0000000000001839
AUTHOR
Claudine Graf
Tissue factor pathway inhibitor primes monocytes for antiphospholipid antibody-induced thrombosis
Antiphospholipid antibodies (aPLs) with complex lipid and/or protein reactivities cause complement-dependent thrombosis and pregnancy complications. Although cross-reactivities with coagulation regulatory proteins contribute to the risk for developing thrombosis in patients with antiphospholipid syndrome, the majority of pathogenic aPLs retain reactivity with membrane lipid components and rapidly induce reactive oxygen species-dependent proinflammatory signaling and tissue factor (TF) procoagulant activation. Here, we show that lipid-reactive aPLs activate a common species-conserved TF signaling pathway. aPLs dissociate an inhibited TF coagulation initiation complex on the cell surface of m…
EPCR Signaling Controls the Activity of Hematopoietic Stem Cells Independent of Coagulation Regulation
Abstract Background All mature blood cells are derived from multipotent hematopoietic stem cells (HSCs) which are activated to meet the demand of the host during inflammation and injury. The endothelial cell protein C receptor (EPCR) is a marker for primitivity and quiescence of HSCs but the relative contributions of EPCR signaling versus anticoagulant functions in HSC maintenance are incompletely defined. Aims We aimed to dissect functions of EPCR by studying anticoagulant and signaling function in HSC of EPCR C/S mice carrying a single intracellular point mutation abolishing normal trafficking of EPCR through endo-lysosomal compartments. We assessed the contributions of EPCR signaling to …
Cognate HLA absence in trans diminishes human NK cell education
NK cells are innate lymphocytes with protective functions against viral infections and tumor formation. Human NK cells carry inhibitory killer cell Ig-like receptors (KIRs), which recognize distinct HLAs. NK cells with KIRs for self-HLA molecules acquire superior cytotoxicity against HLA– tumor cells during education for improved missing-self recognition. Here, we reconstituted mice with human hematopoietic cells from donors with homozygous KIR ligands or with a mix of hematopoietic cells from these homozygous donors, allowing assessment of the resulting KIR repertoire and NK cell education. We found that co-reconstitution with 2 KIR ligand–mismatched compartments did not alter the frequenc…
Myeloid cell-synthesized coagulation Factor X dampens anti-tumor immunity
Immune evasion in the tumor microenvironment (TME) is a crucial barrier for effective cancer therapy, and plasticity of innate immune cells may contribute to failures of targeted immunotherapies. Here, we show that rivaroxaban, a direct inhibitor of activated coagulation factor X (FX), promotes antitumor immunity by enhancing infiltration of dendritic cells and cytotoxic T cells at the tumor site. Profiling FX expression in the TME identifies monocytes and macrophages as crucial sources of extravascular FX. By generating mice with immune cells lacking the ability to produce FX, we show that myeloid cell-derived FX plays a pivotal role in promoting tumor immune evasion. In mouse models of ca…
EPCR/PAR1 Signaling Navigates Long-Term Repopulating Hematopoietic Stem Cell Bone Marrow Homing to Thrombomodulin-Enriched Blood Vessels
Abstract Bone marrow (BM) homing and lodgment of long-term repopulating hematopoietic stem cells (LT-HSCs) is an active and essential first step in clinical stem cell transplantation. EPCR is expressed by murine BM LT-HSCs endowed with the highest repopulation potential and its ligand, activated protein C (aPC), has anticoagulant and anti-sepsis effects in EPCR+/PAR1+ endothelial cells. We recently found that signaling cascades, traditionally viewed as coagulation and inflammation related, also independently control EPCR+ LT-HSC BM retention and recruitment to the blood via distinct PAR1 mediated pathways. EPCR/PAR1 signaling retains LT-HSCs in the BM by restricting nitric oxide (NO) produc…
Strong Expression of Chemokine Receptor CXCR4 by Renal Cell Carcinoma Correlates with Advanced Disease
Diverse chemokines and their receptors have been associated with tumor growth, tumor dissemination, and local immune escape. In different tumor entities, the level of chemokine receptorCXCR4 expression has been linked with tumor progression and decreased survival. The aim of this study was to evaluate the influence ofCXCR4 expression on the progression of human renal cell carcinoma.CXCR4 expression of renal cell carcinoma was assessed by immunohistochemistry in 113 patients. Intensity ofCXCR4 expression was correlated with both tumor and patient characteristics. Human renal cell carcinoma revealed variable intensities ofCXCR4 expression. StrongCXCR4 expression of renal cell carcinoma was si…
Tissue factor as a mediator of coagulation and signaling in cancer and chronic inflammation
Thrombosis is frequently diagnosed as a first symptom in tumor patients and the clinical management of hypercoagulability in cancer patients remains challenging due to concomitant changes in risk factors for severe bleeding. It therefore remains a priority to better understand interactions of the hemostatic system with cancer biology. Specifically, further research is needed to elucidate the details and effects of new anticoagulants on extravascular coagulation and the interplay between cancer progression and chronic inflammation. In addition, it will be important to identify subgroups of cancer patients benefiting from specific modulations of the coagulation system without increasing the b…
A neoepitope generated by an FLT3 internal tandem duplication (FLT3-ITD) is recognized by leukemia-reactive autologous CD8+ T cells.
