Atezolizumab plus bevacizumab versus active surveillance in patients with resected or ablated high-risk hepatocellular carcinoma (IMbrave050): a randomised, open-label, multicentre, phase 3 trial.

BACKGROUND: No adjuvant treatment has been established for patients who remain at high risk for hepatocellular carcinoma recurrence after curative-intent resection or ablation. We aimed to assess the efficacy of adjuvant atezolizumab plus bevacizumab versus active surveillance in patients with high-risk hepatocellular carcinoma. METHODS: In the global, open-label, phase 3 IMbrave050 study, adult patients with high-risk surgically resected or ablated hepatocellular carcinoma were recruited from 134 hospitals and medical centres in 26 countries in four WHO regions (European region, region of the Americas, South-East Asia region, and Western Pacific region). Patients were randomly assigned in a 1:1 ratio via an interactive voice-web response system using permuted blocks, using a block size of 4, to receive intravenous 1200 mg atezolizumab plus 15 mg/kg bevacizumab every 3 weeks for 17 cycles (12 months) or to active surveillance. The primary endpoint was recurrence-free survival by independent review facility assessment in the intention-to-treat population. This trial is registered with ClinicalTrials.gov, NCT04102098. FINDINGS: The intention-to-treat population included 668 patients randomly assigned between Dec 31, 2019, and Nov 25, 2021, to either atezolizumab plus bevacizumab (n=334) or to active surveillance (n=334). At the prespecified interim analysis (Oct 21, 2022), median duration of follow-up was 17·4 months (IQR 13·9-22·1). Adjuvant atezolizumab plus bevacizumab was associated with significantly improved recurrence-free survival (median, not evaluable [NE]; [95% CI 22·1-NE]) compared with active surveillance (median, NE [21·4-NE]; hazard ratio, 0·72 [adjusted 95% CI 0·53-0·98]; p=0·012). Grade 3 or 4 adverse events occurred in 136 (41%) of 332 patients who received atezolizumab plus bevacizumab and 44 (13%) of 330 patients in the active surveillance group. Grade 5 adverse events occurred in six patients (2%, two of which were treatment related) in the atezolizumab plus bevacizumab group, and one patient (<1%) in the active surveillance group. Both atezolizumab and bevacizumab were discontinued because of adverse events in 29 patients (9%) who received atezolizumab plus bevacizumab. INTERPRETATION: Among patients at high risk of hepatocellular carcinoma recurrence following curative-intent resection or ablation, recurrence-free survival was improved in those who received atezolizumab plus bevacizumab versus active surveillance. To our knowledge, IMbrave050 is the first phase 3 study of adjuvant treatment for hepatocellular carcinoma to report positive results. However, longer follow-up for both recurrence-free and overall survival is needed to assess the benefit-risk profile more fully. FUNDING: F Hoffmann-La Roche/Genentech.

Atezolizumab with or without bevacizumab in unresectable hepatocellular carcinoma (GO30140): an open-label, multicentre, phase 1b study.

