TFAIP6 facilitates hepatocellular carcinoma cell glycolysis through upregulating c-myc/PKM2 axis.

Background: Hepatocellular carcinoma (HCC) is the most prevalent liver cancer. Despite of the improvement of therapies, the durable response rate and survival benefit are still limited for HCC patients. It's urgent to clarify the molecular mechanisms and find therapeutic strategies to improve the clinical outcome. TNFα-stimulated gene-6 (TNFAIP6) plays a critical role in the prognosis of various tumors, but its roles in HCC are still unclear. Methods: Quantitative real-time PCR (qRT-PCR) and immunohistochemistry (IHC) analysis were employed to evaluate the clinical relevance of TNFAIP6 expressions in HCC patients. Cell counting kit-8 (CCK-8), Edu assay, and transwell assay were performed to evaluate the malignancy of HCC cells. Glucose uptake, lactate production, ATP production, extracellular acidification rate (ECAR) by Seahorse XF analyzer were employed to evaluate the role of TNFAIP6 in the regulation of aerobic glycolysis. The expressions of key proteins involved in glycolysis were examined by Western blot. Co-immunoprecipitation (Co-IP) and chromatin immunoprecipitation (ChIP) were used for protein-protein interactions or protein-RNA interactions respectively. Knockdown and overexpression of TNFAIP6 in HCC cells were employed for analyzing the functions of TNFAIP6 in HCC. Results: TNFAIP6 was significantly upregulated in HCC and predicted a poor clinical prognosis. Knockdown of TNFAIP6 inhibited in vitro cell proliferation, invasion, migration, as well as glycolysis in HCC cells. Mechanistically, we clarified that TNFAIP6 interacted with heterogeneous nuclear ribonucleoprotein C (HNRNPC), stabilized c-Myc mRNA and upregulated pyruvate kinase M2 (PKM2) to promote glycolysis. Conclusions: Our study reveals a molecular mechanism by which TNFAIP6 promotes aerobic glycolysis, which is beneficial for malignance of HCC and provides a potential clinical therapy for disease management.

Gastrointestinal perforation associated with bevacizumab in metastatic colorectal cancer.

OBJECTIVE: To investigate the risk factors for gastrointestinal perforation in metastatic colorectal cancer patients receiving bevacizumab. METHODS: We retrospectively reviewed 217 patients with metastatic colorectal cancer receiving bevacizumab to investigate the risk factors for gastrointestinal perforation. Three patients occurred intestinal perforation after receiving bevacizumab. We analyzed the clinical characteristics of three patients with intestinal perforation. RESULTS: All patients receiving bevacizumab. Three of 217 patients occurred intestinal perforation after receiving bevacizumab. Patient no. 1 was 70 years old, female, having history of intestinal obstruction. The patient occurred intestinal perforation and ultimately died after receiving bevacizumab. Patient no. 2 was 59 years old, female, having history of intestinal obstruction. The patient occurred intestinal perforation after receiving bevacizumab, and recovered smoothly after symptomatic treatment. Patient no. 3 was 60 years old, female, having history of intestinal obstruction. The patient occurred intestinal perforation and ultimately died after receiving bevacizumab. CONCLUSIONS: Patients with advanced colorectal cancer receiving bevacizumab are at risk of gastrointestinal perforation. The patient's age, gender and history of bowel obstruction may be associated with gastrointestinal perforation.

The surgical outcomes and risk factors of giant hepatic haemangiomas: a single centre experience.

