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1.
Mol Ther ; 32(1): 44-58, 2024 Jan 03.
Artigo em Inglês | MEDLINE | ID: mdl-37952085

RESUMO

Hematopoietic stem cell transplantation (HSCT) is the only approved treatment for presymptomatic infantile globoid cell leukodystrophy (GLD [Krabbe disease]). However, correction of disease is not complete, and outcomes remain poor. Herein we evaluated HSCT, intravenous (IV) adeno-associated virus rh10 vector (AAVrh10) gene therapy, and combination HSCT + IV AAVrh10 in the canine model of GLD. While HSCT alone resulted in no increase in survival as compared with untreated GLD dogs (∼16 weeks of age), combination HSCT + IV AAVrh10 at a dose of 4E13 genome copies (gc)/kg resulted in delayed disease progression and increased survival beyond 1 year of age. A 5-fold increase in AAVrh10 dose to 2E14 gc/kg, in combination with HSCT, normalized neurological dysfunction up to 2 years of age. IV AAVrh10 alone resulted in an average survival to 41.2 weeks of age. In the peripheral nervous system, IV AAVrh10 alone or in addition to HSCT normalized nerve conduction velocity, improved ultrastructure, and normalized GALC enzyme activity and psychosine concentration. In the central nervous system, only combination therapy at the highest dose was able to restore galactosylceramidase activity and psychosine concentrations to within the normal range. These data have now guided clinical translation of systemic AAV gene therapy as an addition to HSCT (NCT04693598, NCT05739643).


Assuntos
Transplante de Células-Tronco Hematopoéticas , Leucodistrofia de Células Globoides , Cães , Animais , Leucodistrofia de Células Globoides/genética , Leucodistrofia de Células Globoides/terapia , Galactosilceramidase/genética , Psicosina , Transplante de Células-Tronco Hematopoéticas/métodos , Terapia Genética/métodos , Modelos Animais de Doenças
2.
J Immunol ; 207(1): 44-54, 2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34162727

RESUMO

Multiple sclerosis (MS) is an idiopathic demyelinating disease in which meningeal inflammation correlates with accelerated disease progression. The study of meningeal inflammation in MS has been limited because of constrained access to MS brain/spinal cord specimens and the lack of experimental models recapitulating progressive MS. Unlike induced models, a spontaneously occurring model would offer a unique opportunity to understand MS immunopathogenesis and provide a compelling framework for translational research. We propose granulomatous meningoencephalomyelitis (GME) as a natural model to study neuropathological aspects of MS. GME is an idiopathic, progressive neuroinflammatory disease of young dogs with a female bias. In the GME cases examined in this study, the meninges displayed focal and disseminated leptomeningeal enhancement on magnetic resonance imaging, which correlated with heavy leptomeningeal lymphocytic infiltration. These leptomeningeal infiltrates resembled tertiary lymphoid organs containing large B cell clusters that included few proliferating Ki67+ cells, plasma cells, follicular dendritic/reticular cells, and germinal center B cell-like cells. These B cell collections were confined in a specialized network of collagen fibers associated with the expression of the lympho-organogenic chemokines CXCL13 and CCL21. Although neuroparenchymal perivascular infiltrates contained B cells, they lacked the immune signature of aggregates in the meningeal compartment. Finally, meningeal B cell accumulation correlated significantly with cortical demyelination reflecting neuropathological similarities to MS. Hence, during chronic neuroinflammation, the meningeal microenvironment sustains B cell accumulation that is accompanied by underlying neuroparenchymal injury, indicating GME as a novel, naturally occurring model to study compartmentalized neuroinflammation and the associated pathology thought to contribute to progressive MS.


Assuntos
Linfócitos B/imunologia , Modelos Animais de Doenças , Meninges/imunologia , Esclerose Múltipla Crônica Progressiva/imunologia , Animais , Linfócitos B/patologia , Cães , Meninges/patologia , Esclerose Múltipla Crônica Progressiva/patologia
3.
Hum Mol Genet ; 28(R1): R119-R131, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31384936

RESUMO

Lysosomal storage diseases (LSDs) are a group of 70 monogenic disorders characterized by the lysosomal accumulation of a substrate. As a group, LSDs affect ~1 in 5000 live births; however, each individual storage disease is rare, limiting the ability to perform natural history studies or to perform clinical trials. Perhaps in no other biomedical field have naturally occurring large animal (canine, feline, ovine, caprine, and bovine) models been so essential for understanding the fundamentals of disease pathogenesis and for developing safe and effective therapies. These models were critical for the development of hematopoietic stem cell transplantation in α- and ß- mannosidosis, fucosidosis, and the mucopolysaccharidoses; enzyme replacement therapy for fucosidosis, the mucopolysaccharidoses, and neuronal ceroid lipofuscinosis; and small molecule therapy in Niemann-Pick type C disease. However, their most notable contributions to the biomedical field are in the development of gene therapy for LSDs. Adeno-associated viral vectors to treat nervous system disease have been evaluated in the large animal models of α-mannosidosis, globoid cell leukodystrophy, GM1 and GM2 gangliosidosis, the mucopolysaccharidoses, and neuronal ceroid lipofuscinosis. This review article will summarize the large animal models available for study as well as their contributions to the development of central and peripheral nervous system dysfunction in LSDs.


