Antibodies are highly specific recognition molecules which are increasingly being applied to target therapy in patients. One type of developmental antibody-based therapy is antibody directed enzyme prodrug therapy (ADEPT) for the treatment of cancer. In ADEPT, an antibody specific to a tumor marker protein delivers a drug-activating enzyme to the cancer. Subsequent intravenous administration of an inactive prodrug results in drug activation and cytotoxicity only within the locale of the tumor. Pilot clinical trials with chemical conjugates of the prodrug activating enzyme carboxypeptidase G2 (CPG2) chemically conjugated with an antibody to and carcinoembryonic antigen (CEA), have shown that CPG2-mediated ADEPT is effective but limited by formation of human antibodies to CPG2 (HACA). We have developed a recombinant fusion protein (termed MFE-CP) of CPG2 with an anti-CEA single chain Fv antibody fragment and we have developed methods to address the immunogenicity of this therapeutic. A HACA-reactive discontinuous epitope on MFE-CP was identified using the crystal structure of CPG2, filamentous phage technology and surface enhanced laser desorption/ionization affinity mass spectrometry. This information was used to create a functional mutant of MFE-CP with a significant reduction (range 19.2 to 62.5%, median 38.5%) in reactivity with the sera of 11 patients with post-therapy HACA. The techniques described here are valuable tools for identifying and adapting undesirable immunogenic sites on protein therapeutics.

中文翻译：

---

映射和中和蛋白质治疗剂上的构象免疫原性位点的策略。

抗体是高度特异性的识别分子，其越来越多地应用于患者的靶向治疗。一种基于发展抗体的疗法是用于癌症治疗的抗体导向酶前药疗法（ADEPT）。在ADEPT中，特异性针对肿瘤标志物蛋白的抗体将药物激活酶传递给癌症。随后，非活性前药的静脉内给药仅在肿瘤区域内导致药物激活和细胞毒性。化学前体活化酶羧肽酶G2（CPG2）与抗癌胚抗原（CEA）抗体化学偶联的化学偶联物的临床试验表明，CPG2介导的ADEPT是有效的，但受人对CPG2（HACA）抗体形成的限制。我们已经开发了具有抗CEA单链Fv抗体片段的CPG2重组融合蛋白（称为MFE-CP），并且已经开发了解决该治疗剂免疫原性的方法。使用CPG2的晶体结构，丝状噬菌体技术和表面增强的激光解吸/电离亲和质谱法鉴定了MFE-CP上的HACA反应性不连续表位。该信息用于创建MFE-CP的功能性突变体，与11例治疗后HACA患者的血清反应性显着降低（范围从19.2至62.5％，中位数38.5％）。此处描述的技术是用于识别和适应蛋白质治疗剂中不良免疫原性位点的宝贵工具。