NEO ANTIGEN AND IMMUNOBIOLOGY OF MEDULLARY BREAST CANCER

这项拟议的研究试图识别和表征在乳腺髓样癌(MC)中引起浆细胞反应的新表达的蛋白质(新抗原)。我们的主要假设是，浆细胞和淋巴成分的显著渗透反映了局部对肿瘤的免疫反应，这是导致仅手术治疗后MIC肿瘤更有利的自然病程印象的原因。我们将应用分子克隆技术从肿瘤浸润性浆细胞中提取抗体，然后使用这些抗体来检索其在肿瘤中的新表达引起反应的蛋白质。我们从浆细胞浸润性MC肿瘤中制备了组合噬菌体Fab文库。我们证明了这些组织表达高度集中的免疫球蛋白抗体谱，支持了我们对肿瘤中表达的新抗原进行特异性免疫反应的前提。此外，肿瘤中主要的类转换免疫球蛋白同种类型和CDR高度突变表明T依赖的抗原，即蛋白质。研究还表明，新抗原既不是P53，也不是两个主要的乳腺癌相关抗原Her2/neu，这使得新抗原更有可能(但尚未证明)是一种以前未描述的蛋白质或一种以前未被认为与肿瘤相关的蛋白质。新抗原可能是存在于肿瘤中的外来(即病毒)蛋白或异常表达或突变的正常蛋白。这些重要的初步数据证明了克隆诱导性抗原的剩余努力是合理的。抗原将通过用患者抗体筛选肿瘤特异性表达文库来克隆，同时寻求一种平行的生化方法以确保克隆成功。然后对克隆和表达的新抗原(S)进行研究和评价，以探讨其在恶性增殖中的可能作用。

The proposed research seeks to identify and characterize newly expressed proteins (neo-antigens) that cause plasma cell reactions in medullary carcinomas (MC) of breast. It is our principal hypothesis that the marked infiltration of plasma cells and lymphoid elements reflects a local immune response against tumor that is responsible for the impression of a more favorable natural course of MIC tumors after surgery-only therapy. We will apply molecular cloning techniques to derive antibodies from tumor-infiltrating plasma cells, and then use these antibodies to retrieve the proteins whose new expression in the tumors caused the response. We prepared combinatorial phage Fab libraries from plasma cell-infiltrated MC tumors. We proved these tissues express highly focussed IgG antibody repertoires, supporting our premise of a specific immune reaction against a neo-antigen expressed in the tumor. Further, the dominant class-switched IgG isotype in the tumor and the high degree of CDR mutations suggest a T-dependent antigen, i.e., a protein. It was also shown that the neo-antigen is neither p53 nor Her2/neu, two dominant breast carcinoma-associated antigens, making it more likely (but not yet proving) that the neo-antigen is a previously undescribed protein or a protein not previously recognized to be tumor-associated. The neo-antigen could be a foreign (i.e., viral) protein or an aberrantly expressed or mutated normal protein that is present in the tumor. These important preliminary data justify the pursuit of the remaining effort to clone the eliciting antigen. Antigen will be cloned by screening of tumor-specific expression libraries with patient antibodies, while pursuing a parallel biochemical approach to ensure success in the cloning. The cloned and expressed neoantigen(s) is(are) then studied and assessed for possible roles in the malignant proliferation.