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.

Early experience with pure laparoscopic donor hepatectomy: comparison with open donor hepatectomy and non-donor laparoscopic hepatectomy.

BACKGROUND: Pure laparoscopic donor hepatectomy (L-DH) has seen a rise in uptake in recent years following the popularization of minimally invasive modality for major hepatobiliary surgery. Our study aimed to determine the safety and compare the perioperative outcomes of L-DH with open donor hepatectomy (O-DH) and laparoscopic non donor hepatectomy (L-NDH) based on our single institution experience. METHODS: Eighty of 113 laparoscopic hemi-hepatectomies performed between 2015 and 2022 met study inclusion criteria. Of these, 11 were L-DH. PSM in a 1:2 ratio of L-DH versus L-NDH and 1:1 ratio of L-DH versus O-DH were performed, identifying patients with similar baseline clinicopathological characteristics. RESULTS: After 2:1 matching, the L-DH cohort were significantly younger (P < 0.001) and had lower ASA scores (P < 0.001) than the L-NDH cohort. L-DH was associated with a longer median operating time (P < 0.001) and shorter median postoperative stay (P < 0.001) than L-NDH. After 1:1 matching, there were no significant differences in baseline demographic between the L-DH and O-DH cohorts. L-DH was associated with lower median blood loss (P = 0.040) and shorter length of stay compared to O-DH (P = 0.004). There were no significant differences in recipient outcomes for both cohorts. CONCLUSION: L-DH can be adopted safely by surgeons experienced in L-NDH and ODH. It is associated with decreased blood loss and shorter length of stay compared to O-DH.

Pcdh19 mediates olfactory sensory neuron coalescence during postnatal stages and regeneration.

The mouse olfactory system regenerates constantly throughout life. While genes critical for the initial projection of olfactory sensory neurons (OSNs) to the olfactory bulb have been identified, what genes are important for maintaining the olfactory map during regeneration are still unknown. Here we show a mutation in Protocadherin 19 (Pcdh19), a cell adhesion molecule and member of the cadherin superfamily, leads to defects in OSN coalescence during regeneration. Surprisingly, lateral glomeruli were more affected and males in particular showed a more severe phenotype. Single cell analysis unexpectedly showed OSNs expressing the MOR28 odorant receptor could be subdivided into two major clusters. We showed that at least one protocadherin is differentially expressed between OSNs coalescing on the medial and lateral glomeruli. Moreover, females expressed a slightly different complement of genes from males. These features may explain the differential effects of mutating Pcdh19 on medial and lateral glomeruli in males and females.

The use and role of digital technology in learning health systems: A scoping review.

OBJECTIVE: The review aimed to identify which digital technologies are proposed or used within learning health systems (LHS) and to analyze the extent to which they support learning processes in LHS. MATERIALS AND METHODS: Multiple databases and grey literature were searched with terms related to LHS. Manual searches and backward searches of reference lists were also undertaken. The review considered publications from 2007 to 2022. Records focusing on LHS, referring to one or more digital technologies, and describing how at least one digital technology could be used in LHS were included. RESULTS: 2046 records were screened for inclusion and 154 records were included in the analysis. Twenty categories of digital technology were identified. The two most common ones across records were data recording and processing and electronic health records. Digital technology was primarily leveraged to support data access and aggregation and data analysis, two of the seven recognized learning processes within LHS learning cycles. DISCUSSION: The results of the review show that a wide array of digital technologies is being leveraged to support learning cycles within LHS. Nevertheless, an over-reliance on a narrow set of technologies supporting knowledge discovery, a lack of direct evaluation of digital technologies and ambiguity in technology descriptions are hindering the realization of the LHS vision. CONCLUSION: Future LHS research and initiatives should aim to integrate digital technology to support practice change and impact evaluation. The use of recognized evaluation methods for health information technology and more detailed descriptions of proposed technologies are also recommended.

