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Patients with chronic hepatitis C without advanced fibrosis and hepatocellular carcinoma: A retrospective clinical–pathological study
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Contents lists available at ScienceDirect
Digestive and Liver Disease journal homepage: www.elsevier.com/locate/dld
Liver, Pancreas and Biliary Tract
Patients with chronic hepatitis C without advanced fibrosis and hepatocellular carcinoma: A retrospective clinical–pathological study Blaise K-Kutala a,b,d,∗ , Pierre Bedossa c , Jeremie Guedj b , Tarik Asselah a,d , Michelle Martinot-Peignoux d , Xavier Duval b , Patrick Marcellin a,d a
Department of Hepatology, Beaujon Hospital, AP-HP, Clichy, France INSERM U1137, University Paris 7, UFR of Medicine, France c Department of Pathology, Beaujon Hospital, AP-HP, Clichy, France d INSERM U1149/CRI, Beaujon Hospital, AP-HP, Clichy, France b
a r t i c l e
i n f o
Article history: Received 25 July 2014 Accepted 17 December 2014 Available online xxx Keywords: Cirrhosis Hepatitis C Hepatocellular carcinoma Nonalcoholic steatohepatitis Screening
a b s t r a c t Background: There are very few studies on the incidence and risk factors of hepatitis C virus (HCV)-induced hepatocellular carcinoma (HCC) in the absence of advanced fibrosis. Our objective was to identify the clinical–pathological features of these patients. Methods: We retrospectively reviewed 162 patients admitted to our hospital for HCV-related HCC between 2000 and 2010. Patients with hepatitis of other aetiologies, human immunodeficiency virus co-infection, or treated with interferon were excluded. We compared demographic, laboratory, clinical and outcome parameters of patients with and without advanced fibrosis. Results: 137 patients had advanced fibrosis (85%). Median age was higher in the advanced fibrosis vs. the non-advanced fibrosis group (62 vs. 65 years, respectively; p = 0.025). Steatosis was significantly more frequent in patients with advanced fibrosis compared to those without advanced fibrosis (43% vs. 20%, respectively; p = 0.032). Independent predictors associated to the occurrence of HCC in patients without advanced fibrosis were hepatitis B core antigen (odds ratio: 3.86; p = 0.044) and duration of hepatitis C infection (odds ratio: 1.21; p = 0.003). Conclusions: Risk factors such as steatosis or diabetes were not frequent in patients without advanced fibrosis. Further studies are needed to evaluate the role of occult hepatitis B and the duration of hepatitis infection in patients with HCC and chronic hepatitis C without advanced fibrosis. © 2014 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.
1. Introduction Hepatocellular carcinoma (HCC) the most common liver cancer; representing 90% of primary liver cancer worldwide, with an increasing incidence in the United States, Western Europe, and Japan [1]. Although HCC usually develops in cirrhotic livers, a proportion also develops in the absence of advanced hepatic fibrosis or cirrhosis. The frequency of this event varies along geographic and ethnic lines and is closely associated with etiological factors [2]. Hepatitis B virus (HBV), for example, is known to have a direct oncogenic effect on the liver by integrating into the host genome [3], and the prevalence of non-cirrhotic HCC is high (20%) in regions
∗ Corresponding author at: Service d’Hépatologie, Hôpital Beaujon AP-HP, 100 Boulevard Général Leclerc, 92110 Clichy, France. Tel.: +33 1 40 87 54 31; fax: +33 1 47 30 94 40. E-mail address: [email protected] (B. K-Kutala).
where this HBV is endemic, such as China and Taiwan [4]. There is also a population of patients that develops HCC in the absence of underlying parenchymal liver disease or exposure to known environmental risk factors, such as aflatoxin B1, nitrosamine or benzopyrenes [5]. Hepatitis C virus (HCV) infection is a well-known cause of HCC; the estimated risk of HCC is 15–20 times higher in patients with chronic hepatitis C (CHC) (30% in Asia and 17% in Western countries) than in those without, and the greatest risk is found in patients with advanced hepatic fibrosis or cirrhosis [6,7]. Unfortunately, there are very few, and sometimes contradictory studies on the incidence of HCC in patients with chronic hepatitis C without cirrhosis [8,9]. Until now, practical clinical guidelines and recommendations have been focused on the screening for HCC in patients with cirrhosis or advanced fibrosis related to chronic hepatitis C but not in those without advanced fibrosis [10]. Thus, the aims of this study were to analyse and compare the main clinical features and tumour characteristics at diagnosis of patients with CHC-related
http://dx.doi.org/10.1016/j.dld.2014.12.010 1590-8658/© 2014 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.
Please cite this article in press as: K-Kutala B, et al. Patients with chronic hepatitis C without advanced fibrosis and hepatocellular carcinoma: A retrospective clinical–pathological study. Dig Liver Dis (2015), http://dx.doi.org/10.1016/j.dld.2014.12.010
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HCC without advanced fibrosis to those of patients with HCC on cirrhosis or advanced fibrosis. 2. Patients and methods 2.1. Study design and data source We retrospectively all consecutive patients in our referral hospital who followed-up for HCC care between January 2000 and December 2010 and fulfilled the following criteria: (1) CHC defined by the association of positive serum anti-HCV antibodies and RNA, with liver histology corresponding to a METAVIR Score of F3-F4 for patients with advanced fibrosis and F0-F2 for those with nonadvanced fibrosis; (2) diagnosis of HCC and staging of the chronic hepatitis, based on the surgical specimen or two adequate liver biopsies (in non-tumorous liver tissue and in HCC nodule) reviewed in our pathology department by an experienced liver pathologist; (3) no other coexisting liver disease, such as HBV, or human immunodeficiency virus (HIV) co-infection (negative serum HBsAg and negative anti-HIV antibodies); (4) daily alcohol consumption <40 g for males and <30 g for females. Thus patients without advanced fibrosis previously treated with interferon (IFN) were excluded. The inclusion period was extended to include more patients without advanced fibrosis. Patients with advanced fibrosis underwent a standard surveillance programme; diagnosis of HCC was based on the following: biopsy from 2001, a combination of imaging and laboratory findings [computed tomography (CT) scan or Magnetic resonance imaging (MRI) and alpha-fetoprotein serum levels (AFP) > 400 ng/ml] between 2001 and 2004 and after 2005; and if necessary, on contrast uptake in the arterial phase and washout in the venous/late phase on CT, MRI or ultrasound or biopsy. Although patients without fibrosis did not undergo routine screening, HCC was usually detected during routine visits for the follow-up of CHC or other comorbid conditions such as arterial hypertension or diabetes mellitus. In some of these cases HCC was diagnosed at the same time as CHC, or was sought because of known risk factors (age >65 years, male gender, elevated serum AFP). 2.2. Clinical and biochemical characteristics We performed a retrospective analysis of a prospective database of patients with CHC-HCC who underwent liver resection in our hospital. Patient files were reviewed and analysed for the following demographic, clinical, and biochemical data collected at admission: age, gender, cause of cirrhosis, HBV markers (AntiHBc, Anti-HBs) and HCV-RNA; aspartate aminotransferase, alanine aminotransferase, total bilirubin, albumin, and serum AFP; prothrombin activity, platelet count, viral load and transmission of HCV infection. Complications were also recorded: Child Pugh score, the presence of ascites, encephalopathy, oesophageal varices, and a history of variceal or gastrointestinal bleeding. The relationship between the metabolic syndrome and HCC in patients without cirrhosis was studied in our series. Metabolic syndrome was defined as a combination of any 3 of the 5 following risk factors: waist circumference ≥102 cm in men and ≥88 cm in women of European descent (≥94 cm and ≥80 for African descent), triglycerides ≥150 mg/dL, HDL-C < 40 mg/dL in men and <50 mg/dL in women, arterial hypertension, and elevated fasting glucose. The estimated duration of infection was defined as the time that elapsed from the presumed date of infection (date of transfusion, first year of intravenous drug use, date of vaccination for African patients, unsterile medical and dental procedures, and traditional medical and cosmetic procedures involving blood exposure) to the date of HCC diagnosis.
