Molecular basis of antibody binding to mucin glycopeptides in lung cancer.
Molecular basis of antibody binding to mucin glycopeptides in lung cancer.
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肺癌中抗体与粘蛋白糖肽结合的分子基础
DOI：
10.3892/ijo.2015.3302
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发表时间：
2016-02
影响因子：
5.2
通讯作者：
Zhou D
中科院分区：
文献类型：
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作者：
Qu J;Yu H;Li F;Zhang C;Trad A;Brooks C;Zhang B;Gong T;Guo Z;Li Y;Ragupathi G;Lou Y;Hwu P;Huang W;Zhou D
携带Tn表位的糖肽是癌症诊断和免疫治疗的新兴靶点。在这项研究中，我们利用公共领域的基因微阵列数据分析了不同类型和分期肺癌患者中含有o -糖基化串联重复序列（TR）序列的膜蛋白。用单克隆IgG抗体14A、16A和B72.3研究了Tn和糖肽表位在肺癌细胞株表面的表达。用表面等离子体共振研究了单克隆抗体与合成糖肽的结合。在肺癌患者中发现了9种粘蛋白mrna的表达，但与健康个体的表达水平相似。在蛋白水平上，与单肽表位（14A）或单糖表位（B72.3）相比，癌细胞表面的糖肽表位更容易被mAb 16A识别。14A和16A更倾向于含有3个以上TR序列的簇状TR，其Kd值比连续两个TR低10倍。B72.3更倾向于识别MUC1上显示的簇状sialyl-Tn，而不是其他o糖蛋白，当MUC1作为糖载体转染时，其结合强度增加100倍，而总糖表位保持不变。这些发现表明，TR骨架和糖的簇对于单抗与粘蛋白糖肽的结合是必不可少的。本文提出了粘蛋白糖肽在分子水平上与抗体结合的三个规律：首先，糖肽的肽主链通过B细胞受体的互补决定区（cdr）突变被B细胞优先识别，重链框架结构突变获得糖结合特异性；其次，多肽和糖肽的连续串联重复序列（TR）更容易被B细胞识别，而B细胞更倾向于包含3个以上TR序列的集群TR；第三，某些糖特异性B细胞识别并容纳粘蛋白表面显示的簇状Tn和sialyl-Tn，而不是其他膜蛋白。
Glycopeptides bearing Tn epitopes are emerging targets for cancer diagnosis and immunotherapy. In this study, we analyzed membrane proteins containing O-glycosylated tandem repeat (TR) sequences in lung cancer patients of different types and stages, using gene microarray data in public domain. The expression of Tn and glycopeptide epitopes on the surface of lung cancer cell lines were studied by monoclonal IgG antibodies 14A, 16A, and B72.3. The binding of mAbs to synthetic glycopeptides were studied by surface plasmon resonance. Nine mucin mRNAs were found to be expressed in lung cancer patients but at similar level to healthy individuals. At protein level, a glycopeptide epitope on cancer cell surface is preferably recognized by mAb 16A, as compared to peptide-alone (14A) or sugar-alone epitopes (B72.3). 14A and 16A favor clustered TR containing more than three TR sequences, with 10-fold lower Kd than two consecutive TR. B72.3 preferrably recognized clustered sialyl-Tn displayed on MUC1 but not other O-glycoproteins, with 100-fold stronger binding when MUC1 is transfected as a sugar carrier, while the total sugar epitopes remain unchanged. These findings indicate that clusters of both TR backbones and sugars are essential for mAb binding to mucin glycopeptides. Three rules of antibody binding to mucin glycopeptides at molecular level are presented here: first, the peptide backbone of a glycopeptide is preferentially recognized by B cells through mutations in complementarity determining regions (CDRs) of B cell receptor, and the sugar-binding specificity is acquired through mutations in frame work of heavy chain; secondly, consecutive tandem repeats (TR) of peptides and glycopeptides are preferentially recognized by B cells, which favor clustered TR containing more than three TR sequences; thirdly, certain sugar-specific B cells recognize and accommodate clustered Tn and sialyl-Tn displayed on the surface of a mucin but not other membrane proteins.