Cilwyn Braganza,1 Lei Chen,1 Jerry R. Mendell,1,3 Zarife Sahenk.1,3 1 Center for Gene Therapy, Nationwide Children’s Hospital, Columbus, OH; 2Department of Physical Medicine and Rehabilitation, The Ohio State University, Columbus, OH; 3 Department of Neurology, The Ohio State University, Columbus, OH. The spontaneous autoimmune peripheral polyneuropathy (SAPP) model in B7-2 knockout non-obese diabetic (NOD) mice demonstrates overlapping clinical features with chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). Secondary axonal loss is prominent in the progressive phase of this neuropathy. Neurotrophin 3 (NT-3) is an important autocrine factor supporting Molecular Therapy Volume 23, Supplement 1, May 2015 Copyright © The American Society of Gene & Cell Therapy
Schwann cell (SC) survival and differentiation and stimulates neurite outgrowth and myelination. Recently, NT-3 delivered in bone marrowderived neural stem cells was shown to exert therapeutic effect, by reducing the inflammation and accelerating remyelination in experimental encephalomyelitis (EAE), an animal model of multiple sclerosis. These findings led to considerations of AAV.NT-3 gene therapy via intramuscular delivery into SAPP mice to attenuate the disease. The results show AAV.NT-3 injection into gastrocnemius muscle of 25 week old SAPP mice produced measurable NT-3 levels in the serum at 7 weeks post gene delivery, sufficient to protect sciatic nerve motor function and compound muscle action potentials (CMAPs). Sciatic nerves of mice, receiving gene transfer had a significantly greater number of remyelinated fibers and decreased infiltration of CD3+ T cells compared to controls. These data correlated with reduced expression of the pro-inflammatory cytokines, interleukin-1β (IL-1β) and tumor necrosis factor (Tnf-α) and increased expression of anti-inflammatory markers, interleukin-10 and forkhead box-P3 (Foxp3) in the sciatic nerve. The treatment also increased the percentage of CD4+CD25+Foxp3+ regulatory T cells (Tregs) in the spleen. This proof-of-principle study established a therapeutic benefit by neuroprotective and immunomodulatory effects of AAV-NT-3 treatment in SAPP. This study demonstrates the potential clinical translational path for AAV delivered NT-3 for refractory cases of CIDP..
598. Fusion of Mesenchymal Stem Cells Exclusively with “Degenerating” Cerebellar Neurons in Spinocerebellar Ataxia Type 1 Model Mice
Fathul Huda,2 Yiping Fan,1 Ayumu Konno,2 Yasunori Matsuzaki,2 Nobutaka Takahashi,2 Jerry Chan,1 Hirokazu Hirai.2 1 Department of Reproductive Medicine, KK Women’s and Children’s Hospital, Singapore, Singapore; 2Department of Neurophysiology, Gunma University, Gunma, Japan. Mesenchymal stem cells (MSCs) have the potential to migrate to damaged tissues where they participate in tissue repair. Previous studies suggest that intravenous or intracortical injections of MSCs results in the emergence of binucleated cerebellar Purkinje cells (PCs) containing a MSC-derived marker protein in mice suggestive of a fusion event. However, transdifferentiation of MSCs into PCs or transfer of a marker protein from a MSC to a PC cannot be completely ruled-out. Here we unequivocally showed a fusion of human fetal MSCs (hMSCs) with mouse PCs by combination of tetracyclineregulated system with Cre-dependent genetic inversion switch (flip-excision; FLEx). We performed intra-cerebellar injection of adeno-associated viral vectors expressing tetracycline transactivator (tTA) and Cre recombinase into either non-symptomatic (4 weekold) or clearly symptomatic (6 – 8 month-old) spinocerebellar ataxia type 1 (SCA1) mice. Two weeks after the treatment, the mice received injection of 50,000 geneticially-engineered hMSCs that expressed GFP only in the presence of Cre recombinase and tTA, which restricted GFP expression exclusively to cerebellar cells fused with hMSCs. We observed GFP-expressing PCs and interneurons in symptomatic, but not non-symptomatic, SCA1 mice as early as 2 weeks after the MSC injection. The frequency of GFP-expressing PCs did not increase when analysed at longer time points (~23 weeks post hMSC grafting), suggesting that most hMSC fusion events occurred within 2 weeks of delivery. Using this model, we show conclusively that hMSC display a tendency to fuse with degenerating PC and interneurons, and not with normal neurons.
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