2.3. Histological data Each liver resection sample was individually coded and processed for histopathological interpretation by a single experienced liver pathologist (P. B.), blinded to the patient’s identity, and clinical and biological data and was routinely processed, formalinfixed, serially sectioned, and stained with haematoxylin/eosin, Masson’s trichrome, and Sirius red to evaluate fibrosis and Perls staining for iron. A detailed scoring sheet was used; including the grading and staging of fibrosis by the METAVIR classification, qualitative evaluation of steatosis (0, absent 1, present), predominant zonal distribution of fat (pericentral, periportal, azonal, diffuse), clarification/ballooning of liver cells (0, absent; 1, mild; 2, marked), perisinusoidal fibrosis (0, absent; 1, mild; 2, marked), Mallory hyaline (0, absent; 1, present) and presence of glycogenated nuclei (0, absent; 1, present) as well as other less common histological features described by Kleiner et al. [11]. The following histological characteristics were analysed for each patient: METAVIR score [12], number and size of nodules, presence of microvascular invasion, presence of steatosis, iron overload, presence of non-alcoholic steatohepatitis (NASH) – based on the presence of steatosis, clarification/ballooning of hepatocytes and lobular inflammation [13]. 2.4. Statistical analysis Data were expressed as medians (range) or percentages. The Chi-square or Fisher exact test was used to compare categorical variables, and the Wilcoxon test was or t-test used to compare continuous variables. A p-value of <0.05 was considered to be significant. To determine which baseline risk factors were independently associated with a diagnosis of HCC in non-advanced fibrosis, binary logistic regression analysis was performed after the Chi square test (or Fisher exact test, when appropriate). In complement, conditional logistic regression was used to investigate the relationship between HCC and a set of baseline factors in a matched case–control cohort. Patients were matched according to age where the case represented patients with CHC-HCC and control was represented patients by CHC without HCC. This multivariate model included the covariates identified as significant in the univariate analysis cohort matched. Survival analysis was performed using Kaplan–Meier estimates and differences were assessed using log rank tests. All the statistical analyses were performed using SAS 9.3 Software (SAS Institute Inc., Cary, NC, USA). 3. Results 3.1. Study population One hundred sixty-two patients with CHC and HCC were included (61% male, median age 62 years, 64% Caucasian, 78% genotype 1). Forty-five patients who had received IFN were excluded (among them 3 patients had non-advanced fibrosis). Of 162 patients, 137 had HCC and advanced fibrosis (84.5%), while the remainder had HCC without advanced fibrosis. Baseline characteristics are reported in Table 1. The overall performance status test (according to the ECOG) was similar in both groups. Median followup was 30 months (range: 1–143). Median age was 62 and 65 years in the advanced fibrosis and non-advanced fibrosis groups respectively (p = 0.025). HBV markers were less prevalent in the advanced fibrosis than in the non-advanced fibrosis group (18% vs. 50% antiHBc-positive, respectively; p = 0.007). ALT, AST, platelet count and prothrombin time significantly differed between the two groups. Median duration of HCV infection was significantly shorter in the advanced fibrosis than in the non-advanced fibrosis group (25 vs. 35 years, respectively; p < 0.001). The proportion of patients with the
Please cite this article in press as: K-Kutala B, et al. Patients with chronic hepatitis C without advanced fibrosis and hepatocellular carcinoma: A retrospective clinical–pathological study. Dig Liver Dis (2015), http://dx.doi.org/10.1016/j.dld.2014.12.010
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Table 1 Baseline characteristics of 162 patients with chronic hepatitis C and hepatocellular carcinoma. Advanced fibrosis (F3-F4) n = 137
Non advanced fibrosis (F0-F2) n = 25
p-Value
Clinical parameters Male gender Median age (years) Duration of infection (years) HCC under surveillance
70 (71%) 62 [54–72] 25 [22–29] 83 (60%)
14 (74%) 65 [60–74] 35 [31–36] 7 (28%)
Ethnicity Caucasian
89 (64%)
13 (52%)
0.658 0.240
Comorbidities BMI (median) Tobacco Diabetes Arterial hypertension History of alcohol use Metabolic syndrome
25 [22–28] 63 (45%) 41 (29%) 37 (27%) 46 (33%) 34 (25%)
24 [22–25] 9 (36%) 3 (12%) 10 (40%) 3 (12%) 5 (20%)
0.059 0.194 0.066 0.180 0.032 0.631
Laboratory data Abnormal total cholesterol Anti-HBc+ Anti-HBs+ Alpha-fetoprotein (ng/ml) ALT (×ULN) AST (×ULN) Genotype 1 HCV-RNA (Log) Ferritin (mg/dL) Platelet count (×109 /L) Prothrombin time (%)
29 (21%) 33 (24%) 28 (20%) 20 [7–105] 2.0 [1.2–3.0] 2.6 [1.8–3.8] 90 (65%) 5.3 [5–6] 282 [166–523] 112 [90–167] 76 [64–88]
3 (12%) 14 (56%) 10 (40%) 16 [10–300] 1.5 [1.2–2.0] 1.6 [1.3–2.2] 18 (72%) 4.7 [4.5–5.2] 138 [87–238] 215 [154–242] 90 [82–97]
0.841 0.025 <0.001 0.001
0.138 0.001 0.032 0.587 0.022 <.001 0.509 0.006 0.007 <.001 0.005
ALT, alanine aminotransferase; AST, aspartate aminotransferase; Anti-HBc, anti-hepatitis B core antigen; IQR, interquartile range; BMI, body mass index; ULN, upper limit of normal. Data are presented as No./Total No. (%) or median [interquartile range], unless otherwise noted. See “Methods”. Baseline parameters were compared between patients with and without advanced fibrosis using the Mann–Whitney test for continuous variables and the fisher or 2 test for categorical variables.
metabolic syndrome was not statistically different between groups (Table 1). 3.2. Histological analysis The proportion of tumours ≥3 cm was significantly lower in the advanced fibrosis than in the non-advanced fibrosis group (31% vs. 48 respectively; p = 0.013). Solitary tumours were less frequent in the advanced fibrosis than in the non-advanced fibrosis group (54% vs. 68% respectively; p = 0.043). Proportion of welldifferentiated tumours was lower in the advanced fibrosis than in the non-advanced fibrosis group (28% vs. 68%, respectively; p < 0.001). Proportion of patients with steatosis was significantly higher in the advanced fibrosis than in the non-advanced fibrosis group (43% vs. 20% respectively; p = 0.003); prevalence of NASH did not differ among the two groups (Table 2). 3.3. Risk factors and survival analysis To identify factors that might be associated with HCC in patients without advanced fibrosis, we performed a multiple logistic regression analysis with HCC as the dependent variable. Parameters that were significant (p-value ≤ 0.02) in the univariate analysis were introduced in the model, with the exception of age and gender which were included regardless of p-value. Anti-HBc (OR: 3.86 [1.03–14.4], p = 0.044), duration of HCV infection (OR: 1.21 [1.09–1.34], p = 0.003), platelet count (1.00 [1.00–1.04], p = 0.006) and prothrombin time (OR: 1.05 [1.01–1.10], p = 0.016) were significantly associated with HCC. Multivariate analysis results are shown in Table 3. The complementary analysis based on a matched cohort demonstrated that, by taking into account the matching factor (age), the duration of
HCV infection (OR: 1.68[1.08–2.69], p = 0.021) is an independent risk factor for HCC in non-advanced fibrosis. History of alcohol intake (p = 0.011), diabetes (p = 0.022) and platelet count were independently associated with the development of HCC in patients with advanced fibrosis, according to multivariate analysis of the case–control matched population (Table 4). The cumulative incidence of death or liver transplantation at 3 and 5 years after liver resection or loco-regional therapies was 42% and 22% for patients in the advanced fibrosis group, and 59% and 35% for patients in the non-advanced fibrosis group. 4. Discussion This series of 162 patients was based on strict inclusion criteria to determine the risk factors for HCC in CHC patients with and without advanced fibrosis. Our results did not demonstrate an independent association of NASH or steatosis with HCC. We have described the association between steatosis and fibrosis and the impact of interferon therapy on fibrosis in HCV infection in prior studies [14–16]. The increased incidence of HCC in patients with chronic HCV infection and non-alcoholic fatty liver disease (NAFLD) worldwide seems to be due to the increase in obesity and an ageing population [17,18]. Although most CHC-HCC occurs in the presence of severe liver fibrosis or cirrhosis, the process leading to the malignant transformation of HCV-infected hepatocytes is still unclear. The frequency of CHC-HCC in the absence of advanced fibrosis or cirrhosis is also unknown [19]. In the present study, HCC was present in 16% of CHC patients without advanced fibrosis. This is similar to the rate found by Yeh et al. [20]. In the HALT-C study the cumulative 5-year incidence of HCC was higher in patients with cirrhosis than in those with bridging fibrosis but this difference was not significant (7.0% vs. 4.1%, respectively; p = 0.08) [21].
Please cite this article in press as: K-Kutala B, et al. Patients with chronic hepatitis C without advanced fibrosis and hepatocellular carcinoma: A retrospective clinical–pathological study. Dig Liver Dis (2015), http://dx.doi.org/10.1016/j.dld.2014.12.010
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Table 2 Pathological characteristics of 162 patients with chronic hepatitis C and hepatocellular carcinoma. Advanced fibrosis (F3-F4) n = 137
Non advanced fibrosis (F0-F2) n = 25
p-Value
Tumour morphology Tumour size ≥3 cm
43 (31%)
12 (48%)
0.013
Tumour type, n (%) Solitary
75 (54%)
17 (68%)
0.043
Differentiation, n (%) Well Moderate Poor
39 (28%) 47 (34%) 52 (38%)
17 (68%) 5 (20%) 3 (12%)
Extension of vascular invasion, n (%) Microscopic Gross None or (unknown)
58 (42%) 14 (10%) 66 (48%)
13 (52%) 4 (16%) 8 (32%)
0.051
Grade of activity, n (%) A0-A1 A2-A3
85 (62%) 53 (38%)
19 (76%) 6 (24%)
0.032
Other parameters Iron Overload Steatosis NASH
25 (18%) 59 (43%) 23 (17%)
3 (12%) 5 (20%) 3 (12%)
0.234 0.004 0.315
0.010 <0.001
NASH, non-alcoholic steatoheaptitis. Data are presented as No./Total No. (%). See “Methods”. p-Value comparing patients with and without advanced fibrosis and the fisher or 2 test was used if necessary.
