FAT10 induces immune suppression by upregulating PD-L1 expression in hepatocellular carcinoma.

The upregulation of programmed death ligand 1 (PD-L1) plays a crucial role in facilitating cancer cells to evade immune surveillance through immunosuppression. However, the precise regulatory mechanisms of PD-L1 in hepatocellular carcinoma (HCC) remain undefined. The correlation between PD-L1 and ubiquitin-like molecules (UBLs) was studied using sequencing data from 20 HCC patients in our center, combined with TCGA data. Specifically, the association between FAT10 and PD-L1 was further validated at both the protein and mRNA levels in HCC tissues from our center. Subsequently, the effect of FAT10 on tumor progression and immune suppression was examined through both in vivo and in vitro experiments. Utilizing sequencing data, qPCR, and Western blotting assays, we confirmed that FAT10 was highly expressed in HCC tissues and positively correlated with PD-L1 expression. Additionally, in vitro experiments demonstrated that the overexpression of FAT10 fostered the proliferation, migration, and invasion of HCC cells. Furthermore, the overexpression of FAT10 in HCC cells led to an increase in PD-L1 expression, resulting in the inhibition of T cell proliferation and the enhancement of HCC cell resistance to T cell-mediated cytotoxicity. Moreover, in vivo experiments utilizing the C57BL/6 mouse model revealed that overexpression of FAT10 effectively suppressed the infiltration of CD8 + GZMB + and CD8 + Ki67 + T cells, as well as reduced serum levels of TNF-α and IFN-γ. Mechanistically, we further identified that FAT10 upregulates PD-L1 expression via activating the PI3K/AKT/mTOR pathway, but not in a ubiquitin-like modification. In conclusion, our findings indicate that FAT10 promotes immune evasion of HCC via upregulating PD-L1 expression, suggesting its potential as a novel target to enhance the efficiency of immunotherapy in HCC.

Identification of FOXP1 as a favorable prognostic biomarker and tumor suppressor in intrahepatic cholangiocarcinoma.

BACKGROUND: Forkhead-box protein P1 (FOXP1) has been proposed to have both oncogenic and tumor-suppressive properties, depending on tumor heterogeneity. However, the role of FOXP1 in intrahepatic cholangiocarcinoma (ICC) has not been previously reported. METHODS: Immunohistochemistry was performed to detect FOXP1 expression in ICC and normal liver tissues. The relationship between FOXP1 levels and the clinicopathological characteristics of patients with ICC was evaluated. Finally, in vitro and in vivo experiments were conducted to examine the regulatory role of FOXP1 in ICC cells. RESULTS: FOXP1 was significantly downregulated in the ICC compared to their peritumoral tissues (p < 0.01). The positive rates of FOXP1 were significantly lower in patients with poor differentiation, lymph node metastasis, invasion into surrounding organs, and advanced stages (p < 0.05). Notably, patients with FOXP1 positivity had better outcomes (overall survival) than those with FOXP1 negativity (p < 0.05), as revealed by Kaplan-Meier survival analysis. Moreover, Cox multivariate analysis showed that negative FOXP1 expression, advanced TNM stages, invasion, and lymph node metastasis were independent prognostic risk factors in patients with ICC. Lastly, overexpression of FOXP1 inhibited the proliferation, migration, and invasion of ICC cells and promoted apoptosis, whereas knockdown of FOXP1 had the opposite role. CONCLUSION: Our findings suggest that FOXP1 may serve as a novel outcome predictor for ICC as well as a tumor suppressor that may contribute to cancer treatment.

Chinese expert recommendations on management of hepatocellular carcinoma during COVID-19 pandemic: a nationwide multicenter survey.

BACKGROUND: This study aimed to investigate the work status of clinicians in China and their management strategy alteration for patients with hepatocellular carcinoma (HCC) during the COVID-19 pandemic. METHODS: A nationwide online questionnaire survey was conducted in 42 class-A tertiary hospitals across China. Experienced clinicians of HCC-related specialties responded with their work status and management suggestions for HCC patients during the pandemic. RESULTS: 716 doctors responded effectively with a response rate of 60.1%, and 664 were included in the final analysis. Overall, 51.4% (341/664) of clinicians reported more than a 60% reduction of the regular workload and surgeons declared the highest proportion of workload reduction. 92.5% (614/664) of the respondents have been using online medical consultation to substitute for the "face-to-face" visits. Adaptive adjustment for the treatment strategy for HCC was made, including the recommendations of noninvasive and minimally invasive treatments such as transcatheter arterial chemoembolization for early and intermediate stage. Targeted therapy has been the mainstay for advanced stage and also as a bridge therapy for resectable HCC. DISCUSSION: During the COVID-19 pandemic, online medical consultation is recommended to avoid social contact. Targeted therapy as a bridge therapy is recommended for resectable HCC considering the possibility of delayed surgery.