Assuntos
Modelos Animais de Doenças , Doenças por Armazenamento dos Lisossomos/complicações , Doenças por Armazenamento dos Lisossomos/genética , Doenças do Sistema Nervoso/etiologia , Doenças do Sistema Nervoso/terapia , Animais , Terapia Combinada , Gerenciamento Clínico , Terapia de Reposição de Enzimas , Terapia Genética , Transplante de Células-Tronco Hematopoéticas , Humanos , Doenças do Sistema Nervoso/diagnóstico , Resultado do Tratamento
4.
Brain ; 143(7): 2058-2072, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32671406

RESUMO

Intravascular injection of certain adeno-associated virus vector serotypes can cross the blood-brain barrier to deliver a gene into the CNS. However, gene distribution has been much more limited within the brains of large animals compared to rodents, rendering this approach suboptimal for treatment of the global brain lesions present in most human neurogenetic diseases. The most commonly used serotype in animal and human studies is 9, which also has the property of being transported via axonal pathways to distal neurons. A small number of other serotypes share this property, three of which were tested intravenously in mice compared to 9. Serotype hu.11 transduced fewer cells in the brain than 9, rh8 was similar to 9, but hu.32 mediated substantially greater transduction than the others throughout the mouse brain. To evaluate the potential for therapeutic application of the hu.32 serotype in a gyrencephalic brain of larger mammals, a hu.32 vector expressing the green fluorescent protein reporter gene was evaluated in the cat. Transduction was widely distributed in the cat brain, including in the cerebral cortex, an important target since mental retardation is an important component of many of the human neurogenetic diseases. The therapeutic potential of a hu.32 serotype vector was evaluated in the cat homologue of the human lysosomal storage disease alpha-mannosidosis, which has globally distributed lysosomal storage lesions in the brain. Treated alpha-mannosidosis cats had reduced severity of neurological signs and extended life spans compared to untreated cats. The extent of therapy was dose dependent and intra-arterial injection was more effective than intravenous delivery. Pre-mortem, non-invasive magnetic resonance spectroscopy and diffusion tensor imaging detected differences between the low and high doses, and showed normalization of grey and white matter imaging parameters at the higher dose. The imaging analysis was corroborated by post-mortem histological analysis, which showed reversal of histopathology throughout the brain with the high dose, intra-arterial treatment. The hu.32 serotype would appear to provide a significant advantage for effective treatment of the gyrencephalic brain by systemic adeno-associated virus delivery in human neurological diseases with widespread brain lesions.


Assuntos
Encéfalo/virologia , Dependovirus , Modelos Animais de Doenças , Terapia Genética/métodos , Vetores Genéticos , alfa-Manosidose/genética , Animais , Encéfalo/patologia , Gatos , Técnicas de Transferência de Genes , Transdução Genética
5.
Mol Cell Neurosci ; 102: 103451, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31794880

RESUMO

Globoid cell leukodystrophy (GLD, Krabbe disease, Krabbe's disease) is caused by genetic mutations in the gene encoding, galactosylceramidase (GALC). Deficiency of this enzyme results in central and peripheral nervous system pathology, and is characterized by loss of myelin and an infiltration of globoid cells. The canine model of GLD provides a translational model which faithfully recapitulates much of the human disease pathology. Targeted lipidomic analysis was conducted in serum and cerebrospinal fluid (CSF) over the lifetime of GLD affected and normal canines, and in brain tissue at humane endpoint to better understand disease progression and identify potential biomarkers of disease. Psychosine, a substrate of GALC and primary contributor to the pathology in GLD, was observed to be significantly elevated in the serum and CSF by 2 or 4 weeks of age, respectively, and steadily increased over the lifetime of affected animals. Importantly, psychosine concentration strongly correlated with disease severity. Galactosylceramide, glucosylceramide, and lactosylceramide were also found to be elevated in the CSF of affected animals and increased with age. Psychosine and galactosylceramide were found to be significantly increased in brain tissue at humane endpoint. This study identified several biomarkers which may be useful in the development of therapeutics for GLD.


Assuntos
Doenças do Cão/líquido cefalorraquidiano , Galactosilceramidas/sangue , Galactosilceramidas/líquido cefalorraquidiano , Leucodistrofia de Células Globoides/veterinária , Psicosina/líquido cefalorraquidiano , Animais , Biomarcadores/sangue , Biomarcadores/líquido cefalorraquidiano , Doenças do Cão/sangue , Doenças do Cão/patologia , Cães , Feminino , Leucodistrofia de Células Globoides/sangue , Leucodistrofia de Células Globoides/líquido cefalorraquidiano , Leucodistrofia de Células Globoides/patologia , Masculino , Psicosina/sangue
6.
Mol Genet Metab ; 129(4): 292-302, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32033912

RESUMO

Niemann-Pick type C (NPC) disease is a rare lysosomal storage disorder caused by mutations in either the NPC1 or the NPC2 gene. A new class of lipids, N-acyl-O-phosphocholineserines were recently identified as NPC biomarkers. The most abundant species in this class of lipid, N-palmitoyl-O-phosphocholineserine (PPCS), was evaluated for diagnosis of NPC disease and treatment efficacy assessment with 2-hydroxypropyl-ß-cyclodextrin (HPßCD) in NPC. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods were developed and validated to measure PPCS in human plasma and cerebrospinal fluid (CSF). A cutoff of 248 ng/mL in plasma provided a sensitivity of 100.0% and specificity of 96.6% in identifying NPC1 patients from control and NPC1 carrier subjects. PPCS was significantly elevated in CSF from NPC1 patients, and CSF PPCS levels were significantly correlated with NPC neurological disease severity scores. Plasma and CSF PPCS did not change significantly in response to intrathetical (IT) HPßCD treatment. In an intravenous (IV) HPßCD trial, plasma PPCS in all patients was significantly reduced. These results demonstrate that plasma PPCS was able to diagnose NPC1 patients with high sensitivity and specificity, and to evaluate the peripheral treatment efficacy of IV HPßCD treatment.


