Identification of a small peptide that inhibits the phosphorylation of ErbB2 and proliferation of ErbB2 overexpressing breast cancer cells
Identification of a small peptide that inhibits the phosphorylation of ErbB2 and proliferation of ErbB2 overexpressing breast cancer cells
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DOI：
10.1002/ijc.20306
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发表时间：
2004-10-10
影响因子：
6.4
通讯作者：
Krag, DN
中科院分区：
文献类型：
--
作者：
Pero, SC;Shukla, GS;Krag, DN
ErbB 2在大约30%的乳腺癌患者中过表达，与预后不良相关。ErbB 2已被鉴定为用于分子靶向的有用受体。利用fUSE 5基因III系统构建了一个20个氨基酸的噬菌体随机肽库。针对ErbB 2外部结构域的2种不同纯化形式淘选文库。这导致在几个ErbB 2结合噬菌体克隆与不同的ErbB 2制剂的可变结合的鉴定。一个克隆（EC-1）结合ErbB 2的所有制备物，包括活细胞和新鲜冷冻的人乳腺癌标本。基于EC-1克隆的推导序列的合成肽及其生物素缀合形式保留了对纯化的ErbB 2和ErbB 2过表达细胞裂解物的结合亲和力。EC-1肽能有效地抑制ErbB 2在Y1248和Y877残基上的磷酸化，并呈剂量和时间依赖性。此外，EC-1肽选择性抑制ErbB 2过表达乳腺癌细胞的增殖。环状EC-1肽的线性部分被证明是必不可少的结合ErbB 2。此外，构建了4个偏向性噬菌体文库，允许EC-1肽的4个不同区域具有随机序列。筛选这些EC-1偏向的文库没有产生更高亲和力的肽，但确实证明了N-末端侧翼臂上位置1-4处的氨基酸和环状环内位置11 - 15处的氨基酸的重要性。有趣的是，EC-1含有与已知ErbB受体家族配体同源的基序。我们已经鉴定了一种小肽，其结合ErbB 2的胞外结构域，抑制ErbB 2自磷酸化并抑制ErbB 2过表达细胞的增殖。这支持了这样的观点，即小肽可以结合到癌症治疗中重要的靶点，即使靶点没有天然配体。对该肽的持续研究包括增加其对ErbB 2的亲和力、评价药代动力学和评价结合抗癌剂的抗增殖作用。(C)2004 Wiley-Liss，Inc.
ErbB2 is overexpressed in approximately 30% of breast cancer patients with a correlation to poor prognosis. ErbB2 has been identified as a useful receptor for molecular targeting. A cyclic 20 amino acid phage display random peptide library was constructed using the fUSE5 gene I I I system. The library was panned against 2 different purified forms of the external domain of ErbB2. This resulted in the identification of several ErbB2-binding phage clones with variable binding to different ErbB2 preparations. One clone (EC-1) bound all preparations of ErbB2 including live cells and fresh frozen human breast cancer specimens. The synthetic peptide based on the deduced sequence of the EC-1 clone and its biotin-conjugated form retained binding affinity for purified ErbB2 and ErbB2 overexpressing cell lysates. EC-1 peptide was able to effectively inhibit the phosphorylation of ErbB2 on residues Y1248 and Y877 in a dose- and time-dependent manner. Furthermore, EC-1 peptide selectively inhibits the proliferation of ErbB2 overexpressing breast cancer cells. The linear portion of the cyclic EC-1 pepticle was shown to be essential for binding ErbB2. In addition, 4 biased phage libraries were constructed allowing 4 different regions of the EC-1 pepticle to have random sequence. Screening these EC-1 biased libraries did not result in higher affinity peptides but did demonstrate the importance of amino acids at position 1-4 on the N-terminal flanking arm and I I-IS within the cyclic ring. Interestingly, EC-1 contains homologous motifs with known ErbB receptor family ligands. We have identified a small peptide that binds to the extracellular domain of ErbB2, inhibits ErbB2 autophosphorylation and inhibits the proliferation of ErbB2 overexpressing cells. This supports the notion that small peptides can bind to targets important in cancer therapy even if a target does not have a natural ligand. Continuing research with this peptide includes increasing its affinity to ErbB2, evaluation of pharmacokinetics and evaluation of anti-proliferative effects with conjugate anti-cancer agents. (C) 2004 Wiley-Liss, Inc.