Bennink Lucas L, Smith Daniel J, Foss Catherine A, Pomper Martin G, Li Yang, Yu S Michael
Department of Bioengineering, University of Utah , Salt Lake City, Utah 84112, United States.
The Russel H. Morgan Department of Radiology and Radiological Science, Johns Hopkins University, School of Medicine , Baltimore, Maryland 21228, United States.
Mol Pharm. 2017 Jun 5;14(6):1906-1915. doi: 10.1021/acs.molpharmaceut.7b00009. Epub 2017 May 8.
Collagen hybridizing peptides (CHPs) have a great potential for use in targeted drug delivery, diagnostics, and regenerative medicine due to their ability to specifically bind to denatured collagens associated with many pathologic conditions. Since peptides generally suffer from poor enzymatic stability, resulting in rapid degradation and elimination in vivo, CHP's serum stability is a critical parameter that may dictate its pharmacokinetic behavior. Here, we report the serum stability of a series of monomeric CHP derivatives and establish how peptide length, amino acid composition, terminal modification, and linker chemistry influence their availability in serum. We show that monomeric CHPs comprised of the collagen-like Gly-Pro-Hyp motif are resistant to common serum proteinases and that their stability can be further increased by simple N-terminal labeling which negates CHP's susceptibility to proline-specific exopeptidases. When fluorescent dyes are conjugated to a CHP via maleimide-thiol reaction, the dye can transfer from CHP onto serum proteins (e.g., albumin), resulting in an unexpected drop in signal during serum stability assays and off-target accumulation during in vivo tests. This work is the crucial first step toward understanding the pharmacokinetic behavior of CHPs, which can facilitate the development of CHP-based theranostics.
胶原杂交肽（CHPs）因其能够特异性结合与多种病理状况相关的变性胶原，在靶向药物递送、诊断和再生医学方面具有巨大的应用潜力。由于肽类通常酶稳定性较差，导致其在体内迅速降解和清除，CHP的血清稳定性是一个关键参数，可能决定其药代动力学行为。在此，我们报告了一系列单体CHP衍生物的血清稳定性，并确定了肽链长度、氨基酸组成、末端修饰和连接子化学结构如何影响它们在血清中的可用性。我们表明，由类胶原Gly-Pro-Hyp基序组成的单体CHP对常见的血清蛋白酶具有抗性，并且通过简单的N端标记可以进一步提高其稳定性，这消除了CHP对脯氨酸特异性外肽酶的敏感性。当荧光染料通过马来酰亚胺-硫醇反应与CHP偶联时，染料可从CHP转移到血清蛋白（如白蛋白）上，导致血清稳定性测定期间信号意外下降以及体内试验期间出现脱靶积累。这项工作是理解CHP药代动力学行为的关键第一步，有助于基于CHP的治疗诊断学的发展。
