LPS-induced expression and release of monocyte tissue factor in patients with haemophilia

In haemophilia, thrombin generation and fibrin deposition upon vascular injury critically depend on the tissue factor (TF)-driven coagulation pathway. TF expression by monocytes/macrophages and circulating microvesicles contributes to haemostasis, thrombosis and inflammation. Inflammation is a hallmark of blood-induced joint disease. The aim of this study is to correlate TF production by whole-blood monocytes with inflammatory markers and clinical parameters in patients with moderate-to-severe haemophilia A or B (n = 43) in comparison to healthy males (n = 23). Monocyte TF antigen and microvesicle-associated TF procoagulant activity (MV TF PCA) were measured immediately after blood draw (baseline) and following incubation of whole blood with buffer or lipopolysaccharide (LPS) using two-colour flow cytometry and chromogenic FXa generation assay, respectively. Patients with HIV or uncontrolled HBV/HCV infections were excluded. TF was hardly detectable and not different in baseline and buffer-treaded samples from both groups. Stimulation with LPS, however, induced monocyte TF production, with increased TF-specific mean fluorescence intensity (P = 0.08) and MV TF PCA (P < 0.05) in patients compared to controls. Patients also had elevated hs-CRP and IL-6 serum levels (P < 0.001), which correlated with LPS-induced TF parameters. Further exploratory analyses revealed that the presence of systemic (low-grade) inflammation and boosted LPS-induced monocyte TF production were mainly restricted to patients with clinically controlled HBV and/or HCV infection (n = 16), who were older and also had a significantly worse orthopaedic joint score than patients with no history of viral hepatitis (P < 0.01). Our study delineates a previously unrecognised link between systemic inflammation and inducible monocyte TF production in patients with haemophilia A or B

Monitoring multiple myeloma by next-generation sequencing of V(D)J rearrangements from circulating myeloma cells and cell-free myeloma DNA

Recent studies suggest that circulating tumor cells and cell-free DNA may represent powerful non-invasive tools for monitoring disease in patients with solid and hematologic malignancies. Here, we conducted a pilot study in 27 myeloma patients to explore the clonotypic V(D)J rearrangement for monitoring circulating myeloma cells and cell-free myeloma DNA. Next-generation sequencing was used to define the myeloma V(D)J rearrangement and for subsequent peripheral blood tracking after treatment initiation. Positivity for circulating myeloma cells/cell-free myeloma was associated with conventional remission status (

Cetuximab Resistance in Head and Neck Cancer Is Mediated by EGFR-K 521 Polymorphism

Head and neck squamous cell carcinomas (HNSCC) exhibiting resistance to the EGFR-targeting drug cetuximab poses a challenge to their effective clinical management. Here, we report a specific mechanism of resistance in this setting based upon the presence of a single nucleotide polymorphism encoding EGFR-K-521 (K-allele), which is expressed in > 40% of HNSCC cases. Patients expressing the K-allele showed significantly shorter progressionfree survival upon palliative treatment with cetuximab plus chemotherapy or radiation. In several EGFR-mediated cancer models, cetuximab failed to inhibit downstream signaling or to kill cells harboring a high K-allele frequency. Cetuximab affinity for EGFR-K-521 was reduced slightly, but ligand-mediated EGFR acti-vation was intact. We found a lack of glycan sialyation on EGFR-K-521 that associated with reduced protein stability, suggesting a structural basis for reduced cetuximab efficacy. CetuGEX, an antibody with optimized Fc glycosylation targeting the same epitope as cetuximab, restored HNSCC sensitivity in a manner associated with antibody-dependent cellular cytotoxicity rather than EGFR pathway inhibition. Overall, our results highlight EGFR-K-521 expression as a key mechanism of cetuximab resistance to evaluate prospectively as a predictive biomarker in HNSCC patients. Further, they offer a preclinical rationale for the use of ADCC-optimized antibodies to treat tumors harboring this EGFR isoform

Thromboembolic events in deceased patients with proven SARS-CoV-2 infection: frequency, characteristics and risk factors

Background: Infection with the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) results in respiratory syndromes but also in vascular complications such as thromboembolism (TE). In this regard, immunothrombosis, resulting from inflammation in SARS-CoV-2 infected tissues, has been described. Data on TE in COVID-19 are mainly based on clinical observational and/or incomplete autopsy studies. The true burden of TE and the relevance of genetic predisposition, however, have not been resolved. Objectives: Here, we report on a consecutive cohort of 100 fully autopsied patients deceased by SARS-CoV-2 infections during the first wave of the pandemic (March to April 2020). We investigated the localization of TE, potential clinical risk factors, and the prothrombotic gene mutations, factor V Leiden and prothrombin G20210A, in postmortem blood or tissue samples. Results: TE was found in 43/100 autopsies. 93 % of TE events were venous occlusions, with 23 patients having pulmonary thromboembolism (PT) with or without lower-extremity deep vein thrombosis. Of these, 70 % showed PT restricted to (sub)segmental arteries, consistent with in situ immunothrombosis. Patients with TE had a significantly higher BMI and died more frequently at an intensive care unit. Hereditary thrombophilia factors were not associated with TE. Conclusions: Our autopsy results show that a significant proportion of SARS-CoV-2 infected patients suffer from TE, affecting predominantly the venous system. Orthotopic peripheral PT was the most frequent finding. Hereditary thrombophilia appears not to be a determinant for TE in COVID-19. However, obesity and the need for intensive care increase the risk of TE in these patients</p