Pham Tran Thu Ha, Tran Quang Binh, Sukasem Chonlaphat, Nguyen Van Dinh, Chu Chi Hieu, Do Thi Quynh Nga, Tran Ngoc Phuong Mai, Phung Thanh Huong
Hanoi University of Pharmacy, Hanoi, Vietnam.
National Institute of Nutrition, Hanoi, Vietnam.
Appl Clin Genet. 2021 Feb 9;14:27-35. doi: 10.2147/TACG.S278652. eCollection 2021.
Allopurinol, a common anti-hyperuricemia drug, is well known as an inducer of severe cutaneous adverse drug reactions (SCARs). One of the most well-defined risk factors of allopurinol-induced SCARs is the presence of polymorphic alleles of human leukocyte antigen () genes, such as and alleles. There is no commercial test or published in-house protocol for the specific detection of the allele. In this article, we established for the first time a simple allele-specific (AS) PCR method to identify allele carriers, and at the same time, determine their zygosities.
A two-step AS-PCR protocol, using four primer sets, was designed to specifically amplify and differentiate the allele from 17 other alleles found in Vietnamese people. The protocol was validated with PCR-sequencing-based typing (SBT) of 100 samples of unknown genotypes.
The PCR protocol can detect the allele and determine the zygosity. The results of this protocol were highly consistent with those of the SBT (ĸ = 0.98, p < 0.001). Regarding the specific detection of the allele, the PCR protocol had a sensitivity of 100% (95% CI: 91.61-100%) and specificity of 98.3% (95% CI: 90.9-99.7%). The protocol was used to determine the distribution of the allele in 810 unrelated Vietnamese Kinh people, 14.2% of which were carriers, the allele frequency was 7.5%.
A novel AS-PCR protocol with a sensitivity of 100% for the detection of the allele was established. The protocol can be used for personalized treatment with allopurinol in order to minimize the risk of SCARs in Vietnamese people as well as in other Asian populations with similar genetic characteristics.
别嘌醇是一种常见的抗高尿酸血症药物，是严重皮肤不良反应（SCARs）的诱导剂。别嘌醇诱导SCARs最明确的危险因素之一是人类白细胞抗原（HLA）基因多态性等位基因的存在，如HLA-B58:01和HLA-A31:01等位基因。目前尚无用于特异性检测HLA-B58:01等位基因的商业检测方法或已发表的内部方案。在本文中，我们首次建立了一种简单的等位基因特异性（AS）PCR方法来鉴定HLA-B58:01等位基因携带者，并同时确定其纯合性。
设计了一种两步AS-PCR方案，使用四组引物，以特异性扩增并区分越南人群中发现的HLA-B*58:01等位基因与其他17种HLA-B等位基因。该方案通过对100个未知基因型样本进行基于PCR测序的分型（SBT）进行验证。
该PCR方案能够检测HLA-B58:01等位基因并确定其纯合性。该方案的结果与SBT的结果高度一致（κ = 0.98，p < 0.001）。关于HLA-B58:01等位基因的特异性检测，该PCR方案的灵敏度为100%（95%CI：91.61 - 100%），特异性为98.3%（95%CI：90.9 - 99.7%）。该方案用于确定810名无亲缘关系的越南京族人群中HLA-B*58:01等位基因的分布，其中14.2%为携带者，等位基因频率为7.5%。
建立了一种新型AS-PCR方案，对HLA-B*58:01等位基因检测的灵敏度为100%。该方案可用于别嘌醇的个体化治疗，以尽量降低越南人群以及其他具有相似遗传特征的亚洲人群中SCARs的风险。
