TNF-α抑制剂治疗银屑病疗效相关基因的筛选与功能研究

Psoriasis is a common chronic inflammatory skin disease with a considerable impact on health-care. While the exact cause of psoriasis is unknown, genetic, immune, and environmental factors play important role in its development. In the last decade, biological drugs targeting tumor necrosis factor-α (TNF-α) have been highly successful in treating severe psoriasis. Nevertheless, the anti-TNFα therapy is very expensive, and there is marked heterogeneity in treatment response to these medications. Not all patients respond well to treatment, and some can develop severe adverse effects. Interindividual differences in several genes may explain this variation in response to treatment. Pharmacogenetics and pharmacogenomics can facilitate more personalized medicine and prevent the adverse effects associated with treatment. To investigate the mechanism of heterogeneous response with anti-TNF agents in Chinese psoriasis patients, we plan to perform a large-scale screening of differential treatment response-related genes using RNA-seq to assay the transcriptome of psoriasis patients with different response to anti-TNF therapy. Then we will utilize quantitative RT–PCR, Western blot, and immunohistochemistry to validate the expression of different genes. Then we will carry out genetic analyses using targeted sequencing and genotyping to find out genetic variants that are associated with the differential treatment responses among the patients. Furthermore, a series of functional studies will be conducted including luciferase assay, RNAi, mouse model and so on to validate the plausible associated key functional variants. This study will not only contribute to personalized treatment of psoriasis, but also help to gain biological insight into the biological mechanism of psoriasis.

银屑病是一种常见的慢性炎症性皮肤病，由遗传、免疫和环境等多因素共同作用所致。TNF-α抑制剂是一种治疗银屑病的有效生物制剂，但其疗效及不良反应存在明显的个体差异，且这种差异的机制不明确。因此，为阐明不同个体对药物不同反应的机制，我们以TNF-α抑制剂治疗的银屑病患者为研究对象，通过转录组测序筛选出TNF-α抑制剂治疗前后差异表达的关键基因；采用实时定量PCR、免疫组化、免疫印迹等方法在mRNA、蛋白质水平验证。同时，结合靶向测序与基因分型的方法搜寻与治疗反应相关的关键基因变异，并通过荧光素酶双报告基因、RNA干扰等方法进行系列功能研究，鉴定出与疗效密切相关的功能性基因变异，为中国人群银屑病患者个体化治疗提供理论指导。以筛选出的关键基因为靶标进一步构建转基因小鼠模型，探讨该基因在银屑病发病中的作用，为发现新的相关基因与治疗靶点奠定基础。

银屑病是一种常见的慢性炎症性皮肤病，由遗传、免疫和环境等多因素共同作用所致。TNF-α抑制剂是一种治疗银屑病的有效生物制剂，但其疗效及不良反应存在明显的个体差异，且这种差异的机制不明确。目前关于其治疗银屑病的药物遗传学研究相对比较局限，研究样本量较小，且存在地域和种族上的异质性。本研究收集了212例依那西普治疗中国汉族斑块型银屑病患者，包括治疗前后的外周血DNA、不同时间点RNA样本及长期随访临床资料，建立了一个较完整的生物制剂治疗银屑病的药物反应个体化多样性数据库，深入开展了系列的药物遗传学研究。.通过对212例接受依那西普治疗的银屑病患者进行全基因组SNP分型实验，利用全基因组关联分析研究这些基因的变异位点与TNF-α抑制剂治疗银屑病疗效的相关性（有效组vs无效组）。结果发现在治疗24周时，依那西普单药治疗组DCHS2基因rs117192735位点与治疗反应呈明显相关性(P=2.57E-07)；而在依那西普联合甲氨蝶呤组，在治疗24周时，发现rs7621900、rs78219077、rs71527245、rs72688609、rs13288811、rs139759817、rs149087125 7个位点与治疗反应呈明显相关性且达到全基因组显著水平。.通过对依那西普治疗的银屑病患者血样（治疗前vs治疗12周）的转录组表达谱深入挖掘，将差异基因表达和共表达分析的整合，优先考虑具有相关功能的DEGs，优化了差异表达基因筛选的生物信息学分析方法，成功发现19个差异表达基因可能与依那西普治疗反应相关。更进一步扩大样本量，在34例接受依那西普治疗的银屑病患者不同时间点（治疗前vs治疗12周、24周）进行转录组差异表达研究，以期鉴定出与依那西普治疗银屑病疗效相关的更多的功能性关键基因。.这是首次在中国汉族银屑病患者中展开较大规模的TNF-α抑制剂药物遗传学研究，研究结果对阐明疗效个体差异的遗传机制具有重要意义，为中国人群银屑病患者个体化治疗提供理论基础。项目实施期间，在Sci Rep、BMC Syst Biol、Exp Dermatol、Indian J Dermatol Venereol Leprol发表4篇SCI论文，1篇中文核心期刊论文；协助培养博士生、硕士生各1名。