Integrating the new systemic treatment landscape and surgical therapy in hepatocellular carcinoma.

The treatment landscape of hepatocellular carcinoma has evolved rapidly within the last decade. Minimally-invasive techniques have reached a new level of safety, affording surgeons to pursue more aggressive treatment strategies to ultimately improve oncological outcomes. These procedures have been increasingly applied to treat patients with more progressed tumors and in select case even patients with advanced stage disease confined to the liver. Concomitantly, a dramatic increase in research into immunotherapy has altered the treatment paradigm in advanced disease stages, where the emerging treatment regimens can provide durable responses in a subset of the patient population for whom prognosis is dramatically improved. These treatments are now tested in early-stage disease to address the pressing unmet need of high recurrence rates after resection and in intermediate stage to complement the proven efficacy of intraarterial embolization in delaying progression. This review provides an in-depth discussion of these trends and describes how the treatment landscape has already changed and which impediments remain.

Short-term postoperative outcomes of lymphadenectomy for cholangiocarcinoma, hepatocellular carcinoma and colorectal liver metastases in the modern era of liver surgery: Insights from the StuDoQ|Liver registry.

INTRODUCTION: The clinical role of lymphadenectomy (LAD) as part of hepatic resection for malignancies of the liver remains unclear. In this study, we aimed to report on the use cases and postoperative outcomes of liver resection and simultaneous LAD for hepatic malignancies (HM). MATERIALS AND METHODS: Clinicopathological data from patients who underwent surgery at 13 German centers from 2017 to 2022 (n = 3456) was extracted from the StuDoQ|Liver registry of the German Society of General and Visceral Surgery. Propensity-score matching (PSM) was performed to account for the extent of liver resection and patient demographics. RESULTS: LAD was performed in 545 (16%) cases. The most common indication for LAD was cholangiocarcinoma (CCA), followed by colorectal liver metastases (CRLM) and hepatocellular carcinoma (HCC). N+ status was found in 7 (8%), 59 (35%), and 56 cases (35%) for HCC, CCA, and CRLM, respectively (p < 0.001). The LAD rate was highest for robotic-assisted resections (28%) followed by open (26%) and laparoscopic resections (13%), whereas the number of resected lymph nodes was equivalent between the techniques (p = 0.303). LAD was associated with an increased risk of liver-specific postoperative complications, especially for patients with HCC. CONCLUSION: In this multicenter registry study, LAD was found to be associated with an increased risk of liver-specific complications. The highest rate of LAD was observed among robotic liver resections.

Robotic versus laparoscopic versus open major hepatectomy - an analysis of costs and postoperative outcomes in a single-center setting.

PURPOSE: In the era of minimal-invasive surgery, the introduction of robotic liver surgery (RS) was accompanied by concerns about the increased financial expenses of the robotic technique in comparison to the established laparoscopic (LS) and conventional open surgery (OS). Therefore, we aimed to evaluate the cost-effectiveness of RS, LS and OS for major hepatectomies in this study. METHODS: We analyzed financial and clinical data on patients who underwent major liver resection for benign and malign lesions from 2017 to 2019 at our department. Patients were grouped according to the technical approach in RS, LS, and OS. For better comparability, only cases stratified to the Diagnosis Related Groups (DRG) H01A and H01B were included in this study. Financial expenses were compared between RS, LS, and OS. A binary logistic regression model was used to identify parameters associated with increased costs. RESULTS: RS, LS and OS accounted for median daily costs of 1,725 €, 1,633 € and 1,205 €, respectively (p < 0.0001). Median daily (p = 0.420) and total costs (16,648 € vs. 14,578 €, p = 0.076) were comparable between RS and LS. Increased financial expenses for RS were mainly caused by intraoperative costs (7,592 €, p < 0.0001). Length of procedure (hazard ratio [HR] = 5.4, 95% confidence interval [CI] = 1.7-16.9, p = 0.004), length of stay (HR [95% CI] = 8.8 [1.9-41.6], p = 0.006) and development of major complications (HR [95% CI] = 2.9 [1.7-5.1], p < 0.0001) were independently associated with higher costs. CONCLUSIONS: From an economic perspective, RS may be considered a valid alternative to LS for major liver resections.

