Expression of growth factor receptors in human brain tumours.

Nl Tuzi,Sl Staddon,Dj Venter,Nr Lemoine,Wj Gullick,S Kumar

doi:10.1038/bjc.1991.54

Abstract

The expression of the EGF receptor, c-erbB-2 and PDGF receptor proteins has been studied in a series of human brain tumour biopsies and cell lines. Western blotting was used to determine the amount of protein present and their intrinsic and ligand promoted enzyme activities were studied by immunoprecipitation followed by autophosphorylation. EGF receptors were found to be expressed at very high levels in 40% of primary tumour biopsies, but at uniformly low levels in tumour derived cell lines. The c-erbB-2 protein was not detected in tumour biopsies, but was present at variable, but low levels in extracts of tumour cell lines. PDGF receptors were also found at moderate to low levels in both primary tumours and cell lines. The EGF receptor gene was amplified in four out of 14 primary tumours and this generally correlated with high levels of protein expression. The c-erbB-2 gene was not amplified. Employing the polymerase chain reaction and sequence specific oligonucleotides as probes there was no evidence of mutations in the c-erbB-2 gene transmembrane region. These results suggest that alterations of expression of the EGF receptor may play a role in human brain tumours. There was however no evidence for aberrant expression of the c-erbB-2 protein. Additional experiments are required to assess the influence of PDGF receptor expression in brain tumour cells.

Highlights

0.1 g of fresh biopsy material was placed in a sterile tube containing Dulbecco's minimum essential medium (DMEM), and an explant culture set up from each sample as follows: The tumour sample was finely minced with a scalpel and trypsinised to further dissociate the cells
In extracts of 14 primary tumours very variable levels of receptor protein expression was observed in the different samples by Western blotting employing either an antipeptide antibody to the receptor's ATP binding site (ISE, Gullick et al, 1986a, data not shown) or an antibody to a region between the kinase domain and the autophosphorylation site domain (2E, Gullick et al, 1986b) (Figure 1)
In this work we confirm that the EGF receptor gene is commonly amplified in glioblastomas of high grade leading to greatly elevated levels of receptor expression

Summary

Methods

Collection, storage and characterisation of brain tumours and cell linesHuman gliomas were removed at surgery and the majority of the sample was immediately snap-frozen in liquid nitrogen and stored until analysis. Collection, storage and characterisation of brain tumours and cell lines. A proportion of the cultured cells were frozen and stored in liquid nitrogen at every 2nd passage. Cells lines were characterised at passage I or 2 to confirm that they contained cells expressing glial markers. Cell lines were employed for analysing receptor expression after between eight and 20 passages, depending on the particular cell line. None of these cell lines were cloned, they were established and the vast majority of the tumour cells in each culture exhibited a similar morphology at the time of lysis

Results

Discussion

Conclusion