nusinersen and Disease-Models--Animal

The new treatments of spinal muscular atrophy (SMA) due by SMN1 gene deletions are reviewed. There are several ways to increase the protein SMN, its activity and persistence in the tissues. Neuroprotective drugs as olesoxime or riluzole, and drugs acting by epigenetic mechanisms, as histone deacetylase inhibitors, have shown positive effects in preclinical studies but no clear efficacy in clinical trials. They might give in the future added benefits when used associated to other genetic modifying drugs. The best improvements in murine models of SMA and in clinical trials have been reached with antisense oligonucleotides, drugs that modify the splicing of SMN2, and they are expected to get better in the near future. Nusinersen, a methoxi-ethyl phosphotioate antisense oligonucleotide has recently approved for treatment of patients with SMA type 1 after having proved its efficacy in clinical trial phase 3. The results of nusinersen are reviewed. New modifications of antisense oligonucleotides with better access to brain, spinal cord and peripheral tissues are on the way. There are data of the efficacy of the genetic therapy with SMN1 gene through adenoassociated virus, now in phase 1 trial. A constant feature of these new treatments is that the earlier the treatment, the best are the results, and they are even better in presymptomatic stage. The general standards of care, particularly nutrition and respiratory management are needed in order to reach optimal results with the new therapies.. Posibilidades de tratamiento en la atrofia espinal infantil.. Se revisan los nuevos tratamientos de la atrofia muscular espinal (AME) producida por delecion del gen SMN1. Se describen las diferentes posibilidades de incrementar la proteina SMN, de su actividad y persistencia en el organismo. Farmacos neuroprotectores, como olesoxime y riluzol, y farmacos que actuan epigeneticamente, como inhibidores de histona deacetilasa, han mostrado cierto efecto positivo en fases preclinicas pero no han conseguido eficacia en los ensayos clinicos. Podrian proporcionar en un futuro un beneficio añadidos a otros farmacos modificadores geneticos. Los mayores cambios en estudios de modelos del raton SMA y en fases clinicas se han encontrado con oligonucleotidos antisentido que modifican el splicing del gen SMN2, y se espera que mejoren en el futuro proximo. Recientemente se ha aprobado el nusinersen, un metoxietilo fosforotioato-oligonucleotido antisentido, para uso en pacientes con AME de tipo I una vez demostrada su eficacia en pacientes en el ensayo en fase 3. Se revisan los resultados de este farmaco. Estan en marcha modificaciones de oligonucleotidos antisentido que amplien la liberacion en el sistema nervioso y en tejidos perifericos. Hay datos que sugieren eficacia de la terapia genica introduciendo el gen SMN1 mediante virus adenoasociados, actualmente en fase clinica 1. Una constante en estos nuevos tratamientos es que los resultados se optimizan en las etapas precoces de la enfermedad y, mejor aun, en estadio presintomatico. Se subraya la importancia de los cuidados generales optimos, especialmente nutricionales y respiratorios, para conseguir los mejores resultados con las nuevas terapias.

Topics: Animals; Child; Clinical Trials as Topic; Dependovirus; Disease Models, Animal; Epigenesis, Genetic; Gene Deletion; Genetic Therapy; Genetic Vectors; Histone Deacetylase Inhibitors; Humans; Mice; Mice, Neurologic Mutants; Multicenter Studies as Topic; Neuroprotective Agents; Oligonucleotides; Oligonucleotides, Antisense; Palliative Care; Pluripotent Stem Cells; Recombinant Proteins; RNA Splicing; Spinal Muscular Atrophies of Childhood; Survival of Motor Neuron 1 Protein; Survival of Motor Neuron 2 Protein; Therapies, Investigational

