A multimodal atlas of hepatocellular carcinoma reveals convergent evolutionary paths and 'bad apple' effect on clinical trajectory.

BACKGROUND & AIMS: Hepatocellular Carcinoma (HCC) is a highly fatal cancer characterized by high intra-tumor heterogeneity (ITH). A panoramic understanding of its tumor evolution, in relation to its clinical trajectory, may provide novel prognostic and treatment strategies. METHODS: Through the Asia-Pacific Hepatocellular Carcinoma (AHCC) trials group (NCT03267641), we recruited one of the largest prospective cohorts of HCC with over 600 whole genome and transcriptome samples from 123 treatment-naïve patients. RESULTS: Using a multi-region sampling approach, we revealed seven convergent genetic evolutionary paths governed by the early driver mutations, late copy number variations and viral integrations, which stratify patient clinical trajectories after surgical resection. Furthermore, such evolutionary paths shaped the molecular profiles, leading to distinct transcriptomic subtypes. Most significantly, although we found the coexistence of multiple transcriptomic subtypes within certain tumors, patient prognosis was best predicted by the most aggressive cell fraction of the tumor, rather than by overall degree of transcriptomic ITH level - a phenomenon we termed the 'bad apple' effect. Finally, we found that characteristics throughout early and late tumor evolution provide significant and complementary prognostic power in predicting patient survival. CONCLUSIONS: Taken together, our study generated a comprehensive landscape of evolutionary history for HCC and provided a rich multi-omics resource for understanding tumor heterogeneity and clinical trajectories. CLINICAL TRIAL NUMBER: NCT03267641 (Observational cohort) IMPACT AND IMPLICATIONS: This prospective study, utilizing comprehensive multi-sector, multi-omics sequencing and clinical data from surgically resected HCC, reveals critical insights into the role of tumor evolution and intra-tumor heterogeneity (ITH) in determining the prognosis of Hepatocellular Carcinoma (HCC). These findings are invaluable for oncology researchers and clinicians, as they underscore the influence of distinct evolutionary paths and the 'bad apple' effect, where the most aggressive tumor fraction dictates disease progression. These insights not only enhance prognostic accuracy post-surgical resection but also pave the way for developing personalized therapies tailored to specific tumor evolutionary and transcriptomic profiles. The co-existence of multiple sub-types within the same tumor prompts a re-appraisal of the utilities of depending on single samples to represent the entire tumor and suggests the need for clinical molecular imaging. This research thus marks a significant step forward in the clinical understanding and management of HCC, underscoring the importance of integrating tumor evolutionary dynamics and multi-omics biomarkers into therapeutic decision-making.

Hypoxia-driven immunosuppression by Treg and type-2 conventional dendritic cells in HCC.

BACKGROUND AND AIMS: Hypoxia is one of the central players in shaping the immune context of the tumor microenvironment (TME). However, the complex interplay between immune cell infiltrates within the hypoxic TME of HCC remains to be elucidated. APPROACH AND RESULTS: We analyzed the immune landscapes of hypoxia-low and hypoxia-high tumor regions using cytometry by time of light, immunohistochemistry, and transcriptomic analyses. The mechanisms of immunosuppression in immune subsets of interest were further explored using in vitro hypoxia assays. Regulatory T cells (Tregs) and a number of immunosuppressive myeloid subsets, including M2 macrophages and human leukocyte antigen-DR isotype (HLA-DRlo ) type 2 conventional dendritic cell (cDC2), were found to be significantly enriched in hypoxia-high tumor regions. On the other hand, the abundance of active granzyme Bhi PD-1lo CD8+ T cells in hypoxia-low tumor regions implied a relatively active immune landscape compared with hypoxia-high regions. The up-regulation of cancer-associated genes in the tumor tissues and immunosuppressive genes in the tumor-infiltrating leukocytes supported a highly pro-tumorigenic network in hypoxic HCC. Chemokine genes such as CCL20 (C-C motif chemokine ligand 20) and CXCL5 (C-X-C motif chemokine ligand 5) were associated with recruitment of both Tregs and HLA-DRlo cDC2 to hypoxia-high microenvironments. The interaction between Tregs and cDC2 under a hypoxic TME resulted in a loss of antigen-presenting HLA-DR on cDC2. CONCLUSIONS: We uncovered the unique immunosuppressive landscapes and identified key immune subsets enriched in hypoxic HCC. In particular, we identified a potential Treg-mediated immunosuppression through interaction with a cDC2 subset in HCC that could be exploited for immunotherapies.

