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The effectiveness of immune checkpoint inhibitor (ICI) therapy for hepatocellular carcinoma (HCC) is limited by treatment resistance. However, the mechanisms underlying immunotherapy resistance remain elusive. We aimed to identify the role of CT10 regulator of kinase-like (CRKL) in resistance to anti-PD-1 therapy in HCC. Gene expression in HCC specimens from 10 patients receiving anti-PD-1 therapy was identified by RNA-sequencing. A total of 404 HCC samples from tissue microarrays were analyzed by immunohistochemistry. Transgenic mice (Alb-Cre/ Trp53 fl/fl) received hydrodynamic tail vein injections of a CRKL -overexpressing vector. Mass cytometry by time of flight was used to profile the proportion and status of different immune cell lineages in the mouse tumor tissues. CRKL was identified as a candidate anti-PD-1-resistance gene using a pooled genetic screen. CRKL overexpression nullifies anti-PD-1 treatment efficacy by mobilizing tumor-associated neutrophils (TANs), which block the infiltration and function of CD8+ T cells. PD-L1+ TANs were found to be an essential subset of TANs that were regulated by CRKL expression and display an immunosuppressive phenotype. Mechanistically, CRKL inhibits APC (adenomatous polyposis coli)-mediated proteasomal degradation of β-catenin by competitively decreasing Axin1 binding, and thus promotes VEGFα and CXCL1 expression. Using human HCC samples, we verified the positive correlations of CRKL/β-catenin/VEGFα and CXCL1. Targeting CRKL using CRISPR-Cas9 gene editing (CRKL knockout) or its downstream regulators effectively restored the efficacy of anti-PD-1 therapy in an orthotopic mouse model and a patient-derived organotypic tumor spheroid model. Activation of the CRKL/β-catenin/VEGFα and CXCL1 axis is a critical obstacle to successful anti-PD-1 therapy. Therefore, CRKL inhibitors combined with anti-PD-1 could be useful for the treatment of HCC. Here, we found that CRKL was overexpressed in anti-PD-1-resistant hepatocellular carcinoma (HCC) and that CRKL upregulation promotes anti-PD-1 resistance in HCC. We identified that upregulation of the CRKL/β-catenin/VEGFα and CXCL1 axis contributes to anti-PD-1 tolerance by promoting infiltration of tumor-associated neutrophils. These findings support the strategy of bevacizumab-based immune checkpoint inhibitor combination therapy, and CRKL inhibitors combined with anti-PD-1 therapy may be developed for the treatment of HCC. [Display omitted] • CRKL is significantly upregulated in anti-PD-1-resistant HCC. • CRKL binds β-catenin and inhibits its proteasomal degradation, contributing to the overexpression of VEGFα and CXCL1. • CRKL/β-catenin/VEGFα and CXCL1 axis upregulation promotes the infiltration and activation of PD-L1+ TANs. • Blocking CRKL/β-catenin/VEGFα and CXCL1 axis using bevacizumab or lenvatinib effectively overcomes anti-PD-1 resistance. [ABSTRACT FROM AUTHOR]

