Kocisko David A, Baron Gerald S, Rubenstein Richard, Chen Jiancao, Kuizon Salomon, Caughey Byron
Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana 59840, USA.
J Virol. 2003 Oct;77(19):10288-94. doi: 10.1128/jvi.77.19.10288-10294.2003.
Transmissible spongiform encephalopathies (TSEs) are fatal, untreatable neurodegenerative diseases associated with the accumulation of a disease-specific form of prion protein (PrP) in the brain. One approach to TSE therapeutics is the inhibition of PrP accumulation. Indeed, many inhibitors of the accumulation of PrP associated with scrapie (PrP(Sc)) in scrapie-infected mouse neuroblastoma cells (ScN(2)a) also have antiscrapie activity in rodents. To expedite the search for potential TSE therapeutic agents, we have developed a high-throughput screening assay for PrP(Sc) inhibitors using ScN(2)a cells in a 96-well format. A library of 2000 drugs and natural products was screened in ScN(2)a cells infected with scrapie strain RML (Chandler) or 22L. Forty compounds were found to have concentrations causing 50% inhibition (IC(50)s) of PrP(Sc) accumulation of <or=10 microM against both strains. Seventeen had IC(50)s of <or=1 microM against both strains. Several classes of compounds were represented in the 17 most potent inhibitors, including naturally occurring polyphenols (e.g., tannic acid and tea extracts), phenothiazines, antihistamines, statins, and antimalarial compounds. These 17 compounds were also evaluated in a solid-phase cell-free hamster PrP conversion assay. Only the polyphenols inhibited the cell-free reaction, and their IC(50)s were near 100 nM. Several of the new PrP(Sc) inhibitors cross the blood-brain barrier and thus have potential to be effective after TSE infection reaches the brain. The fact that many are either approved human drugs or edible natural products should facilitate their use in animal testing and clinical trials.
传染性海绵状脑病（TSEs）是致命的、无法治疗的神经退行性疾病，与一种疾病特异性形式的朊病毒蛋白（PrP）在大脑中的积累有关。TSE治疗的一种方法是抑制PrP的积累。事实上，许多在感染羊瘙痒病的小鼠神经母细胞瘤细胞（ScN(2)a）中抑制与羊瘙痒病相关的PrP（PrP(Sc)）积累的抑制剂在啮齿动物中也具有抗羊瘙痒病活性。为了加快寻找潜在的TSE治疗药物，我们开发了一种使用96孔板格式的ScN(2)a细胞对PrP(Sc)抑制剂进行高通量筛选的试验。在感染羊瘙痒病毒株RML（钱德勒）或22L的ScN(2)a细胞中筛选了一个包含2000种药物和天然产物的文库。发现有40种化合物对两种毒株都具有导致PrP(Sc)积累抑制50%（IC(50)）的浓度≤10 microM。17种化合物对两种毒株的IC(50)≤1 microM。17种最有效的抑制剂中有几类化合物，包括天然存在的多酚（如鞣酸和茶提取物）、吩噻嗪、抗组胺药、他汀类药物和抗疟化合物。这17种化合物还在固相无细胞仓鼠PrP转化试验中进行了评估。只有多酚抑制了无细胞反应，其IC(50)接近100 nM。几种新的PrP(Sc)抑制剂能够穿过血脑屏障，因此在TSE感染到达大脑后有可能有效。许多化合物要么是已批准的人类药物，要么是可食用的天然产物，这一事实应该便于它们用于动物试验和临床试验。