Totally laparoscopic anatomic S7 segmentectomy using in situ split along the right intersectoral and intersegmental planes.

BACKGROUND: The traditional open or laparoscopic segmentectomy of liver segment 7 (S7) requires exposing and controlling the root of the right hepatic vein(RHV)after full mobilization and lifting up of the right liver before liver transection. This approach violates the "no-touch" principle for malignant tumors, and makes laparoscopic resection technically challenging. So reports on isolated totally laparoscopic anatomic S7 segmentectomy have rarely been reported. This study describes our experience in laparoscopic anatomic S7 segmentectomy using in situ split along the right intersectoral and intersegmental planes of the liver. To our knowledge, this is the first description of this novel approach. METHODS: From September 2017 to May 2019, patients who underwent laparoscopic anatomic S7 segmentectomy for hepatocellular carcinoma at the HPB Surgery Department, Sun Yat-Sen Memorial Hospital entered into this retrospective study. This in situ split approach was designed using main vessels as the plane markers of right intersectoral and intersegmental planes, along which liver transection was carried out. There was no need to mobilize the right liver and control the root of RHV. RESULTS: There were 9 women and 15 men. The average diameter of the tumors on preoperative CT/MR was 3.4 cm (range 2-6 cm). All the procedures were successfully carried out laparoscopically. There was no perioperative death. The average operative time was 216.5 min (range 180-310 min). The average blood loss was 320 ml (range 120-620 ml). Pathological study showed all the operations to be R0 resections. CONCLUSION: Laparoscopic anatomic S7 segmentectomy using the in situ split approach resulted in R0 liver resection in all our patients with primary liver cancer. The operation was technically feasible and it provided a better view and increased maneuverability in the cramped operative space compared with the traditional open/laparoscopic approach. The approach also better complies with the "no-touch" principle for malignant tumors. Its long-term oncological outcomes require further studies.

Extensively expanded murine-induced hepatic stem cells maintain high-efficient hepatic differentiation potential for repopulation of injured livers.

BACKGROUND & AIM: Shortage of donor hepatocytes limits hepatocyte transplantation for clinical application. Induced hepatic stem cells (iHepSCs) have capacities of self-renewal and bipotential differentiations. Here, we investigated whether iHepSCs could be extensively expanded, and whether they could differentiate into sufficient functional hepatocytes as donors for transplantation therapy after their extensive expansions. METHODS: Murine extensively expanded iHepSCs (50-55 passages) were induced to differentiate into iHepSC-Heps under a chemically defined condition. iHepSC-Heps were proved for carrying morphological hepatocyte characters and hepatocytic functions including low-density lipoprotein uptake, glycogen storage, CLF secretion, ICG uptake and release, Alb secretion, urea synthesis and metabolism-relative gene expressions respectively. Next, both iHepSCs and iHepSC-Heps were transplanted into Fah-/- mice respectively. Both liver repopulation and alleviation of liver function were compared between two transplantation groups. RESULTS: Murine iHepSCs still maintained the capacities of self-renewal and bipotential differentiations after extensive expansion. The efficiency for the functional hepatocyte differentiation from extensively expanded iHepSCs reached to 72.64%. Transplantations of both extensively expanded iHepSCs and iHepSC-Heps resulted in liver engraftment in Fah-/- mice. Survival rate of Fah-/- mice recipients and level of liver repopulation were 50% and 20.32 ± 4.58% respectively in iHepSC-Heps group, while 33% and 10.4 ± 4.3% in iHepSCs group. CONCLUSIONS: Extensively expanded iHepSCs can efficiently differentiate into hepatocytes in chemical defined medium. Transplantation of iHepSC-Heps was more effective and more efficient than transplantation of iHepSCs in Fah-/- mice. Our results suggested an innovative system to obtain sufficient hepatocytes through hepatic differentiation of iHepSCs generated by lineage reprogramming.

