Single-Chain Antibody-Based Immunotoxins Targeting Her2/neu: Design Optimization and Impact of Affinity on Antitumor Efficacy and Off-Target Toxicity
Single-Chain Antibody-Based Immunotoxins Targeting Her2/neu: Design Optimization and Impact of Affinity on Antitumor Efficacy and Off-Target Toxicity
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DOI：
10.1158/1535-7163.mct-11-0519
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发表时间：
2012-01-01
影响因子：
5.7
通讯作者：
Rosenblum, Michael G.
中科院分区：
文献类型：
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作者：
Cao, Yu;Marks, James D.;Rosenblum, Michael G.
由单链可变区片段（scFv）与多肽毒素基因融合组成的重组免疫毒素代表了用于癌症治疗的潜在有效候选物。我们评估了与重组白树素（rGel）融合的各种抗Her 2/neu scFv的亲和力及其对抗肿瘤功效和脱靶毒性的影响。从人抗Her 2/neu scFv C6.5和各种亲和力突变体（命名为ML 3 -9、MH 3-B1和B1 D3）产生一系列基于rGel的免疫毒素，其亲和力范围为10（-8）至10（-11）mol/L。针对Her 2/neu过表达的肿瘤细胞，具有增加的亲和力的免疫毒素显示出改善的内化和增强的自噬细胞毒性。最高亲和力B1 D3/rGel构建体的靶向指数最高。然而，由于免疫复合物的形成，游离Her 2/neu胞外结构域（ECD）的添加显著降低了B1 D3/rGel的细胞毒性。相比之下，ECD添加对体外较低亲和力构建体几乎没有影响。针对已建立的BT474 M1异种移植物的体内研究显示，所有免疫毒素均抑制生长。令人惊讶的是，B1 D3-rGel治疗诱导了显著的肝毒性，这是因为在循环中与脱落的Her 2/neu抗原形成免疫复合物。具有中等亲和力的MH 3-B1/rGel构建体显示出有效的肿瘤生长抑制，而不诱导肝毒性或复合物形成。这些发现表明，虽然高亲和力构建体可以是有效的抗肿瘤剂，但它们也可能通过与可溶性抗原容易形成复合物而导致显著的脱靶毒性而与误靶相关。由中等亲和力抗体组成的构建体也是对免疫复合物形成更具抗性的有效试剂。因此，在评估靶向治疗剂的设计和功效时，亲和力是一个非常重要的考虑因素。Mol Cancer Ther; 11（1）; 143-53. (C)2011年AACR。
Recombinant immunotoxins, consisting of single-chain variable fragments (scFv) genetically fused to polypeptide toxins, represent potentially effective candidates for cancer therapeutics. We evaluated the affinity of various anti-Her2/neu scFv fused to recombinant gelonin (rGel) and its effect on antitumor efficacy and off-target toxicity. A series of rGel-based immunotoxins were created from the human anti-Her2/neu scFv C6.5 and various affinity mutants (designated ML3-9, MH3-B1, and B1D3) with affinities ranging from 10(-8) to 10(-11) mol/L. Against Her2/neu-overexpressing tumor cells, immunotoxins with increasing affinity displayed improved internalization and enhanced autophagic cytotoxicity. Targeting indices were highest for the highest affinity B1D3/rGel construct. However, the addition of free Her2/neu extracellular domain (ECD) significantly reduced the cytotoxicity of B1D3/rGel because of immune complex formation. In contrast, ECD addition had little impact on the lower affinity constructs in vitro. In vivo studies against established BT474M1 xenografts showed growth suppression by all immunotoxins. Surprisingly, therapy with the B1D3-rGel induced significant liver toxicity because of immune complex formation with shed Her2/neu antigen in circulation. The MH3-B1/rGel construct with intermediate affinity showed effective tumor growth inhibition without inducing hepatotoxicity or complex formation. These findings show that while high-affinity constructs can be potent antitumor agents, they may also be associated with mistargeting through the facile formation of complexes with soluble antigen leading to significant off-target toxicity. Constructs composed of intermediate-affinity antibodies are also potent agents that are more resistant to immune complex formation. Therefore, affinity is an exceptionally important consideration when evaluating the design and efficacy of targeted therapeutics. Mol Cancer Ther; 11(1); 143-53. (C) 2011 AACR.