Journal of Immunology Research / Prophylactic mRNA vaccination against allergy confers long-term memory responses and persistent protection in mice

Recently, mRNA vaccines have been introduced as a safety-optimized alternative to plasmid DNA-based vaccines for protection against allergy. However, it remained unclear whether the short persistence of this vaccine type would limit memory responses and whether the protective immune response type would be maintained during recurrent exposure to allergen. We tested the duration of protective memory responses in mice vaccinated with mRNA encoding the grass pollen allergen Phl p 5 by challenging them with recombinant allergen, 3.5, 6, and 9 months after vaccination. In a second experiment, vaccinated mice were repeatedly challenged monthly with aerosolized allergen over a period of 7 months. Antibody and cytokine responses as well as lung inflammation and airway hyperresponsiveness were assessed. mRNA vaccination induced robust TH1 memory responses for at least 9 months. Vaccination efficiently suppressed TH2 cytokines, IgE responses, and lung eosinophilia. Protection was maintained after repeated exposure to aerosolized allergen and no TH1 associated pathology was observed. Lung function remained improved compared to nonvaccinated controls. Our data clearly indicate that mRNA vaccination against Phl p 5 induces robust, long-lived memory responses, which can be recalled by allergen exposure without side effects. mRNA vaccines fulfill the requirements for safe prophylactic vaccination without the need for booster immunizations.(VLID)179427

Th17 micro-milieu regulates NLRP1-dependent caspase-5 activity in skin autoinflammation.

IL-1β is a potent player in cutaneous inflammation and central for the development of a Th17 micro-milieu in autoinflammatory diseases including psoriasis. Its production is controlled at the transcriptional level and by subsequent posttranslational processing via inflammatory caspases. In this study, we detected inflammatory caspase-5 active in epidermal keratinocytes and in psoriatic skin lesions. Further, interferon-γ and interleukin-17A synergistically induced caspase-5 expression in cultured keratinocytes, which was dependent on the antimicrobial peptide psoriasin (S100A7). However, diseases-relevant triggers for caspase-5 activity and IL-1β production remain unknown. Recently, extranuclear DNA has been identified as danger-signals abundant in the psoriatic epidermis. Here, we could demonstrate that cytosolic double-stranded (ds) DNA transfected into keratinocytes triggered the activation of caspase-5 and the release of IL-1β. Further, interleukin-17A promoted caspase-5 function via facilitation of the NLRP1-inflammasome. Anti-inflammatory vitamin D interfered with the IL-1β release and suppressed caspase-5 in keratinocytes and in psoriatic skin lesions. Our data link the disease-intrinsic danger signals psoriasin (S100A7) and dsDNA for NLPR1-dependent caspase-5 activity in psoriasis providing potential therapeutic targets in Th17-mediated skin autoinflammation

Combined use of expression and CGH arrays pinpoints novel candidate genes in Ewing sarcoma family of tumors

<p>Abstract</p> <p>Background</p> <p>Ewing sarcoma family of tumors (ESFT), characterized by t(11;22)(q24;q12), is one of the most common tumors of bone in children and young adults. In addition to <it>EWS/FLI1 </it>gene fusion, copy number changes are known to be significant for the underlying neoplastic development of ESFT and for patient outcome. Our genome-wide high-resolution analysis aspired to pinpoint genomic regions of highest interest and possible target genes in these areas.</p> <p>Methods</p> <p>Array comparative genomic hybridization (CGH) and expression arrays were used to screen for copy number alterations and expression changes in ESFT patient samples. A total of 31 ESFT samples were analyzed by aCGH and in 16 patients DNA and RNA level data, created by expression arrays, was integrated. Time of the follow-up of these patients was 5–192 months. Clinical outcome was statistically evaluated by Kaplan-Meier/Logrank methods and RT-PCR was applied on 42 patient samples to study the gene of the highest interest.</p> <p>Results</p> <p>Copy number changes were detected in 87% of the cases. The most recurrent copy number changes were gains at 1q, 2, 8, and 12, and losses at 9p and 16q. Cumulative event free survival (ESFT) and overall survival (OS) were significantly better (P < 0.05) for primary tumors with three or less copy number changes than for tumors with higher number of copy number aberrations. In three samples copy number imbalances were detected in chromosomes 11 and 22 affecting the <it>FLI1 </it>and <it>EWSR1 </it>loci, suggesting that an unbalanced t(11;22) and subsequent duplication of the derivative chromosome harboring fusion gene is a common event in ESFT. Further, amplifications on chromosomes 20 and 22 seen in one patient sample suggest a novel translocation type between <it>EWSR1 </it>and an unidentified fusion partner at 20q. In total 20 novel ESFT associated putative oncogenes and tumor suppressor genes were found in the integration analysis of array CGH and expression data. Quantitative RT-PCR to study the expression levels of the most interesting gene, <it>HDGF</it>, confirmed that its expression was higher than in control samples. However, no association between <it>HDGF </it>expression and patient survival was observed.</p> <p>Conclusion</p> <p>We conclude that array CGH and integration analysis proved to be effective methods to identify chromosome regions and novel target genes involved in the tumorigenesis of ESFT.</p

