Assessment of Cancer Antigen 125 in Post-HCV Chronic Liver Disease and Hepatocellular Carcinoma Patients

Short title: serum CA125 and CLD patients with or without HCC. Background and study aim: Most of the morbidity and mortality of chronic liver diseases is due to its progression and complication of cirrhosis as ascites. The cancer antigen (CA) -125 is a high molecular mass glycoprotein produced both by ovarian cancer cells as well as by normal cells derived from coelomic epithelium. This study was conducted to assess the serum and/or ascitic fluid level of CA125 in HCV cirrhotic patients (decompensated or not) with or without Hepatocellular Carcinoma (HCC).


INTRODUCTION
Chronic liver diseases and their complications constitute a major health problem all over the world and especially in our country [1].Most of the morbidity and mortality of chronic liver diseases is due to its progression and complications of cirrhosis [2].Ascites is a frequent complication of advanced liver cirrhosis.Over 50% of cirrhotic patients develop ascites due to increased sodium retention in the kidneys, leading to expansion of extracellular volume and accumulation of fluid in the peritoneum [3].The cancer antigen (CA) -125 is a high molecular mass glycoprotein produced both by ovarian cancer epithelial cells and mesothelial cells [4].
Serum CA-125 levels are used as a marker of tumour activity in patients known to have ovarian carcinoma [5], the severity of liver disease evaluated by MELD score and the presence of ascites were significantly correlated with the elevation of CA-125 level [6].Serum CA-125 levels are generally found to be higher in malignant conditions compared to benign conditions.Levels exceeding 1000 U/ml have been described in benign conditions associated with massive pleural effusion and ascites [7].
Hepatocellular carcinoma (HCC) is a global problem worldwide.Egypt is the sixth country in the Middle East and Arab world and the third one in Africa, with a high prevalence of HCC, the major risk factor for HCC is HCV infection which mainly burden As no enough Egyptian studies were done on the validity of CA-125 and chronic liver diseases or HCC patients, so we aimed to assess the serum and/or ascitic fluid level of CA125 in post HCV cirrhotic patients (decompensated or not) with or without HCC.

Study design: A cross sectional observational study
Patients: patients were recruited from those attending Tropical Medicine Department-Ain Shams University and from Professor Doctor Yassin Abdel Gaffar Charity Center for Liver Disease and Research.The study included number of (120) cases and divided into two groups: Group I; Formed of 60 post HCV liver cirrhosis cases; (A) 30 cases without ascites, (B) 30 cases with ascites.Group II; Formed of 60 HCC on top of post HCV liver cirrhosis cases; (C) 30 cases without ascites, (D) 30 cases with ascites.Inclusion Criteria: Male Patients, Clinical, laboratory and ultrasonographic criteria suggestive of chronic liver disease post HCV liver cirrhosis, HCC cases were diagnosed by; High AFP > cut-off limit (>200ng), Characteristic features of HCC by triphasic spiral abdominal CT [12].Exclusion Criteria: Patients with other causes of liver disease, extrahepatic metastasis or other simultaneous malignancies, Patients with advanced systemic disease and other causes of ascites or serous sac effusion.
Serum and ascetic fluid level of CA-125 were assessed and measured: Specimen collection and preparation: 5ml of venous blood as well as an ascitic sample were taken under complete aseptic conditions.Instrument: The Roche diagnostics cobas e 411 analyzer was used.Steps of test: Sandwich principle, total duration of assay: 18 minutes.Normal range of CA125 serum level: up to 21 u/ml, Ascitic fluid level: up to 35 u/ml [13].

Statistical analysis
IBM SPSS statistics (V.19.0,IBM Corp., USA, 2010) was used for data analysis.Data were expressed as Mean ± SD for quantitative parametric measures in addition to Median Percentiles for quantitative non-parametric measures and both number and percentage for categorized data.
The following tests were done: (1) Comparison between two independent groups for non-parametric data using Wilcoxon Rank Sum test.(2) Ranked Sperman correlation test to study the possible association between each two variables among each group for non-parameteric data.(3) Chisquare test to study the association between each 2 variables or comparison between 2 independent groups as regards the categorized data.The probability of error at 0.05 was considered sig., while at 0.01 was considered highly sig.(4) The ROC was constructed to evaluate the most discriminating markers between the compared groups, AUC can also be calculated.