Multi-region sampling with paired sample sequencing analyses reveals sub-groups of patients with novel patient-specific dysregulation in Hepatocellular Carcinoma.

BACKGROUND: Conventional differential expression (DE) testing compares the grouped mean value of tumour samples to the grouped mean value of the normal samples, and may miss out dysregulated genes in small subgroup of patients. This is especially so for highly heterogeneous cancer like Hepatocellular Carcinoma (HCC). METHODS: Using multi-region sampled RNA-seq data of 90 patients, we performed patient-specific differential expression testing, together with the patients' matched adjacent normal samples. RESULTS: Comparing the results from conventional DE analysis and patient-specific DE analyses, we show that the conventional DE analysis omits some genes due to high inter-individual variability present in both tumour and normal tissues. Dysregulated genes shared in small subgroup of patients were useful in stratifying patients, and presented differential prognosis. We also showed that the target genes of some of the current targeted agents used in HCC exhibited highly individualistic dysregulation pattern, which may explain the poor response rate. DISCUSSION/CONCLUSION: Our results highlight the importance of identifying patient-specific DE genes, with its potential to provide clinically valuable insights into patient subgroups for applications in precision medicine.

Radiofrequency Ablation Versus Repeat Hepatectomy for Recurrent Hepatocellular Carcinoma: A Systematic Review and Meta-Analysis.

BACKGROUND: An updated systematic review and meta-analysis was conducted to compare radiofrequency ablation (RFA) versus repeat hepatectomy (RH) for patients with recurrent hepatocellular carcinoma (rHCC) after a previous liver resection. METHODS: PubMed, EMBASE, and Cochrane databases were searched from inception to October 2021 for randomized controlled trials and propensity-score matched studies. Individual participant survival data of disease-free survival (DFS) and overall survival (OS) were extracted and reconstructed followed by one-stage and two-stage meta-analysis. Secondary outcomes were major complications and length of hospital stay (LOHS). RESULTS: A total of seven studies (1317 patients) were analysed. In both one-stage and two-stage meta-analysis, there was no significant difference in OS between the RFA and RH cohorts (Hazard Ratio (HR) 1.15, 95% CI 0.98-1.36, P = 0.094 and HR 1.12, 95% CI 0.77-1.64, P = 0.474 respectively), while the RFA group had a higher hazard rate of disease recurrence compared to the RH group (HR 1.30, 95% CI 1.13-1.50, P < 0.001 and HR 1.31, 95% CI 1.09-1.57, P = 0.013, respectively). RFA was associated with fewer major complications and shorter LOHS versus RH (Odds Ratio 0.34, 95% CI 0.15-0.76, P = 0.009 and Weighted Mean Difference - 4.78, 95% CI - 6.30 to - 3.26, P < 0.001, respectively). CONCLUSIONS: RH may be associated with superior DFS for rHCC, at the expense of higher morbidity rate and longer LOHS. However, OS is comparable between both modalities. As such, these techniques may be utilized as complementary strategies depending on individual patient and disease factors. Large-scale, randomized, prospective studies are required to corroborate these findings.

Impact of liver cirrhosis on the difficulty of minimally-invasive liver resections: a 1:1 coarsened exact-matched controlled study.

