TMB Signature-Related RCAN2 Promotes Apoptosis by Upregulating EHF/DR5 Pathway in Hepatocellular Carcinoma.

BACKGROUND: The tumour mutation burden (TMB) is a valuable indicator of the accumulation of somatic mutations, and is thought to be associated with the biological behaviour and prognosis of tumours. However, the related genetic mechanism for these association is still unclear. The aim of the present study was to identify the key gene(s) associated with TMB in hepatocellular carcinoma (HCC) and to investigate its biological functions, downstream transcription factors, and mechanism of action. METHODS: Patients in The Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) database were classified according to TMB signature-related genes. Key genes related to the TMB signature and tumour prognosis were identified. Immunohistochemistry and Quantitative Real-Time Polymerase Chain Reaction (qPCR) were then used to assess gene expression in clinical HCC tissues and HCC cells. Cells with altered gene expression were evaluated for the effect on cell proliferation and apoptosis, both in vitro and in vivo. Three independent databases and cell sequencing data were used to identify the mechanisms involved and the downstream transcription factors. The mechanism was also studied by altering the expression of downstream transcription factors in vitro. RESULT: The integrated cluster (IC) 2 group, characterized by 99 TMB signature-related genes, showed a significant different TMB score compared to the IC1 group (p < 0.001), as well as more favourable tumour prognosis (p = 0.031). We identified five key prognostic genes that were differentially expressed between IC2 and IC1 and were associated with overall survival. The expression of one of these key prognostic genes, RCAN2, was negatively correlated with TMB in 18 out of 33 tumour types examined. A high level of RCAN2 was correlated with better overall survival in HCC (p = 0.0009). Overexpression of RCAN2 enhanced apoptosis in vitro and in vivo, whereas knockdown of RCAN2 attenuated apoptosis. The mechanism by which RCAN2 promotes apoptosis may involve upregulation of the expression of ETS homologous factor (EHF) and of death receptor 5 (DR5). CONCLUSIONS: Downregulation of RCAN2 expression was found to correlate with elevated TMB in multiple cancer types. RCAN2 was also found to be a biomarker of HCC prognosis, and to promote the apoptosis of HCC cells through the EHF/DR5 pathway. These findings provide a new perspective on systemic treatment for advanced HCC with a high TMB.

DNA methylation-activated full-length EMX1 facilitates metastasis through EMX1-EGFR-ERK axis in hepatocellular carcinoma.

Altered DNA methylation is a crucial epigenetic event in hepatocellular carcinoma (HCC) development and progression. Through methylation-transcriptomic analysis, we identified a set of sixty potential DNA methylation-based epidriver genes. In this set of genes, we focused on the hypermethylation of EMX1, which is frequently observed in hepatobiliary tumors. Despite of its frequent occurrence, the function of EMX1 remains largely unknown. By utilizing bisulfite-next-generation sequencing, we have detected EMX1 DNA hypermethylation on the gene body, which is positively correlated with EMX1 mRNA expression. Further analysis revealed that EMX1 mRNA terminal exon splicing in HCC generated two protein isoforms: EMX1 full length (EMX1-FL) and alternative terminal exon splicing isoform (EMX1-X1). Cellular functional assays demonstrated that gain-of-function EMX1-FL, but not EMX1-X1, induced HCC cells migration and invasion while silencing EMX1-FL inhibited HCC cells motility. This result was further validated by in vivo tumor metastasis models. Mechanistically, EMX1-FL bound to EGFR promoter, promoting EGFR transcription and activating EGFR-ERK signaling to trigger tumor metastasis. Therefore, EGFR may be a potential therapeutic target for EMX1-high expression HCC. Our work illuminated the crucial role of gene body hypermethylation-activated EMX1-FL in promoting tumorigenesis and metastasis in HCC. These findings pave the way for targeting the EMX1-EGFR axis in HCC tumorigenicity and metastasis.

Hepatic Arterial Infusion of Oxaliplatin, Fluorouracil, and Leucovorin Versus Transarterial Chemoembolization for Large Hepatocellular Carcinoma: A Randomized Phase III Trial.