Biliary microbial patterns in primary sclerosing cholangitis are linked to poorer transplant-free survival.

BACKGROUND: Factors that determine individual disease course of patients with primary sclerosing cholangitis (PSC) are poorly understood. Although an association between gut microbes and disease outcome has been suggested, little is known about the role of microbes in the biliary tract. METHODS: We analyzed microbial cultures from bile specimens obtained during routine endoscopic retrograde cholangiopancreatography (ERCP) and intraoperatively before liver transplantation in 114 patients with PSC in our tertiary academic center. The presence of bacterial and fungal species was correlated with clinical characteristics and outcome data. RESULTS: A total of 87 patients (76%) had positive bile culture results. The presence of concomitant inflammatory bowel disease (IBD) was associated with positive bile culture results in multivariate analysis (OR, 4.707; 95% CI, 1.688-13.128; p=0.003). Enterococcus spp. in the bile was associated with a more frequent occurrence of liver transplantation and/or death (OR, 2.778; 95% CI, 1.147-6.728; p=0.021) and recurrent (≥3) cholangitis episodes (OR, 2.839; 95% CI, 1.037-7.768; p=0.037). Biliary candidiasis was linked to a higher frequency of recurrent (≥3) cholangitis episodes (OR, 5.677; 95% CI, 1.940-16.616; p=0.001). Proton pump inhibitor intake conferred a clinical feature associated with biliary candidiasis in multivariate analysis (OR, 3.559; 95% CI, 1.275-9.937; p=0.016). CONCLUSIONS: Our data indicate that in patients with PSC, presence of Enterococcus spp. and Candida spp. in bile is associated with an adverse outcome. Concomitant IBD is linked to presence of microbes in bile, and proton pump inhibitor intake is a feature associated with biliary candidiasis in patients with PSC.

Identification of IGF2 as Genomic Driver and Actionable Therapeutic Target in Hepatoblastoma.

Management of hepatoblastoma (HB), the most frequent pediatric liver cancer, is based on surgical resection and perioperative chemotherapy regimens. In this study, we aimed to identify actionable targets in HB and assess the efficacy of molecular therapies in preclinical models of HB. Paired tumor and adjacent tissues from 31 HBs and a validation set of 50 HBs were analyzed using RNA-seq, SNP, and methylation arrays. IGF2 overexpression was identified as the top targetable HB driver, present in 71% of HBs (22/31). IGF2high tumors displayed progenitor cell features and shorter recurrence-free survival. IGF2 overexpression was associated in 91% of cases with fetal promoter hypomethylation, ICR1 deregulation, 11p15.5 loss of heterozygosity or miR483-5p overexpression. The antitumor effect of xentuzumab (a monoclonal antibody targeting IGF1/2) alone or in combination with the conventional therapeutic agent cisplatin was assessed in HB cell lines, in PDX-derived HB organoids and in a xenograft HB murine model. The combination of xentuzumab with cisplatin showed strong synergistic antitumor effects in organoids and in IGF2high cell lines. In mice (n = 55), the combination induced a significant decrease in tumor volume and improved survival compared with cisplatin alone. These results suggest that IGF2 is an HB actionable driver and that, in preclinical models of HB, the combination of IGF1/2 inhibition with cisplatin induces superior antitumor effects than cisplatin monotherapy. Overall, our study provides a rationale for testing IGF2 inhibitors in combination with cisplatin in HB patients with IGF2 overexpression.

Inflamed and non-inflamed classes of HCC: a revised immunogenomic classification.