OBJECTIVE: To evaluate the safety of performing surgery on cavernous haemangiomas in the liver larger than 10 cm and establish preoperative predictors of intraoperative blood transfusion and morbidity. METHODS: A total of 373 patients with haemangiomas larger than 10 cm who underwent surgery in our hospital were retrospectively analysed. According to tumour diameter, the patients were divided into a giant haemangioma (GH) group (241 cases) (10 cm ≤ diameter < 15 cm) and an enormous haemangioma (EH) group (132 cases) (diameter ≥ 15 cm). Clinical parameters were then compared between the two groups. RESULTS: Compared with the GH group, the EH group had higher rates of leukopenia (10.6% vs. 4.5%), anaemia (26.5% vs. 15.7%), and thrombocytopenia (13.6% vs. 6.2%). The occlusion time in the EH group was longer than that in the GH group (26.33 ± 14.10 min vs. 31.85 ± 20.09 min, P < 0.01). The blood loss and blood transfusion in the EH group were greater than those in the GH group (P < 0.05). Moreover, the morbidity in the EH group was higher than that in the GH group (17.4% vs. 9.13%, P < 0.05). According to the results of the multivariable analysis, the operation time and size of the haemangioma may be independent risk factors for blood transfusion (P < 0.05). Additionally, the size of the haemangioma may be an independent risk factor associated with complications (P < 0.05). CONCLUSION: Enormous haemangioma is more likely to cause haematologic abnormalities than giant hepatic haemangioma. The risks of the operation and postoperative complications of enormous haemangioma are higher than those of giant hepatic haemangioma.

Adjuvant stereotactic body radiotherapy after marginal resection for hepatocellular carcinoma with microvascular invasion: A randomised controlled trial.

BACKGROUND: Marginal resection frequently occurred in hepatectomy for hepatocellular carcinoma (HCC), leading to increased local recurrence, especially among patients with microvascular invasion (MVI). Stereotactic body radiotherapy (SBRT) showed effectiveness in controlling tumour and tumour thrombosis. This study aimed to investigate the efficacy of SBRT, targeting on suboptimal resection margin, as adjuvant setting in MVI-positive HCC. METHODS: This was a single-centre randomised controlled trial conducted in Eastern Hepatobiliary Surgery Hospital, Shanghai, China. Participants with MVI-positive HCC receiving marginal resection were randomly assigned to the postoperative adjuvant SBRT or surgery alone (SA) group. SBRT was delivered by the CyberKnife® system with marker tracking devices, targeting on resection margin one month after surgery. The disease-free survival (DFS) and overall survival (OS) were compared between the groups, and the adverse events (AEs) were monitored. This trial was registered on ClinicalTrials.gov, NCT04891874. FINDINGS: A total of 76 participants were enrolled, with 38 in each group. The one-, three-, and five-year DFS rates were 92.1%, 65.8%, and 56.1% in SBRT group versus 76.3%, 36.8%, and 26.3% in SA group, respectively (p = 0.005). The one-, three-, and five-year OS rates were 100%, 89.5%, and 75.0% in SBRT group versus 100.0%, 68.4%, and 53.7% in SA group, respectively (p = 0.053). The total dose of SBRT for single participant was 35 Gy, and the biological effective dose (BED) was 59.5 Gy. The overall incidence of radiotherapy-related AE was 31.6% (12/38), and no grade 3 or higher grade AE was developed. INTERPRETATION: SBRT on the resection margin provides a safe therapeutic modality of adjuvant setting in MVI-positive HCC with suboptimal resection margin. It prevents local recurrence and improves DFS. TRIAL REGISTRATION NUMBER: NCT04891874.

Impact of metformin use on risk and mortality of hepatocellular carcinoma in diabetes mellitus.

BACKGROUND: The views regarding the associations between metformin use and hepatocellular carcinoma (HCC) among diabetes mellitus (DM) patients are divisive. Thus we summarized all available published studies evaluating the relationship between metformin therapy and HCC survival and risk, and aim to conduct an updated meta-analysis study to more accurately clarify the association. METHODS: We searched for articles regarding impact of metformin use on risk and mortality of HCC in DM and published before April 2021 in databases (PubMed and Web of Science). We used STATA 12.0 software to compute odds ratios (ORs)/relative risks (RRs) or hazard ratios (HRs) and their 95% confidence intervals (CIs) to generate a computed effect size and 95% CI. RESULTS: The present study showed that metformin use was associated with a decreased risk of HCC in DM with a random effects model (OR/RR = 0.59, 95% CI 0.51-0.68, I2 = 96.5%, p < 0.001). In addition, the study indicated that metformin use was associated with a decreased all-cause mortality of HCC in DM with a random effects model (HR = 0.74, 95% CI 0.66-0.83, I2 = 49.6%, p = 0.037). CONCLUSION: In conclusion, our studies support that the use of metformin in DM patients is significantly associated with reduced risk and all-cause mortality of HCC. And more prospective studies focusing on the metformin therapy as a protective factor for HCC are needed to verify the accuracy of the findings.

