Age related ubiquitin defect on T-cell response to tumor

描述(申请人提供)：这项修订提案的总体目标是确定一种新的包含F-box和WD40结构域的乳腺肿瘤抗原的生物学效应，重点是它在年龄和T细胞依赖的肿瘤生长易感性中的作用。我们已经确定了一种新的人类乳腺肿瘤抗原(HBTA1)，它可以强烈抑制老年小鼠的T细胞增殖，但不是年轻小鼠。用HBTA1免疫2个月龄的BXD 12小鼠可保护16个月龄的小鼠免受肿瘤的攻击。蔗糖梯度纯化小鼠同系物HBTA1(MHBTA1)，然后对纯化的样品进行电子显微镜检查，结果表明mHBTA1包装在TS/A肿瘤的外体中。TS/A外体mHBTA1能强烈抑制老年BXD12小鼠T细胞的增殖，但不能抑制年轻小鼠T细胞的增殖。T细胞在TS/A外切体存在时的增殖能力与外切体mHBTA1的泛素化状态有关。我们的目标是(1)确定mHBTA1泛素化是否与肿瘤生长易感性和肿瘤特异性细胞毒T细胞反应相关。将泛素化外体mHBTA1免疫老龄小鼠，以确定其增强对肿瘤生长的保护作用。同时，我们将确定TS/A外体中内源性泛素化mHBTA1的产生是否与肿瘤易感性有关。(2)利用siRNA技术确定敲除肿瘤细胞外体mHBTA1是否足以逆转TS/A外体对T细胞增殖的抑制，或是否需要其他外体蛋白。(3)为了确定我们最近发现的年龄依赖的T细胞因子是否在外体mHBTA1的泛素化中发挥作用，从而使mHBTA1介导的抑制T细胞激活的失活；我们将进一步确定TS/A外体mHBTA1与T细胞因子相互作用的途径。(4)确定年龄相关T细胞因子(S)是否调节人乳腺肿瘤细胞外体HBTA1的泛素化，并进一步确定其与年龄相关乳腺癌发病的可能联系。产生的数据应该阐明肿瘤外体和外体mHBTA1在年龄和T细胞依赖型乳腺癌发展中所起作用的细胞和分子基础。

DESCRIPTION (provided by applicant): The overall goal of this revised proposal is to determine the biologic effects of a novel breast tumor antigen containing F-box and WD40 domains, with an emphasis on its roles in age- and T cell-dependent susceptibility to tumor growth. We have identified a novel human breast tumor antigen (HBTA1) that strongly inhibits proliferation of T cells from aged, but not young, mice. Immunization of 2-mo-old BXD 12 mice with HBTA1 protects against the tumor challenge at 16-mo-of-age. Sucrose gradient purification of a mouse homologue HBTA1 (mHBTA1) followed by electromicroscope examination of the purified samples led to the realization that mHBTA1 is packed in the exosomes of the TS/A tumors. The TS/A exosomal mHBTA1 strongly inhibits the proliferation of T cells isolated from aged, but not young, BXD12 mice. The ability of T cells to proliferate in the presence of TS/A exosomes is correlated with the ubiquitination status of exosomal mHBTA1. Our aims are (1) To determine if ubiquitination of mHBTA1 is correlated with susceptibility to tumor growth and the tumor-specific cytotoxic T-cell response. Aged-mice will be immunized with ubiquitinated exosomal mHBTA1 to determine its effects on enhancement of protection against the tumor growth. In parallel, we will determine if the production of endogenous ubiquitinated mHBTA1 in TS/A exosomes is correlated with the tumor susceptibility. (2) To use siRNA technology to determine if knockout of exosomal mHBTA1 of tumor cells is sufficient to reverse the inhibition of T cell proliferation mediated by TS/A exosomes or whether other exosomal proteins are required. (3) To determine if the age-dependent T cell factors we recently identified play a role in ubiquitination of exosomal mHBTA1, and thus inactivation of mHBTA1-mediated inhibition of T-cell activation; we will further determine the pathway by which TS/A exosomal mHBTA1 interacts with T-cell factors. (4) To determine if an age-related T-cell factor(s) regulates the ubiquitination of exosomal HBTA1 of human breast tumors, and, further to determine its possible association with the age-related incidence of breast cancers. The data generated should elucidate the cellular and molecular basis for the role of tumor exosomes and exosomal mHBTA1 in the development of age- and T cell-dependent breast cancer.