CD38 marks the exhausted CD8+ tissue-resident memory T cells in hepatocellular carcinoma.

Introduction: Despite recent advances in immunotherapy for hepatocellular carcinoma (HCC), the overall modest response rate underscores the need for a better understanding of the tumor microenvironment (TME) of HCC. We have previously shown that CD38 is widely expressed on tumor-infiltrating leukocytes (TILs), predominantly on CD3+ T cells and monocytes. However, its specific role in the HCC TME remains unclear. Methods: In this current study, we used cytometry time-of-flight (CyTOF), bulk RNA sequencing on sorted T cells, and single-cell RNA (scRNA) sequencing to interrogate expression of CD38 and its correlation with T cell exhaustion in HCC samples. We also employed multiplex immunohistochemistry (mIHC) for validating our findings. Results: From CyTOF analysis, we compared the immune composition of CD38-expressing leukocytes in TILs, non-tumor tissue-infiltrating leukocytes (NIL), and peripheral blood mononuclear cells (PBMC). We identified CD8+ T cells as the dominant CD38-expressing TILs and found that CD38 expression was significantly higher in CD8+ TRM in TILs than in NILs. Furthermore, through transcriptomic analysis on sorted CD8+ TRM from HCC tumors, we observed a higher expression of CD38 along with T cell exhaustion genes, including PDCD1 and CTLA4, compared to the circulating memory CD8 T cells from PBMC. This was validated by scRNA sequencing that revealed co-expression of CD38 with PDCD1, CTLA4, and ITGAE (CD103) in T cells from HCC tumors. The protein co-expression of CD38 and PD-1 on CD8+ T cells was further demonstrated by mIHC on HCC FFPE tissues, marking CD38 as a T cell co-exhaustion marker in HCC. Lastly, the higher proportions of CD38+PD-1+ CD8+ T cells and CD38+PD-1+ TRM were significantly associated with the higher histopathological grades of HCC, indicating its role in the aggressiveness of the disease. Conclusion: Taken together, the concurrent expression of CD38 with exhaustion markers on CD8+ TRM underpins its role as a key marker of T cell exhaustion and a potential therapeutic target for restoring cytotoxic T cell function in HCC.

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.

Preoperative Predictors of Early Recurrence After Liver Resection for Multifocal Hepatocellular Carcinoma.

BACKGROUND: Liver transplantation remains the optimal treatment for multifocal hepatocellular carcinoma (HCC). However, due to resource constrains, other therapeutic modalities such as liver resection (LR), are frequently utilized. LR, however, has to be balanced against potential morbidity and mortality along with the risks of early recurrence leading to futile surgery. In this study, we evaluated preoperative factors, including inflammatory indices, in predicting early (< 1 year) recurrence in patients who underwent LR for multifocal HCC. METHODS: This was a post hoc analysis of 250 consecutive patients with multifocal HCC who underwent LR. RESULTS: After exclusion of 10 patients with 30-day/in-hospital mortality, 240 were included of which 134 (55.8%) developed early recurrence. Hepatitis B/C aetiology, 3/ > more hepatic nodules and elevated alpha-fetoprotein (AFP) ≥ 200 ng/ml were significant independent preoperative predictors of early recurrence. The early recurrence rate was 72.1% when 2 out of 3 significant predictive factors were present. The conglomerate of all 3 factors predicted early recurrence of 100% with a statistically significant association between number of predictive factors and early recurrence (p < 0.001). CONCLUSION: Better patient selection via the use of preoperative predictive factors of early recurrence such as hepatitis B/C aetiology, ≥ 3 nodules and elevated AFP ≥ 200 ng/ml may assist in identifying patients in whom LR is deemed futile and improve resource allocation.

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.

Association of standardized liver volume and body mass index with outcomes of minimally invasive liver resections.