Comprehensive analysis of spatial architecture in primary liver cancer.

Heterogeneity is the major challenge for cancer prevention and therapy. Here, we first constructed high-resolution spatial transcriptomes of primary liver cancers (PLCs) containing 84,823 spots within 21 tissues from seven patients. The progressive comparison of spatial tumor microenvironment (TME) characteristics from nontumor to leading-edge to tumor regions revealed that the tumor capsule potentially affects intratumor spatial cluster continuity, transcriptome diversity, and immune cell infiltration. Locally, we found that the bidirectional ligand-receptor interactions at the 100-µm-wide cluster-cluster boundary contribute to maintaining intratumor architecture and the PROM1+ and CD47+ cancer stem cell niches are related to TME remodeling and tumor metastasis. Last, we proposed a TLS-50 signature to accurately locate tertiary lymphoid structures (TLSs) spatially and unveiled that the distinct composition of TLSs is shaped by their distance to tumor cells. Our study provides previous unknown insights into the diverse tumor ecosystem of PLCs and has potential benefits for cancer intervention.

A combined Cox and logistic model provides accurate predictive performance in estimation of time-dependent probabilities for recurrence of intrahepatic cholangiocarcinoma after resection.

BACKGROUND: Intrahepatic cholangiocarcinoma has heterogeneous outcomes after resection. There remains a need for broadly applicable recurrence-specific tool offering precise evaluation on curativeness of resection. METHODS: A four hospital-based clinical cohort involving 1,655 patients with intrahepatic cholangiocarcinoma who received surgical resection were studied. Cox and logistic models were networked into one system containing risk categories with distinctive probabilities of recurrence. Prediction of time-to-recurrence was performed by formulizing time-dependent risk probabilities. The model was validated in three clinical cohorts (n=332). RESULTS: From the training cohort, 10 and 11 covariates, including diabetes, cholelithiasis, albumin, platelet count, alpha fetoprotein, carbohydrate antigen 19-9, carcinoembryonic antigen, hepatitis B virus infection, tumor size and number, resection type, and lymph node metastasis, from Cox and logistic models were identified significant for recurrence-free survival (RFS). The combined Cox & logistic ranking system (CCLRS)-adjusted time-dependent probabilities were categorized into seven ranks (5-yr RFS for lowest and highest ranks were 75% vs. 0%; hazard ratio 18.5, 95% CI: 14.7-24.9, P<0.0001). The CCLRS was validated with a minimum area under curve value of 0.8086. Prediction of time-to-recurrence was validated to be excellent (Pearson r, 0.8204; P<0.0001). CONCLUSIONS: The CCLRS allows precise estimation on risk of recurrence for intrahepatic cholangiocarcinoma after resection. It could be applicative when estimating time-dependent disease status and stratifying individuals who sole resection of the tumor would not be curative.

Discovery of a Carbamoyl Phosphate Synthetase 1-Deficient HCC Subtype With Therapeutic Potential Through Integrative Genomic and Experimental Analysis.

