Guidelines for Diagnosis and Treatment of Hepatocellular Carcinoma with Portal Vein Tumor Thrombus in China (2021 Edition).

Portal vein tumor thrombus (PVTT) is very common and it plays a major role in the prognosis and clinical staging of hepatocellular carcinoma (HCC). We have published the first version of the guideline in 2016 and revised in 2018. Over the past several years, many new evidences for the treatment of PVTT become available, especially for the advent of new targeted drugs and immune checkpoint inhibitors which have further improved the prognosis of PVTT. So, the Chinese Association of Liver Cancer and Chinese Medical Doctor Association revised the 2018 version of the guideline to adapt to the development of PVTT treatment. Future treatment strategies for HCC with PVTT in China would depend on new evidences from more future clinical trials.

Tumor-derived extracellular vesicles containing long noncoding RNA PART1 exert oncogenic effect in hepatocellular carcinoma by polarizing macrophages into M2.

AIM: We explored whether tumor-derived extracellular vesicles (EVs) could deliver long noncoding RNA (lncRNA) PART1 into macrophage to orchestrate macrophage polarization in the progression of hepatocellular carcinoma (HCC). METHOD: The expression patterns of PART1, microRNA (miR)-372-3p and TLR4 were detected by RT-qPCR in the HCC tissues and HCC cells. PART1 was silenced or overexpressed in HCC cells to assess its effects on the HCC cell process. EVs were isolated from PART1-overexpressed HCC cells, and co-cultured with macrophages, and gain- and loss-of-function assays were implemented in macrophages to evaluate their role in macrophage polarization. Relationship among PART1, miR-372-3p, and TLR4 was evaluated. Effect of EV-PART1 on tumorigenicity in vivo was detected by subcutaneous tumorigenicity test in nude mice. RESULT: PART1 and TLR4 were upregulated while miR-372-3p was downregulated in HCC tissues and cells. PART1 increased HCC cell proliferation, migration, invasion, and EMT. Mechanistically, PART1 bound to miR-372-3p to downregulate its expression, whereas TLR4 was negatively targeted by miR-372-3p in the macrophages. EVs containing PART1, TLR4 overexpression, or miR-372-3p inhibition induced M2 polarization of macrophages. Also, EVs containing PART1 promoted M2 polarization of macrophages and the occurrence of HCC by affecting miR-372-3p/TLR4 axis. CONCLUSION: HCC cell-derived EVs might up-regulate TLR4 by inhibiting miR-372-3p via PART1 delivery to promote macrophage M2 polarization in HCC.

Chinese Expert Consensus on Immunotherapy for Hepatocellular Carcinoma (2021 Edition).

Background: Hepatocellular carcinoma (HCC) is one of the most common malignancies in China. Most HCC patients are first diagnosed at an advanced stage, and systemic treatments are the mainstay of treatment. Summary: In recent years, immune checkpoint inhibitors have made a breakthrough in the systemic treatment of middle-advanced HCC, breaking the single therapeutic pattern of molecular-targeted agents. To better guide the clinical treatment for effective and safe use of immunotherapeutic drugs, the Chinese Association of Liver Cancer and Chinese Medical Doctor Association has gathered multidisciplinary experts and scholars in relevant fields to formulate the "Chinese Clinical Expert Consensus on Immunotherapy for Hepatocellular Carcinoma (2021)" based on current clinical studies and clinical medication experience for reference in China. Key Messages: The consensus contained 17 recommendations, including the preferred regimen for first- and second-line immunotherapy, evaluation and monitoring before/during/after treatment, management of complications, precautions for special patients, and potential population for immunotherapy.

Apatinib as second-line or later therapy in patients with advanced hepatocellular carcinoma (AHELP): a multicentre, double-blind, randomised, placebo-controlled, phase 3 trial.

