Arnaud Lacoste, PhD MBA

Spinal muscular atrophy (SMA), which results from the loss of expression of the survival of motor neuron-1 (SMN1) gene, represents the most common genetic cause of pediatric mortality. A duplicate copy (SMN2) is inefficiently spliced, producing a truncated and unstable protein. We describe herein a potent, orally active, small-molecule enhancer of SMN2 splicing that elevates full-length SMN protein and extends survival in a severe SMA mouse model. We demonstrate that the molecular mechanism of… Show more

Spinal muscular atrophy (SMA), which results from the loss of expression of the survival of motor neuron-1 (SMN1) gene, represents the most common genetic cause of pediatric mortality. A duplicate copy (SMN2) is inefficiently spliced, producing a truncated and unstable protein. We describe herein a potent, orally active, small-molecule enhancer of SMN2 splicing that elevates full-length SMN protein and extends survival in a severe SMA mouse model. We demonstrate that the molecular mechanism of action is via stabilization of the transient double-strand RNA structure formed by the SMN2 pre-mRNA and U1 small nuclear ribonucleic protein (snRNP) complex. The binding affinity of U1 snRNP to the 5' splice site is increased in a sequence-selective manner, discrete from constitutive recognition. This new mechanism demonstrates the feasibility of small molecule-mediated, sequence-selective splice modulation and the potential for leveraging this strategy in other splicing diseases. Show less

Huntington's disease is caused by expanded CAG repeats in HTT, conferring toxic gain of function on mutant HTT (mHTT) protein. Reducing mHTT amounts is postulated as a strategy for therapeutic intervention. We conducted genome-wide RNA interference screens for genes modifying mHTT abundance and identified 13 hits. We tested 10 in vivo in a Drosophila melanogaster Huntington's disease model, and 6 exhibited activity consistent with the in vitro screening results. Among these, negative regulator… Show more

Huntington's disease is caused by expanded CAG repeats in HTT, conferring toxic gain of function on mutant HTT (mHTT) protein. Reducing mHTT amounts is postulated as a strategy for therapeutic intervention. We conducted genome-wide RNA interference screens for genes modifying mHTT abundance and identified 13 hits. We tested 10 in vivo in a Drosophila melanogaster Huntington's disease model, and 6 exhibited activity consistent with the in vitro screening results. Among these, negative regulator of ubiquitin-like protein 1 (NUB1) overexpression lowered mHTT in neuronal models and rescued mHTT-induced death. NUB1 reduces mHTT amounts by enhancing polyubiquitination and proteasomal degradation of mHTT protein. The process requires CUL3 and the ubiquitin-like protein NEDD8 necessary for CUL3 activation. As a potential approach to modulating NUB1 for treatment, interferon-β lowered mHTT and rescued neuronal toxicity through induction of NUB1. Thus, we have identified genes modifying endogenous mHTT using high-throughput screening and demonstrate NUB1 as an exemplar entry point for therapeutic intervention of Huntington's disease. Show less

Unraveling the therapeutic potential of human embryonic stem cells (hESC) requires tools to modify their genome. We have engineered the PiggyBac transposable element to create an efficient system for gene delivery in hESCs. This redesigned system, named "ePiggyBac," can deliver up to 18 Kb inserts, and transgene expression is observed in almost 90% of hES cells. ePiggyBac transposons can also carry insulators, inducible expression cassettes, and short hairpin RNAs for gain- and loss-of-function… Show more

Unraveling the therapeutic potential of human embryonic stem cells (hESC) requires tools to modify their genome. We have engineered the PiggyBac transposable element to create an efficient system for gene delivery in hESCs. This redesigned system, named "ePiggyBac," can deliver up to 18 Kb inserts, and transgene expression is observed in almost 90% of hES cells. ePiggyBac transposons can also carry insulators, inducible expression cassettes, and short hairpin RNAs for gain- and loss-of-function approaches. In hES cells, ePiggyBac's efficiency is superior to that of viral vectors and previously described transposons, including other PiggyBac-based systems. In addition, ePiggyBac transgenes can be removed from the hESC genome without leaving any mutation. We used this system to direct hESC differentiation toward a neuronal phenotype. We then removed the transposons to obtain transgene-free neuronal precursors and neurons. The ability to create fully reversible genetic modifications represents an important step toward clinical applications of hESCs. Show less