INTRODUCTION: The impact of liver cirrhosis on the difficulty of minimal invasive liver resection (MILR) remains controversial and current difficulty scoring systems do not take in to account the presence of cirrhosis as a significant factor in determining the difficulty of MILR. We hypothesized that the difficulty of MILR is affected by the presence of cirrhosis. Hence, we performed a 1:1 matched-controlled study comparing the outcomes between patients undergoing MILR with and without cirrhosis including the Iwate system and Institut Mutualiste Montsouris (IMM) system in the matching process. METHODS: Between 2006 and 2019, 598 consecutive patients underwent MILR of which 536 met the study inclusion criteria. There were 148 patients with cirrhosis and 388 non-cirrhotics. One-to-one coarsened exact matching identified approximately exact matches between 100 cirrhotic patients and 100 non-cirrhotic patients. RESULTS: Comparison between MILR patients with cirrhosis and non-cirrhosis in the entire cohort demonstrated that patients with cirrhosis were associated with a significantly increased open conversion rate, transfusion rate, need for Pringles maneuver, postoperative, stay, postoperative morbidity and postoperative 90-day mortality. After 1:1 coarsened exact matching, MILR with cirrhosis were significantly associated with an increased open conversion rate (15% vs 6%, p = 0.03), operation time (261 vs 238 min, p < 0.001), blood loss (607 vs 314 mls, p = 0.002), transfusion rate (22% vs 9%, p = 0.001), need for application of Pringles maneuver (51% vs 34%, p = 0.010), postoperative stay (6 vs 4.5 days, p = 0.004) and postoperative morbidity (26% vs 13%, p = 0.029). CONCLUSION: The presence of liver cirrhosis affected both the intraoperative technical difficulty and postoperative outcomes of MILR and hence should be considered an important parameter to be included in future difficulty scoring systems for MILR.

Pre-operative Imaging Characteristics in Histology-Proven Resected Intrahepatic Cholangiocarcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC) are the most common primary liver cancers. With the increasing incidence of ICC over the past two decades in Asia, it is essential to differentiate between HCC and ICC. However, ICC may mimic the radiological appearance of HCC on computed tomography scans (CT) and magnetic resonance imaging (MRI), leading to misdiagnosis of ICC. The objective of this study is to evaluate and describe the association of specific pre-operative imaging characteristics (arterial enhancement, portal venous washout) in patients with histologically proven resected ICC in our centre. METHODS: Data on patients with histology-proven ICC and mixed hepatocellular-cholangiocarcinomas (HCC-CC) who had undergone surgical resection at Singapore General Hospital (SGH) were identified from a prospectively maintained database. Pre-operative cross-sectional imaging reports were analysed. RESULTS: Ninety-one patients underwent resection between 1 January 2000 and 31 December 2016. Among those with no risk factors for HCC, a significant percentage of patients with ICC (24.3%) show imaging characteristics of both arterial phase hyperenhancement and non-peripheral venous washout. Among patients with risk factors for HCC, between 20.0 and 33.3% of patients with pure ICC fulfilled the imaging criteria for HCC, and this proportion was generally even higher in the mixed HCC-CC group. CONCLUSIONS: A significant proportion of patients with pure ICC showed pre-operative imaging characteristics which fulfilled the diagnostic criteria for HCC. The differential of ICC should be borne in mind in populations where both malignancies are endemic.

Changing trends and outcomes associated with the adoption of minimally invasive pancreatic surgeries: A single institution experience with 150 consecutive procedures in Southeast Asia.

BACKGROUND: Minimally invasive pancreatic surgeries (MIPS) are increasingly adopted worldwide. However, it remains uncertain if these reported experiences are reproducible throughout the world today. This study examines the safety and evolution of MIPS at a single institution in Southeast Asia. METHODS: This is a retrospective review of 150 consecutive patients who underwent MIPS between 2006 and 2018 of which 135 cases (90%) were performed since 2012. To determine the evolution of MIPS, the study population was stratified into 3 equal groups of 50 patients. Comparison was also made between pancreatoduodenectomies (PD), distal pancreatectomies (DP) and other pancreatic surgeries. RESULTS: One hundred and fifty patients underwent MIPS (103 laparoscopic, 45 robotic and 2 hand-assisted). Forty-three patients underwent PD, 93 DP and 14 other MIPS. There were 21 (14.0%) open conversions. There was an exponential increase in caseload over the study period. Comparison across the 3 time periods demonstrated that patients were significantly more likely to have a higher American Society of Anesthesiologists score, older, undergo PD and a longer operation time. The conversion rate decreased from 28% to 0% and increased again to 14% across the 3 time periods. Comparison between the various types of MIPS demonstrated that patients who underwent PD were significantly older, more likely to have symptomatic tumours, had longer surgery time, increased blood loss, increased frequency of extended pancreatectomies, increased frequency of hybrid procedures, longer post-operative stay, increased post-operative morbidity rate and increased post-operative major morbidity rate. CONCLUSION: The case volume of MIPS increased rapidly at our institution over the study period. Furthermore, although the indications for MIPS expanded to include more complex procedures in higher risk patients, there was no change in key perioperative outcomes.