BACKGROUND: Inhibition of vascular endothelial growth factor receptor (VEGFR) has shown antitumour activity in advanced hepatocellular carcinoma, but few studies of VEGFR inhibitors have been done in populations with a high prevalence of hepatitis B virus infection. The aim of this study was to evaluate the efficacy and safety of apatinib in patients with pretreated advanced hepatocellular carcinoma. METHODS: AHELP was a randomised, double-blind, placebo-controlled, phase 3 trial done at 31 hospitals in China, in patients (aged ≥18 years) with advanced hepatocellular carcinoma who had previously been refractory or intolerant to at least one line of systemic chemotherapy or targeted therapy. Patients were randomly assigned (2:1) to receive apatinib 750 mg or placebo orally once daily in 28-day treatment cycles. Group allocation was done with a central randomisation system, with a block size of six, and was stratified by Eastern Cooperative Oncology Group performance status, previous sorafenib treatment, and presence of vascular invasion or extrahepatic metastasis. The primary endpoint was overall survival, which was defined as time from randomisation to death from any cause, and was analysed in patients who were randomly assigned and received at least one dose of the study drug. Safety analyses were done in patients who received at least one dose of the study treatment and had post-dose safety assessments. This trial is registered with ClinicalTrials.gov, NCT02329860. FINDINGS: Between April 1, 2014, and May 3, 2017, 400 eligible patients were randomly assigned to receive apatinib (n=267) or placebo (n=133). Seven patients (six in the apatinib group and one in the placebo group) did not receive study treatment and were excluded from efficacy analyses. Overall survival was significantly improved in the apatinib group compared with the placebo group (median 8·7 months [95% CI 7·5‒9·8] vs 6·8 months [5·7‒9·1]; hazard ratio 0·785 [95% CI 0·617‒0·998], p=0·048). 387 patients (257 in the apatinib group and 130 in the placebo group) had a safety assessment after study treatment and were included in safety analyses. The most common treatment-related adverse events of grade 3 or 4 were hypertension (71 [28%] patients in the apatinib group vs three [2%] in the placebo group), hand-foot syndrome (46 [18%] vs none), and decreased platelet count (34 [13%] vs one [1%]). 24 (9%) patients in the apatinib group and 13 (10%) in the placebo group died due to adverse events, but none of these deaths were deemed to be related to treatment by investigators. INTERPRETATION: Apatinib significantly improved overall survival in patients with pretreated advanced hepatocellular carcinoma compared with placebo, with a manageable safety profile. FUNDING: Jiangsu Hengrui Medicine.

Identification of key miRNAs in the progression of hepatocellular carcinoma using an integrated bioinformatics approach.

BACKGROUD: It has been shown that aberrant expression of microRNAs (miRNAs) and transcriptional factors (TFs) is tightly associated with the development of HCC. Therefore, in order to further understand the pathogenesis of HCC, it is necessary to systematically study the relationship between the expression of miRNAs, TF and genes. In this study, we aim to identify the potential transcriptomic markers of HCC through analyzing common microarray datasets, and further establish the differential co-expression network of miRNAs-TF-mRNA to screen for key miRNAs as candidate diagnostic markers for HCC. METHOD: We first downloaded the mRNA and miRNA expression profiles of liver cancer from the GEO database. After pretreatment, we used a linear model to screen for differentially expressed genes (DEGs) and miRNAs. Further, we used weighed gene co-expression network analysis (WGCNA) to construct the differential gene co-expression network for these DEGs. Next, we identified mRNA modules significantly related to tumorigenesis in this network, and evaluated the relationship between mRNAs and TFs by TFBtools. Finally, the key miRNA was screened out in the mRNA-TF-miRNA ternary network constructed based on the target TF of differentially expressed miRNAs, and was further verified with external data set. RESULTS: A total of 465 DEGs and 215 differentially expressed miRNAs were identified through differential genes expression analysis, and WGCNA was used to establish a co-expression network of DEGs. One module that closely related to tumorigenesis was obtained, including 33 genes. Next, a ternary network was constructed by selecting 256 pairs of mRNA-TF pairs and 100 pairs of miRNA-TF pairs. Network mining revealed that there were significant interactions between 18 mRNAs and 25 miRNAs. Finally, we used another independent data set to verify that miRNA hsa-mir-106b and hsa-mir-195 are good classifiers of HCC and might play key roles in the progression of HCC. CONCLUSION: Our data indicated that two miRNAs-hsa-mir-106b and hsa-mir-195-are identified as good classifiers of HCC.

Up-regulated microRNA-33b inhibits epithelial-mesenchymal transition in gallbladder cancer through down-regulating CROCC.

