Human hepatocellular carcinomas with “Stemness”-related marker expression: keratin 19 expression and a poor prognosis

Abstract

There is a recently proposed subtype of hepatocellular carcinoma (HCC) that is histologically similar to usual HCC, but characterized by the expression of ''stemness''-related markers. A large-scale study on two different cohorts of HCCs was performed to investigate the clinicopathologic features and epithelial-mesenchymal transition (EMT)-related protein expression status of this subtype of HCCs. The expression status of stemnessrelated (e.g., keratin 19 [K19], cluster of differentiation [CD]133, epithelial cell adhesion molecule [EpCAM], and c-kit) and EMT-related markers (e.g., snail, S100A4, urokinase plasminogen activator receptor [uPAR], ezrin, vimentin, E-cadherin, and matrix metalloproteinase [MMP]2) were examined using tissue microarrays from cohort 1 HCCs (n 5 137). K19 protein expression in cohort 2 HCCs (n 5 237) was correlated with the clinicopathologic parameters and messenger RNA (mRNA) levels of K19, uPAR, VIL2, Snail, Slug, and Twist. K19, EpCAM, c-kit, and CD133 positivity were observed in 18.2%, 35.0%, 34.3%, and 24.8%, respectively. K19 was most frequently expressed in combination with at least one other stemness-related marker (92.0%). K19-positive HCCs demonstrated more frequent major vessel invasion and increased tumor size, compared to K19-negative HCCs (P < 0.05). K19 was most significantly associated with EMT-related protein expression (e.g., vimentin, S100A4, uPAR, and ezrin) (P < 0.05) and a poor prognosis (overall survival: P 5 0.018; disease-free survival: P 5 0.007) in cohort 1. In cohort 2, HCCs with high K19 mRNA levels demonstrated higher mRNA levels of Snail, uPAR, and MMP2 (P < 0.05). K19-positive HCCs demonstrated more frequent microvascular invasion, fibrous stroma, and less tumor-capsule formation, compared to K19-negative HCCs (P < 0.05). K19 expression was a significant independent predictive factor of poor disease-free survival (P 5 0.032). Conclusion: K19 was well correlated with clinicopathologic features of tumor aggressiveness, compared to other stemnessrelated proteins. K19-positive HCCs showed significantly increased EMT-related protein and mRNA expression, suggesting that they may acquire more invasive characteristics, compared to K19-negative HCCs through the up-regulation of EMT-associated genes.

Immunohistochemical expression and clinical significance of suggested stem cell markers in hepatocellular carcinoma

“…15,21 The proportion of tissues expressing EpCAM in this study was a little higher than in earlier series, which reported EpCAM in 15.9%-48.7% of HCCs. [15][16][17][20][21][22][23] Kimura et al 23 have demonstrated that EpCAM expression was observed more often in HCC patients with HBV than in those with other etiologies, and the immunoreactivity of EpCAM has been found in up to 78% of HCC patients with HBV. The high proportion of HCC patients with HBV (74%) examined in this study might be a possible explanation for the higher EpCAMpositive rate.…”

“…20,21 The prognosis of patients with EpCAM-negative HCC is considered to be better than that of those with EpCAM-positive HCC. 11,[14][15][16]20,21 Although EpCAM expression was not found to be an independent predictor of survival in patients with HCC in this study, EpCAM expression was found to be associated with well-known unfavorable prognostic factors in HCC. Thus, the use of EpCAM expression as an unfavorable prognostic factor of HCC is reasonable.…”

“…[3][4][5][6] The expression of several proposed CSC markers in HCC has been reported and might prove to be useful for predicting the prognosis of HCC patients. 11,[14][15][16] However, little is known about the relationships between the IHC expression of CSC markers in paraffin-embedded tissue sections and clinicopathologic factors or clinical outcomes of HCC.…”

“…[14][15][16]18 EpCAM has been validated as a marker of stem cells in the liver, and EpCAM-positive HCC is likely to have originated from hepatic stem cells. 8,15,16,19 In the present study, we observed that EpCAM expression significantly correlated with tumor progression factors of HCC, such as elevated AFP level, vascular invasion, and high tumor grade. These findings are in agreement with previous studies showing that expression level of EpCAM correlates with de-differentiation 11 and vascular invasion and is associated with the high AFP level in HCC.…”

Tissue Diagnosis of Hepatocellular Carcinoma

“…Keratin 19, a progenitor cell/biliary marker, at a cut-off of 5% of positive tumor cells on immunohistochemistry, has been shown to correlate with poor clinical outcome. [53][54][55] …”