INTRODUCTION: While minimally invasive liver resections (MILR) have demonstrated advantages in improved post-operative recovery, widespread adoption is hampered by inherent technical difficulties. Our study attempts to analyze the role of anthropometric measures in MILR-related outcomes. METHODS: Between 2012 and 2020, 676 consecutive patients underwent MILR at the Singapore General Hospital of which 565 met study criteria and were included. Patients were stratified based on Body Mass Index (BMI) as well as Standardized Liver Volumes (SLV). Associations between BMI and SLV to selected peri-operative outcomes were analyzed using restricted cubic splines. RESULTS: A BMI of ≥ 29 was associated with increase in blood loss [Mean difference (MD) 69 mls, 95% CI 2 to 137] as well as operative conversions [Relative Risk (RR) 1.63, 95% CI 1.02 to 2.62] among patients undergoing MILR while a SLV of 1600 cc or higher was associated with an increase in blood loss (MD 30 mls, 95% CI 10 to 49). In addition, a BMI of ≤ 20 was associated with an increased risk of major complications (RR 2.25, 95% 1.16 to 4.35). The magnitude of differences observed in these findings increased with each unit change in BMI and SLV. CONCLUSION: Both BMI and SLV were useful anthropometric measures in predicting peri-operative outcomes in MILR and may be considered for incorporation in future difficulty scoring systems for MILR.

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.

Intraoperative blood transfusion does not impact overall and recurrence-free survival after curative hepatectomy for hepatocellular carcinoma: A propensity-score-matched and inverse probability of treatment-weighted study.

INTRODUCTION: Our primary objective was to determine if receiving intraoperative blood transfusion was a significant prognostic factor for overall and recurrence-free survival after curative resection of hepatic cellular carcinoma (HCC). METHODOLOGY: Between 2001 and 2018, 1092 patients with histologically proven primary HCC who underwent curative liver resection were retrospectively reviewed. Primary study endpoints were recurrence-free survival (RFS) and overall survival (OS). The main analysis was undertaken using propensity-score matching (PSM) to minimize confounding and selection biases in the comparison of patients with or without transfusion. RESULTS: There were 220 patients who received and 666 patients who did not receive intraoperative blood transfusion. The PSM cohort consisted of 163 pairs of patients. After PSM, the only perioperative outcome that appeared to significantly affect whether patients would receive blood transfusion was median blood loss (p = 0.001). In the PSM cohort, whether patients received blood transfusion was neither associated with OS (p = 0.759) nor RFS (p = 0.830). When the volume of blood transfusion was analyzed as a continuous variable, no significant dose-response relationship between blood transfusion volume and HR for OS and RFS was noted. CONCLUSION: Intraoperative blood transfusion had no significant impact on the survival outcomes in patients who receive curative resection in primary HCC.

Poor utility of current nomograms assessing the risk of intraoperative blood transfusion in patients undergoing liver resection for hepatocellular carcinoma and proposal of a new model.

BACKGROUND: The Memorial Sloan Kettering Cancer Center nomogram, the predictive scoring system of Yamamoto et al, and the 3-point transfusion risk score of Lemke et al are models used to determine the probability of receiving intraoperative blood transfusion in patients undergoing liver resection. However, the external validity of these models remains unknown. The objective of this study was to evaluate their predictive performance in an external cohort of patients with hepatocellular carcinoma. We also aimed to identify predictors of blood transfusion and develop a new predictive model for blood transfusion. METHODS: Post hoc analysis of our prospective database of 1,081 patients undergoing liver resection for hepatocellular carcinoma from 2001 to 2018. The predictive performance of current prediction models was evaluated using C statistics. Demographic and clinical variables as predictors of blood transfusion were assessed. Using logistic regression, an alternative model was created. RESULTS: The Lemke transfusion risk score performed better than the Memorial Sloan Kettering Cancer Center nomogram (0.69, 95% confidence interval 0.66-0.73 vs 0.66, 95% liver resection 0.62-0.69) (P < .001). The model from Yamamoto et al performed comparably with no statistically significant differences found through pairwise comparison. In our alternative model, hemoglobin level, albumin level, liver resection type, and tumor size were independent predictors of blood transfusion. The new HATS model obtained a C statistic of 0.74 (95% confidence interval 0.71-0.78), performing significantly better than the previous 3 models (P ≤ 0.001 for all). CONCLUSION: The existing Memorial Sloan Kettering Cancer Center, Yamamoto et al, and Lemke et al had nomograms with the suboptimal accuracy of predicting risk of intraoperative blood transfusion in patients undergoing liver resection for hepatocellular carcinoma. The proposed HATS model was more accurate at predicting patients at risk of blood transfusion.