BACKGROUND: Dual blockade of PD-L1 and VEGF has enhanced anticancer immunity through multiple mechanisms and augmented antitumour activity in multiple malignancies. We aimed to assess the efficacy and safety of atezolizumab (anti-PD-L1) alone and combined with bevacizumab (anti-VEGF) in patients with unresectable hepatocellular carcinoma. METHODS: GO30140 is an open-label, multicentre, multiarm, phase 1b study that enrolled patients at 26 academic centres and community oncology practices in seven countries worldwide. The study included five cohorts, and the two hepatocellular carcinoma cohorts, groups A and F, are described here. Inclusion criteria for these two groups included age 18 years and older; histologically, cytologically, or clinically (per American Association for the Study of Liver Diseases criteria) confirmed unresectable hepatocellular carcinoma that was not amenable to curative treatment; no previous systemic treatment; and Eastern Cooperative Oncology Group performance status of 0 or 1. In group A, all patients received atezolizumab (1200 mg) and bevacizumab (15 mg/kg) intravenously every 3 weeks. In group F, patients were randomly assigned (1:1) to receive intravenous atezolizumab (1200 mg) plus intravenous bevacizumab (15 mg/kg) every 3 weeks or atezolizumab alone by interactive voice-web response system using permuted block randomisation (block size of two) and stratification factors of geographical region; macrovascular invasion, extrahepatic spread, or both; and baseline α-fetoprotein concentration. Primary endpoints were confirmed objective response rate in all patients who received the combination treatment for group A and progression-free survival in the intention-to-treat population in group F, both assessed by an independent review facility according to Response Evaluation Criteria in Solid Tumors version 1.1. In both groups, safety was assessed in all patients who received at least one dose of any study treatment. This study is registered with ClinicalTrials.gov, NCT02715531, and is closed to enrolment. FINDINGS: In group A, 104 patients were enrolled between July 20, 2016, and July 31, 2018, and received atezolizumab plus bevacizumab. With a median follow-up of 12·4 months (IQR 8·0-16·2), 37 (36%; 95% CI 26-46) of 104 patients had a confirmed objective response. The most common grade 3-4 treatment-related adverse events were hypertension (13 [13%]) and proteinuria (seven [7%]). Treatment-related serious adverse events occurred in 25 (24%) patients and treatment-related deaths in three (3%) patients (abnormal hepatic function, hepatic cirrhosis, and pneumonitis). In group F, 119 patients were enrolled and randomly assigned (60 to atezolizumab plus bevacizumab; 59 to atezolizumab monotherapy) between May 18, 2018, and March 7, 2019. With a median follow-up of 6·6 months (IQR 5·5-8·5) for the atezolizumab plus bevacizumab group and 6·7 months (4·2-8·2) for the atezolizumab monotherapy group, median progression-free survival was 5·6 months (95% CI 3·6-7·4) versus 3·4 months (1·9-5·2; hazard ratio 0·55; 80% CI 0·40-0·74; p=0·011). The most common grade 3-4 treatment-related adverse events in group F were hypertension (in three [5%] patients in the atezolizumab plus bevacizumab group; none in the atezolizumab monotherapy group) and proteinuria (in two [3%] patients in the atezolizumab plus bevacizumab group; none in the atezolizumab monotherapy group). Treatment-related serious adverse events occurred in seven (12%) patients in the atezolizumab plus bevacizumab group and two (3%) patients in the atezolizumab monotherapy group. There were no treatment-related deaths. INTERPRETATION: Our study shows longer progression-free survival with a combination of atezolizumab plus bevacizumab than with atezolizumab alone in patients with unresectable hepatocellular carcinoma not previously treated with systemic therapy. Therefore, atezolizumab plus bevacizumab might become a promising treatment option for these patients. This combination is being compared with standard-of-care sorafenib in a phase 3 trial. FUNDING: F Hoffmann-La Roche/Genentech.

IMbrave 050: a Phase III trial of atezolizumab plus bevacizumab in high-risk hepatocellular carcinoma after curative resection or ablation.

Hepatocellular carcinoma recurs in 70-80% of cases following potentially curative resection or ablation and the immune component of the liver microenvironment plays a key role in recurrence. Many immunosuppressive mechanisms implicated in HCC recurrence are modulated by VEGF and/or immune checkpoints such as PD-L1. Atezolizumab (PD-L1 inhibitor) plus bevacizumab (VEGF inhibitor) has been shown to significantly improve overall survival, progression-free survival and overall response rate in unresectable HCC. Dual PD-L1/VEGF blockade may be effective in reducing HCC recurrence by creating a more immune-favorable microenvironment. We describe the rationale and design of IMbrave 050 (NCT04102098), a randomized, open-label, Phase III study comparing atezolizumab plus bevacizumab versus active surveillance in HCC patients at high-risk of recurrence following curative resection or ablation. The primary end point is recurrence-free survival. Clinical Trial Registration: NCT04102098.

DAP12 expression in lung macrophages mediates ischemia/reperfusion injury by promoting neutrophil extravasation.

Neutrophils are critical mediators of innate immune responses and contribute to tissue injury. However, immune pathways that regulate neutrophil recruitment to injured tissues during noninfectious inflammation remain poorly understood. DAP12 is a cell membrane-associated protein that is expressed in myeloid cells and can either augment or dampen innate inflammatory responses during infections. To elucidate the role of DAP12 in pulmonary ischemia/reperfusion injury (IRI), we took advantage of a clinically relevant mouse model of transplant-mediated lung IRI. This technique allowed us to dissect the importance of DAP12 in tissue-resident cells and those that infiltrate injured tissue from the periphery during noninfectious inflammation. Macrophages in both mouse and human lungs that have been subjected to cold ischemic storage express DAP12. We found that donor, but not recipient, deficiency in DAP12 protected against pulmonary IRI. Analysis of the immune response showed that DAP12 promotes the survival of tissue-resident alveolar macrophages and contributes to local production of neutrophil chemoattractants. Intravital imaging demonstrated a transendothelial migration defect into DAP12-deficient lungs, which can be rescued by local administration of the neutrophil chemokine CXCL2. We have uncovered a previously unrecognized role for DAP12 expression in tissue-resident alveolar macrophages in mediating acute noninfectious tissue injury through regulation of neutrophil trafficking.