Assuntos
2-Hidroxipropil-beta-Ciclodextrina/uso terapêutico , Doença de Niemann-Pick Tipo C/diagnóstico , Doença de Niemann-Pick Tipo C/tratamento farmacológico , Fosforilcolina/sangue , Fosforilcolina/líquido cefalorraquidiano , Adolescente , Adulto , Idoso , Animais , Biomarcadores/sangue , Biomarcadores/líquido cefalorraquidiano , Gatos , Criança , Pré-Escolar , Cromatografia Líquida , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Índice de Gravidade de Doença , Espectrometria de Massas em Tandem , Resultado do Tratamento , Adulto Jovem
7.
J Inherit Metab Dis ; 43(3): 618-634, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31707730

RESUMO

2-Hydroxypropyl-ß-cyclodextrin (HP-ß-CD) is an experimental therapy for Niemann-Pick disease type C (NPC) that reduced neuronal cholesterol and ganglioside storage, reduced Purkinje cell death, and increased lifespan in npc1-/- mice and NPC1 cats. In this study, tissue distribution was investigated in normal cats that received a single 120-mg dose of [14 C]-HP-ß-CD (approximately 200 µCi/cat) via the cerebellomedullary cistern (CBMC) and lumbar cistern. One cat was euthanized at each of various time points up to 24 hours postdose for subsequent processing and quantitative whole-body autoradiographic analysis. HP-ß-CD-derived radioactivity absorbed from the CBMC was widely distributed to cat tissues; most tissues were observed to have reached their highest concentration at 1 hour postdose. HP-ß-CD-derived radioactivity penetrated into the deeper parts of the central nervous system with the highest concentration at 4 hours (403 µg Eq/g or 0.28 mM) and remained high (49.7 µg Eq/g or 0.03 mM) at 24 hours. The relatively long half-life (11-30 hours) in cerebral ventricles and the subarachnoid space surrounding the brain and spinal cord might contribute to the efficacy of HP-ß-CD in NPC1 cats. Other tissues with high concentrations of radioactivity were nasal turbinates, pituitary gland, and urinary bladder, while relatively low concentrations were observed in blood and bile.


Assuntos
2-Hidroxipropil-beta-Ciclodextrina/farmacologia , 2-Hidroxipropil-beta-Ciclodextrina/farmacocinética , Proteína C1 de Niemann-Pick/genética , Doença de Niemann-Pick Tipo C/tratamento farmacológico , Animais , Gatos , Colesterol/metabolismo , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Doença de Niemann-Pick Tipo C/metabolismo
8.
J Lipid Res ; 60(8): 1410-1424, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31201291

RESUMO

Niemann-Pick disease type C1 (NPC1) is a fatal, neurodegenerative, cholesterol storage disorder. With new therapeutics in clinical trials, there is an urgency to improve diagnostics and monitor therapeutic efficacy with biomarkers. In this study, we sought to define the structure of an unknown lipid biomarker for NPC1 with [M + H]+ ion at m/z 509.3351, previously designated as lysoSM-509. The structure of N-palmitoyl-O-phosphocholineserine (PPCS) was proposed for the lipid biomarker based on the results from mass spectrometric analyses and chemical derivatizations. As no commercial standard is available, authentic PPCS was chemically synthesized, and the structure was confirmed by comparison of endogenous and synthetic compounds as well as their derivatives using liquid chromatography-tandem mass spectrometry (LC-MS/MS). PPCS is the most abundant species among N-acyl-O-phosphocholineserines (APCS), a class of lipids that have not been previously detected in biological samples. Further analysis demonstrated that all APCS species with acyl groups ranging from C14 to C24 were elevated in NPC1 plasma. PPCS is also elevated in both central and peripheral tissues of the NPC1 cat model. Identification of APCS structures provide an opportunity for broader exploration of the roles of these novel lipids in NPC1 disease pathology and diagnosis.


