Human embryonic stem cell models of huntington’s disease

Huntington's Disease (HD) is a devastating neurodegenerative disorder that typically strikes during the fourth or fifth decade of life causing fatal atrophy of the striatum and other brain regions. A rare feature of this neurodegenerative disorder is the dominant inheritance pattern due to a CAG repeat expansion in exon 1 of the Huntingtin gene. The discovery that ≥35 repeats underlies HD pathology rapidly led to tremendous advances in our understanding of this complex disorder, with the identification from animal models of numerous pathological mechanisms including perturbation of anterograde and retrograde transport, neuronal activity, mitochondrial function, neurotrophic factor production, cholesterol biosynthesis and gene expression. Nonetheless, many disease mechanisms and the interplay between each remain ill defined in HD and particularly within a human cellular environment. As such there exists a need for novel in vitro human models to address these concerns and continue to decipher the etiology of HD. This thesis describes the investigation and detailed characterisation of two human embryonic stem cell (hESC) lines carrying CAG37 and CAG51 repeat expansions to determine whether disease hallmarks are present in undifferentiated or neural differentiated human cells carrying HD mutations. To achieve this, a novel neural differentiation protocol was optimised for the precise comparison of genetically distinct human pluripotent stem cell lines in a high-throughput and chemically defined platform. The application of this protocol revealed HD hESCs possess the capacity to differentiate to various neural lineages with comparable efficiency to wildtype cells consistent with typical neurodevelopment, however, CAG repeat instability, minor gene dysregulation and importantly neuronal functional perturbations were identified. Consequently, HD hESCs carrying typical onset CAG repeat expansions are likely a valid and valuable tool for investigating the pathological events leading to disease onset and elucidating the hierarchical relationship between disease mechanisms. Human neuronal HD cultures additionally provide a promising resource for high-throughput in vitro screening of candidate therapeutics for the advancement of clinical treatment options.