Despite remarkable advances in treatment, most patients with hepatocellular carcinoma (HCC) respond poorly to anti-programmed cell death 1 (anti-PD1) therapy. A deeper insight into the tolerance mechanism of HCC against this therapy is urgently needed. We performed next-generation sequencing, multiplex immunofluorescence, and dual-color immunohistochemistry and constructed an orthotopic HCC xenograft tumor model to identify the key gene associated with anti-PD1 tolerance. A spontaneously tumorigenic transgenic mouse model, an in vitro coculture system, mass cytometry, and multiplex immunofluorescence were used to explore the biological function of zinc finger protein 64 (ZFP64) on tumor progression and immune escape. Molecular and biochemical strategies like RNA-sequencing, chromatin immunoprecipitation-sequencing and mass spectrometry were used to gain insight into the underlying mechanisms of ZFP64. We showed that ZFP64 is frequently upregulated in tumor tissues from patients with anti-PD1-resistant HCC. Elevated ZFP64 drives anti-PD1 resistance by shifting macrophage polarization toward an alternative activation phenotype (M2) and fostering an inhibitory tumor microenvironment. Mechanistically, we primarily demonstrated that protein kinase C alpha (PKCα) directly phosphorylates ZFP64 at S226, leading to its nuclear translocation and the transcriptional activation of macrophage colony-stimulating factor (CSF1). HCC-derived CSF1 transforms macrophages to the M2 phenotype to drive immune escape and anti-PD1 tolerance. Notably, Gö6976, a protein kinase inhibitor, and lenvatinib, a multi-kinase inhibitor, reset the tumor microenvironment and restore sensitivity to anti-PD1 by blocking the PKCα/ZFP64/CSF1 axis. We propose that the PKCα/ZFP64/CSF1 axis is critical for triggering immune evasion and anti-PD1 tolerance. Inhibiting this axis with Gö6976 or lenvatinib overcomes anti-PD1 resistance in HCC. Despite remarkable treatment progress, most patients with hepatocellular carcinoma respond poorly to anti-PD1 therapy (a type of immunotherapy). A deeper insight into the tolerance mechanisms to this therapy is urgently needed. Herein, we unravel a previously unexplored mechanism linking tumor progression, macrophage polarization, and anti-PD1 resistance, and offer an attractive novel target for anti-PD1 combination therapy, which may benefit patients with hepatocellular carcinoma. [Display omitted] • ZFP64 is frequently upregulated in anti-PD1 resistant HCC. • PKCα/ZFP64/CSF1 axis is critical for triggering immune evasion and anti-PD1 tolerance. • Gö6976 and lenvatinib overcome anti-PD1 resistance by blocking the PKCα/ZFP64/CSF1 axis. • Gö6976 combined with anti-PD1 could be an effective new strategy in HCC therapy. [ABSTRACT FROM AUTHOR]

Although the treatment of hepatocellular carcinoma (HCC) has been revolutionized by the advent of effective systemic therapies, the prognosis of patients with HCC remains dismal. Herein, we examined the pathophysiological role of PARG and assessed the utility of targeting dePARylation for HCC therapy. The oncogenic function of PARG was evaluated in 2 orthotopic xenograft models and a Parg flox/flox mice model. The therapeutic efficacy of PARG inhibitors in combination with an anti-PD-1 antibody were assessed in murine orthotopic models. Microarray analysis was used to evaluate the pathological relevance of the PARG/DDB1/c-Myc/MMR axis. High PARG expression was strongly associated with poor HCC prognosis. Hepatocyte-specific PARG deletion significantly impaired liver tumorigenesis. PARG promoted HCC growth and metastasis through DDB1-dependent modulation of c-Myc. Specifically, PARG dePARylated DDB1 and consequently promoted DDB1 autoubiquitination, thus stabilizing the c-Myc protein in HCC cells. PARG downregulation attenuated c-Myc-induced MMR expression and PARG deficiency was correlated with a favorable prognosis in patients with HCC treated with anti-PD-1-based immunotherapy. In addition, PARG inhibitors could act in synergy with anti-PD-1 antibodies in orthotopic mouse models. PARG can act as an oncogene in HCC by modulating PARG/DDB1/c-Myc signaling and could be used as a biomarker to identify patients with HCC who may benefit from anti-PD-1 treatment. Our findings suggest that co-inhibition of PARG and PD-1 is an effective novel combination strategy for patients with HCC. The increase in deaths due to hepatocellular carcinoma (HCC) is a growing concern, with the mechanisms responsible for HCC development still incompletely understood. Herein, we identify a novel mechanism by which the protein PARG contributes to HCC development. Inhibition of PARG increased the efficacy of anti-PD-1 therapy (a type of immunotherapy) in HCC. These findings support the future clinical development of PARG inhibitors, potentially in combination with anti-PD-1 inhibitors. [Display omitted] • PARG is upregulated in HCC tissues and predicts poor prognosis. • PARG regulation of c-MYC expression is dependent on PARG-mediated DDB1 degradation in HCC. • PARG promotes growth and metastasis of HCC via a newly identified PARG/DDB1/c-Myc signaling axis. • PARG promotes MMR expression and could potentially be used to predict which patients might benefit from anti-PD-1 treatment. • PARG inhibition potentiates the efficacy of PD-1 blockade in a murine HCC model. [ABSTRACT FROM AUTHOR]