PURPOSE: In a previous phase II trial, hepatic arterial infusion chemotherapy (HAIC) with infusional fluorouracil, leucovorin, and oxaliplatin (FOLFOX) yielded higher treatment responses than transarterial chemoembolization (TACE) in large unresectable hepatocellular carcinoma. We aimed to compare the overall survival of patients treated with FOLFOX-HAIC versus TACE as first-line treatment in this population. METHODS: In this randomized, multicenter, open-label trial, adults with unresectable hepatocellular carcinoma (largest diameter ≥ 7 cm) without macrovascular invasion or extrahepatic spread were randomly assigned 1:1 to FOLFOX-HAIC (oxaliplatin 130 mg/m2, leucovorin 400 mg/m2, fluorouracil bolus 400 mg/m2 on day 1, and fluorouracil infusion 2,400 mg/m2 for 24 hours, once every 3 weeks) or TACE (epirubicin 50 mg, lobaplatin 50 mg, and lipiodol and polyvinyl alcohol particles). The primary end point was overall survival by intention-to-treat analysis. Safety was assessed in patients who received ≥ 1 cycle of study treatment. RESULTS: Between October 1, 2016, and November 23, 2018, 315 patients were randomly assigned to FOLFOX-HAIC (n = 159) or TACE (n = 156). The median overall survival in the FOLFOX-HAIC group was 23.1 months (95% CI, 18.5 to 27.7) versus 16.1 months (95% CI, 14.3 to 17.9) in the TACE group (hazard ratio, 0.58; 95% CI, 0.45 to 0.75; P < .001). The FOLFOX-HAIC group showed a higher response rate than the TACE group (73 [46%] v 28 [18%]; P < .001) and a longer median progression-free survival (9.6 [95% CI, 7.4 to 11.9] v 5.4 months [95% CI, 3.8 to 7.0], P < .001). The incidence of serious adverse events was higher in the TACE group than in the FOLFOX-HAIC group (30% v 19%, P = .03). Two deaths in the FOLFOX-HAIC group and two in the TACE group were deemed to be treatment-related. CONCLUSION: FOLFOX-HAIC significantly improved overall survival over TACE in patients with unresectable large hepatocellular carcinoma.

Toripalimab Combined With Hepatic Arterial Infusion Chemotherapy Versus Lenvatinib for Advanced Hepatocellular Carcinoma.

Purpose: Immunotherapy combined with chemotherapy have synergistic effects in multiple malignancies. We aimed to compare the efficacy and safety of toripalimab plus hepatic arterial infusion chemotherapy (HAIC) of oxaliplatin, fluorouracil, and leucovorin versus lenvatinib in advanced hepatocellular carcinoma (HCC). Materials and Methods: We conducted this retrospective study at 3 hospitals in China and eligible patients were 18 years or older and had a primary diagnosis of unresectable HCC with macroscopic vascular invasion and/or extrahepatic spread. These patients were treated with toripalimab plus HAIC or lenvatinib monotherapy. The primary endpoint was progression-free survival (PFS) and the secondary endpoints were overall survival (OS), disease control rate per response evaluation criteria in solid tumors (RECIST) 1.1, and objective response rate (ORR) per RECIST 1.1. The results were compared by Student's test or the chi-square test, and the survival curves were calculated by the Kaplan-Meier method, and propensity-score matching (PSM) was used to reduce bias. Results: A total of 118 patients were recruited for this study: 53 in the TorHAIC group and 65 in the lenvatinib group. We found that the TorHAIC group showed a longer PFS (9.3 [95% CI, 7.81-10.8] vs 4.8 months [95% CI, 3.31-6.29]; hazard ratio [HR] = 0.57, 95% CI, 0.38-0.85; p = .006), a longer OS (17.13 [95% CI, 13.99-20.27] vs 10.1 months [95% CI, 8.14-12.06]; HR = 0.5, 95% CI, 0.31 - 0.81; p = .005), a higher disease control rate (86.8% vs 69.2%, p = .002) and a higher ORR (47.2% vs 9.2%, p < .001) by RECIST criteria than the lenvatinib group. Both toripalimab plus HAIC and lenvatinib had acceptable safety profiles. No treatment-related deaths occurred in this study. In the propensity score-matched cohorts (47 pairs), the outcomes in the TorHAIC group were also better than those in the lenvatinib group (p < .05). Conclusion: Toripalimab plus HAIC was tolerable and effective in advanced HCC and the result needs to be confirmed in the phase III trial.

