Clinical Presentation and Outcome of Critically Ill Patients with Inflammatory Bowel Disease.

Introduction: Admission to the intensive care unit severely affects inflammatory bowel disease (IBD) patients. This study aimed to determine factors associated with mortality in IBD patients admitted to the intensive care unit. Methods: A retrospective cohort study was performed, analyzing data of all IBD patients admitted to the Department of Intensive Care Medicine at the University Medical Center Hamburg-Eppendorf between 2013 and 2022. Bivariate comparisons and multivariate regression analyses were performed to identify factors associated with mortality. Results: Overall, 439 IBD patients were admitted to the intensive care unit, representing 0.56% of total admissions. In 98 of these patients, IBD-associated complications were accountable for admission (22.3%). In detail, 39 (40.8%) patients were admitted after IBD-related surgery, 36 (35.7%) due to infections, and 23 (23.5%) due to medical conditions such as bleeding or electrolyte derangement. A total of 16 (16.3%) of these patients died within 90 days after admission. Parameters associated with increased mortality were age (p < 0.001), later age at diagnosis (p 0.026), catecholamine therapy (p 0.003), mechanical ventilation (p < 0.001), renal replacement therapy (p < 0.001), and parenteral nutrition (p 0.002). Prior treatment with anti-TNF therapy was associated with a higher chance of survival (p 0.018). There was no association between prior immunosuppressant therapy and admission because of infections (p 0.294). Conclusions: 16.3% of IBD patients admitted to the intensive care unit died within 90 days after admission. Prior treatment with anti-TNF therapy was associated with a higher chance of survival.

Mutual regulation of CD4+ T cells and intravascular fibrin in infections.

Innate myeloid cells especially neutrophils and their extracellular traps are known to promote intravascular coagulation and thrombosis formation in infections and various other conditions. Innate myeloid cell dependent fibrin formation can support systemic immunity while its dysregulation enhances the severity of infectious diseases. Less is known about the immune mechanisms preventing dysregulation of fibrin homeostasis in infection. During experimental systemic infections local fibrin deposits in the liver microcirculation cause rapid arrest of CD4+ T cells. Arrested T helper cells mostly represent Th17 cells that partially originate from the small intestine. Intravascular fibrin deposits activate mouse and human CD4+ T cells which can be mediated by direct fibrin - CD4+ T cell interactions. Activated CD4+ T cells suppress fibrin deposition and microvascular thrombosis by directly counteracting coagulation activation by neutrophils and classical monocytes. T cell activation, which is initially triggered by IL- 12p40- and MHC-II dependent mechanisms, enhances intravascular fibrinolysis via LFA-1. Moreover, CD4+ T cells disfavor the association of the fibrinolysis inhibitor TAFI with fibrin whereby fibrin deposition is increased by TAFI in the absence but not presence of T cells. In human infections thrombosis development is inversely related to microvascular levels of CD4+ T cells. Thus, fibrin promotes LFA-1 dependent T helper cell activation in infections which drives a negative feedback cycle that rapidly restricts intravascular fibrin and thrombosis development.

Apoptotic cell identity induces distinct functional responses to IL-4 in efferocytic macrophages.

Macrophages are functionally heterogeneous cells essential for apoptotic cell clearance. Apoptotic cells are defined by homogeneous characteristics, ignoring their original cell lineage identity. We found that in an interleukin-4 (IL-4)-enriched environment, the sensing of apoptotic neutrophils by macrophages triggered their tissue remodeling signature. Engulfment of apoptotic hepatocytes promoted a tolerogenic phenotype, whereas phagocytosis of T cells had little effect on IL-4-induced gene expression. In a mouse model of parasite-induced pathology, the transfer of macrophages conditioned with IL-4 and apoptotic neutrophils promoted parasitic egg clearance. Knockout of phagocytic receptors required for the uptake of apoptotic neutrophils and partially T cells, but not hepatocytes, exacerbated helminth infection. These findings suggest that the identity of apoptotic cells may contribute to the development of distinct IL-4-driven immune programs in macrophages.