Innate immune cells in transplantation.

PURPOSE OF REVIEW: To examine the recent literature on the role of innate cells in immunity to transplanted tissue. It specifically addresses the impact of monocytes/macrophages, neutrophils, natural killer cells, and platelets. RECENT FINDINGS: Current research indicates that innate immunity plays a dual role in response to transplanted tissue with the ability to either facilitate rejection or promote tolerance. Intriguingly, some of these cells are even capable of reacting to allogeneic cells, a feature usually only attributed to cells of the adaptive immune system. SUMMARY: This review highlights the new therapeutic targets in the innate immune system that may be useful in the treatment of transplant recipients. It also emphasizes the need to use caution in exploring these new therapeutics.

Ineffective CD8(+) T-cell immunity to adeno-associated virus can result in prolonged liver injury and fibrogenesis.

Chronic viral hepatitis depends on the inability of the T-cell immune response to eradicate antigen. This results in a sustained immune response accompanied by tissue injury and fibrogenesis. We have created a mouse model that reproduces these effects, based on the response of CD8(+) T cells to hepatocellular antigen delivered by an adeno-associated virus (AAV) vector. Ten thousand antigen-specific CD8(+) T cells undergo slow expansion in the liver and can precipitate a subacute inflammatory hepatitis with stellate cell activation and fibrosis. Over time, antigen-specific CD8(+) T cells show signs of exhaustion, including high expression of PD-1, and eventually both inflammation and fibrosis resolve. This model allows the investigation of both chronic liver immunopathology and its resolution.

Brief Report: Safety and Antitumor Activity of Alectinib Plus Atezolizumab From a Phase 1b Study in Advanced ALK-Positive NSCLC.

Introduction: Alectinib is a preferred first-line treatment option for advanced ALK-positive NSCLC. Combination regimens of alectinib with immune checkpoint inhibitors are being evaluated for synergistic effects. Methods: Adults with treatment-naive, stage IIIB/IV, or recurrent ALK-positive NSCLC were enrolled into a two-stage phase 1b study. Patients received alectinib 600 mg (twice daily during cycle 1 and throughout each 21-d cycle thereafter) plus atezolizumab 1200 mg (d8 of cycle 1 and then d1 of each 21-d cycle). Primary objectives were to evaluate safety and tolerability of alectinib plus atezolizumab. Secondary objectives included assessments of antitumor activity. Results: In total, 21 patients received more than or equal to 1 dose of alectinib or atezolizumab. As no dose-limiting toxicities were observed in stage 1 (n = 7), the starting dose and schedule were continued into stage 2 (n = 14). Median duration of follow-up was 29 months (range: 1-39). Grade 3 treatment-related adverse events occurred in 57% of the patients, most often rash (19%). No grade 4 or 5 treatment-related adverse events were reported. Confirmed objective response rate was 86% (18 of 21; 95% confidence interval [CI]: 64-97). Median progression-free survival was not estimable (NE) (95% CI: 13 mo-NE), neither was median overall survival (95% CI: 33 mo-NE). Conclusions: The combination of alectinib and atezolizumab is feasible, but increased toxicity was found compared with the individual agents. With small sample sizes and relatively short follow-up, definitive conclusions regarding antitumor activity cannot be made.

Molecular correlates of clinical response and resistance to atezolizumab in combination with bevacizumab in advanced hepatocellular carcinoma.

Atezolizumab (anti-programmed death-ligand 1 (PD-L1)) and bevacizumab (anti-vascular endothelial growth factor (VEGF)) combination therapy has become the new standard of care in patients with unresectable hepatocellular carcinoma. However, potential predictive biomarkers and mechanisms of response and resistance remain less well understood. We report integrated molecular analyses of tumor samples from 358 patients with hepatocellular carcinoma (HCC) enrolled in the GO30140 phase 1b or IMbrave150 phase 3 trial and treated with atezolizumab combined with bevacizumab, atezolizumab alone or sorafenib (multikinase inhibitor). Pre-existing immunity (high expression of CD274, T-effector signature and intratumoral CD8+ T cell density) was associated with better clinical outcomes with the combination. Reduced clinical benefit was associated with high regulatory T cell (Treg) to effector T cell (Teff) ratio and expression of oncofetal genes (GPC3, AFP). Improved outcomes from the combination versus atezolizumab alone were associated with high expression of VEGF Receptor 2 (KDR), Tregs and myeloid inflammation signatures. These findings were further validated by analyses of paired pre- and post-treatment biopsies, in situ analyses and in vivo mouse models. Our study identified key molecular correlates of the combination therapy and highlighted that anti-VEGF might synergize with anti-PD-L1 by targeting angiogenesis, Treg proliferation and myeloid cell inflammation.

