Molecular detection of EWS-FLI1 chimeric transcripts in Ewing family tumors by nested reverse transcription-polymerase chain reaction: application to archival paraffin-embedded tumor tissues.
Chromosomal translocations generating unique chimeric genes are highly characteristic of specific sarcomas, and their use as diagnostic markers has been suggested. From a diagnostic pathologic point of view, detection of such cytogenetic or molecular aberrations applicable to routinely processed archival tissue specimens is considered a powerful tool for tumor diagnosis. To assess the feasibility and reliability of the molecular detection of the transcript originating from the chimeric gene in paraffin-embedded tumor specimens, we performed a nested reverse transcription-polymerase chain reaction (RT-PCR)-based assay to detect the EWS-FLI1 chimeric message in a series of Ewing family tumors. Of 24 paraffin-embedded tumor specimens from 23 cases analyzed, the chimeric message was detectable in 20 (83%) specimens from 20 cases (87%) by this nested RT-PCR assay, whereas none of 7 small round cell tumors not from this family (3 alveolar rhabdomyosarcomas, 2 neuroblastomas, 2 malignant lymphomas) showed detectable chimeric messages. In the sequence analysis of the PCR products, the amplified chimeric messages contained the junctions between exon 7 of the EWS gene and any one of exons 5, 6 and 8 of the FLI1 gene. The detection process was usually completed within 3 days, except for the subseqent sequence analysis. Our results endorse the use of this molecular assay as an ancillary technique in the diagnosis of Ewing family tumors using paraffin-embedded material.
Hisaoka
M
M
Department of Pathology and Oncology, School of Medicine, University of Occupational and Environmental Health, Kitakyushu, Japan.
Tsuji
S
S
Morimitsu
Y
Y
Hashimoto
H
H
Shimajiri
S
S
Komiya
S
S
Ushijima
M
M
D000293:Adolescent; D000328:Adult; D000368:Aged; D001859:Bone Neoplasms / Q000235:genetics*; D002648:Child; D002675:Child, Preschool; D002880:Chromosomes, Human, Pair 11; D002892:Chromosomes, Human, Pair 22; D005091:Exons; D005260:Female; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007223:Infant; D008297:Male; D008875:Middle Aged; D015514:Oncogene Proteins, Fusion / Q000032:analysis / Q000235:genetics*; D010232:Paraffin; D016133:Polymerase Chain Reaction; D051789:Proto-Oncogene Protein c-fli-1; D034802:RNA-Binding Protein EWS; D012512:Sarcoma, Ewing / Q000235:genetics*; D017422:Sequence Analysis, DNA; D016610:Tissue Embedding; D014157:Transcription Factors / Q000032:analysis / Q000235:genetics*; D014178:Translocation, Genetic
C106031:EWS-FLI fusion protein; D015514:Oncogene Proteins, Fusion; D051789:Proto-Oncogene Protein c-fli-1; D034802:RNA-Binding Protein EWS; D014157:Transcription Factors; D010232:Paraffin