Original Article
Copyright ©2014 Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Dec 28, 2014; 20(48): 18240-18248
Published online Dec 28, 2014. doi: 10.3748/wjg.v20.i48.18240
Circulating tumor and cancer stem cells in hepatitis C virus-associated liver disease
Abeer A Bahnassy, Abdel-Rahman N Zekri, Ahmed El-Bastawisy, Amal Fawzy, Marwa Shetta, Nehal Hussein, Dalia Omran, Abdallah A S Ahmed, Samir S El-Labbody
Abeer A Bahnassy, Molecular Pathology Unit, Pathology Department, National Cancer Institute, Cairo University, Cairo 11796, Egypt
Abdel-Rahman N Zekri, Nehal Hussein, Virology and Immunology Unit, Cancer Biology Department, National Cancer Institute, Cairo University, Cairo 11796, Egypt
Ahmed El-Bastawisy, Medical Oncology Department, National Cancer Institute, Cairo University, Cairo 11796, Egypt
Amal Fawzy, Clinical Pathology Department, National Cancer Institute, Cairo University, Cairo 11796, Egypt
Marwa Shetta, Clinical Pathology, Kasr Al-Aini, Cairo university, Cairo 11782, Egypt
Abdallah A S Ahmed, Samir S El-Labbody, Microbiology Department, El-Azhar University, Cairo 11964, Egypt
Dalia Omran, Tropical Medicine Department, Faculty of Medicine, Cairo University, Cairo 11796, Egypt
Author contributions: Bahnassy AA and Zekri ARN designed the research; Bahnassy AA performed the flow cytometry analysis; Zekri ARN analyzed the data; El-Bastawisy A and Omran D collected the clinical data; Fawzy A and Hussein N performed the polymerase chain reactions; Shetta M performed the statistical analyses; Hussein N assisted in manuscript preparation; Ahmed AAS collected and arranged the samples; El-Labbody SS, Bahnassy AA, Zekri ARN and Fawzy A edited the manuscript.
Correspondence to: Abeer A Bahnassy, MD, PhD, Professor, Molecular Pathology Unit, Pathology Department, National Cancer Institute, Cairo University, Fom El Khalig, Cairo 11796, Egypt. chaya2000@hotmail.com
Telephone: +20-2-23364147 Fax: +20-2-23364720
Received: April 21, 2014
Revised: May 24, 2014
Accepted: July 11, 2014
Published online: December 28, 2014
Processing time: 260 Days and 1.3 Hours
Abstract

AIM: To assess the role of circulating tumor cells (CTCs) and cancer stem cells (CSCs) in hepatitis C virus (HCV)-associated liver disease.

METHODS: Blood and/or tissue samples were obtained from HCV (genotype 4)-associated hepatocellular carcinoma patients (HCC; n = 120), chronic hepatitis C patients (CH; n = 30) and 33 normal control subjects (n = 33). Serum levels of alpha-fetoprotein (AFP), alkaline phosphatase, and alanine and aspartate aminotransferases were measured. Cytokeratin 19 (CK19) monoclonal antibody was used to enumerate CTCs, and CD133 and CD90 were used to enumerate CSCs by flow cytometry. The expression levels of the CSCs markers (CD133 and CD90) as well as telomerase, melanoma antigen encoding gene 1 (MAGE1) and MAGE3 were assessed by RT-PCR and quantitative real-time polymerase chain reactions. The number of CTCs and/or the expression levels of CK19, CD133, telomerase, MAGE1 and MAGE3 were correlated to the standard clinicopathologic prognostic factors and disease progression.

RESULTS: Levels of AFP, alkaline phosphatase and aspartate aminotransferase were significantly different among the HCC, CH and control groups (P < 0.001), whereas alanine aminotransferase differed significantly between patient (HCC and CH) and control groups (P < 0.001). At the specified cutoff values determine by flow cytometry, CK19 (49.8), CD90 (400) and CD133 (73) were significantly higher in the blood of HCC patients compared to those in the CH and control groups (P < 0.001). On the other hand, CD133 at a 69.5 cutoff was significantly higher in the CH compared to the control group (P≤ 0.001). Telomerase, MAGE1 and MAGE3 RNA were expressed in 55.71%, 60.00% and 62.86% of the HCC patients, respectively, but were not detected in patients in the CH or control groups, which were statistically significant (Ps < 0.001). The expression levels of telomerase, CD90, MAGE3, CD133 and CK19 were all significantly associated with high tumor grade and advanced stage in HCC patients (all Ps < 0.05).

CONCLUSION: CTC counts and AFP, CK19, telomerase, and MAGE1/MAGE3 expression predict disease progression in patients with HCV, whereas telomerase, MAGE3, CD90, CD133 and CK19 are prognostic markers in HCC.

Keywords: Cancer stem cells; Circulating tumor cells; Hepatitis C virus genotype-4; Hepatocellular carcinoma

Core tip: Recent studies have shown that cancer stem and circulating tumor cells contribute to tumor development and progression and can predict patient outcome. Although there are various methods for enumeration of circulating tumor cells, this study demonstrates that flow cytometry is a sensitive, rapid and easy technique that can be used to follow chronic hepatitis C virus patients for early detection of hepatocellular carcinoma. Additionally, telomerase, melanoma antigen encoding gene 3 and cancer stem cell markers (CD90, CD133, CK19) are prognostic indicators in hepatocellular carcinoma patients.