F2702 bulk RNA-seq data for manuscript "Therapeutic vaccine containing a novel liposomal adjuvant shifts pathogenic Th17 cells into a Tr1-like phenotype"
Th17 cells can transdifferentiate into other T cell subsets in the context of infections or inflammatory conditions. In many autoimmune diseases, autoantigen-specific Th17 cells play a pivotal role in disease pathogenesis, however, there have been no attempts to target Th17 cell plasticity using vaccines. We aimed to change the phenotype of existing Th17 cells by a protein-in-adjuvant approach and found that antigen formulated in all-trans retinoic acid (ATRA)-containing cationic liposomes (CAF16) effectively inhibited existing Th17 responses. Strikingly, transcriptomic analysis of sorted Th17 cells from IL-17 fate reporter mice revealed a shift of antigen-specific Th17 cells to exTh17 cells, expressing functional markers associated with T cell regulation and tolerance. In addition, vaccination with myelin-specific (MOG) antigen in CAF16 reduced Th17 responses and alleviated disease in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS. This highlights CAF16 as a novel delivery system for ATRA with potential for changing the phenotype of existing Th17 cells in the context of immune mediated diseases. Overall design: Cells from the spleen of IL-17A fate reporter mice were processed and CD4+ T cells were isolated using the EasySep mouse CD4+ T cell isolation kit (STEMCELL Technologies Inc., Canada) according to the manufacturer's recommendation. YFP+ cells were sorted from pools of three mice vaccinated either with CAF01:H56 only or CAF01:H56 followed by CAF16:H56 using a BD FACSMelody™ Cell Sorter (BD BioSciences). Cells were sorted into 100 µl RNase free 1x PBS (Thermo Scientific) with 1 U/µl RNAse inhibitor (Thermo Scientific) and stored at -80 oC before RNA bulk sequencing (performed by CD Genomics (NY, USA)). RNA was extracted from the cell samples using RNeasy Micro Kit (Qiagen). RNA sample quality was assessed by High Sensitivity RNA Tapestation (Agilent Technologies Inc., California, USA) and quantified by Qubit 2.0 RNA HS assay (ThermoFisher, Massachusetts, USA). The SMART-Seq v4 ultra-low input kit was used to synthesize cDNA. Library construction was performed based on the manufacturer's recommendation for Nextera XT kit. The final library quantity was measured by KAPA SYBR® FAST qPCR and library quality was evaluated by TapeStation D1000 ScreenTape (Agilent Technologies, CA, USA). Illumina® 8-nt dual-indices were used. Equimolar pooling of libraries was performed based on QC values and sequenced on an Illumina® NovaSeq S4 (Illumina, California, USA) with a read length configuration of 150 PE for 40 M PE reads per sample (20M in each direction).