Laparoscopic duodenum-preserving total pancreatic head resection: a novel surgical approach for benign or low-grade malignant tumors.

BACKGROUND: Duodenum-preserving total pancreatic head resection (DPPHRt) is an accepted alternative surgical procedure for benign or low-grade malignant tumors of the pancreatic head by preserving the duodenum with its intact blood supply from the pancreatic duodenal arterial arcade. This study describes our experience in laparoscopic DPPHRt (LDPPHRt). To our knowledge, this is the first description of this novel minimally invasive operation. METHODS: From August 2016 to May 2017, all consecutive patients who underwent LDPPHRt for pancreatic head lesions at the HPB Surgery Department, Sun Yat-Sen Memorial Hospital in Guangzhou, China were enrolled into this retrospective study. RESULTS: There were ten women and two men. The average age was 37.3 years (range 8-61 years). The average diameter of the pancreatic head lesions on pre-operative CT/MR was 3.7 cm (range 2-4.8 cm). All the LDPPHRt procedures were performed successfully. There was no peri-operative death. The average operative time was 272.5 min (range 210-320 min). The average blood loss was 215 ml (range 50-450 ml). Post-operative complications included pancreatic fistula grade B (two patients, or 16.7%) and biliary fistula (two patients, or 16.7%). All the complications responded well to conservative treatment. The mean post-operative hospital stay was 11.5 days (range 6-25 days). CONCLUSIONS: LDPPHRt provided a minimally invasive approach with good organ-preservation for benign or low-grade malignant tumors of the pancreatic head. The long-term oncological outcomes, and the exocrine and endocrine pancreatic functions after this operation require further studies.

Is lymph node dissection necessary for resectable intrahepatic cholangiocarcinoma? A systematic review and meta-analysis.

BACKGROUND: The objective of this meta-analysis was to evaluate the effectiveness and safety of lymph node dissection (LND) in patients with intrahepatic cholangiocarcinoma (ICC). METHODS: A literature search with a date range of January 2000 to January 2018 was performed to identify studies comparing lymph node dissection (LND+) with non-lymph node dissection (LND-) for patients with ICC. The LND + group was further divided into positive (LND + N+) and negative (LND + N-) lymph node status groups based on pathological analysis. RESULTS: 13 studies including 1377 patients were eligible. There were no significant differences in overall survival (OS) (HR 1.13, 95% CI 0.94-1.36; P = 0.20), disease-free survival (DFS) (HR 1.23, 95% CI 0.94-1.60; P = 0.13), or recurrence (OR 1.39, 95% CI 0.90-2.15; P = 0.14) between LND + group and LND-group. Postoperative morbidity was significantly higher in the LND + group (OR 2.67, 95% CI 1.74-4.10; P < 0.001). A subset analysis showed that OS was similar between LND + N- and LND-groups (HR 1.13, 95% CI 0.82-1.56; P = 0.450). However when comparing, OS of the LND-group to the LND+N+ group there was a significant increase in OS for the LND-group (HR 3.26, 95% CI 1.85-5.76; P < 0.001). CONCLUSIONS: LND does not seem to positively affect overall survival and is associated with increased post-operative morbidity.

KRAS promotes tumor metastasis and chemoresistance by repressing RKIP via the MAPK-ERK pathway in pancreatic cancer.

Oncogenic KRAS plays a crucial role in pancreatic ductal adenocarcinoma (PDAC) development and progression. However, the mechanism has not been clearly elucidated. RKIP is a tumor repressor, and loss of RKIP has been shown in PDAC. Here, we found that KRAS expression was inversely correlated with RKIP expression in PDAC fresh tissue regardless of the KRAS mutant status. The negative correlation between KRAS and RKIP was further confirmed in our PDAC tissue microarray. KRAS overexpression and RKIP downregulation were associated with poor clinical outcomes. Knockdown or overexpression of KRAS in PDAC cell lines robustly increased or decreased, respectively, RKIP protein and mRNA levels. Furthermore, the MAPK-ERK pathway was involved in the regulation of RKIP. KRAS-regulated RKIP expression, which in turn affected the expression of pivotal epithelial-mesenchymal transition (EMT) and apoptosis factors. The biological function of the KRAS-RKIP axis was demonstrated in human pancreatic cancer cells in vitro and in vivo. KRAS knockdown increased RKIP expression and inhibited metastasis and chemoresistance. Moreover, the feature of metastasis and chemoresistance was rescued in the KRAS-knockdown cells through the inhibition of RKIP by RNA interference. In conclusion, our studies demonstrate how KRAS inhibits the tumor suppressor RKIP, thus offering novel justification for targeting RKIP as a strategy to overcome KRAS-induced tumor metastasis and chemoresistance in PDAC.