Critical Appraisal of the Impact of the Systematic Adoption of Advanced Minimally Invasive Hepatobiliary and Pancreatic Surgery on the Surgical Management of Mirizzi Syndrome.

BACKGROUND: Minimally invasive surgery (MIS) for Mirizzi syndrome (MS) remains a technically challenging procedure with a high open conversion rate. We critically evaluated the impact of the systematic adoption of MI-HBP surgery on the surgical outcomes of MS. METHODS: Ninety-five patients who underwent surgery for MS were retrospectively reviewed. Systematic adoption of advanced MI-HBP surgery started in 2012. The cohort was classified into a preadoption (2002-2012) (Era 1, n = 58) and post-adoption (2013-2017) (Era 2, n = 37). Furthermore, Era 2 was divided into a cohort operated by advanced minimally invasive surgeons (AMIS) (Era 2 AMIS, n = 19) and those by other surgeons (Era 2 others, n = 19). RESULTS: Comparison between Era 2 and Era 1 demonstrated a significant increase in the frequency of MIS attempted (89% vs 33%, p < 0.01), increase in the use of choledochoplasty (24% vs 2%, p < 0.01), increase operation time (180 min vs 150 min, p = 0.03) and significantly lower open conversion rate (24% vs 58%, p < 0.01). Comparison between Era 2 AMIS and Era 2 others demonstrated a significantly greater adoption of MIS (100% vs 78%, p = 0.046) with lower open conversion rate (5% vs 50%, p = 0.005). Comparison between all attempted MIS cases with open procedures demonstrated a significantly higher proportion of subtotal cholecystectomies performed (40% vs 23%, p = 0.04), choledochoplasty (17% vs 2%, p = 0.04) and shorter hospital stay (4 days vs 9 days, p < 0.01). CONCLUSIONS: Systematic adoption of advanced MI-HBP surgery allowed surgeons to perform MIS for MS more frequently and with a significantly lower open conversion rate. Patients who underwent successful MIS had the shortest hospital stay compared to patients who underwent open surgery or required open conversion.

Preoperative Predictors Including the Role of Inflammatory Indices in Predicting Early Recurrence After Re-resection for Recurrent Hepatocellular Carcinoma.

BACKGROUND: Repeat liver resection (RLR) for recurrent HCC (rHCC) is a widely accepted treatment modality. However, early recurrence rate is high, frequently resulting in futile resection. We performed this study to evaluate preoperative factors, including the value of inflammatory indices, in predicting early (<1 year) recurrence in patients who underwent RLR for rHCC. This may help clinicians better select patients for RLR, while excluding cases in which RLR for rHCC would likely be futile. METHODS: This is a retrospective study of 80 patients where 90 operative cases of RLR and 84 cases of early recurrence (<1 year) post-RLR were evaluated. Preoperative predictors of early recurrence and overall survival (OS) were assessed. RESULTS: There were 31 (34.4%) early recurrences with a 5-year OS of 38.9%. Elevated platelet-to-lymphocyte ratio (PLR) >103.6 was a significant independent preoperative predictor of both early recurrence, relative risk (RR) 4.284 (P = 0.001) and OS, RR 2.139 (P = 0.027), while alphafetoprotein (AFP) ≥ 200 was a significant independent preoperative predictor of early recurrence only, RR 11.655 (P = 0.030). Patients were followed-up at a median of 14.3 months with 54.8% developing intrahepatic recurrences and 19.4% developing extrahepatic recurrences. CONCLUSION: Both, elevated PLR and AFP ≥ 200 were independent predictors of early (<1 year) recurrence after RLR for rHCC, while only an elevated PLR was an independent preoperative prognosticators of overall survival. Indication for RLR should be carefully discussed in patients with relapsed HCC with an elevated PLR, due to the potential of early recurrence and poor overall survival.

