156 Abstracts RECURRENT I;17 TRANSLOCATIONS IN HUMAN NEUROBLASTOMA RE- VEAL NON-HOMOLOGOUS MITOTIC RECOMBINATION DURING THE S/G2 PHASE AS A NOVEL MECHANISM FOR LOH. Huib Caron, Peter van Sluis, Nadine van Roy, Jan de Kraker, Frank Speleman, P.A. Vofite, Andries Westerveld, Rosalyn Slater, Rogier Versteeg. Dept.Human Genetics, Univ. Amsterdam, Meibergdreef 15, 1105 AZ AMSTERDAM, The Netherlands. Neuroblastomas often show loss of heterozygosity of chromosome Ip36 (LOH Ip), probably reflecting loss of a tumor suppressor gene. Here we describe three neuroblastoma tumors and two cell lines in which LOH Ip results from an unbalanced translocation between chromosomes Ip and 17q. Southern blot and cytogeoetic analyses shows that in all cases the chromosome 17 homologue from which the I;17 trans- locatlon was derived is still present and intact. This suggests a model where a tranelocation between chromosome Ip and 17q takes place in the S/G2 phase of the cell cycle, resulting in LOH Ip36. This novel mechanism of LOH commonly occurs in neuroblastoma. ONCOGENE AND ANTIONCOGENE FROM THE 22Q12 REGION INVOLVED IN NEUROECTODERMAL TUMOURS G. Thom~*, O. Delattre, J. Zucman, P. Merci, P. Dejong, G. Rouleau, & A. Aurias. *Laboratory of Tumour Genetics, Institut Curie, 26 me d'Ulm, 7.5231 PARIS Cedex 05, FRANCE. The possible occurrence of an oncogenic event in the 22q12 region was first indicated by a recurrent t(11 ;22)(q23,q12) found in Ewing sarcoma (ES), and related neurocctodermaltumours. Second, a t(12;22)(q13,q12) translocation recurrently reported in malignant melanoma of soft parts (MMSP) suggested another oncogenic event which might involve a similar mechanism. Third, germ line alterations associated with the tumour predisposing disease ueurofibromatosis type 2 (NF2) was shown by genetic linkage to reside in 22q12. In order to contribute to the ideatification of the 22q12 genes which may be involved in these processes, a physical map of part of the q12 region was performed and a contiguous regiou of about 1.5 megabase was cloned. Several new genes were identified. One of them, called EWS, encodes a protein with an N- terminal domain which shares distant homology with the C-terminal domain of eukaryotic RNA polymerase 2. The C-terminal half of EWS contains an RNA binding homologous domain. In ES and PN, EWS forms a hybrid gene with either FLI-I (90% of cases) or ERG (10% of cases), two transcription factors of the ETS family. In MMSP, EWS forms a hybrid gene with ATF-1, a transcription factor belonging to the b-zip family. In all cases, the encoded chimeric protein links the N terminal domain of EWS to the DNA bindiug domalu of one of the previous transcription factors. These new oncogenes ,-are believed to act by directly interfering with uanscription regulation. Another gene, called SCII, eucodes a prolein tenned sehwannomin, which demonstrates strong homology with a group of proteill possibly linking the cytoskeleton to membrane proteius. SCIt is the site of germ liue mutation in NF2 patients and of somatic mutatiou in schwanuoma and meningiolna, two tumours known to be predisposed by NF2. Analysis of the SCIf mutation spectrum shows that the mutations predominandy cause the synthesis of a truncated schwannomin protein. Combined with the monitoring of the chromosome 22 allelic status in the tamours, these data indicate that SCH is a tumour suppressor gene. DNA AMPLIFICATION IN HUMAN GLIOMAS : A PARALLEL STUDY BY MODIFIED CGH, FISH AND SOUTHERN METHODS Anna Almeida, Marline Muleris, Anne-Marie Dutrillaux, Bernard Dutrillaux, Bernard Malfoy CNRS URA 620, Institut Curie, Section de Biologie, Paris, France. Gliomas are the most common primary brain tumors and constitute a heterogeneous group in which glioblaslomas are the most malignant. The first evidence of amplification in these tumors came from cytogenetic observations of double minutes chromosomes (dmin) or of homogeneous staining regions (hsr). Four genes have been identified as being recurrently amplified : MYCN, GLI, PDGFRA and EGFR. EGFR is amplified in about 40 % of the glioblastomas. Genomic amplification in nine xenogralted gliomas have been studied. In two cases, the primary tumor was also analysed. Cytogenetic studies showed the presence of dmins in 8 cases and of an hsr in 1 case. The results obtained by a modified comparative genomic hybridization technique (see accompagning communication, Muleris et al.) has been confirmed by in situ hybridization of the suspected amplified sequences, chromosome painting on tumor spreads and gene copy analysis by Southern blot. In some cases, several non-syntenic loci are amplified in the same cell. Bands 7p11, 8q24 and 4q12 corresponding to EGFR, MYC and PDGFRA respectively are involved in these amplifications. However, other chromosomal sites are also found amplified for which candidate genes are not yet known. NEUROBLASTOMA CELLS NEED NORMAL CELLS FOR COMPLETE ORGANOGENIC MATURATION. Inge M. Ambros. Andrea Zellner, Helmut Gadner and Peter F. Ambros. CCRI, St. Anna Kinderspital, A-1090 Vienna (I.MA, A.Z, H.G, P.F.A). Inst. f. Clinical Pathology, Univ. Vienna (LM.A). Neuroblastomas (NBs) are the most heterogeneous neoplastic diseases of infancy and early childhood in terms of clinical behavior, genetic constellation and histomorphology. Maturing stroma-rich NBs, which show a benign clinical behavior, consist of two main cell populations, the ganglionic and the Schwann cells (SC). These cell populations are thought to be derived from a common neoplastic precursor cell, i.e. a pluripotent neural crest cell. Interphase cytogenetic methods are independent from proliferation activity, and, if carried out on histological sections, these techniques enable us for the first time to investigate tumor and normal cells in their histological context. We analyzed 20 formalin-fixed paraffin-embedded NBs which showed spontaneous organogenic maturation, i.e. ganglionic differentiation with the developement of a SC stroma. Up to now these tumors escaped cytogenetic analysis due to lack of proliferation activity. In order to detect numerical and structural aberrations of chromosome 1 and to assign these aberrations to individual, morphologically and immunohistochemically defined cells, we used the centromere-specific probe D1Z1 and the probe D1Z2, specific for the chromosomal region I p36.3. In addition flow cytometric measurements were carried out. The 20 maturing or mature SC-rich NBs showed a triploid DNA content and in none of the tumors a deletion of the short ann of chromosome 1 was found. Furthermore, we show that the numerical chromosomal changes are restricted to the ganglionic cells, and that SCs exhibit a normal diploid karyotype. This finding characterizes the NB associated SC as a normal cell and is incompatible with the current concept that NBs originate from a pluripotent neural crest cell. The SCs obviously have an antiproliferative and differentiation inducing effect on the tumor cells. Moreover, stroma-rich NBs belong to only one of two genetically defined subgroups, i.e. the triploid NBs with the intact short arm of chromosome 1 representing one of the prerequisites for complete maturation.