Combined hepatocellular-cholangiocarcinoma (combined HCC/ CC) is a rare form of liver neoplasms showing both hepatocellular (HCC) and bile duct differentiation (CC). In an attempt to clarify the clonality and genetic/phenotypic relationships in the evolution of these neoplasms, we microdissected multiple HCC and CC foci and studied allelic status of chromosome arms 1p, 1q, 3p, 4q, 5q, 6q, 8p, 9p, 10q, 11q, 13q, 16q, 17p, 17q, 18q, and 22q. Overall, the highest frequency of loss of heterozygosity (LOH) was seen on 4q and 17p, followed by 8p and 16q. Of the 11 cases studied, 3 cases did not show any of the identical allelic losses between HCC and CC foci, indicating the biclonal nature. The remaining 8 cases showed multiple allelic losses shared between both components, strongly suggestive of a single clonal derivation. Moreover, 4 of the 8 cases showed additional or divergent allelic losses at more than 1 chromosomal locus only in HCC and/or CC foci. Thus, this heterogeneity was shown to affect the phenotypic diversity of the tumor. Summarizing the genetic patterns, combined HCC/CC could be classified into the following 3 possibilities: (1) collision tumor in which 2 independent neoplastic clones develop at close proximity; (2) single clonal tumor with homogeneous genetic background in both components--histological diversity is thus a manifestation of divergent differentiation potential of a single clone; (3) single clonal process in which genetic heterogeneity in the process of clonal evolution within the tumor parallels histologic diversity; therefore, the tumor in this category is mainly composed of mosaics of closely related subclones.