In the present study, we characterised risks factors such as NASH, steatosis, diabetes and the metabolic syndrome for the development of HCC in the setting of CHC without advanced fibrosis. NASH has been suggested to be a possible cause of cirrhosis, although most of the histological characteristics of this entity are not present in cryptic cirrhosis [22]. In the absence of reliable clinical features or new biochemical markers, the diagnosis of NASH is only based on histopathology [11,23]. Because studies on NAFLD have shown that only patients with biopsy-proven NASH progress to cirrhosis, it is extremely important to clearly define the histopathological criteria for NAFLD and
NASH in CHC [11,24]. In the presence of CHC, lesions related to the underlying chronic inflammatory disease increase the complexity of identifying NASH, because certain features may overlap (steatosis, lobular inflammation) [14]. Hence, we reviewed files of patients who underwent liver resection or had two biopsies at the time of HCC diagnosis. In this study, only 26 patients in the study population had NASH: 17% in the advanced fibrosis group vs. 12% in the non-advanced fibrosis group (p = 0.315). Steatosis was more frequent in the F3-F4 group than in F0-F2 group (43% vs. 20%). These results do not support the hypothesis of steato-inflammation or NAFLD in patients without advanced fibrosis. As was reported,
Table 3 Univariate and multivariate analysis of predictors of hepatocellular carcinoma in patients with chronic hepatitis C without advanced fibrosis. Parameters
Multivariatea
Univariate OR (95% CI)
p-Value
Age Anti-HBc Anti-HBs ALT AST
1.05 (1.00–1.10) 4.05 (1.67–9.77) 2.61 (1.06–6.45) 0.54 (0.33–0.88) 0.38 (0.22–0.66)
0.024 0.001 0.036 0.015 0.006
Arterial hypertension BMI (kg/m2 ) Caucasian race Diabetes Duration of infection Male gender History of alcohol use Iron hemosiderin NASH
0.86 (0.76–0.99) 0.59 (0.25–1.40) 0.32 (0.92–1.13) 1.17 (1.09–1.26) 0.94 (0.36–2.43) 0.27 (0.17–0.95) 0.76 (0.20–2.76) 0.68 (0.88–2.46)
0.034 0.238 0.078 <.001 0.902 0.042 0.677 0.560
Metabolic syndrome Platelet count Prothrombin time Steatosis Tobacco Total cholesterol (mg/dL)
1.00 (1.01–1.10) 1.05 (1.01–1.08) 0.33 (0.12–0.94) 0.56 (0.22–1.38) 0.51 (0.14–1.83)
<.001 0.001 0.038 0.209 0.303
OR (95% CI)
p-Value
3.86 (1.03–14.4)
0.044
0.46 (0.23–0.94)
0.034
1.21 (1.09–1.34)
0.003
1.00 (1.00–1.04) 1.05 (1.01–1.10)
0.006 0.016
ALT, alanine aminotransferase; AST, aspartate aminotransferase; Anti-HBc, anti-hepatitis B core antigen; Anti-HBs, anti-hepatitis B surface antigen; BMI, body mass index; NASH, non alcoholic steato-hepatitis; OR, odds ratio; CI, confidence interval. a Multivariate included age and gender plus all variables with p ≤ 0.02. The influence of age is already known and AST was not included because ALT was also included. If AST was included willingly, the result did not change.
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B. K-Kutala et al. / Digestive and Liver Disease xxx (2015) xxx–xxx Table 4 Multivariate binary logistic regression evaluating baseline factors associated with hepatocellular carcinoma in 324 matched patients infected with chronic hepatitis C. Multivariate OR
(95% CI)
p-Value
Non-fibrotic (F0-F2) ALT AST Infection duration
0.14 2.14 1.68
(0.20–2.32) (0.29–23.9) (1.08–2.69)
0.178 0.464 0.021
Fibrotic (F3-F4) AST Diabetes Gender History of alcohol use Platelet account Prothrombin time
1.16 3.13 2.36 3.14 1.00 0.95
(0.91–1.49) (1.17–8.38) (0.90–6.15) (1.29–7.62) (1.00–1.04) (0.93–0.98)
0.222 0.022 0.077 0.011 0.013 0.001
ALT, alanine aminotransferase; AST, aspartate aminotransferase; OR, odds ratio; CI, confidence interval. Multivariate including all significant baseline variables with p ≤ 0.02 in univariate analysis.
NAFLD and NASH can progress to cirrhosis and liver failure in 3–15% of patients and lead to HCC [25]. The mechanisms of the pathogenesis of NASH in the development of HCC in patients with CHC infection have not been clarified. Moreover diabetes and obesity have been shown increase the risk of HCC [26]. Despite a lack of strong evidence, this study examines the role of the metabolic syndrome, diabetes and as well as the effect of HCV on the relationship between these risk factors and HCC. However, only 12% and 20% patients had diabetes and metabolic syndrome without visceral obesity in the F0-F2 group.
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Although it has been suggested that the association of NASH and obesity increases the risk of HCC in chronic hepatitis C [27], this association was not found in our series. The most important finding in this study was the association of baseline diabetes and excessive alcohol intake with an increased occurrence of HCC in patients with advanced fibrosis. Although a significant impact of diabetes on overall liver-related outcome may be overlooked, the association of diabetes with an accelerated progression to cirrhosis and HCC in patients with CHC is well known [28] (Tables 4 and 5). Moreover, multivariate analysis revealed that the risk of HCC in patients without advanced fibrosis increased in anti-HBc-positive patients (OR: 3.86) and with increased duration of HCV infection (OR: 1.21). The significantly higher percentage of HBV marker in patients F0-F2 than in F3-F4 (24% vs. 56%, p = 0.001) suggests a possible role of occult HBV infection in the former group. Occult HBV infection has been defined by the European Association for the Study of the Liver (EASL) [29] and the Taormina Consensus Conference in 2008 as the “presence of HBV DNA in the liver of individuals testing HBsAg-negative with currently available assays” and introduced a cut-off value for serum HBV DNA (<200 IU/mL). The relationship between occult HBV infection and serological markers of HBV infection has been extensively studied [30]. There is also significant evidence suggesting that occult HBV is a risk factor for the development of HCC in both patients with HCV-related or not cryptogenic chronic liver disease [31–33]. Moreover Pollicino et al. have shown that many of the potential mechanisms of tumour genesis in overt HBV are maintained in case of occult HBV infection [34]. However, those were cross-sectional case studies examining patients with existing HCC. To our knowledge there are no existing prospective studies investigating the development
Table 5 Baseline characteristics of 324 age-matched chronic hepatitis C patients, with or without hepatocellular carcinoma. All n = 324
Controls n = 162
Cases n = 162
Clinical parameters Male gender Median age (years) Duration of infection (years)
220 (68%) 63 [54–71] 26 [22–32]
102 (63%) 61 [53–69] 26 [20–32]
118 (73%) 63 [55–72] 27 [23–32]
0.056 0.087 0.138
Ethnicity Caucasian
209 (65%)
107 (66%)
102 (63%)
0.561
Comorbidities BMI (median) Tobacco Diabetes Arterial hypertension History of alcohol use
25 [22–27] 125 (39%) 64 (20%) 82 (25%) 72 (22%)
Laboratory data Abnormal total cholesterol Anti-HBc+ Anti-HBs+ Alpha-fetoprotein (ng/ml) ALT (×ULN) AST (×ULN) Genotype 1 HCV-RNA (Log) Ferritin (mg/dL) Platelet count (×109 /L) Prothrombin time (%)
50 (15%) 84 (26%) 63 (19%) 6 [3–13] 2.0 [1.4–3.0] 1.8 [1.1–2.6] 212 (65%) 5.1 [5–6.3] 247 [125–402] 174 [123–219] 90 [79–100]
18 (11%) 38 (23%) 26 (16%) 5 [3–8] 2.0 [1.4–3.1] 1.6 [1.1–2.4] 105 (65%) 5.4 [5–6.4] 241 [122–390] 117 [95–178] 92 [83–100]
32 (20%) 46 (28%) 37 (23%) 16 [8–105] 1.8 [1.2–2.8] 2.2 [1.6–2.9] 107 (66%) 5.0 [4.6–5.5] 269 [134–457] 186 [139–228] 77 [64–90]
Other parameters F0-F2 F3-F4 Steatosis
50 (15%) 247 (85%) 114 (35%)
25 (15%) 137 (85%) 50 (31%)
25 (15%) 137 (85%) 64% (39%)
24 [22–27] 54 (33%) 20 (12%) 35 (22%) 23 (14%)
25 [23–27] 71 (44%) 44 (27%) 47 (29%) 49 (30%)
p-Value
0.143 0.052 0.008 0.125 0.005 0.031 0.310 0.122 <0.001 0.031 <.001 0.815 0.041 0.122 <0.001 <0.001
0.101
Abbreviations: ALT, alanine aminotransferase; AST, aspartate aminotransferase; Anti-HBc+, anti-hepatitis B core antigen positive; Anti-HBs+, anti-hepatitis B surface antigen positive; IQR, interquartile range; ULN, upper limit of normal; BMI, body mass index, calculated as weight in kilograms divided by height in metres squared. Data are presented as No./Total No. (%) or median [interquartile range], unless otherwise noted. See “Methods”. Baseline parameters were compared between patients with and without advanced fibrosis using signed rank Wilcoxon test for continuous variables and the paired t test for categorical variables. Case = with HCC, Control = without CHC matched by the age.
