Mapping tumor specific immunopeptidome for antibody-based targeted therapy

项目摘要 基于抗体的免疫疗法已成为治疗癌症的既定范例。只有一小 然而，一部分患者已经从这些药物中获益，并且许多药物仅对一小部分患者有效 肿瘤。这部分是由于缺乏经验证的细胞表面靶标。只有十几种表面蛋白 已经批准用于抗体干预，并且大多数并不是真正针对恶性细胞的。出现 在寻找肿瘤特异性靶点的细胞免疫肽组方面越来越感兴趣。的 免疫肽组是由细胞内蛋白质的蛋白水解产生并展示在细胞表面的肽库。 细胞表面作为与人类白细胞抗原（HLA）的复合物。据认为，这种肽库 反映了细胞内蛋白质组的状态，并可由驱动癌基因调节。表位窝藏 已经从肿瘤样品中检测到作为肽-HLA复合物（pHLA）的驱动突变或新抗原。 然而，癌基因信号传导如何塑造免疫肽组的机制仍然没有得到表征。 这项提案的目标是开发技术，以绘制独特的免疫肽组的变化， 特定的原基因，以及针对它们的免疫反应的工具。这些发现最终将有助于 开发具有更好安全性和更适用于不同类型癌症的基于抗体的生物制剂。 在此，我们假设增殖性癌基因通过转录调节免疫肽组， 重塑和选择性剪接，并诱导独特的pHLAs超越突变片段的亲本 癌蛋白为了验证这一假设，我们将首先开发一种新的质谱技术， 以等位基因特异性方式表达免疫肽组。接下来，我们将联合收割机这个技术与还原细胞 模型方法研究过表达特定癌基因的同基因细胞系的免疫肽组。的 免疫肽组学数据将通过蛋白质数据库和转录分析来补充，以揭示关键的 癌基因特异性pHLA的贡献者。总的来说，拟议的工作将提供洞察如何一个单一的 遗传改变可影响抗原呈递机制并在细胞表面上产生新抗原。 在这些发现的基础上，我们还将设计抗体，以高亲和力识别感兴趣的pHLA 和肽特异性。这些试剂将进一步阐述成嵌合抗原受体，以武装细胞毒性 T细胞并对展示癌基因特异性pHLA的细胞产生抗肿瘤应答。这些试剂将 在培养的细胞和人源化小鼠模型中测试。从长远来看，基本的生物学见解和抗体 从拟议的研究中产生的工具将支持正在进行的努力，以了解癌症 免疫肽组，并奠定了基础，为未来的治疗靶向新抗原。

Project Summary Antibody-based immunotherapies have become an established paradigm for treating cancers. Only a small portion of patients have benefited from these agents, though, and many are only effective for treating a small set of tumors. This is due, in part, to a lack of validated cell surface targets. Only a dozen of surface proteins has been approved for antibody intervention, and most are not truly exclusive to malignant cells. There has been growing interesting in searching the cellular immunopeptidome for tumor-exclusive targets. The immunopeptidome is a pool of peptides generated from proteolysis of intracellular proteins and displayed on the cell surface as a complex with the human leukocyte antigens (HLA). It is thought that this peptide repertoire reflects the status of the intracellular proteome and can be modulated by driver oncogenes. Epitopes harboring driver mutations, or neo-antigens, have been detected as peptide-HLA complexes (pHLAs) from tumor samples. However, the mechanism of how oncogene signaling shape the immunopeptidome remains uncharacterized. The goals of this proposal are to develop technologies to map changes in the immunopeptidome unique to specific ongene, and tools to mount an immune response against them. These findings will ultimately aid in developing antibody-based biologics with better safety profile and more applicable to diverse types of cancers. Here, we hypothesized that proliferative oncogenes globally alter the immunopeptidome via transcriptional remodeling and alternative splicing, and induce unique pHLAs beyond mutated fragments of parent oncoproteins. To test this hypothesis, we will first develop a new mass spectrometry technique to map diseased immunopeptidome in an allele-specific manner. Next, we will combine this technique with a reductionist cell model approach to study the immunopeptidome of isogenic cell lines overexpressing specific oncogene. The immunopeptidomics data will be complemented by protein databases and transcript analyses to reveal key contributors of oncogene-specific pHLAs. Collectively, the proposed work will provide insight to how a single genetic alteration can impact mechanisms of antigen presentation and produce neo-antigens on the cell surface. To build on these discoveries, we will also engineer antibodies to recognize pHLAs of interest with high affinity and peptide specificity. These reagents will be further elaborated into chimeric antigen receptors to arm cytotoxic T cells and mount antitumor response against cells displaying oncogene-specific pHLAs. These reagents will be tested in cultured cells, and humanized mice models. In the long term, the basic biological insights and antibody tools generated from the proposed studies will bolster ongoing efforts to understanding the cancerous immunopeptidome and lay the foundation for future therapeutics targeting neo-antigens.