Study On The Immunophenotype,Immune Target Screening And Targeted Drug Efficacy Of Vogt-Koyanagi-Harada Disease

Purpose:To interpret the immunophenotype of Vogt-Koyanagi-Harada disease(VKH)from the single-cell transcription level,screen potential immunotherapeutic targets and verify the efficacy of targeted therapeutic drugs,and explore the immunotargeted therapeutic strategy of high efficiency and low toxicity of VKH.Methods: By using single cell sequencing technology,we compared the differences in the transcription of inflammation-related genes in the peripheral blood mononuclear cells of patients with VKH and healthy controls.Through differential expression gene analysis and inflammatory pathway enrichment analysis,we described the differential expression gene heat map and inflammatory enrichment pathway of patients with VKH.We screened the potential immunotherapeutic targets of VKH and preliminarily verified its efficacy.We further evaluated the efficacy and safety of targeted drugs by carrying out prospective,open,longitudinal and single-center clinical studies.Patients who meet the VKH diagnostic criteria(early and late)were included in the group control study.Patients in group A were given targeted drugs combined with methylprednisolone,and patients in group B were given targeted drug monotherapy.The main outcome measures included the best corrected visual acuity(BCVA),the thickness of the subfoveal choroid(SFCT)and the height of subretinal fluid(SRF).The secondary outcome measures included intraocular pressure(IOP),anterior chamber inflammation,fluorescein angiography/indocyanine green angiography(FA/ICGA)score,25 visual function questionnaires(VFQ-25),36 short health questionnaires(SF-36)score Adverse drug reaction(ADR)and recurrence frequency of the drug.Results: Single cell sequencing showed that the proportion of T cells in patients with VKH was significantly increased,and the proportion of CD4+Naive T cells,Th1 cells and Th17 cells in T cell subgroups was significantly different from that in healthy controls.Multiple up-regulated DEGs were detected in CD4+Naive T cells,Th1 cells and Th17 cells that are closely related to classical inflammatory pathways such as immune cell migration and recruitment(CXCR4),immune cell activation(Jun and CD69),and IL-17 signal transduction(Junb).KEGG analysis showed that Th1 and Th2 cell differentiation pathway,IL-17 signal pathway,TNF signal pathway,JAK-STAT signal pathway,chemical factor signal pathway,Th17 cell differentiation related pathway,MAPK signal pathway,HIF-1 signal pathway,inflammatory bowel disease related pathway and rheumatoid arthritis related pathway were activated in different degrees in the cascade inflammatory network of VKH.JAK-STAT signal pathway was significantly activated in all three cell subsets as one of the potential immunotherapeutic targets for VKH.The initial clinical observation of small samples confirmed that the JAK-STAT signal pathway inhibitor baricitinib can effectively treat patients with VKH.The comparative analysis of single cell sequencing showed that the expression of immune-inflammatory-related genes and transcription factors(Fos,Jun,Junb,CXCR4,CD69,etc.)in CD4+Naive T cells,Th1 cells and Th17 cells of patients with VKH could be significantly down-regulated after treatment with baricitinib,and the expression of multiple previously activated inflammatory pathways could be inhibited.In the controlled clinical trial,76 eyes of 38 patients with VKH were included in the study(group A: 42 eyes;group B: 34 eyes),of which 44 eyes were early VKH disease and 32 eyes were late VKH disease.The average age was 43.10±12.32 years old,and the follow-up time was 6 to 18 months.Inflammation of all patients in the combination treatment group and the monotherapy group was effectively controlled.Compared with the baseline data,BCVA,anterior chamber inflammation,SFCT,SRF,FA/ICGA score,VFQ-25,SF-36 score table had statistically significant improvement.However,the patients in group A who received the combined treatment of oral baricitinib and methylprednisolone had more obvious improvement in all indicators than those in monotherapy group at the end of 2 months,while at the end of 6months,there was no statistical difference between the two groups.No serious ADR and disease recurrence were found in all patients during observation.Conclusion: Single cell sequencing showed that VKH has a complex immune phenotype and inflammatory activation network,other than a single inflammatory activation mode.JAK-STAT signal pathway plays an important role in the pathogenesis of VKH and is a potential therapeutic target of VKH.Clinical trials further confirmed that JAK inhibitor is a safe and effective target drug for VKH.JAK inhibitor combined with glucocorticoid at the initial stage of treatment,and JAK inhibitor maintained at the later stage has positive significance in considering the curative effect and reducing glucocorticoid dosage.