Gene dosage and mutational analyses of EGFR in oligodendrogliomas.
We have studied amplification/gene-dosage and sequence variations of the EGFR gene in 41 oligodendroglial tumours graded according to the WHO classification (21 oligodendrogliomas grade II, 13 oligodendrogliomas grade III and 6 oligoastrocytomas grade II-III), using multiplex ligation-dependent probe amplification (MLPA), real-time quantitative PCR, and PCR/SSCP techniques. To determine gene-dose we studied exons 11 (extracellular domain) and 25 (intracellular domain) in the EGFR gene. Overdose (1- to 5-fold increase) was present in exon 11 in 21 of 41 samples (52.5% of cases) and in exon 25, in 7 of 41 samples (17.5% of cases). Gene amplification > 5-fold increase) was present in exon 11, in 17 of 41 samples (42.5% of cases), and in exon 25 in 6 of 41 samples (15% of cases). Three tumours (two grade II oligodendrogliglioma, one mixed oligoastrocytoma) displayed high level amplifications: > 100 gene copies were identified by both real-time quantitative PCR and MLPA analyses. Gene sequence alterations were identified by PCR/SSCP and sequencing in four cases: two missense mutations: G1051A (Ala351Thr) and G2216A (Arg739Hys); one nonsense mutation: C2934T (Asp978Asp); and an 18 bp deletion in position 2423-2441 of E19. These changes were present only in tumoral DNA, not in the corresponding constitutional patients' DNA. We also found four previously unidentified polymorphic variants: G2025A (Ala675Ala), C2233T (Leu745Leu), C2895T (Treo965Treo) and C3168T (Asp1056Asp), and three previously described polymorphic changes: E12+22 Tright curved arrow A, G1748A (Arg583Lys) and A2547T (Glu849Glu). Our findings demonstrate that mutations and amplification/overdose in the EGFR gene are present in low-grade oligodendroglial tumours, and may contribute to the development of these brain neoplasms.
Franco-Hernandez
Carmen
C
Unidad de Investigación, Laboratorio de Oncogenética Molecular, Hospital Universitario La Paz, 28046 Madrid, Spain.
Martinez-Glez
Victor
V
Alonso
M Eva
ME
De Campos
Jose M
JM
Isla
Alberto
A
Vaquero
Jesus
J
Gutierrez
Manuel
M
Rey
Juan A
JA
D019943:Amino Acid Substitution; D001483:Base Sequence; D001706:Biopsy; D001932:Brain Neoplasms / Q000235:genetics* / Q000473:pathology / Q000601:surgery; D004252:DNA Mutational Analysis; D017931:DNA Primers; D066246:ErbB Receptors / Q000235:genetics*; D005091:Exons; D005784:Gene Amplification; D018628:Gene Dosage; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D009837:Oligodendroglioma / Q000235:genetics* / Q000473:pathology / Q000601:surgery; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D018807:Polymorphism, Single-Stranded Conformational; D017384:Sequence Deletion
D017931:DNA Primers; D066246:ErbB Receptors