Establishment of novel monoclonal antibodies KMab-1 and MMab-1 specific for IDH2 mutations
Establishment of novel monoclonal antibodies KMab-1 and MMab-1 specific for IDH2 mutations
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DOI：
10.1016/j.bbrc.2013.01.088
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发表时间：
2013-03-01
影响因子：
3.1
通讯作者：
Kato, Yukinari
中科院分区：
文献类型：
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作者：
Kaneko, Mika Kato;Morita, Shunpei;Kato, Yukinari
已在神经胶质瘤、软骨肿瘤和白血病中检测到异柠檬酸脱氢酶 1/2 (IDH1/2) 突变。 IDH1/2 突变是早期且频繁的遗传改变，特定于 IDH1 中保守且功能重要的精氨酸 132 (R132) 和 IDH2 中的精氨酸 172 (R172) 中的单个密码子。我们之前建立了几种针对 IDH1 突变的单克隆抗体 (mAb)：针对 IDH1-R132H 的克隆 IMab-1 或 HMab-1 或针对 IDH1-R132S 的克隆 SMab-1。然而，针对 IDH2 突变的特异性单克隆抗体尚未见报道。为了建立 IDH2 突变特异性 mAb，我们用含有每种突变的 IDH2 肽（包括 IDH2-R172K 和 IDH2-R172M）对小鼠或大鼠进行免疫。细胞融合后，通过酶联免疫吸附测定 (ELISA) 筛选 IDH2 突变特异性单克隆抗体。在 ELISA 中，已建立的 mAb KMab-1 和 MMab-1 分别与 IDH2-R172K 和 IDH2-R172M 肽发生反应，但不与 IDH2 野生型 (WT) 发生反应。 Western-blot分析还显示KMab-1和MMab-1分别与IDH2-R172K和IDH2-R172M重组蛋白反应，而不与IDH2-WT或其他IDH2突变体反应，表明KMab-1和MMab-1具有IDH2突变特异性。此外，MMab-1在免疫细胞化学中特异性地对表达IDH2-R172M的细胞进行染色，但不对IDH2-WT和其他含有IDH2突变的细胞进行染色。在免疫组织化学分析中，MMab-1 对表达 IDH2-R172M 的神经胶质瘤进行特异性染色。这是第一份建立抗 IDH2 突变特异性单克隆抗体的报告，可用于诊断携带突变的肿瘤。 (C) 2013 Elsevier Inc. 保留所有权利。
Isocitrate dehydrogenase 1/2 (IDH1/2) mutations have been detected in gliomas, cartilaginous tumors, and leukemias. IDH1/2 mutations are early and frequent genetic alterations, are specific to a single codon in the conserved and functionally important Arginine 132 (R132) in IDH1 and Arginine 172 (R172) in IDH2. We previously established several monoclonal antibodies (mAbs), which are specific for IDH1 mutations: clones IMab-1 or HMab-1 against IDH1-R132H or clone SMab-1 against IDH1-R132S. However, specific mAbs against IDH2 mutations have not been reported. To establish IDH2-mutation-specific mAbs, we immunized mice or rats with each mutation-containing IDH2 peptides including IDH2-R172K and IDH2-R172M. After cell fusion, IDH2 mutation-specific mAbs were screened in Enzyme-Linked Immunosorbent Assay (ELISA). Established mAbs KMab-1 and MMab-1 reacted with the IDH2-R172K and IDH2-R172M peptides, respectively, but not with IDH2-wild type (WT) in ELISA. Western-blot analysis also showed that KMab-1 and MMab-1 reacted with the IDH2-R172K and IDH2-R172M recombinant proteins, respectively, not with IDH2-WT or other IDH2 mutants, indicating that KMab-1 and MMab-1 are IDH2-mutation-specific. Furthermore, MMab-1 specifically stained the IDH2-R172M-expressing cells in immunocytochemistry, but did not stain IDH2-WT and other IDH2-mutation-containing cells. In immunohistochemical analysis, MMab-1 specifically stained IDH2-R172M-expressing glioma. This is the first report to establish anti-IDH2-mutation-specific mAbs, which could be useful in diagnosis of mutation-bearing tumors. (C) 2013 Elsevier Inc. All rights reserved.