The prognostic value and clinical relevance of tertiary lymphoid structures (TLSs) in intrahepatic cholangiocarcinoma (iCCA) remain unclear. Thus, we aimed to investigate the prognostic value and functional involvement of TLSs in iCCA. We retrospectively included 962 patients from 3 cancer centers across China. The TLSs at different anatomic subregions were quantified and correlated with overall survival (OS) by Cox regression and Kaplan-Meier analyses. Multiplex immunohistochemistry (mIHC) was applied to characterize the composition of TLSs in 39 iCCA samples. A quaternary TLS scoring system was established for the intra-tumor region (T score) and peri-tumor region (P score) respectively. T scores positively correlated with favorable prognosis (p < 0.001), whereas a high P score signified worse survival (p < 0.001). mIHC demonstrated that both T follicular helper and regulatory T cells were significantly increased in intra-tumoral TLSs compared to peri-tumoral counterparts (p < 0.05), and regulatory T cell frequencies within intra-tumoral TLSs were positively associated with P score (p < 0.05) rather than T score. Collectively, the combination of T and P scores stratified iCCAs into 4 immune classes with distinct prognoses (p < 0.001) that differed in the abundance and distribution pattern of TLSs. Patients displaying an immune-active pattern had the lowest risk, with 5-year OS rates of 68.8%, whereas only 3.4% of patients with an immune-excluded pattern survived at 5 years (p < 0.001). The C-index of the immune class was statistically higher than the TNM staging system (0.73 vs. 0.63, p < 0.001). These results were validated in an internal and 2 external cohorts. The spatial distribution and abundance of TLSs significantly correlated with prognosis and provided a useful immune classification for iCCA. T follicular helper and regulatory T cells may play a critical role in determining the functional orientation of spatially different TLSs. Tertiary lymphoid structures (TLSs) are associated with favorable prognosis in a number of cancers. However, their role in intrahepatic cholangiocarcinoma (iCCA) remains unclear. Herein, we comprehensively evaluated the spatial distribution, abundance, and cellular composition of TLSs in iCCA, and revealed the opposite prognostic impacts of TLSs located within or outside the tumor. This difference could be mediated by the different immune cell subsets present within the spatially distinct TLSs. Based on our analysis, we were able to stratify iCCAs into 4 immune subclasses associated with varying prognoses. [Display omitted] • Tertiary lymphoid structures (TLSs) within and outside the tumor have opposite prognostic impacts on patients with iCCA. • The heterogeneous distribution of Tfh and Treg cells within distinct TLSs might be a determinant of their functional state. • The geographic integration of TLSs stratified iCCAs into 4 immune subclasses with distinct clinical outcomes. [ABSTRACT FROM AUTHOR]