Abstract The FLT3 receptor tyrosine kinase is expressed in more than 90% of acute myelogeneous leukemias (AMLs), up to 30% of which carry an internal tandem duplication (ITD) within the FLT3 gene. Although varying duplication sites exist, most FLT3-ITDs affect a single protein domain. We analyzed the FLT3-ITD of an AML patient for encoding HLA class I–restricted immunogenic peptides. One of the tested peptides (YVDFREYEYY) induced in vitro autologous T-cell responses restricted by HLA-A*0101 that were also detectable ex vivo. These peptide-reactive T cells recognized targets transfected with the patient's FLT3-ITD, but not wild-type FLT3, and recognized the patient's AML cells. Our results …
Erratum to: Cetuximab-induced skin exanthema: prophylactic and reactive skin therapy are equally effective
Purpose Treatment with cetuximab is accompanied by the development of an acneiform follicular skin exanthema in more than 80 % of patients. Severe exanthema (grade III/IV) develops in about 9–19 % of patients with the necessity of cetuximab dose reduction or cessation. Methods The study presented was a retrospective analysis of 50 gastrointestinal cancer patients treated with cetuximab in combination with either FOLFIRI or FOLFOX. One cohort of 15 patients received an in-house reactive skin protocol upon development of an exanthema. A second cohort of 15 patients received a skin prophylaxis starting with the first dose of cetuximab before clinical signs of toxicity. A third historic group o…
Analysis of the expression of SDF-1 splicing variants in human colorectal cancer and normal mucosa tissues
C-X-C motif chemokine ligand 12 (CXCL12), also termed stromal cell-derived factor-1 (SDF-1) is a small protein 8-14 kDa in length that is expressed as six isoforms, consisting of SDF-1α, SDF-1β, SDF-1γ, SDF-1δ, SDF-1ε and SDF-1θ. All six isoforms are encoded by the single CXCL12 gene on chromosome 10. This gene regulates leukocyte trafficking and is variably expressed in a number of normal and cancer tissues. The potential role of the novel CXCL12 splice variants as components of the CXCR4 axis in cancer development is not fully understood. The present study aimed to analyze the expression profile of the various SDF-1 isoforms and SDF-1 polymorphisms, and the association with the clinicopat…
Xenograft models for undifferentiated pleomorphic sarcoma not otherwise specified are essential for preclinical testing of therapeutic agents
Undifferentiated pleomorphic sarcoma not otherwise specified belongs to the heterogeneous group of soft tissue tumors. It is preferentially located in the upper and lower extremities of the body, and surgical resection remains the only curative treatment. Preclinical animal models are crucial to improve the development of novel chemotherapeutic agents for the treatment of undifferentiated pleomorphic sarcoma. However, this approach has been hampered by the lack of reproducible animal models. The present study established two xenograft animal models generated from stable non-clonal cell cultures, and investigated the difference in chemotherapeutic effects on tumor growth between undifferenti…
Cytoskeletal stabilization of inhibitory interactions in immunologic synapses of mature human dendritic cells with natural killer cells
Abstract Human mature dendritic cells (DCs) can efficiently stimulate natural killer (NK)–cell responses without being targeted by their cytotoxicity. To understand this important regulatory crosstalk, we characterized the development of the immunologic synapse between mature DCs and resting NK cells. Conjugates between these 2 innate leukocyte populations formed rapidly, persisted for prolonged time periods and matured with DC-derived f-actin polymerization at the synapse. Polarization of IL-12 and IL-12R to the synapse coincided with f-actin polymerization, while other activating and inhibitory molecules were enriched at the interface between DCs and NK cells earlier. Functional assays re…
Anticoagulation with Factor Xa Inhibitors Is Associated with Improved Overall Response and Progression-Free Survival in Patients with Metastatic Malignant Melanoma Receiving Immune Checkpoint Inhibitors—A Retrospective, Real-World Cohort Study
Immune checkpoint inhibitors (ICI) significantly improved the prognosis of advanced melanoma patients. However, many patients do not derive long-term benefit from ICI therapy due to primary and acquired resistance. In this regard, it has been shown that coagulation factors contribute to cancer immune evasion and might therefore promote resistance to ICI. In particular, recent observations in murine systems demonstrated that myeloid-derived factor Xa (FXa) impedes anti-tumor immunity in the tumor microenvironment and that the oral FXa inhibitor (FXa-i) rivaroxaban synergizes with ICI. The synergistic effect of FXa inhibitors with clinical ICI therapy is unknown. We performed a retrospective …
PAR1 signaling regulates the retention and recruitment of EPCR-expressing bone marrow hematopoietic stem cells