IL-22 promotes liver regeneration after portal vein ligation.

Background: Insufficient remnant liver volume (RLV) after the resection of hepatic malignancy could lead to liver failure and mortality. Portal vein ligation (PVL) prior to hepatectomy is subsequently introduced to increase the remnant liver volume and improve the outcome of hepatic malignancy. IL-22 has previously been reported to promote liver regeneration, while facilitating tumor development in the liver via Steap4 upregulation. Here we performed PVL in mouse models to study the role of IL-22 in liver regeneration post-PVL. Methods: Liver weight and volume was measured via magnetic resonance imaging (MRI). Immunohistochemistry for Ki67 and hepatocyte growth factor (HGF) was performed. IL-22 was analyzed by flow cytometry and quantitative polymerase chain reaction (qPCR) was used for acquisition of Il-33, Steap4, Fga, Fgb and Cebpd. To analyze signaling pathways, mice with deletion of STAT3 and a neutralizing antibody for IL-22 were used. Results: The remnant liver weight and volume increased over time after PVL. Additionally, we found that liver regenerative molecules, including Ki67 and HGF, were significantly increased in remnant liver at day 3 post-PVL, as well as IL-22. Administration of IL-22 neutralizing antibody could reduce Ki67 expression after PVL. The upregulation of IL-22 after PVL was mainly derived from innate cells. IL-22 blockade resulted in lower levels of IL-33 and Steap4 in the remnant liver, which was also the case in mice with deletion of STAT3, the main downstream signaling molecule of IL-22, in hepatocytes. Conclusion: IL-22 promotes liver regeneration after PVL. Thus, a combination of IL-22 supplementation and Steap4 blockade could potentially be applied as a novel therapeutic approach to boost liver regeneration without facilitating tumor progression after PVL.

A protective effect of inflammatory bowel disease on the severity of sclerosing cholangitis.

Background: Primary sclerosing cholangitis (PSC) is a chronic liver disease marked by inflammation of the bile ducts and results in the development of strictures and fibrosis. A robust clinical correlation exists between PSC and inflammatory bowel disease (IBD). At present, published data are controversial, and it is yet unclear whether IBD drives or attenuates PSC. Methods: Mdr2-deficient mice or DDC-fed mice were used as experimental models for sclerosing cholangitis. Additionally, colitis was induced in mice with experimental sclerosing cholangitis, either through infection with Citrobacter rodentium or by feeding with DSS. Lastly, fibrosis levels were determined through FibroScan analysis in people with PSC and PSC-IBD. Results: Using two distinct experimental models of colitis and two models of sclerosing cholangitis, we found that colitis does not aggravate liver pathology, but rather reduces liver inflammation and liver fibrosis. Likewise, people with PSC-IBD have decreased liver fibrosis compared to those with PSC alone. Conclusions: We found evidence that intestinal inflammation attenuates liver pathology. This study serves as a basis for further research on the pathogenesis of PSC and PSC-IBD, as well as the molecular mechanism responsible for the protective effect of IBD on PSC development. This study could lead to the discovery of novel therapeutic targets for PSC.

Corrigendum: PD-1/PD-L1 immune checkpoint therapy demonstrates favorable safety profile in patients with autoimmune and cholestatic liver disease.

[This corrects the article DOI: 10.3389/fimmu.2023.1326078.].

All Three AKT Isoforms Can Upregulate Oxygen Metabolism and Lactate Production in Human Hepatocellular Carcinoma Cell Lines.