OBJECTIVE: We previously reported a characterisation of the hepatocellular carcinoma (HCC) immune contexture and described an immune-specific class. We now aim to further delineate the immunogenomic classification of HCC to incorporate features that explain responses/resistance to immunotherapy. DESIGN: We performed RNA and whole-exome sequencing, T-cell receptor (TCR)-sequencing, multiplex immunofluorescence and immunohistochemistry in a novel cohort of 240 HCC patients and validated our results in other cohorts comprising 660 patients. RESULTS: Our integrative analysis led to define: (1) the inflamed class of HCC (37%), which includes the previously reported immune subclass (22%) and a new immune-like subclass (15%) with high interferon signalling, cytolytic activity, expression of immune-effector cytokines and a more diverse T-cell repertoire. A 20-gene signature was able to capture ~90% of these tumours and is associated with response to immunotherapy. Proteins identified in liquid biopsies recapitulated the inflamed class with an area under the ROC curve (AUC) of 0.91; (2) The intermediate class, enriched in TP53 mutations (49% vs 29%, p=0.035), and chromosomal losses involving immune-related genes and; (3) the excluded class, enriched in CTNNB1 mutations (93% vs 27%, p<0.001) and PTK2 overexpression due to gene amplification and promoter hypomethylation. CTNNB1 mutations outside the excluded class led to weak activation of the Wnt-ßcatenin pathway or occurred in HCCs dominated by high interferon signalling and type I antigen presenting genes. CONCLUSION: We have characterised the immunogenomic contexture of HCC and defined inflamed and non-inflamed tumours. Two distinct CTNNB1 patterns associated with a differential role in immune evasion are described. These features may help predict immune response in HCC.

Novel microenvironment-based classification of intrahepatic cholangiocarcinoma with therapeutic implications.

OBJECTIVE: The diversity of the tumour microenvironment (TME) of intrahepatic cholangiocarcinoma (iCCA) has not been comprehensively assessed. We aimed to generate a novel molecular iCCA classifier that incorporates elements of the stroma, tumour and immune microenvironment ('STIM' classification). DESIGN: We applied virtual deconvolution to transcriptomic data from ~900 iCCAs, enabling us to devise a novel classification by selecting for the most relevant TME components. Murine models were generated through hydrodynamic tail vein injection and compared with the human disease. RESULTS: iCCA is composed of five robust STIM classes encompassing both inflamed (35%) and non-inflamed profiles (65%). The inflamed classes, named immune classical (~10%) and inflammatory stroma (~25%), differ in oncogenic pathways and extent of desmoplasia, with the inflammatory stroma showing T cell exhaustion, abundant stroma and KRAS mutations (p<0.001). Analysis of cell-cell interactions highlights cancer-associated fibroblast subtypes as potential mediators of immune evasion. Among the non-inflamed classes, the desert-like class (~20%) harbours the lowest immune infiltration with abundant regulatory T cells (p<0.001), whereas the hepatic stem-like class (~35%) is enriched in 'M2-like' macrophages, mutations in IDH1/2 and BAP1, and FGFR2 fusions. The remaining class (tumour classical: ~10%) is defined by cell cycle pathways and poor prognosis. Comparative analysis unveils high similarity between a KRAS/p19 murine model and the inflammatory stroma class (p=0.02). The KRAS-SOS inhibitor, BI3406, sensitises a KRAS-mutant iCCA murine model to anti-PD1 therapy. CONCLUSIONS: We describe a comprehensive TME-based stratification of iCCA. Cross-species analysis establishes murine models that align closely to human iCCA for the preclinical testing of combination strategies.

Molecular Markers of Response to Anti-PD1 Therapy in Advanced Hepatocellular Carcinoma.

BACKGROUND & AIMS: Single-agent anti-PD1 checkpoint inhibitors convey outstanding clinical benefits in a small fraction (∼20%) of patients with advanced hepatocellular carcinoma (aHCC) but the molecular mechanisms determining response are unknown. To fill this gap, we herein analyze the molecular and immune traits of aHCC in patients treated with anti-PD1. METHODS: Overall, 111 tumor samples from patients with aHCC were obtained from 13 centers before systemic therapies. We performed molecular analysis and immune deconvolution using whole-genome expression data (n = 83), mutational analysis (n = 72), and histologic evaluation with an endpoint of objective response. RESULTS: Among 83 patients with transcriptomic data, 28 were treated in frontline, whereas 55 patients were treated after tyrosine kinase inhibitors (TKI) either in second or third line. Responders treated in frontline showed upregulated interferon-γ signaling and major histocompatibility complex II-related antigen presentation. We generated an 11-gene signature (IFNAP), capturing these molecular features, which predicts response and survival in patients treated with anti-PD1 in frontline. The signature was validated in a separate cohort of aHCC and >240 patients with other solid cancer types where it also predicted response and survival. Of note, the same signature was unable to predict response in archival tissue of patients treated with frontline TKIs, highlighting the need for fresh biopsies before immunotherapy. CONCLUSION: Interferon signaling and major histocompatibility complex-related genes are key molecular features of HCCs responding to anti-PD1. A novel 11-gene signature predicts response in frontline aHCC, but not in patients pretreated with TKIs. These results must be confirmed in prospective studies and highlights the need for biopsies before immunotherapy to identify biomarkers of response.