Critical appraisal of the impact of individual surgeon experience on the outcomes of laparoscopic liver resection in the modern era: collective experience of multiple surgeons at a single institution with 324 consecutive cases.

BACKGROUND: Most studies analyzing the learning experience of laparoscopic liver resection (LLR) focused on the experience of one or two expert pioneering surgeons. This study aims to critically analyze the impact of individual surgeon experience on the outcomes of LLR based on the contemporary collective experiences of multiple surgeons at single institution. METHODS: Retrospective review of 324 consecutive LLR from 2006 to 2016. The cases were performed by 10 surgeons over various time periods. Four surgeons had individual experience with <20 cases, four surgeons with 20-30 cases, and two surgeons with >90 cases. The cohort was divided into two groups: comparing a surgeon's experience between the first 20, 30, 40, and 50 cases with patients treated thereafter. Similarly, we performed subset analyses for anterolateral lesions, posterosuperior lesions, and major hepatectomies. RESULTS: As individual surgeons gained increasing experience, this was significantly associated with older patients being operated, decreased hand-assistance, larger tumor size, increased liver resections, increased major resections, and increased resections of tumors located at the posterosuperior segments. This resulted in significantly longer operation time and increased use of Pringle maneuver but no difference in other outcomes. Analysis of LLR for tumors in the posterosuperior segments demonstrated that there was a significant decrease in conversion rates after a surgeon had experience with 20 LLR. For major hepatectomies, there was a significant decrease in morbidity, mortality, and length of stay after acquiring experience with 20 LLR. CONCLUSION: LLR can be safely adopted today especially for lesions in the anterolateral segments. LLR for lesions in the difficult posterosuperior segments and major hepatectomies especially in cirrhosis should only be attempted by surgeons who have acquired a minimum experience with 20 LLR.

Changing trends and outcomes associated with the adoption of minimally invasive hepatectomy: a contemporary single-institution experience with 400 consecutive resections.

BACKGROUND: Several studies published mainly from pioneers and early adopters have documented the evolution of minimally invasive hepatectomy (MIH). However, questions remain if these reported experiences are applicable and reproducible today. This study examines the changing trends, safety, and outcomes associated with the adoption of MIH based on a contemporary single-institution experience. METHODS: This is a retrospective review of 400 consecutive patients who underwent MIH between 2006 and 2017 of which 360 cases (90%) were performed since 2012. To determine the evolution of MIH, the study population was stratified into four equal groups of 100 patients. Analyses were also performed of predictive factors and outcomes of open conversion. RESULTS: Four hundred patients underwent MIH of which 379 (94.8%) were totally laparoscopic/robotic. Eighty-eight (22.0%) patients underwent major hepatectomy and 160 (40.0%) had resection of tumors located in the posterosuperior segments. There were 38 (9.5%) open conversions. Comparison across the four groups demonstrated that patients were older, had higher ASA score, and had increased frequency of previous abdominal surgery and repeat liver resections. There was also an increase in the proportion of patients who underwent totally laparoscopic/robotic surgery, major liver resection, resection of ≥ 3 segments, and multiple resections. Comparison of outcomes demonstrated that there was a significant decrease in open conversion rate, longer operation time, and increased use of Pringles maneuver. The presence of cirrhosis and institution experience (1st 100 cases) were independent predictors of open conversion. Patients who required open conversion had significantly increased operation time, blood loss, blood transfusion rate, morbidity, and mortality. CONCLUSION: The case volume of MIH performed increased rapidly at our institution over time. Although the indications of MIH expanded to include higher risk patients and more complex hepatectomies, there was a decrease in open conversion rate and no change in other perioperative outcomes.

