Analysis of TSC Cortical Tubers by Deep Sequencing of TSC1, TSC2 and KRAS Demonstrates that Small Second-Hit Mutations in these Genes are Rare Events

Translational Medicine Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Brain Pathology (Impact Factor: 3.84). 11/2010; 20(6):1096-105. DOI: 10.1111/j.1750-3639.2010.00416.x
Source: PubMed


Tuberous sclerosis complex (TSC) is an often severe neurocutaneous syndrome. Cortical tubers are the predominant neuropathological finding in TSC, and their number and location has been shown to correlate roughly with the severity of neurologic features in TSC. Past studies have shown that genomic deletion events in TSC1 or TSC2 are very rare in tubers, and suggested the potential involvement of the MAPK pathway in their pathogenesis. We used deep sequencing to assess all coding exons of TSC1 and TSC2, and the activating mutation hot spots within KRAS in 46 tubers from TSC patients. Germline heterozygous mutations were identified in 81% of tubers. The same secondary mutation in TSC2 was identified in six tuber samples from one individual. Further study showed that this second hit mutation was widely distributed in the cortex from one cerebral hemisphere of this individual at frequencies up to 10%. No other secondary mutations were found in the other 40 tubers analyzed. These data indicate that small second hit mutations in any of these three genes are very rare in TSC tubers. However, in one TSC individual, a second hit TSC2 point mutation occurred early during brain development, and likely contributed to tuber formation.

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    • "LOH has been convincingly demonstrated in most of the neoplasms associated with TSC including subependymal giant cell astrocytomas (SEGAs), angiomyolipomas, cardiac rhabdomyomas, and lymphangioleiomyomatosis (LAM). Biallelic inactivation has been detected, but in only a small number of cells within cortical tubers (Crino et al., 2010; Qin et al., 2010). "
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    • "This may be because biallelic mutation is necessary to induce severe morphological alteration such as tubers or giant astrocytomas. In fact, so-called “two-hit” mechanisms of germline mutation and somatic mutation have been proposed for human tubers of TSC patients (Crino et al., 2010), although another report argued that the second hit was rare (Qin et al., 2010). The two-hit mechanism has been evaluated with homozygous deletion of TSC2 only in radial glia, and heterozygous deletion in all other cells by crossing Tsc2+/-, Tsc2flox, and GFAP-Cre mice (Way et al., 2009). "
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    • "Other neurodevelopmental disorders occasionally result from somatic mutations, including DCX (Gleeson et al., 2000) and NF1 (Messiaen et al., 2011; Vogt et al., 2011), but these have generally not been mutations limited to brain. In addition, some tubers from individuals with TSC, who carry germline mutations in TSC1 or TSC2, have been demonstrated to have somatic " second hits' in the TSC2 gene (Qin et al., 2010). We hypothesized that the somatic mutations causing HMG might be essentially restricted to the brain and detectable by direct study of affected brain tissue. "
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