A Novel Circulating MicroRNA for the Detection of Acute Myocarditis.

The diagnosis of acute myocarditis typically requires either endomyocardial biopsy (which is invasive) or cardiovascular magnetic resonance imaging (which is not universally available). Additional approaches to diagnosis are desirable. We sought to identify a novel microRNA for the diagnosis of acute myocarditis. To identify a microRNA specific for myocarditis, we performed microRNA microarray analyses and quantitative polymerase-chain-reaction (qPCR) assays in sorted CD4+ T cells and type 17 helper T (Th17) cells after inducing experimental autoimmune myocarditis or myocardial infarction in mice. We also performed qPCR in samples from coxsackievirus-induced myocarditis in mice. We then identified the human homologue for this microRNA and compared its expression in plasma obtained from patients with acute myocarditis with the expression in various controls. We confirmed that Th17 cells, which are characterized by the production of interleukin-17, are a characteristic feature of myocardial injury in the acute phase of myocarditis. The microRNA mmu-miR-721 was synthesized by Th17 cells and was present in the plasma of mice with acute autoimmune or viral myocarditis but not in those with acute myocardial infarction. The human homologue, designated hsa-miR-Chr8:96, was identified in four independent cohorts of patients with myocarditis. The area under the receiver-operating-characteristic curve for this novel microRNA for distinguishing patients with acute myocarditis from those with myocardial infarction was 0.927 (95% confidence interval, 0.879 to 0.975). The microRNA retained its diagnostic value in models after adjustment for age, sex, ejection fraction, and serum troponin level. After identifying a novel microRNA in mice and humans with myocarditis, we found that the human homologue (hsa-miR-Chr8:96) could be used to distinguish patients with myocarditis from those with myocardial infarction. (Funded by the Spanish Ministry of Science and Innovation and others.).Supported by a grant (PI19/00545, to Dr. Martín) from the Ministry of Science and Innovation through the Carlos III Institute of Health–Fondo de Investigación Sanitaria; by a grant from the Biomedical Research Networking Center on Cardiovascular Diseases (to Drs. Martín, Sánchez-Madrid, and Ibáñez); by grants (S2017/BMD-3671-INFLAMUNE-CM, to Drs. Martín and Sánchez-Madrid; and S2017/BMD-3867-RENIM-CM, to Dr. Ibáñez) from Comunidad de Madrid; by a grant (20152330 31, to Drs. Martín, Sánchez-Madrid, and Alfonso) from Fundació La Marató de TV3; by grants (ERC-2011-AdG 294340-GENTRIS, to Dr. Sánchez-Madrid; and ERC-2018-CoG 819775-MATRIX, to Dr. Ibáñez) from the European Research Council; by grants (SAF2017-82886R, to Dr. Sánchez-Madrid; RETOS2019-107332RB-I00, to Dr. Ibáñez; and SAF2017-90604-REDT-NurCaMeIn and RTI2018-095928-BI00, to Dr. Ricote) from the Ministry of Science and Innovation; by Fondo Europeo de Desarrollo Regional (FEDER); and by a 2016 Leonardo Grant for Researchers and Cultural Creators from the BBVA Foundation to Dr. Martín. The National Center for Cardiovascular Research (CNIC) is supported by the Carlos III Institute of Health, the Ministry of Science and Innovation, the Pro CNIC Foundation, and by a Severo Ochoa Center of Excellence grant (SEV-2015-0505). Mr. Blanco-Domínguez is supported by a grant (FPU16/02780) from the Formación de Profesorado Universitario program of the Spanish Ministry of Education, Culture, and Sports. Ms. Linillos-Pradillo is supported by a fellowship (PEJD-2016/BMD-2789) from Fondo de Garantía de Empleo Juvenil de Comunidad de Madrid. Dr. Relaño is supported by a grant (BES-2015-072625) from Contratos Predoctorales Severo Ochoa para la Formación de Doctores of the Ministry of Economy and Competitiveness. Dr. Alonso-Herranz is supported by a fellowship from La Caixa–CNIC. Dr. Caforio is supported by Budget Integrato per la Ricerca dei Dipartimenti BIRD-2019 from Università di Padova. Dr. Das is supported by grants (UG3 TR002878 and R35 HL150807) from the National Institutes of Health and the American Heart Association through its Strategically Focused Research Networks.S

