Identification of the epitope for the epidermal growth factor receptor-specific monoclonal antibody 806 reveals that it preferentially recognizes an untethered form of the receptor
Identification of the epitope for the epidermal growth factor receptor-specific monoclonal antibody 806 reveals that it preferentially recognizes an untethered form of the receptor
复制标题
DOI：
10.1074/jbc.m401218200
复制
发表时间：
2004-07-16
影响因子：
4.8
通讯作者：
Scott, AM
中科院分区：
文献类型：
--
作者：
Johns, TG;Adamas, TE;Scott, AM
表皮生长因子受体（EGFR）在许多上皮癌中过度表达，这一观察结果通常与不良临床结局相关。EGFR的过表达通常由EGFR基因扩增引起，并且有时与在其细胞外结构域中具有内部缺失的变体EGFR（de 2 -7 EGFR或EGFRvIII）的表达相关。单克隆抗体（mAb）806是一种新型EGFR抗体，具有显著的抗肿瘤活性，在过表达时可识别de 2 -7 EGFR和野生型（wt）EGFR的一个子集，但不结合正常组织中表达的wt EGFR。尽管mAb 806仅与A431肿瘤细胞中表达的低比例的wt EGFR结合（接近10%），但mAb 806对裸鼠中生长的A431异种移植物显示出稳健的抗肿瘤活性。为了阐明导致其独特的特异性和抗肿瘤活性模式的机制，我们确定了mAb 806的EGFR结合表位。对mAb 806与酵母表面或免疫印迹形式表达的EGFR片段结合的分析鉴定了含有mAb 806表位的二硫键结合环（氨基酸287-302）。实际上，mAb 806以表观高亲和力（类似于30 nM）结合对应于这些氨基酸的合成EGFR肽。EGFR结构的分析表明，表位仅在受体的过渡形式中完全暴露，所述受体的过渡形式是因为EGFR从无活性的束缚构象变为配体结合的活性形式而发生的。似乎mAb 806结合了这一小部分瞬时受体，阻止了它们的激活，这反过来又产生了强烈的抗肿瘤作用。最后，我们的观察表明，产生的抗体过渡形式的生长因子受体可能代表了一种新的方式，减少正常组织的目标，但保留抗肿瘤活性。
The epidermal growth factor receptor (EGFR) is over-expressed in many epithelial cancers, an observation often correlated with poor clinical outcome. Overexpression of the EGFR is commonly caused by EGFR gene amplification and is sometimes associated with expression of a variant EGFR (de2-7 EGFR or EGFRvIII) bearing an internal deletion in its extracellular domain. Monoclonal antibody (mAb) 806 is a novel EGFR antibody with significant antitumor activity that recognizes both the de2-7 EGFR and a subset of the wild type (wt) EGFR when overexpressed but does not bind the wt EGFR expressed in normal tissues. Despite only binding to a low proportion of the wt EGFR expressed in A431 tumor cells (similar to10%), mAb 806 displays robust antitumor activity against A431 xenografts grown in nude mice. To elucidate the mechanism leading to its unique specificity and mode of antitumor activity, we have determined the EGFR binding epitope of mAb 806. Analysis of mAb 806 binding to EGFR fragments expressed either on the surface of yeast or in an immunoblot format identified a disulfide-bonded loop (amino acids 287-302) that contains the mAb 806 epitope. Indeed, mAb 806 binds with apparent high affinity (similar to30 nM) to a synthetic EGFR peptide corresponding to these amino acids. Analysis of EGFR structures indicates that the epitope is fully exposed only in the transitional form of the receptor that occurs because EGFR changes from the inactive tethered conformation to a ligand-bound active form. It would seem that mAb 806 binds this small proportion of transient receptors, preventing their activation, which in turn generates a strong antitumor effect. Finally, our observations suggest that the generation of antibodies to transitional forms of growth factor receptors may represent a novel way of reducing normal tissue targeting yet retaining antitumor activity.