Robotic Liver Surgery: Technical Advantages Over Laparoscopic Technique Based on Parameters of Surgical Complexity and Perioperative Outcomes.

Introduction: In view of the limited availability, our study addresses the issue of optimal case selection for robotic liver surgery over standard laparoscopy offering an in-detail analysis of intra- and postoperative outcomes. Materials and Methods: Clinical and technical data of all consecutive cases of robotic liver surgery of a single high-volume center from 2018 to 2020 were collected prospectively. Second, we performed a retrospective analysis of all laparoscopic liver resections from 2015 to 2020. Parameters of surgical complexity were extracted and descriptive analysis and statistical hypothesis testing were performed to assess parameters of intraoperative and postoperative outcomes. Results: A total of 121 robotic resections were compared with 435 laparoscopic resections. Shorter robotic operating times were shown for segmentectomies of the right liver lobe compared with laparoscopic procedures (P = .003) with an according trend for extended resections. A shorter duration of applied Pringle's maneuver was observed for robotic procedures. This advantage was further enhanced in cases with close proximity of the tumor to major vessels. There were no significant differences in postoperative morbidity and mortality between both groups. Conclusion: Our study offers the first in-detail analysis of intraoperative and postoperative outcomes of robotic liver surgery depending on established parameters of surgical complexity. The results indicate potential technical advantages of robotic technology in liver surgery based on parameters that can be studied before the operation. When evaluating robotic technology, future studies should focus not only at overall postoperative outcomes, but rather at potential technical intraoperative advantages to allow optimal case selection for robotic liver surgery. Clinical Trial Registration Number: DRKS00017229.

Hepatocellular Carcinoma in Mongolia Delineates Unique Molecular Traits and a Mutational Signature Associated with Environmental Agents.

PURPOSE: Mongolia has the world's highest incidence of hepatocellular carcinoma (HCC), with ∼100 cases/100,000 inhabitants, although the reasons for this have not been thoroughly delineated. EXPERIMENTAL DESIGN: We performed a molecular characterization of Mongolian (n = 192) compared with Western (n = 187) HCCs by RNA sequencing and whole-exome sequencing to unveil distinct genomic and transcriptomic features associated with environmental factors in this population. RESULTS: Mongolian patients were younger, with higher female prevalence, and with predominantly HBV-HDV coinfection etiology. Mongolian HCCs presented significantly higher rates of protein-coding mutations (121 vs. 70 mutations per tumor in Western), and in specific driver HCC genes (i.e., APOB and TSC2). Four mutational signatures characterized Mongolian samples, one of which was novel (SBS Mongolia) and present in 25% of Mongolian HCC cases. This signature showed a distinct substitution profile with a high proportion of T>G substitutions and was significantly associated with a signature of exposure to the environmental agent dimethyl sulfate (71%), a 2A carcinogenic associated with coal combustion. Transcriptomic-based analysis delineated three molecular clusters, two not present in Western HCC; one with a highly inflamed profile and the other significantly associated with younger female patients. CONCLUSIONS: Mongolian HCC has unique molecular traits with a high mutational burden and a novel mutational signature associated with genotoxic environmental factors present in this country.

CXCR2 inhibition enables NASH-HCC immunotherapy.