Construction of a single nucleotide variant score-related gene-based prognostic model in hepatocellular carcinoma: analysis of multi-independent databases and validation in vitro.

BACKGROUND: The accumulation of single nucleotide variants (SNVs) and the emergence of neoantigens can affect tumour proliferation and the immune microenvironment. However, the SNV-related immune microenvironment characteristics and key genes involved in hepatocellular carcinoma (HCC) are still unclear. We aimed to evaluate differences in the SNV-related immune microenvironment, construct a prognostic model and validate the key genes in vitro. METHODS: The categories of samples were defined by the expression of SNV score-related genes to evaluate the differences in mutational features, immune environment and prognosis. The survival model was constructed with survival-associated genes and verified in two independent test datasets. RCAN2, the key gene screened out for biofunction, was validated in vitro. RESULTS: IC2, among the three integrated clusters (IC1, IC2, IC3) classified by the 82 SNV score-related genes, was distinct from the rest in SNV score and immune cell infiltration, showing a better prognosis. Seven prognostic markers, HTRA3, GGT5, RCAN2, LGALS3, CXCL1, CLEC3B, and CTHRC1, were screened to construct a prognostic model. The survival model distinguished high-risk patients with poor prognoses in three independent datasets (log-rank P < 0.0001, 0.011, and 0.0068, respectively) with acceptable sensitivity and specificity. RCAN2 was inversely correlated with NK cell infiltration, and knockdown of RCAN2 promoted proliferation in HCC. CONCLUSIONS: This study revealed the characteristics of the HCC SNV-associated subgroup and screened seven latent markers for their accuracy of prognosis. Additionally, RCAN2 was preliminarily proven to influence proliferation in HCC and it had a close relationship with NK cell infiltration in vitro. With the capability to predict HCC outcomes, the model constructed with seven key differentially expressed genes offers new insights into individual therapy.

Hepatic Arterial Infusion Chemotherapy of Oxaliplatin, Fluorouracil, and Leucovorin With or Without Sorafenib as Initial Treatment for Advanced Hepatocellular Carcinoma.

PURPOSE: Our previous study showed that hepatic arterial infusion chemotherapy (HAIC) using oxaliplatin, fluorouracil, and leucovorin (FOLFOX) plus sorafenib provided a significant survival benefit over sorafenib for advanced hepatocellular carcinoma. However, it is unclear whether the survival benefit should be attributed to the synergism between HAIC and sorafenib or just HAIC alone. We aim to compare HAIC using FOLFOX plus sorafenib with HAIC alone in patients with advanced hepatocellular carcinoma. MATERIALS AND METHODS: This was a retrospective study including 225 eligible patients treated with HAIC using FOLFOX (HAIC alone group, n=126, oxaliplatin 85 mg/m², leucovorin 400 mg/m², fluorouracil bolus 400 mg/m² and 2400 mg/m² for 46 hours, every 3 weeks) alone or HAIC plus sorafenib (soraHAIC group, n=99, sorafenib 400 mg twice daily). Survival curves were calculated by the Kaplan-Meier method, and propensity-score matching was used to reduce bias. RESULTS: The soraHAIC group showed a longer overall survival (12.9 [95% CI, 10.4-15.4] vs. 10.5 [95% CI, 9.5-11.5] months, HR=0.71 [95% CI, 0.53-0.96]; P=0.025), a better progression free survival (7.0 [95% CI, 5.3-8.8] vs. 5.3 [95% CI, 3.5-7.1] months, HR=0.76 [95% CI, 0.58-0.99]; P=0.046), and a higher disease control rate (RECIST 1.1: 74.8% vs. 61.1%, P=0.030) than the HAIC alone group. In multivariate analysis, soraHAIC was an independent favor factor for survival. In terms of the grade 3/4 adverse event, hand-foot skin reaction was more frequent in the soraHAIC group than the HAIC alone group. In the propensity-score matched cohorts (93 pairs), the overall survival, the progression free survival and disease control rates in the soraHAIC group were also better than those in the HAIC group (P<0.05). CONCLUSION: HAIC plus sorafenib may improve overall survival and progression free survival compared with HAIC alone as initial treatment for advanced hepatocellular carcinoma.