BACKGROUND AND AIMS: Metabolic reprogramming plays an important role in tumorigenesis. However, the metabolic types of different tumors are diverse and lack in-depth study. Here, through analysis of big databases and clinical samples, we identified a carbamoyl phosphate synthetase 1 (CPS1)-deficient hepatocellular carcinoma (HCC) subtype, explored tumorigenesis mechanism of this HCC subtype, and aimed to investigate metabolic reprogramming as a target for HCC prevention. APPROACH AND RESULTS: A pan-cancer study involving differentially expressed metabolic genes of 7,764 tumor samples in 16 cancer types provided by The Cancer Genome Atlas (TCGA) demonstrated that urea cycle (UC) was liver-specific and was down-regulated in HCC. A large-scale gene expression data analysis including 2,596 HCC cases in 7 HCC cohorts from Database of HCC Expression Atlas and 17,444 HCC cases from in-house hepatectomy cohort identified a specific CPS1-deficent HCC subtype with poor clinical prognosis. In vitro and in vivo validation confirmed the crucial role of CPS1 in HCC. Liquid chromatography-mass spectrometry assay and Seahorse analysis revealed that UC disorder (UCD) led to the deceleration of the tricarboxylic acid cycle, whereas excess ammonia caused by CPS1 deficiency activated fatty acid oxidation (FAO) through phosphorylated adenosine monophosphate-activated protein kinase. Mechanistically, FAO provided sufficient ATP for cell proliferation and enhanced chemoresistance of HCC cells by activating forkhead box protein M1. Subcutaneous xenograft tumor models and patient-derived organoids were employed to identify that blocking FAO by etomoxir may provide therapeutic benefit to HCC patients with CPS1 deficiency. CONCLUSIONS: In conclusion, our results prove a direct link between UCD and cancer stemness in HCC, define a CPS1-deficient HCC subtype through big-data mining, and provide insights for therapeutics for this type of HCC through targeting FAO.

Single-Cell Transcriptome Analysis Uncovers Intratumoral Heterogeneity and Underlying Mechanisms for Drug Resistance in Hepatobiliary Tumor Organoids.

Molecular heterogeneity of hepatobiliary tumor including intertumoral and intratumoral disparity always leads to drug resistance. Here, seven hepatobiliary tumor organoids are generated to explore heterogeneity and evolution via single-cell RNA sequencing. HCC272 with high status of epithelia-mesenchymal transition proves broad-spectrum drug resistance. By examining the expression pattern of cancer stem cells markers (e.g., PROM1, CD44, and EPCAM), it is found that CD44 positive population may render drug resistance in HCC272. UMAP and pseudo-time analysis identify the intratumoral heterogeneity and distinct evolutionary trajectories, of which catenin beta-1 (CTNNB1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and nuclear paraspeckle assembly transcript 1 (NEAT1) advantage expression clusters are commonly shared across hepatobiliary organoids. CellphoneDB analysis further implies that metabolism advantage organoids with enrichment of hypoxia signal upregulate NEAT1 expression in CD44 subgroup and mediate drug resistance that relies on Jak-STAT pathway. Moreover, metabolism advantage clusters shared in several organoids have similar characteristic genes (GAPDH, NDRG1 (N-Myc downstream regulated 1), ALDOA, and CA9). The combination of GAPDH and NDRG1 is an independent risk factor and predictor for patient survival. This study delineates heterogeneity of hepatobiliary tumor organoids and proposes that the collaboration of intratumoral heterogenic subpopulations renders malignant phenotypes and drug resistance.

Falcarindiol Enhances Cisplatin Chemosensitivity of Hepatocellular Carcinoma via Down-Regulating the STAT3-Modulated PTTG1 Pathway.

Hepatocellular carcinoma (HCC) is the most frequent primary liver malignancy globally and the third leading cause of cancer-related death. Chemotherapy is one of the main methods in treating HCC, while recent studies have found that the resistance of HCC to chemotherapeutic drugs reduces the efficacy of the chemotherapy. Falcarindiol (FAD) is a cytotoxic and anti-inflammatory polyacetylenic oxylipin found in food plants of the carrot family (Apiaceae), while its role in HCC remains to be explored. Here, HCC cells (Huh7 and LM3) were treated with FAD at different doses. Cell proliferation was tested by the cell counting kit-8 (CCK-8) method and colony formation assay, while the apoptosis was monitored by flow cytometry. The profiles of apoptosis-related proteins (Bax, bcl2, and Caspase-3), DNA repair proteins (Rad51, BRCA1, and MDC1), and the signal transducer and activator of transcription 3 (STAT3)/Pituitary Tumor Transforming Gene 1 (PTTG1) were verified by western blot (WB) or quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The interaction between STAT3 and PTTG1 was verified by immunoprecipitation (IP). In addition, a xenograft tumor model was constructed in mice to explore the anti-tumor effects of FAD in vivo, and immunohistochemistry (IHC) was performed to count the number of Ki67-stained cells. As a result, FAD inhibited HCC cell proliferation and DNA repair, facilitated their apoptosis, and also enhanced cisplatin (DDP) chemosensitivity. The Combination Index (CI) evaluation showed that FAD and DDP had synergistic effects in repressing HCC cell proliferation. Besides, FAD dampened the STAT3/PTTG1 pathway expression. Further studies revealed that inhibiting STAT3 enhanced the inhibitive effect of FAD on HCC cells, whereas overexpressing PTTG1 attenuated the anti-tumor effect of FAD. Overall, our study illustrated that FAD is a potential anticancer drug and strengthens the chemosensitivity of HCC cells to DDP by inhibiting the STAT3/PTTG1 pathway.