Background & Aims: Neutrophil infiltration has been linked to clinical outcome of various cancer types. However, its role in hepatocellular carcinoma (HCC) is unclear. In this study, we investigated prognostic values for intratumoral and peritumoral neutrophils in HCC patients undergoing curative resection. Methods: The expression of CD66b, CD8, TGF-beta, and CD34 was assessed by immunohistochemistry in tissue microarrays containing paired intratumoral and peritumoral tissues from 197 patients receiving curative resection for HCC. Prognostic values for these and other clinicopathologic factors were evaluated. Results: Intratumoral CD66b+ neutrophils significantly correlated with CD8+ T cells (r =0.240, p =0.004), TGF-beta expression (p =0.012), BCLC stage (p =0.016), and early recurrence (p =0.041). Increased intratumoral neutrophils were significantly associated with decreased RFS/OS (p =0.001 and p <0.001, respectively) in univariate analysis and were identified as an independent prognostic factor (HR=1.845, 95% CI=1.169–2.911, p =0.008 for RFS; HR=2.578, 95% CI=1.618–4.106, p <0.001 for OS) in multivariate analysis. Intratumoral neutrophil-to-CD8+ T cell ratio (iNTR) better predicted the outcome in terms of minimum p values. Intratumoral neutrophils were also demonstrated to be statistically predictive for RFS/OS in the normal AFP subgroup, small HCC subgroup, and validation cohort. However, peritumoral neutrophils were not associated with the outcome of HCC. Conclusions: The presence of intratumoral neutrophils was a poor prognostic factor for HCC after resection. Intratumoral neutrophil-to-CD8+ T cell ratio was a better predictor of outcome. [Copyright &y& Elsevier]

• 182 Chinese hepatocellular carcinomas were sequenced. • WNK2 , RUNX1T1 , CTNNB1 , TSC1 , and TP53 somatic mutations correlated with early tumor recurrence after curative resection. • WNK2 displayed somatic mutation, copy number loss, and downregulated expression in HCC. • WNK2 deficiency leads to ERK1/2 signalling activation, TAM infiltration, and tumor growth and metastasis. Early recurrence of hepatocellular carcinoma (HCC) after curative resection is common. However, the association between genetic mechanisms and early HCC recurrence, especially in Chinese patients, remains largely unknown. We performed whole-genome sequencing (49 cases), whole-exome sequencing (18 cases), and deep targeted sequencing (115 cases) on 182 primary HCC samples. Focusing on WNK2 , we used Sanger sequencing and qPCR to evaluate all the coding exons and copy numbers of that gene in an additional 554 HCC samples. We also explored the functional effect and mechanism of WNK2 on tumor growth and metastasis. We identified 5 genes (WNK2 , RUNX1T1 , CTNNB1 , TSC1 , and TP53) harboring somatic mutations that correlated with early tumor recurrence after curative resection in 182 primary HCC samples. Focusing on WNK2 , the overall somatic mutation and copy number loss occurred in 5.3% (39/736) and 27.2% (200/736), respectively, of the total 736 HCC samples. Both types of variation were associated with lower WNK2 protein levels, higher rates of early tumor recurrence, and shorter overall survival. Biofunctional investigations revealed a tumor-suppressor role of WNK2: its inactivation led to ERK1/2 signaling activation in HCC cells, tumor-associated macrophage infiltration, and tumor growth and metastasis. Our results delineate genomic events that characterize Chinese HCCs and identify WNK2 as a driver of early HCC recurrence after curative resection. We applied next-generation sequencing and conducted an in-depth genomic analysis of hepatocellular carcinomas from a Chinese patient cohort. The results delineate the genomic events that characterize hepatocellular carcinomas in Chinese patients and identify WNK2 as a driver associated with early tumor recurrence after curative resection. [ABSTRACT FROM AUTHOR]

