Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 6 de 6
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
J Clin Invest ; 129(11): 4817-4831, 2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-31589162

RESUMO

BACKGROUNDSpinal muscular atrophy (SMA) is caused by deficient expression of survival motor neuron (SMN) protein. New SMN-enhancing therapeutics are associated with variable clinical benefits. Limited knowledge of baseline and drug-induced SMN levels in disease-relevant tissues hinders efforts to optimize these treatments.METHODSSMN mRNA and protein levels were quantified in human tissues isolated during expedited autopsies.RESULTSSMN protein expression varied broadly among prenatal control spinal cord samples, but was restricted at relatively low levels in controls and SMA patients after 3 months of life. A 2.3-fold perinatal decrease in median SMN protein levels was not paralleled by comparable changes in SMN mRNA. In tissues isolated from nusinersen-treated SMA patients, antisense oligonucleotide (ASO) concentration and full-length (exon 7 including) SMN2 (SMN2-FL) mRNA level increases were highest in lumbar and thoracic spinal cord. An increased number of cells showed SMN immunolabeling in spinal cord of treated patients, but was not associated with an increase in whole-tissue SMN protein levels.CONCLUSIONSA normally occurring perinatal decrease in whole-tissue SMN protein levels supports efforts to initiate SMN-inducing therapies as soon after birth as possible. Limited ASO distribution to rostral spinal and brain regions in some patients likely limits clinical response of motor units in these regions for those patients. These results have important implications for optimizing treatment of SMA patients and warrant further investigations to enhance bioavailability of intrathecally administered ASOs.FUNDINGSMA Foundation, SMART, NIH (R01-NS096770, R01-NS062869), Ionis Pharmaceuticals, and PTC Therapeutics. Biogen provided support for absolute real-time RT-PCR.


Assuntos
Envelhecimento , Neurônios Motores , Atrofia Muscular Espinal , Oligodesoxirribonucleotídeos Antissenso/administração & dosagem , Medula Espinal , Envelhecimento/genética , Envelhecimento/metabolismo , Envelhecimento/patologia , Autopsia , Sobrevivência Celular , Feminino , Humanos , Masculino , Neurônios Motores/metabolismo , Neurônios Motores/patologia , Atrofia Muscular Espinal/tratamento farmacológico , Atrofia Muscular Espinal/genética , Atrofia Muscular Espinal/metabolismo , Atrofia Muscular Espinal/patologia , Medula Espinal/metabolismo , Medula Espinal/patologia , Proteína 2 de Sobrevivência do Neurônio Motor/antagonistas & inibidores , Proteína 2 de Sobrevivência do Neurônio Motor/genética , Proteína 2 de Sobrevivência do Neurônio Motor/metabolismo
2.
Int J Mol Sci ; 19(1)2018 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-29316633

RESUMO

Spinal muscular atrophy (SMA) is an autosomal-recessive childhood motor neuron disease and the main genetic cause of infant mortality. SMA is caused by deletions or mutations in the survival motor neuron 1 (SMN1) gene, which results in SMN protein deficiency. Only one approved drug has recently become available and allows for the correction of aberrant splicing of the paralogous SMN2 gene by antisense oligonucleotides (ASOs), leading to production of full-length SMN protein. We have already demonstrated that a sequence of an ASO variant, Morpholino (MO), is particularly suitable because of its safety and efficacy profile and is both able to increase SMN levels and rescue the murine SMA phenotype. Here, we optimized this strategy by testing the efficacy of four new MO sequences targeting SMN2. Two out of the four new MO sequences showed better efficacy in terms of SMN protein production both in SMA induced pluripotent stem cells (iPSCs) and SMAΔ7 mice. Further, the effect was enhanced when different MO sequences were administered in combination. Our data provide an important insight for MO-based treatment for SMA. Optimization of the target sequence and validation of a treatment based on a combination of different MO sequences could support further pre-clinical studies and the progression toward future clinical trials.