Development and Optimization of Logic Gated Small Interfering RNAs for Operation Inside Mammalian Cells

In 1900, Paul Ehrlich put forth the concept of a pharmacologic “magic bullet”, a combination of a disease selective chemical agent conjugated to a non-selective toxin that could target drug activity to disease causing cells without harming healthy tissues in the body. For the past two decades, various researchers in DNA and RNA nanotechnology have proposed ideas for riboswitch regulated magic bullets in which the activities of oligonucleotide drugs are switched ON or OFF according to the presence or absence of specific biomarkers in tissues or cells. Although elegant in concept, practical implementation of these “logical therapeutics” for mammalian cells has proven elusive. We have now developed a conditionally activated small interfering RNA (Cond-siRNA) in which an RNA sensor regulates the RNAi activity of a conjugated siRNA post-transfection into mammalian cells. The sensor switches ON RNAi activity when it base-pairs to an RNA transcript from a designated “trigger gene”. In cells without trigger gene expression, the sensor keeps RNAi activity OFF. The identities of the trigger and target genes are encoded by two entirely independent, easily programmable RNA sequences, giving the Cond-siRNAs the ability to target a specific population of cells for the silencing of an arbitrary gene. We will present experimental data for Cond-siRNAs with different trigger and target gene combinations in different mammalian cell lines, explain the design principles and optimizations that enable easy programmability and correct functioning in mammalian cells, and demonstrate an early example of therapeutic application development in the treatment of cardiac hypertrophy. As small, chemically modified, unimolecular RNA complexes, Cond-siRNAs are compatible with existing methods of siRNA delivery and practical for development into clinically viable RNAi drugs. By making possible post-delivery targeting of RNAi activity to specific populations of disease driving cells, they could enable paradigm shifting development of RNAi drugs that can safely target pleiotropic genes whose dysregulated activities drive the progression of many chronic diseases lacking effective treatments today

A novel circulating microRNA for the detection of acute myocarditis

BACKGROUND The diagnosis of acute myocarditis typically requires either endomyocardial biopsy (which is invasive) or cardiovascular magnetic resonance imaging (which is not universally available). Additional approaches to diagnosis are desirable. We sought to identify a novel microRNA for the diagnosis of acute myocarditis. METHODS To identify a microRNA specific for myocarditis, we performed microRNA microarray analyses and quantitative polymerase-chain-reaction (qPCR) assays in sorted CD4+ T cells and type 17 helper T (Th17) cells after inducing experimental autoimmune myocarditis or myocardial infarction in mice. We also performed qPCR in samples from coxsackievirus-induced myocarditis in mice. We then identified the human homologue for this microRNA and compared its expression in plasma obtained from patients with acute myocarditis with the expression in various controls. RESULTS We confirmed that Th17 cells, which are characterized by the production of interleukin-17, are a characteristic feature of myocardial injury in the acute phase of myocarditis. The microRNA mmu-miR-721 was synthesized by Th17 cells and was present in the plasma of mice with acute autoimmune or viral myocarditis but not in those with acute myocardial infarction. The human homologue, designated hsa-miR-Chr8:96, was identified in four independent cohorts of patients with myocarditis. The area under the receiver-operating-characteristic curve for this novel microRNA for distinguishing patients with acute myocarditis from those with myocardial infarction was 0.927 (95% confidence interval, 0.879 to 0.975). The microRNA retained its diagnostic value in models after adjustment for age, sex, ejection fraction, and serum troponin level. CONCLUSIONS After identifying a novel microRNA in mice and humans with myocarditis, we found that the human homologue (hsa-miR-Chr8:96) could be used to distinguish patients with myocarditis from those with myocardial infarction