NCALD Antisense Oligonucleotide Therapy in Addition to Nusinersen further Ameliorates Spinal Muscular Atrophy in Mice

Abstract

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.

Twenty-Five Years of Spinal Muscular Atrophy Research: From Phenotype to Genotype to Therapy, and What Comes Next

“…Splice-modifying ASOs were first tested in an SMA mouse model with SMN2 to confirm their effectiveness in vivo (78,80). Recently, combinatorial therapy with two different ASOs-an SMN2 splice corrector and an inhibitor of the NCALD genetic modifier-was tested in an SMA mouse model and showed promising outcomes (169). Gene therapy with self-complementary adenoassociated virus (scAAV) delivery of SMN1 alone or with other genetic modifiers, including IGF, PLS3, and STMN1, has also been tested in SMA mouse models (45,55,87,121,134,170,176).…”

Spinal muscular atrophy: From approved therapies to future therapeutic targets for personalized medicine

“…125 Similar therapeutic benefits were obtained by decreasing levels of NCALD. 126 Furthermore, PLS3 interacts with coronin 1C-an actin-bundling protein, with calcineurin-like EF-hand protein 1 (CHP1)-a calcium sensor and calcineurin inhibitor, 127 and with members of the hnRNP F/H family of proteins. 128 Modulation of their respective expression improved impaired endocytic pathways and neuromuscular pathology in SMA models.…”

The Identification of Novel Biomarkers Is Required to Improve Adult SMA Patient Stratification, Diagnosis and Treatment

“…Moreover, reducing NCALD levels either in vitro or in vivo significantly ameliorated SMA pathology across SMA species [ 10 ]. Based on these encouraging results, Torres-Benito et al [ 89 ] developed an ASO-based therapy to target Ncald in mouse spinal cord and used it in combination with low-dose SMN splice switching ASOs to design an efficient combinatorial therapy in SMA mice [ 89 ]. Therefore, repression of NCALD is considered protective for SMA.…”