Disclosed are methods of proteolytically converting a precursor protein (e.g. tau) to a product fragment (e.g. a 12 kd fragment) in a stable cell line, wherein the precursor protein is associated with a disease state in which the precursor protein aggregates pathologically (e.g. a tauopathy), and the methods comprise:(a) providing a stable cell line transfected with nucleic acid encoding: (i) a template fragment of the precursor protein such that the template fragment is constitutively expressed in the cell at a level which is not toxic to the cell; and (ii) the precursor protein, which protein is inducibly expressed in the cell in response to a stimulus, whereby interaction of the template fragment with the precursor protein causes a conformational change in the precursor protein such as to cause aggregation and proteolytic processing of the precursor protein to the product fragment. The method is preferably used to screen for modulators of the aggregation process by monitoring production (or modulation of production) of the product band or bands. Also provided are materials for used in the assays, plus medicaments, and related uses and processes, based on compounds which show high activity in the assay of the invention e.g. reduced diaminophenothiazines.
公开了在稳定
细胞系中将前体蛋白(如 tau)蛋白
水解转化为产物片段(如 12 kd 片段)的方法,其中前体蛋白与疾病状态相关,在疾病状态中,前体蛋白病理性聚集(如 tau 病)。(a) 提供转染有编码核酸的稳定
细胞系:(i) 前体蛋白的模板片段,使模板片段在细胞中以对细胞无毒性的
水平组成性表达;以及 (ii) 前体蛋白,该蛋白在刺激下在细胞中诱导性表达,模板片段与前体蛋白的相互作用导致前体蛋白的构象变化,从而引起前体蛋白的聚集和蛋白
水解加工为产物片段。该方法最好用于通过监测一条或多条产物带的产生(或对产生的调节)来筛选聚集过程的调节剂。此外,还提供了用于检测的材料,以及基于在本发明检测中显示高活性的化合物(如还原型二
氨基
吩噻嗪)的药物及相关用途和工艺。