Assuntos
Oligonucleotídeos Antissenso/metabolismo , Animais , Encéfalo/metabolismo , Células HeLa , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Camundongos , Camundongos Transgênicos , Neurônios Motores/metabolismo , Atrofia Muscular Espinal/tratamento farmacológico , Atrofia Muscular Espinal/metabolismo , Atrofia Muscular Espinal/patologia , Oligonucleotídeos Antissenso/uso terapêutico , Medula Espinal/metabolismo , Proteína 2 de Sobrevivência do Neurônio Motor/antagonistas & inibidores , Proteína 2 de Sobrevivência do Neurônio Motor/genética , Proteína 2 de Sobrevivência do Neurônio Motor/metabolismo , Regulação para Cima
3.
Nucleic Acids Res ; 45(1): 395-416, 2017 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-27557711

RESUMO

Spinal Muscular Atrophy (SMA) is a neuromuscular disorder caused by insufficient levels of the Survival of Motor Neuron (SMN) protein. SMN is expressed ubiquitously and functions in RNA processing pathways that include trafficking of mRNA and assembly of snRNP complexes. Importantly, SMA severity is correlated with decreased snRNP assembly activity. In particular, the minor spliceosomal snRNPs are affected, and some U12-dependent introns have been reported to be aberrantly spliced in patient cells and animal models. SMA is characterized by loss of motor neurons, but the underlying mechanism is largely unknown. It is likely that aberrant splicing of genes expressed in motor neurons is involved in SMA pathogenesis, but increasing evidence indicates that pathologies also exist in other tissues. We present here a comprehensive RNA-seq study that covers multiple tissues in an SMA mouse model. We show elevated U12-intron retention in all examined tissues from SMA mice, and that U12-dependent intron retention is induced upon siRNA knock-down of SMN in HeLa cells. Furthermore, we show that retention of U12-dependent introns is mitigated by ASO treatment of SMA mice and that many transcriptional changes are reversed. Finally, we report on missplicing of several Ca2+ channel genes that may explain disrupted Ca2+ homeostasis in SMA and activation of Cdk5.


Assuntos
Íntrons , Atrofia Muscular Espinal/genética , Splicing de RNA , RNA Mensageiro/genética , Ribonucleoproteínas Nucleares Pequenas/genética , Proteína 1 de Sobrevivência do Neurônio Motor/genética , Animais , Cálcio/metabolismo , Canais de Cálcio/deficiência , Canais de Cálcio/genética , Modelos Animais de Doenças , Feminino , Células HeLa , Humanos , Masculino , Camundongos , Neurônios Motores/metabolismo , Neurônios Motores/patologia , Atrofia Muscular Espinal/metabolismo , Atrofia Muscular Espinal/patologia , Atrofia Muscular Espinal/terapia , Oligonucleotídeos Antissenso/administração & dosagem , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo , RNA Mensageiro/metabolismo , Ribonucleoproteínas Nucleares Pequenas/metabolismo , Análise de Sequência de RNA , Medula Espinal/metabolismo , Medula Espinal/patologia , Proteína 1 de Sobrevivência do Neurônio Motor/antagonistas & inibidores , Proteína 1 de Sobrevivência do Neurônio Motor/metabolismo , Proteína 2 de Sobrevivência do Neurônio Motor/antagonistas & inibidores , Proteína 2 de Sobrevivência do Neurônio Motor/genética , Proteína 2 de Sobrevivência do Neurônio Motor/metabolismo
4.
Mol Ther ; 22(4): 854-61, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24317636

RESUMO

Spinal muscular atrophy (SMA) is a fatal autosomal recessive disease caused by survival motor neuron (SMN) protein insufficiency due to SMN1 mutations. Boosting SMN2 expression is a potential therapy for SMA. SMN2 has identical coding sequence as SMN1 except for a silent C-to-T transition at the 6th nucleotide of exon 7, converting a splicing enhancer to a silencer motif. Consequently, most SMN2 transcripts lack exon 7. More than ten putative splicing regulatory elements (SREs) were reported to regulate exon 7 splicing. To investigate the relative strength of each negative SRE in inhibiting exon 7 inclusion, antisense oligonucleotides (AONs) were used to mask each element, and the fold increase of full-length SMN transcripts containing exon 7 were compared. The most potent negative SREs are at intron 7 (in descending order): ISS-N1, 3' splice site of exon 8 (ex8 3'ss) and ISS+100. Dual-targeting AONs were subsequently used to mask two nonadjacent SREs simultaneously. Notably, masking of both ISS-N1 and ex8 3'ss induced the highest fold increase of full-length SMN transcripts and proteins. Therefore, efforts should be directed towards the two elements simultaneously for the development of optimal AONs for SMA therapy.