Please cite this article in press as: K-Kutala B, et al. Patients with chronic hepatitis C without advanced fibrosis and hepatocellular carcinoma: A retrospective clinical–pathological study. Dig Liver Dis (2015), http://dx.doi.org/10.1016/j.dld.2014.12.010
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of HCC in patients with CHC in relation to their occult HBV status. Squadrito et al. [32] have also shown that occult HBV may be associated with the development of HCC in chronic liver disease and confirmed that, in HBsAg-negative individuals, those with the HBV genomic sequences have a higher risk of developing HCC. Additionally, the HCV virus contains some hydrophobic and hydrophilic proteins, such as proteins NS3, NS4B and NS5A that have a regulatory effect on cell promoters and seem to interact with some of the cellular proteins involved in the mechanism of carcinogenesis under certain specific conditions [35]. Our findings show that the duration of HCV infection (OR: 1.21, p = 0.003) is an independent factor and may influence development of HCC in CHC patients without advanced fibrosis. This factor is well recognised (at least for HCV-induced cirrhosis) as strongly correlated to HCC development. However, most studies on HCC risk factors in cirrhosis or advanced fibrosis did not adequately take into account duration of infection as a confounding factor. This duration of infection in patients without advanced fibrosis may suggest that a few years of inflammation can lead to HCC. More data are needed to clarify this issue. Other baseline factors significantly associated with HCC occurrence in patients without advanced fibrosis at multivariate analysis were ALT, platelet count, and prothrombin level. Those factors are already known to have an impact on disease progression and they are surrogate markers of inflammatory activity of the liver, and portal hypertension. The complementary analysis, including matched patients, revealed implications of some variables known as risk factors of HCC or progression of liver disease – such as, alcohol intake, diabetes and platelet count. Although optimal screening protocols and monitoring tools are still a subject of debate, the particularly poor survival of HCV patients without advanced fibrosis in our series (35% at 5 years, Log-rank, p = 0.038) is alarming and underscores the importance of surveillance in the HCV-infected population without advanced fibrosis. Of note, in our study 48% of patients without advanced fibrosis had a tumour ≥3 cm vs. 31% of patients in the advanced fibrosis group. The effectiveness of screening programmes will depend on external factors, such as physician and patient acceptance and adherence to the organization’s programme, economic considerations, and a host of other factors, including availability and accessibility to diagnostic tools, treatment and adequate follow-up [36]. Although confounding factors were taken into account, this study has several limitations. Firstly, the study population was small, which might induce a selection bias and can have an impact on the interpretation of our findings. However compelling our findings may be, the absence of RT-PCR on liver tissue samples limits the power of this study. This study suggests that anti-HBc, diabetes and alcohol intake remain the main risk factors for HCC in patients with advanced fibrosis. These results could modify the clinical management of these patients. To understand the precise mechanisms of the development of HCC in CHC without advanced fibrosis, the pathogenic role of each risk factor and the epidemiology of occult HBV infection must be more clearly defined. Thus, testing patients with CHC for HBV genomes could be a powerful tool to identify subjects who require careful screening to obtain an early diagnosis of HCC. Conflict of interest None declared. References [1] El-Serag HB. Hepatocellular carcinoma. New England Journal of Medicine 2011;365:1118–27.
[2] El-Serag HB, Rudolph KL. Hepatocellular carcinoma: epidemiology and molecular carcinogenesis. Gastroenterology 2007;132:2557–76. [3] Sherman M. Epidemiology of hepatocellular carcinoma. Oncology 2010;78(Suppl. 1):7–10. [4] Beasley RP, Hwang LY, Lin CC, et al. Hepatocellular carcinoma and hepatitis B virus. A prospective study of 22 707 men in Taiwan. Lancet 1981;2: 1129–33. [5] Buendia MA. Hepatitis B viruses and hepatocellular carcinoma. Advances in Cancer Research 1992;59:167–226. [6] Fattovich G, Stroffolini T, Zagni I, et al. Hepatocellular carcinoma in cirrhosis: incidence and risk factors. Gastroenterology 2004;127:S35–50. [7] Gomaa AI, Khan SA, Toledano MB, et al. Hepatocellular carcinoma: epidemiology, risk factors and pathogenesis. World Journal of Gastroenterology: WJG 2008;14:4300–8. [8] Yoshida H, Shiratori Y, Moriyama M, et al. Interferon therapy reduces the risk for hepatocellular carcinoma: national surveillance program of cirrhotic and noncirrhotic patients with chronic hepatitis C in Japan. IHIT Study Group. Inhibition of Hepatocarcinogenesis by Interferon Therapy. Annals of Internal Medicine 1999;131:174–81. [9] Paradis V, Zalinski S, Chelbi E, et al. Hepatocellular carcinomas in patients with metabolic syndrome often develop without significant liver fibrosis: a pathological analysis. Hepatology 2009;49:851–9. [10] Davila JA, Henderson L, Kramer JR, et al. Utilization of surveillance for hepatocellular carcinoma among hepatitis C virus-infected veterans in the United States. Annals of Internal Medicine 2011;154:85–93. [11] Kleiner DE, Brunt EM, Van Natta M, et al. Design and validation of a histological scoring system for nonalcoholic fatty liver disease. Hepatology 2005;41:1313–21. [12] Bedossa P, Poynard T. An algorithm for the grading of activity in chronic hepatitis C. The METAVIR Cooperative Study Group. Hepatology 1996;24: 289–93. [13] Clouston AD, Jonsson JR, Purdie DM, et al. Steatosis and chronic hepatitis C: analysis of fibrosis and stellate cell activation. Journal of Hepatology 2001;34:314–20. [14] Asselah T, Boyer N, Guimont MC, et al. Liver fibrosis is not associated with steatosis but with necroinflammation in French patients with chronic hepatitis C. Gut 2003;52:1638–43. [15] Marcellin P, Asselah T, Boyer N. Fibrosis and disease progression in hepatitis C. Hepatology 2002;36:S47–56. [16] Cardoso AC, Moucari R, Figueiredo-Mendes C, et al. Impact of peginterferon and ribavirin therapy on hepatocellular carcinoma: incidence and survival in hepatitis C patients with advanced fibrosis. Journal of Hepatology 2010;52: 652–7. [17] Zimmet P, Magliano D, Matsuzawa Y, et al. The metabolic syndrome: a global public health problem and a new definition. Journal of Atherosclerosis and Thrombosis 2005;12:295–300. [18] Alberti KG, Zimmet P, Shaw J, Group IDFETFC. The metabolic syndrome – a new worldwide definition. Lancet 2005;366:1059–62. [19] Nash KL, Woodall T, Brown AS, et al. Hepatocellular carcinoma in patients with chronic hepatitis C virus infection without cirrhosis. World Journal of Gastroenterology: WJG 2010;16:4061–5. [20] Yeh MM, Daniel HD, Torbenson M. Hepatitis C-associated hepatocellular carcinomas in non-cirrhotic livers. Modern Pathology 2010;23:276–83. [21] Lok AS, Seeff LB, Morgan TR, et al. Incidence of hepatocellular carcinoma and associated risk factors in hepatitis C-related advanced liver disease. Gastroenterology 2009;136:138–48. [22] Bugianesi E, Leone N, Vanni E, et al. Expanding the natural history of nonalcoholic steatohepatitis: from cryptogenic cirrhosis to hepatocellular carcinoma. Gastroenterology 2002;123:134–40. [23] Brunt EM. Nonalcoholic steatohepatitis. Seminars in Liver Disease 2004;24:3–20. [24] Leandro G, Mangia A, Hui J, et al. Relationship between steatosis, inflammation, and fibrosis in chronic hepatitis C: a meta-analysis of individual patient data. Gastroenterology 2006;130:1636–42. [25] Angulo P. Nonalcoholic fatty liver disease. New England Journal of Medicine 2002;346:1221–31. [26] Welzel TM, Graubard BI, Zeuzem S, El-Serag HB, et al. Metabolic syndrome increases the risk of primary liver cancer in the United States: a study in the SEER-Medicare database. Hepatology 2011;54:463–71. [27] El-Serag HB, Mason AC. Risk factors for the rising rates of primary liver cancer in the United States. Archives of Internal Medicine 2000;160: 3227–30. [28] El-Serag HB. Epidemiology of viral hepatitis and hepatocellular carcinoma. Gastroenterology 2012;142:1264–73, e1. [29] Raimondo G, Navarra G, Mondello S, et al. Occult hepatitis B virus in liver tissue of individuals without hepatic disease. Journal of Hepatology 2008;48: 743–6. [30] Marcellin P, Thiers V, Degott C, et al. Hepatocellular carcinoma with normal adjacent liver. Hepatitis B virus DNA status. Journal of Hepatology 1989;8:249–53. [31] Cacciola I, Pollicino T, Squadrito G, Cerenzia G, Orlando ME, Raimondo G. Occult hepatitis B virus infection in patients with chronic hepatitis C liver disease. New England Journal of Medicine 1999;341:22–6. [32] Squadrito G, Pollicino T, Cacciola I, et al. Occult hepatitis B virus infection is associated with the development of hepatocellular carcinoma in chronic hepatitis C patients. Cancer 2006;106:1326–30.
Please cite this article in press as: K-Kutala B, et al. Patients with chronic hepatitis C without advanced fibrosis and hepatocellular carcinoma: A retrospective clinical–pathological study. Dig Liver Dis (2015), http://dx.doi.org/10.1016/j.dld.2014.12.010
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[33] Squadrito G, Cacciola I, Aldibrandi A, et al. Impact of occult hepatitis B virus infection on the outcome of chronic hepatitis C. Journal of Hepatology 2013;59:696–700. [34] Pollicino T, Squadrito G, Cerenzia G, et al. Hepatitis B virus maintains its prooncogenic properties in the case of occult HBV infection. Gastroenterology 2004;126:102–10.
