HFE -related hemochromatosis (HC) is caused by hepcidin dysregulation and is characterized by excessive iron absorption and accumulation in the liver, heart, and endocrine glands, which leads to complications including arthropathy, cirrhosis, and hepatocellular carcinoma. Standard of care (SoC) typically involves phlebotomy, with iron chelators used less frequently. However, these therapies may cause side effects and negatively impact quality of life, making them troublesome for some patients. Using the Delphi methodology, a survey was developed to identify unmet needs and clinical challenges within the current therapeutic landscape of HC, and to establish consensus statements for its management. Consensus was defined as ≥75% agreement. Thirty-two HC specialists from Europe, Australia and the USA responded to the survey. After three rounds of survey refinement, final consensus statements were compiled. The Delphi process identified key unmet needs in HC, including lack of alternatives to phlebotomy, persistent symptoms, and burden related to the use of current SoC. The consensus process helped establish definitions for high phlebotomy treatment burden, intolerance, and suboptimal response to phlebotomy. The process also identified the subgroup of patients overly burdened by phlebotomy. Patient-reported outcomes were considered key to assessing phlebotomy’s impact, although they are rarely measured in clinical practice. The Delphi study highlighted the limitations of phlebotomy, with respondents identifying a high unmet need for patients who cannot be managed with or do not tolerate this approach. The study suggested exploring alternative therapy options for patients who experience high treatment burden or intolerance to phlebotomy.

Sickle cell disease-associated pulmonary hypertension (SCD-PH) affects approximately 10% of adults with SCD and markedly increases mortality, yet mechanistic and haemodynamic heterogeneity complicates classification, trial design, and treatment selection. We propose an integrated framework linking five interacting axes-anaemia/high-output, haemolysis/haem/iron toxicity, hypoxia, inflammation, and thrombosis-to clinically defined phenotypes (post-capillary, pre-capillary, combined, chronic thromboembolic PH [CTEPH], and acute cor pulmonale). Chronic anaemia drives high-output physiology, left ventricular diastolic dysfunction, and post-capillary PH. Intravascular and erythrophagocytic haemolysis cause convergent inside-out and outside-in pulmonary vascular injury via nitric oxide depletion and oxidative damage, promoting pre-capillary PH; hypoxia, inflammation, and thrombosis amplify remodelling, helping explain why combined phenotypes predominate. Management prioritises hydroxyurea and transfusion, while PDE5 inhibition (sildenafil) has shown harm. Emerging avenues include soluble guanylate cyclase stimulation, L-arginine, haemoglobin/haem scavenging (haptoglobin, hemopexin), anti-inflammatory strategies, and iron-targeted interventions. This mechanism-to-phenotype map supports phenotype-stratified, mechanism-guided trials in SCD-PH.

