Poster abstracts

Poster number 3 submitted by Thomas Bebee

Generation of a new mouse model to help determine the therapeutic time-point for SMN replacement therapies: an update

Thomas Bebee (The Center for Childhood Cancer, Columbus Children’s Research Institute and the Department of Pediatrics, The Ohio State University School of Medicine and Public Health, Columbus, Ohio), Jordan Gladman (The Center for Childhood Cancer, Columbus Children’s Research Institute and the Department of Pediatrics, The Ohio State University School of Medicine and Public Health, Columbus, Ohio), Dawn S. Chandler (The Center for Childhood Cancer, Columbus Children’s Research Institute and the Department of Pediatrics, The Ohio State University School of Medicine and Public Health, Columbus, Ohio)

Abstract:
Proximal Spinal Muscular Atrophy (SMA), a leading genetic cause of infant mortality in humans, is caused by deletion or mutation of the survival of motor neuron gene-1 (SMN1). Homozygous deletion of the homologous gene (SMN) in mouse models leads to early embryonic lethality, arguing the necessity of SMN during early development. Humans carry a nearly identical SMN2 gene that is capable of rescuing the embryonic lethality and disease phenotype in mouse models, in a dose dependent manner. The SMN2 gene differs from SMN1 primarily in a C>T point mutation in the exon splicing enhancer (ESE) of exon 7, leading to exon 7 skipping and a truncated non-functional protein. A fraction of SMN2 transcripts include exon 7, therefore, correction of SMN2 splicing to produce increased exon 7 inclusion is an intriguing target for SMA therapies. However, the timing of SMN replacement therapies will be crucial. A temporally inducible SMN-transgenic mouse when placed in the SMA disease background will allow for the determination of this window. To achieve an inducible SMN-transgenic mouse we will place SMN expression under the control of tamoxifen inducible Cre recombination, allowing for controlled expression of SMN at varying time points during development. In our transgene construct, both drug resistance and reporter gene expression are expressed prior to Cre recombination, whereas, SMN expression is permitted only after Cre recombination. The non-recombined transgene construct was electroporated into ES cells. We screened the resultant ES cells for adequate expression by staining for the reporter gene and for a single integration event by Southern Blot. Three selected ES cell lines were then electroporated with a plasmid expressing Cre recombinase, and expression of the transgene and Cre specific recombination were assessed by PCR. These lines have been injected into Mouse blastocysts and chimeras from these lines are being tested for germline transmission.

Keywords: Spinal Muscular Atrophy, Survival Motor Neuron, Mouse Model