Assuntos
Doença de Niemann-Pick Tipo C/metabolismo , Fosforilcolina/metabolismo , Animais , Biomarcadores/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Doença de Niemann-Pick Tipo C/genética
9.
Lancet ; 390(10104): 1758-1768, 2017 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-28803710

RESUMO

BACKGROUND: Niemann-Pick disease, type C1 (NPC1) is a lysosomal storage disorder characterised by progressive neurodegeneration. In preclinical testing, 2-hydroxypropyl-ß-cyclodextrins (HPßCD) significantly delayed cerebellar Purkinje cell loss, slowed progression of neurological manifestations, and increased lifespan in mouse and cat models of NPC1. The aim of this study was to assess the safety and efficacy of lumbar intrathecal HPßCD. METHODS: In this open-label, dose-escalation phase 1-2a study, we gave monthly intrathecal HPßCD to participants with NPC1 with neurological manifestation at the National Institutes of Health (NIH), Bethesda, MD, USA. To explore the potential effect of 2-week dosing, three additional participants were enrolled in a parallel study at Rush University Medical Center (RUMC), Chicago, IL, USA. Participants from the NIH were non-randomly, sequentially assigned in cohorts of three to receive monthly initial intrathecal HPßCD at doses of 50, 200, 300, or 400 mg per month. A fifth cohort of two participants received initial doses of 900 mg. Participants from RUMC initially received 200 or 400 mg every 2 weeks. The dose was escalated based on tolerance or safety data from higher dose cohorts. Serum and CSF 24(S)-hydroxycholesterol (24[S]-HC), which serves as a biomarker of target engagement, and CSF protein biomarkers were evaluated. NPC Neurological Severity Scores (NNSS) were used to compare disease progression in HPßCD-treated participants relative to a historical comparison cohort of 21 NPC1 participants of similar age range. FINDINGS: Between Sept 21, 2013, and Jan 19, 2015, 32 participants with NPC1 were assessed for eligibility at the National Institutes of Health. 18 patients were excluded due to inclusion criteria not met (six patients), declined to participate (three patients), pursued independent expanded access and obtained the drug outside of the study (three patients), enrolled in the RUMC cohort (one patient), or too late for the trial enrolment (five patients). 14 patients were enrolled and sequentially assigned to receive intrathecal HPßCD at a starting dose of 50 mg per month (three patients), 200 mg per month (three patients), 300 mg per month (three patients), 400 mg per month (three patients), or 900 mg per month (two patients). During the first year, two patients had treatment interrupted for one dose, based on grade 1 ototoxicity. All 14 patients were assessed at 12 months. Between 12 and 18 months, one participant had treatment interrupted at 17 months due to hepatocellular carcinoma, one patient had dose interruption for 2 doses based on caregiver hardship and one patient had treatment interrupted for 1 dose for mastoiditis. 11 patients were assessed at 18 months. Between Dec 11, 2013, and June 25, 2014, three participants were assessed for eligibility and enrolled at RUMC, and were assigned to receive intrathecal HPßCD at a starting dose of 200 mg every 2 weeks (two patients), or 400 mg every two weeks (one patient). There were no dropouts in this group and all 3 patients were assessed at 18 months. Biomarker studies were consistent with improved neuronal cholesterol homoeostasis and decreased neuronal pathology. Post-drug plasma 24(S)-HC area under the curve (AUC8-72) values, an indicator of neuronal cholesterol homoeostasis, were significantly higher than post-saline plasma 24(S)-HC AUC8-72 after doses of 900 mg (p=0·0063) and 1200 mg (p=0·0037). CSF 24(S)-HC concentrations in three participants given either 600 or 900 mg of HPßCD were increased about two fold (p=0·0032) after drug administration. No drug-related serious adverse events were observed. Mid-frequency to high-frequency hearing loss, an expected adverse event, was documented in all participants. When managed with hearing aids, this did not have an appreciable effect on daily communication. The NNSS for the 14 participants treated monthly increased at a rate of 1·22, SEM 0·34 points per year compared with 2·92, SEM 0·27 points per year (p=0·0002) for the 21 patient comparison group. Decreased progression was observed for NNSS domains of ambulation (p=0·0622), cognition (p=0·0040) and speech (p=0·0423). INTERPRETATION: Patients with NPC1 treated with intrathecal HPßCD had slowed disease progression with an acceptable safety profile. These data support the initiation of a multinational, randomised, controlled trial of intrathecal HPßCD. FUNDING: National Institutes of Health, Dana's Angels Research Trust, Ara Parseghian Medical Research Foundation, Hope for Haley, Samantha's Search for the Cure Foundation, National Niemann-Pick Disease Foundation, Support of Accelerated Research for NPC Disease, Vtesse, Janssen Research and Development, a Johnson & Johnson company, and Johnson & Johnson.


Assuntos
2-Hidroxipropil-beta-Ciclodextrina/administração & dosagem , Progressão da Doença , Doença de Niemann-Pick Tipo C/tratamento farmacológico , 2-Hidroxipropil-beta-Ciclodextrina/efeitos adversos , Adolescente , Biomarcadores/sangue , Biomarcadores/líquido cefalorraquidiano , Calbindinas/líquido cefalorraquidiano , Criança , Pré-Escolar , Relação Dose-Resposta a Droga , Proteína 3 Ligante de Ácido Graxo/líquido cefalorraquidiano , Feminino , Perda Auditiva de Alta Frequência/induzido quimicamente , Humanos , Hidroxicolesteróis/sangue , Hidroxicolesteróis/líquido cefalorraquidiano , Injeções Espinhais , Masculino , Doença de Niemann-Pick Tipo C/sangue , Doença de Niemann-Pick Tipo C/líquido cefalorraquidiano , Doenças Raras/tratamento farmacológico , Adulto Jovem
10.
Mol Ther ; 24(1): 26-33, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26354342