Spinal muscular atrophy (SMA) is a devastating neuromuscular disorder characterized by loss of spinal cord motor neurons, muscle atrophy and infantile death or severe disability. It is caused by severe reduction of the ubiquitously expressed survival motor neuron (SMN) protein, owing to loss of the SMN1 gene. This would be completely incompatible with survival without the presence of a quasi-identical duplicated gene, SMN2, specific to humans. SMN2 harbours a silent point mutation that favours the production of transcripts lacking exon 7 and a rapidly degraded non-functional SMNΔ7 protein, but from which functional full length SMN protein is produced at very low levels (∼10%). Since the seminal discovery of the SMA-causing gene in 1995, research has focused on the development of various SMN replacement strategies culminating, in December 2016, in the approval of the first precise molecularly targeted therapy for SMA (nusinersen), and a pivotal proof of principle that therapeutic antisense oligonucleotide (ASO) treatment can effectively target the central nervous system (CNS) to treat neurological and neuromuscular disease. Nusinersen is a steric block ASO that binds the SMN2 messenger RNA and promotes exon 7 inclusion and thus increases full length SMN expression. Here, we consider the implications of this therapeutic landmark for SMA therapeutics and discuss how future developments will need to address the challenges of delivering ASO therapies to the CNS, with appropriate efficiency and activity, and how SMN-based therapy should be used in combination with complementary strategies to provide an integrated approach to treat CNS and peripheral pathologies in SMA.

Topics: Animals; Central Nervous System; Disease Models, Animal; Exons; Humans; Mice; Motor Neurons; Muscular Atrophy, Spinal; Oligodeoxyribonucleotides, Antisense; Oligonucleotides; Oligonucleotides, Antisense; RNA, Messenger; Spinal Cord; Survival of Motor Neuron 1 Protein; Survival of Motor Neuron 2 Protein

Spinal muscular atrophy (SMA) is a devastating autosomal recessive neuromuscular disease characterized by degeneration of spinal cord alpha motor neurons (αMNs). SMA is caused by the homozygous deletion or mutation of the survival motor neuron 1 (SMN1) gene, resulting in reduced expression of SMN protein, which leads to αMN degeneration and muscle atrophy. The majority of transcripts of a second gene (SMN2) generate an alternative spliced isoform that lacks exon 7 and produces a truncated nonfunctional form of SMN. A major function of SMN is the biogenesis of spliceosomal snRNPs, which are essential components of the pre-mRNA splicing machinery, the spliceosome. In recent years, new potential therapies have been developed to increase SMN levels, including treatment with antisense oligonucleotides (ASOs). The ASO-nusinersen (Spinraza) promotes the inclusion of exon 7 in SMN2 transcripts and notably enhances the production of full-length SMN in mouse models of SMA. In this work, we used the intracerebroventricular injection of nusinersen in the SMN∆7 mouse model of SMA to evaluate the effects of this ASO on the behavior of Cajal bodies (CBs), nuclear structures involved in spliceosomal snRNP biogenesis, and the cellular distribution of polyadenylated mRNAs in αMNs. The administration of nusinersen at postnatal day (P) 1 normalized SMN expression in the spinal cord but not in skeletal muscle, rescued the growth curve and improved motor behavior at P12 (late symptomatic stage). Importantly, this ASO recovered the number of canonical CBs in MNs, significantly reduced the abnormal accumulation of polyadenylated RNAs in nuclear granules, and normalized the expression of the pre-mRNAs encoding chondrolectin and choline acetyltransferase, two key factors for αMN homeostasis. We propose that the splicing modulatory function of nusinersen in SMA αMN is mediated by the rescue of CB biogenesis, resulting in enhanced polyadenylated pre-mRNA transcription and splicing and nuclear export of mature mRNAs for translation. Our results support that the selective restoration of SMN expression in the spinal cord has a beneficial impact not only on αMNs but also on skeletal myofibers. However, the rescue of SMN expression in muscle appears to be necessary for the complete recovery of motor function.

Topics: Active Transport, Cell Nucleus; Animals; Coiled Bodies; Disease Models, Animal; Mice; Mice, Knockout; Motor Neurons; Muscular Atrophy, Spinal; Oligonucleotides; RNA, Messenger; Survival of Motor Neuron 2 Protein