Extending Surgical Resection for Hepatocellular Carcinoma Beyond Barcelona Clinic for Liver Cancer (BCLC) Stage A: A Novel Application of the Modified BCLC Staging System.

Objective: We aimed to prognosticate survival after surgical resection of HCC stratified by stage with amalgamation of the modified Barcelona Clinic Liver Cancer (BCLC) staging system and location of tumour. Methods: This single-institutional retrospective cohort study included patients with HCC who underwent surgical resection between 1st January 2000 to 30th June 2016. Participants were divided into 6 different subgroups: A-u) Within MC with Unilobar lesions; A-b) Within MC + Bilobar lesions; B1-u) Out of MC + within Up-To-7 + Unilobar lesions; B1-b) Out of MC + within Up-to-7 + Bilobar lesions; B2-u) Out of MC + Out of Up-To-7 + Unilobar lesions; B2-b) Out of MC + Out of Up-To-7 + Bilobar lesions. A separate survival analysis was conducted for solitary HCC lesions according to three subgroups: A-S (Within MC); B1-S (Out of MC + within Up-To-7); B2-S (Out of MC + out of Up-To-7). Results: A total of 794 of 1043 patients with surgical resection for HCC were analysed. Groups A-u (64.6%), A-b (58.4%) and B1-u (56.2%) had 5-year cumulative overall survival (OS) rates above 50% after surgical resection and median OS exceeding 60 months (P = 0.0001). The 5-year cumulative recurrence-free survival rates (RFS) were 40.4% (group A-u), 38.2% (group A-b), 36.3% (group B1-u), 24.6% (group B2-u), and 7.3% (group B2-b)(P=0.0001). For solitary lesions, the 5-year OS for the subgroups were A-S (65.1%), B1-S (56.0%) and B2-S (47.1%) (P = 0.0003). Compared to A-S, there was also a significant trend towards relatively poorer OS as the lesion sizes increased in B1-S (HR 1.46, 95% CI 1.03-2.08) and B2-S (HR 1.65, 95% CI 1.25-2.18). Conclusion: We adopted a novel approach combining the modified BCLC B sub-classification and dispersion of tumour to show that surgical resection in intermediate stage HCC can be robustly prognosticated. We found that size prognosticates resection outcomes in solitary tumours.

Laparoscopic versus open caudate lobe liver resections: a 1:2 propensity score-matched controlled study based on a single institution experience.

BACKGROUND: Despite the wide use of laparoscopy for liver resection, laparoscopic caudate lobe resections(L-CLR) remain technically challenging, only attempted by experts in the field. The primary objective of this study was to determine the safety and compare the perioperative outcomes of L-CLR with O-CLR based on our single institution experience in a 1:2 propensity score-matched controlled study based on our single institution experience. METHODS: Between 2004 and 2020, 67 consecutive patients who underwent CLR at Singapore General Hospital were identified. Propensity score matching (PSM) of laparoscopic versus open caudate lobe resections(O-CLR) was performed in a 1:2 ratio with no replacements using nearest neighbour matching method. RESULTS: L-CLR was associated with a significantly decreased median blood loss (150 mL versus 500 mL, P = 0.001) and a decreased median post-operative stay (3 days versus 7.5 days, P = <0.01) in the unmatched cohorts. After 1:2 propensity score matching, these results were again demonstrated with a significantly lower blood loss (150 mL versus 400 mL, P = 0.016) and a shorter postoperative stay (3 days versus 7 days, P = <0.01) in favour of L-CLR. 30-day readmission and major morbidity (Clavien-Dindo grade > 2) rates were all in favour of L-CLR as well but could not reach statistical significance. CONCLUSION: L-CLR can be safely performed by experienced surgeons. It is associated with decreased blood loss and shorter perioperative stay compared to O-CLR.

