Multiple molecular markers MAGE-1, MAGE-3 and AFP mRNAs expression nested PCR assay for sensitive and specific detection of circulating hepatoma cells: Enhanced detection of hepatocellular carcinoma

Hepatocellular Carcinoma is a multifactorial, multistep and complex process. Its prognosis is poor and early diagnosis and monitoring of metastasis of HCC is of utmost importance. Circulating alpha-fetoprotien mRNA has been proposed as a marker of HCC cells disseminated into the circulation but the specificity of this molecular marker and its correlation with the main HCC clinico-pathological parameters remain controversial. In recent years; several different multi-marker assays have been developed for the detection of hepatoma cells in the peripheral blood of patients with HCC.In this study 58 patients and 15 matched healthy volunteers were included; the patients were divided into three groups; group A: patients with primary HCC (n = 32), group B: patients with cirrhosis with no evidence of HCC (n = 12), group C: patients with metastatic cancer in liver (n = 14). Group D: 15 healthy volunteers age and sex matched. The staging of HCC was carried out according to the Tumor/Node/Metastasis (TNM) classification.Peripheral blood samples were obtained from all subjects; MAGE-1 and MAGE-3 and AFP mRNAs were detected by nested RT-PCR.The positive rates of MAGE-1, MAGE-3 and AFP mRNAs were 18/32 (56.3%), 15/32 (46.9%) and 19/32 (59.4%) respectively in the primary HCC patients.In the cirrhotic group only 4/12 (33.3%) patients were positive for AFP mRNA, whereas in the metastatic group 5/14 (35.7%) and 4/14 (28.6%) were positive to MAGE-1 and MAGE-3 mRNAs respectively. MAGE-1 and MAGE-3 mRNAs were correlated with TNM clinical stages; tumor number and tumor size (p < 0.05).Our results indicate that a multi-marker nested RT-PCR assay with cancer-specific markers such as MAGE-1 and MAGE-3 in combination with a hepatocyte-specific AFP marker may be a promising diagnostic tool for monitoring hepatocellular carcinoma patients. Nested PCR exhibits higher sensitivity, stronger specificity and lower false-positive occurrence as compared to single RT.