Lenvatinib, toripalimab, plus hepatic arterial infusion chemotherapy versus lenvatinib alone for advanced hepatocellular carcinoma.

BACKGROUND: Lenvatinib is the first-line treatment for advanced hepatocellular carcinoma, but prognosis is still unsatisfactory. Recently, hepatic arterial infusion chemotherapy (HAIC), and immune checkpoint inhibitors showed promising results for advanced hepatocellular carcinoma. Considering different anti-malignancy mechanisms, combining these three treatments may improve outcomes. This study aimed to compare the efficacy and safety of lenvatinib, toripalimab, plus HAIC versus lenvatinib for advanced hepatocellular carcinoma. METHODS: This was a retrospective study including patients treated with lenvatinib [8 mg (⩽60 kg) or 12 mg (>60 kg) once daily] or lenvatinib, toripalimab plus HAIC [LeToHAIC group, lenvatinib 0-1 week prior to initial HAIC, 240 mg toripalimab 0-1 day prior to every HAIC cycle, and HAIC with FOLFOX regimen (oxaliplatin 85 mg/m2, leucovorin 400 mg/m2, 5-fluorouracil bolus 400 mg/m2 on day 1, and 5-fluorouracil infusion 2400 mg/m2 for 46 h, every 3 weeks)]. Progression-free survival, overall survival, objective response rate, and treatment-related adverse events were compared. RESULTS: From February 2019 to August 2019, 157 patients were included in this study: 71 in the LeToHAIC group and 86 in the lenvatinib group. The LeToHAIC group showed longer progression-free survival (11.1 versus 5.1 months, p < 0.001), longer overall survival (not reached versus 11 months, p < 0.001), and a higher objective response rate (RECIST: 59.2% versus 9.3%, p < 0.001; modified RECIST: 67.6% versus 16.3%, p < 0.001) than the lenvatinib group. In addition, 14.1% and 21.1% of patients in the LeToHAIC group achieved complete response of all lesions and complete response of the intrahepatic target lesions per modified RECIST criteria, respectively. Grade 3/4 treatment-related adverse events that were more frequent in the LeToHAIC group than in the lenvatinib group included neutropenia (8.5% versus 1.2%), thrombocytopenia (5.6% versus 0), and nausea (5.6% versus 0). CONCLUSIONS: Lenvatinib, toripalimab, plus HAIC had acceptable toxic effects and might improve survival compared with lenvatinib alone in advanced hepatocellular carcinoma.

Comparison of HBV reactivation between patients with high HBV-DNA and low HBV-DNA loads undergoing PD-1 inhibitor and concurrent antiviral prophylaxis.