Evolution of minimally invasive distal pancreatectomies at a single institution.

INTRODUCTION: This study aims to study the changing trends and outcomes associated with the adoption of minimally invasive distal pancreatectomy (MIDP) at a single centre. MATERIALS AND METHODS: Retrospective review of sixty consecutive patients who underwent MIDP from September 2006 to November 2016 at a single institution. To study the evolution of MIDP, the study population was divided into three groups consisting of twenty patients (Group I, Group II and Group III). RESULTS: Sixty patients underwent MIDP with 11 (18.3%) requiring open conversions. The median operation time was 305 (range: 85-775) min and the median post-operative stay was 6 (range: 3-73) days. Fifteen procedures were spleen-saving pancreatectomies. Major post-operative morbidity (>Grade 2) occurred in 12 (20.0%) patients and there was no mortality or reoperations. There were 33 (55.0%) pancreatic fistulas, of which 15 (25.0%) were Grade B fistulas of which 12 (20.0%) required percutaneous drainage. Comparison between the three groups demonstrated a statistically significant increase in the frequency of procedures performed, increase in robotic-assisted procedures and proportion of asymptomatic tumours resected. There also tended to be non-significant decrease in open conversion rates from 25% to 5% between the three groups and increase in tumour size resected from 24 to 40 mm. CONCLUSION: Comparison between the three groups demonstrated that MIDP was performed with increased frequency. There was a statistically significant increase in the frequency of resections performed for asymptomatic tumours and resections performed through robotic assistance. There was also a non-significant trend towards a decrease in open conversions and increase in the size of tumours resected.

Prediction of Hepatocellular Carcinoma Recurrence Beyond Milan Criteria After Resection: Validation of a Clinical Risk Score in an International Cohort.

OBJECTIVE: This study aims to validate a previously reported recurrence clinical risk score (CRS). SUMMARY OF BACKGROUND DATA: Salvage transplantation after hepatocellular carcinoma (HCC) resection is limited to patients who recur within Milan criteria (MC). Predicting recurrence patterns may guide treatment recommendations. METHODS: An international, multicenter cohort of R0 resected HCC patients were categorized by MC status at presentation. CRS was calculated by assigning 1 point each for initial disease beyond MC, multinodularity, and microvascular invasion. Recurrence incidences were estimated using competing risks methodology, and conditional recurrence probabilities were estimated using the Bayes theorem. RESULTS: From 1992 to 2015, 1023 patients were identified, of whom 613 (60%) recurred at a median follow-up of 50 months. CRS was well validated in that all 3 factors remained independent predictors of recurrence beyond MC (hazard ratio 1.5-2.1, all P < 0.001) and accurately stratified recurrence risk beyond MC, ranging from 19% (CRS 0) to 67% (CRS 3) at 5 years. Among patients with CRS 0, no other factors were significantly associated with recurrence beyond MC. The majority recurred within 2 years. After 2 years of recurrence-free survival, the cumulative risk of recurrence beyond MC within the next 5 years for all patients was 14%. This risk was 12% for patients with initial disease within MC and 17% for patients with initial disease beyond MC. CONCLUSIONS: CRS accurately predicted HCC recurrence beyond MC in this international validation. Although the risk of recurrence beyond MC decreased over time, it never reached zero.

Significance of neutrophil-to-lymphocyte ratio, platelet-to-lymphocyte ratio and prognostic nutrition index as preoperative predictors of early mortality after liver resection for huge (≥10 cm) hepatocellular carcinoma.