Background & Aims Intrahepatic cholangiocarcinoma (ICC) is the second-most lethal primary liver cancer. Little is known about intratumoral heterogeneity (ITH) and its impact on ICC progression. We aimed to investigate the ITH of ICC in the hope of helping to develop new therapeutic strategies. Methods We obtained 69 spatially distinct regions from six operable ICCs. Patient-derived primary cancer cells (PDPCs) were established for each region, followed by whole-exome sequencing (WES) and multi-level validation. Results We observed widespread ITH for both somatic mutations and clonal architecture, shaped by multiple mechanisms, like clonal “illusion”, parallel evolution and chromosome instability. A median of 60.3% of mutations were heterogeneous, among which 85% of the driver mutations were located on the branches of tumor phylogenetic trees. Many truncal and clonal driver mutations occurred in tumor suppressor genes, such as TP53 , SMARCB1 and PBRM1 that are involved in DNA repair and chromatin-remodeling. Genome doubling occurred in most cases (5/6) after the accumulation of truncal mutations and was shared by all intratumoral sub-regions. In all cases, ongoing chromosomal instability is evident throughout the evolutionary trajectory of ICC. The recurrence of ICC1239 provided evidence to support the polyclonal metastatic seeding in ICC. The change of mutation landscape and internal diversity among subclones during metastasis, such as the loss of chemoresistance mediator, can be used for new treatment strategies. Targeted therapy against truncal alterations, such as IDH1, JAK1, and KRAS mutations and EGFR amplification, was developed in 5/6 patients. Conclusions Integrated investigations of spatial ITH and clonal evolution may provide an important molecular foundation for enhanced understanding of tumorigenesis and progression in ICC. Lay summary We applied multiregional whole-exome sequencing to investigate the evolution of intrahepatic cholangiocarcinoma (ICC). The results revealed that many factors, such as parallel evolution and chromosome instability, may participate and promote the branch diversity of ICC. Interestingly, in one patient with primary and recurrent metastatic tumors, we found evidence of polyclonal metastatic seeding, indicating that symbiotic communities of multiple clones existed and were maintained during metastasis. More realistically, some truncal alterations, such as IDH1, JAK1, and KRAS mutations and EGFR amplification, could be promising treatment targets in patients with ICC. [ABSTRACT FROM AUTHOR]

Background & Aims Identifying target genetic mutations in hepatocellular carcinoma (HCC) for therapy is made challenging by intratumoral heterogeneity. Circulating cell-free DNAs (cfDNA) may contain a more complete mutational spectrum compared to a single tumor sample. This study aimed to identify the most efficient strategy to identify all the mutations within heterogeneous HCCs. Methods Whole exome sequencing (WES) and targeted deep sequencing (TDS) were carried out in 32 multi-regional tumor samples from five patients. Matched preoperative cfDNAs were sequenced accordingly. Intratumoral heterogeneity was measured using the average percentage of non-ubiquitous mutations (present in parts of tumor regions). Profiling efficiencies of single tumor specimen and cfDNA were compared. The strategy with the highest performance was used to screen for actionable mutations. Results Variable levels of heterogeneity with branched and parallel evolution patterns were observed. The heterogeneity decreased at higher sequencing depth of TDS compared to measurements by WES (28.1% vs. 34.9%, p <0.01) but remained unchanged when additional samples were analyzed. TDS of single tumor specimen identified an average of 70% of the total mutations from multi-regional tissues. Although genome profiling efficiency of cfDNA increased with sequencing depth, an average of 47.2% total mutations were identified using TDS, suggesting that tissue samples outperformed it. TDS of single tumor specimen in 66 patients and cfDNAs in four unresectable HCCs showed that 38.6% (26/66 and 1/4) of patients carried mutations that were potential therapeutic targets. Conclusions TDS of single tumor specimen could identify actionable mutations targets for therapy in HCC. cfDNA may serve as secondary alternative in profiling HCC genome. Lay summary Targeted deep sequencing of single tumor specimen is a more efficient method to identify mutations in hepatocellular carcinoma made from mixed subtypes compared to circulating cell-free DNA in blood. cfDNA may serve as secondary alternative in profiling HCC genome. Identifying mutations may help clinicians choose targeted therapy for better individual treatments. [ABSTRACT FROM AUTHOR]