Gallbladder cancer (GBC) is a relatively rare but fatal gastrointestinal tumor. The microRNA-33b (miR-33b), a member of miR-33 family, is reported to function as a tumor suppressor in various cancers. Notably, miR-33 was predicted to target CROCC based on microarray-based analysis. Hereby, we aimed to characterize the effect of miR-33b on epithelial-mesenchymal transition (EMT) in GBC and the potential mechanism involved with the regulation of CROCC. In GBC cell lines, miR-33b expressed at low levels, and CROCC expressed at high levels, with enhanced EMT process. To further examine the specific mechanism of miR-33b and CROCC in GBC, the GBC cells were treated with the miR-33b mimic/inhibitor or siRNA-CROCC to assess the expression alteration of EMT-related genes and cell proliferation, migration, and invasion. MiR-33b was verified to target and down-regulate the expression of CROCC. The miR-33b up-regulation or CROCC silencing was observed to increase the level of E-cadherin but decrease the levels of N-cadherin and Vimentin, corresponding to impeded cell proliferation, migration, invasion, EMT, and tumor growth. The findings suggest that miR-33b up-regulation hinders GBC development through down-regulating CROCC, which was achieved by inhibition of EMT. The present study may provide an insight on a novel target for GBC treatment.

Retrospective analysis of sorafenib efficacy and safety in Chinese patients with high recurrence rate of post-hepatic carcinectomy.

Background: There is no guideline recommendation for preventing hepatocellular carcinoma (HCC) recurrence after hepatic resection. Moreover, an unmet need exists on the effectiveness of sorafenib therapy in recurrent HCC. Purpose: We therefore assessed the efficacy and safety of sorafenib in Chinese HCC patients with high risk of recurrence. Patients and methods: Data were collected retrospectively from 15 Chinese research centers from January 1, 2012 to November 15, 2013, by chart reviews of patients with moderate-advanced HCC who received hepatic carcinectomy. The primary end point was recurrence-free survival rate at 1 year in patients with a high recurrence risk. Secondary end points included 1-year survival rate, time to recurrence and safety assessment. Results: A total of 209 high-risk patients (sorafenib, n=98; control, n=111) who underwent carcinectomy were analyzed. There was no significant difference in the proportion of patients with recurrence-free survival at 1 year between the sorafenib and control (70.43% vs 68.90%: χ2=0.007, P=0.934). One-year survival rate was significantly higher with sorafenib than observed with control (95.5% vs 83.35%; χ2=7.441, P=0.006). Time to recurrence between sorafenib and control groups was similar. Incidences of all the adverse events (AEs) were similar in both the groups and transaminase elevation was most common in both groups (20.37% vs 24.79%). Thrombocytopenia incidence was significantly lower with the sorafenib group than with control (1.85% vs 9.40%; P=0.015). Conclusion: Sorafenib may be considered as a feasible option in the treatment of HCC recurrence.

MicroRNA-1305 Inhibits the Stemness of LCSCs and Tumorigenesis by Repressing the UBE2T-Dependent Akt-Signaling Pathway.

MicroRNAs (miRNAs) are involved in the maintenance of the cancer stem cell (CSC) phenotype by binding to genes and proteins that modulate cell proliferation and/or cell apoptosis. In our study, we aimed to investigate the role of miR-1305 in the proliferation and self-renewal of liver CSCs (LCSCs) via the ubiquitin-conjugating enzyme E2T (UBE2T)-mediated Akt-signaling pathway. Differentially expressed genes in human hepatocellular carcinoma (HCC) were obtained by in silico analysis. The relationship between miR-1305 and UBE2T was verified by dual luciferase reporter gene assay. qRT-PCR and western blot analysis were performed to determine the expression of UBE2T, the Akt-signaling pathway, and stemness-related factors in LCSCs. In addition, miR-1305 disrupted the activation of the Akt-signaling pathway by targeting UBE2T, and, ultimately, it repressed the sphere formation, colony formation, and proliferation, as well as tumorigenicity of LCSCs. In summary, miR-1305 targeted UBE2T to inhibit the Akt-signaling pathway, thereby suppressing the self-renewal and tumorigenicity of LCSCs. Those findings may provide an enhanced understanding of miR-1305 as a therapeutic target to limit the progression of LCSCs.

miR­21 inhibitor facilitates the anticancer activity of doxorubicin loaded nanometer in melanoma.