BACKGROUND: Programmed cell death protein-1 (PD-1) inhibitor is recommended to treat advanced hepatocellular carcinoma (HCC). However, the safety of PD-1 inhibitor in patients with high HBV-DNA load is unknown because of the potential risk of hepatitis B virus (HBV) reactivation. This study was to compare the HBV reactivation between patients with low HBV-DNA loads and high HBV-DNA loads undergoing antiviral prophylaxis and PD-1 inhibitor. METHODS: This was a retrospective study including consecutive hepatitis B surface antigen-positive HCC patients who received PD-1 inhibitor and concurrent antiviral prophylaxis for prevention of clinical hepatitis. Patients were divided into low HBV-DNA group (low group, ≤ 500 IU/ml) and high HBV-DNA group (high group, > 500 IU/ml) according to the baseline HBV-DNA level. The incidences of HBV reactivation, HBV-associated hepatitis, and PD-1 inhibitor disruption were compared between the two groups. RESULTS: Two hundred two eligible patients were included: 94 in the low group and 108 in the high group. Seven patients (5 in the low group and 2 in the high group) developed HBV reactivation, and all recovered from HBV reactivation and HBV-associated hepatitis. The incidence of HBV reactivation in the two groups was low (5.3% vs 1.9%, P = 0.34). There was also no difference in the incidence of HBV-associated hepatitis (P = 0.56), or PD-1 inhibitor disruption (P = 0.82). The multivariable analysis showed PD-1 inhibitor with hepatic arterial infusion chemotherapy was the only significant risk factor for HBV reactivation (P = 0.04) and hepatitis (P = 0.002). CONCLUSION: With concurrent antiviral prophylaxis, HBV-DNA load higher than 500 IU/ml should not be a contraindication for PD-1 inhibitor.

NAP1L1 is a prognostic biomarker and contribute to doxorubicin chemotherapy resistance in human hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most fatal cancers, and its molecular basis needs to be delineated to identify biomarkers for its potential treatment. The purpose of this study was to identify a novel gene, nucleosome assembly proteins 1-like 1 protein (NAP1L1), associated with aggressive phenotypes of HCC. METHODS: Immunohistochemical staining was used to detect NAP1L1 protein expression in HCC tissues. The prognostic value of NAP1L1 expression was determined using Kaplan-Meier analysis and the Cox proportional hazards model. CCK-8 and apoptosis assays were used to detect the chemosensitivity in vitro. Xenograft tumor models were used to evaluate tumor cell proliferation and chemosensitivity in vivo. RESULTS: NAP1L1 expression was significantly upregulated in tumor tissues as compared to adjacent non-tumor tissues. High NAP1L1 expression in HCC tissues was associated with aggressive clinicopathologic features, such as serum AFP levels, tumor size and tumor number. Patients with high NAP1L1 expression had poor overall survival in our cohort and in the extra-validation cohort analyzed by TCGA microarray dataset and was further identified as an independent prognostic factor in HCC patients treated with radical resection. Both in vitro and in vivo assays showed that NAP1L1 promoted HCC cell proliferation and contribute to chemotherapy resistance. Further analyses found that some certain stemness associated genes were decreased concurrently with NAP1L1 down-regulation in HCC cell lines. CONCLUSIONS: Our findings support that NAP1L1 is a prognostic biomarker and may contribute to chemotherapy resistance in human hepatocellular carcinoma.

Transarterial Chemoembolization related to Good Survival for Selected Patients with advanced Hepatocellular Carcinoma.

Background & aims: It remains controversial whether patients with advanced-stage hepatocellular carcinoma could be benefit from transarterial chemoembolization (TACE) treatment. The purpose of the present study is to identify predictors of survival following TACE in patients with advanced HCC. Methods: Overall, 303 patients with Barcelona Clinic Liver Cancer (BCLC) stage C HCC who were first treated with TACE from Sun Yat-sen University Cancer Centre between January 2009 and December 2013 were reviewed and enrolled in this study. We carried out Kaplan-Meier and Cox proportional hazard model analyses of prognostic factors. Results: The median survival of the whole cohort was 8.4 months. Multivariable Cox regression analyses confirmed that four risk factors, high serum levels of gamma-glutamyl transpeptidase (GGT), C-reactive protein (CRP), alkaline phosphatase (ALP) and presence of portal vein tumour thrombosis (PVTT), were independent prognostic factors for overall survival. The expected median survival among patients with 0-1 and 2-4 risk factors were 18.1 (95% CI: 15.5-20.7) and 6.8 (95% CI: 5.8-7.8) months, respectively. Objective tumor response among patients with 0-1 and 2-4 risk factors were 38.9% and 17.3%, respectively. Conclusion: We found four risk factors were associated with dismal overall survival for advanced HCC patients: serum GGT level, serum CRP, serum ALP and presence of PVTT. TACE may be recommended for patients with advanced HCC with 0-1 risk factors due to the favourable prognosis.