RESUMO

Lysosomal storage diseases (LSDs) are debilitating neurometabolic disorders for most of which long-term effective therapies have not been developed. Gene therapy is a potential treatment but a critical barrier to treating the brain is the need for global correction. We tested the efficacy of cisterna magna infusion of adeno-associated virus type 1 (AAV1) expressing feline alpha-mannosidase gene in the postsymptomatic alpha-mannosidosis (AMD) cat, a homologue of the human disease. Lysosomal alpha-mannosidase (MANB) activity in the cerebrospinal fluid (CSF) and serum were increased above the control values in untreated AMD cats. Clinical neurological signs were delayed in onset and reduced in severity. The lifespan of the treated cats was significantly extended. Postmortem histopathology showed resolution of lysosomal storage lesions throughout the brain. MANB activity in brain tissue was significantly above the levels of untreated tissues. The results demonstrate that a single cisterna magna injection of AAV1 into the CSF can mediate widespread neuronal transduction of the brain and meaningful clinical improvement. Thus, cisterna magna gene delivery by AAV1 appears to be a viable strategy for treatment of the whole brain in AMD and should be applicable to many of the neurotropic LSDs as well as other neurogenetic disorders.


Assuntos
Doenças do Gato/terapia , Cisterna Magna/metabolismo , Dependovirus/genética , alfa-Manosidase/genética , alfa-Manosidose/veterinária , Idade de Início , Animais , Encéfalo/enzimologia , Doenças do Gato/patologia , Gatos , Modelos Animais de Doenças , Terapia Genética , Vetores Genéticos/administração & dosagem , Humanos , Injeções , Lisossomos/metabolismo , alfa-Manosidase/sangue , alfa-Manosidase/líquido cefalorraquidiano , alfa-Manosidase/metabolismo , alfa-Manosidose/patologia , alfa-Manosidose/terapia
11.
Mol Ther ; 24(2): 206-216, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26447927

RESUMO

Mucopolysaccharidosis VII (MPS VII) is a lysosomal storage disease arising from mutations in ß-d-glucuronidase (GUSB), which results in glycosaminoglycan (GAG) accumulation and a variety of clinical manifestations including neurological disease. Herein, MPS VII dogs were injected intravenously (i.v.) and/or intrathecally (i.t.) via the cisterna magna with AAV9 or AAVrh10 vectors carrying the canine GUSB cDNA. Although i.v. injection alone at 3 days of age resulted in normal cerebrospinal fluid (CSF) GUSB activity, brain tissue homogenates had only ~1 to 6% normal GUSB activity and continued to have elevated GAG storage. In contrast, i.t. injection at 3 weeks of age resulted in CSF GUSB activity 44-fold normal while brain tissue homogenates had >100% normal GUSB activity and reduced GAGs compared with untreated dogs. Markers for secondary storage and inflammation were eliminated in i.t.-treated dogs and reduced in i.v.-treated dogs compared with untreated dogs. Given that i.t.-treated dogs expressed higher levels of GUSB in the CNS tissues compared to those treated i.v., we conclude that i.t. injection of AAV9 or AAVrh10 vectors is more effective than i.v. injection alone in the large animal model of MPS VII.


Assuntos
Doenças do Sistema Nervoso Central/terapia , Terapia Genética/métodos , Glucuronidase/genética , Mucopolissacaridose VII/terapia , Animais , Animais Recém-Nascidos , Doenças do Sistema Nervoso Central/genética , Doenças do Sistema Nervoso Central/metabolismo , Dependovirus/genética , Modelos Animais de Doenças , Cães , Vetores Genéticos/administração & dosagem , Glucuronidase/líquido cefalorraquidiano , Glicosaminoglicanos/metabolismo , Injeções Intravenosas , Injeções Espinhais , Masculino , Mucopolissacaridose VII/complicações , Mucopolissacaridose VII/genética , Mucopolissacaridose VII/metabolismo
12.
Yale J Biol Med ; 90(3): 417-431, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28955181

RESUMO

For many lethal or debilitating genetic disorders in patients there are no satisfactory therapies. Several barriers exist that hinder the developments of effective therapies including the limited availability of clinically relevant animal models that faithfully recapitulate human genetic disease. In 1974, the Referral Center for Animal Models of Human Genetic Disease (RCAM) was established by Dr. Donald F. Patterson and continued by Dr. Mark E. Haskins at the University of Pennsylvania with the mission to discover, understand, treat, and maintain breeding colonies of naturally occurring hereditary disorders in dogs and cats that are orthologous to those found in human patients. Although non-human primates, sheep, and pig models are also available within the medical community, naturally occurring diseases are rarely identified in non-human primates, and the vast behavioral, clinicopathological, physiological, and anatomical knowledge available regarding dogs and cats far surpasses what is available in ovine and porcine species. The canine and feline models that are maintained at RCAM are presented here with a focus on preclinical therapy data. Clinical studies that have been generated from preclinical work in these models are also presented.