Retention of long-term repopulating hematopoietic stem cells (LT-HSCs) in the bone marrow is essential for hematopoiesis and for protection from myelotoxic injury. We report that signaling cascades that are traditionally viewed as coagulation related also control retention of endothelial protein C receptor-positive (EPCR(+)) LT-HSCs in the bone marrow and their recruitment to the blood via two pathways mediated by protease activated receptor 1 (PAR1). Thrombin-PAR1 signaling induces nitric oxide (NO) production, leading to EPCR shedding mediated by tumor necrosis factor-α-converting enzyme (TACE), enhanced CXCL12-CXCR4-induced motility and rapid stem and progenitor cell mobilization. Conver…
Regulation of long-term repopulating hematopoietic stem cells by EPCR/PAR1 signaling
The common developmental origin of endothelial and hematopoietic cells is manifested by coexpression of several cell surface receptors. Adult murine bone marrow (BM) long-term repopulating hematopoietic stem cells (LT-HSCs), endowed with the highest repopulation and self-renewal potential, express endothelial protein C receptor (EPCR), which is used as a marker to isolate them. EPCR/protease-activated receptor-1 (PAR1) signaling in endothelial cells has anticoagulant and anti-inflammatory roles, while thrombin/PAR1 signaling induces coagulation and inflammation. Recent studies define two new PAR1-mediated signaling cascades that regulate EPCR(+) LT-HSC BM retention and egress. EPCR/PAR1 sig…
Rapid molecular dissection of viral and bacterial immunomes
The development of preventive or therapeutic recombinant vaccines and the generation of serodiagnostic assays for infectious diseases depend essentially on the availability of molecularly defined antigens. A major bottleneck for the identification of suitable target antigens for many pathogens is the isolation of sufficient amounts of material for subsequent genomic or proteomic screening. Applying a highly efficient expression cloning strategy to the human pathogens vaccinia virus (VV) and Chlamydia pneumoniae (CP), we demonstrate that sub-nanogram amounts of isolated nucleic acids can be utilized to determine comprehensive sets of immunodominant antigens. Remarkably, the approach not only…
Cetuximab-induced skin exanthema: prophylactic and reactive skin therapy are equally effective
Purpose Treatment with cetuximab is accompanied by the development of an acneiform follicular skin exanthema in more than 80 % of patients. Severe exanthema (grade III/IV) develops in about 9–19 % of patients with the necessity of cetuximab dose reduction or cessation.
Macrophage Factor Xa Signaling Promotes Cancer Immune Evasion
Abstract Coagulation signaling through protease activated receptors (PARs) participates in inflammation and immunity. In cancer, tissue factor (TF) driven signaling via PAR2 promotes tumor progression, but effective pharmacological strategies to inhibit the PAR2 activating proteases for clinical anti-cancer benefit are currently unknown. To gain a better understanding of signaling by coagulation proteases, we generated PAR2 mouse strains with mutations that abolish canonical proteolysis by all proteases including FVIIa (PAR2 R38E) or create specific resistance to cleavage by the TF-FVIIa-Xa signaling complex (PAR2 G37I) that requires the endothelial cell protein C receptor (EPCR, Procr). As…
PDGFRa/β expression correlates with the metastatic behavior of human colorectal cancer: A possible rationale for a molecular targeting strategy
As new multi-target tyrosine kinase inhibitors are emerging in the therapy of various malignancies, our aim was to define the co-expression pattern of receptor-tyrosine-kinase platelet-derived growth factor receptors alpha and beta (PDGFRalpha/beta) in human colorectal cancer. The co-expression pattern of PDGFRalpha/beta was analyzed by RT-PCR in 99 histologically confirmed human colorectal carcinomas and five colorectal cancer cell lines. In addition, immunohistochemical (IHC) staining was applied for confirmation of expression and analysis of receptor tyrosine kinase (RTK) localisation. The colorectal cancer cell lines that were analysed revealed varying expression intensities of PDGFRalp…
Identification of T cell epitopes by the use of rapidly generated mRNA fragments.
Abstract Although the number of defined T cell epitopes of clinically relevant antigens is constantly increasing, there is still an enormous need to identify further peptides, processed from new antigens or presented by rare HLA molecules, respectively. Here we introduce a novel two-step approach for the rapid identification of T cell epitopes. It was established in the CMV infection model. From the peripheral blood of healthy donors sharing HLA-A1 according to HLA serotyping we isolated CD8 + T lymphocytes and generated dendritic cells (DCs). DCs were electroporated with CMV pp65 mRNA and tested for recognition by autologous CD8 + T lymphocytes in IFN-γ ELISPOT assays. In all 10 CMV-seropo…
Coagulation signaling and cancer immunotherapy.
The last decades have delineated many interactions of the hemostatic system with cancer cells that are pivotal for cancer-associated thrombosis, angiogenesis and metastasis. Expanding evidence shows that platelets, the tissue factor pathway, and proteolytic signaling involving protease-activated receptors (PARs) are also central players in innate and adaptive immunity. Recent studies in immune-competent mice have uncovered new immune-evasive roles of coagulation signaling networks in the development and growth of different preclinical tumor models. Tumor-type specific PAR1 signaling facilitates the escape from immune surveillance by cytotoxic T cells. In addition, tumor-associated macrophag…