Human embryonic stem cell-derived hepatoblasts are an optimal lineage stage for hepatitis C virus infection.

Maturation of hepatic cells can be gradually acquired through multiple stages of hepatic lineage specification, while it is unclear whether hepatitis C virus (HCV) infection is maturationally lineage-dependent. We investigated the susceptibility to HCV at multiple stages of human embryonic stem cells, definitive endodermal cells, hepatic stem cells, hepatoblasts (hHBs), and mature hepatocytes. Susceptibility to infection occurred initially at the stage of human hepatic stem cells; however, hHBs proved to have the highest permissiveness and infectivity compared with all other stages. The hHBs' susceptibility to HCV correlated with the translocation of occludin, an HCV receptor, from cytoplasm to plasma membrane of HBs. Vascular endothelial cell growth factor enhanced the HCV susceptibility of hHBs through rearrangement of occludin by dephosphorylation; this minimized hHB polarization and prevented hHBs from further maturation. The transcription profiles of different hepatic lineage stages indicated that expression of innate immune response genes was correlated with hepatic maturation; interferon ß played an important role in protecting hHBs from HCV infection. HCV-infected hHBs were able to engraft and integrate into the livers of Fah-/- Rag2-/- mice and maintained an hHB phenotype for over 12 weeks during the time when HCV antigen was evident. After suppression of interferon ß in hHBs, HCV infection was significantly enhanced in the engrafted humanized liver tissue of host mice. CONCLUSION: Human embryonic stem cell-derived hHBs are the optimal hosts for HCV infectivity; the realization that HCV entry and replication occur primarily at a particular hepatic lineage stage enables us to understand the HCV infection factors, life cycle, and infection dynamics that are facets of the pathogenesis as well as suggesting targets for anti-HCV treatment. (Hepatology 2017;66:717-735).

Pericentral hepatocytes produce insulin-like growth factor-2 to promote liver regeneration during selected injuries in mice.

Liver regeneration (LR) happens after various types of injuries. Unlike the well-studied LR caused by partial hepatectomy (PHx), there is accumulating evidence suggesting that LR during other injuries may result from unknown mechanisms. In this study, we found that insulin-like growth factor 2 (IGF-2) was drastically induced following the liver injuries caused by tyrosinemia or long-term treatments of CCl4 . However, this was not observed during the early phase of acute liver injuries after PHx or single treatment of CCl4 . Remarkably, most IGF-2-expressing hepatocytes were located at the histological area around the central vein of the liver lobule after the liver injuries caused either in fumarylacetoacetate hydrolase-deficient mice or in CCl4 chronically treated mice. Hepatocyte proliferation in vivo was significantly promoted by induced IGF-2 overexpression, which could be inhibited by adeno-associated virus-delivered IGF-2 short hairpin RNAs or linsitinib, an inhibitor of IGF-2 signaling. Proliferating hepatocytes in vivo responded to IGF-2 through both insulin receptor and IGF-1 receptor. IGF-2 also significantly promoted DNA synthesis of primary hepatocytes in vitro. More interestingly, the significantly induced IGF-2 was also found to colocalize with glutamine synthetase in the region enriched with proliferating hepatocytes for the liver samples from patients with liver fibrosis. CONCLUSION: IGF-2 is produced by pericentral hepatocytes to promote hepatocyte proliferation and repair tissue damage in the setting of chronic liver injury, which is distinct from the signaling that occurs post-PHx. (Hepatology 2017;66:2002-2015).

Vps4A functions as a tumor suppressor by regulating the secretion and uptake of exosomal microRNAs in human hepatoma cells.