Background & Aims: The immunosuppressive network within the tumor microenvironment is one of the major obstacles to the success of cancer immunotherapy. γδ T cells are attractive effectors for cancer immunotherapy. Nevertheless, the promising anti-tumor effect in vitro is partially if not totally mitigated in vivo. Thus, understanding the immune status of tumor-infiltrating γδ T cells is essential for orchestrating effective immunotherapy strategies. In this study, we have investigated the immunophenotype and function of γδ T cells in hepatocellular carcinoma (HCC) patients. Methods: The phenotype of γδ T cells in peripheral blood, and peritumoral and tumoral tissues of HCC patients (n=61) was characterized by flow cytometry. Functional analysis of the HCC-infiltrating γδ T cells was conducted directly after γδ T cell isolation. Results: The infiltration of γδ T cells in tumoral tissues was significantly reduced compared to paired peritumoral tissues. Impairment in degranulation of the granule pathway and downregulation of IFN-γ secretion were also demonstrated in HCC-infiltrating γδ T cells, which was in agreement with the results of gene microarray analysis, and further strengthened by the compromised specific cytotoxicity and IFN-γ secretion in vitro. Moreover, isolated HCC-infiltrating CD4+CD25+ regulatory T cells (Treg cells) directly suppressed the cytotoxic function and IFN-γ secretion of γδ T cells in a TGFβ- and IL-10-dependent manner. Conclusions: The effector function of γδ T cells was substantially impaired in HCC, which is partially mediated by Treg cells. We propose a new mechanism by which immune privilege develops within the tumor milieu. [Copyright &y& Elsevier]

Background & Aims: Fibroblast growth factor 21 (FGF21), a hormone primarily secreted by the liver in response to peroxisome proliferator-activated receptor-α (PPARα) activation, has recently been shown to possess beneficial effects on lipid metabolism and hepatic steatosis in animal models. This study investigated the association of FGF21 with nonalcoholic fatty liver disease (NAFLD) in Chinese patients. Methods: Serum FGF21 levels were determined by enzyme-linked immunosorbent assay (ELISA) in 224 NAFLD and 124 control subjects, and their association with parameters of adiposity, glucose, and lipid profiles and levels of liver injury markers was studied. Besides serum concentrations, the mRNA expression of FGF21 in the liver tissue was also quantified by real-time PCR in 17 subjects with different degrees of steatosis, and was correlated with the levels of intrahepatic lipid. The protein levels of FGF21 were determined by quantitative ELISA. Results: Serum FGF21 levels in patients with NAFLD (402.38pg/ml [242.03, 618.25]) were significantly higher than those in control subjects (198.62pg/ml [134.96, 412.62]) (p <0.01). In human liver tissues, FGF21 mRNA expression increased with the degree of steatosis. Both FGF21 mRNA expression and serum FGF21 concentrations were positively correlated with intrahepatic triglyceride (TG) having r =0.692 and r =0.662, respectively, at p <0.01. Furthermore, the increased expression of FGF21 was accompanied by elevated protein levels in liver tissues. Conclusions: These results support the role of FGF21 as a key regulator of hepatic lipid metabolism in humans, and suggest that serum FGF21 can be potentially used as a biomarker for NAFLD. [ABSTRACT FROM AUTHOR]