Aberrant MCT4 and GLUT1 expression is correlated with early recurrence and poor prognosis of hepatocellular carcinoma after hepatectomy.

The tumor microenvironment is a key determinant of cancer cell biology. The microenvironment is a complex mixture of tumor cells, stromal cells, and proteins, extracellular matrix, oxygen tension, and pH levels surrounding the cells that regulate the tumor progress. This study identified the prognostic factors associated with hepatocellular carcinoma (HCC) and MCT4 and GLUT1 expression levels in HCC specimens. In this study, we analyzed MCT4 and GLUT1 expression levels in tissue samples from 213 patients with HCC by immunohistochemical analyses and in HCC tumor tissues and matched adjacent nonneoplastic tissues by quantitative real-time PCR. We conducted a prognostic analysis of the overall survival (OS) and time to recurrence (TTR) using immunoreactivity and other common clinical and pathological parameters. All variables with prognostic impact were further analyzed by multivariate analysis. We found that MCT4 and GLUT1 expression levels were significantly higher in tumor tissues than in adjacent nontumor tissues, and they were positively correlated with tumor size. Survival analysis showed that patients with high expression levels of MCT4 or GLUT1 had a poor OS and TTR. In patients with HCC, MCT4 expression was an independent negative prognostic factor for OS (hazard ratio [HR] = 1.617; 95% confidence interval [CI] = 1.102-2.374; P = 0.014), and metabolic indicators were independent prognostic factors for OS (HR = 1.617, 95% CI = 1.102-2.374, P = 0.006) and TTR (HR = 1.348, 95% CI = 1.079-1.685, P = 0.009). Interestingly, patients with positive metabolic indicator expression in tumor cells had a significantly shorter OS and earlier TTR than those with negative metabolic indicator expression in tumor cells in the ≤5 cm and >5 cm subgroups. In summary, using the expression of MCT4 and GLUT1 and their metabolic parameters to determine the metabolic status of tumors is promising for predicting the prognosis of patients with HCC.

Comparison of Stable and Unstable Ethiodized Oil Emulsions for Transarterial Chemoembolization of Hepatocellular Carcinoma: Results of a Single-Center Double-Blind Prospective Randomized Controlled Trial.

PURPOSE: To compare the stability of stable and unstable water-in-oil emulsions and the efficacy and safety of these emulsions in a single-center, prospective double-blind trial of transarterial chemoembolization for hepatocellular carcinoma (HCC). MATERIALS AND METHODS: A total of 812 patients with inoperable HCC were randomized (stable emulsion, n = 402; unstable emulsion, n = 410). The 2 emulsions were prepared by using the same protocol except that different solvents were used for chemotherapy agents, including epirubicin, lobaplatin, and mitomycin C. The solvent in the stable emulsion arm was contrast medium and distilled water, and the solvent in the unstable emulsion arm was distilled water. The primary endpoint was overall survival (OS), and secondary endpoints were time to progression (TTP), tumor response, adverse events (AEs), and plasma epirubicin concentrations. RESULTS: In vitro, stable emulsions did not occur until 1 day, and unstable emulsions, with a lower peak plasma concentration (P = .001) in vivo, exhibited rapid separation of the oil and aqueous phases after 10 minutes. Median OS times in the stable and unstable emulsion arms were 17.7 and 19.2 months, respectively (P = .81). No differences were found in TTP, tumor response, and AEs except for myelosuppression (anemia, 3.5% vs 7.6%; thrombocytopenia, 11.5% vs 17.7%), which was significantly more severe and frequent in the unstable emulsion arm (P = .013). CONCLUSIONS: Chemoembolization is equally effective with the use of stable and unstable emulsions, but the use of a stable emulsion has the advantage of less myelosuppression and a favorable pharmacokinetic profile.