UNLABELLED: The deregulation of microRNAs (miRNAs) plays an important role in human hepatocarcinogenesis. In this study, we highlight exosomes as mediators involved in modulating miRNA profiles in hepatocellular carcinoma (HCC) cells. First, we examined the different miRNA expression profiles in HCC cells and HCC cell-derived exosomes. Next, coculture experiments indicated that HCC cell-derived exosomes promoted the cell growth, migration, and invasion of HCC cells and had the ability to shuttle miRNAs to recipient cells. Further, our data showed that Vps4A, a key regulator of exosome biogenesis, was frequently down-regulated in HCC tissues. The reduction of Vps4A in HCC tissues was associated with tumor progression and metastasis. In vitro studies revealed that Vps4A repressed the growth, colony formation, migration, and invasion of HCC cells. We further investigated the role and involvement of Vps4A in suppressing the bioactivity of exosomes and characterized its ability to weaken the cell response to exosomes. By small RNA sequencing, we demonstrated that Vps4A facilitated the secretion of oncogenic miRNAs in exosomes as well as accumulation and uptake of tumor suppressor miRNAs in cells. A subset of Vps4A-associated miRNAs was identified. Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that the phosphatidylinositol-3-kinase/Akt signaling pathway was the most likely candidate pathway for modulation by these miRNAs. Indeed, we proved that the phosphatidylinositol-3-kinase/Akt pathway was inactivated by Vps4A overexpression. CONCLUSION: Exosome-mediated miRNA transfer is an important mechanism of self-modulation of the miRNA expression profiles in HCC cells, and Vps4A may function as a tumor suppressor, which utilizes exosomes as mediators to regulate the secretion and uptake of miRNAs in hepatoma cells; these observations provide new insights into the development of HCC.

Cabozantinib reverses multidrug resistance of human hepatoma HepG2/adr cells by modulating the function of P-glycoprotein.

BACKGROUND & AIMS: Cabozantinib, a small-molecule multitargeted tyrosine kinase inhibitor, has entered into a phase III clinical trial for the treatment of hepatocellular carcinoma (HCC). This study assessed the mechanistic effect of cabozantinib on the reversal of P-glycoprotein (P-gp)-mediated multidrug resistance (MDR). METHODS: CCK-8 assays and tumour xenografts were used to investigate the reversal of MDR in vitro and in vivo respectively. Substrate retention assays were evaluated by fluorescence microscope and flow cytometry. Western blotting was used to detect protein expression levels. mRNA expression was determined by qPCR. The ATPase activity of P-gp was investigated using Pgp-Glo(™) assay systems. The binding mechanism of cabozantinib to P-gp at the molecular level was evaluated using docking analysis. RESULTS: Cabozantinib enhanced the cytotoxicity of P-gp substrate drugs in HepG2/adr and HEK293-MDR1 cells but had no effect on non-P-gp substrates. In addition, cabozantinib increased the accumulation of P-gp substrates in HepG2/adr cells but had no effect in HepG2 cells. Furthermore, cabozantinib did not alter the expression of P-gp mRNA or protein but did stimulate the activity of P-gp ATPase. The docking study indicated that cabozantinib and verapamil may partially share a binding site on P-gp. The reversal concentrations of cabozantinib did not affect the expression of MET, AKT and ERK1/2. Significantly, cabozantinib increased the inhibitory efficacy of doxorubicin in P-gp-overexpressing HepG2/adr cell xenografts in nude mice. CONCLUSION: Cabozantinib reverses P-gp-mediated MDR by directly inhibiting the efflux function of P-gp, indicating that cabozantinib may help to reverse P-gp-mediated MDR in HCC and other cancer chemotherapy.

Laparoscopic surgery minimizes the release of circulating tumor cells compared to open surgery for hepatocellular carcinoma.

BACKGROUND: The aim of this study was to determine whether tumor manipulation enhances cancer cell release from the primary tumor in HCC patients and which surgical approach, open surgery or laparoscopic resection, is superior with respect to preventing tumor cells from scattering in the blood. METHODS: A total of 26 HCC patients were prospectively randomized to receive either open surgery (n = 14) or laparoscopic surgery (n = 12). Blood samples were obtained at three time points: preoperative, postoperative, and 24 h after surgery. The CD45(-)/CD44(+)/CD90(+) cells were obtained and counted using quantitative flow cytometry. The serum levels of interleukin-6 (IL-6), interleukin-8 (IL-8), and tumor necrosis factor (TNF-α) were also compared between the two groups. RESULTS: There was no significant difference between the laparoscopic and open groups in terms of patient characteristics. The levels of CCSCs increased immediately after surgical manipulation, and the laparoscopy group released fewer tumor cells into the blood stream. The amount of CCSCs in both groups decreased to reach a similar level 24 h after surgery. Both IL-6 and IL-8 increased after surgery, and the mean postoperative increases in IL-6 and IL-8 serum levels were significantly less in the laparoscopic group than in the open group. The TNF-α levels showed no differences at any time point. CONCLUSIONS: Our results showed that patients with laparoscopic surgery have lower IL-6, IL-8 secretion and less CTCs, which may suggest an advantage by restricting CTCs release and a preserved immune response. Further studies are needed to investigate the relationship between the number of CCSCs after surgery and long-term survival rates.