OBJECTIVE: Hepatocellular carcinoma (HCC) is increasingly associated with non-alcoholic steatohepatitis (NASH). HCC immunotherapy offers great promise; however, recent data suggests NASH-HCC may be less sensitive to conventional immune checkpoint inhibition (ICI). We hypothesised that targeting neutrophils using a CXCR2 small molecule inhibitor may sensitise NASH-HCC to ICI therapy. DESIGN: Neutrophil infiltration was characterised in human HCC and mouse models of HCC. Late-stage intervention with anti-PD1 and/or a CXCR2 inhibitor was performed in murine models of NASH-HCC. The tumour immune microenvironment was characterised by imaging mass cytometry, RNA-seq and flow cytometry. RESULTS: Neutrophils expressing CXCR2, a receptor crucial to neutrophil recruitment in acute-injury, are highly represented in human NASH-HCC. In models of NASH-HCC lacking response to ICI, the combination of a CXCR2 antagonist with anti-PD1 suppressed tumour burden and extended survival. Combination therapy increased intratumoural XCR1+ dendritic cell activation and CD8+ T cell numbers which are associated with anti-tumoural immunity, this was confirmed by loss of therapeutic effect on genetic impairment of myeloid cell recruitment, neutralisation of the XCR1-ligand XCL1 or depletion of CD8+ T cells. Therapeutic benefit was accompanied by an unexpected increase in tumour-associated neutrophils (TANs) which switched from a protumour to anti-tumour progenitor-like neutrophil phenotype. Reprogrammed TANs were found in direct contact with CD8+ T cells in clusters that were enriched for the cytotoxic anti-tumoural protease granzyme B. Neutrophil reprogramming was not observed in the circulation indicative of the combination therapy selectively influencing TANs. CONCLUSION: CXCR2-inhibition induces reprogramming of the tumour immune microenvironment that promotes ICI in NASH-HCC.

Treatment of Intrahepatic Cholangiocarcinoma-A Multidisciplinary Approach.

Intrahepatic cholangiocarcinoma (iCC) is distinguished as an entity from perihilar and distal cholangiocarcinoma and gallbladder carcinoma. Recently, molecular profiling and histopathological features have allowed further classification. Due to the frequent delay in diagnosis, the prognosis for iCC remains poor despite major technical advances and multimodal therapeutic approaches. Liver resection represents the therapeutic backbone and only curative treatment option, with the functional residual capacity of the liver and oncologic radicality being deciding factors for postoperative and long-term oncological outcome. Furthermore, in selected cases and depending on national guidelines, liver transplantation may be a therapeutic option. Given the often advanced tumor stage at diagnosis or the potential for postoperative recurrence, locoregional therapies have become increasingly important. These strategies range from radiofrequency ablation to transarterial chemoembolization to selective internal radiation therapy and can be used in combination with liver resection. In addition, adjuvant and neoadjuvant chemotherapies as well as targeted therapies and immunotherapies based on molecular profiles can be applied. This review discusses multimodal treatment strategies for iCC and their differential use.

Objective Response Predicts Survival in Advanced Hepatocellular Carcinoma Treated with Systemic Therapies.

PURPOSE: Because of the increased number of sequential treatments used for advanced hepatocellular carcinoma (HCC), there is a need for surrogate endpoints of overall survival (OS). We analyze whether objective response (OR) is an independent predictor and surrogate endpoint of OS. PATIENTS AND METHODS: A systematic review of randomized clinical trials (RCT) in advanced HCC published between 2010 and 2020 was conducted to explore OS surrogacy of OR by Response Evaluation Criteria in Solid Tumors (RECIST) and modified RECIST (mRECIST). In parallel, RCTs exploring the impact of OR on OS in a time-dependent multivariate analysis were integrated in a meta-analysis. RESULTS: Of 65 RCTs identified in advanced HCC, we analyzed 34 studies including 14,056 patients that reported OS and OR by either RECIST (n = 23), mRECIST (n = 5), or both (n = 6). When exploring surrogacy, the trial-level correlation between OR odds ratio and OS HR was R = 0.677 by mRECIST and R = 0.532 by RECIST. Meta-analysis of five RCTs assessing predictors of survival in multivariate analysis found that patients with OR by mRECIST presented a pooled HR for OS of 0.44 (95% confidence interval, 0.27-0.70; P < 0.001) compared with nonresponders. Responses to atezolizumab-bevacizumab had a greater impact on OS than tyrosine kinase inhibitor responses. CONCLUSIONS: OR-mRECIST is an independent predictor of OS in patients with advanced HCC. Although correlation of OR-mRECIST and OS is better than with OR-RECIST, the level of surrogacy is modest. Thus, it can be used as endpoint in proof-of-concept phase II trials, but the data do not support its use as a primary endpoint of phase III investigations assessing systemic therapies.