OBJECTIVES:To validate whether cerebrospinal fluid oxyhaemoglobin (CSF-Hb), measured from external ventricular or lumbar drains, is associated with secondary brain injury (SAH-SBI) after aneurysmal subarachnoid haemorrhage (aSAH) and to assess its value as a real-time monitoring biomarker. DESIGN:Pre-registered multicentre prospective observational cohort study. SETTING:Eight neurosurgical tertiary centres in Switzerland, Germany and Austria between August 2021 and June 2024. PARTICIPANTS:366 patients with aSAH (mean age 58 years; 65% women). Of these, 260 provided cerebrospinal fluid (CSF) samples via external ventricular drain (EVD; 2467 samples, median 10 days per patient) and 66 via lumbar drain (LD; 379 samples, median 6 days). INTERVENTIONS:Daily CSF samples were collected via EVD or LD from day 1 to day 14 after haemorrhage; no therapeutic interventions were tested. MAIN OUTCOME MEASURES:CSF-Hb and its metabolites were analysed post hoc in a blinded manner. The primary outcome was SAH-SBI, defined as a composite of angiographic vasospasm (aVSP), delayed cerebral ischaemia (DCI) and delayed ischaemic neurological deficits (DIND), assessed daily over 14 days. Secondary outcomes included temporal CSF-Hb profiles and associations with aneurysm location, haematoma volume, intraventricular haemorrhage, chronic hydrocephalus and 3 month functional outcome. RESULTS:CSF-Hb showed a delayed peak pattern: concentrations were low after aSAH, rose to a maximum on day 10 (EVD-derived CSF-Hb median 11.3 µM, IQR 2.64 to 25.90) and then declined. Larger haematoma volume (p<0.001) and intraventricular haemorrhage (p<0.001) were associated with higher EVD-derived CSF-Hb. SAH-SBI occurred in 209/366 patients (57%). Daily EVD-derived CSF-Hb showed no association with SAH-SBI (p=0.25) and only poor prognostic potential for same-day SAH-SBI (area under the curve 0.59, 95% CI 0.56 to 0.63), with substantial between-centre heterogeneity. In a post-hoc exploratory analysis, higher CSF methaemoglobin showed a positive point-estimate of association with SAH-SBI (OR 1.18 per log(µM), 95% CI 1.02 to 1.36). Higher acute-phase EVD-derived CSF-Hb was associated with chronic hydrocephalus and a poor 3 month functional outcome. Catheter-related infection rates were low (2.2%). CONCLUSIONS:In this preregistered multicentre validation study, EVD-derived CSF-Hb did not perform as a robust real-time monitoring biomarker for SAH-SBI, showing limited same-day discrimination and substantial between-centre heterogeneity. These findings argue against clinical implementation of CSF-Hb point-measurement as a single-parameter biomarker. Higher CSF methaemoglobin was associated with SAH-SBI; this hypothesis-generating observation requires prospective confirmation and motivates continued investigation of haemolysis-related pathways. Future work using the HeMoVal biobank will apply multi-marker, pathway-level analyses to define haemolysis-related biomarker signatures and provide a platform for robust external validation of future candidates. TRIAL REGISTRATION NUMBER:NCT04998370.

Abstract Tumor-associated macrophages can either promote or suppress cancer, but therapeutic targeting remains challenging because we lack a predictive framework for macrophage function. The prevailing M1/M2 paradigm oversimplifies how macrophage developmental origin (ontogeny) and local cytokines shape antitumor versus protumor behavior. We systematically map eight reference macrophage states by differentiating mouse bone marrow cells with M-CSF or GM-CSF and polarizing them with four key cytokines (IFN‑γ, IL‑4, IL‑10, TGF‑β). Using integrated transcriptomic profiling, 3D tumor spheroids, and experimental metastasis models, we find that macrophage ontogeny determines whether cytokines promote or suppress tumor progression. Most notably, IL-4 induces opposite effects depending on ontogeny: promoting tumor growth, invasion, and metastasis in M-CSF-derived macrophages, while suppressing these processes in GM-CSF-derived macrophages. A similar ontogeny-dependent divergence was observed for IL-10, whereas IFN-γ consistently exerted antitumor effects and TGF-β protumor effects across both lineages. These findings define an ontogeny‑cytokine interaction framework that determines macrophage function based on developmental origin and cytokine context. By identifying ontogeny as a key determinant of cytokine responses, this work provides a conceptual basis for more precise macrophage-directed cancer immunotherapy strategies.

