Phamacogenomic studies for the genetic basis for lethal arrhythmias

对120例正常志愿者、先天性长QT综合征患者和20例正常对照者的基因组DNA进行了遗传筛查，（cLQTS ; n=30），获得性长QT综合征（aLQTS ; n=4），无症状Brugada综合征（aBS ; n=30），症状性Brugada综合征结果发现cLQTS中存在心脏possatium通道基因KCNH 2突变。在心脏Na通道基因（SCN 5A）中发现9个不同的突变：cLQTS（2/30）、aLQTS（1/4）、aBS（1/30）、sBS（5/37）、PCCD（1/2）和AS（1/2）。使用哺乳动物培养细胞tsA-201，通过定点诱变和异源表达系统确定8个SCN 5A突变的生物物理异常。通过瞬时转染这些突变Na通道记录全细胞Na电流。负责cLQTS和aLQTS的3个突变体通道显示出持续的Na电流，这是其他LQT 3突变体通道的特征。sBS和AS突变体通道的5个突变体中的2个突变体通道未显示可观察到的电流。aBS的突变是在SCN 5A的同一个外显子的双突变，并表现出BS的功能特性的损失。我们已经确定了两个新的单核苷酸多态性（SNP）在SCN 5A。V1951 L和L1988 R。TaqMan PCR法检测的等位基因频率在正常组和Brugada综合征组之间差异无统计学意义。这些SNP的氟卡尼敏感性与野生型Na通道几乎相同，表明这些SNP不太可能是对Na通道阻滞剂敏感性差异的基础。在aBS病例中鉴定的双重突变是导致aBS的第一个SCN 5A突变（Yokoi et al. Heart Rhythm 2005）。导致AS的突变是迄今为止报道的第三个SCN 5A突变。我们发现在这种情况下，在连接蛋白40的伴随SNP，现在调查的发病机制，心房特异性的性质，这种心律失常。

Genetic screening was performed in the genomic DNAs from normal volunteers (n=120), congenital long QT syndrome (cLQTS ; n=30), acquired long QT syndrome (aLQTS ; n=4), asymptomatic Brugada syndrome (aBS ; n=30), symptomatic Brugada syndrome (sBS ; n=37), progressive cardiac conduction disturbance (PCCD ; n=2), and atrial standstill (AS ; n=2).We found a cardiac possatium channel gene KCNH2 mutation in cLQTS. Nine distinct mutations were found in cardiac Na channels genes (SCN5A) ; cLQTS(2/30), aLQTS(1/4), aBS(1/30), sBS(5/37), PCCD(1/2), and AS(1/2). Biophysical abnormalities of the eight SCN5A mutations were determined by site-directed mutagenesis and heterologous expression system using mammalian cultured cell tsA-201. Whole cell Na current were recorded by transient transfection of these mutant Na channels. 3 mutant channels responsible for cLQTS and aLQTS showed persistent Na current that is characteristic for other LQT3 mutant channels. 2 mutant channels out of 5 mutation of sBS and the AS mutant channel showed no observable current. The mutation responsible for aBS was a double mutation at the same exon of SCN5A, and showed loss-of function properties characteristic for BS.We have identified two novel single nucleotide polymorphisms (SNP) in SCN5A. V1951L and L1988R. Allele frequencies determined by TaqMan PCR strategies was not significantly difference between normal group and Brugada syndrome group. Flecainide sensitivities of these SNPs were nearly identical to wild type Na channel, suggesting that these SNPs are unlikely to underlie differences in the sensitivities to Na channel blockers.The double mutation identified in an aBS case is the first SCN5A mutation responsible for aBS (Yokoi et al. Heart Rhythm 2005). The mutation responsible for AS was the third SCN5A mutation reported so far. We found a concomitant SNP in the connexin 40 in this case, and now investigating the pathogenesis of atria-specific nature of this arrhythmia.