Faculty Bibliography

By screening a human fetal brain cDNA expression library using a monoclonal antiphosphotyrosine antibody and by 5' RACE procedures, we have isolated overlapping cDNAs encoding a receptor-type tyrosine kinase belonging to the EPH family, DRT (Developmentally Regulated EPH-related Tyrosine kinase gene). The DRT gene is expressed in three different size transcripts (i.e. 4, 5 and 11 kb). DRT transcripts are expressed in human brain and several other tissues, including heart, lung, kidney, placenta, pancreas, liver and skeletal muscle, but the 11 kb DRT transcript is preferentially expressed in fetal brain. Steady-state levels of DRT mRNA in several tissues, including brain, heart, lung and kidney, are greater in the midterm fetus than those in the adult. DRT transcripts are detectable at low levels in a human teratocarcinoma cell line (NTera-2), but its expression is greatly increased after the NTera-2 cells are induced to become postmitotic neurons (NTera-2N) by retinoic acid treatment. These data suggest that DRT plays a part in human neurogenesis. A large number of tumor cell lines derived from neuroectoderm express DRT transcripts, including 12 neuroblastomas, two medulloblastomas, one primitive neuroectodermal tumor and six small cell lung carcinomas (SCLC). Interestingly, several neuroblastoma cell lines with 1p deletion and one SCLC cell line express DRT transcripts of aberrant size (i.e. 3, 6 and 8 kb) in addition to those found in normal tissues. We mapped the DRT gene to human chromosome 1p35-1p36.1 by PCR screening of human-rodent somatic cell hybrid panels and by fluorescence in situ hybridization. As the distal end of chromosome 1p is often deleted in neuroblastomas and altered in some cases in SCLCs, these chromosomal abnormalities may have resulted in the generation of aberrant size transcripts. Thus, the DRT gene may play a part in neuroblastoma and SCLC tumorigenesis.

We analyzed 156 primary neuroblastoma tumor samples for loss of heterozygosity at the distal short arm of chromosome 1 (1p LOH). We also compared 1p LOH with known clinical and genetic prognostic variables as well as patient outcome. 1p LOH was detected in 30 of 156 tumors (19%) and was strongly associated with adverse clinical and biological features. 1p LOH was also strongly predictive of a poor outcome in univariate analyses (estimated 4-year survival, 32 +/- 10% SE versus 76 +/- 5% SE; P < 0.001). However, the prognostic value of 1p LOH was equivocal when stratified for amplification of the MYCN oncogene (P = 0.16). We conclude that 1p LOH is an important component of a pattern of genetic abnormalities in neuroblastoma associated with an aggressive clinical course.

Mutations of CDKN2 have been found recently in melanoma and many other tumor types. Neuroblastoma shares with melanoma a neuroectodermal origin and a high incidence of deletions of the short arm of chromosome 1. Therefore, we analyzed 18 primary neuroblastomas and 9 tumor-derived cell lines for mutations in CDKN2. We used PCR-single-strand conformation polymorphism to examine exons 1 and 2 of the CDKN2 gene for mutations, but none were detected. Furthermore, no homozygous deletions were detected and there was no loss of heterozygosity at the closely linked IFNA locus. We conclude that disruption of the CDKN2 gene is not required for malignant transformation of human neuroblastomas.