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. 1997;76(5):628–633. doi: 10.1038/bjc.1997.436

Semiquantification of circulating hepatocellular carcinoma cells by reverse transcriptase polymerase chain reaction.

I H Wong 1, T Leung 1, S Ho 1, W Y Lau 1, M Chan 1, P J Johnson 1
PMCID: PMC2228015  PMID: 9303362

Abstract

Hepatocellular carcinoma (HCC) is one of the most common and rapidly fatal malignancies worldwide. Treatment options are severely limited by the frequent presence of metastases. If hepatocyte-specific mRNAs are detected in the circulation, it is possible to infer the presence of circulating, presumably malignant, liver cells. If these can be quantified, it is possible to predict the likelihood of haematogenous metastasis. In this investigation, we have attempted to gain an index of the mass of circulating HCC cells (with reference to the number of hepatoblastoma cells) by measuring the amounts of PCR products for albumin (alb) mRNA and alpha-fetoprotein (afp) mRNA by reverse transcriptase polymerase chain reaction (RT-PCR) and Southern blot analysis. For calibration, total RNA from 1-10(6) HepG2 cells was mixed with total RNA from 10(6) normal peripheral mononuclear cells. A linear relationship was demonstrated between the amount of alb- or afp PCR product and the level of HepG2 total RNA spiked. The assay is sensitive down to a detection level of one HepG2 cell. Alb mRNA was detected in 50% of 18 normal subjects and afp mRNA in only two normal subjects. The alb mRNA cut-off level for the normal was exceeded by seven normal subjects and 34 out of 64 HCC patients, and that for afp mRNA was exceeded by six HCC patients but none of the normal subjects. The level of alb mRNA detected was not linearly proportional to the amount of afp mRNA detected in peripheral blood of the same patients, suggesting heterogeneous expression of alb and afp genes in different circulating tumour cells. In addition, no significant linear association between the levels of afp mRNA and serum AFP was observed. Semiquantification of both mRNA markers for HCC cell detection may prove useful in prediction of metastases.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aden D. P., Fogel A., Plotkin S., Damjanov I., Knowles B. B. Controlled synthesis of HBsAg in a differentiated human liver carcinoma-derived cell line. Nature. 1979 Dec 6;282(5739):615–616. doi: 10.1038/282615a0. [DOI] [PubMed] [Google Scholar]
  2. Burchill S. A., Bradbury F. M., Smith B., Lewis I. J., Selby P. Neuroblastoma cell detection by reverse transcriptase-polymerase chain reaction (RT-PCR) for tyrosine hydroxylase mRNA. Int J Cancer. 1994 Jun 1;57(5):671–675. doi: 10.1002/ijc.2910570510. [DOI] [PubMed] [Google Scholar]
  3. Burchill S. A., Bradbury M. F., Pittman K., Southgate J., Smith B., Selby P. Detection of epithelial cancer cells in peripheral blood by reverse transcriptase-polymerase chain reaction. Br J Cancer. 1995 Feb;71(2):278–281. doi: 10.1038/bjc.1995.56. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  5. Datta Y. H., Adams P. T., Drobyski W. R., Ethier S. P., Terry V. H., Roth M. S. Sensitive detection of occult breast cancer by the reverse-transcriptase polymerase chain reaction. J Clin Oncol. 1994 Mar;12(3):475–482. doi: 10.1200/JCO.1994.12.3.475. [DOI] [PubMed] [Google Scholar]
