High-Resolution Expression Profiling of Peripheral Blood CD8+ Cells in Patients with Multiple Sclerosis Displays Fingolimod-Induced Immune Cell Redistribution

Abstract

Fingolimod, a sphingosine-1-phosphate (S1P) receptor modulator, is an oral drug approved for the treatment of active relapsing-remitting multiple sclerosis (RRMS). It selectively inhibits the egress of lymphocytes from lymph nodes. We studied the changes in the transcriptome of peripheral blood CD8 cells to unravel the effects at the molecular level during fingolimod therapy. We separated CD8 cells from the blood of RRMS patients before the first dose of fingolimod as well as 24 h and 3 months after the start of therapy. Changes in the expression of coding and non-coding genes were measured with high-density Affymetrix Human Transcriptome Array (HTA) 2.0 microarrays. Differentially expressed genes in response to therapy were identified by t test and fold change and analyzed for their functions and molecular interactions. No gene was expressed at significantly higher or lower levels 24 h after the first administration of fingolimod compared to baseline. However, after 3 months of therapy, 861 transcripts were found to be differentially expressed, including interleukin and chemokine receptors. Some of the genes are associated to the S1P pathway, such as the receptor S1P5 and the kinase MAPK1, which were significantly increased in expression. The fingolimod-induced transcriptome changes reflect a shift in the proportions of CD8 T cell subsets, with CCR7 effector memory T cells being relatively increased in frequency in the blood of fingolimod-treated patients. In consequence, CCR7 mRNA levels were reduced by >80 % and genes involved in T cell activation and lymphocyte cytotoxicity were increased in expression. Gene regulatory programs caused by downstream S1P signaling had only minor effects.

Transcriptome profiling of peripheral blood immune cell populations in multiple sclerosis patients before and during treatment with a sphingosine‐1‐phosphate receptor modulator

“…The huge transcriptome shifts in CD4+ cells and CD8+ cells clearly display the selective homing of T cell subpopulations to lymphoid tissues in response to sustained fingolimod therapy . On the contrary, the gene expression changes in CD19+ B cells were quite subtle considering the fact that their cell count was also remarkably reduced in the peripheral blood by almost 90% after 3 months (Table ).…”

“…Additional gene regulatory effects independent of S1P receptor binding, however, could not be evidenced in these cells in our study. The transcriptome shift in T cells and its impact on biological processes and S1P‐related pathways are presented in detail in our previous publications . In the following, we will focus on the results for the subset of B cells, which recently gained increasing attention in MS research …”

“…The strengths of our study are that we analyzed the gene expression signatures of distinct blood cell populations longitudinally in a well‐characterized cohort of RRMS patients . Moreover, we used 150 high‐resolution HTA 2.0 microarrays to obtain very accurate and comprehensive snapshots of the cellular transcriptomes.…”

“…Details about study design and analysis methods were published previously . Briefly, a large dataset comprising 150 high‐resolution microarrays was obtained for specific cell populations from blood samples of MS patients.…”

“…The probes were summarized in the Expression Console 1.3.1 software (Affymetrix) to 70523 gene‐level probe sets and 914585 exon‐level probe sets. Quality control and data normalization by the robust multiarray average algorithm including a log2 transformation were performed as described elsewhere . The processing of the microarray data was performed separately for each cell population.…”