Dynamic phenotypic heterogeneity and the evolution of multiple RNA subtypes in hepatocellular carcinoma: the PLANET study.

Intra-tumor heterogeneity (ITH) is a key challenge in cancer treatment, but previous studies have focused mainly on the genomic alterations without exploring phenotypic (transcriptomic and immune) heterogeneity. Using one of the largest prospective surgical cohorts for hepatocellular carcinoma (HCC) with multi-region sampling, we sequenced whole genomes and paired transcriptomes from 67 HCC patients (331 samples). We found that while genomic ITH was rather constant across stages, phenotypic ITH had a very different trajectory and quickly diversified in stage II patients. Most strikingly, 30% of patients were found to contain more than one transcriptomic subtype within a single tumor. Such phenotypic ITH was found to be much more informative in predicting patient survival than genomic ITH and explains the poor efficacy of single-target systemic therapies in HCC. Taken together, we not only revealed an unprecedentedly dynamic landscape of phenotypic heterogeneity in HCC, but also highlighted the importance of studying phenotypic evolution across cancer types.

Trajectory of immune evasion and cancer progression in hepatocellular carcinoma.

Immune evasion is key to cancer initiation and later at metastasis, but its dynamics at intermediate stages, where potential therapeutic interventions could be applied, is undefined. Here we show, using multi-dimensional analyses of resected tumours, their adjacent non-tumour tissues and peripheral blood, that extensive immune remodelling takes place in patients with stage I to III hepatocellular carcinoma (HCC). We demonstrate the depletion of anti-tumoural immune subsets and accumulation of immunosuppressive or exhausted subsets along with reduced tumour infiltration of CD8 T cells peaking at stage II tumours. Corresponding transcriptomic modification occur in the genes related to antigen presentation, immune responses, and chemotaxis. The progressive immune evasion is validated in a murine model of HCC. Our results show evidence of ongoing tumour-immune co-evolution during HCC progression and offer insights into potential interventions to reverse, prevent or limit the progression of the disease.

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.

Effect of age on the short- and long-term outcomes of patients undergoing curative liver resection for HCC.

BACKGROUND: Few studies have evaluated the outcomes of curative liver resection (LR) in octogenarian patients, analysed cancer-specific survival (CSS) with HCC-related death or explored the age-varying effect of HCC-related death in elderly patients undergoing LR. We aim to determine the effect of age on the short and long-term outcomes of LR for HCC. METHODOLOGY: Between 2000 and 2018, 1,092 patients with primary HCC who underwent LR with curative intent were retrospectively reviewed. The log-rank test and Gray's test were used to assess the equality of survivor functions and competing risk-adjusted cumulative incidence functions between patients in the three age categories respectively. Regression adjustment was used to control for confounding bias via a Principal Component Analysis. Quantile, Firth logistic, Cox, and Fine-Gray competing risk regression were used to analyse continuous, binary, time-to-event, and cause-specific survival respectively. Restricted cubic splines were used to illustrate the dose-effect relationship between age and patient outcomes. RESULTS: The study comprised of 764 young patients (<70 years), 278 septuagenarians (70-79 years old) and 50 octogenarians (≥80 years). Compared to young patients, octogenarians had significantly lower 5-year OS(62.1% vs 37.7%, p < 0.001). However, there was no significant difference in 1-year RFS(73.1% vs 67.0%, p = 0.774) or 5-year CSS (5.4% vs 15.2%, p = 0.674). Every 10-year increase in age was significantly associated with an increase length of stay (p < 0.001), postoperative complications (p = 0.004) and poorer OS(p = 0.018) but not significantly associated with major complications (p = 0.279), CSS(p = 0.338) or RFS(p = 0.941). CONCLUSION: Age by itself was associated with OS after LR for HCC but was not a significant risk factor for HCC-related death.