Assuntos
Doenças Genéticas Inatas , Doenças Raras , Animais , Gatos , Modelos Animais de Doenças , Cães , Humanos , Ovinos , Suínos
13.
Hum Mol Genet ; 23(22): 6022-33, 2014 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-24964810

RESUMO

Niemann-Pick C1 (NPC1) disease is a rare, neurodegenerative lysosomal cholesterol storage disorder, typified by progressive cognitive and motor function impairment. Affected individuals usually succumb to the disease in adolescence. 2-Hydroxypropyl-ß-cyclodextrin (HP-ß-CD) has emerged as a promising intervention that reduces lipid storage and prolongs survival in NPC1 disease animal models. A barrier to the development of HP-ß-CD and other treatments for NPC disease has been the lack of validated biochemical measures to evaluate efficacy. Here we explored whether cholesterol homeostatic responses resulting from HP-ß-CD-mediated redistribution of sequestered lysosomal cholesterol could provide biomarkers to monitor treatment. Upon direct CNS delivery of HP-ß-CD, we found increases in plasma 24(S)-HC in two independent NPC1 disease animal models, findings that were confirmed in human NPC1 subjects receiving HP-ß-CD. Since circulating 24(S)-HC is almost exclusively CNS-derived, the increase in plasma 24(S)-HC provides a peripheral, non-invasive measure of the CNS effect of HP-ß-CD. Our findings suggest that plasma 24(S)-HC, along with the other cholesterol-derived markers examined in this study, can serve as biomarkers that will accelerate development of therapeutics for NPC1 disease.


Assuntos
Colesterol/sangue , Doença de Niemann-Pick Tipo C/tratamento farmacológico , beta-Ciclodextrinas/administração & dosagem , 2-Hidroxipropil-beta-Ciclodextrina , Adolescente , Animais , Biomarcadores/sangue , Criança , Modelos Animais de Doenças , Monitoramento de Medicamentos/métodos , Feminino , Homeostase , Humanos , Masculino , Camundongos Endogâmicos BALB C , Doença de Niemann-Pick Tipo C/sangue , Adulto Jovem
14.
J Neurosci Res ; 94(11): 1007-17, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27638585

RESUMO

Globoid cell leukodystrophy (GLD), or Krabbe's disease, is a debilitating and always fatal pediatric neurodegenerative disease caused by a mutation in the gene encoding the hydrolytic enzyme galactosylceramidase (GALC). In the absence of GALC, progressive loss of myelin and accumulation of a neurotoxic substrate lead to incapacitating loss of motor and cognitive function and death, typically by 2 years of age. Currently, there is no cure. Recent convincing evidence of the therapeutic potential of combining gene and cell therapies in the murine model of GLD has accelerated the requirement for validated markers of disease to evaluate therapeutic efficacy. Here we demonstrate clinically relevant and quantifiable measures of central (CNS) and peripheral (PNS) nervous system disease progression in the naturally occurring canine model of GLD. As measured by brainstem auditory-evoked response testing, GLD dogs demonstrated a significant increase in I-V interpeak latency and hearing threshold at all time points. Motor nerve conduction velocities (NCVs) in GLD dogs were significantly lower than normal by 12-16 weeks of age, and sensory NCV was significantly lower than normal by 8-12 weeks of age, serving as a sensitive indicator of peripheral nerve dysfunction. Post-mortem histological evaluations confirmed neuroimaging and electrodiagnostic assessments and detailed loss of myelin and accumulation of storage product in the CNS and the PNS. Additionally, cerebrospinal fluid psychosine concentrations were significantly elevated in GLD dogs, demonstrating potential as a biochemical marker of disease. These data demonstrate that CNS and PNS disease progression can be quantified over time in the canine model of GLD with tools identical to those used to assess human patients. © 2016 Wiley Periodicals, Inc.


Assuntos
Potenciais Evocados Auditivos do Tronco Encefálico/genética , Leucodistrofia de Células Globoides/complicações , Leucodistrofia de Células Globoides/genética , Doenças do Sistema Nervoso , Animais , Modelos Animais de Doenças , Cães , Estimulação Elétrica , Feminino , Galactosilceramidase/genética , Humanos , Leucodistrofia de Células Globoides/diagnóstico por imagem , Leucodistrofia de Células Globoides/veterinária , Imageamento por Ressonância Magnética , Masculino , Mutação de Sentido Incorreto/genética , Sistema Nervoso/diagnóstico por imagem , Sistema Nervoso/patologia , Sistema Nervoso/fisiopatologia , Doenças do Sistema Nervoso/diagnóstico por imagem , Doenças do Sistema Nervoso/etiologia , Doenças do Sistema Nervoso/metabolismo , Doenças do Sistema Nervoso/terapia , Condução Nervosa/genética , Psicosina/líquido cefalorraquidiano
15.
Epilepsia ; 57(12): 1949-1957, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27807850

RESUMO

OBJECTIVE: Epilepsy is a chronic disorder, but seizure recordings are usually obtained in the acute setting. The chronic behavior of seizures and the interictal bursts that sometimes initiate them is unknown. We investigate the variability of these electrographic patterns over an extended period of time using chronic intracranial recordings in canine epilepsy. METHODS: Continuous, yearlong intracranial electroencephalography (iEEG) recordings from four dogs with naturally occurring epilepsy were analyzed for seizures and interictal bursts. Following automated detection and clinician verification of interictal bursts and seizures, temporal trends of seizures, burst count, and burst-burst similarities were determined. One dog developed status epilepticus, the recordings of which were also investigated. RESULTS: Multiple seizure types, determined by onset channels, were observed in each dog, with significant temporal variation between types. The first 14 days of invasive recording, analogous to the average duration of clinical invasive recordings in humans, did not capture the entirety of seizure types. Seizures typically occurred in clusters, and isolated seizures were rare. The count and dynamics of interictal bursts form distinct groups and do not stabilize until several weeks after implantation. SIGNIFICANCE: There is significant temporal variability in seizures and interictal bursts after electrode implantation that requires several weeks to reach steady state. These findings, comparable to those reported in humans implanted with the NeuroPace Responsive Neurostimulator System (RNS) device, suggest that transient network changes following electrode implantation may need to be taken into account when interpreting or analyzing iEEG during evaluation for epilepsy surgery. Chronic, ambulatory iEEG may be better suited to accurately map epileptic networks in appropriate individuals.


