Transcriptomic analysis of fumarate compounds identifies unique effects of isosorbide di-(methyl fumarate) on NRF2, NF-kappaB and IRF1 pathway genes. Transcriptomic analysis of fumarate compounds identifies unique effects of isosorbide di-(methyl fumarate) on NRF2, NF-kappaB and IRF1 pathway genes

Dimethyl fumarate (DMF) has emerged as first-line therapy for relapsing-remitting multiple scle-rosis (RRMS). This treatment, however, has been limited by adverse effects, which has prompted development of novel derivatives with improved tolerability. We compared effects of fumarates on gene expression in astrocytes. Our analysis included diroximel fumarate (DRF) and its metabolite monomethyl fumarate (MMF), along with a novel compound isosorbide di-(methyl fumarate) (IDMF). Treatment with IDMF resulted in the largest number of differentially expressed genes. The effects of DRF and MMF were consistent with NRF2 activation and NF-kappaB inhibition, respectively. IDMF responses, however, were concordant with both NRF2 activation and NF-kappaB inhibition, and we confirmed IDMF-mediated NF-kappaB inhibition using a reporter assay. IDMF also down-regulated IRF1 expression and IDMF-decreased gene promoters were enriched with IRF1 recognition se-quences. Genes altered by each fumarate overlapped significantly with those near loci from MS genetic association studies, but IDMF had the strongest overall effect on MS-associated genes. These results show that next-generation fumarates such as DRF and IDMF have effects differing from those of the MMF metabolite. Our findings support a model in which IDMF attenuates oxidative stress via NRF2 activation, with suppression of NF-kappaB and IRF1 contributing to mitigation of in-flammation and pyroptosis. Overall design: We treated human fetal astrocytes with sterile distilled water (CTL), MMF (2.5 µM), DRF (2.5 µM) or IDMF (2.5 µM) for 24 hours (n = 3 per treatment) and evaluated gene expression using Clariom S microarray assays.

富马酸二甲酯（Dimethyl fumarate, DMF）已成为复发缓解型多发性硬化（relapsing-remitting multiple sclerosis, RRMS）的一线治疗方案。然而，该疗法受限于不良反应，因此推动了耐受性更优的新型富马酸酯衍生物的研发。本研究对比了各类富马酸酯对星形胶质细胞基因表达的调控作用。本次分析涵盖的受试物包括富马酸地洛西美（diroximel fumarate, DRF）及其代谢物单甲基富马酸酯（monomethyl fumarate, MMF），以及新型化合物异山梨醇二(富马酸甲酯)（isosorbide di-(methyl fumarate), IDMF）。
经IDMF处理的细胞中，差异表达基因的数量最多。DRF与MMF的调控效应分别对应NRF2激活与NF-κB抑制；而IDMF的细胞应答则同时匹配NRF2激活与NF-κB抑制，我们还通过报告基因实验验证了IDMF介导的NF-κB抑制作用。此外，IDMF可下调干扰素调节因子1（IRF1）的表达，且受IDMF抑制的基因其启动子区域富集有IRF1识别序列。
各富马酸酯调控的基因均与多发性硬化遗传关联研究中定位的基因座邻近基因存在显著重叠，但IDMF对多发性硬化相关基因的整体调控效应最为显著。上述结果表明，新一代富马酸酯（如DRF与IDMF）的作用模式与MMF代谢物存在显著差异。本研究结果支持如下作用模型：IDMF通过激活NRF2减轻氧化应激，同时抑制NF-κB与IRF1，从而缓解炎症反应与细胞焦亡。
总体实验设计：本研究将人胎星形胶质细胞分别用无菌蒸馏水（对照组, CTL）、2.5 μM MMF、2.5 μM DRF或2.5 μM IDMF处理24小时（每组生物学重复数n=3），并通过Clariom S基因芯片检测评估基因表达水平。