Relevant Degree Programs
Affiliations to Research Centres, Institutes & Clusters
Graduate Student Supervision
Doctoral Student Supervision (Jan 2008 - April 2022)
Frontotemporal dementia (FTD) is a devastating and progressive disorder and a common form of early-onset dementia. There are currently no disease-modifying therapies available, signifying the need for new therapeutic approaches. Progranulin (PGRN) haploinsufficiency due to autosomal dominant mutations in the progranulin gene (GRN) is a major cause of familial FTD (FTD-GRN), with nearly a quarter of these genetic cases resulting from a nonsense mutation. Nonsense mutations introduce premature termination codons (PTCs) that can be therapeutically targeted by compounds allowing readthrough, and aminoglycoside antibiotics are known to be potent PTC readthrough drugs. When this research project was initiated, restoring PGRN through PTC readthrough had not been explored as a therapeutic intervention in FTD-GRN. We used human induced pluripotent cell (hiPSC) lines bearing clinical nonsense mutations spanning the GRN coding region (S116X⁺/˗, R418X⁺/˗, R493X-/- KI) to evaluate G418 and novel PTC readthrough enhancer (CDX-series) combination treatments. Our aim was to demonstrate proof-of-concept GRN PTC readthrough and lower the required dose of G418 to address the known toxicity of traditional aminoglycoside PTC readthrough agents.Screening in HEK293 cells expressing nonsense mutant (S116X, R418X, R493X) GRN expression constructs found PTC readthrough combination treatment with G418 and CDX5-288 enhancer most potently induced GRN readthrough. We demonstrated in vivo proof-of-concept GRN PTC readthrough by performing single intracerebroventricular (ICV) injections of G418 with or without CDX5-288 enhancer in our GRN R493X adeno-associated virus-based mouse model. Combination treatment in hiPSC-derived isogenic R493X-/- KI cortical neurons significantly restored PGRN levels and normalized overexpression of the mature form of the lysosomal enzyme cathepsin D. We attempted to achieve in vivo PTC readthrough of GrnR493X through repeated ICV administrations of G418 in GrnR493X/R493X mice. However, G418 doses capable of eliciting PTC readthrough in these mice were associated with significant neurotoxicity. We next conducted further neuropathological characterization of lysosomal dysfunction, neuroinflammation, and neurodegeneration in GrnR493X/R493X mice, identifying several phenotypes recently reported in Grn-/- mice, including decreased thalamic excitatory neuronal density. Taken together, our findings suggest that PTC readthrough may be a potential therapeutic strategy for FTD caused by GRN nonsense mutations and support further investigations into novel readthrough drugs with improved tolerability.
Master's Student Supervision (2010 - 2021)
Frontotemporal dementia (FTD) refers to a group of neurodegenerative disorders that are characterized by pathology predominantly localized to the frontal and temporal lobes. FTD is the second leading cause of early-onset dementia behind Alzheimer’s Disease, and there are currently no disease-modifying treatments available. Approximately 40% of FTD cases are familial, and 25% of these are caused by heterozygous loss of function mutations in the gene encoding for progranulin (PGRN), GRN. Since its discovery almost 15 years ago, a plethora of research has attempted to explain the mechanisms for how loss of PGRN leads to FTD, but an entire picture remains unclear. Evidence from murine models has in the past suggested that astrocytes, the main supporting cells of the brain, do not secrete PGRN; however, more recent analysis has shown astrocyte PGRN expression in both murine and human cells. It has also been shown that mutations in MAPT – another leading cause of familial FTD – greatly alters astrocyte gene expression which leads to subsequent non-cell autonomous effects on neurons. In this study, we utilized human induced pluripotent stem cell (hiPSC)-derived neural tissue carrying a homozygous GRN R493X-/- knock-in mutation to investigate in vitro whether GRN mutant astrocytes have a non-cell autonomous effect on neurons. Using microelectrode array analysis, we demonstrated that GRN R493X-/- astrocytes impact neuron maturation by significantly delaying excitatory electrical development. Histologically, neurons during the delay showed a decrease in GABAergic synaptic markers while also showing an increase in glutaminergic synaptic markers. We also demonstrate that this effect is due in-part due to soluble factors, and that GRN mutation alters neurotrophic factor secretion in astrocytes. Overall, this work represents the first study investigating astrocyte-induced neuronal pathology in GRN mutant hiPSCs, and supports the hypothesis of astrocyte involvement in the progression of FTD.