BACKGROUND: This study aimed to determine preoperative predictors of early (<1 year) mortality from disease recurrence after liver resection (LR) for huge (≥10 cm) HCC, with special emphasis on the importance of neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), and prognostic nutrition index (PNI). METHODS: Between 2000 to 2013, 166 patients underwent LR for huge HCC. Optimal cut-offs for alpha fetoprotein (AFP), NLR, PLR, and PNI were determined by plotting the receiver operator curves (ROC) in predicting early mortality and utilizing the Youden index. RESULTS: The 30-day/in-hospital postoperative mortality rate was 4.2%. The 5-year overall survival (OS) and the 5-year recurrence-free survival (RFS) was 43% and 24%, respectively. Early mortality from disease recurrence occurred in 35 of 159 (22%) patients. Multivariate analyses demonstrated that tumor rupture and high AFP (>1,085 ng/ml) were independent preoperative predictors of early mortality after LR for HCC, and both a low PNI (<41) and high AFP were independent predictors of early mortality for non-ruptured HCC. In 51 patients who had none of these three factors, only four (7.8%) patients experienced early mortality from disease recurrence. CONCLUSIONS: Spontaneous rupture, high AFP, and low PNI were predictors of early mortality from disease recurrence after LR for huge HCC. J. Surg. Oncol. 2016;113:621-627. © 2016 Wiley Periodicals, Inc.

Histone-lysine N-methyltransferase EHMT2 (G9a) inhibition mitigates tumorigenicity in Myc-driven liver cancer.

Hepatocellular carcinoma (HCC) is the third deadliest and sixth most common cancer in the world. Histone-lysine N-methyltransferase EHMT2 (also known as G9a) is a histone methyltransferase frequently overexpressed in many cancer types, including HCC. We showed that Myc-driven liver tumours have a unique H3K9 methylation pattern with corresponding G9a overexpression. This phenomenon of increased G9a was further observed in our c-Myc-positive HCC patient-derived xenografts. More importantly, we showed that HCC patients with higher c-Myc and G9a expression levels portend a poorer survival with lower median survival months. We demonstrated that c-Myc interacts with G9a in HCC and cooperates to regulate c-Myc-dependent gene repression. In addition, G9a stabilises c-Myc to promote cancer development, contributing to the growth and invasive capacity in HCC. Furthermore, combination therapy between G9a and synthetic-lethal target of c-Myc, CDK9, demonstrates strong efficacy in patient-derived avatars of Myc-driven HCC. Our work suggests that targeting G9a could prove to be a potential therapeutic avenue for Myc-driven liver cancer. This will increase our understanding of the underlying epigenetic mechanisms of aggressive tumour initiation and lead to improved therapeutic and diagnostic options for Myc-driven hepatic tumours.

IFNγ-IL-17+ CD8 T cells contribute to immunosuppression and tumor progression in human hepatocellular carcinoma.

IL-17-producing CD8 (Tc17) T cells have been shown to play an important role in infection and chronic inflammation, however their implications in hepatocellular carcinoma (HCC) remain elusive. In this study, we performed cytometry by time-of-flight (CyTOF) and revealed the distinctive immunological phenotypes of two IFNγ+ and IFNγ- Tc17 subsets that were preferentially enriched in human HCC. Single-cell RNA-sequencing analysis further revealed regulatory circuits governing the different phenotypes of these Tc17 subsets. In particular, we discovered that IFNγ- Tc17 subset demonstrated pro-tumoral characteristics and expressed higher levels of CCL20. This corresponded to increased tumor infiltration of T regulatory cells (Treg) validated by immunohistochemistry in another independent HCC cohort, demonstrating the immunosuppressive functions of IFNγ- Tc17 subset. Most importantly, higher intra-tumoral proportions of IFNγ- Tc17 were associated with poorer prognosis in patients with HCC and this was further validated in The Cancer Genome Atlas (TCGA) HCC cohort. Taken together, this compendium of transcriptomic and proteomic data of Tc17 subsets sheds light on the immunosuppressive phenotypes of IFNγ- Tc17 and its implications in HCC progression.

A genomic enhancer signature associates with hepatocellular carcinoma prognosis.

