Investigation into pathology of CAG repeat diseases and development of therapies

脊髓和球性肌萎缩症（SBMA）是一种遗传性运动神经元疾病。SBMA的分子基础是雄激素受体（AR）基因中编码聚谷氨酰胺（polyQ）通道的三核苷酸CAG重复序列的扩增。我们在巨细胞病毒增强子和鸡β-肌动蛋白启动子的控制下，建立了表达含有24或97个CAG重复序列的全长人AR的转基因小鼠模型（AR- 97q小鼠）。首先，我们阉割雄性AR-97Q小鼠。去势雄鼠的运动功能、体重、寿命和突变AR的核积累均有显著改善。另一方面，睾酮给药使其症状显著加重，并增强了雌性小鼠突变AR的核积累。突变蛋白的核积累与多q束的扩展可能是在大多数多q疾病中诱导神经元细胞功能障碍和变性的重要原因。阉割后的雄鼠通过降低睾丸激素水平来抑制突变体AR的核积累，并改善其运动功能。同样，睾酮阻断治疗，使用leuprorelin（一种LHRH类似物，可以减少睾丸激素的释放），显示出突变AR的运动功能和核积累的显著改善。基于这些结果，我们开始使用leuprorelin进行双盲临床试验。接下来，我们将AR-97Q小鼠与过表达人HSP70诱导型的小鼠杂交。高表达HSP70可显著改善AR-97Q小鼠的运动功能。在双转基因小鼠中，核突变体AR的积累，特别是大复合形式的积累，显著减少。HSP70过表达也减少了单个突变AR的数量，表明突变AR的降解增强。这些发现表明，HSP70过表达通过减少核定位突变AR来改善小鼠SBMA表型，这可能是由于突变AR降解增强所致。少

Spinal and bulbar muscular atrophy (SBMA) is an inherited motor neuron disease. The molecular basis of SBMA is the expansion of a trinucleotide CAG repeat, which encodes the polyglutamine (polyQ) tract, in the androgen receptor (AR) gene. We generated transgenic mouse model expressing the full-length human AR containing either 24 or 97 CAG repeats under the control of a cytomegalovirus enhancer and a chicken β-actin promoter (AR-97Q mice). First, we castrated male AR-97Q mice. Castrated males showed profound improvement of their motor function, body weight, lifespan and nuclear accumulation of the mutant AR. On the other hand, testosterone administration caused a significant aggravation of their symptoms and enhanced nuclear accumulation of the mutant AR in female mice. Nuclear accumulation of the mutant protein with an expended polyQ tract is likely to be important in inducing neuronal cell dysfunction and degeneration in the majority of the polyQ diseases. Castration of the males pre … More vented the nuclear accumulation of the mutant AR by reducing the testosterone level, and improved their motor function. Similarly, testosterone blockade therapy, using leuprorelin, a LHRH analogue which reduces testosterone release from the testis, showed a marked amelioration of motor function and nuclear accumulation of the mutant AR. On the basis of these results, we started the double-blind clinical trial using leuprorelin.Next, we cross-bred AR-97Q mice with mice over-expressing the inducible form of human HSP70. High expression of HSP70 markedly ameliorated motor function of AR-97Q mice. In double transgenic mice, nuclear mutant AR accumulation, particularly that of the large complex form, was significantly reduced. Monomeric mutant AR was also reduced in amount by HSP70 over-expression, suggesting the enhanced degradation of mutant AR. These findings suggest that HSP70 over-expression ameliorates SBMA phenotypes in mice by reducing nuclear-localized mutant AR, which probably due to enhanced mutant AR degradation. Less