Transcriptomic profile of multiple sclerosis patients treated with dimethyl fumarate and their age and sex-matched healthy controls
Dimethyl fumarate (DMF) is an immunomodulatory drug approved for the therapy of multiple sclerosis (MS). The identification of response biomarkers to DMF is a necessity in the clinical practice. With this aim, we studied the transcriptomic changes produced by DMF in peripheral blood mononuclear cells (PBMCs) and its association with clinical response. DMF induced a mild transcriptional effect, with only 328 differentially expressed genes (DEGs) after 12 months of treatment. The overall effect was a downregulation of pro-inflammatory genes, chemokines, and activators of the NF-kB pathway. At baseline, no DEGs were found between responders and non-responders. During DMF treatment a differential transcriptomic response was observed, with responders presenting a higher number of DEGs (902 genes) compared to non-responders (189 genes). Responder patients to DMF exhibit a distinguishable transcriptomic response compared to non-responders that should be further studied for the validation of biomarkers of treatment response to DMF. Overall design: We extracted blood from 10 multiple sclerosis patients (5 responders to DMF and 5 non-responders to DMF) immediately before starting treatment and at 12 months of therapy with DMF. Blood was also extracted from 10 healthy donors, which were matched for age and sex with the patients, at a single timepoint. Peripheral blood mononuclear cells (PBMCs) were isolated from the blood using Ficoll density gradient centrifugation. Total RNA from PBMCs was extracted using the Maxwell 16 LEV simply RNA Cells kit (Promega Biotech Ibérica) on the Maxwell 16 instrument, according to the manufacturer's instructions. The quality and quantity of the RNA was determined in Bioanalyzer 2100 (Agilent Technologies) and Qubit 3.0 (Thermo Fisher Scientific), respectively. Poly(A)+ mRNA fraction was isolated from total RNA and cDNA libraries were obtained following Illumina's recommendations. Briefly, poly(A)+ RNA was isolated on poly-T oligo-attached magnetic beads and chemically fragmented prior to reverse transcription and cDNA generation. The cDNA went through an end repair process, addition of a single “A” base to the 3' end and ligation to the adapters. Finally, the products were purified and enriched with PCR to create the indexed final double stranded DNA library. The quality and quantity of the libraries were analysed in tapeStation 4200, using the High Sensitivity assay (Agilent Technologies). The pool of libraries was sequenced by pair-end sequencing (150x2) in Novaseq 6000 (Illumina) at a sequencing depth of 40 million reads per sample.