■肿瘤内B细胞介导过多的免疫效应机制,在抗肿瘤免疫中起关键作用,并在多种实体瘤类型中充当积极的预后指标,包括上皮性卵巢癌(EOC)。肿瘤内B细胞的几个方面仍不清楚。比如他们的激活状态,抗原库,和成熟成浆细胞的能力。
■B淋巴细胞从原发性EOC组织和恶性腹水中分离并维持在细胞培养基中。用流式细胞术和B细胞受体测序对稳定维持的细胞系进行剖析。分泌的抗体用包含超过21,000种蛋白质的人类蛋白质组阵列进行测试。然后用ELISA进行验证。起始肿瘤样品用于芯片细胞计数的空间分析。
■从四个不同的EOC患者的卵巢肿瘤微环境(TME)中分离出分泌抗体的B淋巴细胞。高度克隆的细胞群体在体外经历了自发永生化,稳定地维持在抗体分泌状态,并显示存在爱泼斯坦-巴尔病毒(EBV)蛋白。所有来源的肿瘤都有高频率的肿瘤浸润B细胞,作为淋巴聚集体存在,或三级淋巴结构。四个细胞系中的三个所识别的抗原是含有蛋白155(CCDC155)的卷曲-卷曲结构域,生长因子受体结合蛋白2(GRB2),和丙酮酸脱氢酶磷酸酶2(PDP2),分别。在20例EOC患者中的9例(45%)血清中检测到了抗CCDC155循环IgG抗体。多参数芯片细胞计数的组织分析表明,这些新型人B细胞系分泌的抗体将其同源抗原结合在肿瘤细胞上。
这些研究表明,在EOC的肿瘤浸润淋巴细胞群体中,存在自然暴露于EBV的低频率抗体分泌B细胞群体。一旦稳定保持,这些新的细胞系为肿瘤内B细胞生物学和新的靶抗原识别的未来研究提供了独特的机会,并用于非EBV相关实体瘤如EOC的TME中EBV潜伏期和/或病毒再激活的研究。
UNASSIGNED: Intra-tumoral B cells mediate a plethora of immune effector mechanisms with key roles in anti-tumor immunity and serve as positive prognostic indicators in a variety of solid tumor types, including epithelial ovarian cancer (EOC). Several aspects of intra-tumoral B cells remain unclear, such as their state of activation, antigenic repertoires, and capacity to mature into plasma cells.
UNASSIGNED: B lymphocytes were isolated from primary EOC tissue and malignant ascites and were maintained in cell culture medium. The stably maintained cell lines were profiled with flow cytometry and B cell receptor sequencing. Secreted antibodies were tested with a human proteome array comprising more than 21,000 proteins, followed by ELISA for validation. Originating tumor samples were used for spatial profiling with chip cytometry.
UNASSIGNED: Antibody-secreting B lymphocytes were isolated from the ovarian tumor microenvironment (TME) of four different EOC patients. The highly clonal cell populations underwent spontaneous immortalization in vitro, were stably maintained in an antibody-secreting state, and showed presence of Epstein-Barr viral (EBV) proteins. All originating tumors had high frequency of tumor-infiltrating B cells, present as lymphoid aggregates, or tertiary lymphoid structures. The antigens recognized by three of the four cell lines are coil-coil domain containing protein 155 (CCDC155), growth factor receptor-bound protein 2 (GRB2), and pyruvate dehydrogenase phosphatase2 (PDP2), respectively. Anti-CCDC155 circulating IgG antibodies were detected in 9 of 20 (45%) of EOC patients\' sera. Tissue analyses with multiparameter chip cytometry shows that the antibodies secreted by these novel human B cell lines engage their cognate antigens on tumor cells.
UNASSIGNED: These studies demonstrate that within the tumor-infiltrating lymphocyte population in EOC resides a low frequency population of antibody-secreting B cells that have been naturally exposed to EBV. Once stably maintained, these novel cell lines offer unique opportunities for future studies on intratumor B cell biology and new target antigen recognition, and for studies on EBV latency and/or viral reactivation in the TME of non-EBV related solid tumors such as the EOC.