Timing of hepatectomy in resectable synchronous colorectal liver metastases (SCRLM): Simultaneous or delayed?

UNLABELLED: The optimal surgical strategy for treatment of patients with synchronous colorectal liver metastases (SCLRM) remains controversial. We conducted a systematic review and meta-analysis of all observational studies to define the safety and efficacy of simultaneous versus delayed resection of the colon and liver. A search for all major databases and relevant journals from inception to April 2012 without restriction on languages or regions was performed. Outcome measures were the primary parameters of postoperative survival, complication, and mortality, as well as other parameters of blood loss, operative time, and length of hospitalization. The test of heterogeneity was performed with the Q statistic. A total of 2,880 patients were included in the meta-analysis. Long-term oncological pooled estimates of overall survival (hazard ratio [HR]: 0.96; 95% confidence interval [CI]: 0.81-1.14; P = 0.64; I(2) = 0) and recurrence-free survival (HR: 1.04; 95% CI: 0.76-1.43; P = 0.79; I(2) = 53%) all showed similar outcomes for both simultaneous and delayed resections. A lower incidence of postoperative complication was attributed to the simultaneous group as opposed to that in the delayed group (modified relative ratio [RR] = 0.77; 95% CI: 0.67-0.89; P = 0.0002; I(2) = 10%), whereas in terms of mortality within the postoperative 60 days no statistical difference was detected (RR = 1.12; 95% CI: 0.61-2.08; P = 0.71; I(2) = 32%). Finally, selection criteria were recommended for SCRLM patients suitable for a simultaneous resection. CONCLUSION: Simultaneous resection is as efficient as a delayed procedure for long-term survival. There is evidence that in SCRLM patients simultaneous resection is an acceptable and safe option with carefully selected conditions. Due to the inherent limitations of the present study, future randomized controlled trials will be useful to confirm this conclusion. (HEPATOLOGY 2013;57:2346-2357).

A prospective, randomized, controlled, trial comparing occult-scar incision laparoscopic cholecystectomy and classic three-port laparoscopic cholecystectomy.

BACKGROUND: This study was designed to evaluate the outcome of laparoscopic cholecystectomy by comparing a new technique using occult-scar incision for laparoscopic cholecystectomy (OSLC) with classic three-port laparoscopic cholecystectomy (CLC). In the occult-scar incision, we moved the subcostal and subxiphoid trocar insertion sites to the suprapubic area so that operative scars were hidden in the pubic hairs and below umbilicus. METHODS: Between July 2009 and 2012, patients undergoing laparoscopic cholecystectomy were randomized to the OSLC or CLC approach after obtaining informed consent. Outcome was measured by operative time, operative complications, hospital length of stay, cost, analgesia required after surgery, and cosmetic outcomes. The patient satisfaction score (PSS) and visual analog score (VAS) also were used to evaluated the level of cosmetic result and postoperative pain. RESULTS: A total of 75 patients were randomized into CLC (n = 35) and OSLC (n = 40) groups. No patient was converted to an open procedure in either the CLC or OSLC group. No operative complications were reported within 30 days in either group. The PSS of 7 and 30 days after surgery were both significantly higher in the OSLC group than in the CLC group (5.8 ± 1.5 vs. 8.0 ± 1.1, P = 0.03; 6.5 ± 1.2 vs. 9.2 ± 0.8, P = 0.02). The VAS for pain was significantly lower in the OSLC group on postoperative day 3 compared with the CLC group (2.6 ± 1.2 vs. 6.3 ± 0.9, P = 0.01). There was no significant difference in operative time, hospital stay, and cost between the two groups. CONCLUSIONS: The OSLC is a safe and feasible alternative compared with CLC in experienced hands, and it is superior for outcomes regarding pain control and cosmesis.