Hepatocellular carcinoma (HCC), the main pathological type of liver cancer, is related to risk factors such as viral hepatitis, alcohol intake, and non-alcoholic fatty liver disease (NAFLD). The constitutive activation of the PI3K/AKT signaling pathway is common in HCC and has essential involvement in tumor progression. The serine/threonine kinase AKT has several downstream substrates, which have been implicated in the regulation of cellular metabolism. However, the contribution of each of the three AKT isoforms, i.e., AKT1, AKT2 and AKT3, to HCC metabolism has not been comprehensively investigated. In this study, we analyzed the functional role of AKT1, AKT2 and AKT3 in HCC metabolism. The overexpression of activated AKT1, AKT2 and AKT3 isoforms in the human HCC cell lines Hep3B and Huh7 resulted in higher oxygen consumption rate (OCR), ATP production, maximal respiration and spare respiratory capacity in comparison to vector-transduced cells. Vice versa, lentiviral vector-mediated knockdowns of each AKT isoform reduced OCR in both cell lines. Reduced OCR rates observed in the three AKT isoform knockdowns were associated with reduced extracellular acidification rates (ECAR) and reduced lactate production in both analyzed cell lines. Mechanistically, the downregulation of OCR by AKT isoform knockdowns correlated with an increased phosphorylation of the pyruvate dehydrogenase on Ser232, which negatively regulates the activity of this crucial gatekeeper of mitochondrial respiration. In summary, our data indicate that each of the three AKT isoforms is able to upregulate OCR, ECAR and lactate production independently of each other in human HCC cells through the regulation of the pyruvate dehydrogenase.

Obesity and diabetes mellitus are associated with SARS-CoV-2 outcomes without influencing signature genes of extrapulmonary immune compartments at the RNA level.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) which is responsible for eliciting Coronavirus disease 2019 (COVID-19) still challenges healthcare services worldwide. While many patients only suffer from mild symptoms, patients with some pre-existing medical conditions are at a higher risk for a detrimental course of disease. However, the underlying mechanisms determining disease course are only partially understood. One key factor influencing disease severity is described to be immune-mediated. In this report, we describe a post-mortem analysis of 45 individuals who died from SARS-CoV-2 infection. We could show that although sociodemographic factors and premedical conditions such as obesity and diabetes mellitus reduced survival time in our cohort, they were not associated with changes in the expression of immune-related signature genes at the RNA level in the blood, the gut, or the liver between these different groups. Our data indicate that obesity and diabetes mellitus influence SARS-CoV-2-related mortality, without influencing the extrapulmonary gene expression of immunity-related signature genes at the RNA level.

Mouse models of spontaneous liver and lung metastasis for colorectal cancer.

To investigate underlying mechanisms for cancer metastasis and promising therapies in animal models, spontaneous metastasis models can be used to recreate metastasis development. Here, we present three mouse models of spontaneous lung and/or liver metastasis induction. We describe steps for cancer cell preparation, mouse analgesia, and three injection techniques (subcutaneous, intracecal, and intramucosal). We then detail procedures for evaluating metastasis. Most of these models generate metastasis in a time span of 4 weeks in the majority of injected mice. For complete details on the use and execution of this protocol, please refer to Giannou et al.1.

Protocol for generating lung and liver metastasis in mice using models that bypass intravasation.

Forced metastasis models, those in which the step of intravasation is bypassed, can be used to investigate the mechanisms underlying metastasis and evaluate potential therapeutic targets. Here, we present a protocol for using three forced models of lung and liver metastasis to generate metastasis within 3-4 weeks in approximately 99% of injected mice. We describe steps for cancer cell preparation, mouse analgesia and anesthesia; injecting through intrasplenic, intraportal, and intravenous techniques; and daily evaluation of metastasis. For complete details on the use and execution of this protocol, please refer to Giannou et al.1.

Intestinal epithelia and myeloid immune cells shape colitis severity and colorectal carcinogenesis via High-mobility group box protein 1.