Minimally Invasive vs Open Major Hepatectomies for Liver Malignancies: a Propensity Score-Matched Analysis.

BACKGROUND: The majority of evidence with regards to minimally invasive liver resection (MILR) favors its application in minor hepatectomies. We conducted a propensity score-matched (PSM) analysis to determine its feasibility and safety in major hepatectomies (MIMH) for liver malignancies. METHODS: Retrospective review of 130 patients who underwent MIMH and 490 patients who underwent open major hepatectomy (OMH) for malignant pathologies was performed. PSM in a 1:1 ratio identified two groups of patients with similar baseline clinicopathological characteristics. Perioperative outcomes were then compared. Major hepatectomies included traditional major (>3 segments) and technical major hepatectomies (right anterior and right posterior sectionectomies). RESULTS: Both cohorts were well-matched for baseline characteristics after PSM. Of 130 MIMH cases, there were 12 conversions to open. Comparison of perioperative outcomes demonstrated a significant association of MIMH with longer operation time and more frequent application of Pringle's maneuver (PM), but decreased postoperative stay. These results were consistent on a subgroup analysis that only included patients undergoing traditional major hepatectomies. A second subgroup analysis restricted to cirrhotic patients demonstrated that while perioperative outcomes were equivalent, MIMH was similarly associated with a longer operative time. Subset analyses of resections performed after 2015 demonstrated that MIMH was additionally associated with a lower postoperative morbidity compared to OMH. CONCLUSION: Comparison of perioperative and short-term oncological outcomes between MIMH and OMH for malignancies demonstrated that MIMH is feasible and safe. It is associated with a shorter hospital stay at the expense of a longer operation time compared to OMH.

Impact of non-liver-related previous abdominal surgery on the difficulty of minimally invasive liver resections: a propensity score-matched controlled study.

INTRODUCTION: The presence of previous abdominal surgery (PAS) has traditionally been considered to add difficulty to and increase risk of complications of laparoscopic procedures. This study aims to analyse the impact of non-liver-related PAS on the difficulty of minimally invasive liver resections (MILRs). MATERIALS AND METHODS: After exclusion of patients with concomitant major surgical procedures as well as previous liver resections, 515 consecutive patients undergoing MILR in Singapore General Hospital from 2006 to 2019 were analysed, consisting of 161 MILR in patients with previous abdominal surgery (WPAS) and 354 MILR in patients without previous abdominal surgery (WOPAS). Propensity score-matched (PSM) comparison was performed between WPAS and WOPAS groups. In addition, subgroup analysis was made comparing previous upper or lower abdominal surgery and open versus minimally invasive approach of PAS. Outcomes measured include those associated with operative difficulty such as open conversion rates, operative time, blood loss, as well as morbidity and mortality rates. RESULTS: MILR outcomes in patients WPAS are not inferior to those WOPAS. Overall open conversion rate was 8.2%, higher in patients WOPAS compared to patients WPAS (11.9% versus 3.5%, p = 0.015). Operating time (p = 0.942), blood loss (p = 0.063), intraoperative blood transfusion (p = 0.750), length of hospital stay (p = 0.206), morbidity (p = 0.217) and 30- and 90-day mortality (p = 1 & p = 0.367) were comparable between the two groups and subgroup analysis. CONCLUSION: Outcomes of MILR in patients with previous non-liver-related abdominal surgery are not inferior to patients without previous abdominal surgery.