Efficacy and Safety of Central Memory T Cells Combined With Adjuvant Therapy to Prevent Recurrence of Hepatocellular Carcinoma With Microvascular Invasion: A Pilot Study

Abstract

Background: Postoperative adjuvant transcatheter arterial chemoembolization (TACE) following curative hepatectomy has been reported to improve the clinical outcomes of hepatocellular carcinoma (HCC) patients with microvascular invasion (MVI), but more endeavors are required to achieve greater clinical benefit. Central memory T-cell (Tcm) self-transfusion has shown superior antitumor activity in several preclinical studies; however, clinical studies are rare. The aim of this study was to evaluate the clinical benefit and safety of combination treatment with Tcm self-transfusion and TACE as adjuvant treatment in HCC patients with MVI after curative hepatectomy.

Methods: From October 2016 to September 2018, primary HCC patients with histologically confirmed MVI who underwent curative hepatectomy at the Cancer Hospital of the Chinese Academy of Medical Sciences were recruited for this study. The patients were divided into a Tcm group (combined Tcm self-transfusion with TACE treatment) or a control group (TACE treatment alone) according to their willingness. The recurrence-free survival (RFS), quality-of-life (QOL) score, and adverse events of each patient were recorded within 2 years.

Results: A total of 52 patients were enrolled, and 48 were eligible for the final data analysis. The median follow-up time was 20.5 months (95% CI: 17.05-22.55 months). The median RFS time was 9.5 months in the control group; the cutoff date was not reached in the Tcm group (when the follow-up duration was 12 months, p = 0.049, HR = 0.40; 95% CI: 0.16-0.99). Compared with the control group, 1- and 2-year RFS rates were higher in the Tcm group (72.0% vs. 46.4% and 58.18% vs. 39.14%, respectively). Multivariate analysis did not indicate that Tcm treatment was an independent prognostic factor associated with HCC recurrence (p = 0.107, HR = 2.312; 95% CI: 0.835-6.400), which might be due to the small sample size of this study. Nevertheless, Tcm treatment effectively improved a reduced QOL due to HCC and liver function injury. Finally, the safety profile of Tcm treatment in this study was good, without any serious adverse events.

Conclusions: This pilot study showed that Tcm self-transfusion combined with TACE treatment might be a beneficial adjuvant therapy with good safety for primary HCC patients with MVI after curative hepatectomy.

Trial registration number: NCT03575806.

Keywords: Tcm treatment; adjuvant therapy; clinical study; hepatocellular carcinoma; microvascular invasion.

Conflict of interest statement

DL and HQ are current employees of Newish Technology (Beijing) Co., LTD. The authors declare that this study received funding from Newish Technology Co. LTD. The funder had the following involvement in the study: DL and HQ were involved in the preclinical study design, collection, analysis, interpretation of the preclinical data and the revision of the manuscript. Tcm cells were manufactured in the GMP laboratory of Newish Technology. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Copyright © 2021 Cai, Zhao, Liu, Xie, Qi, Ma, Sun and Zhao.

Figures

Figure 1

Phenotypes, cytotoxicity, and safety profiles of Tcms in the preclinical study, which was performed as described in the Supplementary Methods. (A) Representative flow plots of Tcm (central memory T cell), Tem (effector memory T cell), and Tn (naïve T cell) surface marker expression in cultured monocytes on D0 and D10. (B) Percentage of the indicated T-cell subsets among monocytes cultured from D0 to D10. Data are shown as the mean ± SD (n = 3). (C) Percentage of Tcms, Tems, and Tns in the CD3+ T-cell population among monocytes cultured from D0 to D10. Data are shown as the mean ± SD (n = 3). (D) Proliferation rate of cultured Tcms on D14 stimulated with anti-CD3/CD28 antibody, shown as carboxyfluorescein succinimidyl ester (CFSE) staining and cultured for over 4 consecutive days; unstimulated Tcms are shown as a control. (E) Cytokine secretion of cultured Tcms on D14 shown as the percentage of IFNγ+ and IL2+ cells measured at 0, 2, and 4 h after stimulation with phorbol myristate acetate/ionomycin (P/I) or anti-CD3/CD28 beads. Data are shown as the mean ± SD (n = 3). (F, G) Cytotoxicity (in vitro killing) of cultured Tcms on D14 against human hepatocellular carcinoma cell line QGY-7703, as measured by targeted cell stained with 7-AAD and Annexin V, detected by flow cytometry. Tcms and QGY-7703 cells were coincubated for 6 h at an effector:target ratio of 1:5. Representative flow plots and the percentage of Annexin V+ cells in target cells are shown in (F, G), respectively. (H) Representative immunohistochemistry-paraffin (IHC-P) images of tumor tissue slices immunostained with antibodies against CD4, CD8, and CD45RO and DAPI. Mice were inoculated subcutaneously with QGY-7703 tumor cells on D0 and then treated with Tcms or PBS (control) by tail vein injection, and on D42, the subcutaneous tumor was stripped for IHC-P. (I) Tcm persistence (shown as copies/μg DNA) in the tumor mass and peripheral blood of NPG mouse recipients measured by qPCR at the indicated time after Tcm transfusion by tail vein injection. Data are shown as the mean ± SD (n = 6 mice). (J–L) Different doses of Tcms were transfused into QGY-7703 tumor-bearing mice, and the (J) tumor volume and (K) body weight were measured on the indicated days. (L) The tumor weight of each mouse was measured on D42 posttumor cell injection. Animals in the control group received only PBS. Data are shown as the mean ± SEM (n = 6 mice). ns, not significant; *p < 0.5; **p < 0.01; ***p < 0.001; ****p < 0.0001.

Figure 2

Comparison of RFS between the Tcm group and the control group at (A) 12 months (p = 0.049) and (B) 24 months (p = 0.060). p-values were calculated with the Gehan-Breslow-Wilcoxon test.

Figure 3

Comparison of FACT-Hep scores according to Tcm treatment. ***p < 0.001.

Figure 4

Mean laboratory results of (A–F) liver and kidney function tests and (G–K) blood tests in the Tcm group and control group during the follow-up period. The upper and lower dotted lines in each graph are the upper and lower limits of normal, respectively; for graph with only one dotted line, the lower limits of normal is 0. ( ALB, albumin; ALT, alanine aminotransferase; AST, glutamic oxaloacetic aminotransferase; TBIL, total bilirubin; CRE, creatinine; UREA, urea; WBC, white blood cell; PLT, platelet; HGB, hemoglobin; NEU, neutrophil; LYMPH, lymphocyte; ULN, upper limit of normal).