Development of targeted therapeutic agents for the treatment of FXTAS (2 of 6)

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Research under Project 1 of the Consortium will involve further study of neural cell models of the neurodegenerative disorder, fragile X-associated tremor/ataxia syndrome (FXTAS). The pathogenesis of FXTAS is the direct result of a toxic gain-of-function of excess FMR1 RNA that is produced from premutation CGG-repeat expansions of the fragile X mental retardation (FMR1) gene. The principal objective of this project will be the identification and assessment of various candidate therapeutic agents that m.ight attenuate the effects of the pathogenic RNA; through antisense-DNA- (ASO) or siRNA-mediated knock-down (Aims 1 and 2); through downstream effects of the neuropathic process, such as impaired glutamate regulation/uptake leading to glutamate toxicity (Aim 3); or through more proximal effects of the expanded- CGG-repeat RNA (Aim 4). Aims 1 and 2 will test the hypothesis that elimination of the RNA, either in neural cell models or in transgenic mouse models (Project 2), will at least partially reverse the pathogenic process. Studies using inducible cell models will test this hypothesis and, with Project 2, will develop expectations for future oligonucleotide-based therapies in humans. Based on clinical/pathological evidence (Projects 1 and 3) of clear astrocyte dysfunction in FXTAS, we will test our working hypothesis that glutamate dysregulation contributes to the neurodegeneration in FXTAS, as one component of these studies, we will test the effect of memantine in mixed neuronal/astrocyte murine cell models of FXTAS, in part to provide a cellular basis for the clinical trial of memantine in Project 3. The development of oligonucleotide-based therapeutic agents (ASO, siRNA) will involve two important elements. First, a systematic determination of the necessity of using ASOs, which can target nuclear as well as cytoplasmic RNA, or whether it is sufficient to use siRNA-based targeting to the cytoplasm. Second, together with Project 2, we will develop non-viral, CNS-targeted immunoliposomal delivery methods to deliver candidate ASO or siRNA oligos in vivo in mouse models (Project 2). During the course of these experiments, and in conjunction with Project 2, we will examine the timing and reversibility of the pathogenic response; this issue is important when considering the timing of therapeutic intervention for FXTAS. Finally, together with Projects 2 and 4, we will investigate the nature of developmental problems in children who are carriers of premutation alleles, to determine whether, as we suspect, their involvement represents a novel developmental phenotype due to RNA toxicity, represents an effect of slightly lower protein levels that would characterize a broad fragile X syndrome spectrum effect, or reflects a combination of both pathogenic mechanisms. This last issue provides an example of the true power of the interdisciplinary approach.