Establishment and characterization of a human cell line from paediatric cerebellar glioblastoma multiforme

Permanent glioma cell lines are invaluable tools in understanding the biolo gy of glioblastomas. The present study reports the establishment of a clona l human cell line, GBM6840, derived from a biopsy of paediatric cerebellar glioblastoma multiforme. GBM6840 had a doubling time of 32 h and grew as a monolayer of large round cells that retained immunopositivity for glial fib rillary acidic protein and vimentin. Karyotypic analysis revealed a modal c hromosome number of 68 and polysomies of chromosomes 3, 5 and 20, as well a s the presence of 3-4 marker chromosomes. GBM6840 also showed anchorage-ind ependent growth in soft agar and tumour formation in nude mice. The p16(CDK N2A) gene was transcriptionally silenced by hypermethylation, consistent wi th the lack of protein expression observed in the original tumour and cultu red cells. Western blot analysis revealed normal protein expression of pRb and CDK4. It appears that p16 is the major component altered in the cell cy cle pathway and may confer these cells unrestrained proliferation potential . Neither EGFR gene amplification nor over-expression of the protein was de tected in the cultured cells. Over-expression of the p53 protein was observ ed in the majority of cells, despite undetectable mutation (exons 5-8) in t he gene. One allele of the PTEN gene was found to be mutated during in vitr o cultivation. Telomerase activity was demonstrated in the cultured cells b ut not in the original tumour, supporting the hypothesis that telomerase is required for the in vitro immortalization process.