[Laparoscopic hepatectomy for recurrent hepatocellular carcinoma after previous open hepatectomy].

OBJECTIVE: To evaluate the feasibility, safety and long-term outcomes of laparoscopic repeat hepatectomy for recurrent hepatocellular carcinoma (HCC) following previous resection. METHODS: Between January 2003 and January 2011, 14 patients with recurrent HCC were carefully selected to undergo repeat laparoscopic hepatectomy, among which 9 patients were male, 5 patients were female, and the average age was 54 years. Prior to re-resection, all patients had undergone at least one open hepatectomy for HCC. The perioperative and long-term outcomes of these patients were retrospectively analyzed. RESULTS: Repeat laparoscopic hepatectomy for these 14 patients were successfully performed without major perioperative complications. The mean operative time, intraoperative blood loss and hospital stay were (124 ± 82) minutes, (112 ± 43) ml and (7 ± 4) days, respectively. The mean follow-up period was 23 months (range 14 to 42 months). At the time of follow-up, 11 patients were still alive, among which 3 patients developed recurrent disease and 8 patients remained disease free. One patient died of liver dysfunction at 21 months, and another 2 patients died of tumor recurrence at 17, 31 months, respectively. CONCLUSION: Laparoscopic surgery for recurrent HCC remains a viable option for repeat hepatectomy in selected patients who have undergone open hepatectomy.

CircNUP54 promotes hepatocellular carcinoma progression via facilitating HuR cytoplasmic export and stabilizing BIRC3 mRNA.

Circular RNAs (circRNAs) have been implicated in tumorigenesis and progression of various cancers. However, the underlying mechanisms of circRNAs in hepatocellular carcinoma (HCC) have not been fully elucidated. Herein, a new oncogenic circRNA, hsa_circ_0070039 (circNUP54), was identified to be significantly upregulated in HCC through circRNA sequencing. As verified in 68 HCC samples, circNUP54 overexpression was correlated with aggressive cancerous behaviors and poor outcomes. Moreover, the function experiments showed that knockdown of circNUP54 inhibited the malignant progression of HCC in vitro and in vivo, whereas overexpression of circNUP54 had the opposite role. Mechanistic investigations carried out by RNA pull-down, RNA immunoprecipitation, and immunofluorescence revealed that circNUP54 interacted with the RNA-binding protein Hu-antigen R (HuR) and promoted its cytoplasmic export. The cytoplasmic accumulation of HuR stabilized the downstream BIRC3 mRNA through its binding to the 3' UTR region. Consequently, the encoded protein of BIRC3, cellular inhibitor of apoptosis 2 (cIAP2), proceeded to activate the NF-κB signal pathway and ultimately contributed to HCC progression. In addition, depletion of BIRC3 rescued the pro-tumorigenic effect of circNUP54 on HCC cells. Overall, this study demonstrated that circNUP54 facilitates HCC progression via regulating the HuR/BIRC3/NF-κB axis, which may serve as a promising therapeutic target for HCC treatment.

A novel risk score system based on immune subtypes for identifying optimal mRNA vaccination population in hepatocellular carcinoma.

PURPOSE: Although mRNA vaccines have shown certain clinical benefits in multiple malignancies, their therapeutic efficacies against hepatocellular carcinoma (HCC) remains uncertain. This study focused on establishing a novel risk score system based on immune subtypes so as to identify optimal HCC mRNA vaccination population. METHODS: GEPIA, cBioPortal and TIMER databases were utilized to identify candidate genes for mRNA vaccination in HCC. Subsequently, immune subtypes were constructed based on the candidate genes. According to the differential expressed genes among various immune subtypes, a risk score system was established using machine learning algorithm. Besides, multi-color immunofluorescence of tumor tissues from 72 HCC patients were applied to validate the feasibility and efficiency of the risk score system. RESULTS: Twelve overexpressed and mutated genes associated with poor survival and APCs infiltration were identified as potential candidate targets for mRNA vaccination. Three immune subtypes (e.g. IS1, IS2 and IS3) with distinct clinicopathological and molecular profiles were constructed according to the 12 candidate genes. Based on the immune subtype, a risk score system was developed, and according to the risk score from low to high, HCC patients were classified into four subgroups on average (e.g. RS1, RS2, RS3 and RS4). RS4 mainly overlapped with IS3, RS1 with IS2, and RS2+RS3 with IS1. ROC analysis also suggested the significant capacity of the risk score to distinguish between the three immune subtypes. Higher risk score exhibited robustly predictive ability for worse survival, which was further independently proved by multi-color immunofluorescence of HCC samples. Notably, RS4 tumors exhibited an increased immunosuppressive phenotype, higher expression of the twelve potential candidate targets and increased genome altered fraction, and therefore might benefit more from vaccination. CONCLUSIONS: This novel risk score system based on immune subtypes enabled the identification of RS4 tumor that, due to its highly immunosuppressive microenvironment, may benefit from HCC mRNA vaccination.