Direct, help-independent priming of CD8+ T cells by adeno-associated virus-transduced hepatocytes.

UNLABELLED: Both hepatitis B and C viruses frequently establish chronic infection, raising the question whether T cells are poorly primed in the liver. To determine the role of different cell types in the activation of CD8+ T cells against hepatocellular antigens, we used an Adeno-associated virus to deliver ovalbumin to hepatocytes. In contrast to CD8+ T cells, CD4+ T cells were not activated. The CD8+ T cells were activated even in the absence of endogenous CD4+ T cells; however, in the liver, these cells were high in the programmed death-1 protein and low in CD127. Chimera experiments revealed that these CD8+ T cells were activated on a solid tissue cell. CONCLUSION: Priming of CD8+ T cells directly on nonhematopoietic cells, in the absence of CD4+ T cell help, results in suboptimal T cell activation. This could explain the impaired function of CD8+ T cells seen in chronic liver infection.

Atezolizumab plus bevacizumab combination enables an unresectable hepatocellular carcinoma resectable and links immune exclusion and tumor dedifferentiation to acquired resistance.

We reported a patient with unresectable hepatocellular carcinoma (HCC) who initially received 15 cycles of atezolizumab plus bevacizumab combination and had best tumor response of partial response, but later experienced disease progression. After subsequent surgical resection, the patient enjoyed long-term disease-free status at the last follow-up 19 months after surgery. By investigating paired tumor tissues (pretreatment and post-progression samples) with immunohistochemistry, multiplex immunofluorescence, RNA sequencing, and DNA sequencing, we explored the dynamic changes in the tumor microenvironment (TME) and potential mechanisms underlying acquired resistance to the combination. In the post-progression HCC tissue compared with the baseline tissue, the expression of PD-L1 in tumor-infiltrating immune cells and the abundance of CD8+ T cells in the tumor area had decreased, and an immune-excluded TME had emerged. Transcriptomic analysis revealed a gene expression signature representing progenitor/hepatoblast features in the post-progression tumor tissue, with an increased expression of imprinted genes and decreased expression of cytochrome P450 family genes. Finally, tumor mutational burden and MHC class I expression in tumor cells were both increased in the post-progression tissue, suggesting that neoantigen depletion or loss-of-antigen presentation were unlikely causes of acquired resistance in this patient. Atezolizumab plus bevacizumab combination therapy enabled our patient to receive hepatectomy and achieve long-term remission. A comparison of paired tumor tissues suggested that immune-excluded TME and tumor dedifferentiation may have contributed to acquired resistance to the combination.

Safety and Clinical Activity of Atezolizumab Plus Bevacizumab in Patients with Ovarian Cancer: A Phase Ib Study.

PURPOSE: Atezolizumab has shown antitumor activity in patients with ovarian cancer. Dual blockade of programmed death-ligand 1 (PD-L1) and VEGF enhances anticancer immunity and augments antitumor activity in several cancers. The safety and efficacy of atezolizumab plus bevacizumab were evaluated in patients with ovarian cancer. PATIENTS AND METHODS: In this open-label, multicenter phase Ib study, patients with platinum-resistant ovarian cancer received intravenous atezolizumab (1,200 mg) and bevacizumab (15 mg/kg) once every 3 weeks. The primary endpoint was safety; secondary endpoints included overall response rate (ORR), duration of response (DOR), progression-free survival (PFS), and overall survival (OS). Exploratory biomarkers were also evaluated. RESULTS: Twenty patients received treatment. Treatment-related adverse events occurred in 19 patients (95%); seven (35%) had grade 3/4 events. No grade 5 events occurred. The safety profile of atezolizumab plus bevacizumab was consistent with those of the individual agents. Two patients (10%) discontinued treatment because of pneumonitis and small bowel obstruction. Three patients had partial responses of 11.3-18.9 months' duration; the ORR was 15%. Eight patients (40%) had stable disease, hence the disease control rate was 55%. The median DOR was not reached (95% confidence interval, 11.3-not reached). Median PFS was 4.9 months (range, 1.2-20.2); median OS was 10.2 months (range, 1.2-26.6). No association was seen between treatment response and PD-L1 expression, tumor histology, or number of prior therapies. CONCLUSIONS: Atezolizumab plus bevacizumab led to durable responses and/or disease stabilization in some patients with platinum-resistant ovarian cancer; the safety profiles were consistent with those of each agent.

Spleen-derived classical monocytes mediate lung ischemia-reperfusion injury through IL-1ß.

