Pathogenic mechanisms of C9orf72 GGGGCC repeat expansions in amyotrophic lateral sclerosis and frontotemporaldementia and development of therapeutic strategies

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DESCRIPTION (provided by applicant): Expanded GGGGCC hexanucleotide repeats in a non-coding region of the C9orf72 gene were recently identified as the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two neurodegenerative conditions with genetic and pathological overlap. The pathogenic mechanisms of this expansion are not understood, but initial observations point to either a loss of function of the endogenous C9orf72 gene, and/or a toxic gain of function of the expanded RNA, mediated either by sequestration of RNA binding proteins or by production of aberrant polypeptide(s) through repeat-associated non-ATG-dependent (RAN) translation. Here, I propose to use genetically modified mouse models to determine contribution of RNA-mediated gain of toxicity and/or C9orf72 loss of function to ALS/FTD pathogenesis caused by C9orf72 repeat expansions and to develop therapeutic strategies. I have already established multiple lines of BAC transgenic mice expressing a repeat-containing human C9orf72 gene with different repeat lengths and expression levels, including ones with a similar RNA level but with hexanucleotide repeats between ~100 and ~450, and those that have ~450 repeats with a 4-fold range of RNA expression. I have also documented abnormal, pathologic RNA foci containing both sense and antisense repeat RNAs in several brain regions and spinal cords of all transgenic mice tested so far (4 lines), similar to what have been observed in human C9orf72 patients, supporting that ALS/FTD pathogenesis is driven, at least in part, by RNA-mediated gain of toxicity. In Aim 1A & 1B, I will determine if any age-dependent pathology (RNA foci or repeat-associated non-ATG (RAN) translation), RNA signature change and/or phenotype is developed in C9orf72 repeat expressing transgenic mice and if such pathology/phenotype is repeat length and/or expression dependent. Recognizing that a contribution of C9orf72 haploinsufficiency to ALS and FTD pathogenesis is supported by reduced C9orf72 mRNAs in tissues from C9orf72-ALS/FTD patients, I will determine the consequence (if any) of diminished C9orf72 expression in mice with a disrupted C9orf72 allele and if the transgene-dependent ALS/FTD related phenotype is exacerbated by C9orf72 loss of function (Aim 1C). Finally, I will use transgenic mice to identify antisense oligonucleotides (ASOs) that mediate degradation of C9orf72 RNAs carrying repeat expansions in an effort to develop a therapeutic approach to diminish the consequences of any toxic gain of function (Aim 2). I believe that the combination of Aims 1 and 2 that I propose here have substantial promise to identify disease mechanism from hexanucleotide expansion in C9orf72 and facilitate development of ASO infusion therapy as an approach for human clinical trial.