BACKGROUND: HMGB1 is a ubiquitous nucleoprotein with immune-regulatory properties following cellular secretion or release in sterile and infectious inflammation. Stool and serum HMGB1 levels correlate with colitis severity and colorectal cancer (CRC) progression, yet recent reports indicated HMGB1 to mainly operate as an intracellular determinant of enterocyte fate during colitis, and investigations into the roles of HMGB1 in CRC are lacking. Using mice with conditional HMGB1-knockout in enterocytes (Hmgb1ΔIEC) and myeloid cells (Hmgb1ΔLysM), respectively, we explored functions of HMGB1 in pathogenetically diverse contexts of colitis and colitis-associated CRC. RESULTS: HMGB1 is overexpressed in human inflammatory bowel disease and gastrointestinal cancers, and HMGB1 protein localizes in enterocytes and stromal cells in colitis and CRC specimens from humans and rodents. As previously described, enterocyte HMGB1 deficiency aggravates severe chemical-induced intestinal injury, but not Citrobacter rodentium or T cell transfer colitis in mice. HMGB1-deficient enterocytes and organoids do not exhibit deviant apoptotic or autophagic activity, altered proliferative or migratory capacity, abnormal intestinal permeability or aberrant DSS-induced organoid inflammation in vitro. Instead, we observed altered in vivo-reprogramming of both intestinal epithelia and infiltrating myeloid cells in Hmgb1ΔIEC early during colitis, suggesting HMGB1-mediated paracrine injury signaling. Hmgb1ΔIEC had higher CRC burden than wildtypes in the Apc+/min model, whereas inflammatory CRC was attenuated in Hmgb1ΔLysM. Cellular and molecular phenotyping of Hmgb1ΔIEC and Hmgb1ΔLysM cancers indicates context-dependent transcriptional modulation of immune signaling and extracellular matrix remodeling via HMGB1. CONCLUSION: Enterocytes and myeloid cells context-dependently regulate host responses to severe colitis and maladaptive intestinal wound healing via HMGB1.

Nmes1 is a novel regulator of mucosal response influencing intestinal healing potential.

The initiation of tissue remodeling following damage is a critical step in preventing the development of immune-mediated diseases. Several factors contribute to mucosal healing, leading to innovative therapeutic approaches for managing intestinal disorders. However, uncovering alternative targets and gaining mechanistic insights are imperative to enhance therapy efficacy and broaden its applicability across different intestinal diseases. Here we demonstrate that Nmes1, encoding for Normal Mucosa of Esophagus-Specific gene 1, also known as Aa467197, is a novel regulator of mucosal healing. Nmes1 influences the macrophage response to the tissue remodeling cytokine IL-4 in vitro. In addition, using two murine models of intestinal damage, each characterized by a type 2-dominated environment with contrasting functions, the ablation of Nmes1 results in decreased intestinal regeneration during the recovery phase of colitis, while enhancing parasitic egg clearance and reducing fibrosis during the advanced stages of Schistosoma mansoni infection. These outcomes are associated with alterations in CX3CR1+ macrophages, cells known for their wound-healing potential in the inflamed colon, hence promising candidates for cell therapies. All in all, our data indicate Nmes1 as a novel contributor to mucosal healing, setting the basis for further investigation into its potential as a new target for the treatment of colon-associated inflammation.

Does gamma-glutamyltransferase correlate with liver tumor burden in neuroendocrine tumors?