Ischemia-reperfusion injury, a form of sterile inflammation, is the leading risk factor for both short-term mortality following pulmonary transplantation and chronic lung allograft dysfunction. While it is well recognized that neutrophils are critical mediators of acute lung injury, processes that guide their entry into pulmonary tissue are not well understood. Here, we found that CCR2+ classical monocytes are necessary and sufficient for mediating extravasation of neutrophils into pulmonary tissue during ischemia-reperfusion injury following hilar clamping or lung transplantation. The classical monocytes were mobilized from the host spleen, and splenectomy attenuated the recruitment of classical monocytes as well as the entry of neutrophils into injured lung tissue, which was associated with improved graft function. Neutrophil extravasation was mediated by MyD88-dependent IL-1ß production by graft-infiltrating classical monocytes, which downregulated the expression of the tight junction-associated protein ZO-2 in pulmonary vascular endothelial cells. Thus, we have uncovered a crucial role for classical monocytes, mobilized from the spleen, in mediating neutrophil extravasation, with potential implications for targeting of recipient classical monocytes to ameliorate pulmonary ischemia-reperfusion injury in the clinic.

Monocytes in sterile inflammation: recruitment and functional consequences.

Monocytes play an important role in initiating innate immune responses. Three subsets of these cells have been defined in mice including classical, nonclassical and intermediate monocytes. Each of these cell types has been extensively studied for their role in infectious diseases. However, their role in sterile injury as occurs during ischemia-reperfusion injury, atherosclerosis, and trauma has only recently been the focus of investigations. Here, we review mechanisms of monocyte recruitment to sites of sterile injury, their modes of action, and their effect on disease outcome in murine models with some references to human studies. Therapeutic strategies to target these cells must be developed with caution since each monocyte subset is capable of mediating either anti- or pro-inflammatory effects depending on the setting.

Experimental models of lung transplantation.

Lung transplantation is a life saving treatment for end stage pulmonary diseases. The development and refinement of this therapy required the utilization of various animal models, without which this procedure would not have become a clinical reality. Canine models were critical in the initial breakthroughs in surgical technique and immunosuppressive regimens, which directly led to the first successful human lung transplantation. Orthotopic lung transplant models in the rat provided a platform for more detailed investigation of immune responses to pulmonary grafts. Investigation of chronic rejection of lungs has significantly been advanced through the use of mouse tracheal transplant experiments. And finally, the advent of orthotopic, vascularized lung transplantation in the mouse opened the door to the use of genetic and molecular tools that are necessary for the rigorous mechanistic study of alloimmune and non-alloimmune factors contributing to lung graft failure. Taken together, animal models will continue to be a cornerstone in the advancement of clinical success in lung transplantation.

Surgical technique for lung retransplantation in the mouse.

Microsurgical cuff techniques for orthotopic vascularized murine lung transplantation have allowed for the design of studies that examine mechanisms contributing to the high failure rate of pulmonary grafts. Here, we provide a detailed technical description of orthotopic lung retransplantation in mice, which we have thus far performed in 144 animals. The total time of the retransplantation procedure is approximately 55 minutes, 20 minutes for donor harvest and 35 minutes for the implantation, with a success rate exceeding 95%. The mouse lung retransplantation model represents a novel and powerful tool to examine how cells that reside in or infiltrate pulmonary grafts shape immune responses.

Intravital 2-photon imaging of leukocyte trafficking in beating heart.

Two-photon intravital microscopy has substantially broadened our understanding of tissue- and organ-specific differences in the regulation of inflammatory responses. However, little is known about the dynamic regulation of leukocyte recruitment into inflamed heart tissue, largely due to technical difficulties inherent in imaging moving tissue. Here, we report a method for imaging beating murine hearts using intravital 2-photon microscopy. Using this method, we visualized neutrophil trafficking at baseline and during inflammation. Ischemia reperfusion injury induced by transplantation or transient coronary artery ligation led to recruitment of neutrophils to the heart, their extravasation from coronary veins, and infiltration of the myocardium where they formed large clusters. Grafting hearts containing mutant ICAM-1, a ligand important for neutrophil recruitment, reduced the crawling velocities of neutrophils within vessels, and markedly inhibited their extravasation. Similar impairment was seen with the inhibition of Mac-1, a receptor for ICAM-1. Blockade of LFA-1, another ICAM-1 receptor, prevented neutrophil adherence to endothelium and extravasation in heart grafts. As inflammatory responses in the heart are of great relevance to public health, this imaging approach holds promise for studying cardiac-specific mechanisms of leukocyte recruitment and identifying novel therapeutic targets for treating heart disease.