**3. Proteantigen**

#### **3.1 Glypican-3 (GPC3)**

Glypican-3 (GPC3) is a member of the glycican family of heparan sulfate proteoglycans linked to cell membranes by glycosyl-phosphatidylinositol [17]. It is a fetal glycoprotein that exists on the cell surface to help regulate cell growth during pregnancy. GPC3 is associated with the malignant proliferation of cells but there are currently no studies to prove its association with healthy people and benign conditions. Quite a number of studies have proven the overexpression of GPC3 in malignant diseases such as breast cancer, ovarian cancer, or lung adenocarcinoma [18, 19]. With HCC, its expression is increased through the autocrine/paracrine regulator in conjunction with the Wnt signaling pathway [20]. Some studies have concluded that the sensitivity of GPC3 in HCC diagnosis ranges from 40 to 53%, which is interesting considering that in about 33% of cases, both AFP and DCP serum were within normal limits [21, 22]. GPC3 has been detected in HCC tumor but not in benign liver tissues, so it is likely a marker for early detection of HCC [23]. GPC3 expression does not depend on some clinical features such as tumor size, GPC3 sensitivity in early HCC diagnosis (size <3 cm) was 56% [23]. In a meta-analysis, the sensitivity and specificity of serum GPC3 to diagnose HCC were 55.2% and 84.2%, respectively [24]. A smaller analysis of the early-stage HCC group (BCLC 0 and A or TNM phase I) showed a sensitivity and specificity of GPC3 of 55.1% and 97%, respectively, which are higher than the those obtained with the AFP serum in

the same study, that were 34.7% and 87.6%. Combining GPC3 and AFP increased the sensitivity to 76% for early-stage tumors [24]. In short, GPC3 might be a marker for HCC, especially in the early stages, but GPC3 expression also increases in some other malignancies, so the specificity for HCC diagnosis is not high. It can still increase diagnostic sensitivity when combined with other valuable serum markers (**Table 1**).

#### **3.2 Heat shock protein 70 (HSP70)**

Heat shock protein (HSP) is an antiapoptotic protein whose overexpression allows cell survival. It protects cells and stimulates the reparation of tissue damage. A study indicated the positive rate of HSP70 and HSP27 in HCC tissues at 56.3% and 61.9%, respectively [25]. There was a correlation between the stained intensity of HSP70 and tumor size, portal vein invasion, and tumor stage, while HSP27 was only associated with hepatitis B virus (HBV) related HCC. In addition, the overexpression of HSP70 and HSP27 in HCC tumors may lead to increased tumor growth and metastasis (**Table 1**) [26].

Data suggest that HSP70 can be used as a prognosis indicator for HCC. Its expression was detected in 282 of 392 HCC cases (71.9%), compared to 14 of 115 non-neoplastic liver tissues [27]. The sensitivity and specificity in the detection of HCC have been measured at 57.5% and 85%, respectively [28]. The expression of HSP70 is also correlated with the differentiation and apoptosis of tumor cells. HSP70 promotes cancer cell growth by stabilizing cyclin D1 and suppressing apoptosis in cancer cells by inhibiting the p53 pathway [29, 30]. This information makes HSP70 and HSP27 potential markers of HCC that should be further investigated.

#### **3.3 Golgi protein 73 (GP73)**

Golgi protein 73 (GP73) is a type II Golgi-specific membrane protein, which is normally expressed in epithelial cells of many human tissue types, but not hepatocytes [31]. A study showed that serum GP73 levels of patients with HBV-related HCC were significantly increased compared to patients with HBV and healthy adults [32, 33]. The sensitivity of diagnosis of HCC (76.9%) was significantly higher than that of AFP (48.6%), suggesting that GP73 can be an effective serum biomarker for the diagnosis of HCC [34]. The combination of GP73 and AFP further increased the sensitivity and specificity to 89.2% and 85.2%, respectively, with an AUC of 0.96 (**Table 1**).

FC-GP73 further improves the HCC diagnostic performance made with GP73 from 65 to 90 to 90–100%, respectively. Even when GP73 is at a very low level or absent, FC-GP73 is still detectable [35]. These are encouraging data but there is still a lot of work to be done regarding the correlation between GP73 and tumor size, stage, recurrence, and prognosis before this marker can be used.

#### **3.4 Squamous cell carcinoma antigen (SCCA)**

Squamous cell carcinoma antigen (SCCA) belongs to the high molecular weight protease inhibitor family found in the squamous and granular layers of the normal squamous epithelium. It consists of two different isomers, encoded by two highly homologous genes: SCCA1 being neutral, and SCCA2 acid [36]. SCCA2 has been detected in many malignancies such as cervical, lung, head and neck carcinoma, and it has been used as a valuable diagnostic biomarker in clinical practice [37].

Giannelli et al. showed that SCCA expression was higher in the HCC group than in the cirrhotic group. The sensitivity of SCCA is 84.2%, but the specificity is low at

#### *Circulating Biomarkers for Early Diagnosis of Hepatocellular Carcinoma DOI: http://dx.doi.org/10.5772/intechopen.98483*

48.9%. In the small tumor group (≤ 3 cm) the sensitivity and specificity of SCCA were 56.1% and 74.9% with a cut-off of 3.2 ng/ml. In their study of SCCA expression in cells, using immunohistochemistry, Guido et al. demonstrated that SCCA expression in cancerous tissues and dysplasia nodules was much higher than that of newly formed nodules in early HCC diagnosis [38]. SCCA was highly sensitive, but its specificity was quite low. Its expression in early HCC tissue and in dysplasia nodules makes SCCA a valuable complementary marker for HCC diagnosis. An alternative biomarker is an immune complex between SCCA and IgM, SCCA-IgM, whose expression increases in early HCC. The immune complex SCCA-IgM has a higher diagnostic performance than the free SCCA and is also more relevant since it is not found in the serum of healthy people. However, the detection rate of SCCA-IgM immune complex is 18% for chronic hepatitis, 26% for cirrhosis and 70% for HCC [39]. Its sensitivity and specificity for HCC diagnosis are 89% and 50% [40]. The concentration of SCCA-IgM immune complex is constantly increasing in patients with cirrhosis who tend to progress to HCC. Sensitivity and specificity were of higher value than AFP in the studies of Pontisso et al. [37].

Increased serum SCCA in patients with liver disease can be considered a valuable marker for early diagnosis of HCC. Especially the SCCA-IgM immune complex, which is highly sensitive. However, since its specificity is quite low, it must be combined with other markers such as serum AFP or DCP to increase its diagnostic value.
