By screening a meningioma expression library with autologous serum we identified four cDNA clones representing a novel gene with striking homology to Caenorhabditis elegans hyaluronidase as indicated by BLASTP analysis. In humans hyaluronidase has been implicated in cancer development and three human genes are known to encode proteins with hyaluronidase activity. None of the human genes, however, showed any homology at the nucleotide or amino acid sequence level to the newly isolated antigen we termed meningioma expressed antigen 5 (MGEA5). Somatic cell hybrid mapping and fluorescence in situ hybridization mapped the gene for MGEA5 to chromosomal band 10q24.1-q24.3. Reverse transcription (RT)-PCR and northern blot hybridization revealed expression of the gene encoding MGEA5 in several meningioma and additional human tissues. Expression analysis also indicated an alternative splicing event giving rise to a shorter and altered transcript termed MGEA5s. The expression of MGEA5 and MGEA5s as fusion proteins revealed an approximate molecular weight of 92 and 54 kDa, respectively. Using heterologous sera we found antibodies against MGEA5s in five out of 23 meningioma patients, whereas no immune reaction was detected in 12 control sera from healthy individuals. Confirmation of hyaluronidase activity was independently achieved by turbidometric analysis and a gel matrix assay. A model for involvement of the novel hyaluronidase gene in meningioma development is proposed.

Gene amplification, which is generally considered to occur late in tumor development, is a common feature of high grade glioma. Up until now, there have been no reports on amplification in astrocytoma grade I. In this study, we report cloning and sequencing of a cDNA termed glioma-amplified sequence (GAS41) which was identified recently in a glioblastoma cell line by microdissection-mediated cDNA capture. This technique is tailored to isolate amplified genes from human tumors. An increased copy number of GAS41 was found in glioblastoma multiforme and astrocytoma III, and at a high frequency in astrocytoma grades I and II. Sequence comparison indicates a high homology between the GAS41 protein, the yeast and human AF-9 and the human ENL proteins. Both AF-9 and ENL belong to a new class of transcription factors, indicating that GAS41 might also represent a transcription factor. With GAS41 being the first gene found with increased copy number in low grade glioma, this study provides the first evidence that gene amplification can occur in early tumor development.

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There is increasing evidence that tumor expressed genes induce immune responses in cancer patients. To identify meningioma expressed antigens, we established a meningioma expression library which was screened with autologous serum. Out of 20 positive cDNA clones eight share high sequence homologies as determined by sequence analysis. These eight clones can be grouped into three classes which differ in length and which are characterized by specific sequence variations. The longest open reading frame was found to be 2412 bp encoding an immunoreactive antigen termed meningioma expressed antigen 6 (MEA6). Using five sequence specific primer pairs, somatic hybrid panel mapping revealed locations of the three classes on several human chromosomes including chromosomes 2, 3, 6, 7, 9, 13 and 14. The mapping results were confirmed by fluorescence in situ hybridization. RT-PCR showed consistent expression of all classes in several meningiomas and additional tissues using the same set of primer pairs as for chromosomal mapping. The expression data were confirmed by northern blot analysis. For the predicted amino acid sequence BLASTX revealed a homology to a human C219-reactive peptide which was previously isolated by an antibody directed against p-glycoprotein. Sequence properties of the MEA protein include an acidic activation domain, a proline-rich region and two coiled-coil domains indicating protein binding and activation functions.

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