PURPOSE: In patients with neuroendocrine tumors (NETs) and liver metastases, increased gamma-glutamyltransferase (GGT) is commonly assumed as an indicator for progressive disease. To date, however, empirical data are lacking. This study aimed to investigate associations between GGT and liver tumor burden. In longitudinal analyses, associations of GGT and radiographic responses of liver metastases under therapy were investigated. METHODS: The cross-sectional sample consisted of 104 patients who were treated at the University Medical Center Hamburg-Eppendorf from 2008 to 2021 (mean age 62.3 ± 12.6 years, 58.7% male). GGT and liver imaging were identified in a time range of 3 months. Radiologic reassessments were performed to estimate liver tumor burden. In a separate longitudinal sample (n = 15), the course of GGT levels under chemotherapy was analyzed. Data were retrospectively analyzed with a univariate ANOVA, linear regression analyses, and Wilcoxon tests. RESULTS: Of 104 cross-sectionally analyzed patients, 54 (51.9%) showed a GGT elevation. GGT levels and liver tumor burden were positively correlated (p < 0.001), independently from age, gender, primary tumor location, grading, and cholestasis. Notably, GGT increase was associated with a liver tumor burden of >50%. In the longitudinal sample, 10 of 11 patients with progressive disease showed increasing GGT, whereas 4 of 4 patients with regressive disease showed declining GGT. CONCLUSION: Our findings indicate that GGT is associated with liver tumor burden. Over the course of therapy, GGT appears to change in line with radiographic responses. Further longitudinal studies with larger sample sizes are required to define GGT as a reliable marker for tumor response.

IFNγ and CTLA-4 Drive Hepatic CD4 T-Cell Tolerance and Protection From Autoimmunity in Mice.

BACKGROUND & AIMS: The liver has a distinct capacity to induce immune tolerance to hepatic antigens. Although liver tolerance can be advantageous for preventing autoimmune and inflammatory diseases, it also can be detrimental by preventing immune surveillance of infected or malignant cells. Here, we investigated the immune mechanisms that establish hepatic tolerance. METHODS: Tolerance was investigated in C-reactive protein (CRP)-myelin basic protein (MBP) mice expressing the neuroantigen MBP in hepatocytes, providing profound resistance to MBP-induced neuroinflammation. Tolerance induction was studied after transfer of MBP-specific CD4 T cells into CRP-MBP mice, and tolerance mechanisms were tested using depleting or blocking antibodies. RESULTS: Although tolerant CRP-MBP mice display increased numbers of forkhead box P3+ regulatory T cells, we here found them not essential for the maintenance of hepatic tolerance. Instead, upon MBP recognition in the liver, MBP-specific T cells became activated to produce interferon (IFN)γ, which, in turn, induced local up-regulation of recruitment molecules, including Chemokine (C-X-C motif) ligand9 and its receptor C-X-C motif chemokine receptor3, facilitating endothelial translocation and redirection of MBP-specific T cells into the hepatic parenchyma. There, the translocated MBP-specific CD4 T cells partly converted into interleukin 10-producing type 1 regulatory T cells, and significantly up-regulated the expression of immune checkpoint molecules, notably cytotoxic T-lymphocyte-associated protein 4 (CTLA-4). Intriguingly, although liver tolerance was not affected by impairment of interleukin 10 signaling, concomitant blockade of IFNγ and CTLA-4 abrogated hepatic tolerance induction to MBP, resulting in neuroinflammatory autoimmune disease in these mice. CONCLUSIONS: IFNγ-mediated redirection of autoreactive CD4 T cells into the liver and up-regulation of checkpoint molecules, including CTLA-4, were essential for tolerance induction in the liver, hence representing a potential treatment target for boosting or preventing liver tolerance.

IL-10 dampens antitumor immunity and promotes liver metastasis via PD-L1 induction.

