Somatic Mutations in TSC1 and TSC2 Cause Focal Cortical Dysplasia

Abstract

Focal cortical dysplasia (FCD) is a major cause of the sporadic form of intractable focal epilepsies that require surgical treatment. It has recently been reported that brain somatic mutations in MTOR account for 15%-25% of FCD type II (FCDII), characterized by cortical dyslamination and dysmorphic neurons. However, the genetic etiologies of FCDII-affected individuals who lack the MTOR mutation remain unclear. Here, we performed deep hybrid capture and amplicon sequencing (read depth of 1003-20,0123) of five important mTOR pathway genes-PIK3CA, PIK3R2, AKT3, TSC1, and TSC2-by using paired brain and saliva samples from 40 FCDII individuals negative for MTOR mutations. We found that 5 of 40 individuals (12.5%) had brain somatic mutations in TSC1 (c.64C>T [p.Arg22Trp] and c.610C>T [p.Arg204Cys]) and TSC2 (c.4639G>A [p.Val1547Ile]), and these results were reproducible on two different sequencing platforms. All identified mutations induced hyperactivation of the mTOR pathway by disrupting the formation or function of the TSC1-TSC2 complex. Furthermore, in utero CRISPR-Cas9-mediated genome editing of Tsc1 or Tsc2 induced the development of spontaneous behavioral seizures, as well as cytomegalic neurons and cortical dyslamination. These results show that brain somatic mutations in TSC1 and TSC2 cause FCD and that in utero application of the CRISPR-Cas9 system is useful for generating neurodevelopmental disease models of somatic mutations in the brain.

Somatic Mutations Activating the mTOR Pathway in Dorsal Telencephalic Progenitors Cause a Continuum of Cortical Dysplasias

“…While we acknowledge the age-dependence of syndromic TSC manifestations, based on available records and specialized pediatric neurology examinations, both patients lacked any signs of syndromic TSC, and brain MRIs did not show other TSC hallmarks. These findings are remarkable since TSC mutations have not previously been associated with isolated HME, and since capturing both germline and somatic mutations in TSC-associated cortical dysplasia is uncommon (Lim et al, 2017). The percentage of cells carrying the pathogenic mutations ranged from 6.2–41.2% for the eight patients with HME and identified somatic mutations (Figure 1A).…”

“…We did not identify germline mutations in TSC1 or TSC2 in either of these cases, further suggesting that both patients had isolated FCD and not syndromic TSC. Somatic TSC mutation without detectable germline mutation has recently been reported in FCD (Lim et al, 2017). …”

“…Similarly, a second hit has only once been identified in a FCD patient who carried germline and somatic mutations in DEPDC5 (Baulac et al, 2015). Curiously, a recent report of somatic mutations in TSC1 and TSC2 in FCD cases failed to identify associated germline mutations (Lim et al, 2017), which would seem in principle easier to detect. We demonstrate germline and somatic mutations 1) in TSC2 from non-TSC patients and 2) associated with HME.…”

“…Several new genes have also been described that act analogously to TSC1 and TSC2 as negative regulators of MTOR , causing FCD and HME via germline loss-of-function mutation coupled to demonstrated or inferred somatic loss of the second allele (Baulac et al, 2015; D’Gama et al, 2015; Lim et al, 2017; Scheffer et al, 2014; Sim et al, 2016; Weckhuysen et al, 2016). Surprisingly, TSC mutations can cause isolated FCD or HME in the absence of widespread hamartomas (D’Gama et al, 2015; Hoelz et al, 2017; Lim et al, 2017). …”

Hypervascularization in mTOR‐dependent focal and global cortical malformations displays differential rapamycin sensitivity

“…[1][2][3] A large subset of MCDs result from de novo somatic mutations occurring during brain development affecting mammalian target of rapamycin (mTOR) pathway genes [4][5][6][7][8][9][10][11][12][13] and share similar cytoarchitectural abnormalities. [1][2][3] A large subset of MCDs result from de novo somatic mutations occurring during brain development affecting mammalian target of rapamycin (mTOR) pathway genes [4][5][6][7][8][9][10][11][12][13] and share similar cytoarchitectural abnormalities.…”