PROJECT ABSTRACT
The GGGGCC (G4C2) hexanucleotide repeat expansion (HRE) in the first intron of the gene C9orf72, is
the most common genetic abnormality associated with frontotemporal dementia (FTD) and amyotrophic lateral
sclerosis (ALS). The disease pathogenesis ultimately leads to the concurrent degeneration of cortical forebrain
and spinal motor neurons, and result in the clinical deficits of motor function and dementia. While the C9orf72-
FTD/ALS disease pathogenesis has been well characterized in spinal motor neurons and a contribution of the
observed neurodegeneration has been attributed to spinal cord astrocytes, there is little known about the
pathobiology in cortical astrocytes and their role in cortical neurodegeneration, which is proposed to contribute
to the dementia symptoms in this patient population. Here, we hypothesize that cortical astrocytes play an
integral role in the non-cell autonomous disease pathology contributing to the degeneration of cortical
neurons in C9orf72-FTD/ALS. To test this hypothesis, we will investigate hiPSC-derived C9orf72-FTD/ALS
cortical astrocytes in monoculture and in co-culture with cortical neurons (Aim 1). We will characterize C9orf72-
FTD/ALS hiPSC-cortical astrocytes by assessing astrocyte function and determine C9orf72 HRE-specific
pathobiology. Furthermore, we will establish the relationship between cortical astrocytes and cortical neurons
using co-cultures of control and C9orf72-FTD/ALS lines. The co-cultures will be assessed for changes in
astrocyte function, neuronal function and viability, and C9orf72-disease pathobiology. In addition, we will
examine transcriptomic alterations in the diseased hiPSC-derived cortical astrocytes (Aim 2). Transcriptomic
profiles of the diseased and control cortical astrocytes in both monoculture and co-culture conditions will be
generated using RNA-sequencing. In addition, we will analyze existing single nuclei RNA seq data already
generated in the lab and identify overlapping candidate genes that are specifically dysregulated in cortical
astrocytes in C9orf72-FTD/ALS patients and hiPSC lines. We will validate these candidate genes on the RNA
and protein level in postmortem patient tissue samples via RNAscope and immunohistochemistry, respectively.
Select validated top hits will undergo preliminary mechanistic validation through genetic manipulation of these
candidate genes in the hiPSC in vitro model. These studies will for the first time elucidate the contributing role of
cortical astrocytes in the neurodegeneration of cortical neurons in C9orf72-FTD/ALS, addressing the disease
mechanisms of dementia in this spectrum disorder. Additionally, this work will provide novel opportunities for
drug target identification with the hope of identifying novel therapeutics for the affected patient populations.