Spinal muscular atrophy (SMA)
Background
Spinal muscular atrophy (SMA) is a neuromuscular disease that is the leading genetic cause of infant mortality in humans, affecting 1 in 6000 live births in the UK. SMA primarily causes muscle wasting and mobility impairment, and despite a full understanding of the genetic cause of the disease (mutations in a single gene known as survival motor neuron 1, SMN1, resulting in loss of the SMN protein), there is currently no cure. Severe SMA patients will usually die within 2 years.
Huge progress has been made in the past few years in the development of SMA therapies. Nusinersen, or Spinraza, is an anti-sense oligonucleotide that has recently been given approval to treat SMA patients and treated patients are showing vast health and survival improvements. There remain a number of additional therapeutics, including gene therapy, at varying stages of development in the pipeline.
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One of the major concerns in therapy development for SMA is that all pathologies that develop due to low levels of SMN are treated. The main pathology in SMA is the neuromuscular system, but we know that many other tissues and organs are affected (systemic pathology). Nusinersen specifically targets the central nervous system and there is not data yet on currently treated patients to determine whether any secondary pathology is developing in these patients. Many of the therapies under development are targeting systemic pathology too.
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A number of roles for SMN have been proposed, but more basic research is required to determine exactly what SMN does in normal healthy individuals and why reduced levels of SMN causes pathology in so many different systems. Understanding more about the role of SMN in different tissues will enable better design of therapeutics.
Previous work
Recent research carried out by us and our collaborators has shown that a gene downstream of SMN1, Ubiquitin-like modifier-activating enzyme-1 (Uba1) is downregulated in SMA leading to increased levels of β-catenin (Wishart et al., 2014). We have demonstrated that therapeutic targeting of downstream effectors of SMN1 (for example β-catenin) can lead to the amelioration of neuromuscular, but not systemic, pathology (Wishart et al., 2014). In further work, we showed that Uba1 gene therapy rescues a number of, but not all, phenotypes in the Taiwanese SMA mouse model (Powis et al, 2016).
Our work has also expanded on the tissues affected by reduced levels of SMN (Hamilton and Gillingwater, 2013). In 2014, we established that SMN-dependent intrinsic defects in Schwann cells contributed to pathogenesis in SMA and that this was linked to ubiquitin homestasis (Hunter et al.., 2014; Aghamaleky Sarvestany et al., 2014) and in 2016 we described partial rescue of some phenotypes in our novel mouse model expressing SMN solely in Schwann cells (Hunter et al., 2016).
We advocate that a greater understanding of the molecular pathways underlying SMA will enable the development of therapeutics targeting all aspects of SMA pathology as soon as possible to produce the best healthcare outcomes for patients. As part of this, we believe that the development of SMN-Plus, or combinatorial therapies, may provide the best chance of this happening (Groen et al., 2018).
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Restoring SMN expression in Schwann cells rescues myelination defects in SMA mice (Hunter et al., 2016).
UBA1 and mono-/multi-meric ubiquitin is reduced in SMA mice (Wishart et al., 2014).
Rescue of zebrafish motor neuron pathology following Uba1 restoration (Powis et al., 2016).
Improved body weight and survival in SMA mice treated with AAV9-UBA1 (Powis et al., 2016).
Current projects
The contribution of epigenetics to SMA pathology
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There is some evidence that epigenetic modifications contribute to SMA pathogenesis. A recent comparison of lymphoblasts from SMA patients compared to those from healthy individuals identified a number of genes that were differentially methylated (Zheleznykova et al., 2013).
In our lab, we have carried out preliminary work to determine whether any of the genes identified from the patient study have altered levels of gene expression in clinically relevant tissue from the Taiwanese SMA mouse model. Our preliminary data demonstrates that there are significant changes in the expression levels of at least three genes in SMA muscle tissue (Figure 1).
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Figure 1. Significantly altered expression of CDK2AP1, CYTSB and RPL9 in spinal muscular atrophy.
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We are also interested in linking epigenetic change in SMA to known biological mechanisms such as reduced levels of UBA1. UBA1 has an important role in protein ubiquitination and we are currently exploring whether there is any link in SMA between altered UBA1, histone ubiquitination and gene expression. A greater understanding of the biological mechanisms underlying SMA provides greater scope for the development of successful therapeutics for SMA.
