Abstracts

The [italic]GLI3[/italic] gene (MIM 165240) encodes a DNA-binding transcription factor involved in downstream regulation of Sonic Hedgehog-mediated CNS development. Truncating (frameshift and nonsense) mutations located 3-prime of the zinc finger-encoding regions of [italic]GLI3[/italic] cause familial Pallister-Hall syndrome (PHS; MIM 146510), a rare entity characterized by hypothalamic hamartoma, central polydactyly and other malformations. Hypothalamic hamartoma also occurs as an isolated CNS malformation in patients with an increasingly recognized clinical presentation incorporating early onset gelastic seizures and often leading to secondary generalized epilepsy. We aimed to investigate the role of [italic]GLI3[/italic] mutation in the etiology of non-syndromal (sporadic) hypothalamic hamartoma associated with epilepsy.
24 patients with hypothalamic hamartoma and refractory epilepsy were studied; none met clinical criteria for PHS. Whole blood samples were obtained and DNA extracted using the QIAGEN Blood DNA Kit. In 12 patients DNA from hypothalamic hamartoma tissue collected during transcallosal resection was extracted using DNAzol (Invitrogen). Single-stranded conformation polymorphism screening (SSCP) of samples was performed: HEX-labeled primers were designed to amplify all 14 exons of [italic]GLI3[/italic]; a 30ng sample of patient DNA was amplified in a total volume of 10[micro]l; products were separated on nondenaturing 4% polyacrylamide gels containing 2% glycerol (GelScan 2000; Corbett Research). PCR products showing conformational changes were re-amplified, using unlabelled primers, from 100ng of genomic DNA, and then sequenced (ABI PRISM BigDye Terminator v3.0 Ready Reaction Cycle Sequencing Kit; Perkin Elmer).
No[italic] GLI3[/italic] mutations were found in peripheral blood samples. In hypothalamic hamartoma tissue of one patient, a truncating mutation (Gln926Stop) was found in the CBP-binding domain of[italic] GLI3[/italic] that was absent in the blood sample. Although several gene polymorphisms were detected, disease-causing mutations were not identified in the other patients by SSCP, but may be revealed by sequencing experiments, currently underway.
The location and type of the[italic] GLI3[/italic] mutation found in the hamartoma of our patient is similar to that described in some families with PHS, strongly suggesting that sporadic hypothalamic hamartomas can be due to somatic mutations in the Sonic Hedgehog pathway. This raises the possibility that other [quot]sporadic[quot] malformations of cortical development may be due to somatic mutations.