Safety of intermittent Pringle maneuver during minimally invasive liver resection in patients with hepatocellular carcinoma with and without cirrhosis.

PURPOSE: The aim of this study was to analyze the impact of minimally invasive intermittent Pringle maneuver (IPM) on postoperative outcomes in patients with hepatocellular carcinoma (HCC) and liver cirrhosis. METHODS: In this retrospective cohort study, we evaluated the safety of IPM in patients with HCC who underwent minimally invasive liver resection during five years at our center. Factors influencing the use of IPM were examined in univariate and multivariate regression analysis. Cases with use of IPM (IPM) and those without use of IPM (no IPM) were then compared regarding intraoperative and postoperative outcomes after propensity score matching (PSM) for surgical difficulty. RESULTS: One hundred fifty-one patients underwent liver resection for HCC at our center and met inclusion criteria. Of these, 73 patients (48%) received IPM with a median duration of 18 min (5-78). One hundred patients (66%) had confirmed liver cirrhosis. In multivariate analysis, patients with large tumors (≥ 3 cm) and difficult tumor locations (segments VII or VIII) were more likely to undergo IPM (OR 1.176, p = 0.043, and OR 3.243, p = 0.001, respectively). After PSM, there were no differences in intraoperative blood transfusion or postoperative complication rates between the IPM and no IPM groups. Neither did we observe any differences in the subgroup analysis for cirrhotic patients. Postoperative serum liver function tests were not affected by the use of IPM. CONCLUSIONS: Based on our findings, we conclude that the use of IPM in minimally invasive liver resection is safe and feasible for patients with HCC, including those with compensated liver cirrhosis.

Complexity-Adjusted Learning Curves for Robotic and Laparoscopic Liver Resection: A Word of Caution.

Background: Minimally invasive liver surgery (MILS) has a high variance in the type of resection and complexity, which has been underestimated in learning curve studies in the past. The aim of this work was to evaluate complexity-adjusted learning curves over time for laparoscopic liver resection (LLR) and robotic liver resection (RLR). Methods: Cumulative sum analysis (CUSUM) and complexity adjustment were performed using the Iwate score for LLR and RLR (n = 647). Lowest point of smoothed data was used to capture the cutoff of the increase in complexity. Data were collected retrospectively at the Department of Surgery of the Charité-Universitätsmedizin Berlin. Results: A total of 132 RLR and 514 LLR were performed. According to the complexity-adjusted CUSUM analysis, the initial learning phase was reached after 117 for LLR and 93 procedures for RLR, respectively. With increasing experience, the rate of (extended) right hemihepatectomy multiplied from 8.4% to 18.9% for LLR (P = 0.031) and from 21.6% to 58.3% for RLR (P < 0.001). Complication rates remained comparable between both episodes for LLR and RLR (T1 vs T2, P > 0.05). The complexity-adjusted CUSUM analysis demonstrated for blood transfusion, conversion, and operative time an increase during the learning phase (T1), while a steady state was reached in the following (T2). Conclusions: The learning phase for MILS after adjusting for complexity is about 4 times longer than assumed in previous studies, which should urge caution.

Unannotated small RNA clusters associated with circulating extracellular vesicles detect early stage liver cancer.

