Mapping a clinically significant internalizing tumor epitope space.

这项提议的目标是开发基于抗体库的方法，以促进有效的鉴定 临床上相关的肿瘤细胞表面抗原，适用于鉴定的方法 一般情况下，细胞类型特定的谱系标记。肿瘤细胞的异常生理在一定程度上反映在 它们细胞表面的化学和分子组成发生变化。表面的渐变 分子表达使肿瘤细胞能够有效地对外部信号做出反应，以促进生长和生存 与宿主组织相互作用，以实现转移，并避免免疫监视。身份识别和 然而，肿瘤特异性细胞表面抗原的靶向性受到表位复杂性的阻碍。 肿瘤细胞表面的空隙。除了蛋白质，相关的抗原还包括碳水化合物和其他 翻译后修饰产物不能从基因组拷贝数或 M RNA表达水平。该建议旨在开发基于组合抗体库的策略， 允许有效地识别用于诊断、预后和治疗的肿瘤特异性细胞表面抗原 申请。具体地说，我们的目标是(1)开发一种基于以下公式的高通量减法选择策略 流式细胞仪分选显著提高选择效率，获得更多肿瘤- 靶向噬菌体抗体。(2)通过筛选噬菌体抗体鉴定临床相关的肿瘤抗原 在适当的间质环境中，原位对抗前列腺癌细胞。我们建议将抗体 用激光捕获显微切割(LCM)鉴定抗肿瘤抗体的文库技术 组织载片上的细胞。由此产生的抗体将有很高的可能性在临床上识别 相关肿瘤抗原。(3)建立识别的肿瘤细胞表面抗原的分子鉴定 肿瘤特异性抗体。我们开发了一种新型的组合酵母cDNA展示文库。特定的 流式分选后，将从该文库中鉴定抗原-抗体对。同时，我们会继续推行 翻译后分析的质谱学方法分析肿瘤抗原 修改。这项研究将增加我们对肿瘤生理学的认识，并将有助于设计 有效的治疗；它还将有助于确定翻译后修改对 肿瘤表位空间的最终组成。

The goal of this proposal is to develop antibody library-based methods which promote efficient identification of clinically relevant tumor specific cell surface antigens, methods which are applicable to the identification of cell type-specific lineage markers in general. The abnormal physiology of tumor cells is reflected in part in the altered chemical and molecular composition of their cell surface. Progressive changes in surface molecule expression allow tumor cells to respond efficiently to external signals for growth and survival, to interact with host tissues, to achieve metastasis, and to avoid immune surveillance. The identification and targeting of tumor-specific cell surface antigens, however, is hampered by the complexity of the epitope space at the tumor cell surface. In addition to proteins, relevant antigens include carbohydrates and other post-translational modification products that cannot be predicted from studies of genomic copy number or mRNA expression levels. This proposal aims to develop combinatiroal antibody library-based strategies that allow efficient identification of tumor specific cell surface antigens for diagnostic, prognostic and therapeutic applications. Specifically, we aim (1) To develop a high throughput subtractive selection strategy based on flow cytometric sorting to significantly improve selection efficiency and to obtain greater numbers of tumor- targeting phage antibodies. (2) To identify clinically relevant tumor antigens by selecting phage antibodies against prostate cancer cells in situ, within their proper stromal contexts. We propose to combine antibody library technology with laser capture microdissection (LCM) to identify antibodies against individual tumor cells on tissue slides. The resulting antibodies will have a very high likelihood of recognizing clinically relevant tumor antigens. (3) To establish the molecular identify of tumor cell surface antigens recognized by tumor-specific antibodies. We have developed a novel combinatorial yeast cDNA display library. Specific antigen-antibody pairs will be identified from this library following flow sorting. In parallel, we will pursue the analysis of tumor antigens by mass spectrometry methods which are capable of analyzing post-translational modifications. This study will increase our knowledge of tumor physiology and will facilitate the design of effective therapy; it will also help determine the exact contribution of post-translational modifications to the final makeup of the tumor epitope space.