BACKGROUND:Tumor-associated macrophages (TAMs) can switch between immune-activating and cancer-promoting states; yet, the stress pathways that lock them into procancerous states remain obscure. Here we defined the role of transcription factor NRF2 as a mediator of procancerous macrophages. METHODS:We combined spatial transcriptomics, single-cell RNA sequencing, three-dimensional (3D) cell culture and in vivo tumor models to explore how NRF2 activation status in tumor-associated macrophages modifies responses to immunotherapy. RESULTS:In MC38 colon tumors, repeated anti-CD40 or radiotherapy created necrosis that split TAMs into peripheral Cxcl9+ and peri-necrotic Spp1+ subsets. Spatial transcriptomics, single-cell RNA sequencing, and Keap1-deficient mice showed that the latter are NRF2-imprinted "stress-TAMs", with immunosuppressive and tumor-promoting activity. The same NRF2 activation gradient separates pro-inflammatory CXCL9+ and anti-inflammatory SPP1+TAMs across diverse human cancers. NRF2-imprinted TAMs silence IFN-STAT1 programs, lose major histocompatibility complex-II and chemokine expression, fail to expand T cells, drive tumor cell invasion in 3D co-cultures, and foster metastasis. Constitutive hematopoietic NRF2 activation accelerated the growth of therapy-naïve MMTV-PyMT breast tumors and markedly impaired the efficacy of agonistic anti-CD40 antibody therapy in MC38 subcutaneous and lung-metastasis models. Conversely, macrophage-specific Nrf2 deletion restored immunogenic TAMs and potentiated anti-CD40 and anti-programmed cell death protein-1 treatments. CONCLUSIONS:Our data pinpoint a previously underappreciated cytoprotective mechanism, which inadvertently sustains immunosuppressive macrophages and confers therapy resistance. These results define stress-induced TAMs as an untapped driver of macrophage-based immune evasion. Inhibiting NRF2 activity alongside standard immunotherapies could restore a pro-inflammatory macrophage-T-cell amplification loop, potentially improving patient responses to T-cell-and macrophage-directed immunotherapies.

Hemolytic anemias involve premature red blood cell (RBC) destruction and present complex phenotypes, including disturbances in iron metabolism, extramedullary erythropoiesis, and systemic organ involvement. To guide the selection of appropriate murine models for studying pathophysiology and pharmacologic treatments of human hemolytic disorders, we systematically characterized three genetic mouse models commonly used to investigate such conditions: sickle cell disease (SCD), β-thalassemia (THAL), and hereditary spherocytosis (SPH). We sought to clarify how these models differ in the severity and nature of hemolysis, the balance between erythropoietic responses and iron regulation, and the long-term patterns of iron distribution. Our findings reveal that SPH mice exhibit severe intravascular hemolysis and suppressed hepcidin levels, leading to unopposed intestinal iron absorption and extensive tissue iron loading, especially in the liver. In contrast, SCD and THAL mice display predominantly extravascular hemolysis, moderate anemia, relatively stable hepcidin levels, and balanced erythropoiesis with partially regulated iron overload. Single-cell ribonucleic acid (RNA) sequencing of spleens highlighted distinct erythropoietic progenitor distributions, whereas iron-isotope tracing experiments confirmed divergent RBC turnover kinetics and tissue distribution. This study defines distinct disease trajectories for common hemolytic disease models by providing a unique comparative framework. Our work will support more informed model selection and refined experimental design to investigate hemolytic anemia pathobiology and therapeutics.

Hemorrhagic stroke triggers secondary brain injury through the red blood cell toxins hemoglobin (Hb) and heme, which fuel iron-driven lipid peroxidation and neuronal injury. We sought to use organotypic brain-slice cultures to dissect how the high-affinity scavenger proteins haptoglobin (Hp) and hemopexin (Hpx) modulate this cascade. By day 7 of culture, slices remained structurally intact, metabolically active, and responsive to oxidative stress, enabling precise toxin exposure studies. Isotopic 58Fe tracing revealed that upon cell-free Hb and heme exposures, heme-iron accumulated in brain slices and heightened lipid peroxidation. In contrast, Hpx neutralized heme, nearly abolishing iron deposition, while Hp partially reduced Hb-driven iron accumulation. Both scavengers attenuated lipid peroxidation and reduced neuronal cell death. Transcriptomic profiling revealed that free toxins increased oxidative stress and neuroinflammatory activation markers, whereas Hpx suppressed the expression of heme-induced genes. Remarkably, HbHp complexes triggered a strong Nrf2-centered adaptive program that enhanced iron metabolism and glutathione synthesis. Integrating five readouts─iron accumulation, lipid peroxidation, neuronal cell death, heme-stress transcripts, and Nrf2/metabolic transcripts─via bootstrap-based principal component analysis yielded two orthogonal axes. An oxidative toxicity axis (PC1) captured iron-driven reactive oxygen species and cell death, while a metabolic adaptation axis (PC2) reflected Nrf2-mediated reprogramming. Free toxins clustered at the toxic extreme on PC1, and heme-Hpx aligned near baseline. HbHp shifted slices upward on PC2, reducing neuronal loss through safe adaptation. These findings establish that Hpx neutralizes free heme, whereas Hp stabilizes Hb and elicits cytoprotective gene expression, offering a rational, dual-scavenger strategy to mitigate secondary brain injury in hemorrhagic stroke.