Assuntos
Ondas Encefálicas/fisiologia , Encéfalo/fisiopatologia , Epilepsia/fisiopatologia , Epilepsia/veterinária , Animais , Cães , Eletrodos Implantados , Eletroencefalografia , Feminino , Estudos Longitudinais , Masculino
16.
Epilepsia ; 56(6): 882-7, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25953073

RESUMO

OBJECTIVES: There are a limited number of marketed intravenous antiepileptic drugs (AEDs) available to treat status epilepticus (SE). All were first developed for chronic therapy of epilepsy, not specifically for SE. Epilepsy and canine SE (CSE) occur naturally in dogs, with prevalence, presentation, and percentage of refractory cases similar to human epilepsy. The objective of this study was to determine if CSE treated with fosphenytoin (FOS) results in a similar responder rate as for people. METHODS: A randomized clinical trial was performed for dogs with CSE. Dogs who presented during a seizure or who had additional seizures after enrolling received intravenous (i.v.) benzodiazepine (BZD) followed immediately by intravenous infusion of 15 mg/kg phenytoin equivalent (PE) of fosphenytoin (FOS) or saline placebo (PBO). If seizures continued, additional AEDs were administered per the standard of care for veterinary patients. Total and unbound plasma phenytoin (PHT) concentrations were measured. RESULTS: Consent was obtained for 50 dogs with CSE. Thirty-one had additional motor seizures and were randomized to the study intervention (22 FOS and 9 PBO). There was a statistically significant difference in the 12 h responder rate, with 63% in the FOS group versus 22% in the placebo group (p = 0.043) having no further seizures. The unbound PHT concentrations at 30 and 60 min were within the therapeutic concentrations for people (1-2 µg/ml) with the exception of one dog. There was mild vomiting in 36% of the FOS group (7/22) within 20 min of FOS administration and none of the placebo group (0/9) (p = 0.064). SIGNIFICANCE: This proof of concept study provides the first evidence that FOS is tolerated and effective in canine SE at PHT concentrations clinically relevant for human SE. Furthermore, naturally occurring CSE can be utilized as a translational platform for future studies of novel SE compounds.


Assuntos
Anticonvulsivantes/uso terapêutico , Fenitoína/análogos & derivados , Estado Epiléptico/tratamento farmacológico , Estado Epiléptico/veterinária , Animais , Benzodiazepinas/administração & dosagem , Cães , Método Duplo-Cego , Feminino , Infusões Intravenosas , Masculino , Fenitoína/sangue , Fenitoína/uso terapêutico , Estudos Prospectivos , Estado Epiléptico/sangue , Fatores de Tempo , Resultado do Tratamento
17.
Gene ; 893: 147941, 2024 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-37913889

RESUMO

A 6-month-old cat of unknown ancestry presented for a neurologic evaluation due to progressive motor impairment. Complete physical and neurologic examinations suggested the disorder was likely to be hereditary, although the signs were not consistent with any previously described inherited disorders in cats. Due to the progression of disease signs including severely impaired motor function and cognitive decline, the cat was euthanized at approximately 10.5 months of age. Whole genome sequence analysis identified a homozygous missense variant c.2506G > A in MANBA that predicts a p.Gly836Arg alteration in the encoded lysosomal enzyme ß -mannosidase. This variant was not present in the whole genome or whole exome sequences of any of the 424 cats represented in the 99 Lives Cat Genome dataset. ß -Mannosidase enzyme activity was undetectable in brain tissue homogenates from the affected cat, whereas α-mannosidase enzyme activities were elevated compared to an unaffected cat. Postmortem examination of brain and retinal tissues revealed massive accumulations of vacuolar inclusions in most cells, similar to those reported in animals of other species with hereditary ß -mannosidosis. Based on these findings, the cat likely suffered from ß -mannosidosis due to the abolition of ß -mannosidase activity associated with the p.Gly836Arg amino acid substitution. p.Gly836 is located in the C-terminal region of the protein and was not previously known to be involved in modulating enzyme activity. In addition to the vacuolar inclusions, some cells in the brain of the affected cat contained inclusions that exhibited lipofuscin-like autofluorescence. Electron microscopic examinations suggested these inclusions formed via an autophagy-like process.