circFN1 Mediates Sorafenib Resistance of Hepatocellular Carcinoma Cells by Sponging miR-1205 and Regulating E2F1 Expression.

In recent years, circular RNAs (circRNAs) have been shown to have critical regulatory roles in the resistance to anti-cancer drugs. However, the contributions of circRNAs to sorafenib resistance in hepatocellular carcinoma (HCC) remain largely unknown. The present study aims to explore the involvement of circFN1 in sorafenib resistance and how circFN1 is associated with the miR-1205/E2F1 pathway, which have been demonstrated to mediate this resistance in HCC cells. We investigated the expression of circRNAs in five paired sorafenib-sensitive HepG2 cells and sorafenib-resistant (SR)-HepG2 cells by microarray analysis. The quantitative real-time PCR analysis was used to investigate the expression pattern of circFN1 in HCC patient tissues and cell lines. Then, the effects of circFN1 on sorafenib resistance, cell proliferation, and apoptosis were assessed in HCC in vitro and in vivo. In this study, circFN1 was observed to be upregulated in HCC patient tissues and cell lines. Overexpression of circFN1 in HCC was significantly correlated with aggressive characteristics and served as an independent risk factor for overall survival in patients with HCC. Our in vivo and in vitro data indicated that inhibition of circFN1 enhances the sorafenib sensitivity of HCC cells. Mechanistically, we found that circFN1 could promote the expression of E2F1 by sponging miR-1205. In summary, our study demonstrated that circFN1 contributes to sorafenib resistance by regulating the miR-1205/E2F1 signaling pathway. These results indicate that circFN1 may represent a potentially valuable target for overcoming sorafenib resistance for HCC.

Trajectory and Functional Analysis of PD-1high CD4+CD8+ T Cells in Hepatocellular Carcinoma by Single-Cell Cytometry and Transcriptome Sequencing.

The spatial heterogeneity of immune microenvironment in hepatocellular carcinoma (HCC) remains elusive. Here, a single-cell study involving 17 432 600 immune cells of 39 matched HCC (T), nontumor (N), and leading-edge (L) specimens by mass cytometry is conducted. The tumor-associated CD4/CD8 double-positive T (DPT) cells are found enriched in L regions with synergetic expression of PD-1/HLA-DR/ICOS/CD45RO and exhibit a higher level of IFN-γ, TNF-α, and PD-1 upon stimulation. The enrichment of DPT and PD-1+DPT in L regions indicates favorable prognosis. These tumor-associated DPT cells with similar phenotype are also verified in other tumors and HCC animal models. Single-cell RNA-seq further characterizes the molecular features of DPT cells and uncovers 11 clusters with different cytotoxicity, exhaustion, and activation scores. TCR-based trajectory analysis reveals that tumor-associated DPT clusters share separated ancestries with local CD4+ or CD8+SPT cells rather than CD3+PBMC cells. TCR clones with frequency above 10 are mainly found coexisting in DPT and CD8+SPT cells. Specifically, PD-1highDPT cluster (TDPT_10) shares the same ancestry with exhausted CD8+SPT cluster (TCD8T_2) and shows higher expression similarity and closer pathological location to PD-1+CD8+ than PD-1+CD4+T cells. Together, this study systematically characterizes the unique distribution of PD-1+DPTs in HCC and puts forward new insights for the function and origin of tumor-associated DPT cells.

LncRNA ST8SIA6-AS1 promotes hepatocellular carcinoma cell proliferation and resistance to apoptosis by targeting miR-4656/HDAC11 axis.

