Alpha-1-acid glycoprotein as potential biomarker for alpha-fetoprotein-low hepatocellular carcinoma

Bachtiar et al. BMC Research Notes 2010, 3:319 http://www.biomedcentral.com/1756-0500/3/319 RESEARCH ARTICLE Open Access Alpha-1-acid glycoprotein as potential biomarker for alpha-fetoprotein-low hepatocellular carcinoma Indra Bachtiar1*, Valentine Kheng1, Gunawan A Wibowo1, Rino A Gani2, Irsan Hasan2, Andri Sanityoso2, Unggul Budhihusodo2, Syafruddin AR Lelosutan3, Ruswhandi Martamala3, Wenny A Achwan4, Soewignyo Soemoharjo4, Ali Sulaiman5, Laurentius A Lesmana2, Susan Tai1 Abstract Background: The outcome of patients with hepatocellular carcinoma (HCC) remains poor because of late diagnosis. We determined the performances of a -1-acid glycoprotein (AAG) and des-g-carboxy prothrombin (DCP) for the diagnosis of HCC, especially for a-fetoprotein (AFP)-low HCC. Methods: Of the 220 patients included in this retrospective study, 124 had HCC, and 61 (49%) of these were AFPlow HCC (AFP ≤ 20 ng/mL). The remaining 96 patients, including 49 with chronic hepatitis B or C and 47 with cirrhosis, were considered as control. Plasma AAG was analyzed using high performance liquid chromatography (HPLC) and confirmed using Western blot technique. Results: When all patients with HCC were evaluated, the area under receiver operating characteristic (ROC) curves for AAG (0.94, 95% CI: 0.91-0.97) and DCP (0.92, 95% CI: 0.88-0.95) were similar (P = 0.40). AAG had better area under ROC curve (0.96, 95% CI: 0.94-0.99) than DCP (0.87, 95% CI: 0.81-0.93) for AFP-low HCC (P < 0.05). At the specificity 95%, the sensitivity of AAG was higher in AFP-low HCC than in AFP-high HCC (82% and 62%, respectively). In contrast, higher sensitivity was obtained from DCP in discriminating HCC patients with low AFP than that in high AFP (57% and 90%, respectively). Conclusion: Our cross-sectional study showed that AAG was better performance in diagnosing HCC patients with low AFP, while DCP did better in those with high AFP. Background Hepatocellular carcinoma (HCC) is one of the most common malignancies in the world [1]. Until recently, AFP has been the most widely used plasma marker for diagnosis, surveillance, and as a prognostic indicator of HCC patients’ survival [2]. Several studies indicated that high plasma levels of AFP are related to poor prognosis, as well as histologic grade of malignancy [3]. Those with high plasma AFP level at the time of HCC diagnosis have more unfavourable outcomes compared to patients with low AFP level [4]. However, it has been recognized that AFP has a low sensitivity in detection of HCC, and that AFP level often increases in the absence of HCC [5]. * Correspondence: 1 Proteomic Division, Mochtar Riady Institute for Nanotechnology, Lippo Karawaci, Tangerang, 15811, Indonesia Full list of author information is available at the end of the article To date, several tumor markers have been proposed as complement or substitute for AFP in HCC diagnosis. Recently, Lens culinaris agglutinin-reactive fraction of AFP (AFP-L3) [6] and DCP [7] have been approved by the Food and Drug Administration as plasma markers for HCC. AFP-L3 can be detected in the plasma of patients with small tumors [8,9]. However, for the diagnosis of early stage of HCC, AFP-L3 [10] and DCP [10,11] are less sensitive than AFP for the diagnosis of early and very early stage HCC. DCP has not been used in Indonesia and it could be a potential biomarker for diagnosis of HCC. Therefore a cross-sectional study is required to determine the role of DCP in the diagnosis of HCC in the Indonesian population. Another potential biomarker for HCC is AAG. AAG is an acute phase protein, synthesized predominantly in the liver. Cytokines can cause plasma AAG level to increase as part of an inflammatory response [12]. The plasma level of © 2010 Bachtiar et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Bachtiar et al. BMC Research Notes 2010, 3:319 http://www.biomedcentral.com/1756-0500/3/319 AAG has been suggested to be a potential marker for diagnosing cirrhosis and HCC [13]. Recently we have shown that combination of AAG and AFP improves the accuracy of HCC diagnosis [14]. Given the rising incidence of HCC in Indonesia and lack of data on the clinical utility of these two tumor markers (AAG and DCP) for detecting the presence of HCC, we selected 220 chronic liver patients, with and without HCC. This study is a further step in order to assess the feasibility of AAG to diagnose AFP-low HCC. This study has limitations, and not designed to assess the treatment and follow-up of the screen-detected patients. Our specific aims were to: (i) define the level of each tumor marker with the best sensitivity, specificity, accuracy, positive predictive value (PPV) and negative predictive value (NPV) for diagnosis of HCC; and (ii) to evaluate the clinical utilities of AAG and DCP in HCC patients with low AFP (≤ 20 ng/mL) and high AFP (> 20 ng/mL), especially in attempt to find the best biomarker for diagnosing HCC patients with low AFP. Methods Patients A total of 220 plasma samples were collected between January 2006 and January 2009 from patients in four participating hospitals, Cipto Mangunkusumo Hospital, Gatot Subroto Army Hospital, Ali Sulaiman Liver Clinic Center (Jakarta, West Indonesia) and Hepatitis Laboratory Mataram (West Nusa Tenggara, East Indonesia). Demographic and clinical information of each patient including age, gender, and cause of disease were obtained from chart review. All patients gave informed consents to participate in the study, and the Institutional Review Board of each participating institution approved the protocol. The patients were divided into three groups: (i) control (non-malignant disease), either chronic hepatitis patients with HBV/HCV infection (n = 49) or cirrhotic patients (n = 47); (ii) AFP-low HCC (n = 61); and (iii) AFP-high HCC (n = 63). Etiology of underlying liver disease was attributed to HBV based on detection of HBsAg in plasma via a commercial assay and HCV based on detection of hepatitis C antibody/hepatitis C virus RNA in plasma. The presence of cirrhosis was defined by histological measurement or non-histological by evidence of portal hypertension in the presence of chronic liver disease. Diagnosis of HCC relied on the presence of a malignant liver nodule, as established by imaging techniques. Fine needle aspiration biopsy procedure reconfirmed the diagnosis of HCC for sample in which the AFP concentration was low. Subjects with HIV co-infection or autoimmune hepatitis were excluded in this study. None of the subjects was taking vitamin K at time of the study. Page 2 of 8 Biomarkers measurements Blood samples were collected in pyrogen-free tube (BD Vacutainer, Plymouth, UK) with ethylene diamine tetraacetic acid as an anti (...truncated)