Assuntos
beta-Manosidose , Gatos , Animais , beta-Manosidose/complicações , beta-Manosidose/diagnóstico , beta-Manosidose/genética , beta-Manosidase/genética , beta-Manosidase/metabolismo , Mutação de Sentido Incorreto
18.
Mol Ther Methods Clin Dev ; 32(2): 101272, 2024 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-38946937

RESUMO

Alpha-mannosidosis is caused by a genetic deficiency of lysosomal alpha-mannosidase, leading to the widespread presence of storage lesions in the brain and other tissues. Enzyme replacement therapy is available but is not approved for treating the CNS, since the enzyme does not penetrate the blood-brain barrier. However, intellectual disability is a major manifestation of the disease; thus, a complimentary treatment is needed. While enzyme replacement therapy into the brain is technically feasible, it requires ports and frequent administration over time that are difficult to manage medically. Infusion of adeno-associated viral vectors into the cerebrospinal fluid is an attractive route for broadly targeting brain cells. We demonstrate here the widespread post-symptomatic correction of the globally distributed storage lesions by infusion of a high dose of AAV1-feline alpha-mannosidase (fMANB) into the CSF via the cisterna magna in the gyrencephalic alpha-mannosidosis cat brain. Significant improvements in clinical parameters occurred, and widespread global correction was documented pre-mortem by non-invasive magnetic resonance imaging. Postmortem analysis demonstrated high levels of MANB activity and reversal of lysosomal storage lesions throughout the brain. Thus, CSF treatment by adeno-associated viral vector gene therapy appears to be a suitable complement to systemic enzyme replacement therapy to potentially treat the whole patient.

19.
Hum Gene Ther ; 34(3-4): 94-111, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-36606687

RESUMO

Adeno-associated virus (AAV)-based gene therapies, exemplified by the approved therapy for spinal muscular atrophy, have the potential to deliver disease-course-altering treatments for central nervous system (CNS) indications. However, several clinical trials have reported severe adverse events, including patient deaths following high-dose systemic administration for muscle-directed gene transfer, highlighting the need to explore approaches utilizing lower doses when targeting the CNS. Animal models of disease provide insight into the response to new AAV therapies. However, translation from small to larger animals and eventually to humans is hampered by anatomical and biological differences across the species and their impact on AAV delivery. We performed a literature review of preclinical studies of AAV gene therapy biodistribution following cerebrospinal fluid (CSF) delivery (intracerebroventricular, intra-cisterna magna, and intrathecal lumbar). The reviewed literature varies greatly in the reported biodistribution of AAV following administration into the CSF. Differences between studies, including animal model, vector serotype used, method used to assess biodistribution, and route of administration, among other variables, contribute to differing outcomes and difficulties in translating these preclinical results. For example, only half of the published AAV-based gene therapy studies report vector copy number, the most direct readout following administration of a vector; none of these studies reported details such as the empty:full capsid ratio and quality of encapsidated genome. Analysis of the last decade's literature focusing on AAV-based gene therapies targeting the CNS underscores limitations of the body of knowledge and room for continued research. In particular, there is a need to understand the biodistribution achieved by different CSF-directed routes of administration and determining if specific cell types/structures of interest will be transduced. Our findings point to a clear need for a more systematic approach across the field to align the assessments and elements reported in preclinical research to enable more reliable translation across animal models and into human studies.


Assuntos
Dependovirus , Terapia Genética , Animais , Humanos , Dependovirus/genética , Distribuição Tecidual , Terapia Genética/métodos , Sistema Nervoso Central , Modelos Animais , Vetores Genéticos/genética , Técnicas de Transferência de Genes
20.
Genes (Basel) ; 14(9)2023 08 31.
Artigo em Inglês | MEDLINE | ID: mdl-37761886

RESUMO

A 7-month-old Doberman Pinscher dog presented with progressive neurological signs and brain atrophy suggestive of a hereditary neurodegenerative disorder. The dog was euthanized due to the progression of disease signs. Microscopic examination of tissues collected at the time of euthanasia revealed massive accumulations of vacuolar inclusions in cells throughout the central nervous system, suggestive of a lysosomal storage disorder. A whole genome sequence generated with DNA from the affected dog contained a likely causal, homozygous missense variant in MAN2B1 that predicted an Asp104Gly amino acid substitution that was unique among whole genome sequences from over 4000 dogs. A lack of detectable α-mannosidase enzyme activity confirmed a diagnosis of a-mannosidosis. In addition to the vacuolar inclusions characteristic of α-mannosidosis, the dog exhibited accumulations of autofluorescent intracellular inclusions in some of the same tissues. The autofluorescence was similar to that which occurs in a group of lysosomal storage disorders called neuronal ceroid lipofuscinoses (NCLs). As in many of the NCLs, some of the storage bodies immunostained strongly for mitochondrial ATP synthase subunit c protein. This protein is not a substrate for α-mannosidase, so its accumulation and the development of storage body autofluorescence were likely due to a generalized impairment of lysosomal function secondary to the accumulation of α-mannosidase substrates. Thus, it appears that storage body autofluorescence and subunit c accumulation are not unique to the NCLs. Consistent with generalized lysosomal impairment, the affected dog exhibited accumulations of intracellular inclusions with varied and complex ultrastructural features characteristic of autophagolysosomes. Impaired autophagic flux may be a general feature of this class of disorders that contributes to disease pathology and could be a target for therapeutic intervention. In addition to storage body accumulation, glial activation indicative of neuroinflammation was observed in the brain and spinal cord of the proband.


Assuntos
Doenças por Armazenamento dos Lisossomos , alfa-Manosidose , Animais , Cães , alfa-Manosidase/genética , alfa-Manosidose/genética , alfa-Manosidose/veterinária , Doenças por Armazenamento dos Lisossomos/genética , Doenças por Armazenamento dos Lisossomos/veterinária , Lisossomos , Mutação de Sentido Incorreto , Vacúolos , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/veterinária
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