To identify genes that might be involved in the development and growth of medulloblastoma, rhabdomyosarcoma or both, we made Serial Analysis of Gene Expression (SAGE) libraries and compared the expression profiles of the tumors with those of normal tissue. In medulloblastoma we found OTX2 highly expressed and specific for this type of tumor. SAGE data were confirmed by northern blot analysis, and the majority of the medulloblastomas tested were positive for either OTX1 or OTX2. The OTX genes are orthologues of the Drosophila otd gene and are highly conserved in mammals. Immunohistochemistry with OTX2 antibodies confirmed the RNA data and showed that OTX expression is a good marker for medulloblastomas. We also identified HES6 expression in medulloblastomas and rhabdomyosarcomas. HES genes are orthologues of hairy, a transcription factor in Drosophila. Analysis of the CGAP SAGE libraries showed that expression of HES6 is rare in other tumors and tissues. Northern blot analysis of ten additional medulloblastomas and nine rhabdomyosarcomas confirmed the high rate of expression of HES-6 in the majority of these tumors. Expression of HES6 might be another marker for medulloblastoma and rhabdomyosarcoma.