RNA ImmunoGenic Assay: A Method to Detect Immunogenicity of in vitro Transcribed mRNA in Human Whole Blood
AKM Haque,
Petra Weinmann,
Sumit Biswas,
Rupert Handgretinger,
Markus Mezger,
Michael Kormann,
Justin Antony
Affiliations
AKM Haque
Department of Pediatrics I, Pediatric Infectiology and Immunology, Translational Genomics and Gene Therapy in Pediatrics, University of Tuebingen, Tuebingen, Germany
Petra Weinmann
Department of Pediatrics I, Pediatric Infectiology and Immunology, Translational Genomics and Gene Therapy in Pediatrics, University of Tuebingen, Tuebingen, Germany
Sumit Biswas
Department of Pediatrics I, Pediatric Infectiology and Immunology, Translational Genomics and Gene Therapy in Pediatrics, University of Tuebingen, Tuebingen, Germany
Rupert Handgretinger
University Children's Clinic Department of Paediatrics I, Hematology and Oncology, University of Tuebingen, Tuebingen, Germany
Markus Mezger
University Children's Clinic Department of Paediatrics I, Hematology and Oncology, University of Tuebingen, Tuebingen, Germany
Michael Kormann
Department of Pediatrics I, Pediatric Infectiology and Immunology, Translational Genomics and Gene Therapy in Pediatrics, University of Tuebingen, Tuebingen, Germany
Justin Antony
Department of Pediatrics I, Pediatric Infectiology and Immunology, Translational Genomics and Gene Therapy in Pediatrics, University of Tuebingen, Tuebingen, GermanyUniversity Children's Clinic Department of Paediatrics I, Hematology and Oncology, University of Tuebingen, Tuebingen, Germany
The mRNA therapeutics is a new class of medicine to treat many various diseases. However, in vitro transcribed (IVT) mRNA triggers immune responses due to recognition by human endosomal and cytoplasmic RNA sensors, but incorporation of modified nucleosides have been shown to reduce such responses. Therefore, an assay signifying important aspects of the human immune system is still required. Here, we present a simple ex vivo method called ‘RNA ImmunoGenic Assay’ to measure immunogenicity of IVT-mRNAs in human whole blood. Chemically modified and unmodified mRNA are complexed with a transfection reagent (TransIT), and co-incubated in human whole blood. Specific cytokines are measured (TNF-α, INF-α, INF-γ, IL-6 and IL-12p70) using ELISAs. The qPCR analysis is performed to reveal the activation of specific immune pathways. The RNA ImmunoGenic Assay provides a simple and fast method to detect donor specific - immune response against mRNA therapeutics.Graphic abstract:Schematic representation of RNA ImmunoGenic Assay
