Lydia Greenlees

Inhibition of Myogenic MicroRNAs 1, 133, and 206 by Inflammatory Cytokines Links Inflammation and Muscle Degeneration in Adult Inflammatory Myopathies

The molecular basis of inflammatory myopathies such as dermatomyositis (DM), polymyositis, and inclusion body myositis, which share the characteristics of chronic muscle inflammation and skeletal muscle wasting, are poorly understood. As such, effective targeted treatments for these diseases are lacking, resulting in critical unmet medical needs for these devastating diseases. The purpose of this study was to identify possible new targets for drug development by exploring the mechanism by which inflammation may play a role in the pathology of the inflammatory myopathies.

MicroRNA‐206 induces G1 arrest in melanoma by inhibition of CDK4 and Cyclin D

Expression profiling of microRNAs in melanoma lesional skin biopsies compared with normal donor skin biopsies, as well as melanoma cell lines compared with normal melanocytes, revealed that hsa‐miR‐206 was down‐regulated in melanoma (−75.4‐fold, P = 1.7 × 10−4). MiR‐206 has been implicated in a large number of cancers, including breast, lung, colorectal, ovarian, and prostate cancers; however, its role in tumor development remains largely unknown, its biologic function is poorly characterized, and its targets affecting cancer cells are largely unknown. MiR‐206 reduced growth and migration/invasion of multiple melanoma cell lines. Bioinformatics identified cell cycle genes CDK2, CDK4, Cyclin C, and Cyclin D1 as strong candidate targets. Western blots and 3′UTR reporter gene assays revealed that miR‐206 inhibited translation of CDK4, Cyclin D1, and Cyclin C. Additionally, hsa‐miR‐206 transfection induced G1 arrest in multiple melanoma cell lines. These observations support hsa‐miR‐206 as a tumor suppressor in melanoma and identify Cyclin C, Cyclin D1, and CDK4 as miR‐206 targets.

Genomic signatures characterize leukocyte infiltration in myositis muscles

BackgroundLeukocyte infiltration plays an important role in the pathogenesis and progression of myositis, and is highly associated with disease severity. Currently, there is a lack of: efficacious therapies for myositis; understanding of the molecular features important for disease pathogenesis; and potential molecular biomarkers for characterizing inflammatory myopathies to aid in clinical development.MethodsIn this study, we developed a simple model and predicted that 1) leukocyte-specific transcripts (including both protein-coding transcripts and microRNAs) should be coherently overexpressed in myositis muscle and 2) the level of over-expression of these transcripts should be correlated with leukocyte infiltration. We applied this model to assess immune cell infiltration in myositis by examining mRNA and microRNA (miRNA) expression profiles in muscle biopsies from 31 myositis patients and 5 normal controls.ResultsSeveral gene signatures, including a leukocyte index, type 1 interferon (IFN), MHC class I, and immunoglobulin signature, were developed to characterize myositis patients at the molecular level. The leukocyte index, consisting of genes predominantly associated with immune function, displayed strong concordance with pathological assessment of immune cell infiltration. This leukocyte index was subsequently utilized to differentiate transcriptional changes due to leukocyte infiltration from other alterations in myositis muscle. Results from this differentiation revealed biologically relevant differences in the relationship between the type 1 IFN pathway, miR-146a, and leukocyte infiltration within various myositis subtypes.ConclusionsResults indicate that a likely interaction between miR-146a expression and the type 1 IFN pathway is confounded by the level of leukocyte infiltration into muscle tissue. Although the role of miR-146a in myositis remains uncertain, our results highlight the potential benefit of deconvoluting the source of transcriptional changes in myositis muscle or other heterogeneous tissue samples. Taken together, the leukocyte index and other gene signatures developed in this study may be potential molecular biomarkers to help to further characterize inflammatory myopathies and aid in clinical development. These hypotheses need to be confirmed in separate and sufficiently powered clinical trials.

Identification of unique proteomic signatures in allergic and non-allergic skin disease

Atopic dermatitis (AD), psoriasis (PS), and contact dermatitis (CD) are common skin diseases, characterized by barrier disruption and systemic inflammation, with unique epidermal signatures and common inflammatory pathways identified by transcriptomic profiling. This study profiled proteomic signatures in serum from subjects with AD, PS, and CD compared with healthy controls (HC).

