| Background Psoriasis(Ps O)is a common chronic inflammatory skin disease with the participation of genetic and environmental factors and mediated by immunoreaction.About 2% of the global population is affected by this disease.Intense itching,visible skin changes,possible disability,increased risk of chronic diseases and recurrent course of disease have brought serious physical and mental injury to patients with psoriasis,that make a great burden of disease.In the World Health Assembly resolution,psoriasis has been recognized as a serious non-communicable disease with important public health impact by member states.Compared with mild psoriasis,patients with moderateto-severe psoriasis suffering the greater disease burden and their quality of life is more seriously affected.Besides,it is more difficult to achieve and maintain the remission of skin lesions in treatment.As the first routine systemic medicine for treating moderateto-severe psoriasis,methotrexate(MTX)has been used for more than 50 years.However,high quality clinical research about MTX in treating psoriasis is still relatively limited.Even in the strictly clinical trials,significant differences in drug responses were observed among different individuals.Genetic factors are the internal basis of these differences in individualized responses.With the completion of the Human Genome Project at the beginning of 21 th century and the advancement of high throughput genotyping and gene sequencing technology with low-cost,the pharmacogenomics,which studies the relationship between genetic polymorphisms and individual drug response,has begun to flourish.The analysis of genetic polymorphisms can partly explain the reasons for the various response in patients,and lay a theoretical foundation for the exploitation of genetic markers that can be used to predict drug effectiveness and adverse reactions.At present,some specific genetic markers have been found to be related to the response of MTX in the treatment of psoriasis.However,these studies have mostly aimed at identifying genes which directly related to drug distribution or action,and ignored other markers within the genome including susceptibility genes for psoriasis.From candidate gene research to the application of genome-wide research strategies,it is possible to comprehensively identify the underlying causes associated with individualized differences in drug responses,and to provide new insights into disease pathophysiology and molecular pharmacology.Object To provide real and high-quality clinical data on the effectiveness and safety of MTX in the treatment of psoriasis by performing a prospective observational study.To explore the genetic mechanism that can explain the difference response of MTX through candidate gene research and Genome-wide association study(GWAS)analysis.And to lay a theoretical foundation for the development of precision medicine in the future.Methods(1)We used a prospective,single-center and cross-section study to recruit patients who had received MTX for 12 weeks.Basic clinical characteristics and the change of various indicators such as PASI(psoriasis area index),indicators of blood routine test,liver function test etc.were recorded.These indicators were analyzed in all patients and further analyzed in subgroup according by patients with or without psoriasis arthritis;(2)Patients were divided into two groups(responser and nonresponser)by PASI.After using MTX,improvements in PASI more than 75% were defined as responser,in contrast,less than 50% were defined as non-responser.18 susceptibility SNPs for psoriasis were selected into our study.The genotype of 18 SNPs and the status of HLA-Cw*0602 in 90 patients were detected by using Agena Mass Array genotyping method and HLA-C direct sequencing typing method.ABI SNa Pshot genotyping platform were used to validate 2 significant SNPs.Multivariate analysis was performed to adjust for confounding factors such as age at onset,disease duration and body mass index(BMI)etc.;(3)GWAS method were used into our pharmacogenomics study after expanding sample size.In first filter stage,Illumina Omni Zhong Hua 8 Human Bead Chips V1.3 and V1.4 were used to screen out SNPs sites that might be related to MTX response.The SNPs which had strong signal in first stage were further verified by Mass Array genotyping method in independent samples.The susceptibility SNPs that related to MTX response were investigated by combining the data from the two stages.Bioinformatics analysis were used to find the susceptible genes and their corresponding biological pathways that might associated with SNPs signals.Results(1)After 12 weeks of medication,the average PASI score decreased by 62.2(32.6)%.The patients who reached PASI50,PASI75,and PASI90 were 166(70.6%),107(45.5%),and 46(19.6%);(2)MTX affects most of blood routine and liver function indicators(P <0.05),but this medicine has no significant effect on renal function indicators such as uric acid,creatinine,and urea nitrogen;(3)Adverse events related to MTX were found in 45% of patients.Gastrointestinal discomfort(45%)and abnormal liver function(26%)were the most common.No serious adverse events occurred in all patients;(4)The improvement of skin lesions in patients without psoriatic arthritis(Ps O)is generally better than patients with psoriatic arthritis(Ps A).Liver function were more severely affected by MTX and adverse reactions are also occur more often in patients with Ps A;(5)In the preliminary screening of candidate SNPs,it was found that the CC genotype of rs4112788 on LCE3D(P = 0.002)and the TT genotype of rs10036748 on TNIP1(P = 0.011)were significantly different between responser and non-responser.TT genotype in rs10036748 was further validated to be associated with better drug response.(6)Except for genetic factors,after adjusting for the confounding factors,patients with low BMI and without psoriatic arthritis would have a better response to MTX(P < 0.05);(7)We enrolled 441 samples into our GWAS analysis.In first stage,we made an association study by using whole genome genotyping data of 185 responser and 150 non-responser,after quality control,41 SNPs were selected out.Significant differences in the allele frequencies of rs4405344(P = 0.02945)and rs4713429(P = 0.04795)were found in verification stage in 108 independent samples.By combining the results of two-stage,after quality control,a suggestive signal rs4713429 was found to be related to the response of MTX(P = 7.93 × 10-6,OR = 2.06).Rs4713429 located in the upstream of 5' untranslated region of HCG22 which significantly affects the m RNA expression of HLA-C and VARS2.Conclusions This is a first a complete series of pharmacogenomics studies of MTX in patients with psoriasis was conducted in a Chinese population based on a prospective cohort collection,candidate gene analysis and genome-wide association analysis.This study found that the effective rate of MTX in treating psoriasis was about 46%,and the genotype of rs10036748 which is the susceptibility marker for psoriasis was also related to the effectiveness of MTX in treating psoriasis.In this study,we first used GWAS method to identify a suggestive SNP rs4713429 related to MTX response in 441 Chinese patients with psoriasis.Bioinformatics analysis further suggested that HLA-C and VARS2 might have a potential to be novel drug targets for the treatment of psoriasis.This study will help to improve the genetic mechanism related to MTX response and lay a theoretical foundation for the future development of precision medicine. |
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