Genome-wide association study identifies the GLDC/IL33 locus associated with survival of osteosarcoma patients

Survival rates for osteosarcoma, the most common primary bone cancer, have changed little over the past three decades and are particularly low for patients with metastatic disease. We conducted a multi‐institutional genome‐wide association study (GWAS) to identify germline genetic variants associated with overall survival in 632 patients with osteosarcoma, including 523 patients of European ancestry and 109 from Brazil. We conducted a time‐to‐event analysis and estimated hazard ratios (HR) and 95% confidence intervals (CI) using Cox proportional hazards models, with and without adjustment for metastatic disease. The results were combined across the European and Brazilian case sets using a random‐effects meta‐analysis. The strongest association after meta‐analysis was for rs3765555 at 9p24.1, which was inversely associated with overall survival (HR = 1.76; 95% CI 1.41–2.18, p = 4.84 × 10−7). After imputation across this region, the combined analysis identified two SNPs that reached genome‐wide significance. The strongest single association was with rs55933544 (HR = 1.9; 95% CI 1.5–2.4; p = 1.3 × 10−8), which localizes to the GLDC gene, adjacent to the IL33 gene and was consistent across both the European and Brazilian case sets. Using publicly available data, the risk allele was associated with lower expression of IL33 and low expression of IL33 was associated with poor survival in an independent set of patients with osteosarcoma. In conclusion, we have identified the GLDC/IL33 locus on chromosome 9p24.1 as associated with overall survival in patients with osteosarcoma. Further studies are needed to confirm this association and shed light on the biological underpinnings of this susceptibility locus

Genome-wide Association Study Identifies Two Susceptibility Loci for Osteosarcoma

Osteosarcoma is the most common primary bone malignancy of adolescents and young adults. In order to better understand the genetic etiology of osteosarcoma, we performed a multi-stage genome-wide association study (GWAS) consisting of 941 cases and 3,291 cancer-free adult controls of European ancestry. Two loci achieved genome-wide significance: rs1906953 at 6p21.3, in the glutamate receptor metabotropic 4 [GRM4] gene (P = 8.1 ×10-9), and rs7591996 and rs10208273 in a gene desert on 2p25.2 (P = 1.0 ×10-8 and 2.9 ×10-7). These two susceptibility loci warrant further exploration to uncover the biological mechanisms underlying susceptibility to osteosarcoma

Secretion of MCP-1 and other paracrine factors in a novel tumor-bone coculture model

BackgroundThe bone-tumor microenvironment encompasses unique interactions between the normal cells of the bone and marrow cavity and the malignant cells from a primary or metastasized cancer. A multitude of paracrine factors within this microenvironment such as the growth factor, TGF-beta, and the chemokine, MCP-1, are secreted by many of these cell types. These factors can act in concert to modulate normal and malignant cell proliferation, malignant cell migration and invasion and, often, mediate bone cancer pain. Although many valuable in vitro and in vivo models exist, identifying the relevant paracrine factors and deciphering their interactions is still a challenge. The aim of our study is to test an ex vivo coculture model that will allow monitoring of the expression, release and regulation of paracrine factors during interactions of an intact femur explant and tumor cells.MethodsIntact or marrow-depleted neonatal mouse femurs and select murine and human sarcoma or carcinoma cell lines were incubated singly or in coculture in specialized well plates. Viability of the bone and cells was determined by immunohistochemical stains, microscopy and marrow cytopreps. Secretion and mRNA expression of paracrine factors was quantitated by ELISA and real-time RT-PCR.ResultsCompartments of the bone were optimally viable for up to 48 h in culture and tumor cells for up to 4 days. Bone was the major contributor of TGF-beta and MMP2 whereas both bone and sarcoma cells secreted the chemokine MCP-1 in cocultures. Synergistic interaction between the femur and sarcoma resulted in enhanced MCP-1 secretion and expression in cocultures and was dependent on the presence of the hematopoietic component of the bone as well as other bone cells. In contrast, coculturing with breast carcinoma cells resulted in reduction of TGF-beta and MCP-1 secretion from the bone.ConclusionThese studies illustrate the feasibility of this model to examine paracrine interactions between intact bone and tumor cells. Further study of unique regulation of MCP-1 secretion and signaling between these cell types in different types of cancer will be possible using this simulated microenvironment

Genome-wide association study identifies two susceptibility loci for osteosarcoma

Osteosarcoma is the most common primary bone malignancy of adolescents and young adults. To better understand the genetic etiology of osteosarcoma, we performed a multistage genome-wide association study consisting of 941 individuals with osteosarcoma (cases) and 3,291 cancer-free adult controls of European ancestry. Two loci achieved genome-wide significance: a locus in the GRM4 gene at 6p21.3 (encoding glutamate receptor metabotropic 4; rs1906953; P = 8.1 × 10⁻⁹) and a locus in the gene desert at 2p25.2 (rs7591996 and rs10208273; P = 1.0 × 10⁻⁸ and 2.9 × 10⁻⁷, respectively). These two loci warrant further exploration to uncover the biological mechanisms underlying susceptibility to osteosarcoma