BACKGROUND & AIMS: The liver is one of the organs most commonly affected by metastasis. The presence of liver metastases has been reported to be responsible for an immunosuppressive microenvironment and diminished immunotherapy efficacy. Herein, we aimed to investigate the role of IL-10 in liver metastasis and to determine how its modulation could affect the efficacy of immunotherapy in vivo. METHODS: To induce spontaneous or forced liver metastasis in mice, murine cancer cells (MC38) or colon tumor organoids were injected into the cecum or the spleen, respectively. Mice with complete and cell type-specific deletion of IL-10 and IL-10 receptor alpha were used to identify the source and the target of IL-10 during metastasis formation. Programmed death ligand 1 (PD-L1)-deficient mice were used to test the role of this checkpoint. Flow cytometry was applied to characterize the regulation of PD-L1 by IL-10. RESULTS: We found that Il10-deficient mice and mice treated with IL-10 receptor alpha antibodies were protected against liver metastasis formation. Furthermore, by using IL-10 reporter mice, we demonstrated that Foxp3+ regulatory T cells (Tregs) were the major cellular source of IL-10 in liver metastatic sites. Accordingly, deletion of IL-10 in Tregs, but not in myeloid cells, led to reduced liver metastasis. Mechanistically, IL-10 acted on Tregs in an autocrine manner, thereby further amplifying IL-10 production. Furthermore, IL-10 acted on myeloid cells, i.e. monocytes, and induced the upregulation of the immune checkpoint protein PD-L1. Finally, the PD-L1/PD-1 axis attenuated CD8-dependent cytotoxicity against metastatic lesions. CONCLUSIONS: Treg-derived IL-10 upregulates PD-L1 expression in monocytes, which in turn reduces CD8+ T-cell infiltration and related antitumor immunity in the context of colorectal cancer-derived liver metastases. These findings provide the basis for future monitoring and targeting of IL-10 in colorectal cancer-derived liver metastases. IMPACT AND IMPLICATIONS: Liver metastasis diminishes the effectiveness of immunotherapy and increases the mortality rate in patients with colorectal cancer. We investigated the role of IL-10 in liver metastasis formation and assessed its impact on the effectiveness of immunotherapy. Our data show that IL-10 is a pro-metastatic factor involved in liver metastasis formation and that it acts as a regulator of PD-L1. This provides the basis for future monitoring and targeting of IL-10 in colorectal cancer-derived liver metastasis.

Preoperative TIPS and in-hospital mortality in patients with cirrhosis undergoing surgery.

Background & Aims: Cirrhosis is associated with an increased surgical morbidity and mortality. Portal hypertension and the surgery type have been established as critical determinants of postoperative outcome. We aim to evaluate the hypothesis that preoperative transjugular intrahepatic portosystemic shunt (TIPS) placement in patients with cirrhosis is associated with a lower incidence of in-house mortality/liver transplantation (LT) after surgery. Methods: A retrospective database search for the years 2010-2020 was carried out. We identified 64 patients with cirrhosis who underwent surgery within 3 months after TIPS placement and 131 patients with cirrhosis who underwent surgery without it (controls). Operations were categorised into low-risk and high-risk procedures. The primary endpoint was in-house mortality/LT. We analysed the influence of high-risk surgery, preoperative TIPS placement, age, sex, baseline creatinine, presence of ascites, Chronic Liver Failure Consortium Acute Decompensation (CLIF-C AD), American Society of Anesthesiologists (ASA), and model for end-stage liver disease (MELD) scores on in-house mortality/LT by multivariable Cox proportional hazards regression. Results: In both the TIPS and the control cohort, most patients presented with a Child-Pugh B stage (37/64, 58% vs. 70/131, 53%) at the time of surgery, but the median MELD score was higher in the TIPS cohort (14 vs. 11 points). Low-risk and high-risk procedures amounted to 47% and 53% in both cohorts. The incidence of in-house mortality/LT was lower in the TIPS cohort (12/64, 19% vs. 52/131, 40%), also when further subdivided into low-risk (0/30, 0% vs. 10/61, 16%) and high-risk surgery (12/34, 35% vs. 42/70, 60%). Preoperative TIPS placement was associated with a lower rate for postoperative in-house mortality/LT (hazard ratio 0.44, 95% CI 0.19-1.00) on multivariable analysis. Conclusions: A preoperative TIPS might be associated with reduced postoperative in-house mortality in selected patients with cirrhosis. Impact and implications: Patients with cirrhosis are at risk for more complications and a higher mortality after surgical procedures. A transjugular intrahepatic portosystemic shunt (TIPS) is used to treat complications of cirrhosis, but it is unclear if it also helps to lower the risk of surgery. This study takes a look at complications and mortality of patients undergoing surgery with or without a TIPS, and we found that patients with a TIPS develop less complications and have an improved survival. Therefore, a preoperative TIPS should be considered in selected patients, especially if indicated by ascites.