OBJECTIVE: Surveillance tools for early cancer detection are suboptimal, including hepatocellular carcinoma (HCC), and biomarkers are urgently needed. Extracellular vesicles (EVs) have gained increasing scientific interest due to their involvement in tumour initiation and metastasis; however, most extracellular RNA (exRNA) blood-based biomarker studies are limited to annotated genomic regions. DESIGN: EVs were isolated with differential ultracentrifugation and integrated nanoscale deterministic lateral displacement arrays (nanoDLD) and quality assessed by electron microscopy, immunoblotting, nanoparticle tracking and deconvolution analysis. Genome-wide sequencing of the largely unexplored small exRNA landscape, including unannotated transcripts, identified and reproducibly quantified small RNA clusters (smRCs). Their key genomic features were delineated across biospecimens and EV isolation techniques in prostate cancer and HCC. Three independent exRNA cancer datasets with a total of 479 samples from 375 patients, including longitudinal samples, were used for this study. RESULTS: ExRNA smRCs were dominated by uncharacterised, unannotated small RNA with a consensus sequence of 20 nt. An unannotated 3-smRC signature was significantly overexpressed in plasma exRNA of patients with HCC (p<0.01, n=157). An independent validation in a phase 2 biomarker case-control study revealed 86% sensitivity and 91% specificity for the detection of early HCC from controls at risk (n=209) (area under the receiver operating curve (AUC): 0.87). The 3-smRC signature was independent of alpha-fetoprotein (p<0.0001) and a composite model yielded an increased AUC of 0.93. CONCLUSION: These findings directly lead to the prospect of a minimally invasive, blood-only, operator-independent clinical tool for HCC surveillance, thus highlighting the potential of unannotated smRCs for biomarker research in cancer.

Immunomodulatory Effects of Lenvatinib Plus Anti-Programmed Cell Death Protein 1 in Mice and Rationale for Patient Enrichment in Hepatocellular Carcinoma.

BACKGROUND AND AIMS: Lenvatinib is an effective drug in advanced HCC. Its combination with the anti-PD1 (programmed cell death protein 1) immune checkpoint inhibitor, pembrolizumab, has generated encouraging results in phase Ib and is currently being tested in phase III trials. Here, we aimed to explore the molecular and immunomodulatory effects of lenvatinib alone or in combination with anti-PD1. APPROACH AND RESULTS: We generated three syngeneic models of HCC in C57BL/6J mice (subcutaneous and orthotopic) and randomized animals to receive placebo, lenvatinib, anti-PD1, or combination treatment. Flow cytometry, transcriptomic, and immunohistochemistry analyses were performed in tumor and blood samples. A gene signature, capturing molecular features associated with the combination therapy, was used to identify a subset of candidates in a cohort of 228 HCC patients who might respond beyond what is expected for monotherapies. In mice, the combination treatment resulted in tumor regression and shorter time to response compared to monotherapies (P < 0.001). Single-agent anti-PD1 induced dendritic and T-cell infiltrates, and lenvatinib reduced the regulatory T cell (Treg) proportion. However, only the combination treatment significantly inhibited immune suppressive signaling, which was associated with the TGFß pathway and induced an immune-active microenvironment (P < 0.05 vs. other therapies). Based on immune-related genomic profiles in human HCC, 22% of patients were identified as potential responders beyond single-agent therapies, with tumors characterized by Treg cell infiltrates, low inflammatory signaling, and VEGFR pathway activation. CONCLUSIONS: Lenvatinib plus anti-PD1 exerted unique immunomodulatory effects through activation of immune pathways, reduction of Treg cell infiltrate, and inhibition of TGFß signaling. A gene signature enabled the identification of ~20% of human HCCs that, although nonresponding to single agents, could benefit from the proposed combination.

Evidence-Based Management of Hepatocellular Carcinoma: Systematic Review and Meta-analysis of Randomized Controlled Trials (2002-2020).

BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related mortality, with a rapidly changing landscape of treatments. In the past 20 years, numerous randomized controlled trials (RCTs) have aimed at improving outcomes across disease stages. We aimed to analyze the current evidence and identify potential factors influencing response to therapies. METHODS: We conducted a systematic review of phase III RCTs (2002-2020) across disease stages. A meta-analysis was designed to examine the relationship between etiology and outcome after systemic therapies with either tyrosine-kinase inhibitor (TKI)/antiangiogenic or immune checkpoint inhibitor (ICI) therapy. RESULTS: Out of 10,100 studies identified, 76 were phase III RCTs. Among them, a rigorous screening algorithm identified 49 with high quality including a total of 22,113 patients undergoing adjuvant (n = 7) and primary treatment for early (n = 2), intermediate (n = 7), and advanced (first-line, n = 21; second-line, n = 12) stages of disease. Nine of these trials were positive, 6 treatments have been adopted in guidelines (sorafenib [2 RCTs], lenvatinib, atezolizumab+bevacizumab, regorafenib, cabozantinib and ramucirumab), but 2 were not (adjuvant CIK cells and sorafenib plus hepatic arterial infusion with FOLFOX). Meta-analysis of 8 trials including 3739 patients revealed ICI therapy to be significantly more effective in patients with viral hepatitis compared with nonviral-related HCC, whereas no differences related to etiology were observed in patients treated with TKI/anti-vascular endothelial growth factor. CONCLUSIONS: Among 49 high-quality RCTs conducted in HCC during 2002-2020, 9 resulted in positive results. A meta-analysis of systemic therapies suggests that immunotherapies may be more effective in viral etiologies.

