Genotyping assays for the most common mutations associated with abacavir hypersen

描述（由申请人提供）：我们建议开发一种近患者测定法，以对与阿巴卡韦-严重皮肤不良反应（SCAR）相关的2个SNP进行评分;即，HLA B*5701和Hsp 70-Hom M493 T等位基因（也称为Hsp 70-Hom M493 T等位基因）。rs2227956CT）。将用Trimgen's BuccalQuick”（Sparks，MD）从颊拭子获得的细胞中分离核酸模板。将使用我们专有的解旋酶依赖性扩增（HDA）开发两种基因分型检测。HDA产品将使用装在扩增子密闭容器中的侧向流装置进行检测，以防止实验室污染。在第一阶段，我们将实施设计控制，并确保BioHestival获得ISO 13485认证。我们还将使用该技术平台开发上述SNP位点的分型测定。最后，我们将与PGXL合作，使用PGXL研究人员收集的一组50份临床标本，相对于金标准（DNA测序）验证这些检测试剂盒的性能。在第二阶段，我们将进行一项前瞻性、多中心的临床研究，旨在验证这些检测方法作为一种手段来预测服用阿巴卡韦治疗艾滋病的患者发生SJS和TEN的风险。公共卫生相关性：阿巴卡韦（ABC）是一种人类免疫缺陷病毒-1（HIV-1）核苷类似物逆转录酶抑制剂，在Trizivir中与拉米夫定（3 TC）和齐多夫定（ZDV）共同配制;每日一片，无食物限制，药物相互作用有限，耐药性良好。Trizivir与其他HIV药物联合，是美国卫生和人类服务部共识小组指南推荐的抗逆转录病毒初治患者的可能初始治疗。在Epzicom（R）中，ABV也与拉米夫定联合使用。此外，ABC（作为单一药物以Ziagen（R）的名称出售）由于其成本低，是发展中国家使用的一线疗法的一部分（www.who.int/hiv/pub/guidelines/adult/en/index.html）。两个单核苷酸多态性（SNP）与阿巴卡韦严重皮肤不良反应（SCAR）高度相关。HLA B*5701是高加索人ABC超敏反应的已知遗传风险因素（Martin et al. 2005）。Hsp 70-Hom M493 T等位基因（又称Hsp 70-Hom M493 T等位基因）是Hsp 70-Hom M493 T等位基因（又称Hsp 70-Hom M493 T等位基因）。rs 2227956 CT）与HLA-B*5701的组合与ABC超敏反应强烈相关。虽然存在HLA分型的测定，但这些测定是昂贵的，并且产生更多的信息，而这些信息对于检测ABC超敏反应的风险是严格必要的。例如，靶向DNA测序用于高分辨率分型，基于序列特异性引物（SSP）或序列特异性寡核苷酸探针（SSOP）的测定用于低分辨率HLA分型。SSP和SOPP测定通常对每个基因座使用20至100个不同的引物组或探针。DNA测序仍然是一种复杂且昂贵的DNA诊断方法。我们建议开发的方法将是成本较低，易于在附近的病人设置实施。

DESCRIPTION (provided by applicant): We propose to develop a near patient assays to score 2 SNPs associated with abacavir-severe cutaneous adverse reactions (SCAR); i.e., HLA B*5701 and the Hsp70-Hom M493T allele (a.k.a. rs2227956CT). Nucleic acid templates will be isolated from cells obtained from cheek swabs with the Trimgen's BuccalQuick" (Sparks, MD). Two genotyping assays will be developed using our proprietary helicase dependent amplification (HDA). HDA products will be detected with a lateral flow device encased in an amplicon containment vessel that prevents contamination of the laboratory. In Phase I we will Implement design control, and secure ISO 13485 certification for BioHelix. We will also develop typing assays for the aforementioned SNP loci using this technology platform. Finally we will collaborate with PGXL to validate the performance of these assays relative to a gold standard (DNA sequencing) using a set of 50 clinical specimens collected by PGXL researchers. In Phase II, we would perform a prospective, multi-site clinical study aimed at validating the assays as a means of predicting the risk of SJS and TEN in patients being prescribed abacavir for the treatment of AIDS. PUBLIC HEALTH RELEVANCE: Abacavir (ABC), a human immunodeficiency virus-1 (HIV-1) nucleoside- analogue reverse transcriptase inhibitor, is coformulated with lamivudine (3TC ) and zidovudine (ZDV) in Trizivir; a single one a day pill with no food restrictions, limited drug-drug interactions, and a favorable resistance profile. Trizivir in combination with other HIV drugs, is recommended by the US Department of Health and Human Services Consensus Panel Guidelines as a possible initial treatment in antiretroviral-naive patients. ABV is also used in a combo with lamivudine in Epzicom(R). In addition, ABC (sold as a single drug under the name Ziagen(R)) is part of the first-line regimens used in developing countries because of its low cost (www.who.int/hiv/pub/guidelines/adult/en/index.html). Two single nucleotide polymorphisms (SNP) are highly association with abacavir-severe cutaneous adverse reactions (SCAR). HLA B*5701 is a known genetic risk factor for ABC hypersensitivity in Caucasians (Martin et al. 2005). The Hsp70-Hom M493T allele (a.k.a. rs2227956CT) in combination with HLA-B*5701 is strongly associated with ABC hypersensitivity. Although assays exist for HLA typing, these are costly, and yield more information that the strict necessary for detecting the risk of ABC hypersensitivity. For example, targeted DNA sequencing is used for high resolution typing, and sequence-specific primer (SSP), or sequence specific oligonucleotide probe (SSOP) based assays are used for low resolution HLA typing. SSP and SOPP assays typically 20 to 100 different primer sets or probes are used for each locus. DNA sequencing remains a complex and costly method for DNA diagnostics. The method we propose to develop would be lower cost and easy to implement in near patient settings.