Targeting and therapy of carcinoembryonic antigen-expressing tumors in transgenic mice with an antibody-interleukin 2 fusion protein.
Targeting and therapy of carcinoembryonic antigen-expressing tumors in transgenic mice with an antibody-interleukin 2 fusion protein.
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DOI：
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发表时间：
2000-08
期刊：
影响因子：
11.2
通讯作者：
Xiaochuan Xu;P. Clarke;G. Szalai;J. Shively;L. Williams;Y. Shyr;Ergang Shi;F. Primus
中科院分区：
文献类型：
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作者：
Xiaochuan Xu;P. Clarke;G. Szalai;J. Shively;L. Williams;Y. Shyr;Ergang Shi;F. Primus
本研究的目的是设计一种双价单链抗癌胚抗原(CEA)抗体和一种白介素2(IL-2)融合蛋白衍生物，用于选择性靶向肿瘤细胞因子。利用高亲和力抗CEA抗体T84.66的可变区形成单基因编码抗体[单链可变区与结晶区Fc(ScFvFc)连接]。融合蛋白(scFvFc.IL-2)由小鼠IL-2与scFvFc的COOH末端融合而成。重组蛋白被组装成完整的分子，在抗原结合特性上与完整的抗CEA单抗相似。根据融合蛋白中IL-2的含量，其支持CTLL-2细胞增殖的能力与IL-2相同。尽管分子大小与完整的单抗相似，但融合蛋白的血液清除速度明显快于完整的单抗或scFvFc。用放射性标记的scFvFc.IL-2在小鼠血清中孵育，而不是完整的或scFvFc抗体在小鼠血清中孵育时伴随着复合体的出现，提示后者可能有助于加速融合蛋白的清除。研究了CEA转基因小鼠体内CEA阳性小鼠肿瘤和抗原阴性亲本肿瘤的生物分布和肿瘤靶向性。二价抗CEA scFvFc具有与完整单抗相似的肿瘤定位特性。虽然将IL-2融合到双价ScFvFc的COOH末端改变了其药代动力学性质，但融合抗体能够特异性地靶向肿瘤。CEA阳性肿瘤对完整单抗、scFvFc和scFvFc.IL-2的最大摄取分别为29.3+/-5.0、19.5+/-2.1和6.6+/-0.9%注射剂量/g。三种抗体的最大肿瘤定位比(CEA阳性/CEA阴性肿瘤)相似(4.6~6.0)，表明肿瘤靶向的抗原特异性。未观察到明显的抗原特异性靶向转基因小鼠CEA阳性的正常组织。尽管融合蛋白的肿瘤靶向性较低，但用抗CEA-IL-2抗体处理转基因小鼠后，CEA表达(P=0.01)而非抗原无关(P=0.22)的同基因肿瘤细胞的生长受到抑制。静脉注射后CEA表达肿瘤的治疗得到改善。给予融合蛋白(P=0.0001)。这些研究表明，抗CEA抗体导向的细胞因子靶向可能为CEA表达的肿瘤提供一种有效的治疗方法。具有免疫功能的CEA转基因小鼠模型的可获得性也将有助于确定这些融合蛋白的免疫治疗特性。
The purpose of this study was to engineer a bivalent single-chain anticarcinoembryonic antigen (CEA) antibody and an interleukin 2 (IL-2) fusion protein derivative for selective tumor targeting of cytokines. The variable domains of a high affinity anti-CEA antibody, T84.66, were used to form a single-gene-encoded antibody [single-chain variable fragment joined to the crystallizable fragment, Fc (scFvFc)]. The fusion protein (scFvFc.IL-2) consisted of mouse IL-2-fused to the COOH-terminal end of the scFvFc. The engineered proteins were assembled as complete molecules and were similar to the intact anti-CEA monoclonal antibody (Mab) in antigen-binding properties. Based on IL-2 content of the fusion protein, its ability to support proliferation of CTLL-2 cells was identical with that of IL-2. Despite a molecular size similar to that of the intact Mab, the blood clearance of the fusion protein was markedly faster than that of the intact Mab or scFvFc. Incubation of radiolabeled scFvFc.IL-2 but not the intact or scFvFc antibodies in mouse serum was accompanied by the appearance of complexes, suggesting that the latter may contribute to the accelerated clearance of the fusion protein. Biodistribution and tumor targeting studies were carried out in CEA-transgenic mice bearing CEA-positive murine tumors as well as the antigen-negative parental tumor. The bivalent anti-CEA scFvFc had tumor localization properties similar to those of the intact Mab. Although fusion of IL-2 to the COOH-terminal end of the bivalent scFvFc altered its pharmacokinetic properties, the fusion antibody was able to target tumors specifically. Maximum uptake of the intact Mab, scFvFc, and scFvFc.IL-2 in CEA-positive tumors was 29.3 +/- 5.0, 19.5 +/- 2.1, and 6.6 +/- 0.9% injected dose/g, respectively. Maximum tumor localization ratios (CEA-positive/CEA-negative tumor) were similar for all three antibody types (4.6-6.0), demonstrating the antigen specificity of the tumor targeting. Significant antigen-specific targeting to CEA-positive normal tissues of transgenic mice was not observed. Although the tumor-targeting properties of the fusion protein were low, the growth of CEA-expressing (P = 0.01) but not antigen-irrelevant (P = 0.22) syngeneic tumor cells was inhibited after treatment of transgenic mice with the anti-CEA-IL-2 antibody. Therapy of CEA-expressing tumors was improved after i.v. administration of the fusion protein (P = 0.0001). These studies indicate that anti-CEA antibody-directed cytokine targeting may offer an effective treatment for CEA-expressing carcinomas. The availability of an immunocompetent CEA transgenic mouse model will also help to determine the immunotherapeutic properties of these fusion proteins.