  6. Di Bisceglie A. M., Dusheiko G. M., Paterson A. C., Alexander J., Shouval D., Lee C. S., Beasley R. P., Kew M. C. Detection of alpha-foetoprotein messenger RNA in human hepatocellular carcinoma and hepatoblastoma tissue. Br J Cancer. 1986 Nov;54(5):779–785. doi: 10.1038/bjc.1986.240. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gerhard M., Juhl H., Kalthoff H., Schreiber H. W., Wagener C., Neumaier M. Specific detection of carcinoembryonic antigen-expressing tumor cells in bone marrow aspirates by polymerase chain reaction. J Clin Oncol. 1994 Apr;12(4):725–729. doi: 10.1200/JCO.1994.12.4.725. [DOI] [PubMed] [Google Scholar]
  8. Hillaire S., Barbu V., Boucher E., Moukhtar M., Poupon R. Albumin messenger RNA as a marker of circulating hepatocytes in hepatocellular carcinoma. Gastroenterology. 1994 Jan;106(1):239–242. doi: 10.1016/s0016-5085(94)95705-3. [DOI] [PubMed] [Google Scholar]
  9. Kar S., Carr B. I. Detection of liver cells in peripheral blood of patients with advanced-stage hepatocellular carcinoma. Hepatology. 1995 Feb;21(2):403–407. [PubMed] [Google Scholar]
  10. Knowles B. B., Howe C. C., Aden D. P. Human hepatocellular carcinoma cell lines secrete the major plasma proteins and hepatitis B surface antigen. Science. 1980 Jul 25;209(4455):497–499. doi: 10.1126/science.6248960. [DOI] [PubMed] [Google Scholar]
  11. Komeda T., Fukuda Y., Sando T., Kita R., Furukawa M., Nishida N., Amenomori M., Nakao K. Sensitive detection of circulating hepatocellular carcinoma cells in peripheral venous blood. Cancer. 1995 May 1;75(9):2214–2219. doi: 10.1002/1097-0142(19950501)75:9<2214::aid-cncr2820750905>3.0.co;2-x. [DOI] [PubMed] [Google Scholar]
  12. Liotta L. A., Kleinerman J., Saidel G. M. Quantitative relationships of intravascular tumor cells, tumor vessels, and pulmonary metastases following tumor implantation. Cancer Res. 1974 May;34(5):997–1004. [PubMed] [Google Scholar]
  13. Matsumura M., Niwa Y., Hikiba Y., Okano K., Kato N., Shiina S., Shiratori Y., Omata M. Sensitive assay for detection of hepatocellular carcinoma associated gene transcription (alpha-fetoprotein mRNA) in blood. Biochem Biophys Res Commun. 1995 Feb 15;207(2):813–818. doi: 10.1006/bbrc.1995.1259. [DOI] [PubMed] [Google Scholar]
  14. Matsumura M., Niwa Y., Kato N., Komatsu Y., Shiina S., Kawabe T., Kawase T., Toyoshima H., Ihori M., Shiratori Y. Detection of alpha-fetoprotein mRNA, an indicator of hematogenous spreading hepatocellular carcinoma, in the circulation: a possible predictor of metastatic hepatocellular carcinoma. Hepatology. 1994 Dec;20(6):1418–1425. doi: 10.1002/hep.1840200607. [DOI] [PubMed] [Google Scholar]
  15. Mattano L. A., Jr, Moss T. J., Emerson S. G. Sensitive detection of rare circulating neuroblastoma cells by the reverse transcriptase-polymerase chain reaction. Cancer Res. 1992 Sep 1;52(17):4701–4705. [PubMed] [Google Scholar]
  16. McLeod J. F., Cooke N. E. The vitamin D-binding protein, alpha-fetoprotein, albumin multigene family: detection of transcripts in multiple tissues. J Biol Chem. 1989 Dec 25;264(36):21760–21769. [PubMed] [Google Scholar]
  17. Otsuru A., Nagataki S., Koji T., Tamaoki T. Analysis of alpha-fetoprotein gene expression in hepatocellular carcinoma and liver cirrhosis by in situ hybridization. Cancer. 1988 Sep 15;62(6):1105–1112. doi: 10.1002/1097-0142(19880915)62:6<1105::aid-cncr2820620613>3.0.co;2-z. [DOI] [PubMed] [Google Scholar]

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