Verification of Potential Glioma-Associated Antigens by Quantitative RT-PCR
Heilbrun, Noah C.
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Gliomas constitute more than 80% of the neoplasms of the adult central nervous system (Chakravarti, et al. 2001). In spite of advances in cancer research over the past twenty years, the diagnosis of glioma carries with it limited treatment options and poor prognosis (Mahler, et al. 2001). Emerging therapies, such as gene therapy and immunotherapy, seek to take advantage of our expanding molecular and genetic understanding which has accompanied the sequencing of the human genome. In house computer-generated serial analysis of gene expression (SAGE) data indicate several glioma-associated proteins have immunotherapeutic potential (Sloan, et al. in preparation). This study sought to verify the presence of 14 of these potential antigens in glioblastoma multiforme and oligodendroglioma tumors using gene-specific primers (GSPs) and quantitative reverse-transcription PCR (RT-PCR). Data collected during PCR was validated using native polyacrylamide gel electrophoresis (PAGE). Although quantitative PCR results indicated elevated expression levels of all fourteen selected genes across both tumor types, conclusions could not be reached. Ectopic products were observed upon PAGE analysis of RT-PCR products, either drowning out any signal from the expected products or accounting for the entire observed signal collected during the amplification process, indicating experimental conditions of the quantitative PCR reactions must be further optimized. Results from the PAGE study clearly indicated the use of a high-resolution separation method to accurately analyze and verify quantitative PCR products is a necessity.