gastrin-releasing-peptide and Neuroectodermal-Tumors--Primitive--Peripheral

The Ewing tumor family of peripheral primitive neuroectodermal tumors (pPNETs) are characterized by chromosomal translocations leading to EWS-ETS gene fusions. These hybrid genes express chimeric proteins that are thought to act as aberrant transcription factors. We therefore used differential display-PCR to compare gene expression patterns in pPNET cell lines with those of other small round cell tumors (SRCTs) of childhood. This technique detected differential expression of sequences corresponding to human gastrin-releasing peptide (GRP) in pPNET cell lines but not in other SRCT cell lines. Subsequent Northern and reverse transcription-PCR analysis of SRCT cell lines confirmed GRP positivity in all pPNET lines tested. Of primary tumors tested by reverse transcription-PCR, GRP expression was found in 7 (44%) of 16 pPNETs but in no other primary SRCTs examined. Expression of the GRP receptor gene was demonstrable in 55% of pPNET cell lines and 25% of primary pPNET tumors but also in several other SRCTs. Radioimmunoassays and immunohistochemistry confirmed expression of bioactive GRP peptide in pPNET cell lines and primary tumors, respectively. Moreover, in vitro growth of a pPNET cell line was slowed by treatment with a GRP receptor antagonist and accelerated by a GRP receptor agonist. GRP is a known autocrine growth factor in small cell lung cancer and other neuroendocrine tumors. Its expression in pPNETs provides further evidence for a neuroectodermal histogenesis of these tumors and suggests that autocrine growth of this family of tumors may be at least partially regulated by GRP.

Topics: Artificial Gene Fusion; Base Sequence; Bone Neoplasms; Carcinoma, Small Cell; Cloning, Molecular; Gastrin-Releasing Peptide; Humans; Molecular Sequence Data; Neuroectodermal Tumors, Primitive, Peripheral; Peptides; Polymerase Chain Reaction; Protein Precursors; Receptors, Bombesin; Sarcoma, Ewing; Sarcoma, Small Cell; Tumor Cells, Cultured

Regulation of co-expression of three neuropeptide genes, i.e. genes encoding enkephalin, cholecystokinin, and gastrin-releasing peptide, was studied in human neuroepithelioma cells. In nondifferentiated state, the continuous cell line SK-N-MC displayed an equally high level of expression of the enkephalin, cholecystokinin, and gastrin-releasing peptide genes. By culturing in medium containing endothelial cell growth supplement the SK-N-MC cells differentiated morphologically into a cell type with neurite-like processes. After 3 days the expression of the enkephalin gene in endothelial cell growth supplement-differentiated cells was significantly reduced by 75% as compared to the nondifferentiated cells, while there was no change in the expression of the cholecystokinin and gastrin-releasing peptide genes during differentiation. The results show that the enkephalin gene is selectively down-regulated during differentiation of neuroepithelioma cells. It is suggested that the down-regulation is related to the transient expression of the enkephalin gene in developing brain and other organs. Thus the neuroepithelioma cell line may provide a cellular model to study the underlying molecular mechanism.

Topics: Blotting, Northern; Cell Differentiation; Cell Line; Cholecystokinin; Enkephalins; Gastrin-Releasing Peptide; Gene Expression Regulation, Neoplastic; Humans; In Vitro Techniques; Neuroectodermal Tumors, Primitive, Peripheral; Peptides; Protein Precursors; RNA, Messenger; RNA, Neoplasm

The cholinergic human neuroepithelioma cell line SK-N-MCIXC expressed mRNAs for the neuropeptides cholecystokinin (CCK), neuropeptide Y, gastrin-releasing peptide (GRP) and enkephalin. The CCK transcript of about 800 nt was present at very high levels and CCK-like peptides immunoreactive to a C-terminal CCK octapeptide antiserum were present in the cell line and its medium. This clonal neuronal cell line provides a unique model system to identify cis- and trans-acting factors responsible for neuron-specific expression and regulation of the CCK gene. Furthermore, the pluripotent properties of the undifferentiated cell line may open studies on neuronal differentiation at the level of co-expression of neuropeptides and transmitters.

Topics: Blotting, Northern; Cholecystokinin; Enkephalins; Gastrin-Releasing Peptide; Gene Expression; Humans; Neuroectodermal Tumors, Primitive, Peripheral; Neuropeptide Y; Neuropeptides; Peptides; RNA, Messenger; Tumor Cells, Cultured