Toripalimab Plus Bevacizumab as First-line Treatment for Advanced Hepatocellular Carcinoma: A Prospective, Multicenter, Single-Arm, Phase II Trial.

PURPOSE: This phase II, multicenter, prospective, single-arm study aimed to evaluate the efficacy and safety of toripalimab plus bevacizumab in treating advanced hepatocellular carcinoma (HCC). PATIENTS AND METHODS: Treatment-naïve patients with advanced HCC received toripalimab 240 mg plus bevacizumab 15 mg/kg every 3 weeks. Primary endpoints included safety and tolerability, and objective response rate (ORR) assessed by the investigator per Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1. RESULTS: Fifty-four patients were enrolled between Apr 17, 2020 and Dec 11, 2020. As assessed by the investigator according to RECIST v1.1, the ORR was 31.5% [95% confidence interval (CI), 19.5-45.6] and the lower bound of the 95% CI was above the pre-specified boundary of 10%. The independent review committee (IRC) assessed ORR according to modified RECIST (mRECIST) was 46.3% (95% CI, 32.6-60.4). The median progression-free survival were 8.5 months (95% CI, 5.5-11.0) and 9.8 months (95% CI, 5.6-not evaluable) assessed by the investigator according to RECIST v1.1 and IRC according to mRECIST criteria, respectively. The median overall survival (OS) was not reached, and the 12- and 24-month OS rates were 77.3% and 63.5%, respectively. Grade 3 or higher treatment-emergent adverse events (TEAEs) occurred in 27 patients (50.0%). The most common TEAEs were proteinuria (59.3%), hypertension (38.9%), aspartate aminotransferase increased (33.3%), amylase increased (29.6%), platelet count decreased (27.8%), and bilirubin increased (27.8%). CONCLUSIONS: Toripalimab plus bevacizumab showed a favorable efficacy and safety profile, supporting further studies of this combination regimen as a first-line treatment of advanced HCC.

Anticancer drugs cause release of exosomes with heat shock proteins from human hepatocellular carcinoma cells that elicit effective natural killer cell antitumor responses in vitro.

Failure of immune surveillance related to inadequate host antitumor immune responses has been suggested as a possible cause of the high incidence of recurrence and poor overall survival outcome of hepatocellular carcinoma. The stress-induced heat shock proteins (HSPs) are known to act as endogenous "danger signals" that can improve tumor immunogenicity and induce natural killer (NK) cell responses. Exosome is a novel secretory pathway for HSPs. In our experiments, the immune regulatory effect of the HSP-bearing exosomes secreted by human hepatocellular carcinoma cells under stress conditions on NK cells was studied. ELISA results showed that the production of HSP60, HSP70, and HSP90 was up-regulated in both cell lines in a stress-specific manner. After exposure to hepatocellular carcinoma cell-resistant or sensitive anticancer drugs (hereafter referred to as "resistant" or "sensitive" anticancer drug), the membrane microvesicles were actively released by hepatocellular carcinoma cells, differing in their ability to present HSPs on the cell surface, which were characterized as exosomes. Acting as a decoy, the HSP-bearing exosomes efficiently stimulated NK cell cytotoxicity and granzyme B production, up-regulated the expression of inhibitory receptor CD94, and down-regulated the expression of activating receptors CD69, NKG2D, and NKp44. Notably, resistant anticancer drugs enhanced exosome release and generated more exosome-carried HSPs, which augmented the activation of the cytotoxic response. In summary, our findings demonstrated that exosomes derived from resistant anticancer drug-treated HepG2 cells conferred superior immunogenicity in inducing HSP-specific NK cell responses, which provided a clue for finding an efficient vaccine for hepatocellular carcinoma immunotherapy.