Detection of alphafetoprotein-expressing cells in the blood of patients with hepatoma and hepatitis

Abstract

Summary The presence of tumour cells in the blood circulation may predict disease recurrence and metastasis. We have evaluated the specificity and sensitivity of detecting hepatoma cells in blood using nested polymerase chain reaction with primers specific for the alphafetoprotein (AFP) gene. The nested polymerase chain reaction amplified a 270-base pair AFP DNA fragment from cDNA of Hep 3B hepatoma cells. In a reconstitution experiment, AFP mRNA was detected from peripheral mononuclear cells isolated from 10 ml of blood containing as few as ten Hep 3B cells. Peripheral mononuclear cells from the blood of 20 hepatoma patients were analysed, and 19 patients showed positive AFP mRNA expression. Seven of 13 samples from hepatitis patients also showed positive AFP mRNA expression. All five paired samples of peripheral blood or umbilical cord blood from pregnant mothers and their babies, respectively, showed positive AFP expression. None of 22 control samples was positive. The presence of AFP mRNA in the blood of hepatitis or hepatoma patients suggests the presence of circulating hepatoma cells or hepatocytes in the circulation. The high incidence of AFP mRNA in the blood of hepatoma patients supports the notion of early haematogenous spreading of the disease.Keywords: alphafetoprotein; hepatocellular carcinoma; hepatitis; polymerase chain reaction Hepatocellular carcinoma (HCC) is one of the most common malignancies in Taiwan, mainland China and southern Africa. The disease is highly associated with hepatitis B and C virus infections as well as with the carcinogen aflatoxin (Chen, 1987). Despite improved survival of some cancer patients in recent years, the therapeutic efficacy for HCC remains very limited. Survival after the onset of symptoms is only a few months (Lai et al, 1987). It is therefore important to establish techniques for the early diagnosis of HCC.Alphafetoprotein (AFP) is a glycoprotein that is normally expressed during embryogenesis. The concentration of AFP in serum decreases as the liver develops and matures. However, AFP levels can become elevated in some disease states, particularly in HCC (Bellet et al, 1985;Chen, 1987;Di Bisceglie and Hoofnagle, 1989). Elevated serum AFP is employed as a highly specific and sensitive marker for the diagnosis of HCC, as about 82% of HCC patients have elevated serum AFP levels (Bellet et al, 1985). However, AFP can also be elevated in non-malignant forms of liver disease, such as acute and chronic hepatitis and cirrhosis (Chen, 1987;Di Bisceglie and Hoofnagle, 1989). In addition, increasing AFP levels have also have been associated with liver regeneration (Silver et al, 1974).A major characteristic of cancer is the ability of tumour cells to metastasize to other sites. The process of tumour metastasis involves multiple host-tumour interactions, and it is thought that (Chen, 1987). To understand stage tumour progression better, several laboratories have reported using the polymerase chain reaction (PCR) to detect metastatic cells in peripheral blood or lymph nod...

Evaluation of MAGE-1 and MAGE-3 as tumour-specific markers to detect blood dissemination of hepatocellular carcinoma cells

“…Though elevation of serum a-FP has been routinely applied as a parameter for HCC diagnosis, a-FP transcripts were detected in normal liver cells, the liver cirrhosis and the liver infectious diseases such as HBV/HCV infection (Jiang et al, 1997). Furthermore, a-FP transcripts was detected even in the blood samples from patients without HCC after the surgical injury of the liver, which results in the shedding of liver cells into circulation under surgical operation (Lemoine et al, 1997;Malek et al, 1999).…”

“…Then, the gene transcripts of both tissue-specific and tumour specific markers have been applied in RT -PCR based diagnosis of micrometastasis of tumour cells in peripheral blood, such as the tyrosinase and MAGE-3 transcripts in melanoma (Brossart et al, 1993;Foss et al, 1995;Hoon et al, 1995;Smith et al, 1991), prostate specific antigen (PSA) and prostate specific membrane antigen (PSMA) encoding gene transcripts in prostate cancer (Israeli et al, 1994(Israeli et al, , 1995, carcinoembryonic antigen (CEA) in colon cancer , CD44 and CK-19 transcripts in breast cancer (Dall et al, 1995;Kruger et al, 1996) and tyrosinase hydroxylase in neuroectodermal tumour (Miyajima et al, 1996;Naito et al, 1991). With respect to HCC, both albumin and a-FP mRNA are widely used as tumour markers for HCC cells in circulation (Chou et al, 1994;Hillaire et al, 1994;Jiang et al, 1997;Kar and Carr, 1995;Komeda et al, 1995;Lemoine et al, 1997;Malek et al, 1999;Matsumura et al, 1994;Wong et al, 1997). However, the reliability using them as tumour markers have been challenged, because both albumin and a-FP are abundantly expressed in normal liver cells, they are released to the peripheral blood by either surgical injury of the liver for the disease other than HCC or by hepatitis virus infection (Chou et al, 1994;Jiang et al, 1997; Lemoine et al, 1997;Malek et al, 1999;Wong et al, 1997).…”

“…With respect to HCC, both albumin and a-FP mRNA are widely used as tumour markers for HCC cells in circulation (Chou et al, 1994;Hillaire et al, 1994;Jiang et al, 1997;Kar and Carr, 1995;Komeda et al, 1995;Lemoine et al, 1997;Malek et al, 1999;Matsumura et al, 1994;Wong et al, 1997). However, the reliability using them as tumour markers have been challenged, because both albumin and a-FP are abundantly expressed in normal liver cells, they are released to the peripheral blood by either surgical injury of the liver for the disease other than HCC or by hepatitis virus infection (Chou et al, 1994;Jiang et al, 1997; Lemoine et al, 1997;Malek et al, 1999;Wong et al, 1997).…”

Detection of alphafetoprotein-expressing cells in the blood of patients with hepatoma and hepatitis

“…We read with great interest the article by Jiang et al (1997). Their findings may have a significant impact on studies predicting recurrence and metastasis of hepatocellular carcinoma (HCC) by detecting alphafetoprotein (AFP) mRNA in circulating cells (Komeda et al, 1995;Wong et al, 1997).…”

Does surgical resection of hepatocellular carcinoma accelerate cancer dissemination?

“…With the use of PCR-based method which permits detection of 1 tumor cell among 10 7 normal peripheral, mononuclear blood cells, the blood-borne dispersion of tumor cells during surgical manipulation has been reported in humans with prostatic carcinoma, melanoma, breast carcinoma and pancreatic carcinoma [9][10][11][12] . For detecting HCC cells in circulation, reverse transcriptase (RT) PCR targeting human alphafetoprotein (hAFP) messenger RNA (mRNA) or human albumin mRNA has been proposed [13][14][15][16][17][18][19][20][21][22] . Lack of the specificity of albumin mRNA has been emphasized [21,22] .…”