Matrix metalloproteinase 12 expression is associated with tumor FOXP3+ regulatory T cell infiltration and poor prognosis in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is among the most fatal types of cancer worldwide due to its high rates of recurrence and metastasis. The molecular processes involved in HCC progression require further investigation to identify biomarkers for use in diagnosis and treatment. In the present study, the significance and prognostic value of matrix metallopeptidase 12 (MMP12) expression in human HCC was investigated. MMP12 mRNA expression was investigated using reverse transcription-quantitative polymerase chain reaction analysis of 42 pairs of tumor and non-tumor liver tissues obtained from patients with HCC following surgical treatment. Immunohistochemical staining was used to detect MMP12 and forkhead box P3 (FOXP3) expression in 158 paraffin-embedded HCC tissues. The prognostic value of MMP12 expression was determined using Kaplan-Meier analysis and the Cox proportional hazards model. MMP12 mRNA levels were significantly higher in liver tumor tissues compared with matched non-tumor liver tissues. MMP12 expression and FOXP3+ regulatory T cell (Treg) infiltration was positively correlated (r=0.302; P<0.001). MMP12 protein overexpression was positively correlated with tumor size (P=0.018), high serum alpha-fetoprotein levels (P=0.005) and poor overall survival time (P=0.012) in patients with HCC. Furthermore, MMP12 protein level was an independent predictive factor for overall survival time of patients with HCC who underwent curative resection. In conclusion, these results suggest that MMP12 may increase FOXP3+ Treg infiltration into tumor tissues, and promote tumor progression and immune evasion of HCC. The overexpression of MMP12 protein is, therefore, a valuable prognostic indicator in patients with HCC.

Prognostic significance of sodium-potassium ATPase regulator, FXYD3, in human hepatocellular carcinoma.

The clinical significance of the sodium-potassium ATPase regulator FXYD domain-containing ion transport regulator 3 (FXYD3) has been demonstrated in a number of types of cancer. However, the role of this protein in human hepatocellular carcinoma (HCC) remains to be elucidated. In the present study, 217 HCC tissue samples were analyzed to evaluate the expression and prognostic significance of FXYD3 in HCC. Reverse transcription-quantitative polymerase chain reaction was used to analyze the mRNA expression of FXYD3 in 80 primary HCC specimens and paired non-cancerous liver tissue samples, while western blotting was used to analyze the protein expression level of FXYD3 in another 24 pairs. These analyses demonstrated that the expression level of FXYD3 was significantly increasedb at the mRNA and protein levels in HCC tumor tissues compared with adjacent non-cancerous tissues. Immunohistochemical analysis of 137 paraffin-embedded HCC tissue samples indicated that the expression of FXYD3 was associated with HCC clinicopathological characteristics. Kaplan-Meier analysis demonstrated that patients with high FXYD3 protein expression (n=60) experienced significantly poorer overall survival time compared with patients with low FXYD3 protein expression (n=77) (P<0.001). Multivariate analysis demonstrated that FYXD3 protein expression level (hazard ratio, 2.137; 95% confidence interval, 1.224-3.732; P=0.008) was an independent prognostic factor in patients with HCC. Overall, the results indicated that FXYD3 expression levels were higher in HCC tumor tissues than in adjacent non-cancerous tissues, and that the FXYD3 protein may serve as a prognostic marker for HCC.

Phase II Study of Sorafenib Combined with Concurrent Hepatic Arterial Infusion of Oxaliplatin, 5-Fluorouracil and Leucovorin for Unresectable Hepatocellular Carcinoma with Major Portal Vein Thrombosis.

