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Using the recently developed technique of digital karyotyping, we performed a high resolution genome-wide study of medulloblastoma specimens and found specific genetic changes that are expected to be an important key to unraveling the molecular pathogenesis of these tumors. A 28-fold amplification located on chromosome 14q22.3 was identified from a medulloblastoma cell line. A single gene, a developmentally regulated gene, orthodenticle homolog-2, or OTX2, was identified in the amplified segment. OTX2 gene amplification was also identified in primary medulloblastoma specimens by fluorescence in situ hybridization and real-time quantitative PCR. OTX2 is expressed during cerebellum development and shut down in the normal adult tissues. We observed that OTX2 was exclusively expressed at high levels in >50% medulloblastomas by performing real-time quantitative PCR, serial analysis of gene expression (SAGE) and immunohistochemistry on clinical samples. This indicates that a mis-programming of OTX2 expression drives the medulloblastoma tumorigenesis. To determine that OTX2 promotes medulloblastoma cell proliferation, we transfected medulloblastoma cell lines and a non neoplastic cell line with OTX2 expression vector and found that OTX2 significantly enhanced the cell proliferation. To further determine whether OTX2 expression is essential for medulloblastoma cell survival, we disrupted OTX2 expression in multiple medulloblastoma cells using OTX2 gene- specific siRNAs against OTX2. Repression of OTX2 expression by siRNAs inhibited cell growth in the OTX2-expressing cell lines, whereas there was no effect using the same treatment on the OTX2-nonexpressing cell line. Through a combination of comprehensive genomic, expression and function studies, we identified OTX2 as an oncogene in medulloblastomas. This study will advance our knowledge of the pathogenesis of medulloblastoma which could make significant contributions to the molecular classification and therapeutic intervention in this frequently lethal pediatric cancer.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]