Abstract
Epilepsy is a major health burden, calling for new mechanistic insights and therapies. CRISPR-mediated gene editing shows promise to cure genetic pathologies, although hitherto it has mostly been applied ex vivo. Its translational potential for treating non-genetic pathologies is still unexplored. Furthermore, neurological diseases represent an important challenge for the application of CRISPR, because of the need in many cases to manipulate gene function of neurons in situ. A variant of CRISPR, CRISPRa, offers the possibility to modulate the expression of endogenous genes by directly targeting their promoters. We asked if this strategy can effectively treat acquired focal epilepsy, focusing on ion channels because their manipulation is known be effective in changing network hyperactivity and hypersynchronziation. We applied a doxycycline-inducible CRISPRa technology to increase the expression of the potassium channel gene Kcna1 (encoding Kv1.1) in mouse hippocampal excitatory neurons. CRISPRa-mediated Kv1.1 upregulation led to a substantial decrease in neuronal excitability. Continuous video-EEG telemetry showed that AAV9-mediated delivery of CRISPRa, upon doxycycline administration, decreased spontaneous generalized tonic-clonic seizures in a model of temporal lobe epilepsy, and rescued cognitive impairment and transcriptomic alterations associated with chronic epilepsy. The focal treatment minimizes concerns about off-target effects in other organs and brain areas. This study provides the proof-of-principle for a translational CRISPR-based approach to treat neurological diseases characterized by abnormal circuit excitability.
Highlights
Epilepsy affects up to 1% of the population, and 30% of patients continue to experience seizures despite the use of current medication (Kwan et al, 2011; Tang et al, 2017)
We first asked if CRISPR activation (CRISPRa) can be exploited to increase endogenous gene expression in glutamatergic neurons and decrease their excitability
We have shown that CRISPRa can be used to increase endogenous Kcna1 expression to modulate neuronal activity, to reduce seizure initiation and to rescue behavioural and transcriptomic abnormalities in a mouse model of chronic temporal lobe epilepsy
Summary
Epilepsy affects up to 1% of the population, and 30% of patients continue to experience seizures despite the use of current medication (Kwan et al, 2011; Tang et al, 2017). Current experimental gene therapies mainly rely on viral vector-mediated expression of genes encoding normal CNS proteins or exogenous non-mammalian proteins (Richichi et al, 2004; Noe et al, 2008; Wykes et al, 2012, 2016; Krook-Magnuson et al, 2013; Katzel et al, 2014; Lieb et al, 2018; Agostinho et al, 2019) This approach has several potential limitations, including a finite packaging capacity of viral vectors, difficulty in ensuring normal splicing and post-transcriptional processing, and, in the case of nonmammalian membrane proteins, concerns about immunogenicity. Modulating the expression of endogenous genes, in contrast, would represent an important step toward safe and rational treatment of intractable epilepsy and other neurological diseases
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