Development of a new ARCHITECT automated periostin immunoassay

BACKGROUND Periostin is being investigated as a potential biomarker for T-helper-2 (Th2)-driven asthma or eosinophilic inflammation and may help to identify patients more likely to benefit from interleukin-13-targeted treatments. We report the development and analytic performance of the investigational use only ARCHITECT Periostin Immunoassay, a new automated assay developed to detect serum periostin concentrations. METHODS We assessed assay performance in terms of precision, sensitivity, linearity, interference from classical immunoassay interferents and representatives of common asthma medications, specimen handling, and isoform reactivity. The assay was also used to assess the biological variability of serum periostin concentrations in samples from healthy volunteers and from subjects with uncontrolled asthma (the intended use population). RESULTS The percentage CVs for 5-day total precision, assessed using two instruments, was <6% across 2 controls and one serum-based panel. Limit of quantitation was 4ng/mL (dilution adjusted concentration), suiting the needs for this application. Dilution analysis yielded linear results and no endogenous sample or drug interferences were observed. All known periostin isoforms expressed in the mature human lung were detected by the assay. CONCLUSION Our studies provide support that the ARCHITECT Periostin Immunoassay is a reliable and robust test for measuring serum periostin concentrations.

Development and analytical performance of a new ARCHITECT automated dipeptidyl peptidase-4 immunoassay

Background Dipeptidyl peptidase-4 (DPP-4) may be a suitable biomarker to identify people with severe asthma who have greater activation of the interleukin-13 (IL-13) pathway and may therefore benefit from IL-13-targeted treatments. We report the analytical performance of an Investigational Use Only immunoassay and provide data on the biological range of DPP-4 concentrations. Methods We assessed assay performance, utilising analyses of precision, linearity and sensitivity; interference from common endogenous assay interferents, and from asthma and anti-diabetic medications, were also assessed. The assay was used to measure the range of serum DPP-4 concentrations in healthy volunteers and subjects with diabetes and severe, uncontrolled asthma. Results The total precision of DPP-4 concentration measurement (determined using percentage coefficient of variation) was ≤5% over 20 days. Dilution analysis yielded linear results from 30 to 1305 ng/mL; the limit of quantitation was 19.2 ng/mL. No notable endogenous or drug interferences were observed at the expected therapeutic concentration. Median DPP-4 concentrations in healthy volunteers and subjects with asthma or Type 1 diabetes were assessed, with concentrations remaining similar in subjects with diabetes and asthma across different demographics. Conclusion These analyses indicate that the ARCHITECT DPP-4 Immunoassay is a reliable and robust method for measuring serum DPP-4 concentration.

SAT0004 Genomic signatures characterize leukocyte infiltration in myositis muscles

Background Leukocyte infiltration plays an important role in the pathogenesis and progression of myositis, and is highly associated with disease severity. Objectives We employed a genomics approach to assess the inflammatory cell infiltration in myositis. Methods Muscle biopsies from 31 myositis patients and 5 normal healthy donors were profiled by microarray in parallel with microRNA (miRNA) expression analyses. Results Several gene signatures, such as the leukocyte index, type 1 interferon (IFN), MHC-1 and immunoglobulin signatures, were developed to characterize myositis patients at the molecular level. The leukocyte index was comprised of genes predominantly associated with the immune function and displayed a strong concordance with the pathological assessment of immune cell infiltration. This leukocyte index was subsequently utilized to evaluate expression changes of transcripts due to leukocyte infiltration in myositis muscle biopsies. The ability to distinguish different sources of altered gene expression in heterogeneous tissues increased our understanding of the complex interactions crucial to the pathogenesis of myositis. Such approach can be applied to various other studies and allowed more accurate interpretation of the underlying biology with gene expression data from heterogenous tissue samples. One application of the leukocyte index comparing miRNA and mRNA expression profiles revealed a complex interaction between miR-146a expression and the regulation of the type 1 IFN pathway in dermatomyositis. Conclusions Collectively, the distinct miRNA and mRNA signatures identified in this study may contribute to the development of new therapeutic targets and provide utility as molecular biomarkers for characterizing inflammatory myopathies. Disclosure of Interest W. Zhu Shareholder of: Astra Zeneca, Employee of: MedImmune, K. Streicher Shareholder of: Astra Zeneca, Employee of: MedImmune, N. Shen Consultant for: MedImmune, B. Higgs Shareholder of: Astra Zeneca, Employee of: MedImmune, C. Morehouse Shareholder of: Astra Zeneca, Employee of: MedImmune, L. Greenlees Shareholder of: Astra Zeneca, Employee of: MedImmune, K. Ranade Shareholder of: Astra Zeneca, Employee of: MedImmune, L. Richman Shareholder of: Astra Zeneca, Employee of: MedImmune, D. Fiorentino: None Declared, B. Jallal Shareholder of: Astra Zeneca, Employee of: MedImmune, S. Greenberg Consultant for: MedImmune, Y. Yao Shareholder of: Astra Zeneca, Employee of: MedImmune