Background & Aims: Lifestyle and environmental-related exposures are important risk factors for hepatocellular carcinoma (HCC), suggesting that epigenetic dysregulation significantly underpins HCC. We profiled 30 surgically resected tumours and the matched adjacent normal tissues to understand the aberrant epigenetic events associated with HCC. Methods: We identified tumour differential enhancers and the associated genes by analysing H3K27 acetylation (H3K27ac) chromatin immunoprecipitation sequencing (ChIP-seq) and Hi-C/HiChIP data from the resected tumour samples of 30 patients with early-stage HCC. This epigenome dataset was analysed with previously reported genome and transcriptome data of the overlapping group of patients from the same cohort. We performed patient-specific differential expression testing using multiregion sequencing data to identify genes that undergo both enhancer and gene expression changes. Based on the genes selected, we identified two patient groups and performed a recurrence-free survival analysis. Results: We observed large-scale changes in the enhancer distribution between HCC tumours and the adjacent normal samples. Many of the gain-in-tumour enhancers showed corresponding upregulation of the associated genes and vice versa, but much of the enhancer and gene expression changes were patient-specific. A subset of the upregulated genes was activated in a subgroup of patients' tumours. Recurrence-free survival analysis revealed that the patients with a more robust upregulation of those genes showed a worse prognosis. Conclusions: We report the genomic enhancer signature associated with differential prognosis in HCC. Findings that cohere with oncofoetal reprogramming in HCC were underpinned by genome-wide enhancer rewiring. Our results present the epigenetic changes in HCC that offer the rational selection of epigenetic-driven gene targets for therapeutic intervention or disease prognostication in HCC. Impact and Implications: Lifestyle and environmental-related exposures are the important risk factors of hepatocellular carcinoma (HCC), suggesting that tumour-associated epigenetic dysregulations may significantly underpin HCC. We profiled tumour tissues and their matched normal from 30 patients with early-stage HCC to study the dysregulated epigenetic changes associated with HCC. By also analysing the patients' RNA-seq and clinical data, we found the signature genes - with epigenetic and transcriptomic dysregulation - associated with worse prognosis. Our findings suggest that systemic approaches are needed to consider the surrounding cellular environmental and epigenetic changes in HCC tumours.

Outcomes of laparoscopic, robotic, and open pancreatoduodenectomy: A network meta-analysis of randomized controlled trials and propensity-score matched studies.

BACKGROUND: This network meta-analysis was performed to determine the optimal surgical approach for pancreatoduodenectomy by comparing outcomes after laparoscopic pancreatoduodenectomy, robotic pancreatoduodenectomy and open pancreatoduodenectomy. METHODS: A systematic search of the PubMed, EMBASE, Scopus, and Web of Science databases was conducted to identify eligible randomized controlled trials and propensity-score matched studies. RESULTS: Four randomized controlled trials and 23 propensity-score matched studies comprising a total of 4,945 patients were included for analysis. Operation time for open pancreatoduodenectomy was shorter than both laparoscopic pancreatoduodenectomy (mean difference -57.35, 95% CI 26.25-88.46 minutes) and robotic pancreatoduodenectomy (mean difference -91.08, 95% CI 48.61-133.56 minutes), blood loss for robotic pancreatoduodenectomy was significantly less than both laparoscopic pancreatoduodenectomy (mean difference -112.58, 95% CI 36.95-118.20 mL) and open pancreatoduodenectomy (mean difference -209.87, 95% CI 140.39-279.36 mL), both robotic pancreatoduodenectomy and laparoscopic pancreatoduodenectomy were associated with reduced rates of delayed gastric emptying compared with open pancreatoduodenectomy (odds ratio 0.59, 95% CI 0.39-0.90 and odds ratio 0.69, 95% CI 0.50-0.95, respectively), robotic pancreatoduodenectomy was associated with fewer wound infections compared with open pancreatoduodenectomy (odds ratio 0.35, 95% CI 0.18-0.71), and laparoscopic pancreatoduodenectomy patients enjoyed significantly shorter length of stay compared with open pancreatoduodenectomy (odds ratio 0.43, 95% CI 0.28-0.95). There were no differences in other outcomes. CONCLUSION: This network meta-analysis of high-quality studies suggests that when laparoscopic pancreatoduodenectomy and robotic pancreatoduodenectomy are performed in high-volume centers, short-term perioperative and oncologic outcomes are largely comparable, if not slightly improved, compared with traditional open pancreatoduodenectomy. These findings should be corroborated in further prospective randomized studies.