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[35] Zemel R, Gerechet S, Greif H, et al. Cell transformation induced by hepatitis C virus NS3 serine protease. Journal of Viral Hepatitis 2001;8: 96–102. [36] Andersson KL, Salomon JA, Goldie SJ, Chung RT. Cost effectiveness of alternative surveillance strategies for hepatocellular carcinoma in patients with cirrhosis. Clinical Gastroenterology and Hepatology 2008;6:1418–24.
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Contents lists available at ScienceDirect
Digestive and Liver Disease journal homepage: www.elsevier.com/locate/dld
Liver, Pancreas and Biliary Tract
Patients with chronic hepatitis C without advanced fibrosis and hepatocellular carcinoma: A retrospective clinical–pathological study Blaise K-Kutala a,b,d,∗ , Pierre Bedossa c , Jeremie Guedj b , Tarik Asselah a,d , Michelle Martinot-Peignoux d , Xavier Duval b , Patrick Marcellin a,d a
Department of Hepatology, Beaujon Hospital, AP-HP, Clichy, France INSERM U1137, University Paris 7, UFR of Medicine, France c Department of Pathology, Beaujon Hospital, AP-HP, Clichy, France d INSERM U1149/CRI, Beaujon Hospital, AP-HP, Clichy, France b
a r t i c l e
i n f o
Article history: Received 25 July 2014 Accepted 17 December 2014 Available online xxx Keywords: Cirrhosis Hepatitis C Hepatocellular carcinoma Nonalcoholic steatohepatitis Screening
a b s t r a c t Background: There are very few studies on the incidence and risk factors of hepatitis C virus (HCV)-induced hepatocellular carcinoma (HCC) in the absence of advanced fibrosis. Our objective was to identify the clinical–pathological features of these patients. Methods: We retrospectively reviewed 162 patients admitted to our hospital for HCV-related HCC between 2000 and 2010. Patients with hepatitis of other aetiologies, human immunodeficiency virus co-infection, or treated with interferon were excluded. We compared demographic, laboratory, clinical and outcome parameters of patients with and without advanced fibrosis. Results: 137 patients had advanced fibrosis (85%). Median age was higher in the advanced fibrosis vs. the non-advanced fibrosis group (62 vs. 65 years, respectively; p = 0.025). Steatosis was significantly more frequent in patients with advanced fibrosis compared to those without advanced fibrosis (43% vs. 20%, respectively; p = 0.032). Independent predictors associated to the occurrence of HCC in patients without advanced fibrosis were hepatitis B core antigen (odds ratio: 3.86; p = 0.044) and duration of hepatitis C infection (odds ratio: 1.21; p = 0.003). Conclusions: Risk factors such as steatosis or diabetes were not frequent in patients without advanced fibrosis. Further studies are needed to evaluate the role of occult hepatitis B and the duration of hepatitis infection in patients with HCC and chronic hepatitis C without advanced fibrosis. © 2014 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.
1. Introduction Hepatocellular carcinoma (HCC) the most common liver cancer; representing 90% of primary liver cancer worldwide, with an increasing incidence in the United States, Western Europe, and Japan [1]. Although HCC usually develops in cirrhotic livers, a proportion also develops in the absence of advanced hepatic fibrosis or cirrhosis. The frequency of this event varies along geographic and ethnic lines and is closely associated with etiological factors [2]. Hepatitis B virus (HBV), for example, is known to have a direct oncogenic effect on the liver by integrating into the host genome [3], and the prevalence of non-cirrhotic HCC is high (20%) in regions
∗ Corresponding author at: Service d’Hépatologie, Hôpital Beaujon AP-HP, 100 Boulevard Général Leclerc, 92110 Clichy, France. Tel.: +33 1 40 87 54 31; fax: +33 1 47 30 94 40. E-mail address: [email protected] (B. K-Kutala).
where this HBV is endemic, such as China and Taiwan [4]. There is also a population of patients that develops HCC in the absence of underlying parenchymal liver disease or exposure to known environmental risk factors, such as aflatoxin B1, nitrosamine or benzopyrenes [5]. Hepatitis C virus (HCV) infection is a well-known cause of HCC; the estimated risk of HCC is 15–20 times higher in patients with chronic hepatitis C (CHC) (30% in Asia and 17% in Western countries) than in those without, and the greatest risk is found in patients with advanced hepatic fibrosis or cirrhosis [6,7]. Unfortunately, there are very few, and sometimes contradictory studies on the incidence of HCC in patients with chronic hepatitis C without cirrhosis [8,9]. Until now, practical clinical guidelines and recommendations have been focused on the screening for HCC in patients with cirrhosis or advanced fibrosis related to chronic hepatitis C but not in those without advanced fibrosis [10]. Thus, the aims of this study were to analyse and compare the main clinical features and tumour characteristics at diagnosis of patients with CHC-related
http://dx.doi.org/10.1016/j.dld.2014.12.010 1590-8658/© 2014 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.
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HCC without advanced fibrosis to those of patients with HCC on cirrhosis or advanced fibrosis. 2. Patients and methods 2.1. Study design and data source We retrospectively all consecutive patients in our referral hospital who followed-up for HCC care between January 2000 and December 2010 and fulfilled the following criteria: (1) CHC defined by the association of positive serum anti-HCV antibodies and RNA, with liver histology corresponding to a METAVIR Score of F3-F4 for patients with advanced fibrosis and F0-F2 for those with nonadvanced fibrosis; (2) diagnosis of HCC and staging of the chronic hepatitis, based on the surgical specimen or two adequate liver biopsies (in non-tumorous liver tissue and in HCC nodule) reviewed in our pathology department by an experienced liver pathologist; (3) no other coexisting liver disease, such as HBV, or human immunodeficiency virus (HIV) co-infection (negative serum HBsAg and negative anti-HIV antibodies); (4) daily alcohol consumption <40 g for males and <30 g for females. Thus patients without advanced fibrosis previously treated with interferon (IFN) were excluded. The inclusion period was extended to include more patients without advanced fibrosis. Patients with advanced fibrosis underwent a standard surveillance programme; diagnosis of HCC was based on the following: biopsy from 2001, a combination of imaging and laboratory findings [computed tomography (CT) scan or Magnetic resonance imaging (MRI) and alpha-fetoprotein serum levels (AFP) > 400 ng/ml] between 2001 and 2004 and after 2005; and if necessary, on contrast uptake in the arterial phase and washout in the venous/late phase on CT, MRI or ultrasound or biopsy. Although patients without fibrosis did not undergo routine screening, HCC was usually detected during routine visits for the follow-up of CHC or other comorbid conditions such as arterial hypertension or diabetes mellitus. In some of these cases HCC was diagnosed at the same time as CHC, or was sought because of known risk factors (age >65 years, male gender, elevated serum AFP). 2.2. Clinical and biochemical characteristics We performed a retrospective analysis of a prospective database of patients with CHC-HCC who underwent liver resection in our hospital. Patient files were reviewed and analysed for the following demographic, clinical, and biochemical data collected at admission: age, gender, cause of cirrhosis, HBV markers (AntiHBc, Anti-HBs) and HCV-RNA; aspartate aminotransferase, alanine aminotransferase, total bilirubin, albumin, and serum AFP; prothrombin activity, platelet count, viral load and transmission of HCV infection. Complications were also recorded: Child Pugh score, the presence of ascites, encephalopathy, oesophageal varices, and a history of variceal or gastrointestinal bleeding. The relationship between the metabolic syndrome and HCC in patients without cirrhosis was studied in our series. Metabolic syndrome was defined as a combination of any 3 of the 5 following risk factors: waist circumference ≥102 cm in men and ≥88 cm in women of European descent (≥94 cm and ≥80 for African descent), triglycerides ≥150 mg/dL, HDL-C < 40 mg/dL in men and <50 mg/dL in women, arterial hypertension, and elevated fasting glucose. The estimated duration of infection was defined as the time that elapsed from the presumed date of infection (date of transfusion, first year of intravenous drug use, date of vaccination for African patients, unsterile medical and dental procedures, and traditional medical and cosmetic procedures involving blood exposure) to the date of HCC diagnosis.
