In HCC, an elevated plasma level of OPN is regarded as a potential prognostic biomarker and overexpression of OPN is closely correlated with intrahepatic metastasis, early recurrence, and a worse prognosis [14]. ANXA2 is an inducible, calcium-dependent phospholipid-binding protein that is overexpressed in a variety of human malignancies [13]. So the purpose of this study was to evaluate the role of ANXA2 and OPN for early diagnosis of HCC in hepatitis C virus patients.
In our study there were statistically significant differences in AST, GGT, serum bilirubin (total and direct), international normalized ratio, and serum albumin levels between the HCC group and the chronic hepatitis C group, all these parameters were higher in the HCC group than the chronic hepatitis C group which is in agreement with the results by Fuoad et al. who explained this difference by the progression of the underlying liver cirrhosis caused by HCC with a subsequent decreased albumin and protein synthesis and poor utilization of vitamin K in advanced parenchymal liver disease [15].
Our results showed that serum AFP was found to be significantly higher in the HCC group in comparison to the chronic hepatitis C group and the cutoff point for predicting the probability for HCC was 6.0 (ng/ml) with sensitivity of 77.50%, specificity of 82.50%, positive predictive value of 81.60%, negative predictive value of 78.6%, and accuracy of 80%. These results are comparable with those of Marrero et al. who performed a large case-control study involving 836 patients. There was a significant difference between the early HCC and the cirrhotic patient group as regards AFP [16].
Also, these results were in agreement with that of El-Tayeh et al. who explained his results by an increase in the selective transcriptional activation of the AFP gene in malignant hepatocytes, which resulted in the increased secretion of AFP during the development of HCC [17]. On the other hand, these results are incompatible with El-Gezawy et al. who postulated that there was a similarity and no significant difference between the early HCC and cirrhotic groups as regards AFP [18].
In the present study, OPN level was found to be significantly higher in HCC patients than chronic hepatitis C patients. These results are compatible with the study performed by Shang et al. who reported that OPN was significantly higher in early HCC patients than cirrhotic patients [19].
In the HCC group, there was significant positive correlation between OPN level and serum bilirubin (total and direct); these results suggested that the OPN was correlated with the progression of liver disease. However, no significant correlation was noted between OPN level and other parameters. The correlation coefficient between serum OPN and AFP values was not significant. Hodeib et al. reported that OPN levels were significantly correlated with AFP levels but no significant correlation between OPN level and other parameters in HCC patients [20].
The ROC curve for OPN for detection of HCC was significant, the cutoff point was 13.2 (ng/ml) with sensitivity of 65.0% and specificity of 90.0%. These results are compatible with the study performed by Lee et al. who reported that the accuracy achieved by using plasma OPN levels for diagnosis of HCC was inferior to the accuracy achieved using AFP. At a cutoff value of 6 (ng/ml), plasma AFP showed high sensitivity (63.9%) and specificity (95%). Plasma OPN at a cutoff value of 557 (ng/ml) showed a high specificity (92.5%) but a lower sensitivity (26.1%) [21].
These results are incompatible with the study performed by Shang et al. who reported that OPN at higher threshold of 91 (ng/ml), its diagnostic performance higher than AFP (0.739, 0.680, respectively) for discriminating between early HCC and cirrhosis. OPN demonstrated 75% sensitivity and 62% specificity for early stage HCC, compared to 46% sensitivity and 93% specificity for AFP [19]. The exact reason for these differences as regards cutoffs is not clear, but these discrepancies may be in consequence of the different assay systems and conditions of sample collection used in different studies.
The binding of secreted OPN from HCV-infected cells to integrinαvβ3 and CD44 leads to elevation of reactive oxygen species and activation of Ca2+ signaling and downstream cellular kinases; all of which promote epithelial-mesenchymal transition, cell migration, and invasion to enhance tumor progression and metastasis in HCC [22]. The role of OPN in metastasis is more prominent because OPN expression facilitates recurrence and reduces patient survival after liver transplantation for HCC. Thus, OPN may be a useful marker for detecting early recurrence of HCC after surgery [23].
In the present study, ANXA2 level was found to be significantly higher in early HCC patients than chronic hepatitis C patients. These results are compatible with El-Gezawy et al. who pastulated that ANXA2 was significantly higher in early HCC patients than cirrhotic patients [18]. Shaker et al. also reported that ANXA2 was significantly higher in early HCC patients than chronic liver disease (CLD) patients [24].
Interestingly, in the HCC group, there was significant positive correlation between ANXA2 level and alkaline phosphatase and there was significant relation as regards fatigue, abdominal pain, and vomiting. However, no significant correlation was noted between annexin level and other parameters. There was no significant correlation between ANXA2 level and AFP; this agrees with the studies done by El-Gezawy et al. [18], Shaker MK et al. [24], and Sun et al. [13]. The correlation coefficient between serum ANXA2 and AFP values was not significant, indicating that measuring both markers (AFP and ANXA2) in serum can improve the diagnostic value.
The ROC curve for ANXA2 for detection of HCC was significant, the cutoff point was 10.1 (ng/ml) with sensitivity of 85.0% and specificity of 85.0%. These results are compatible with El-Gezawy et al. who reported for early stage HCC, ANXA2 (optimal cutoff of 24.99 IU/ml), higher sensitivity, and specificity (79.34% and 85.56% respectively) than those of AFP (optimal cutoff of 5.96 IU/ml) (67.78% and 59.85% respectively) [18]. The ANXA2 mRNA expression level was obviously, significantly higher and over expressed in HCC tissues rather than in the other patient groups. One explanation showed that ANXA2 synthesis is induced in transformed hepatocytes [25].
These results are also, more or less, compatible with the study performed by Shaker et al. who reported that ANXA2 was significantly higher in HCC patients than chronic liver disease patients, the cutoff point for predicting the probability for early HCC was 18 ng/mL, the diagnostic sensitivity was 74%, the specificity was 88%, the PPV was 92.5%, the NPV was 62.9%, and the efficacy was 78.7% which is higher than AFP (cutoff value was 19.8 ng/ml) as regards diagnostic sensitivity (70%), but similar to AFP as regards specificity, positive and negative predictive values, and efficacy [24].
Shaker et al. stated that there was a significant difference observed between patients with CLD and healthy people with respect to AFP, who declared that one of the limitations in the use of AFP for the diagnosis of HCC is its increase in patients who have hepatitis and CLD but who do not have HCC, but found that ANXA2 levels were highly and significantly increased in patients with HCC compared with the levels in patients with CLD and in controls; however, no statistical significance was found between patients with CLD and the healthy people with respect to ANXA2 expression [24].
This was explained by Zhang et al. who stated that the ANXA2 gene is upregulated in HCV-associated HCC [26]. In addition, Mohammad et al. stated that ANXA2 is rarely detected in either normal or chronic hepatic tissues but is over expressed at both the mRNA and protein levels HCC [27].
Wang et al. stated that, one of the possible mechanisms explaining the relationship between ANXA2 and HCC is promotion of HCC cell migration and invasion by ANXA2 pseudogene [28]. A recent study done by Lou et al. who reported that ANXA2 binds with Lung cancer associated transcript 1 (LUCAT1), which plays a key role in tumorigenesis, progression of HCC and a better therapeutic target for HCC patients. LUCAT1 inhibits the phosphorylation of ANXA2 and increase the secretion of plasminogen into plasmin [29].