Cardiopulmonary bypass–induced hemolysis is linked to acute kidney injury in cardiac surgery. Emerging therapies targeting cell-free hemoglobin (CFHb), like haptoglobin, nitric oxide (NO), and antioxidants, show promise in reducing kidney injury, highlighting the need for further research.

Microscopic hemorrhage is a common aspect of cancers, yet its potential role as an independent factor influencing both cancer progression and therapeutic response is largely ignored. Recognizing the essential function of macrophages in red blood cell disposal, we explored a pathway that connects intratumoral hemorrhage with the formation of cancer-promoting tumor-associated macrophages (TAMs). Using spatial transcriptomics, we found that NRF2-activated myeloid cells possessing characteristics of procancerous TAMs tend to cluster in perinecrotic hemorrhagic tumor regions. These cells resembled antiinflammatory erythrophagocytic macrophages. We identified heme, a red blood cell metabolite, as a pivotal microenvironmental factor steering macrophages toward protumorigenic activities. Single-cell RNA-Seq and functional assays of TAMs in 3D cell culture spheroids revealed how elevated intracellular heme signals via the transcription factor NRF2 to induce cancer-promoting TAMs. These TAMs stabilized epithelial-mesenchymal transition, enhancing cancer invasiveness and metastatic potential. Additionally, NRF2-activated macrophages exhibited resistance to reprogramming by IFN-γ and anti-CD40 antibodies, reducing their tumoricidal capacity. Furthermore, MC38 colon adenocarcinoma-bearing mice with NRF2 constitutively activated in leukocytes were resistant to anti-CD40 immunotherapy. Overall, our findings emphasize hemorrhage-activated NRF2 in TAMs as a driver of cancer progression, suggesting that targeting this pathway could offer new strategies to enhance cancer immunity and overcome therapy resistance.

Secondary brain injury (SBI) occurs with a lag of several days post-bleeding in patients with aneurysmal subarachnoid hemorrhage (aSAH) and is a strong contributor to mortality and long-term morbidity. aSAH-SBI coincides with cell-free hemoglobin (Hb) release into the cerebrospinal fluid. This temporal association and convincing pathophysiological concepts suggest that CSF-Hb could be a targetable trigger of SBI. However, sparse experimental evidence for Hb’s neurotoxicity in vivo defines a significant research gap for clinical translation. We modeled the CSF-Hb exposure observed in aSAH patients in conscious sheep, which allowed us to assess neurological functions in a gyrencephalic species. Twelve animals were randomly assigned for 3-day bi-daily intracerebroventricular (ICV) injections of either Hb or Hb combined with the high-affinity Hb scavenger protein haptoglobin (Hb-Hp, CSL888). Repeated CSF sampling confirmed clinically relevant CSF-Hb concentrations. This prolonged CSF-Hb exposure over 3 days resulted in disturbed movement activity, reduced food intake, and impaired observational neuroscores. The Hb-induced neurotoxic effects were significantly attenuated when Hb was administered with equimolar haptoglobin. Preterminal magnetic resonance imaging (MRI) showed no CSF-Hb-specific structural brain alterations. In both groups, histology demonstrated an inflammatory response and revealed enhanced perivascular histiocytic infiltrates in the Hb-Hp group, indicative of adaptive mechanisms. Heme exposure in CSF and iron deposition in the brain were comparable, suggesting comparable clearance efficiency of Hb and Hb-haptoglobin complexes from the intracranial compartment. We identified a neurological phenotype of CSF-Hb toxicity in conscious sheep, which is rather due to neurovascular dysfunction than structural brain injury. Haptoglobin was effective at attenuating CSF-Hb-induced neurological deterioration, supporting its therapeutic potential.

