Case 3 -

Combined Hepatocellular-Cholangiocarcinoma Ricardo V. Lloyd and Arie Perry

Clinical History This 36-year-old man presented with an enlarged right upper quadrant mass. Case 3 - Figure 1 Case 3 - Figure 2 Case 3 - Figure 3 Microscopic Description The biopsy showed carcinoma composed of cells with abundant eosinophilic cytoplasm which grew in sheets and nests. Other areas of the tumor showed glandular differentiation. Immunohistochemical staining was diffusely positive for keratin (CAM 5.2). There was positive staining for polyclonal CEA in a focal canalicular pattern. Hepatocyte paraffin 1 immunostaining was positive in the hepatocellular carcinoma areas but not in the cholangiocarcinoma areas. In situ hybridization for albumin mRNA was diffusely positive in both areas. Discussion Combined hepatocellular-cholangiocarcinoma (HCC-CC) is a rare primary tumor of the liver showing features of both hepatocellular and biliary epithelial differentiation [1, 2, 3, 4] . In a review of 24 cases of these tumors, Goodman et al, [1] classified the tumors into three histologic types. Type I or collision tumors (4 cases) had a coincidental occurrence of both tumors in the same patients. Type II or transitional tumors (n = 12) had intermediate differentiation and an identifiable transition between hepatocellular and cholangiocarcinoma. Type III or fibrolamellar tumors (n = 8) resembled fibrolamellar variant of hepatocellular carcinoma which also contained mucin-producing pseudoglands. Type III tumors occurred at a younger age in the absence of cirrhosis and had a slightly larger survival. In a series from Hong Kong (n = 21), most of the tumors (n = 18) were Type II. Two patients had Type I tumors, and one had Type III or fibrolamellar variant [2]. These combined HCC-CC had clinical and pathological features more similar to regular HCC. Cirrhosis or chronic hepatitis was present in 77.8 % of patients; 75% of patients had hepatitis B surface antigen positivity. Elevated serum AFP was present in 61.5% of patients, while 55% of the tumors had immunoreactive AFP [2]. Molecular studies of these combined tumors had shown loss of heterozygosity on 4 q and 17 p most commonly followed by 8 p and 16 q. Of the 11 cases analyzed, 8 showed identical allelic classes in both areas [3]. In addition to alpha-fetoprotein and polyclonal CEA, newer immunohistochemical markers that are useful in the diagnosis of HCC include hepatocyte paraffin 1 and CD10 [5, 6, 7, 8] . Albumin protein is present in high concentrations in the serum, so immunohistochemical staining for albumin normally results in strong non-specific background staining. However, albumin mRNA is largely restricted to a few cells and tumors with hepatocytes and hepatocellular neoplasms being the principal synthesizers of albumin. The albumin gene was cloned many years ago [8, 9] , and most molecular biology studies showed no difference between the normal liver and hepatocellular carcinomas in the use of multiple sites for transcriptional initiation and polyadenylation of the albumin gene [9, 10] . Murray et al. used a digoxigenin-labeled 51 bp oligonucleotide probe to study albumin expression in the liver and reported that 9/12 (75 percent) hepatocellular carcinomas examined were positive [10]. Other investigators have utilized oligonucleotide and RNA probes to detect albumin mRNA in primary and metastatic liver lesions [11, 12, 13, 14, 15] (Table 1}. Table 1. Oligonucleotide Sequences of Albumin Probe Cocktail Probe Sequence Nucleotides 1 51 GAG AAA AAG GGA AAT AAA GGT TAC CCA CT 42-70 2 51 GCA TCT CGA CGA AAC ACA CCC CTG GAA AT 87-115 3 51 CAA CCT CTG GTC TCA CCA ATC GGG GGA GGT 441-470 4 51 AGC AGG CAT CTC ATC ATT TTC CAC TTC GGC 981-1010 5 51 CTC GGC AAA GCA GGT CTC CTT ATC GTC AGC 1791-1820 Because albumin synthesis is largely restricted to the liver, detection of albumin mRNA in a poorly differentiated neoplasm, can be very useful in distinguishing a primary, and a metastatic lesion in the liver or in