BRAF-V600E expression is identified in hematopoietic progenitor and precursor myeloid dendritic cells in patients with high-risk LCH, and enforced expression of BRAF-V600E in CD11c+ cells recapitulates a high-risk LCH-like phenotype in mice.

Pre-eclampsia affects approximately 5% of pregnancies and remains a leading cause of maternal and neonatal mortality and morbidity in the United States and the world1,2. The clinical hallmarks of this maternal disorder include hypertension, proteinuria, endothelial dysfunction and placental defects. Advanced-stage clinical symptoms include cerebral hemorrhage, renal failure and the HELLP (hemolysis, elevated liver enzymes and low platelets) syndrome. An effective treatment of pre-eclampsia is unavailable owing to the poor understanding of the pathogenesis of the disease. Numerous recent studies3–5 have shown that women with pre-eclampsia possess autoantibodies, termed AT1-AAs, that bind and activate the angiotensin II receptor type 1a (AT1 receptor). We show here that key features of pre-eclampsia, including hypertension, proteinuria, glomerular endotheliosis (a classical renal lesion of pre-eclampsia), placental abnormalities and small fetus size appeared in pregnant mice after injection with either total IgG or affinity-purified AT1-AAs from women with pre-eclampsia. These features were prevented by co-injection with losartan, an AT1 receptor antagonist, or by an antibody neutralizing seven–amino-acid epitope peptide. Thus, our studies indicate that pre-eclampsia may be a pregnancy-induced autoimmune disease in which key features of the disease result from autoantibody-induced angiotensin receptor activation. This hypothesis has obvious implications regarding pre-eclampsia screening, diagnosis and therapy.

IMPORTANCE Whole-exome sequencing (WES) has the potential to reveal tumor and germline mutations of clinical relevance, but the diagnostic yield for pediatric patients with solid tumors is unknown. OBJECTIVE To characterize the diagnostic yield of combined tumor and germline WES for children with solid tumors. DESIGN Unselected children with newly diagnosed and previously untreated central nervous system (CNS) and non-CNS solid tumors were prospectively enrolled in the BASIC3 study at a large academic children’s hospital during a 23-month period from August 2012 through June 2014. Blood and tumor samples underwent WES in a certified clinical laboratory with genetic results categorized on the basis of perceived clinical relevance and entered in the electronic health record. MAIN OUTCOMES AND MEASURES Clinical categorization of somatic mutations; frequencies of deleterious germline mutations related to patient phenotype and incidental medically-actionable mutations. RESULTS Of the first 150 participants (80 boys and 70 girls, mean age, 7.4 years), tumor samples adequate for WES were available from 121 patients (81%). Somatic mutations of established clinical utility (category I) were reported in 4 (3%) of 121 patients, with mutations of potential utility (category II) detected in an additional 29 (24%) of 121 patients. CTNNB1 was the gene most frequently mutated, with recurrent mutations in KIT, TSC2, and MAPK pathway genes (BRAF, KRAS, and NRAS) also identified. Mutations in consensus cancer genes (category III) were found in an additional 24 (20%) of 121 tumors. Fewer than half of somatic mutations identified were in genes known to be recurrently mutated in the tumor type tested. Diagnostic germline findings related to patient phenotype were discovered in 15 (10%) of 150 cases: 13 pathogenic or likely pathogenic dominant mutations in adult and pediatric cancer susceptibility genes (including 2 each in TP53, VHL, and BRCA1), 1 recessive liver disorder with hepatocellular carcinoma (TJP2), and 1 renal diagnosis (CLCN5). Incidental findings were reported in 8 (5%) of 150 patients. Most patients harbored germline uncertain variants in cancer genes (98%), pharmacogenetic variants (89%), and recessive carrier mutations (85%). CONCLUSIONS AND RELEVANCE Tumor and germline WES revealed mutations in a broad spectrum of genes previously implicated in both adult and pediatric cancers. Combined reporting of tumor and germline WES identified diagnostic and/or potentially actionable findings in nearly 40% of newly diagnosed pediatric patients with solid tumors.

• Recurrent somatic mutations in MAP2K1 were identified in 33% of LCH lesions with wild-type BRAF. The mutant MAPK kinase 1 proteins activate ERK.• The ability of MAPK pathway inhibitors to suppress MAPK kinase and ERK phosphorylation in vitro was dependent on the specific LCH mutation.Langerhans cell histiocytosis (LCH) is a myeloproliferative disorder characterized by lesions composed of pathological CD207 1 dendritic cells with an inflammatory infiltrate.BRAFV600E remains the only recurrent mutation reported in LCH. In order to evaluate the spectrum of somatic mutations in LCH, whole exome sequencing was performed on matched LCH and normal tissue samples obtained from 41 patients. Lesions from other histiocytic disorders, juvenile xanthogranuloma, Erdheim-Chester disease, and RosaiDorfman disease were also evaluated. All of the lesions from histiocytic disorders were characterized by an extremely low overall rate of somatic mutations. Notably, 33% (7/21) of LCH cases with wild-type BRAF and none (0/20) with BRAFV600E harbored somatic mutations in MAP2K1 (6 in-frame deletions and 1 missense mutation) that induced extracellular signal-regulated kinase (ERK) phosphorylation in vitro. Single cases of somatic mutations of the mitogen-activated protein kinase (MAPK) pathway genes ARAF and ERBB3 were also detected. The ability of MAPK pathway inhibitors to suppress MAPK kinase and ERK phosphorylation in cell culture and primary tumor models was dependent on the specific LCH mutation. The findings of this study support a model in which ERK activation is a universal end point in LCH arising from pathological activation of upstream signaling proteins. (Blood. 2014;124(19):3007-3015)