2.3. Histological data Each liver resection sample was individually coded and processed for histopathological interpretation by a single experienced liver pathologist (P. B.), blinded to the patient’s identity, and clinical and biological data and was routinely processed, formalinfixed, serially sectioned, and stained with haematoxylin/eosin, Masson’s trichrome, and Sirius red to evaluate fibrosis and Perls staining for iron. A detailed scoring sheet was used; including the grading and staging of fibrosis by the METAVIR classification, qualitative evaluation of steatosis (0, absent 1, present), predominant zonal distribution of fat (pericentral, periportal, azonal, diffuse), clarification/ballooning of liver cells (0, absent; 1, mild; 2, marked), perisinusoidal fibrosis (0, absent; 1, mild; 2, marked), Mallory hyaline (0, absent; 1, present) and presence of glycogenated nuclei (0, absent; 1, present) as well as other less common histological features described by Kleiner et al. [11]. The following histological characteristics were analysed for each patient: METAVIR score [12], number and size of nodules, presence of microvascular invasion, presence of steatosis, iron overload, presence of non-alcoholic steatohepatitis (NASH) – based on the presence of steatosis, clarification/ballooning of hepatocytes and lobular inflammation [13]. 2.4. Statistical analysis Data were expressed as medians (range) or percentages. The Chi-square or Fisher exact test was used to compare categorical variables, and the Wilcoxon test was or t-test used to compare continuous variables. A p-value of <0.05 was considered to be significant. To determine which baseline risk factors were independently associated with a diagnosis of HCC in non-advanced fibrosis, binary logistic regression analysis was performed after the Chi square test (or Fisher exact test, when appropriate). In complement, conditional logistic regression was used to investigate the relationship between HCC and a set of baseline factors in a matched case–control cohort. Patients were matched according to age where the case represented patients with CHC-HCC and control was represented patients by CHC without HCC. This multivariate model included the covariates identified as significant in the univariate analysis cohort matched. Survival analysis was performed using Kaplan–Meier estimates and differences were assessed using log rank tests. All the statistical analyses were performed using SAS 9.3 Software (SAS Institute Inc., Cary, NC, USA). 3. Results 3.1. Study population One hundred sixty-two patients with CHC and HCC were included (61% male, median age 62 years, 64% Caucasian, 78% genotype 1). Forty-five patients who had received IFN were excluded (among them 3 patients had non-advanced fibrosis). Of 162 patients, 137 had HCC and advanced fibrosis (84.5%), while the remainder had HCC without advanced fibrosis. Baseline characteristics are reported in Table 1. The overall performance status test (according to the ECOG) was similar in both groups. Median followup was 30 months (range: 1–143). Median age was 62 and 65 years in the advanced fibrosis and non-advanced fibrosis groups respectively (p = 0.025). HBV markers were less prevalent in the advanced fibrosis than in the non-advanced fibrosis group (18% vs. 50% antiHBc-positive, respectively; p = 0.007). ALT, AST, platelet count and prothrombin time significantly differed between the two groups. Median duration of HCV infection was significantly shorter in the advanced fibrosis than in the non-advanced fibrosis group (25 vs. 35 years, respectively; p < 0.001). The proportion of patients with the
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Table 1 Baseline characteristics of 162 patients with chronic hepatitis C and hepatocellular carcinoma. Advanced fibrosis (F3-F4) n = 137
Non advanced fibrosis (F0-F2) n = 25
p-Value
Clinical parameters Male gender Median age (years) Duration of infection (years) HCC under surveillance
70 (71%) 62 [54–72] 25 [22–29] 83 (60%)
14 (74%) 65 [60–74] 35 [31–36] 7 (28%)
Ethnicity Caucasian
89 (64%)
13 (52%)
0.658 0.240
Comorbidities BMI (median) Tobacco Diabetes Arterial hypertension History of alcohol use Metabolic syndrome
25 [22–28] 63 (45%) 41 (29%) 37 (27%) 46 (33%) 34 (25%)
24 [22–25] 9 (36%) 3 (12%) 10 (40%) 3 (12%) 5 (20%)
0.059 0.194 0.066 0.180 0.032 0.631
Laboratory data Abnormal total cholesterol Anti-HBc+ Anti-HBs+ Alpha-fetoprotein (ng/ml) ALT (×ULN) AST (×ULN) Genotype 1 HCV-RNA (Log) Ferritin (mg/dL) Platelet count (×109 /L) Prothrombin time (%)
29 (21%) 33 (24%) 28 (20%) 20 [7–105] 2.0 [1.2–3.0] 2.6 [1.8–3.8] 90 (65%) 5.3 [5–6] 282 [166–523] 112 [90–167] 76 [64–88]
3 (12%) 14 (56%) 10 (40%) 16 [10–300] 1.5 [1.2–2.0] 1.6 [1.3–2.2] 18 (72%) 4.7 [4.5–5.2] 138 [87–238] 215 [154–242] 90 [82–97]
0.841 0.025 <0.001 0.001
0.138 0.001 0.032 0.587 0.022 <.001 0.509 0.006 0.007 <.001 0.005
ALT, alanine aminotransferase; AST, aspartate aminotransferase; Anti-HBc, anti-hepatitis B core antigen; IQR, interquartile range; BMI, body mass index; ULN, upper limit of normal. Data are presented as No./Total No. (%) or median [interquartile range], unless otherwise noted. See “Methods”. Baseline parameters were compared between patients with and without advanced fibrosis using the Mann–Whitney test for continuous variables and the fisher or 2 test for categorical variables.
metabolic syndrome was not statistically different between groups (Table 1). 3.2. Histological analysis The proportion of tumours ≥3 cm was significantly lower in the advanced fibrosis than in the non-advanced fibrosis group (31% vs. 48 respectively; p = 0.013). Solitary tumours were less frequent in the advanced fibrosis than in the non-advanced fibrosis group (54% vs. 68% respectively; p = 0.043). Proportion of welldifferentiated tumours was lower in the advanced fibrosis than in the non-advanced fibrosis group (28% vs. 68%, respectively; p < 0.001). Proportion of patients with steatosis was significantly higher in the advanced fibrosis than in the non-advanced fibrosis group (43% vs. 20% respectively; p = 0.003); prevalence of NASH did not differ among the two groups (Table 2). 3.3. Risk factors and survival analysis To identify factors that might be associated with HCC in patients without advanced fibrosis, we performed a multiple logistic regression analysis with HCC as the dependent variable. Parameters that were significant (p-value ≤ 0.02) in the univariate analysis were introduced in the model, with the exception of age and gender which were included regardless of p-value. Anti-HBc (OR: 3.86 [1.03–14.4], p = 0.044), duration of HCV infection (OR: 1.21 [1.09–1.34], p = 0.003), platelet count (1.00 [1.00–1.04], p = 0.006) and prothrombin time (OR: 1.05 [1.01–1.10], p = 0.016) were significantly associated with HCC. Multivariate analysis results are shown in Table 3. The complementary analysis based on a matched cohort demonstrated that, by taking into account the matching factor (age), the duration of
HCV infection (OR: 1.68[1.08–2.69], p = 0.021) is an independent risk factor for HCC in non-advanced fibrosis. History of alcohol intake (p = 0.011), diabetes (p = 0.022) and platelet count were independently associated with the development of HCC in patients with advanced fibrosis, according to multivariate analysis of the case–control matched population (Table 4). The cumulative incidence of death or liver transplantation at 3 and 5 years after liver resection or loco-regional therapies was 42% and 22% for patients in the advanced fibrosis group, and 59% and 35% for patients in the non-advanced fibrosis group. 4. Discussion This series of 162 patients was based on strict inclusion criteria to determine the risk factors for HCC in CHC patients with and without advanced fibrosis. Our results did not demonstrate an independent association of NASH or steatosis with HCC. We have described the association between steatosis and fibrosis and the impact of interferon therapy on fibrosis in HCV infection in prior studies [14–16]. The increased incidence of HCC in patients with chronic HCV infection and non-alcoholic fatty liver disease (NAFLD) worldwide seems to be due to the increase in obesity and an ageing population [17,18]. Although most CHC-HCC occurs in the presence of severe liver fibrosis or cirrhosis, the process leading to the malignant transformation of HCV-infected hepatocytes is still unclear. The frequency of CHC-HCC in the absence of advanced fibrosis or cirrhosis is also unknown [19]. In the present study, HCC was present in 16% of CHC patients without advanced fibrosis. This is similar to the rate found by Yeh et al. [20]. In the HALT-C study the cumulative 5-year incidence of HCC was higher in patients with cirrhosis than in those with bridging fibrosis but this difference was not significant (7.0% vs. 4.1%, respectively; p = 0.08) [21].
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Table 2 Pathological characteristics of 162 patients with chronic hepatitis C and hepatocellular carcinoma. Advanced fibrosis (F3-F4) n = 137
Non advanced fibrosis (F0-F2) n = 25
p-Value
Tumour morphology Tumour size ≥3 cm
43 (31%)
12 (48%)
0.013
Tumour type, n (%) Solitary
75 (54%)
17 (68%)
0.043
Differentiation, n (%) Well Moderate Poor
39 (28%) 47 (34%) 52 (38%)
17 (68%) 5 (20%) 3 (12%)
Extension of vascular invasion, n (%) Microscopic Gross None or (unknown)
58 (42%) 14 (10%) 66 (48%)
13 (52%) 4 (16%) 8 (32%)
0.051
Grade of activity, n (%) A0-A1 A2-A3
85 (62%) 53 (38%)
19 (76%) 6 (24%)
0.032
Other parameters Iron Overload Steatosis NASH
25 (18%) 59 (43%) 23 (17%)
3 (12%) 5 (20%) 3 (12%)
0.234 0.004 0.315
0.010 <0.001
NASH, non-alcoholic steatoheaptitis. Data are presented as No./Total No. (%). See “Methods”. p-Value comparing patients with and without advanced fibrosis and the fisher or 2 test was used if necessary.