Background & aims: Phosphatidylethanolamine-binding protein 1 (PEBP1, also RKIP) plays a pivotal role in cancer by regulating multiple cellular signaling processes and suppressing metastasis in animal models. We examined whether PEBP1 expression in hepatocellular carcinoma (HCC) correlated with the risk of recurrence and survival after resection. Methods: A randomly selected cohort of 240 Chinese HCC patients, predominantly hepatitis B related, formed the basis of the study. PEBP1 expression levels were evaluated by immunohistochemistry and real-time reverse-transcriptase PCR. Survival analysis was performed by univariate and multivariate analyses. The results were further validated in an independent series of 403 patients. The relevance of PEBP1 to phospho-ERK was determined by Western blot analysis on clinical samples and hepatoma cell lines. Results: PEBP1, prevalently down-regulated in HCC, was significantly associated with tumor invasive characteristics (such as vascular invasion, lack of encapsulation, poor differentiation and large size). Both PEBP1 protein and mRNA levels were independent predictors for tumor recurrence (hazard ratio (HR)=1.877, p =0.001; HR=2.633, p =0.001; respectively), and patient survival (HR=1.796, p =0.004; HR=1.730, p =0.044; respectively). The prognostic value of PEBP1 was then confirmed in the validation cohort. In addition, Western blot suggested that loss of PEBP1 led to hyperactivity of MAPK signaling. Conclusions: Down-regulation of PEBP1 in HCC indicated aggressive tumor behaviors and predicted a worse clinical outcome, which may be a useful biomarker to identify the patients at high risk of post-operative recurrence. [ABSTRACT FROM AUTHOR]

Background/Aims: To assess the value of Dickkopf-1 (DKK1) for predicting clinical outcome of human hepatocellular carcinoma (HCC) in patients with HCC. Methods: Expression of DKK1 and β-catenin was investigated in HCC cell lines using qRT-PCR, Western blotting and immunofluorescence. Tissue microarrays representing 314 HCC patients were used to determine the expression patterns of DKK1 and β-catenin by immunohistochemistry, and prognostic significance was assessed by using Kaplan–Meier survival estimates and log-rank tests. Results: The expression level of DKK1 was associated with the staining pattern of β-catenin in HCC cell lines, and DKK1 overexpression correlated with β-catenin cytoplasmic/nuclear accumulation in clinical HCC samples (P =0.011, correlation coefficient=0.144). High DKK1 expression predicted unfavorable prognosis in HCC patients, especially in early stage patients and those with normal AFP levels. In multivariate analyses, DKK1 was an independent predictor for overall survival (OS) (P =0.002) and disease-free survival (DFS) (P =0.002) of HCC patients. Furthermore, the HCC patients with high DKK1 expression and cytoplasmic/nuclear β-catenin accumulation had very poor prognosis. Conclusions: Elevated expression of DKK1 is a critical event in patients with HCC that indicates poor clinical outcome. DKK1, alone or combined with β-catenin, is a novel prognostic predictor for HCC patients. [Copyright &y& Elsevier]

Background/Aims: To study the impact of hepatitis B e antigen on tumor recurrence and patients’ survival after curative resection of hepatocellular carcinoma. Methods: Two hundred and three patients with small hepatocellular carcinomas (⩽3cm) who had undergone curative resection were divided into HBeAg-positive group and HBeAg-negative group. Clinicopathological factors and postoperative outcomes were compared between groups, and risk factors for survival and recurrence were studied. Results: The median follow-up time was 32.9months. Patients with negative HBeAg had higher 5-year overall survival rates (76% vs 53.9%, hazards ratio (HR): 2.363, 95% CI: 1.335–4.181, p =0.002) and 5-year disease-free survival rates (52.9% vs 37.4%, HR: 1.603, 95% CI: 1.00–2.561, p =0.046).There was no significant difference in operative morbidity and tumor factors between the two groups, but younger age, higher serum alanine aminotransferase levels, and more macronodular cirrhosis were found in the HBeAg-positive group. Multivariate analysis revealed that age >50years, HBeAg positivity and macronodular cirrhosis were independent factors for overall survival, and HBeAg positivity and multiple tumor nodules were independent factors for disease-free survival. Positive serum HBeAg was associated with a higher risk of early recurrence (within 1year). Conclusions: HBeAg is associated with a higher risk of early recurrence and poorer survival in patients after curative resection of small HCC. [Copyright &y& Elsevier]

A correction to the article "Circumventing Intratumoral Heterogeneity to Identify Potential Therapeutic Targets in Hepatocellular Carcinoma," published in a previous issue of the journal is presented.