Chromosomal rearrangements unraveled by spectral karyotyping (SKY) indicated frequent chromosome 19 translocations in hepatocellular carcinoma (HCC). In an effort to characterize the aberrant 19 rearrangements in HCC, we performed positional mapping by fluorescence in-situ hybridization (FISH) in 10 HCC cell lines. SKY analysis indicated structural rearrangements of chromosome 19 in 6 cell lines, 4 of which demonstrated recurring 19p translocations with different partner chromosomes. Using fluorescence-labeled BAC probes, physical mapping indicated a breakpoint cluster between 19p13.12 and 19p12. A corresponding transcriptional mapping by cDNA array on 19p suggested the differential expression of a single downregulated gene ACP5 (tartrate-resistant acid phosphatase type 5). Quantitative RT-PCR confirmed the reduced expression of ACP5 and indicated a strong correlation of its repressed expression only in cell lines that contain a 19p rearrangement (p = 0.004). We further examined the expression of ACP5 in a cohort of 82 primary tumors and 74 matching nonmalignant liver tissues. In the primary HCC examined, a reduction of ACP5 transcripts by 2 to as much as 1,000-fold was suggested in 67% of tumors (55/82 cases). When compared to adjacent nonmalignant tissues, 46% of tumors (34/74 cases) demonstrated a lower expression level (p = 0.015). On closer examination, a high significance of ACP5 repression was suggested in the cirrhotic HCC subgroup that was derived from chronic hepatitis B infected patients (55%; 30/54 cases; p = 0.001). Functional examination of ACP5 ectopic expression in HCC cells further demonstrated a significant growth inhibitory effect of ACP5 on tumor cell survival (p < 0.001). In our study, the novel finding of common ACP5 downregulation in HCC may provide basis for further investigations on the role of acid phosphatase in hepatocarcinogenesis.