In the present study, we characterised risks factors such as NASH, steatosis, diabetes and the metabolic syndrome for the development of HCC in the setting of CHC without advanced fibrosis. NASH has been suggested to be a possible cause of cirrhosis, although most of the histological characteristics of this entity are not present in cryptic cirrhosis [22]. In the absence of reliable clinical features or new biochemical markers, the diagnosis of NASH is only based on histopathology [11,23]. Because studies on NAFLD have shown that only patients with biopsy-proven NASH progress to cirrhosis, it is extremely important to clearly define the histopathological criteria for NAFLD and
NASH in CHC [11,24]. In the presence of CHC, lesions related to the underlying chronic inflammatory disease increase the complexity of identifying NASH, because certain features may overlap (steatosis, lobular inflammation) [14]. Hence, we reviewed files of patients who underwent liver resection or had two biopsies at the time of HCC diagnosis. In this study, only 26 patients in the study population had NASH: 17% in the advanced fibrosis group vs. 12% in the non-advanced fibrosis group (p = 0.315). Steatosis was more frequent in the F3-F4 group than in F0-F2 group (43% vs. 20%). These results do not support the hypothesis of steato-inflammation or NAFLD in patients without advanced fibrosis. As was reported,
Table 3 Univariate and multivariate analysis of predictors of hepatocellular carcinoma in patients with chronic hepatitis C without advanced fibrosis. Parameters
Multivariatea
Univariate OR (95% CI)
p-Value
Age Anti-HBc Anti-HBs ALT AST
1.05 (1.00–1.10) 4.05 (1.67–9.77) 2.61 (1.06–6.45) 0.54 (0.33–0.88) 0.38 (0.22–0.66)
0.024 0.001 0.036 0.015 0.006
Arterial hypertension BMI (kg/m2 ) Caucasian race Diabetes Duration of infection Male gender History of alcohol use Iron hemosiderin NASH
0.86 (0.76–0.99) 0.59 (0.25–1.40) 0.32 (0.92–1.13) 1.17 (1.09–1.26) 0.94 (0.36–2.43) 0.27 (0.17–0.95) 0.76 (0.20–2.76) 0.68 (0.88–2.46)
0.034 0.238 0.078 <.001 0.902 0.042 0.677 0.560
Metabolic syndrome Platelet count Prothrombin time Steatosis Tobacco Total cholesterol (mg/dL)
1.00 (1.01–1.10) 1.05 (1.01–1.08) 0.33 (0.12–0.94) 0.56 (0.22–1.38) 0.51 (0.14–1.83)
<.001 0.001 0.038 0.209 0.303
OR (95% CI)
p-Value
3.86 (1.03–14.4)
0.044
0.46 (0.23–0.94)
0.034
1.21 (1.09–1.34)
0.003
1.00 (1.00–1.04) 1.05 (1.01–1.10)
0.006 0.016
ALT, alanine aminotransferase; AST, aspartate aminotransferase; Anti-HBc, anti-hepatitis B core antigen; Anti-HBs, anti-hepatitis B surface antigen; BMI, body mass index; NASH, non alcoholic steato-hepatitis; OR, odds ratio; CI, confidence interval. a Multivariate included age and gender plus all variables with p ≤ 0.02. The influence of age is already known and AST was not included because ALT was also included. If AST was included willingly, the result did not change.
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B. K-Kutala et al. / Digestive and Liver Disease xxx (2015) xxx–xxx Table 4 Multivariate binary logistic regression evaluating baseline factors associated with hepatocellular carcinoma in 324 matched patients infected with chronic hepatitis C. Multivariate OR
(95% CI)
p-Value
Non-fibrotic (F0-F2) ALT AST Infection duration
0.14 2.14 1.68
(0.20–2.32) (0.29–23.9) (1.08–2.69)
0.178 0.464 0.021
Fibrotic (F3-F4) AST Diabetes Gender History of alcohol use Platelet account Prothrombin time
1.16 3.13 2.36 3.14 1.00 0.95
(0.91–1.49) (1.17–8.38) (0.90–6.15) (1.29–7.62) (1.00–1.04) (0.93–0.98)
0.222 0.022 0.077 0.011 0.013 0.001
ALT, alanine aminotransferase; AST, aspartate aminotransferase; OR, odds ratio; CI, confidence interval. Multivariate including all significant baseline variables with p ≤ 0.02 in univariate analysis.
NAFLD and NASH can progress to cirrhosis and liver failure in 3–15% of patients and lead to HCC [25]. The mechanisms of the pathogenesis of NASH in the development of HCC in patients with CHC infection have not been clarified. Moreover diabetes and obesity have been shown increase the risk of HCC [26]. Despite a lack of strong evidence, this study examines the role of the metabolic syndrome, diabetes and as well as the effect of HCV on the relationship between these risk factors and HCC. However, only 12% and 20% patients had diabetes and metabolic syndrome without visceral obesity in the F0-F2 group.
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Although it has been suggested that the association of NASH and obesity increases the risk of HCC in chronic hepatitis C [27], this association was not found in our series. The most important finding in this study was the association of baseline diabetes and excessive alcohol intake with an increased occurrence of HCC in patients with advanced fibrosis. Although a significant impact of diabetes on overall liver-related outcome may be overlooked, the association of diabetes with an accelerated progression to cirrhosis and HCC in patients with CHC is well known [28] (Tables 4 and 5). Moreover, multivariate analysis revealed that the risk of HCC in patients without advanced fibrosis increased in anti-HBc-positive patients (OR: 3.86) and with increased duration of HCV infection (OR: 1.21). The significantly higher percentage of HBV marker in patients F0-F2 than in F3-F4 (24% vs. 56%, p = 0.001) suggests a possible role of occult HBV infection in the former group. Occult HBV infection has been defined by the European Association for the Study of the Liver (EASL) [29] and the Taormina Consensus Conference in 2008 as the “presence of HBV DNA in the liver of individuals testing HBsAg-negative with currently available assays” and introduced a cut-off value for serum HBV DNA (<200 IU/mL). The relationship between occult HBV infection and serological markers of HBV infection has been extensively studied [30]. There is also significant evidence suggesting that occult HBV is a risk factor for the development of HCC in both patients with HCV-related or not cryptogenic chronic liver disease [31–33]. Moreover Pollicino et al. have shown that many of the potential mechanisms of tumour genesis in overt HBV are maintained in case of occult HBV infection [34]. However, those were cross-sectional case studies examining patients with existing HCC. To our knowledge there are no existing prospective studies investigating the development
Table 5 Baseline characteristics of 324 age-matched chronic hepatitis C patients, with or without hepatocellular carcinoma. All n = 324
Controls n = 162
Cases n = 162
Clinical parameters Male gender Median age (years) Duration of infection (years)
220 (68%) 63 [54–71] 26 [22–32]
102 (63%) 61 [53–69] 26 [20–32]
118 (73%) 63 [55–72] 27 [23–32]
0.056 0.087 0.138
Ethnicity Caucasian
209 (65%)
107 (66%)
102 (63%)
0.561
Comorbidities BMI (median) Tobacco Diabetes Arterial hypertension History of alcohol use
25 [22–27] 125 (39%) 64 (20%) 82 (25%) 72 (22%)
Laboratory data Abnormal total cholesterol Anti-HBc+ Anti-HBs+ Alpha-fetoprotein (ng/ml) ALT (×ULN) AST (×ULN) Genotype 1 HCV-RNA (Log) Ferritin (mg/dL) Platelet count (×109 /L) Prothrombin time (%)
50 (15%) 84 (26%) 63 (19%) 6 [3–13] 2.0 [1.4–3.0] 1.8 [1.1–2.6] 212 (65%) 5.1 [5–6.3] 247 [125–402] 174 [123–219] 90 [79–100]
18 (11%) 38 (23%) 26 (16%) 5 [3–8] 2.0 [1.4–3.1] 1.6 [1.1–2.4] 105 (65%) 5.4 [5–6.4] 241 [122–390] 117 [95–178] 92 [83–100]
32 (20%) 46 (28%) 37 (23%) 16 [8–105] 1.8 [1.2–2.8] 2.2 [1.6–2.9] 107 (66%) 5.0 [4.6–5.5] 269 [134–457] 186 [139–228] 77 [64–90]
Other parameters F0-F2 F3-F4 Steatosis
50 (15%) 247 (85%) 114 (35%)
25 (15%) 137 (85%) 50 (31%)
25 (15%) 137 (85%) 64% (39%)
24 [22–27] 54 (33%) 20 (12%) 35 (22%) 23 (14%)
25 [23–27] 71 (44%) 44 (27%) 47 (29%) 49 (30%)
p-Value
0.143 0.052 0.008 0.125 0.005 0.031 0.310 0.122 <0.001 0.031 <.001 0.815 0.041 0.122 <0.001 <0.001
0.101
Abbreviations: ALT, alanine aminotransferase; AST, aspartate aminotransferase; Anti-HBc+, anti-hepatitis B core antigen positive; Anti-HBs+, anti-hepatitis B surface antigen positive; IQR, interquartile range; ULN, upper limit of normal; BMI, body mass index, calculated as weight in kilograms divided by height in metres squared. Data are presented as No./Total No. (%) or median [interquartile range], unless otherwise noted. See “Methods”. Baseline parameters were compared between patients with and without advanced fibrosis using signed rank Wilcoxon test for continuous variables and the paired t test for categorical variables. Case = with HCC, Control = without CHC matched by the age.