Preclinical studies indicate that the systemic application of C1-inhibitor, clinically used to treat hereditary angioedema, reduces secondary brain injury after ischemic stroke. This study assessed the effect of C1-inhibitor on secondary brain injury after hemorrhagic stroke. We used an established striatal whole-blood injection mouse model to mimic intracerebral hemorrhage-related secondary brain injury. Based on the spatiotemporal dynamics in our model, we calculated the necessary sample size (n = 24) and determined the most sensitive time point to detect potential group differences (48 h) prior to the experiments. The experimental setup, tissue processing and image analysis adhered to our published protocol. We randomized mice into three groups: C1-inhibitor treatment, placebo, and sham. Histology was standardized by taking eight anatomically predefined slices across the entire lesion. Lesion size, vascular leakage, and inflammatory responses were assessed using automated thresholding and dextran/ICAM1/CD45 intensity mapping. Investigators were blinded to group allocation during the experiment, tissue processing, and image analysis. Whole blood injection resulted in significantly larger lesion size and more pronounced vascular leakage and cellular inflammation compared to the sham group. However, there was no difference in lesion size or inflammatory markers between the C1-inhibitor and placebo groups. In addition, there was no difference in the inflammatory response of the choroid plexus, which has been identified as a central organ orchestrating inflammation after intracerebral hemorrhage. The protective effect of C1-inhibitor might be isolated to pathophysiological processes with a predominant thromboinflammatory component, as in ischemia-reperfusion, but less so in permanent ischemia or intracerebral hemorrhage.

Abstract We characterized the transcriptional profiles of erythroid cells differentiated from peripheral blood mononuclear cells (PBMCs) from peripheral blood collected from patients diagnosed with Sickle Cell Disease (SCD), which have been treated with Hydroxyurea (HU) in comparison to untreated SCD patients and healthy controls (HC) using bulk RNAseq. We identified 398 differentially expressed genes (DEGs) in SCD non-treated-derived erythroid cells and 65 DEGs in SCD HU-treated patient-derived erythroid cells compared to HC. We found biological processes such as oxidative phosphorylation pathway, proteasome, autophagy, natural killer cell (NK) cytotoxicity, adaptive immune response or inflammatory response to be significantly enriched in our patient study groups by using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Our findings collectively suggest different as well as common molecular signatures between our groups. We could validate 12 of our top DEGs in treated patients by qRT-PCR. We performed additional experiments to compare the mRNA levels of mutS homolog 5- Suppressor APC Domain Containing 1 (MSH5-SAPCD1), G protein subunit gamma 4 (GNG4), stabilin 1/ clever-1 (STAB1) and Fas Binding Factor 1 (FBF1) from the bone marrow cells and spleen tissue from the Berkely SCD mouse model to the expressions observed in the transcriptome.

