Molecular Pharmacology and Chemistry Program and High Throughput Screening Core Facility, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA.
Proc Natl Acad Sci U S A. 2009 Dec 1;106(48):20228-33. doi: 10.1073/pnas.0910757106. Epub 2009 Nov 11.
gamma-Secretase cleaves multiple substrates within the transmembrane domain that include the amyloid precursor protein as well as the Notch family of receptors. These substrates are associated with Alzheimer disease and cancer. Despite extensive investigation of this protease, little is known regarding the regulation of gamma-secretase specificity. To discover selective inhibitors for drug development and for probing the mechanisms of gamma-secretase specificity, we screened chemical libraries and consequently developed a di-coumarin family of inhibitors that preferentially inhibit gamma-secretase-mediated production of Abeta42 over other cleavage activities. These coumarin dimer-based compounds interact with gamma-secretase by binding to an allosteric site. By developing a multiple photo-affinity probe approach, we demonstrate that this allosteric binding causes a conformational change within the active site of gamma-secretase at the S2 and S1 sub-sites that leads to selective inhibition of Abeta42. In conclusion, by using these di-coumarin compounds, we reveal a mechanism by which gamma-secretase specificity is regulated and provide insights into the molecular basis by which familial presenilin mutations may affect the active site and specificity of gamma-secretase. Furthermore, this class of selective inhibitors provides the basis for development of Alzheimer disease therapeutic agents.
γ-分泌酶在跨膜结构域内切割多种底物，包括淀粉样前体蛋白和 Notch 受体家族。这些底物与阿尔茨海默病和癌症有关。尽管对这种蛋白酶进行了广泛的研究，但对 γ-分泌酶特异性的调节知之甚少。为了发现选择性抑制剂用于药物开发和探究 γ-分泌酶特异性的机制，我们筛选了化学文库，随后开发了一类二香豆素抑制剂，该抑制剂优先抑制 γ-分泌酶介导的 Abeta42 的产生，而对其他切割活性的抑制作用较弱。这些基于香豆素二聚体的化合物通过与别构位点结合来与 γ-分泌酶相互作用。通过开发多光亲和探针方法，我们证明这种别构结合导致 γ-分泌酶的活性位点在 S2 和 S1 亚位点发生构象变化，从而选择性地抑制 Abeta42 的产生。总之，通过使用这些二香豆素化合物，我们揭示了 γ-分泌酶特异性调节的机制，并提供了有关家族早老素突变如何影响 γ-分泌酶的活性位点和特异性的分子基础的见解。此外，这类选择性抑制剂为开发治疗阿尔茨海默病的药物提供了基础。
