Abstract

The recombinant proteins E6-CBD and E7-CBD, which are antigens E6 and E7 of human papillomavirus type 16 (HPV16), were connected by a glycine-serine spacer with the cellulose-binding domain (CBD) of Anaerocellum thermophilum in preparations obtained via synthesis in a heterologous expression system in Escherichia coli. A simple, effective, one-step technology for the isolation and purification of recombinant proteins E6-CBD and E7-CBD has been developed. The presence of antigenic properties in the obtained protein preparations was confirmed via Western blot. Three variants of immunogenic compositions with the recombinant proteins E6-CBD and E7-CBD and a three-dose immunization scheme have been developed. The immunogenicity was assessed in the BALB/c mice, and the antitumor effect of immunogenic compositions was studied on an experimental model of BALB/c mice with HPV16-associated tumors. It is shown that E6-CBD and E7-CBD are highly immunogenic and have antitumor effects in vivo. Both proteins, E6-CBD and E7-CBD, must be present to achieve the highest efficiency as part of the immunogenic composition. The obtained HPV16 recombinant antigens E6-CBD and E7-CBD can be used in two ways: (a) in clinical practice as the main specific components of therapeutic vaccines for the treatment of HPV16-associated neoplastic lesions of various localizations (cervical cancer, carcinoma of the anogenital tract, oropharynx) and (b) in diagnostics for the development of diagnostic kits to detect antibodies to E6 and E7 oncoproteins in patients with HPV16-associated malignant neoplasms.

中文翻译：

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人乳头瘤病毒 16 型的重组抗原 E6 和 E7：制备、纯化和体内免疫原性和抗肿瘤作用的评估

摘要

重组蛋白 E6-CBD 和 E7-CBD 是 16 型人乳头瘤病毒 (HPV16) 的抗原 E6 和 E7，通过甘氨酸-丝氨酸间隔区与嗜热厌氧菌的纤维素结合结构域 (CBD) 连接，通过大肠杆菌异源表达系统的合成. 已经开发出一种用于分离和纯化重组蛋白 E6-CBD 和 E7-CBD 的简单、有效的一步技术。通过蛋白质印迹证实所获得的蛋白质制剂中存在抗原特性。已经开发了具有重组蛋白 E6-CBD 和 E7-CBD 的三种免疫原性组合物变体以及三剂量免疫方案。在 BALB/c 小鼠中评估免疫原性，并在具有 HPV16 相关肿瘤的 BALB/c 小鼠的实验模型上研究免疫原性组合物的抗肿瘤作用。表明E6-CBD和E7-CBD具有高度免疫原性并在体内具有抗肿瘤作用。作为免疫原性组合物的一部分，E6-CBD 和 E7-CBD 两种蛋白质都必须存在才能实现最高效率。a ) 在临床实践中作为治疗性疫苗的主要特定成分，用于治疗各种部位的 HPV16 相关肿瘤病变（宫颈癌、肛门生殖道癌、口咽部）和 ( b ) 在诊断中用于开发诊断试剂盒检测 HPV16 相关恶性肿瘤患者的 E6 和 E7 癌蛋白抗体。