Association of preoperative EpCAM Circulating Tumor Cells and peripheral Treg cell levels with early recurrence of hepatocellular carcinoma following radical hepatic resection

Abstract

Background: This study was carried out to determine the prognostic significance of preoperative peripheral epithelial cell adhesion molecule- positive (EpCAM (+)) circulating tumor cell (CTC) and T regulatory (Treg) cell levels in hepatocellular carcinoma (HCC) patients for the prediction of postoperative recurrence following curative resection.

Methods: A total of 49 patients about to undergo curative resection for HCC were recruited into the study. PCR and FACS were used to detect the preoperative levels of EpCAM (mRNA+) CTCs and CD4(+)CD25(+)Foxp3(+) Treg cells. The prognostic value of EpCAM (mRNA+) CTCs, CD4(+)CD25(+)Foxp3(+) Treg cells, and other clinicopathological factors were analyzed by applying the Kaplan-Meier method and the multivariate Cox proportional hazards model.

Results: The number of EpCAM (mRNA+) CTCs and Treg/CD4(+) cells showed significant correlation as prognostic factors of postoperative HCC recurrence: EpCAM (mRNA+) CTC ≥ 2.22 (P = 0.001) and Treg/CD4(+) ≥ 5.07 (P = 0.045), with EpCAM (mRNA+) CTC ≥ 2.22 (P = 0.003, HR = 6.668) being the most important indicator. Patients with high CTC/Treg levels showed a significantly higher risk of developing postoperative HCC recurrence than those with low CTC/Treg levels (66.7 % vs. 10.3 %, P < 0.001). The high CTC/low Treg group also presented higher 1-year recurrence rates compared with the low CTC/low Treg level group (50.0 % vs. 10.3 %, P = 0.004).

Conclusions: Elevated EpCAM (mRNA+) CTC and Treg/CD4(+) levels were associated with early recurrence of HCC, indicative of poor clinical outcome. The combined detection of EpCAM (mRNA+) CTC and Treg/CD4(+) may therefore provide a novel prognostic predictor for HCC patients.

Keywords: Circulating tumor cells; Epithelial cell adhesion molecule; Hepatocellular carcinoma; Regulatory T cells; Tumor recurrence.

Figures

Fig. 1

Expression of EpCAMmRNA+ CTC and Treg/CD4+ in HCC recurrence vs no recurrence group. a, Based on 2−ΔΔCq algorithm transformation, the relative EpCAM mRNA expression of CTC in HCC patients with or without tumor recurrence after resection. EpCAMmRNA+ CTC showed significantly higher in recurrence patients (p = 0.001); b, The high ratio of Treg/CD4+ was observed d in recurrence group (p = 0.0197)

Fig. 2

Association between EpCAM+ CTC and Treg/CD4+ ratio. Distribution of Treg/CD4+ ratio in EpCAMmRNA+ -positive and negative group. The cutoff to discriminate between CTC-positive and -negative was set at 2.0 [9]

Fig. 3

The Prognostic significance of EpCAM mRNA+ CTC and Treg/CD4+ ratio in HCC patients. a. Kaplan–Meier analysis of HCC patients receiving curative resection treatment according to relative preoperative EpCAM expression <2.22 or ≥2.22 (2−ΔΔC q algorithm transformation). b. Kaplan–Meier analysis of patients with HCC undergoing curative resection according to preoperative Treg/CD4+ <5.07 % or ≥5.07 %. c. Prognostic significance of combination of EpCAMmRNA+ CTC and Treg/CD4+ load with respect to time to recurrence in patients with high EpCAMmRNA+ CTC (≥2.22) and high Treg/CD4+(≥5.07 %), high EpCAMmRNA+ CTC(<2.22) and low Treg/CD4+(<5.07 %), low/no EpCAMmRNA+ CTC (<2.22) and high Treg/CD4+(≥5.07 %), and low/no EpCAMmRNA+ CTC (<2.22) and low Treg/CD4+(<5.07 %)