No abstract

The expression of leukocyte and endothelial cell adhesion molecules (CAMs) is essential for the emigration of leukocytes during an inflammatory response. The importance of the inflammatory response in the development of atherosclerosis is indicated by the increased expression of adhesion molecules, proinflammatory cytokines, and growth factors in lesions and lesion-prone areas and by protection in mice deficient in various aspects of the inflammatory response. We have quantitated the effect of deficiency for intercellular adhesion molecule (ICAM)-1, P-selectin, or E-selectin on atherosclerotic lesion formation at 20 wk of age in apolipoprotein (apo) E−/− (deficient) mice fed a normal chow diet. All mice were apo E−/− and CAM+/+ or CAM−/− littermates, and no differences were found in body weight or cholesterol levels among the various genotypes during the study. ICAM-1−/− mice had significantly less lesion area than their ICAM-1+/+ littermates: 4.08 ± 0.70 mm2 for −/− males vs. 5.87 ± 0.66 mm2 for +/+ males, and 3.95 ± 0.65 mm2 for −/− females vs. 5.59 ± 1.131 mm2 for +/+ females, combined P < 0.0001. An even greater reduction in lesion area was observed in P-selectin−/− mice: 3.06 ± 1.04 mm2 for −/− males vs. 5.09 ± 1.22 mm2 for +/+ males, and 2.85 ± 1.26 mm2 for −/− females compared with 5.60 ± 1.19 mm2 for +/+ females, combined P < 0.001. The reduction in lesion area for the E-selectin null mice, although less than that seen for ICAM-1 or P-selectin, was still significant (4.54 ± 2.14 mm2 for −/− males vs. 5.92 ± 0.63 mm2 for +/+ males, and 4.38 ± 0.85 mm2 for −/− females compared with 5.94 ± 1.44 mm2 for +/+ females, combined P < 0.01). These results, coupled with the closely controlled genetics of this study, indicate that reductions in the expression of P-selectin, ICAM-1, or E-selectin provide direct protection from atherosclerotic lesion formation in this model.

SummaryDuring the initial phase of the inflammatory response, leukocytes marginate and roll along the endothelial surface, a process mediated largely by the selectins and their ligands. Mice with mutations in individual selectins show no spontaneous disease and have mild or negligible deficiencies of inflammatory responses. In contrast, we find that mice with null mutations in both endothelial selectins (P and E) develop a phenotype of leukocyte adhesion deficiency characterized by mucocutaneous infections, plasma cell proliferation, hypergammaglobulinemia, severe deficiencies of leukocyte rolling in cremaster venules with or without addition of TNF-cx, and an absence of neutrophil emigration at 4 h in response to intraperitoneal Streptococcus pneumoniae peritonitis. These mice provide strong evidence for the functional importance of selectins in vivo.L eukocyte and endothelial cell adhesion molecules play an important role in inflammatory and immune responses (1, 2). The initial steps in leukocyte emigration in response to inflammatory stimuli involve leukocyte rolling that is mediated primarily by interactions between selectins and selectin ligand molecules (3). P-selectin is expressed on endothelium and platelets, E-selectin on endothelium, and L-selectin on the majority of leukocytes. Selectins bind to carbohydrate portions of glycoproteins that serve as selectin ligands, many of which are mucin-like proteins (4-7). Leukocyte rolling is followed by firm attachment and emigration, processes largely dependent on the interaction of the [32 leukocyte integrins and immunoglobulin family members including ICAM-1 and related molecules (1,8). The leukocyte integrins are heterodimeric proteins and include LFA-1 (Cr , p150,95 (OLX~2) , and (~'d~2" The functional importance of these adhesion molecules is demonstrated by the occurrence of two human genetic disorders, leukocyte adhesion deficiency types I and II (LAD I/II) 1 (9). LAD I is caused by genetic deficiency of 1Abbreviations used in this paper: ES, embryonic stem; LAD, leukocyte adhesion deficiency.Drs. Kubo, Doerschuk, and Doyle's current address is

Langerhans-cell histiocytosis (LCH) is a rare disease characterized by heterogeneous lesions containing CD207+ Langerhans cells and lymphocytes that can arise in almost any tissue and cause significant morbidity and mortality. After decades of research, the cause of LCH remains speculative. A prevailing model suggests that LCH arises from malignant transformation and metastasis of epidermal Langerhans cells. In this study, CD207+ cells and CD3+ T cells were isolated from LCH lesions to determine cell-specific gene expression. Compared to control epidermal CD207+ cells, the LCH CD207+ cells yielded 2113 differentially-expressed genes (FDR<0.01). Surprisingly, expression of many genes previously associated with LCH, including cell-cycle regulators, pro-inflammatory cytokines and chemokines were not significantly different from control LCs in our study. However, several novel genes whose products activate and recruit T cells to sites of inflammation, including SPP1 (osteopontin), were highly over-expressed in LCH CD207+ cells. Furthermore, several genes associated with immature myeloid dendritic cells were over-expressed in LCH CD207+ cells. Compared to the peripheral CD3+ cells from LCH patients, the LCH lesion CD3+ cells yielded only 162 differentially-regulated genes (FDR<0.01), and the expression profile of the LCH lesion CD3+ cells was consistent with an activated regulatory T cell phenotype with increased expression of FOXP3, CTLA4 as well as SPP1. Results from this study support a model of LCH pathogenesis in which lesions do not arise from epidermal Langerhans cells, but from accumulation of bone-marrow derived immature myeloid dendritic cells that recruit activated lymphocytes.