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The role of neural crest cells in SMA pathology
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The majority of SMA researchers acknowledge that lack of SMN protein leads to pathology in multiple organs, for example, the heart, liver, spleen and brain, albeit to different extents (Hamilton and Gillingwater 2013). Current therapeutics for SMA are provided to affected individuals following diagnosis, but the age at which patients receive this can vary dependent on circumstances. Newborn screening is being strongly advocated for, particularly in the United States, and this will ensure that children with SMA will obtain life-changing drugs as soon as possible after diagnosis.
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We believe that both the timing and targeting of SMA therapeutic intervention is crucial to patient benefit. Recent work demonstrated that restoring SMN at embryonic or early postnatal time points in SMA mouse models substantially improved survival, arrested motor neuron loss, and mitigated neuromuscular pathology; later restoration has little effect (reviewed in Hamilton and Gillingwater, 2013). A greater understanding of the cellular consequences of SMN loss in early development is vital in determining the best time to deliver a therapy with the most benefit.
One unexplored cell type in SMA research, as far as we are aware, are neural crest cells (NCCs). The NC is a multipotent and migratory cell type that forms transiently in developing vertebrate embryos, however, the effect of reduced SMN levels on actual NCC development has not yet been evaluated. We have recently started a project using a zebrafish model of SMA (Figure 2) to investigate whether NCC development is affected in this disease. Zebrafish are an excellent model to study neural crest development and in particular we are investigating two types of NCC, pigment cells and jaw cartilage, both of which can be quantified with relative ease. We also hope to use this model to determine whether earlier (pre-natal) therapeutic intervention might be beneficial to SMA patients.
Figure 2. Zebrafish pigmentation patterns in control fish compared to fish injected with a morpholino targeting Smn.
References
Aghamaleky Sarvestany A, Hunter G, Tavendale A, Lamont DJ, Llavero Hurtado M, Graham LC, Wishart TM, Gillingwater TH (2014) Label-free quantitative proteomic profiling identifies disruption of ubiquitin homeostasis as a key driver of schwann cell defects in spinal muscular atrophy. Journal of Proteome Research, 13(11):4546-57
Groen EJN, Talbot K, Gillingwater, TH (2018) Advances in therapy for spinal muscular atrophy: promises and challenges. Nat Rev Neurol 14, 214-224
Hamilton G and Gillingwater TH (2013) Spinal muscular atrophy: going beyond the motor neuron. Trends in Molecular Medicine, Jan;19(1):40-50
Hunter G, Powis RA, Jones RA, Groen EJN, Shorrock HK, Lane FM, Zheng Y, Sherman DL, Brophy PJ, Gillingwater TH (2016) Restoration of SMN in Schwann cell reverses myelination defects and improves neuromuscular function in spinal muscular atrophy. Human Molecular Genetics, 25(13):2853-2861
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Hunter G, Aghamaleky Sarvestany A*, Roche SL, Syme RC, Gillingwater TH (2014) SMN-dependent intrinsic defects in Schwann cells in mouse models of spinal muscular atrophy. Human Molecular Genetics, 23(9):2235-50
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Powis RA, Karyka E, Boyd P, Côme J, Jones RA, Yinan Z, Szunyogova E, Groen EJN, Hunter G, Thomson D, Wishart TM, Becker CG, Parson SH, Martinat C, Azzouz M, Gillingwater TH (2016) Systemic restoration of UBA1 ameliorates disease in spinal muscular atrophy.
Journal of Clinical Investigation Insight, 1(11):e87908
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Wishart TM, Mutsaers CA, Riessland M, Reimer MM, Hunter G, Hannam ML, Eaton S, Fuller HR, Roche SL, Somers E, Morse R, Young PJ, Lamont DJ, Hammerschmidt M, Morris GE, Parson SH, Skehel PA, Becker T, Robinson IM, Becker CG, Wirth B, Gillingwater TH (2014) Dysregulation of ubiquitin homeostasis and β-catenin signalling promote spinal muscular atrophy. Journal of Clinical Investigation, 124(4):1821-34​
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Zheleznyakova GY, Viosin S, Kiselev AV, Sallman Almen M, Xavier MJ, Maretina MA, Tishchenko LI, Fredriksson R, Baranov VS, Schoth HB (2013) Genome-wide analysis shows association of epigenetic changes in regulators of Rab and Tho GTPases with spinal muscular atrophy severity. European Journal of Human Genetics, 21(9):988-93