High-throughput identification of putative receptors for cancer-binding peptides using biopanning and microarray analysis.
High-throughput identification of putative receptors for cancer-binding peptides using biopanning and microarray analysis.
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DOI：
10.1039/c2ib20187a
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发表时间：
2013-02
期刊：
影响因子：
--
通讯作者：
Hallahan DE
中科院分区：
文献类型：
--
作者：
Ferraro DJ;Bhave SR;Kotipatruni RP;Hunn JC;Wildman SA;Hong C;Dadey DY;Muhoro LK;Jaboin JJ;Thotala D;Hallahan DE
噬菌体展示肽生物筛选已成功用于多种模型的癌症靶向肽鉴定。对于癌症结合肽，肽受体的鉴定是必要的，以证明其作用机制，并进一步优化特异性和靶向结合。受体鉴定的过程可能是缓慢的，一些肽可能会结合普遍存在的蛋白质，不适合进一步的药物开发。在本报告中，我们描述了一种高通量方法，用于平行筛选大量肽以识别肽受体，我们称之为“反向生物筛选”，然后可以根据其肽受体选择进一步开发。为了证明这种方法，我们筛选了先前在我们实验室鉴定的39个肽库，以特异性结合辐照后的癌症。反向生物筛选过程确定了2个肽，RKFLMTTRYSRV和KTAKKNVFFCSV，作为蛋白质税收相互作用蛋白1 （TIP-1）的候选配体，该蛋白先前在我们的实验室发现，在肿瘤细胞表面表达，并在暴露于电离辐射后上调。我们使用计算模型作为快速验证肽- tip -1结合的初始方法。HVGGSSV、TTRYSRV和NVFFCSV的赝结合能分别为−360.645 kcal/mol、−487.239 kcal/mol和−595.328 kcal/mol，表明与已知的与TIP-1结合的肽HVGGSSV相比，它们与TIP-1的结合至少相似，甚至更强。我们在体外通过电泳迁移位移实验验证了肽，结果表明RKFLMTTRYSRV和截断形式的TTRYSRV具有很强的结合。该方法允许识别许多肽受体和随后的肽选择，以进一步基于肽受体的药物开发。
Phage-display peptide biopanning has been successfully used to identify cancer-targeting peptides in multiple models. For cancer-binding peptides, identification of the peptide receptor is necessary to demonstrate mechanism of action and to further optimize specificity and target binding. The process of receptor identification can be slow and some peptides may turn out to bind ubiquitous proteins not suitable for further drug development. In this report, we describe a high-throughput method for screening a large number of peptides in parallel to identify peptide receptors, which we have termed “reverse biopanning,” which can then be selected for further development based on their peptide receptor. To demonstrate this method, we screened a library of 39 peptides previously identified in our laboratory to bind specifically cancers after irradiation. The reverse biopanning process identified 2 peptides, RKFLMTTRYSRV and KTAKKNVFFCSV, as candidate ligands for the protein tax interacting protein 1 (TIP-1), a protein previously identified in our laboratory to be expressed in the cell surface in tumors and upregulated after exposure to ionizing radiation. We used computational modeling as the initial method for rapid validation of peptide-TIP-1 binding. Pseudo-binding energies were calculated to be −360.645 kcal/mol, −487.239 kcal/mol, and −595.328 kcal/mol for HVGGSSV, TTRYSRV, and NVFFCSV respectively, suggesting that the peptides would have at least similar, if not stronger, binding to TIP-1 compared to the known TIP-1 binding peptide HVGGSSV. We validated peptide in vitro via electrophoretic mobility shift assay, which showed strong binding of RKFLMTTRYSRV and the truncated form TTRYSRV. This method allows for the identification of many peptide receptors and subsequent selection of peptides for further drug development based on the peptide receptor.
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影响因子：
11.2
作者：
Passarella RJ;Spratt DE;van der Ende AE;Phillips JG;Wu H;Sathiyakumar V;Zhou L;Hallahan DE;Harth E;Diaz R
通讯作者：
Diaz R
影响因子：
16.6
作者：
通讯作者：
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DOI：
10.1158/1078-0432.ccr-09-0945
发表时间：
2009-10-15
期刊：
Clinical cancer research : an official journal of the American Association for Cancer Research
影响因子：
--
作者：
Passarella RJ;Zhou L;Phillips JG;Wu H;Hallahan DE;Diaz R
通讯作者：
Diaz R
影响因子：
8.8
作者：
通讯作者：
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影响因子：
50.3
作者：
Hallahan, D;Geng, L;Clanton, J
通讯作者：
Clanton, J