DETECTION OF MYCN GENE AMPLIFICATION AND DELETIONS OF CHROMOSOME 1P IN NEUROBLASTOMA BY IN-SITU HYBRIDIZATION USING ROUTINE HISTOLOGIC SECTIONS

BACKGROUND: Amplification of the MYCN oncogene and partial deletion of chromosome 1p are genetic changes frequently seen in neuroblastoma th at are indicators of poor prognosis. The identification techniques usu ally used-Southern blotting for MYCN gene amplification, and karyotypi c analysis of viable tumor cells for large 1p deletions-are time-consu ming and suited for specialized laboratories. EXPERIMENTAL DESIGN: We have developed an immunofluorescence in situ hybridization assay suita ble for detection of MYCN amplification and lp deletion in formalin-fi xed, paraffin-embedded tissue sections. The technique is rapid, does n ot involve the use of radioactivity, and can be carried out in laborat ories already familiar with conventional in situ hybridization and imm unohistochemical detection methods. RESULTS: MYCN gene amplification a ppeared as multiple signals per nucleus, corresponding to double minut e chromosomes. Deletion of 1p was detected by loss of one of the norma l signals present within the nucleus of a neuroblastoma cell line. Use of different fluorophores enabled the simultaneous detection of MYCN gene amplification and 1p deletion at the individual cell level. Detec tion was found to be enhanced by RNase and pepsin treatment of tissue sections before hybridization and by a novel microwave denaturation me thod. CONCLUSIONS: Application of this methodology to formalin-fixed s amples of neuroblastoma will permit comprehensive retrospective studie s of these two genetic markers using archived tumors. In situ hybridiz ation surveys will have widespread applications for studying known gen etic aberrations in individual cells of a variety of solid tumors and for determining interrelationships and clinical significance in relati on to tumor progression and patient outcome.