Karin Hartmann

Standards and standardization in mastocytosis: Consensus Statements on Diagnostics, Treatment Recommendations and Response Criteria

Although a classification for mastocytosis and diagnostic criteria are available, there remains a need to define standards for the application of diagnostic tests, clinical evaluations, and treatment responses. To address these demands, leading experts discussed current issues and standards in mastocytosis in a Working Conference. The present article provides the resulting outcome with consensus statements, which focus on the appropriate application of clinical and laboratory tests, patient selection for interventional therapy, and the selection of appropriate drugs. In addition, treatment response criteria for the various clinical conditions, disease‐specific symptoms, and specific pathologies are provided. Resulting recommendations and algorithms should greatly facilitate the management of patients with mastocytosis in clinical practice, selection of patients for therapies, and the conduct of clinical trials.

Definitions, Criteria and Global Classification of Mast Cell Disorders with Special Reference to Mast Cell Activation Syndromes: A Consensus Proposal

Activation of tissue mast cells (MCs) and their abnormal growth and accumulation in various organs are typically found in primary MC disorders also referred to as mastocytosis. However, increasing numbers of patients are now being informed that their clinical findings are due to MC activation (MCA) that is neither associated with mastocytosis nor with a defined allergic or inflammatory reaction. In other patients with MCA, MCs appear to be clonal cells, but criteria for diagnosing mastocytosis are not met. A working conference was organized in 2010 with the aim to define criteria for diagnosing MCA and related disorders, and to propose a global unifying classification of all MC disorders and pathologic MC reactions. This classification includes three types of ‘MCA syndromes’ (MCASs), namely primary MCAS, secondary MCAS and idiopathic MCAS. MCA is now defined by robust and generally applicable criteria, including (1) typical clinical symptoms, (2) a substantial transient increase in serum total tryptase level or an increase in other MC-derived mediators, such as histamine or prostaglandin D2, or their urinary metabolites, and (3) a response of clinical symptoms to agents that attenuate the production or activities of MC mediators. These criteria should assist in the identification and diagnosis of patients with MCAS, and in avoiding misdiagnoses or overinterpretation of clinical symptoms in daily practice. Moreover, the MCAS concept should stimulate research in order to identify and exploit new molecular mechanisms and therapeutic targets.

Monocyte Chemoattractant Protein-1 Enhances Gene Expression and Synthesis of Matrix Metalloproteinase-1 in Human Fibroblasts by an Autocrine IL-1α Loop

Monocyte chemoattractant protein-1 (MCP-1), a member of the C-C chemokine superfamily, has recently been shown to be involved in the pathogenesis of tissue fibrosis. In vitro studies demonstrated that MCP-1 up-regulates type I collagen gene expression via endogenous production of TGF-β in rat lung fibroblasts. We here show that recombinant human MCP-1 affects gene expression of interstitial collagenase (matrix metalloproteinase-1 (MMP-1)) in primary human skin fibroblasts and a stable fibroblast cell line. MMP-1 mRNA was induced by MCP-1 (10 ng/ml) as early as 6 h and reached a maximal expression at 24 h. MCP-1 also caused an increase of MMP-2 mRNA expression in both types of fibroblasts at 48 h. Interestingly, tissue inhibitor of metalloproteinase-1 (TIMP-1) mRNA was also up-regulated by MCP-1, and TIMP-1 mRNA expression peaked at 48 h in both types of fibroblasts. Immunoblot analysis demonstrated increased levels of MMP-1 and TIMP-1 protein in the culture supernatants of primary fibroblasts stimulated with MCP-1. In addition, MCP-1 strongly induced IL-1α mRNA expression in dermal fibroblasts in parallel with the induction of MMP-1. Preincubation with IL-1 receptor antagonist almost completely abrogated the expression of MMP-1 mRNA, and partially inhibited MMP-1 synthesis induced by MCP-1. Transient transfection of primary skin fibroblasts with a MMP-1 promoter-reporter construct indicated a dose-dependent increase in promoter activity by MCP-1 stimulation. These data demonstrate that MCP-1 up-regulates MMP-1 mRNA expression and synthesis in human skin fibroblasts at a transcriptional level and provide evidence that this is mediated by an IL-1α autocrine loop.

Mast cell and macrophage chemokines CXCL1/CXCL2 control the early stage of neutrophil recruitment during tissue inflammation.