diagnosing metastatic hepatocellular carcinoma as in the current case. Staining for albumin mRNA is highly specific (Table 2). Table 2. Analysis of Albumin mRNA by In Situ Hybridization   % Positive Diagnosis Yamaguchi et al. [12] Papotti et al. [13] Salomao et al. [14] Krishna et al. [15] Hepatocellular Carcinoma 50/53 (94%) 26/30 (87%) 27/33 (82%) 22/23 (96%) Adenoma 1/1 (100%) -- -- 7/7 (100%) FNH 1/1 (100%) -- -- 7/7 (100%) Cholangiocarcinoma 0/8 (0%) -- 0/7 (0%) 0/9 (0%) Metastatic Adenocarcinoma 0/14 (0%) 0/10 (0%) 0/13 (0%) 0/16 (0%) In the two studies done at the Mayo Clinic [14, 15] , the only other neoplasm that had a positive ISH signal was a malignant mixed germ cell tumor which had hepatoid areas. Hepatoblastomas are also positive for albumin mRNA [15], so this ISH test should assist in distinguishing hepatoblastomas from other childhood neoplasms in the differential diagnosis. Albumin mRNA has also been useful in separating clear cell carcinoma of the liver from other clear cell neoplasms in the abdomen [16]. References Goodman ZD, Ishak KG, Langloss JM, Sesterhenn IA, Robin L. Combined hepatocellular-cholangiocarcinoma. A histologic and immunohistochemical study. Cancer 55:124-135, 1985 Ng IO, Shek TW, Nicholls J, Ma LT. Combined hepatocellular-cholangiocarcinoma: A clinicopathologic study. J Gastroenterol Hepatol 13:34-40, 1998 Fujii H, Zhu XG, Matsumoto T, et al. Genetic classification of combined hepatocellular-cholangiocarcinoma. Human Pathol 31:1011-1017, 2000 Ding S-F, Delhanty JDA, Bowles L, et al. Loss of constitutional heterozygosity on chromosomes 5 and 17 in cholangiocarcinoma. Br J Cancer 67:1007-1016, 1993 Zimmerman RL, Burke MA, Young NA, Solomides CC, Bibbo M. Diagnostic value of hepatocyte paraffin 1 antibody to disseminate hepatocellular carcinoma from metastatic carcinoma in fine-needle aspiration biopsies of the liver. Cancer 93:288-291, 2001 Maitra A, Murakata LA, Albores-Saavedra J. Immunoreactivity for hepatocyte paraffin 1 antibody in hepatoid adenocarcinomas of the gastrointestinal tract. Am J Clin Pathol 115:689-694, 2001. Boscheri N, Roessner A, Röcken C. Canalicular immunostaining of neprilysin (CD10) as a diagnostic marker for hepatocellular carcinomas. Am J Surg Pathol 25:1297-1303, 2001 Dragovic T, Deddish PA, Tran F, et al. Increased expression of the prilysin (neutral endopeptidase 24.11) in rate and human hepatocellular carcinomas. Lab Invest 70:107-113, 1994 Dugaiczyk A, Law SW, Dennison OE: Nucleotide sequence and the encoded amino acids of human serum albumin mRNA. Proc Natl Acad Sci USA 79:71-75, 1982 Urano Y, Watanabe K, Sakai M, Tamaoki T: The human albumin gene. Characterization of the 5' and 3' flanking regions and the polymorphic gene transcripts. J Biol Chem 261:3244-3251, 1986 Murray GI, Paterson PJ, Ewen SWB, Melvin WT: In situ hybridization of albumin mRNA in normal liver and hepatocellular carcinoma with a digoxigenin-labeled oligonucleotide probe. J Clin Pathol 45:21-24, 1992 Yamaguchi K, Nalesnik MA, Corr BI: In situ hybridization of albumin mRNA in normal liver and liver tumors: identification of hepatocellular origin. Virch Archiv B Cell Pathol 64:361-365, 1993 Papotti M, Paccicni D, Negro F, Bonino F, Bussolati G: Albumin gene expression in Mol Pathol 7:271-275, 1994 Salomao DR, Lloyd RV, Goellner JR: Hepatocellular carcinoma: needle biopsy findings in 74 cases. Acta Cytologia 16:8-13, 1996 Krishna M, Lloyd RV, Batts K: Detection of albumin messenger RNA in hepatic and extrahepatic neoplasms. Am J Surg Path 21:147-152, 1997 Olivera AM, Erickson LA, Borgert LJ, Lloyd RV. Differentiation of primary and metastatic clear cell tumors in the liver by in situ hybridization for albumin messenger RNA. Am J Surg Pathol 24:177-182, 2000 Tickoo SK, Zee SY, Obiekwe S, Xiao H, Koea J, Robiou C, Blumgart LH, Jarnagin W, Ladanyi M, Klimstra DS. Combined hepatocellular - cholangiocarcinoma. A histopathologic, immunohistochemical and in situ hybridization study. Am J Surg Pathol 26:989-997, 2002