Covalent Fluorescent Probes for Cancer Cell Detection

摘要：荧光成像是一种强大的工具，可以可视化种群中特定的细胞状态；然而，现有的荧光标记方法对于许多生物系统来说并不是实验上可用的。此外，与现有的荧光成像方法相比，细胞状态的荧光小分子传感器可能提供一种有价值的替代方案。的总目标是 这个项目是创造荧光小分子三磷酸腺苷定点探针，可以选择性地标记特定的激酶，并作为正常和病理细胞状态的成像探针。蛋白激酶在许多方面都是开发用于癌症生物学的选择性小分子成像探针的理想靶点。 这是因为蛋白激酶参与了大多数细胞过程，它们的定位、可及性和丰度的变化与细胞状态的变化有关。蛋白激酶已被用作癌症生物学中的生物标志物，因为点突变、基因缺失、基因扩增和染色体重排导致的内源性激酶调节机制的丧失已得到充分证实。 作为许多癌症的关键事件。本项目的具体目标是：1)合成能够与三磷酸腺苷半胱氨酸残基形成共价键的荧光标记的激酶抑制剂；2)利用显微镜筛选目标1中制备的化合物，以鉴定作为正常和病理细胞状态选择性探针的化合物；以及3)鉴定目标2中确定的活性化合物的细胞内靶标(S)。 这项提议是Dana Farber癌症研究所的Nathanael Gray博士、哈佛医学院的Priscilla Yang博士和波士顿大学的张伟博士之间正在进行的合作的延伸。这三人也都是达纳·法伯/哈佛癌症中心的成员。这项建议的优点之一是，每个合作伙伴负责三个具体目标中的一个，使项目受益于每个人及其后续机构为伙伴关系带来的平衡技能和专门知识。因为这三个人 调查人员的职业生涯刚刚起步，格雷、杨和张博士将受益于职业发展活动和内在和外在的导师。此外，培训核心将对这一项目作出重大贡献。

Abstract: Fluorescence imaging is a powerful tool that permits visualization of specific cell states within a population; however, existing methods for fluorescence labeling are not experimentally accessible for many biological systems. Furthermore, fluorescent small-molecule sensors of cell state may provide a valuable alternative with significant benefits relative to existing methods for fluorescence imaging. The overall goal of this project is to create fluorescent small molecule ATP-site directed probes that can selectively label particular kinases and serve as imaging probes of normal versus pathological cell state. Protein kinases are in many ways ideal targets for the development of selective small molecule imaging probes for use in cancer biology. This is because protein kinases are involved in most cellular processes and changes in their localization, accessibility, and abundance are associated with changes in cellular state. Protein kinases have been used as biomarkers in cancer biology because the loss of endogenous kinase regulatory mechanisms by point mutations, gene deletions, gene amplifications, and chromosomal rearrangements has been well-established as crucial events in many cancers. The specific aims of this project are to: 1) Synthesize fluorescently-tagged kinase inhibitors capable of forming covalent bonds with ATP-site cysteine residues; 2) Use microscopy-based screening ofthe compounds prepared in Aim 1 to identify compounds that are selective-probes of normal and pathological cellular states and 3) Identify the intracellular target(s) of active compounds identified in Aim 2. This proposed is the extension of an ongoing collaboration between Drs. Nathanael Gray (Dana Farber Cancer Institute), Priscilla Yang (Harvard Medical School) and Wei Zhang (UMass Boston). All three are also members of the Dana Farber/Harvard Cancer Center. One of the strong points of this proposal is that each Co-Pl is responsible for one of the three Specific Aims allowing the project to be benefited by the balanced skills and expertise that each person and their subsequent institution brings to the partnership. Because all three investigators are eariy in their careers, Drs. Gray, Yang and Zhang will benefit from career development activities and mentors who are intemal and extemal. Additionally, the Training Core will significantiy contribute to this project.