Research for development of therapy of Machado-Joseph disease by siRNA

Machado-Joseph病是一种进行性遗传退行性疾病，是由位于染色体14q32.1的MJD中CAG重复序列不稳定扩增引起的。MJD的基因产物ataxin-3突变体携带扩展的聚谷氨酰胺拉伸，获得不良功能，导致神经元功能障碍，导致神经元死亡。目前还没有有效的治疗方法。如果能抑制突变蛋白的表达，我们就能很好地治疗这种疾病。短干扰RNA （siRNA）将是建立有效治疗方法的良好候选者。亨廷顿氏病也是一种多谷氨酰胺疾病，由HD基因1外显子CAG重复序列异常扩增引起。研究了sirna对HD基因的作用，以降低其在培养细胞中的表达。一种名为siRNA-HD- exon1的siRNA靶向CAG重复序列的上游区域，可以有效地特异性抑制HD外显子1-EGFP融合迷你基因的表达。它也能有效抑制人神经母细胞瘤培养细胞系中内源性亨廷顿蛋白的表达（Proc Japan Acad 79(Ser B): 293-298, 2003）。MJD基因存在多个单核苷酸多态性（SNPs）。在日本人群中，这些snp有两种单倍型。一个是c527T-c669A-c987C-c1118A，另一个是c527C-c669G-c987G-c1118C。携带异常扩增CAG重复序列的疾病染色体只显示前一单倍型。如果携带该单倍型的等位基因的表达可以被合适的siRNA特异性有效地抑制，这将是开发MJD根治性治疗的第一步。两个研究小组报道了成功抑制单倍型特异性表达的siRNA，其目标是c987C/G SNP周围的序列。

Machado-Joseph disease is a progressive heredodegenerative disease that is caused by unstable expansions of CAG repeats in MJD which is located at chromosome 14q32.1. The mutant ataxin-3, the gene product of MJD, carrying the expanded polyglutamine stretch gains an adverse function and causes dysfunction of neurons, leading to neuronal death. No effective therapeutics has been established. If the expression of the mutant protein could be inhibited, we could treat the disease very well. Short interfering RNA (siRNA) would be a good candidate to establish an effective therapeutics.Huntington's disease is also one of the polyglutamine diseases and cased by an abnormally expanded CAG repeats in exon 1 of the HD gene. It was investigated that the effects of siRNAs directed against the HD gene in order to knock down its expression in cultured cells. One siRNA, named as siRNA-HD-Exon1, which is targeted against the region at immediate upstream of the CAG repeats, can efficiently and specifically inhibit the expression of HD exon 1-EGFP fusion mini-gene. It also did efficiently suppress the endogenous huntingtin expression in cultured cell lines derived from human neuroblastoma (Proc Japan Acad 79(Ser B) : 293-298, 2003).There are several single nucleotide polymorphisms (SNPs) in the MJD gene. In Japanese population, there are two haplotypes of those SNPs. One is c527T-c669A-c987C-c1118A and the other is c527C-c669G-c987G-c1118C.Disease chromosomes which carry the abnormally expanded CAG repeats show exclusively the former haplotype. If the expression of the allele carrying that haplotype could be specifically and effectively inhibited by a suitable siRNA, it would be the first step toward the development of a radical therapy for MJD. Two research groups reported successful suppression of the haplotype-specific expression by the siRNA of which the target is the sequence surrounding c987C/G SNP.