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of HCC in patients with CHC in relation to their occult HBV status. Squadrito et al. [32] have also shown that occult HBV may be associated with the development of HCC in chronic liver disease and confirmed that, in HBsAg-negative individuals, those with the HBV genomic sequences have a higher risk of developing HCC. Additionally, the HCV virus contains some hydrophobic and hydrophilic proteins, such as proteins NS3, NS4B and NS5A that have a regulatory effect on cell promoters and seem to interact with some of the cellular proteins involved in the mechanism of carcinogenesis under certain specific conditions [35]. Our findings show that the duration of HCV infection (OR: 1.21, p = 0.003) is an independent factor and may influence development of HCC in CHC patients without advanced fibrosis. This factor is well recognised (at least for HCV-induced cirrhosis) as strongly correlated to HCC development. However, most studies on HCC risk factors in cirrhosis or advanced fibrosis did not adequately take into account duration of infection as a confounding factor. This duration of infection in patients without advanced fibrosis may suggest that a few years of inflammation can lead to HCC. More data are needed to clarify this issue. Other baseline factors significantly associated with HCC occurrence in patients without advanced fibrosis at multivariate analysis were ALT, platelet count, and prothrombin level. Those factors are already known to have an impact on disease progression and they are surrogate markers of inflammatory activity of the liver, and portal hypertension. The complementary analysis, including matched patients, revealed implications of some variables known as risk factors of HCC or progression of liver disease – such as, alcohol intake, diabetes and platelet count. Although optimal screening protocols and monitoring tools are still a subject of debate, the particularly poor survival of HCV patients without advanced fibrosis in our series (35% at 5 years, Log-rank, p = 0.038) is alarming and underscores the importance of surveillance in the HCV-infected population without advanced fibrosis. Of note, in our study 48% of patients without advanced fibrosis had a tumour ≥3 cm vs. 31% of patients in the advanced fibrosis group. The effectiveness of screening programmes will depend on external factors, such as physician and patient acceptance and adherence to the organization’s programme, economic considerations, and a host of other factors, including availability and accessibility to diagnostic tools, treatment and adequate follow-up [36]. Although confounding factors were taken into account, this study has several limitations. Firstly, the study population was small, which might induce a selection bias and can have an impact on the interpretation of our findings. However compelling our findings may be, the absence of RT-PCR on liver tissue samples limits the power of this study. This study suggests that anti-HBc, diabetes and alcohol intake remain the main risk factors for HCC in patients with advanced fibrosis. These results could modify the clinical management of these patients. To understand the precise mechanisms of the development of HCC in CHC without advanced fibrosis, the pathogenic role of each risk factor and the epidemiology of occult HBV infection must be more clearly defined. Thus, testing patients with CHC for HBV genomes could be a powerful tool to identify subjects who require careful screening to obtain an early diagnosis of HCC. Conflict of interest None declared. References [1] El-Serag HB. Hepatocellular carcinoma. New England Journal of Medicine 2011;365:1118–27.
[2] El-Serag HB, Rudolph KL. Hepatocellular carcinoma: epidemiology and molecular carcinogenesis. Gastroenterology 2007;132:2557–76. [3] Sherman M. Epidemiology of hepatocellular carcinoma. Oncology 2010;78(Suppl. 1):7–10. [4] Beasley RP, Hwang LY, Lin CC, et al. Hepatocellular carcinoma and hepatitis B virus. A prospective study of 22 707 men in Taiwan. Lancet 1981;2: 1129–33. [5] Buendia MA. Hepatitis B viruses and hepatocellular carcinoma. Advances in Cancer Research 1992;59:167–226. [6] Fattovich G, Stroffolini T, Zagni I, et al. Hepatocellular carcinoma in cirrhosis: incidence and risk factors. Gastroenterology 2004;127:S35–50. [7] Gomaa AI, Khan SA, Toledano MB, et al. Hepatocellular carcinoma: epidemiology, risk factors and pathogenesis. World Journal of Gastroenterology: WJG 2008;14:4300–8. [8] Yoshida H, Shiratori Y, Moriyama M, et al. Interferon therapy reduces the risk for hepatocellular carcinoma: national surveillance program of cirrhotic and noncirrhotic patients with chronic hepatitis C in Japan. IHIT Study Group. Inhibition of Hepatocarcinogenesis by Interferon Therapy. Annals of Internal Medicine 1999;131:174–81. [9] Paradis V, Zalinski S, Chelbi E, et al. Hepatocellular carcinomas in patients with metabolic syndrome often develop without significant liver fibrosis: a pathological analysis. Hepatology 2009;49:851–9. [10] Davila JA, Henderson L, Kramer JR, et al. Utilization of surveillance for hepatocellular carcinoma among hepatitis C virus-infected veterans in the United States. Annals of Internal Medicine 2011;154:85–93. [11] Kleiner DE, Brunt EM, Van Natta M, et al. Design and validation of a histological scoring system for nonalcoholic fatty liver disease. Hepatology 2005;41:1313–21. [12] Bedossa P, Poynard T. An algorithm for the grading of activity in chronic hepatitis C. The METAVIR Cooperative Study Group. Hepatology 1996;24: 289–93. [13] Clouston AD, Jonsson JR, Purdie DM, et al. Steatosis and chronic hepatitis C: analysis of fibrosis and stellate cell activation. Journal of Hepatology 2001;34:314–20. [14] Asselah T, Boyer N, Guimont MC, et al. Liver fibrosis is not associated with steatosis but with necroinflammation in French patients with chronic hepatitis C. Gut 2003;52:1638–43. [15] Marcellin P, Asselah T, Boyer N. Fibrosis and disease progression in hepatitis C. Hepatology 2002;36:S47–56. [16] Cardoso AC, Moucari R, Figueiredo-Mendes C, et al. Impact of peginterferon and ribavirin therapy on hepatocellular carcinoma: incidence and survival in hepatitis C patients with advanced fibrosis. Journal of Hepatology 2010;52: 652–7. [17] Zimmet P, Magliano D, Matsuzawa Y, et al. The metabolic syndrome: a global public health problem and a new definition. Journal of Atherosclerosis and Thrombosis 2005;12:295–300. [18] Alberti KG, Zimmet P, Shaw J, Group IDFETFC. The metabolic syndrome – a new worldwide definition. Lancet 2005;366:1059–62. [19] Nash KL, Woodall T, Brown AS, et al. Hepatocellular carcinoma in patients with chronic hepatitis C virus infection without cirrhosis. World Journal of Gastroenterology: WJG 2010;16:4061–5. [20] Yeh MM, Daniel HD, Torbenson M. Hepatitis C-associated hepatocellular carcinomas in non-cirrhotic livers. Modern Pathology 2010;23:276–83. [21] Lok AS, Seeff LB, Morgan TR, et al. Incidence of hepatocellular carcinoma and associated risk factors in hepatitis C-related advanced liver disease. Gastroenterology 2009;136:138–48. [22] Bugianesi E, Leone N, Vanni E, et al. Expanding the natural history of nonalcoholic steatohepatitis: from cryptogenic cirrhosis to hepatocellular carcinoma. Gastroenterology 2002;123:134–40. [23] Brunt EM. Nonalcoholic steatohepatitis. Seminars in Liver Disease 2004;24:3–20. [24] Leandro G, Mangia A, Hui J, et al. Relationship between steatosis, inflammation, and fibrosis in chronic hepatitis C: a meta-analysis of individual patient data. Gastroenterology 2006;130:1636–42. [25] Angulo P. Nonalcoholic fatty liver disease. New England Journal of Medicine 2002;346:1221–31. [26] Welzel TM, Graubard BI, Zeuzem S, El-Serag HB, et al. Metabolic syndrome increases the risk of primary liver cancer in the United States: a study in the SEER-Medicare database. Hepatology 2011;54:463–71. [27] El-Serag HB, Mason AC. Risk factors for the rising rates of primary liver cancer in the United States. Archives of Internal Medicine 2000;160: 3227–30. [28] El-Serag HB. Epidemiology of viral hepatitis and hepatocellular carcinoma. Gastroenterology 2012;142:1264–73, e1. [29] Raimondo G, Navarra G, Mondello S, et al. Occult hepatitis B virus in liver tissue of individuals without hepatic disease. Journal of Hepatology 2008;48: 743–6. [30] Marcellin P, Thiers V, Degott C, et al. Hepatocellular carcinoma with normal adjacent liver. Hepatitis B virus DNA status. Journal of Hepatology 1989;8:249–53. [31] Cacciola I, Pollicino T, Squadrito G, Cerenzia G, Orlando ME, Raimondo G. Occult hepatitis B virus infection in patients with chronic hepatitis C liver disease. New England Journal of Medicine 1999;341:22–6. [32] Squadrito G, Pollicino T, Cacciola I, et al. Occult hepatitis B virus infection is associated with the development of hepatocellular carcinoma in chronic hepatitis C patients. Cancer 2006;106:1326–30.
Please cite this article in press as: K-Kutala B, et al. Patients with chronic hepatitis C without advanced fibrosis and hepatocellular carcinoma: A retrospective clinical–pathological study. Dig Liver Dis (2015), http://dx.doi.org/10.1016/j.dld.2014.12.010
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ARTICLE IN PRESS B. K-Kutala et al. / Digestive and Liver Disease xxx (2015) xxx–xxx
[33] Squadrito G, Cacciola I, Aldibrandi A, et al. Impact of occult hepatitis B virus infection on the outcome of chronic hepatitis C. Journal of Hepatology 2013;59:696–700. [34] Pollicino T, Squadrito G, Cerenzia G, et al. Hepatitis B virus maintains its prooncogenic properties in the case of occult HBV infection. Gastroenterology 2004;126:102–10.
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[35] Zemel R, Gerechet S, Greif H, et al. Cell transformation induced by hepatitis C virus NS3 serine protease. Journal of Viral Hepatitis 2001;8: 96–102. [36] Andersson KL, Salomon JA, Goldie SJ, Chung RT. Cost effectiveness of alternative surveillance strategies for hepatocellular carcinoma in patients with cirrhosis. Clinical Gastroenterology and Hepatology 2008;6:1418–24.
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