BACKGROUND: Dysregulation of long non-coding RNAs (lncRNAs) results in development of human diseases including hepatocellular carcinoma (HCC). Although several HCC related lncRNAs have been reported, the biological functions of many lncRNAs during the development of HCC remains unknown. METHODS: The expression of ST8SIA6-AS1 was studied by realtime PCR (RT-qPCR) and bioinformatic analysis. The biological functions of ST8SIA6-AS1 was examined by CCK-8 assay and flow cytometry analysis. The target of ST8SIA6-AS1 was analyzed by bioinformatic analysis and validated by dual luciferase reporter assay, western blotting and RT-qPCR. RESULTS: In this study we demonstrated that ST8SIA6-AS1 was an upregulated lncRNA in hepatocellular carcinoma. SiRNA-mediated knockdown of ST8SIA6-AS1 repressed cell proliferation and induced cell apoptosis in HCC cells. Bioinformatic analysis and RT-qPCR further showed that ST8SIA6-AS1 mainly located in cytoplasm. Dual luciferase reporter assay further revealed that ST8SIA6-AS1 interacted with miR-4656 in HCC cells. In addition, HDAC11 was identified as a target gene in HCC cells and ST8SIA6-AS1 could upregulate HDAC11 via sponging miR-4656. Transfection of recombinant HDAC11 partially rescued the inhibition of cell proliferation and increase of cell apoptosis inducing by knockdown of ST8SIA6-AS1. CONCLUSION: In conclusion, our findings suggested that ST8SIA6-AS1 was a novel upregulated lncRNA in HCC and could facilitate cell proliferation and resistance to cell apoptosis via sponging miR-4656 and elevation of HDAC11, which might be a promising biomarker for patients with HCC.

Long noncoding RNA LINC00460 conduces to tumor growth and metastasis of hepatocellular carcinoma through miR-342-3p-dependent AGR2 up-regulation.

Hepatocellular carcinoma (HCC) is the fifth most common malignant tumor in the world. It ranks third among cancer-induced deaths worldwide and has the characteristics of high metastasis and high recurrence rate. Long non-coding RNA (LncRNA) LINC00460 is significantly up-regulated in multiple types of cancers and is closely related to the progression of tumors. However, effects of LINC00460 and corresponding regulatory path in HCC are still poorly investigated.In our study, we found that expression of LINC00460 was up-regulated in HCC tissues and cell lines compared with the control. Then we revealed that knockdown of LINC00460 suppressed cell proliferation and cell mobility and induced cell apoptosis in HCC cells. Further study demonstrated that knockdown of LINC00460 suppressed the progression of HCC by elevating the expression of microRNA (miRNA, miR)-342-3p. Besides that, metastasis marker, Anterior gradient homolog 2 (AGR2) was found to be a target of miR-342-3p and overexpression of AGR2 promoted the progression of HCC. Finally, the in vivo experiments further verified the anti-tumor effects of LINC00460 / miR-342-3p / AGR2 axis in HCC.The LINC00460 / miR-342-3p / AGR2 axis exerts anti-tumor effect in HCC in vitro and in vivo, consolidating and expanding the research about targeted gene therapy for early diagnosis and treatment of HCC.

Long Non-Coding RNA SNHG14 Contributes to the Development of Hepatocellular Carcinoma via Sponging miR-217.

BACKGROUND: Thousands of long non-coding RNAs (lncRNAs) have been functionally verified as crucial regulators of physiological processes and disease progressions, yet their roles in hepatocellular carcinoma (HCC) have not been clearly illuminated. METHODS: We analyzed the expression of lncRNA-SNHG14 in TCGA data via bioinformatic analysis and detected its expression in HCC specimens by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Loss-of-function experiments were used to study the biological function of SNHG14 in HCC cells. RT-qPCR, Western blotting and dual-luciferase reporter assay were carried out to investigate the molecular mechanism of SNHG14 in HCC. RESULTS: The upregulation of lncRNA-SNHG14 was observed in HCC tissues compared with normal tissues via RT-qPCR and bioinformatic analysis of TCGA data. Silencing of SNHG14 inhibited cell proliferation and induced cell apoptosis in HCC cells. microRNA-217 (miR-217), the tumor-suppressive miRNA in HCC, was predicted and confirmed as a miRNA sponged by SNHG14 in HCC cells. Via downregulation of miR-217, SNHG14 increased the expression of several miR-217-related oncogenes and subsequently activated oncogene-related signaling pathways in HCC cells. In addition, inhibition of miR-217 reversed SNHG14 silencing induced decrease of cell proliferation and increase of cell apoptosis. Their association was verified in the published microarray dataset and the collected HCC samples. CONCLUSION: In summary, SNHG14 is involved in the development of HCC via sponging miR-217 and it may be a biomarker for patients with HCC.

