Chemical Probes of Immunoproteasome Function in Cancer

描述（由申请人提供）：泛素-蛋白酶体系统（UPS）在许多细胞过程中的重要作用现在已经得到了很好的认识。然而，许多问题仍未得到解答。令人感兴趣但尚未完全理解的是免疫蛋白酶体的功能，这是组成蛋白酶体的另一种形式。研究表明，免疫蛋白酶体在细胞中的作用不仅仅局限于产生抗原肽，而是多方面的。在这方面，最近的研究表明，免疫蛋白酶体的催化亚基与包括癌症在内的多种疾病有关。尽管有这些治疗意义，但免疫蛋白酶体在癌症药物发现方面仍未得到很大程度的开发。更好地了解免疫蛋白酶体的功能对于开发最佳的LMP2抑制剂至关重要。为了更好地了解免疫蛋白酶体在细胞中的作用，我们最近开发了一种小分子探针（UK-101），专门针对免疫蛋白酶体的主要催化亚基LMP2。此外，我们发现LMP2在几种类型的原发肿瘤组织和癌细胞系中高表达。有趣的是，高表达LMP2的癌细胞比低表达LMP2的癌细胞对UK-101明显更敏感。与这些体外结果一致，UK-101在高表达LMP2的人前列腺肿瘤小鼠异种移植模型中抑制肿瘤生长，而对低表达LMP2的肿瘤则没有抑制作用。这些观察结果进一步支持了LMP2在癌细胞生长和存活中的潜在作用。本应用程序的目的是确定免疫蛋白酶体在前列腺癌中的作用，并探讨免疫蛋白酶体靶向方法在癌症治疗中的治疗潜力。为了实现这一目标，我们建议使用创新的化学探针，选择性地靶向和可视化免疫蛋白酶体的单个催化亚基。为了实现这一目标，提出了四个具体目标：1)验证免疫蛋白酶体作为UK-101的主要药理学靶点；2)确定催化活性免疫蛋白酶体的动态细胞分布；3)确定免疫蛋白酶体与组成型蛋白酶体相比的独特功能；4)通过构建结构多样化的小分子文库来开发最佳的LMP2抑制剂。拟议的工作结合了化学和生物方法。该项目的成功完成将大大扩展我们对免疫蛋白酶体作用的认识。此外，该研究将为开发免疫蛋白酶体特异性抑制剂作为潜在治疗剂提供关键信息。最后，小分子探针的应用将为泛素-蛋白酶体生物学提供有价值的研究工具。

DESCRIPTION (provided by applicant): The essential roles of the ubiquitin-proteasome system (UPS) in many cellular processes are now well recognized. However, many questions still remain unanswered. Intriguing yet not fully understood are the functions of the immunoproteasome, an alternative form of the constitutive proteasome. It has been shown that the roles of the immunoproteasome in cells are not just limited to the generation of antigenic peptides, but far more multifaceted. In this regard, recent studies have shown that catalytic subunits of the immunoproteasome are implicated in various types of diseases including cancer. Despite these therapeutic implications, the immunoproteasome remains largely untapped for cancer drug discovery. A better understanding of immunoproteasome functions would be crucial in developing the optimum LMP2 inhibitor(s). In our continuing efforts to better understand the role of the immunoproteasome in cells, we have recently developed a small molecule probe ('UK-101') that specifically targets LMP2, a major catalytic subunit of the immunoproteasome. Further, we found that LMP2 is highly expressed in several types of primary tumor tissues and cancer cell lines. Interestingly, cancer cells that highly express LMP2 are markedly more sensitive to UK-101 than those expressing low level of LMP2. Consistent with these in vitro results, UK-101 inhibits tumor growth in a mouse xenograft model of human prostate tumor that highly expresses LMP2, but not that of tumor that expresses low level of LMP2. These observations further support a potential role of LMP2 in cancer cell growth and survival. The goals of this application are to determine the roles of the immunoproteasome in prostate cancer and to investigate therapeutic potentials of the immunoproteasome-targeting approaches in cancer therapy. Towards this goal, we propose to use innovative chemical probes that selectively target and visualize individual catalytic subunits of the immunoproteasome. In order to accomplish this goal, four specific aims are proposed: 1) To validate the immunoproteasome as the major pharmacological target of UK-101, 2) To determine dynamic cellular distribution of the catalytically active immunoproteasome, and 3) To determine the unique functions of the immunoproteasome compared to the constitutive proteasome and 4) To develop an optimum LMP2 inhibitor(s) by generating a structurally diversified small molecule library. The proposed work integrates both chemical and biological approaches. Successful completion of the project will significantly expand our knowledge on the role of the immunoproteasome. In addition, the study will yield critical information in developing immunoproteasome-specific inhibitors as a potential therapeutic agent. Finally, small molecule probes that will be used in this application will provide valuable research tools for the ubiquitin-proteasome biology community.