Modelling neurodegeneration using human isogenic system: A next generation approach to study Huntington’s Disease

The development of therapies to treat patients with neuronal indications is currently hampered by the use of animal models as less than 10% of findings derived from these preclinical models can be translated to humans (1). Patient-derived induced pluripotent stem cells (iPSCs) offer the possibility to generate in vitro systems to model neurological diseases that can recapitulate relevant human disease phenotypes. However, conventional human iPSC differentiation protocols are often lengthy, inconsistent, and difficult to scale. More importantly, the lack of genetically matched controls for patient-derived models further complicates the investigation of disease-relevant phenotype and study of molecular mechanisms underlying neurodegeneration. 

To overcome these problems, we developed a proprietary gene-targeting strategy, opti-ox(TM), that enables highly controlled expression of transcription factors to rapidly reprogram human iPSCs (hiPSCs) into pure somatic cell types in a scalable manner (2). 

Combined with CRISPR/Cas9-mediated genetic engineering enabled us to introduce specific mutations in these iPSC-lines and create isogenic disease models that will improve screen specificity and accelerate drug development. 

opti-ox based isogenic ioGlutamatergic Neurons Huntington’s disease (HD) model carrying a 50CAG expansion in the huntingtin (HTT) gene have been developed using this approach. Mutant huntingtin proteins containing elongated polyglutamine (PolyQ) stretches are aggregation-prone and have been reported to affect a range of neuronal subtypes, including the cortical glutamatergic neurons. Characterisation of these neurons by immunochemistry and RT-qPCR showed that the expression profile of pan-neuronal (MAP2 and TUBB3) and glutamatergic (VGLUT1 and VGLUT2) marker genes as well as of the HTT transcript itself are highly similar between ioGlutamatergic Neurons HTT 50CAG and the isogenic control. We are currently performing an in-depth phenotypic characterisation of this disease model and the genetically matched  control to determine the differences in their transcriptome, neuronal activity and mitochondrial functions. Beside the 50CAG mutation in HTT, we have generated mutations in the MAPT, TARDBP, GBA and PRKN to provide isogenic disease models for FTD, FTD/ALS and Parkinson’s disease. Our novel strategy to use the opti-ox technology for the scalable and consistent production of iPSC-derived isogenic disease models, offers new avenues into drug discovery and can accelerate research and the development of new therapeutics.

1 Pawlowski M, et al. Stem Cell Reports. 8(4), 803-812, 2017

2 Vargas-Caballero M, et al., Expert Opin Drug Dis, 2016