Systems biology investigation of cAMP modulation to increase SMN levels for the treatment of spinal muscular atrophy.

Sean G Mack,Matthew E R Butchbach,Prasad Dhurjati,Daniel J Cook,Thomas H Gillingwater

doi:10.1371/journal.pone.0115473

Abstract

Spinal muscular atrophy (SMA), a leading genetic cause of infant death worldwide, is an autosomal recessive disorder caused by the loss of SMN1 (survival motor neuron 1), which encodes the protein SMN. The loss of SMN1 causes a deficiency in SMN protein levels leading to motor neuron cell death in the anterior horn of the spinal cord. SMN2, however, can also produce some functional SMN to partially compensate for loss of SMN1 in SMA suggesting increasing transcription of SMN2 as a potential therapy to treat patients with SMA. A cAMP response element was identified on the SMN2 promoter, implicating cAMP activation as a step in the transcription of SMN2. Therefore, we investigated the effects of modulating the cAMP signaling cascade on SMN production in vitro and in silico. SMA patient fibroblasts were treated with the cAMP signaling modulators rolipram, salbutamol, dbcAMP, epinephrine and forskolin. All of the modulators tested were able to increase gem formation, a marker for SMN protein in the nucleus, in a dose-dependent manner. We then derived two possible mathematical models simulating the regulation of SMN2 expression by cAMP signaling. Both models fit well with our experimental data. In silico treatment of SMA fibroblasts simultaneously with two different cAMP modulators resulted in an additive increase in gem formation. This study shows how a systems biology approach can be used to develop potential therapeutic targets for treating SMA.

Highlights

Spinal muscular atrophy (SMA) is an autosomal recessive, neurodegenerative disorder characterized by the progressive loss of a-motor neurons in the anterior horn of the spinal cord; this loss leads to progressive muscle weakness and atrophy [1]
Fibroblasts derived from a type II SMA patient (GM03813; [26]) were treated with increasing doses (n53/dose) of one of the following modulators of cyclic adenosine monophosphate (cAMP) signaling: epinephrine, salbutamol, forskolin, dibutyryl cAMP (dbcAMP) and rolipram
While none of the compounds attained gem counts similar to those observed in fibroblasts (GM03814) derived from the mother of GM03813—i.e. carrier fibroblasts, the number of gems/100 nuclei in SMA fibroblasts treated with the highest doses of dbcAMP, forskolin, salbutamol and rolipram reached at least 50% of the gem counts found in carrier fibroblasts

Summary

Introduction

Spinal muscular atrophy (SMA) is an autosomal recessive, neurodegenerative disorder characterized by the progressive loss of a-motor neurons in the anterior horn of the spinal cord; this loss leads to progressive muscle weakness and atrophy [1]. SMA is a leading genetic cause of infant death worldwide with 1 in 5000– 10,000 children born with the disease [2, 3]. There is a single nucleotide change (CRT) within SMN2 exon 7 that causes most of SMN2 mRNAs to lack exon 7 (SMND7). The resultant SMND7 protein is unstable and not fully functional [7, 8]. The number of SMN2 copies modifies disease severity in SMA patients [9,10,11,12,13,14,15,16]. In transgenic mouse models for SMA, the copy number of human SMN2 modulates the phenotypic severity [17,18,19]. SMN2 is, an endogenous genetic modifier of disease severity in SMA

Methods

Results

Conclusion