Malignant mesothelioma associated antigens recognized by tumor infiltrating B cells. -usefulness of antibody titer to the antigens -

恶性胸膜间皮瘤早期诊断困难，尚未建立普遍接受的治疗方法。本研究试图通过异种移植SCID小鼠模型获得恶性间皮瘤肿瘤浸润B淋巴细胞（TIB）衍生的肿瘤特异性抗体，并通过重组表达克隆法（SEREX）将TIB产生的IgG用于抗原的血清学鉴定。我们尝试从5例恶性间皮瘤患者的手术标本中建立肿瘤细胞系，并将这些间皮瘤组织植入SCID小鼠皮下，异种移植后每2周采集一次血液样本。建立了2个间皮瘤细胞系（K921MS0和L324MSO）。从恶性间皮瘤细胞系K921MS0构建cDNA文库。使用移植间皮瘤组织的SCID小鼠血清对文库进行免疫筛选。对免疫反应性cDNA克隆进行亚克隆，并利用数据库进行同源性分析。分析了恶性间皮瘤患者血清中对所鉴定抗原的抗体滴度。免疫组化染色显示肿瘤组织中有B淋巴细胞浸润。在所有SCID小鼠血清中均检测到人IgG抗体（134 ~ 24000μg/ml）。利用来自TIB的抗体，鉴定出11种抗原。6种抗原在恶性间皮瘤中呈过表达。在SCID小鼠血清和间皮瘤患者血清中均能检测到6种抗原的抗体。其中，12例间皮瘤患者中有7例检测到Gene-X抗体，11例间皮瘤患者检测到Thrombospondin-2抗体。我们的研究结果表明浸润在肿瘤组织中的B细胞已被致敏并产生针对肿瘤相关抗原的抗体。在SEREX定义的抗原中，2种抗原（Gene-X和Thrombosponddin-2）在恶性间皮瘤中过表达，间皮瘤患者中检测到抗体的比例高于正常健康人。这两种抗原可作为恶性间皮瘤患者的肿瘤标志物。少

Malignant pleural mesothelioma is difficult to be diagnosed at early stage and universally accepted therapy has not been established. In this study, we tried to obtain tumor specific antibody derived from tumor infiltrated B lymphocytes (TIB) in malignant mesothelioma by using xenotransplanted SCID mice model, and applied TIB-produced IgG to serological identification of antigens by recombinant expression cloning method (SEREX).Surgical specimens from 5 patients with malignant mesothelioma have been tried to establish tumor cell lines, and also these mesothelioma tissues were engrafted subcutaneously in SCID mice and blood samples were obtained every 2 weeks after xenotransplantation. Two mesothelioma cell lines have been established (K921MS0 and L324MSO). cDNA library was constructed from a tumor cell line (K921MS0) of malignant mesothelioma. Immunoscreening of the library was performed using the serum from SCID mice transplanted with mesothelioma tissues. Immuno-reactive cDNA clones … More were subcloned, and analyzed using database for homology search. Antibodies titers against the identified antigens were analyzed in sera of patients with malignant mesothelioma.Immunohistochemical staining revealed that B lymphocytes infiltrated in tumor tissues. In all SCID mouse serum, human IgG antibody could be detected (ranged from 134 to 24000μg/ml). By using antibodies derived from TIB, 11 antigens were identified. Six antigens showed over expression for malignant mesothelioma. Antibodies against the 6 antigens could be detected not only in serum of SCID mouse but also in sera of mesothelioma patients. Among them, antibody against Gene-X was detected in 7 out of 12 mesothelioma patients, and antibody against Thrombospondin-2 was detected in 11 out of 12 mesothelioma patients.Our results suggest that the B cells infiltrated in the tumor tissues have been sensitized and produced antibody against tumor associate antigens. Among the SEREX defined antigens, 2 antigens (Gene-X and Thrombosponddin-2) were overexpressed in malignant mesothelioma, and antibodies against them could be detected in higher proportion of mesothelioma patients than normal healthy persons. These two antigens could be used as tumor markers for diagnosis of patients with malignant mesothelioma. Less