Targeted sequencing reveals the mutational landscape responsible for sorafenib therapy in advanced hepatocellular carcinoma.

Rationale: The existence of primary and acquired drug resistance is the main obstacle for the effect of multi-kinase inhibitor sorafenib and regorafenib in advanced hepatocellular carcinoma (HCC). However, plenty of patients did not significantly benefit from sorafenib treatment and little is known about the mechanism of drug resistance. Methods: Laser capture microdissection was used to acquire matched normal liver and tumor tissues on formalin-fixed paraffin-embedded specimens collected before sorafenib therapy from the first surgery of 119 HCC patients. Ultra-deep sequencing (~1000×) targeting whole exons of 440 genes in microdissected specimens and siRNA screen in 7 cell lines were performed to find mutations associated with differential responses to sorafenib. Patient-derived xenograft models were employed to determine the role of TP53 in response to sorafenib. Lentiviruses harboring wild-type and c.G52C-mutant OCT4 were applied to explore the function of OCT4 in resistance to sorafenib. ChIP-PCR assay for analysis of OCT4 transcriptional activity was performed to explore the affinity with the KITLG promoter. Statistical analyses were used to associate levels of p53 and OCT4 with tumor features and patient outcomes. Results: Total 1,050 somatic mutations and 26 significant driver genes were identified. SiRNA screening in 7 HCC cell lines was further performed to identify mutations associated with differential responses to sorafenib. A recurrent nonsynonymous mutation c.G52C in OCT4 (OCT4mut) was strongly associated with good response to sorafenib, whereas the stop-gain mutation in TP53 showed the opposite outcome both in vitro and in vivo. OCT4wt-induced stem cell factor (encoded by KITLG gene, SCF) expression and cross-activation of c-KIT/FLT3-BRAF signals were identified indispensably for sorafenib resistance, which could be reversed by the combination of c-KIT tyrosine kinase inhibitors or neutralizing antibody against SCF. Mechanistically, an OCT4 binding site in upstream of KITLG promoter was identified with a higher affinity to wildtype of OCT4 rather than G52C-mutant form, which is indispensable for OCT4-induced expression of KITLG and sorafenib resistance. Conclusion: Our study reported a novel somatic mutation in OCT4 (c.G52C) responsible for the sorafenib effect, and also shed new light on the treatment of HCC through the combination of specific tyrosine kinase inhibitors according to individual genetic patterns.

Transarterial chemoembolization versus percutaneous microwave coagulation therapy for recurrent unresectable intrahepatic cholangiocarcinoma: Development of a prognostic nomogram.

BACKGROUND: Transarterial chemoembolization (TACE) and percutaneous microwave coagulation therapy (PMCT) are commonly used to treat intrahepatic recurrent liver cancers. However, there is no information regarding their effectiveness in patients with recurrent intrahepatic cholangiocarcinoma (ICC) after resection. METHODS: A total of 275 patients with localized recurrent ICC who received either TACE (n = 183) or PMCT (n = 92) were studied. A propensity score matching analysis was performed to compare prognostic impact of TACE and PMCT. Prognostic factors for TACE and PMCT were identified respectively. Predictive nomograms for each TACE and PMCT were developed using the Cox independent prognostic factors and were validated in independent patient groups by receiver operating characteristic curves and area under curve values. RESULTS: Both TACE and PMCT provided curativeness in partial patients (5-year overall survival: 21.4% and 6.1%, respectively), but TACE provided better survival benefit in both overall patients (hazard ratio [HR] = 0.71; 95% confidence interval [CI]: 0.50-0.97; P = 0.034) and propensity score matching analysis (HR = 0.69; 95% CI: 0.47-0.98; P = 0.041). Independent prognostic factors for TACE were tumor size >5 cm, poor differentiation, and major resection, whereas poor differentiation, hepatitis B virus infection, cholelithiasis, and lymph node metastasis were identified for PMCT. Both predictive nomograms for TACE and PMCT were validated to be effective with area under curve values of 0.77 and 0.70, respectively. CONCLUSIONS: TACE provided better survival benefits compared to PMCT. However, there was a disparity in prognostic factors, suggesting evaluation of the two nomograms may be supportive in modality selection. Further prospective validation studies are required for the results to be applied in clinical medicine.