EpCAM-positive circulating tumor cells and serum AFP levels predict outcome after curative resection of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) has high recurrence rates exceeding 50% despite curative resection. The serum biomarker alpha-fetoprotein (AFP) is a well-known prognostic marker for HCC. EpCAM-positive circulating tumor cells (CTC) have a high predictive value for early HCC recurrence after curatively intended resection, most likely indicating micro-metastases at the time of resection. However, sensitivity remains low. The objective of this study was to evaluate a composite test comprising both CTC and AFP to identify patients at high risk for early HCC recurrence. We prospectively enrolled 58 patients undergoing curative intended resection for HCC at a tertiary referral center. Blood specimens were obtained prior to resection and analyzed for EpCAM-positive CTC and serum AFP levels. A positive result was defined as either detection of CTC or AFP levels ≥ 400 ng/ml. Eight patients tested positive for CTC, seven for AFP, and two for both markers. A positive composite test was significantly associated with shorter early recurrence-free survival (5 vs. 16 months, p = 0.005), time to recurrence (5 vs. 16 months, p = 0.011), and overall survival (37 vs. not reached, p = 0.034). Combining CTC and AFP identified patients with poor outcome after surgical resection, for whom adjuvant or neoadjuvant therapies may be particularly desirable.

Protocol for orthotopic single-lung transplantation in mice as a tool for lung metastasis studies.

The transplantation model provides the opportunity to assess the relevance of a molecule of interest for tumor cell extravasation by using a respective genetically modified donor animal. Here, we present a protocol for orthotopic single-lung transplantation in mice as a tool for lung metastasis studies. We describe steps for animal preparation, lung transplantation, and tumor cell injection. We then detail procedures for the direct comparison of tumor cell spreading between the genetically modified left lung and the naive right lung parenchyma. For complete details on the use and execution of this protocol, please refer to Giannou et al. (2023).1.

Efficacy and safety of atezolizumab/bevacizumab in patients with HCC after prior systemic therapy: A global, observational study.

BACKGROUND: Since the introduction of the combination treatment of anti-programmed death-ligand 1 antibody atezolizumab and anti-VEGF antibody bevacizumab (AB), median overall survival in HCC has drastically improved. However, evidence on the efficacy and safety of the novel treatment standard in patients with prior exposure to systemic treatment is scarce. The aim of this global, multicenter, observational study was to evaluate the efficacy and safety of AB in patients after previous systemic therapy. METHODS: We screened our global, multicenter, prospectively maintained registry database for patients who received any systemic therapy before AB. The primary end point was overall survival; secondary end points were time-to-progression, progression-free survival, objective response rate, and safety (rate and severity of adverse events). RESULTS: Among 493 patients who received AB for unresectable HCC, 61 patients received prior systemic therapy and were included in this analysis. The median age of the study population was 66 years, with 91.8% males. Predominant risk factors for HCC were viral hepatitis (59%) and alcohol (23%). Overall survival for AB was 16.2 (95% CI, 14.5-17.9) months, time-to-progression and progression-free survival were 4.1 (95% CI, 1.5-6.6) and 3.1 (95% CI, 1.1-5.1) months, respectively. The objective response rate was 38.2% (7.3% with complete and 30.9% with partial response). Overall survival was not influenced by treatment line (2nd vs. >2nd) or previous systemic treatment modality (tyrosine kinase inhibitors vs. immune checkpoint inhibitors). Treatment-related adverse events of all grades according to Common Terminology Criteria for Adverse Events were documented in 42.6% of patients, with only 13.1% of grade ≥3, including one death. CONCLUSION: In this observational study, AB emerges as a safe and efficacious treatment option in patients with HCC previously treated with other systemic therapy.