Human endogenous retroviruses (HERVs) account for approximately 8% of the human genome. Since the majority of HERV elements have accumulated inactivating mutations in the viral genes, only few expressed viral open reading frames (ORFs) have been described. In this study, we have analyzed the expression of a HERV-H copy located on Xp22.3 encompassing a potential ORF immediately downstream of the viral promoter. Conventional and real time RT-PCR based expression analysis of this specific HERV-H sequence showed overexpression in 16 of 34 (47%) colorectal, 25 of 63 (40%) gastric and 2 of 12 (17%) pancreatic cancers, whereas no overexpression was detected in bronchial and cervical cancers. Normal human testis, placenta and breast tissue did not show expression of this sequence. CpG methylation analysis of the viral promoter revealed a loss of methylation in cell lines expressing the HERV-H sequence as compared to nonexpressing cell lines and lymphocyte DNA derived from healthy individuals. Further investigations of the HERV-H long terminal repeat and the HERV-H RNA are necessary to assess the functional relevance of the HERV-H expression. © 2007 Wiley-Liss, Inc.