Latent Risk Intrahepatic Cholangiocarcinoma Susceptible to Adjuvant Treatment After Resection: A Clinical Deep Learning Approach.

Background: Artificial Intelligence (AI) frameworks have emerged as a novel approach in medicine. However, information regarding its applicability and effectiveness in a clinical prognostic factor setting remains unclear. Methods: The AI framework was derived from a pooled dataset of intrahepatic cholangiocarcinoma (ICC) patients from three clinical centers (n = 1,421) by applying the TensorFlow deep learning algorithm to Cox-indicated pathologic (four), serologic (six), and etiologic (two) factors; this algorithm was validated using a dataset of ICC patients from an independent clinical center (n = 234). The model was compared to the commonly used staging system (American Joint Committee on Cancer; AJCC) and methodology (Cox regression) by evaluating the brier score (BS), integrated discrimination improvement (IDI), net reclassification improvement (NRI), and area under curve (AUC) values. Results: The framework (BS, 0.17; AUC, 0.78) was found to be more accurate than the AJCC stage (BS, 0.48; AUC, 0.60; IDI, 0.29; NRI, 11.85; P < 0.001) and the Cox model (BS, 0.49; AUC, 0.70; IDI, 0.46; NRI, 46.11; P < 0.001). Furthermore, hazard ratios greater than three were identified in both overall survival (HR; 3.190; 95% confidence interval [CI], 2.150-4.733; P < 0.001) and disease-free survival (HR, 3.559; 95% CI, 2.500-5.067; P < 0.001) between latent risk and stable groups in validation. In addition, the latent risk subgroup was found to be significantly benefited from adjuvant treatment (HR, 0.459; 95% CI, 0.360-0.586; P < 0.001). Conclusions: The AI framework seems promising in the prognostic estimation and stratification of susceptible individuals for adjuvant treatment in patients with ICC after resection. Future prospective validations are needed for the framework to be applied in clinical practice.

Expression of Programmed Death Ligand 1 Is Associated with the Prognosis of Intrahepatic Cholangiocarcinoma.

BACKGROUND: Programmed death ligand 1 (PD-L1) is expressed in many malignancies and plays a critical role in escape from immune surveillance through inhibition of its receptor programmed death 1. The role of PD-L1 in intrahepatic cholangiocarcinoma (ICC) and mechanisms of its regulation, however, remain largely unknown. AIMS: To analyze the expression and prognostic significance of PD-L1 in ICC and to study the regulatory mechanisms of PD-L1. METHODS: Samples were obtained from 125 patients diagnosed with ICC in the Eastern Hepatobiliary Surgery Hospital from January 2012 to January 2013. The records of each patient were analyzed to examine the relationship between PD-L1 and clinical data. In vitro experiments were performed to investigate the relationship between PD-L1 and the IL-6/mTOR signaling pathway and the feedback mechanism pathway of PD-L1. RESULTS: Expression of PD-L1 is closely related to tumor vascular invasion, lymphatic metastasis and TNM staging. High PD-L1 expression is closely related to poor prognosis in ICC. Mechanically, IL-6 induces PD-L1 expression through mTOR signaling in ICC cells. In addition, PD-L1 has a negative feedback inhibition effect on AKT signaling. CONCLUSIONS: In summary, high PD-L1 expression was found to be associated with poor prognosis. The IL-6/mTOR pathway upregulates expression of PD-L1, thus promoting tumor invasion, and PD-L1 negatively inhibits the AKT pathway.