OBJECTIVES:Cerebrospinal fluid hemoglobin has been positioned as a potential biomarker and drug target for aneurysmal subarachnoid hemorrhage-related secondary brain injury (SAH-SBI). The maximum amount of hemoglobin, which may be released into the cerebrospinal fluid, is defined by the initial subarachnoid hematoma volume (ISHV). In patients without external ventricular or lumbar drain, there remains an unmet clinical need to predict the risk for SAH-SBI. The aim of this study was to explore automated segmentation of ISHV as a potential surrogate for cerebrospinal fluid hemoglobin to predict SAH-SBI.METHODS:This study is based on a retrospective analysis of imaging and clinical data from 220 consecutive patients with aneurysmal subarachnoid hemorrhage collected over a five-year period. 127 annotated initial non-contrast CT scans were used to train and test a convolutional neural network to automatically segment the ISHV in the remaining cohort. Performance was reported in terms of Dice score and intraclass correlation. We characterized the associations between ISHV and baseline cohort characteristics, SAH-SBI, ventriculoperitoneal shunt dependence, functional outcome, and survival. Established clinical (World Federation of Neurosurgical Societies, Hunt & Hess) and radiological (modified Fisher, Barrow Neurological Institute) scores served as references.RESULTS:A strong volume agreement (0.73 Dice, range 0.43 - 0.93) and intraclass correlation (0.89, 95% CI, 0.81-0.94) were shown. While ISHV was not associated with the use of antithrombotics or cardiovascular risk factors, there was strong evidence for an association with a lower Glasgow Coma Scale at hospital admission. Aneurysm size and location were not associated with ISHV, but the presence of intracerebral or intraventricular hemorrhage were independently associated with higher ISHV. Despite strong evidence for a positive association between ISHV and SAH-SBI, the discriminatory ability of ISHV for SAH-SBI was insufficient. The discriminatory ability of ISHV was, however, higher regarding ventriculoperitoneal shunt dependence and functional outcome at three-months follow-up. Multivariate survival analysis provided strong evidence for an independent negative association between survival probability and both ISHV and intraventricular hemorrhage.CONCLUSIONS:The proposed algorithm demonstrates strong performance in volumetric segmentation of the ISHV on the admission CT. While the discriminatory ability of ISHV for SAH-SBI was similar to established clinical and radiological scores, it showed a high discriminatory ability for ventriculoperitoneal shunt dependence and functional outcome at three-months follow-up.

ABSTRACTPurposeCell-free hemoglobin in the cerebrospinal fluid (CSF-Hb) may be one of the main drivers of secondary brain injury after aneurysmal subarachnoid hemorrhage. Haptoglobin scavenging of CSF-Hb has been shown to mitigate cerebrovascular disruption. Using digital subtraction angiography (DSA) and blood oxygenation-level dependent cerebrovascular reactivity imaging (BOLD-CVR) the aim was to assess the acute toxic effect of CSF-Hb on cerebral blood flow and autoregulation, as well as to test the protective effects of haptoglobin.MethodsDSA imaging was performed in eight anesthetized and ventilated sheep (mean weight: 80.4 kg) at baseline, 15, 30, 45 and 60 minutes after infusion of hemoglobin (Hb) or co-infusion with haptoglobin (Hb:Haptoglobin) into the left lateral ventricle. Additionally, 10 ventilated sheep (mean weight: 79.8 kg) underwent BOLD-CVR imaging to assess the cerebrovascular reserve capacity.ResultsDSA imaging did not show a difference in mean transit time or cerebral blood flow. Wholebrain BOLD-CVR compared to baseline decreased more in the Hb group after 15 minutes (Hb vs Hb:Haptoglobin: −0.03 ±0.01 vs −0.01 ±0.02) and remained diminished compared to Hb:Haptoglobin group after 30 minutes (Hb vs Hb:Haptoglobin: −0.03 ±0.01 vs 0.0 ±0.01), 45 minutes (Hb vs Hb:Haptoglobin: −0.03 ±0.01 vs 0.01 ±0.02) and 60 minutes (Hb vs Hb:Haptoglobin: −0.03 ±0.02 vs 0.01 ±0.01).ConclusionIt is demonstrated that CSF-Hb toxicity leads to rapid cerebrovascular reactivity impairment, which is blunted by haptoglobin co-infusion. BOLD-CVR may therefore be further evaluated as a monitoring strategy for CSF-Hb toxicity after aSAH.

Effective public health measures against SARS-CoV-2 require granular knowledge of population-level immune responses. We developed a Tripartite Automated Blood Immunoassay (TRABI) to assess the IgG response against three SARS-CoV-2 proteins. We used TRABI for continuous seromonitoring of hospital patients and blood donors (n = 72'250) in the canton of Zurich from December 2019 to December 2020 (pre-vaccine period). We found that antibodies waned with a half-life of 75 days, whereas the cumulative incidence rose from 2.3% in June 2020 to 12.2% in mid-December 2020. A follow-up health survey indicated that about 10% of patients infected with wildtype SARS-CoV-2 sustained some symptoms at least twelve months post COVID-19. Crucially, we found no evidence of a difference in long-term complications between those whose infection was symptomatic and those with asymptomatic acute infection. The cohort of asymptomatic SARS-CoV-2-infected subjects represents a resource for the study of chronic and possibly unexpected sequelae.

