Anti-human HB-EGF monoclonal antibodies inhibiting ectodomain shedding of HB-EGF and diphtheria toxin binding
Anti-human HB-EGF monoclonal antibodies inhibiting ectodomain shedding of HB-EGF and diphtheria toxin binding
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DOI：
10.1093/jb/mvq033
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发表时间：
2010-07-01
影响因子：
2.7
通讯作者：
Mekada, Eisuke
中科院分区：
文献类型：
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作者：
Hamaoka, Miki;Chinen, Ichino;Mekada, Eisuke
HB-EGF是EGF家族中的一员，能结合并激活EGF受体。HB-EGF作为膜锚定蛋白（proHB-EGF）合成，然后蛋白水解切割，产生促有丝分裂活性的可溶形式。ProHB-EGF作为白喉毒素（DT）的受体发挥作用。HB-EGF在包括癌症在内的病理生理过程中起着关键作用。HB-EGF的特异性单克隆抗体（mAb）是研究HB-EGF的重要工具。然而，迄今为止，很少有这样的mAb被建立。在这项研究中，我们新产生的7个克隆的杂交瘤衍生的单克隆抗体免疫HB-EGF裸小鼠与重组人HB-EGF蛋白。所有mAb均特异性结合人HB-EGF，但不结合小鼠HB-EGF。表位作图分析表明，大多数单克隆抗体识别EGF样结构域。虽然没有一个新分离的单克隆抗体直接抑制HB-EGF对EGFR表达细胞的促有丝分裂活性，但有些抗体强烈抑制DT结合。有趣的是，一些mAb有效地抑制proHB-EGF的胞外结构域脱落，并因此阻止EGFR表达细胞在与proHB-EGF表达细胞共培养系统中的细胞生长。因此，这些新的抗HB-EGF单克隆抗体可能会推进HB-EGF的临床和基础研究。
HB-EGF is a member of the EGF family of growth factors that bind and activate the EGF receptor. HB-EGF is synthesized as a membrane-anchored protein (proHB-EGF), and then proteolytically cleaved, resulting in the mitogenically active soluble form. ProHB-EGF functions as the receptor for the diphtheria toxin (DT). HB-EGF plays pivotal roles in pathophysiological processes, including cancer. Monoclonal antibodies (mAbs) specific for HB-EGF could be an important tool in HB-EGF research. However, few such mAbs have been established to date. In this study, we newly generated seven clones of hybridoma-derived mAbs by immunizing HB-EGF null mice with recombinant human HB-EGF protein. All mAbs specifically bound to human HB-EGF but not to mouse HB-EGF. Epitope mapping analysis showed that most of the mAbs recognized the EGF-like domain. Although none of the newly isolated mAbs directly inhibited the mitogenic activity of HB-EGF for EGFR-expressing cells, some strongly inhibited DT-binding. Interestingly, some of the mAbs efficiently inhibited ectodomain shedding of proHB-EGF, and consequently prevented the cell growth of the EGFR-expressing cells in a co-culture system with proHB-EGF-expressing cells. Hence, these new anti-HB-EGF mAbs may advance clinical as well as basic research on HB-EGF.