High-throughput screening platform for cancer drug discovery

 描述（由申请人提供）：靶向治疗在癌症治疗中显示出巨大的前景。高通量筛选（HTS）活动通常依赖于使用标记的配体或酶底物的测定。与标记相关的伪影导致错误识别作用于标记底物而不是预期靶点的化合物。无标记、基于溶液的HTS方法将有助于鉴定对预期靶点特异性起作用的化合物。这项拟议研究的目标是开发一种光谱-光学量热技术，该技术将使量热法能够用于针对癌症治疗靶点的化合物库的高通量筛选。为了实现这一目标，热致变色材料，它将转换为吸收波长的变化的温差将被选择，一个微流控装置，以产生反应液滴，并提供反应物的快速混合将被制造，一个光学检测系统，能够检测温度变化的500 pl反应将建成，该技术将通过测量一套良好的表征反应进行验证。将通过测量已知抑制剂对组蛋白脱乙酰酶1和6的结合和抑制来证明该方法。光谱-光学量热法技术将使结合和酶促反应的完全热力学表征成为可能，当考虑到新靶点标记HTS检测开发的需求时，其通量显著优于1536孔板检测。更高的通量和更低的样品消耗的结合将改变研究人员看待量热法的方式：而不是被视为有限数量的高价值测量的技术，它将被视为主要的筛选方法。

 DESCRIPTION (provided by applicant): Targeted therapies have shown great promise in the treatment of cancer. High-throughput screening (HTS) campaigns have often relied on assays using labeled ligands or enzyme substrates. Artifacts associated with labeling have led to the erroneous identification of compounds that act in on the labeled substrate rather than the intended target. A label-free, solution-based HTS method will facilitate identification of compounds acting specifically on the intended targets. The goal of this proposed research is to develop a spectro-optical calorimetry technology that will enable the use of calorimetry in high-throughput screening of compound libraries against cancer therapeutic targets. To achieve this goal, a thermochromic material, which converts temperature differences to absorbance wavelength changes will be selected, a microfluidic device to generate reaction droplets and provide rapid mixing of reactants will be fabricated, an optical detection system capable of detecting temperature changes in 500 pl reactions will be built, and the technique will be validated by measuring a suite of well-characterized reactions. The method will be demonstrated by measuring the binding to and inhibition of histone deacetylases 1 and 6 by known inhibitors. The spectro-optical calorimetry technology will enable full thermodynamic characterization of binding and enzymatic reactions with significantly better than the throughput of 1536-well plate assays when accounting for the need for labeled HTS assay development for new targets. The combination of higher throughput and lower sample consumption will change the way researchers view calorimetry: rather than being seen as a technique for a limited number of high-value measurements, it will be viewed as a primary screening method.