T Cell Responses to Dystrophin in a Natural History Study of Duchenne Muscular Dystrophy

Duchenne muscular dystrophy (DMD) is caused by the lack of dystrophin, but many patients have rare revertant fibres that express dystrophin. The skeletal muscle pathology of DMD patients includes immune cell infiltration and inflammatory cascades. There are several strategies to restore dystrophin in skeletal muscles of patients, including exon skipping and gene therapy. There is some evidence that dystrophin restoration leads to a reduction in immune cells, but dystrophin epitopes expressed in revertant fibres or following genome editing, cell therapy or microdystrophin delivery after AAV gene therapy may elicit T cell production in patients. This may affect the efficacy of the therapeutic intervention, and potentially lead to serious adverse events. To confirm and extend previous studies, we performed annual Enzyme- Linked Immunospot interferon-gamma assays on peripheral blood mononuclear cells from 77 paediatric boys with DMD recruited into a natural history study, 69 of whom (89.6%) were treated with corticosteroids. T cell responses to dystrophin were quantified using a total of 368 peptides spanning the entire dystrophin protein, organized into nine peptide pools. Peptide mapping pools were used to further localize the immune response in one positive patient. Six (7.8%) patients had a T cell-mediated immune response to dystrophin at at least one timepoint. All patients that had a positive result had been treated with corticosteroids, either prednisolone or prednisone. Our results show that ~8% of DMD individuals in our cohort have a pre-existing T cell-mediated immune response to dystrophin despite steroid treatment. Although these responses are relatively low-level, this information should be considered as a useful immunological baseline before undertaking clinical trials and future DMD studies. We further highlight the importance for a robust, reproducible standard operating procedure for collecting, storing and shipping samples from multiple centres to minimise the number of inconclusive data

Elucidation of pathways driving asthma pathogenesis: development of a systems-level analytic strategy.

Asthma is a genetically complex, chronic lung disease defined clinically as episodic airflow limitation and breathlessness that is at least partially reversible, either spontaneously or in response to therapy. Whereas asthma was rare in the late 1800s and early 1900s, the marked increase in its incidence and prevalence since the 1960s points to substantial gene × environment interactions occurring over a period of years, but these interactions are very poorly understood (1-6). It is widely believed that the majority of asthma begins during childhood and manifests first as intermittent wheeze. However, wheeze is also very common in infancy and only a subset of wheezy children progress to persistent asthma for reasons that are largely obscure. Here, we review the current literature regarding causal pathways leading to early asthma development and chronicity. Given the complex interactions of many risk factors over time eventually leading to apparently multiple asthma phenotypes, we suggest that deeply phenotyped cohort studies combined with sophisticated network models will be required to derive the next generation of biological and clinical insights in asthma pathogenesis

Blood-derived biomarkers correlate with clinical progression in Duchenne muscular dystrophy

Background: Duchenne Muscular Dystrophy is a severe, incurable disorder caused by mutations in the dystrophin gene. The disease is characterized by decreased muscle function, impaired muscle regeneration and increased inflammation. In a clinical context, muscle deterioration, is evaluated using physical tests and analysis of muscle biopsies, which fail to accurately monitor the disease progression. Objectives: This study aims to confirm and asses the value of blood protein biomarkers as disease progression markers using one of the largest longitudinal collection of samples. Methods: A total of 560 samples, both serum and plasma, collected at three clinical sites are analyzed using a suspension bead array platform to assess 118 proteins targeted by 250 antibodies in microliter amount of samples. Results: Nine proteins are confirmed as disease progression biomarkers in both plasma and serum. Abundance of these biomarkers decreases as the disease progresses but follows different trajectories. While carbonic anhydrase 3, microtubule associated protein 4 and collagen type I alpha 1 chain decline rather constantly over time, myosin light chain 3, electron transfer flavoprotein A, troponin T, malate dehydrogenase 2, lactate dehydrogenase B and nestin plateaus in early teens. Electron transfer flavoprotein A, correlates with the outcome of 6-minutes-walking-test whereas malate dehydrogenase 2 together with myosin light chain 3, carbonic anhydrase 3 and nestin correlate with respiratory capacity. Conclusions: Nine biomarkers have been identified that correlate with disease milestones, functional tests and respiratory capacity. Together these biomarkers recapitulate different stages of the disorder that, if validated can improve disease progression monitoring

Novel free-circulating and extracellular vesicle-derived miRNAs dysregulated in Duchenne muscular dystrophy

Item does not contain fulltextAim: To perform cross-sectional and longitudinal miRNA profiling in plasma from Duchenne muscular dystrophy (DMD) subjects and find non-invasive biomarkers in DMD. Subjects/materials & methods: Plasma was collected from 14 age and sex matched controls and 46 DMD subjects. Free-circulating and extracellular vesicle (EV)-derived miRNA expression was measured by RT-qPCR. Results: Free-circulating and EVs derived miR-29c-3p and miR-133a-3p are dysregulated in DMD subjects. Free-circulating and EV-derived miR-29c-3p are reduced in DMD subjects undergoing daily corticosteroid treatment. Free-circulating miR-1-3p and miR-122-5p are longitudinally upregulated in ambulant DMD subjects. Conclusion: We detected novel free-circulating and EV-derived dysregulated miRNAs in plasma from DMD subjects and characterized the longitudinal profile of free-circulating miRNA on plasma from DMD subjects

T Cell Responses to Dystrophin in a Natural History Study of Duchenne Muscular Dystrophy

Duchenne muscular dystrophy (DMD) is caused by the lack of dystrophin, but many patients have rare revertant fibres that express dystrophin. The skeletal muscle pathology of DMD patients includes immune cell infiltration and inflammatory cascades. There are several strategies to restore dystrophin in skeletal muscles of patients, including exon skipping and gene therapy. There is some evidence that dystrophin restoration leads to a reduction in immune cells, but dystrophin epitopes expressed in revertant fibres or following genome editing, cell therapy or microdystrophin delivery after AAV gene therapy may elicit T cell production in patients. This may affect the efficacy of the therapeutic intervention, and potentially lead to serious adverse events. To confirm and extend previous studies, we performed annual Enzyme- Linked Immunospot interferon-gamma assays on peripheral blood mononuclear cells from 77 paediatric boys with DMD recruited into a natural history study, 69 of whom (89.6%) were treated with corticosteroids. T cell responses to dystrophin were quantified using a total of 368 peptides spanning the entire dystrophin protein, organized into nine peptide pools. Peptide mapping pools were used to further localize the immune response in one positive patient. Six (7.8%) patients had a T cell-mediated immune response to dystrophin at at least one timepoint. All patients that had a positive result had been treated with corticosteroids, either prednisolone or prednisone. Our results show that ~8% of DMD individuals in our cohort have a pre-existing T cell-mediated immune response to dystrophin despite steroid treatment. Although these responses are relatively low-level, this information should be considered as a useful immunological baseline before undertaking clinical trials and future DMD studies. We further highlight the importance for a robust, reproducible standard operating procedure for collecting, storing and shipping samples from multiple centres to minimise the number of inconclusive data