AbstractIn early 2022, a 77-year-old man presented with weight loss and recurrent subfebrile temperatures since 6 months. Workup with CT revealed a lung infiltrate. Despite antibiotic treatment, serum inflammation markers remained high. The patient further developed eczematous skin changes, uveitis (sequentially on both eyes), and macrocytic anemia. Finally, an autoinflammatory disease was suspected, and FDG PET/CT was performed. The examination revealed metabolically active foci in several tissues (tracheal cartilage, bone marrow, muscles). Bone marrow aspiration revealed anUBA1mutation, which is pathognomonic for VEXAS syndrome.

Abstract Aim Aneurysmal subarachnoid haemorrhage (aSAH) is a devastating form of stroke with an unmet need to improve outcome. This study focusses on the development of intrathecal haptoglobin supplementation as a treatment by reviewing current knowledge and progress and arriving at a Delphi-based global consensus regarding the role of extracellular haemoglobin and research priorities for clinical translation of haemoglobin-scavenging strategies. Method A narrative review of the topic by searching PubMed with relevant keywords and through discussions with leading expert in the field of aSAH. A modified Delphi processes. Four virtual meetings were held, two prior to starting, and two during the first round. Otherwise, the study was conducted electronically. Results Clinicians (n = 72) and scientific experts (n = 28) from five continents participated in the Delphi study. Microvascular spasm, iInflammation, microvascular spasm, and initial intracranial pressure rise were deemed the most important pathophysiological pathways determining outcome. Extracellular haemoglobin was thought to play an important role mostly in iron toxicity, oxidative stress, nitric oxide homeostasis and inflammation. While useful, there was consensus that further preclinical work was not a priority, with most believing the field to be ready for early phase trial. The highest priorities were related to confirming haptoglobin’s anticipated safety, individualized versus standard dosing, timing of treatment, pharmacokinetics, pharmacodynamics and outcome measure selection. Conclusions These results highlight the pressing need for early phase trials of intracranial haptoglobin for aSAH, and the immense value of early input from clinical disciplines on a global scale during the early stages of clinical translation.

BackgroundAgonistic anti-CD40 monoclonal antibodies (mAbs) have emerged as promising immunotherapeutic compounds with impressive antitumor effects in mouse models. However, preclinical and clinical studies faced dose-limiting toxicities mediated by necroinflammatory liver disease. An effective prophylactic treatment for liver immune-related adverse events that does not suppress specific antitumor immunity remains to be found.MethodsWe used different mouse models and time-resolved single-cell RNA-sequencing to characterize the pathogenesis of anti-CD40 mAb induced liver toxicity. Subsequently, we developed an antibody-based treatment protocol to selectively target red blood cells (RBCs) for erythrophagocytosis in the liver, inducing an anti-inflammatory liver macrophage reprogramming.ResultsWe discovered that CD40 signaling in Clec4f(+) Kupffer cells is the non-redundant trigger of anti-CD40 mAb-induced liver toxicity. Taking advantage of the highly specific functionality of liver macrophages to clear antibody-tagged RBCs from the blood, we hypothesized that controlled erythrophagocytosis and the linked anti-inflammatory signaling by the endogenous metabolite heme could be exploited to reprogram liver macrophages selectively. Repeated low-dose administration of a recombinant murine Ter119 antibody directed RBCs for selective phagocytosis in the liver and skewed the phenotype of liver macrophages into a Hmox(high)/Marco(high)/MHCIIlow anti-inflammatory phenotype. This unique mode of action prevented necroinflammatory liver disease following high-dose administration of anti-CD40 mAbs. In contrast, extrahepatic inflammation, antigen-specific immunity, and antitumor activity remained unaffected in Ter119 treated animals.ConclusionsOur study offers a targeted approach to uncouple CD40-augmented antitumor immunity in peripheral tissues from harmful inflammatoxicity in the liver.

