Gene Expression Profiling of Peripheral Tissues in Amyotrophic Lateral Sclerosis

Background: Amyotrophic Lateral Sclerosis, in which cortical and spinal motor neurons degenerate, is a late onset neurodegenerative condition that accounts for ~1 in 400 UK deaths, typically within 3-5 years from the initial manifestations of disease. It forms part of a broad spectrum of clinically, genetically as well as pathologically heterogeneous disorders that include behavioural variant frontotemporal lobar degeneration (bvFTLD). A large intronic hexanucleotide G4C2 repeat expansion of >30 copies was recently identified, in 2011, in the previously uncharacterised chromosome 9 open reading frame 72 (C9ORF72) gene which is now thought to explain up to 43% of familial ALS (~20-30% of familial FTLD) and around 7% of sporadic cases. Rationale & Hypothesis: The principle aim of the PhD was to perform gene expression profiling of peripheral tissues in ALS. In the first instance whole blood was trialled. However, this proved unreliable, owing to the shear abundance of erythrocyte derived alpha and beta haemoglobin transcripts that are contained within the sample and the variability in the efficiency of its removal using the Ambion® GLOBINClear or NuGEN Ovation® WB reduction strategies. Instead disease related changes in transcription/alternative splicing were detected in a large bank (n=820) of patient and control lymphoblastoid cell lines (LCL’s) with the main purpose of: 1) elucidating further the mechanism(s) of neurotoxicity associated with the C9ORF72 G4C2 repeat expansion and, 2) establishing within this specific genetic subtype, modifiers of a fast (<2yrs) versus slow (≥4yrs) disease progression in order to identify potential new areas of therapeutic research. Methodology: Biotinylated, sense-strand cDNA targets of ~40 -70nt were hybridized onto Human Exon 1.0ST GeneChip® Arrays. A GC-RMA normalisation procedure was carried out in Partek® Genomics Suite and differentially expressed or alternatively spliced transcripts were detected at the 5% significance level (p