The deficiency of survival motor neuron (SMN) protein can result in the onset of spinal muscular atrophy (SMA), an autosomal recessive disorder characterized by a progressive loss of motor neurons and skeletal muscle atrophy. The mechanism underlying SMA pathology remains unclear. Here, we demonstrate that SMN protein regulates oxidative stress and inflammatory response in microglia. Antisense oligonucleotide, which increases SMN protein expression (SMN-ASO), attenuated SMA model mice phenotypes and suppressed the activation of microglia in the spinal cord. The expression of oxidative stress marker in microglia was decreased by SMN-ASO injection in SMA model mice. Increased reactive oxygen species production and subsequent antioxidative stress reaction was observed in SMN protein-depleted RAW264.7. Furthermore, nuclear factor kappa B (NFκB) and c-Jun amino terminal kinase (JNK) signaling, which mainly mediate the inflammatory response, are activated in SMN protein-depleted RAW264.7. Tumor necrosis factor-α (TNF-α) production is also increased in SMN protein-depleted RAW264.7. These findings suggest that SMN protein regulates oxidative stress and inflammatory response in microglia, supporting current claims that microglia can be an effective target for SMA therapy.

Topics: Animals; Disease Models, Animal; Gene Expression; Inflammation; MAP Kinase Signaling System; Mice; Mice, Transgenic; Microglia; Molecular Targeted Therapy; Muscular Atrophy, Spinal; NF-kappa B; Oligonucleotides; Oligonucleotides, Antisense; Oxidative Stress; RAW 264.7 Cells; Spinal Cord; Survival of Motor Neuron 1 Protein; Tumor Necrosis Factor-alpha

Spinal muscular atrophy (SMA) is a neuromuscular disease causing the most frequent genetic childhood lethality. Recently, nusinersen, an antisense oligonucleotide (ASO) that corrects SMN2 splicing and thereby increases full-length SMN protein, has been approved by the FDA and EMA for SMA therapy. However, the administration of nusinersen in severe and/or post-symptomatic SMA-affected individuals is insufficient to counteract the disease. Therefore, additional SMN-independent therapies are needed to support the function of motoneurons and neuromuscular junctions. We recently identified asymptomatic SMN1-deleted individuals who were protected against SMA by reduced expression of neurocalcin delta (NCALD). NCALD reduction is proven to be a protective modifier of SMA across species, including worm, zebrafish, and mice. Here, we identified Ncald-ASO3-out of 450 developed Ncald ASOs-as the most efficient and non-toxic ASO for the CNS, by applying a stepwise screening strategy in cortical neurons and adult and neonatal mice. In a randomized-blinded preclinical study, a single subcutaneous low-dose SMN-ASO and a single intracerebroventricular Ncald-ASO3 or control-ASO injection were presymptomatically administered in a severe SMA mouse model. NCALD reduction of >70% persisted for about 1 month. While low-dose SMN-ASO rescues multiorgan impairment, additional NCALD reduction significantly ameliorated SMA pathology including electrophysiological and histological properties of neuromuscular junctions and muscle at P21 and motoric deficits at 3 months. The present study shows the additional benefit of a combinatorial SMN-dependent and SMN-independent ASO-based therapy for SMA. This work illustrates how a modifying gene, identified in some asymptomatic individuals, helps to develop a therapy for all SMA-affected individuals.

Topics: Animals; Disease Models, Animal; Gene Expression Regulation; Mice; Muscular Atrophy, Spinal; Neurocalcin; Oligonucleotides; Oligonucleotides, Antisense; Survival of Motor Neuron 1 Protein

Spinal muscular atrophy (SMA) is the most common genetic cause of infantile death caused by mutations in the SMN1 gene. Nusinersen (Spinraza), an antisense therapy-based drug with the 2'-methoxyethoxy (2'MOE) chemistry approved by the FDA in 2016, brought antisense drugs into the spotlight. Antisense-mediated exon inclusion targeting SMN2 leads to SMN protein expression. Although effective, 2'MOE has weaknesses such as the inability to cross the blood-brain barrier and the high cost of treatment. To investigate new chemistries of antisense oligonucleotides (ASOs), SMA mouse models can serve as an important source. Here we describe methods to test the efficacy of ASOs, such as phosphorodiamidate morpholino oligomers (PMOs), in a severe SMA mouse model.

Topics: Animals; Disease Models, Animal; Exons; Gene Expression Regulation; Genotype; Infusions, Intraventricular; Mice; Mice, Knockout; Morpholinos; Mutation; Oligonucleotides; Oligonucleotides, Antisense; RNA Splicing; Survival of Motor Neuron 2 Protein