MicroRNA­21 (miR­21) is a potential therapeutic target for melanoma. Whether miR­21 inhibitor affects the anti­cancer activity of doxorubicin assisted by c(RGDyK)­modified liposome (DLN) in melanoma and the underlying mechanisms are largely unknown. In this study, in vitro and animal models were used to explore the effect of DLN combined with miR­21 inhibitor on melanoma cells. The data demonstrated that treatment with 5 µl DLN (final concentration of doxorubicin 5 mg/ml) for 72 h effectively inhibited melanoma cell growth (~75% inhibition). The experiments were then divided into five groups: Control group, vector group, DLN group, miR­21 inhibitor group and miR­21 inhibitor + DLN group. Compared with the control group, DLN (5 µl) or miR­21 inhibitor significantly reduced migration and invasion of melanoma cells, promoted apoptosis and arrested cells at the G1 phase. Notably, the combined application of DLN with miR­21 inhibitor further promoted the anti­cancer effects (reducing migration and invasion of melanoma cells, promoting apoptosis and arresting cells at G1 phase) compared with individual application of DLN or miR­21 inhibitor. Mechanically, DLN did not function by reducing miR­21 expression, whereas DLN and miR­21 inhibitor downregulated B­cell lymphoma-2 (BCL­2) expression, and facilitated BCL­2­associated X protein (Bax) and P53 expression in melanoma cells. DLN and miR­21 inhibitor together displayed stronger effects on Bcl­2, Bax and P53 expression that each alone. In vivo data further demonstrated that DLN inhibited tumor growth further than a similar dose of doxorubicin only. Furthermore, miR­21 inhibitor and DLN exerted the optimal anti­cancer effect compared with single application of DLN or miR­21 inhibitor. Together, the findings demonstrated miR­21 inhibitor facilitated the anti­cancer activity of DLN in melanoma, and the mechanisms involved Bcl­2, Bax and P53 expression.

c[RGDyk]-coated liposomes loaded with adriamycin and miR-21 mimics inhibit the growth of hepatoma cell line SMCC-7721 via up-regulating Bax and p53.

BACKGROUND: This study was conducted to investigate the effects of c[RGDyk]-coated liposomes loaded with Adriamycin (nanodrug) and miR-21 mimics on hepatoma cell line SMCC-7721. METHODS: SMCC-7721 cells were divided into five groups: control (receiving no treatment), nanodrug, miR-21 mimics + nanodrug and miR-21 mimics and empty vector. The concentration and duration of treatments were determined using the MTT assay. Cell apoptosis was detected using flow cytometer. The expression of Bax, Bcl-2 and p53 was measured using qPCR and Western blot analysis. RESULTS: MTT showed that the nanodrug inhibited cell proliferation. Nanodrug and miR-21 led to cell arrest at S phase and apoptosis. qPCR showed that cells treated with nanodrug and miR-21 increased the expression of Bax and p53. Western blot analysis indicated that Bcl-2 expression was significantly reduced. CONCLUSIONS: Our work demonstrates that nanodrug and miR-21 have inhibitory effect on SMCC-7721 cells via up-regulating Bax and p53.

A novel missense mutation in the SLC6A19 gene in a Chinese family with Hartnup disorder.

BACKGROUND: Hartnup disease is a rare autosomal-recessive abnormality of renal and gastrointestinal neutral amino acid transport associated with neurologic, psychiatric, and dermatologic symptoms. Mutations in the SLC6A19 gene have been proposed to be responsible for the underlying changes in this disorder. AIM: To investigate a pedigree with Hartnup disorder and to search for the mutation in the SLC6A19 gene in this pedigree. METHODS: The encoding exons of the SLC6A19 gene were amplified and sequenced from genomic DNA samples. Amino acids were determined in urine samples from the proband and her family members. RESULTS: The proband and her brother had a homozygous mutation of c.850G > A in the SLC6A19 gene, causing G284R in the transmembrane domain of the SLC6A19 transporter, inherited from their parents who were heterozygous carriers. Their urine samples showed increased values of eight neutral amino acids. CONCLUSION: We found a novel homozygous mutation of G284R in the transmembrane domain of the SLC6A19 transporter in the proband, with typical dermatologic and neurologic manifestations and increased levels of urinary neutral amino acids.