肿瘤微环境中异常表达的免疫卡控点B7-H1、B7-H3和B7-H4等是参与肿瘤免疫逃逸的关键分子，但其在肿瘤演进中的表达规律及涉及的关键细胞和机制尚待进一步探讨。我们自主研制了系列抗人B7-H1、B7-H3和B7-H4单抗，发现B7-H1、B7-H3在结直肠息肉阶段表达上调，B7-H4仅在癌变阶段表达，且表达与浸润免疫细胞相关。鉴此，本项目（1）对已获得的B7-H1、B7-H3和B7-H4免疫组化抗体进一步鉴定，按临床病理诊断试剂要求建立质控参数；（2）建立新型的免疫组化多重染色方法，实现结直肠癌组织中3种分子的精确定位；（3）利用特异性单抗动态分析上述分子在人结直肠正常和癌变组织的表达规律及其与浸润免疫细胞的相关性和可能的调控机制。本研究不仅为免疫卡控点治疗提供伴随诊断试剂，还通过多重染色对人结直肠癌演进组织进行动态分析，为疾病免疫评价提供多参数指标，有助于阐明肿瘤免疫逃逸的机制。

Recently, we found that B7-H1(PD-L1), B7-H3 and B7-H4 were highly expressed in colorectal cancer tissues and showed differential expression levels during disease progression. B7-H1, B7-H3 and B7-H4 expression in colorectal cancer tissues can be heterogeneous in staining intensity and have variable distribution throughout an individual tissue. We have succeeded in developing anti-human B7-H1, B7-H3 and B7-H4 mAbs suitable for immunohistochemical staining in formalin-fixed tissue sections with reliable clarity and intensity. The purpose of the project is to apply multiplex immunohistochemistry and objective assessment parameters to identify B7-H1, B7-H3 and B7-H4 that correlate with colorectal cancer status, T cell infiltrate, and patient survival. B7-H1, B7-H3 and B7-H4 monoclonal antibodies were developed for IHC detection providing a consistent and objective assessment for tumor diagnosis and prognosis on a multi-analysis basis. Three-color IHC detection will be used in a single tissue sample with B7-H1, B7-H3 and B7-H4 monoclonal antibodies achieving a high level of sensitivity and specificity. We will systematically determine the expression pattern of B7-H1, B7-H3 and B7-H4 molecules at different stages of colorectal cancer and its correlation with immune cell subsets in tumor microenvironment, as well as clinic pathological parameters. Relationships analysis among B7-H1, B7-H3, B7-H4, CD8+ TILs and macrophage will be performed to examine the dynamic interactions that occur within the tumor microenvironment, which could be significant to determine the immune classification (immunoscore) in cancer patients. These mAbs are potentially useful as companion diagnostic agents to analyze B7-H1, B7-H3 and B7-H4 expression in the clinical setting while the expression pattern in the corresponding stage will be important to clarify tumor immune escape mechanism at different stages of colorectal cancer.