Recurrence of Hepatocellular Carcinoma After Liver Transplantation is Associated with Episodes of Acute Rejections.

PURPOSE: The impact of acute rejection (AR) after liver transplantation (LT) for hepatocellular carcinoma (HCC) on patient outcome is uncertain. This aim of this study is to investigate whether AR is associated with HCC relapse and overall survival. PATIENTS AND METHODS: Patients undergoing LT for HCC between 2001 and 2015 were retrospectively analyzed with regard to histopathological proven AR within the median time until recurrence. Cox's regression analysis was conducted revealing risk factors for HCC recurrence. RESULTS: HCC recurred in 47 of 252 analyzed patients with a median time to recurrence of 20 months. Patients with AR (28.6%) had a significantly higher frequency of recurrence compared to patients without AR (13.0%, p=0.002). Multiple Cox regression analyses identified AR within 20 months to be an independent risk factor for HCC recurrence both as dichotomized (HR=2.91, 95%CI: 1.30-6.53; p=0.009) and as a continuous variable (HR=1.81, 95%CI: 1.28-2.54; p=0.001). HCC recurrence and AR were associated with higher grades of liver fibrosis one year after LT, when compared to patients without AR (p=0.019). CONCLUSION: Our results demonstrate an association of AR with HCC recurrence after LT with implications for intervals of monitoring in tumor surveillance. Graft fibrosis and immune mechanisms are potentially related and causal interactions are worth further investigation.

NASH limits anti-tumour surveillance in immunotherapy-treated HCC.

Hepatocellular carcinoma (HCC) can have viral or non-viral causes1-5. Non-alcoholic steatohepatitis (NASH) is an important driver of HCC. Immunotherapy has been approved for treating HCC, but biomarker-based stratification of patients for optimal response to therapy is an unmet need6,7. Here we report the progressive accumulation of exhausted, unconventionally activated CD8+PD1+ T cells in NASH-affected livers. In preclinical models of NASH-induced HCC, therapeutic immunotherapy targeted at programmed death-1 (PD1) expanded activated CD8+PD1+ T cells within tumours but did not lead to tumour regression, which indicates that tumour immune surveillance was impaired. When given prophylactically, anti-PD1 treatment led to an increase in the incidence of NASH-HCC and in the number and size of tumour nodules, which correlated with increased hepatic CD8+PD1+CXCR6+, TOX+, and TNF+ T cells. The increase in HCC triggered by anti-PD1 treatment was prevented by depletion of CD8+ T cells or TNF neutralization, suggesting that CD8+ T cells help to induce NASH-HCC, rather than invigorating or executing immune surveillance. We found similar phenotypic and functional profiles in hepatic CD8+PD1+ T cells from humans with NAFLD or NASH. A meta-analysis of three randomized phase III clinical trials that tested inhibitors of PDL1 (programmed death-ligand 1) or PD1 in more than 1,600 patients with advanced HCC revealed that immune therapy did not improve survival in patients with non-viral HCC. In two additional cohorts, patients with NASH-driven HCC who received anti-PD1 or anti-PDL1 treatment showed reduced overall survival compared to patients with other aetiologies. Collectively, these data show that non-viral HCC, and particularly NASH-HCC, might be less responsive to immunotherapy, probably owing to NASH-related aberrant T cell activation causing tissue damage that leads to impaired immune surveillance. Our data provide a rationale for stratification of patients with HCC according to underlying aetiology in studies of immunotherapy as a primary or adjuvant treatment.