BACKGROUND: Sorafenib is recommended for the first-line treatment of advanced hepatocellular carcinoma (HCC). However, the median progression-free survival (PFS) of patients with HCC and major portal vein tumor thrombosis treated with sorafenib monotherapy is no more than 3 months. A prospective single-arm phase II study was conducted to determine whether adding hepatic arterial infusion chemotherapy of oxaliplatin, 5-fluorouracil and leucovorin to sorafenib could improve on these results. METHODS: Thirty five patients were treated with sorafenib 400 mg orally twice a day, oxaliplatin 85 mg/m2 HAI on day 1, leucovorin 400 mg/m2 HAI on days 1, and 5-fluorouracil 2800 mg/m2 on days 1 and 2, repeated every 21 days. The primary end point was the 3-month PFS rate. RESULTS: The 3-, 6-, and 12-month PFS rates were 82.9, 51.4, and 22.9%, respectively. The median PFS and overall survival was 6.7 and 13.2 months, respectively. The objective response rate was 40%, and the disease control rate was 77.1% by RECIST criteria. Five (14.3%) patients achieved conversion to complete resection after the study treatment, and one of them experienced a pathological complete response. Treatment-related deaths did not occur. Grade 3-4 toxicities consisted of increases in aspartate aminotransferase (31.4%), hand-foot syndrome (17.1%), thrombocytopenia (14.3%), and neutropenia (8.6%). CONCLUSIONS: The combination treatment met the pre-specified end point of a 3-month progression free survival rate exceeding 65% and was clinical tolerable. The merits of this approach need to be established with a phase III trial. Clinical trial number http://ClinicalTrials.gov (No. NCT02981498).

Hepatic artery infusion chemotherapy using mFOLFOX versus transarterial chemoembolization for massive unresectable hepatocellular carcinoma: a prospective non-randomized study.

BACKGROUND: Transarterial chemoembolization (TACE) is recommended as the standard care for unresectable hepatocellular carcinoma (HCC) at Barcelona Clinic Liver Cancer (BCLC) stage A-B. However, the efficacy of TACE on large (≥ 10 cm) stage A-B HCC is far from satisfactory, and it is proposed that hepatic artery infusion chemotherapy (HAIC) might be a better first-line treatment of this disease. Hence, we compared the safety and efficacy of HAIC with the modified FOLFOX (mFOLFOX) regimen and those of TACE in patients with massive unresectable HCC. METHODS: A prospective, non-randomized, phase II study was conducted on patients with massive unresectable HCC. The protocol involved HAIC with the mFOLFOX regimen (oxaliplatin, 85 mg/m2 intra-arterial infusion; leucovorin, 400 mg/m2 intra-arterial infusion; and fluorouracil, 400 mg/m2 bolus infusion and 2400 mg/m2 continuous infusion) every 3 weeks and TACE with 50 mg of epirubicin, 50 mg of lobaplatin, 6 mg of mitomycin, and lipiodol and polyvinyl alcohol particles. The tumor responses, time-to-progression (TTP), and safety were assessed. RESULTS: A total of 79 patients were recruited for this study: 38 in the HAIC group and 41 in the TACE group. The HAIC group exhibited higher partial response and disease control rates than did the TACE group (52.6% vs. 9.8%, P < 0.001; 83.8% vs. 52.5%, P = 0.004). The median TTPs for the HAIC and TACE groups were 5.87 and 3.6 months (hazard radio [HR] = 2.35, 95% confidence interval [CI] = 1.16-4.76, P = 0.015). More patients in the HAIC group than in the TACE group underwent resection (10 vs. 3, P = 0.033). The proportions of grade 3-4 adverse events (AE) and serious adverse events (SAE) were lower in the HAIC group than in the TACE group (grade 3-4 AEs: 13 vs. 27, P = 0.007; SAEs: 6 vs. 15, P = 0.044). More patients in the TACE group than in the HAIC group had the study treatment terminated early due to intolerable treatment-related adverse events or the withdrawal of consent (10 vs. 2, P = 0.026). CONCLUSIONS: HAIC with mFOLFOX yielded significantly better treatment responses and less serious toxicity than did TACE. HAIC might represent a feasible and promising first-line treatment for patients with massive unresectable HCC.