Immune checkpoint inhibitors (ICI) have rapidly changed the treatment landscape in oncology. However, the majority of cancer patients do not benefit from immunotherapies (primary resistance) or present with only short-term benefit (secondary resistance). IMMUcan is a public-private European consortium funded by the Innovative Medicines Initiative, set-up in 2019, to elucidate the relationship between host and cancer cells within the tumor microenvironment (TME) and the impact of current standard of care treatments in up to 3,000 cancer patients. By integrating molecular (whole exome sequencing and total RNA sequencing) and cellular data (multiplex immunofluorescence (IF) and imaging mass cytometry (IMC)) of patient tumors with longitudinal clinical information, the tumor-host interaction and the response to therapies can be dissected. This represents a unique multi-modal dataset facilitating the discovery of novel biomarkers and resistance mechanisms. Thus, IMMUcan arguably represents one of Europe's largest TME-profiling efforts to date. More than 100 investigators from 17 European countries are prospectively recruiting patients with non-small cell lung cancer (NSCLC), head and neck squamous cell carcinoma (HNSCC), triple negative breast cancer (TNBC), renal cell carcinoma and colorectal cancer, using the EORTC SPECTA platform (NCT02834884). To date, more than half of the targeted patients have been recruited, including almost 400 HNSCC and NSCLC as well as more than 600 breast cancer patients. The first analysis on these cohorts will be presented during the ESMO-IO conference highlighting the novel biomarkers identified in TNBC (early and metastatic settings), the resistance mechanisms of HNSCC patients treated with immunotherapies, as well as the integration of molecular and cellular modalities to gain an understanding of advanced NSCLC disease.

Aneurysmal subarachnoid hemorrhage (aSAH) is a devastating form of stroke frequently affecting young to middle-aged adults, with an unmet need to improve outcome. This special report focusses on the development of intrathecal haptoglobin supplementation as a treatment by reviewing current knowledge and progress, arriving at a Delphi-based global consensus regarding the pathophysiological role of extracellular hemoglobin and research priorities for clinical translation of hemoglobin-scavenging therapeutics. After aneurysmal subarachnoid hemorrhage, erythrocyte lysis generates cell-free hemoglobin in the cerebrospinal fluid, which is a strong determinant of secondary brain injury and long-term clinical outcome. Haptoglobin is the body’s first-line defense against cell-free hemoglobin by binding it irreversibly, preventing translocation of hemoglobin into the brain parenchyma and nitric oxide-sensitive functional compartments of cerebral arteries. In mouse and sheep models, intraventricular administration of haptoglobin reversed hemoglobin-induced clinical, histological, and biochemical features of human aneurysmal subarachnoid hemorrhage. Clinical translation of this strategy imposes unique challenges set by the novel mode of action and the anticipated need for intrathecal drug administration, necessitating early input from stakeholders. Practising clinicians (n=72) and scientific experts (n=28) from 5 continents participated in the Delphi study. Inflammation, microvascular spasm, initial intracranial pressure increase, and disruption of nitric oxide signaling were deemed the most important pathophysiological pathways determining outcome. Cell-free hemoglobin was thought to play an important role mostly in pathways related to iron toxicity, oxidative stress, nitric oxide, and inflammation. While useful, there was consensus that further preclinical work was not a priority, with most believing the field was ready for an early phase trial. The highest research priorities were related to confirming haptoglobin’s anticipated safety, individualized versus standard dosing, timing of treatment, pharmacokinetics, pharmacodynamics, and outcome measure selection. These results highlight the need for early phase trials of intracranial haptoglobin for aneurysmal subarachnoid hemorrhage, and the value of early input from clinical disciplines on a global scale during the early stages of clinical translation.