RESULTS
In order to fulfill our aim, (120) cases were included and divided into two groups; Group I: included 60 post HCV liver cirrhoses cases; (A) (30) patients with compensated cirrhoses.(B) (30) patients with decompensated cirrhosis.Group II: included 60 HCC of post HCV liver cirrhosis cases; (C) 30 cases with compensated cirrhoses, (D) 30 cases with decompensated cirrhosis.
Concerning serum level of CA-125; there was a significant difference between groups (A) & (B) being higher in group (B) (P<0.01), also a significant difference between groups (C) & (D) being higher in group (D) (P<0.01) as in Table (1).
In Table (2) and (3): There were a positive correlation between serum level of CA-125 and AFP in group (A) and (C) (P>0.05).Regarding Group (B) and (D), there was a positive correlation between serum level of CA-125 and each of AFP level (P>0.05) and ascitic level of CA-125 (P<0.01).Table (4) shows that there was a significant difference between groups B (decompensated non HCC) & D (decompensated HCC) concerning ascitic level of Ca 125 being higher in group D (P<0.01).Regarding the serum level of CA125, there was a significant elevation in both compensated group (A) and decompensated group (B) post HCV cirrhotic cases but more higher in decompensated group (B), this in agreement with each of; Piekarska et al. [14], who stated that the mean serum levels of CA-125 in patients with cirrhosis (with and without ascites) were elevated above the normal level, Xiao and Liu [15] who found the elevation of CA-125 in sera of cirrhotic patients without ascites and Assmar et al. [16] who reported that CA-125 was not a tissue or tumor specific antigen, elevated level could be detected in patient without malignant transformation and indicate presence of cirrhosis in benign liver disease cases.

Regarding the diagnostic performance of AFP and serum CA125 in differentiating compensated HCC group (C) from non-HCC group (A), as in
There was a significant positive correlation between serum level and ascetic level of CA125 in both non HCC (B) and HCC (D) decompensated groups and this was in a harmony with Hussain and Camilleri [17] who suggested that simultaneous assay of CA-125 in serum and body fluid and/or with a panel of other tumor marker could be accurate in diagnosing malignant and non malignant cases, also significant elevation of CA-125 could occur in HCC cases.Also Tuzun et al. [18] reported a positive correlation between serum and ascitic fluid levels of CA-125, however ascitic fluid levels were higher than serum levels, which explained that the antigen is mesothelial rather than tumoral origin.However our results reported that the ascitic level of Ca 125 was elevated in decompensated HCC group (D) than non HCC group (B), this supported that the antigen origin may be from both mesothelial and tumoral cells.
Regarding the correlation between AFP and CA125 serum and ascitic level; Only AFP had significant diagnostic performance in differentiating compensated and decompensated HCC groups (C) & (D) from non-HCC groups (A) &(B).However Lopez et al. [19] who stated that AFP-CA-125 combination had the highest sensitivity to diagnose 96% of the HCC patients, also both could be combined for HCC screening due to their excellent sensitivity and specificity, respectively: a negative result for both, or even CA 125 alone, would discourage HCC diagnosis, while positive results of both would make HCC presence highly probable.A positive CA 125 and negative AFP would be equivocal for HCC.Also Elias and Kew [20] reported that CA 125 has a highly sensitivity but lacks specificity for HCC diagnosis.
Fig. (1); Only AFP had significant diagnostic performance (P<0.01) in differentiating compensated HCC group (C) from non-HCC group (A).However, diagnostic performance of AFP, serum CA125 and ascetic fluid CA125 in differentiating decompensated HCC (D) group from non-HCC (B) group as in Figure (2); Only AFP had significant diagnostic performance in differentiating decompensated HCC (D) group from non-HCC (B).

Table (
2): Correlation between CA-125 serum level and other parameters in group A and B

Table ( 3
): Correlation between CA-125 serum levels and other parameters in group C and D

Table ( 4
): Comparison between group B and group D concerning ascitic levels of CA-125