Neutrophil recruitment is an important early step in controlling tissue infections or injury. Here, we report that this influx depends on both tissue-resident mast cells and macrophages. Mice with mast cell deficiency recruit reduced numbers of neutrophils in the first few hours of intraperitoneal lipopolysaccharide (LPS) stimulation. Conversely, in mice with clodronate-ablated macrophages, neutrophils extravasate, but have limited ability to reach the peritoneal fluid. Tissue macrophages synthesize neutrophil chemoattractants CXCL1/CXCL2 (CXC chemokine ligands 1/2) in response to LPS. Mast cells also produce these chemokines of which a proportion are preformed in granules. Release of the granules and new CXCL1/CXCL2 synthesis is Toll-like receptor 4-dependent. Both in vivo studies with blocking monoclonal antibodies and in vitro chemotaxis experiments show the neutrophil response to mast cells and macrophages to be CXCL1/CXCL2-dependent. The data are in keeping with the model that mast cells, optimally positioned in close proximity to the vasculature, initiate an early phase of neutrophil recruitment by releasing the chemoattractants CXCL1/CXCL2. Having arrived within the stimulated tissue, neutrophils penetrate further in a macrophage-dependent manner. Therefore, we demonstrate a positive role for mast cells in tissue inflammation and define how this comes about with contribution from a second tissue cell, the macrophage.

Pathogenic role for skin macrophages in a mouse model of keratinocyte-induced psoriasis-like skin inflammation

Psoriasis is a common skin disease, the pathogenesis of which has not yet been resolved. In mice, epidermis-specific deletion of inhibitor of NF-kappaB (IkappaB) kinase 2 (IKK2) results in a skin phenotype that mimics human psoriasis in several aspects. Like psoriasis, this skin disease shows pronounced improvement when mice are treated with a TNF-neutralizing agent. We have found previously that this phenotype does not depend on the presence of alphabeta T lymphocytes. In order to evaluate contributions of other immune cell populations to the skin disease, we selectively eliminated macrophages and granulocytes from the skin of mice with epidermis-specific deletion of IKK2 (K14-Cre-IKK2fl/fl mice). Elimination of skin macrophages by subcutaneous injection of clodronate liposomes was accompanied by inhibition of granulocyte migration into the skin and resulted in a dramatic attenuation of psoriasis-like skin changes. The hyperproliferative, inflammatory skin disease in K14-Cre-IKK2fl/fl mice was a direct consequence of the presence of macrophages in the skin, as targeted deletion of CD18, which prevented accumulation of granulocytes but not macrophages, did not lead to major changes in the phenotype. Targeted deletion of the receptor for IFN-gamma revealed that the pathogenesis of the skin disease does not depend on classical IFN-gamma-mediated macrophage activation. Our results demonstrate that in mice epidermal keratinocytes can initiate a hyperproliferative, inflammatory, IFN-gamma-independent, psoriasis-like skin disease whose development requires essential contributions from skin macrophages but not from granulocytes or alphabeta T lymphocytes.

The receptor tyrosine kinase c-Kit controls IL-33 receptor signaling in mast cells

Members of the Toll/interleukin-1 receptor (TIR) family are of importance for host defense and inflammation. Here we report that the TIR-family member interleukin-33R (IL-33R) cross-activates the receptor tyrosine kinase c-Kit in human and murine mast cells. The IL-33R-induced activation of signal transducer and activator of transcription 3 (STAT3), extracellular signal-regulated kinase 1/2 (Erk1/2), protein kinase B (PKB), and Jun NH(2)-terminal kinase 1 (JNK1) depends on c-Kit and is required to elicit optimal effector functions. Costimulation with the c-Kit ligand stem cell factor (SCF) is necessary for IL-33-induced cytokine production in primary mast cells. The structural basis for this cross-activation is the complex formation between c-Kit, IL-33R, and IL-1R accessory protein (IL-1RAcP). We found that c-Kit and IL-1RAcP interact constitutively and that IL-33R joins this complex upon ligand binding. Our findings support a model in which signals from seemingly disparate receptors are integrated for full cellular responses.

Efficacy and Safety of Midostaurin in Advanced Systemic Mastocytosis

BACKGROUND Advanced systemic mastocytosis comprises rare hematologic neoplasms that are associated with a poor prognosis and lack effective treatment options. The multikinase inhibitor midostaurin inhibits KIT D816V, a primary driver of disease pathogenesis. METHODS We conducted an open-label study of oral midostaurin at a dose of 100 mg twice daily in 116 patients, of whom 89 with mastocytosis-related organ damage were eligible for inclusion in the primary efficacy population; 16 had aggressive systemic mastocytosis, 57 had systemic mastocytosis with an associated hematologic neoplasm, and 16 had mast-cell leukemia. The primary outcome was the best overall response. RESULTS The overall response rate was 60% (95% confidence interval [CI], 49 to 70); 45% of the patients had a major response, which was defined as complete resolution of at least one type of mastocytosis-related organ damage. Response rates were similar regardless of the subtype of advanced systemic mastocytosis, KIT mutation status, or exposure to previous therapy. The median best percentage changes in bone marrow mast-cell burden and serum tryptase level were -59% and -58%, respectively. The median overall survival was 28.7 months, and the median progression-free survival was 14.1 months. Among the 16 patients with mast-cell leukemia, the median overall survival was 9.4 months (95% CI, 7.5 to not estimated). Dose reduction owing to toxic effects occurred in 56% of the patients; re-escalation to the starting dose was feasible in 32% of those patients. The most frequent adverse events were low-grade nausea, vomiting, and diarrhea. New or worsening grade 3 or 4 neutropenia, anemia, and thrombocytopenia occurred in 24%, 41%, and 29% of the patients, respectively, mostly in those with preexisting cytopenias. CONCLUSIONS In this open-label study, midostaurin showed efficacy in patients with advanced systemic mastocytosis, including the highly fatal variant mast-cell leukemia. (Funded by Novartis Pharmaceuticals and others; ClinicalTrials.gov number, NCT00782067.).