Assuntos
Atrofia Muscular Espinal/terapia , Oligonucleotídeos Antissenso/uso terapêutico , Splicing de RNA/genética , Elementos Facilitadores Genéticos/genética , Éxons/genética , Terapia Genética , Humanos , Neurônios Motores/metabolismo , Neurônios Motores/patologia , Atrofia Muscular Espinal/genética , Atrofia Muscular Espinal/patologia , Sítios de Splice de RNA , Proteína 1 de Sobrevivência do Neurônio Motor/genética , Proteína 2 de Sobrevivência do Neurônio Motor/antagonistas & inibidores , Proteína 2 de Sobrevivência do Neurônio Motor/genética , Proteína 2 de Sobrevivência do Neurônio Motor/uso terapêutico , Transcrição Gênica
5.
EMBO Mol Med ; 5(10): 1586-601, 2013 10.
Artigo em Inglês | MEDLINE | ID: mdl-24014320

RESUMO

Loss-of-function mutations in SMN1 cause spinal muscular atrophy (SMA), a leading genetic cause of infant mortality. The related SMN2 gene expresses suboptimal levels of functional SMN protein, due to a splicing defect. Many SMA patients reach adulthood, and there is also adult-onset (type IV) SMA. There is currently no animal model for adult-onset SMA, and the tissue-specific pathogenesis of post-developmental SMN deficiency remains elusive. Here, we use an antisense oligonucleotide (ASO) to exacerbate SMN2 mis-splicing. Intracerebroventricular ASO injection in adult SMN2-transgenic mice phenocopies key aspects of adult-onset SMA, including delayed-onset motor dysfunction and relevant histopathological features. SMN2 mis-splicing increases during late-stage disease, likely accelerating disease progression. Systemic ASO injection in adult mice causes peripheral SMN2 mis-splicing and affects prognosis, eliciting marked liver and heart pathologies, with decreased IGF1 levels. ASO dose-response and time-course studies suggest that only moderate SMN levels are required in the adult central nervous system, and treatment with a splicing-correcting ASO shows a broad therapeutic time window. We describe distinctive pathological features of adult-onset and early-onset SMA.


Assuntos
Atrofia Muscular Espinal/patologia , Splicing de RNA , Animais , Sequência de Bases , Sistema Nervoso Central/metabolismo , Humanos , Fator de Crescimento Insulin-Like I/metabolismo , Fígado/patologia , Camundongos , Camundongos Transgênicos , Atrofia Muscular Espinal/metabolismo , Miocárdio/patologia , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/farmacologia , Splicing de RNA/efeitos dos fármacos , Proteína 1 de Sobrevivência do Neurônio Motor/genética , Proteína 1 de Sobrevivência do Neurônio Motor/metabolismo , Proteína 2 de Sobrevivência do Neurônio Motor/antagonistas & inibidores , Proteína 2 de Sobrevivência do Neurônio Motor/genética , Proteína 2 de Sobrevivência do Neurônio Motor/metabolismo , Fatores de Tempo
6.
J Cell Biol ; 199(1): 21-5, 2012 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-23027901

RESUMO

One of the greatest thrills a biomedical researcher may experience is seeing the product of many years of dedicated effort finally make its way to the patient. As a team, we have worked for the past eight years to discover a drug that could treat a devastating childhood neuromuscular disease, spinal muscular atrophy (SMA). Here, we describe the journey that has led to a promising drug based on the biology underlying the disease.


Assuntos
Atrofia Muscular Espinal/tratamento farmacológico , Oligonucleotídeos Antissenso/farmacologia , Animais , Humanos , Camundongos , Camundongos Transgênicos , Atrofia Muscular Espinal/genética , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/genética , Proteína 1 de Sobrevivência do Neurônio Motor/antagonistas & inibidores , Proteína 1 de Sobrevivência do Neurônio Motor/genética , Proteína 2 de Sobrevivência do Neurônio Motor/antagonistas & inibidores , Proteína 2 de Sobrevivência do Neurônio Motor/genética
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...