Proposed diagnostic algorithm for patients with suspected mastocytosis: a proposal of the European Competence Network on Mastocytosis.

Mastocytosis is an emerging differential diagnosis in patients with more or less specific mediator‐related symptoms. In some of these patients, typical skin lesions are found and the diagnosis of mastocytosis can be established. In other cases, however, skin lesions are absent, which represents a diagnostic challenge. In the light of this unmet need, we developed a diagnostic algorithm for patients with suspected mastocytosis. In adult patients with typical lesions of mastocytosis in the skin, a bone marrow (BM) biopsy should be considered, regardless of the basal serum tryptase concentration. In adults without skin lesions who suffer from mediator‐related or other typical symptoms, the basal tryptase level is an important parameter. In those with a slightly increased tryptase level, additional investigations, including a sensitive KIT mutation analysis of blood leucocytes or measurement of urinary histamine metabolites, may be helpful. In adult patients in whom (i) KIT D816V is detected and/or (ii) the basal serum tryptase level is clearly increased (>25–30 ng/ml) and/or (iii) other clinical or laboratory features suggest the presence of ‘occult’ mastocytosis or another haematologic neoplasm, a BM investigation is recommended. In the absence of KIT D816V and other signs or symptoms of mastocytosis or another haematopoietic disease, no BM investigation is required, but the clinical course and tryptase levels are monitored in the follow‐up. In paediatric patients, a BM investigation is usually not required, even if the tryptase level is increased. Although validation is required, it can be expected that the algorithm proposed herein will facilitate the management of patients with suspected mastocytosis and help avoid unnecessary referrals and investigations.

KIT mutation analysis in mast cell neoplasms: recommendations of the European Competence Network on Mastocytosis.

Although acquired mutations in KIT are commonly detected in various categories of mastocytosis, the methodologies applied to detect and quantify the mutant type and allele burden in various cells and tissues are poorly defined. We here propose a consensus on methodologies used to detect KIT mutations in patients with mastocytosis at diagnosis and during follow-up with sufficient precision and sensitivity in daily practice. In addition, we provide recommendations for sampling and storage of diagnostic material as well as a robust diagnostic algorithm. Using highly sensitive assays, KIT D816V can be detected in peripheral blood leukocytes from most patients with systemic mastocytosis (SM) that is a major step forward in screening and SM diagnosis. In addition, the KIT D816V allele burden can be followed quantitatively during the natural course or during therapy. Our recommendations should greatly facilitate diagnostic and follow-up investigations in SM in daily practice as well as in clinical trials. In addition, the new tools and algorithms proposed should lead to a more effective screen, early diagnosis of SM and help to avoid unnecessary referrals.

Cutaneous manifestations in patients with mastocytosis: Consensus report of the European Competence Network on Mastocytosis; the American Academy of Allergy, Asthma & Immunology; and the European Academy of Allergology and Clinical Immunology.

Cutaneous lesions in patients with mastocytosis are highly heterogeneous and encompass localized and disseminated forms. Although a classification and criteria for cutaneous mastocytosis (CM) have been proposed, there remains a need to better define subforms of cutaneous manifestations in patients with mastocytosis. To address this unmet need, an international task force involving experts from different organizations (including the European Competence Network on Mastocytosis; the American Academy of Allergy, Asthma & Immunology; and the European Academy of Allergology and Clinical Immunology) met several times between 2010 and 2014 to discuss the classification and criteria for diagnosis of cutaneous manifestations in patients with mastocytosis. This article provides the major outcomes of these meetings and a proposal for a revised definition and criteria. In particular, we recommend that the typical maculopapular cutaneous lesions (urticaria pigmentosa) should be subdivided into 2 variants, namely a monomorphic variant with small maculopapular lesions, which is typically seen in adult patients, and a polymorphic variant with larger lesions of variable size and shape, which is typically seen in pediatric patients. Clinical observations suggest that the monomorphic variant, if it develops in children, often persists into adulthood, whereas the polymorphic variant may resolve around puberty. This delineation might have important prognostic implications, and its implementation in diagnostic algorithms and future mastocytosis classifications is recommended. Refinements are also suggested for the diagnostic criteria of CM, removal of telangiectasia macularis eruptiva perstans from the current classification of CM, and removal of the adjunct solitary from the term solitary mastocytoma.