Quantitative methods for developing Fc mutants with extended half-lives
Quantitative methods for developing Fc mutants with extended half-lives
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DOI：
10.1002/bit.20624
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发表时间：
2005-12-20
影响因子：
3.8
通讯作者：
Lauffenburger, DA
中科院分区：
文献类型：
--
作者：
Kamei, DT;Lao, BJ;Lauffenburger, DA
对新生儿受体FcRn具有增加的结合亲和力的Fc突变体表现出增加的体内半衰期，并且代表了用于延长治疗性抗体的半衰期的有吸引力的手段。其他治疗分子的半衰期（例如，蛋白质）也可以通过将它们缀合至Fc片段来延伸，从而降低患者注射的频率并允许施用低且潜在无毒浓度的治疗剂。为了研究进一步增加Fc半衰期的可能性，提出了一对定量方法来补充组合筛选和体内测试。具体而言，开发了一种简单的分子建模程序，以预测不同突变体和物种之间Fc和FcRn之间结合值的相对吉布斯自由能（Δ Δ G（结合））。发现该程序合理地再现了来自我们的实验和文献的实验Δ Δ G（结合）值，并且可以用作在实验测试之前更充分地探索Fc序列空间的初始筛选。此外，制定了Fc运输的数学模型，并与细胞水平的脉冲追踪测定相结合，以获得人T84细胞中的定量再循环参数。发现该Fc再循环参数与结合亲和力相关，但捕获了Fc和FcRn之间相互作用的pH依赖性性质，并且可以用作组合实验后的额外筛选。(c)2005 Wiley Periodicals，Inc.
Fc mutants with increased binding affinity for the neonatal receptor, FcRn, exhibit increased half-lives in vivo, and represent an attractive means for extending the half-lives of therapeutic antibodies. The half-lives of other therapeutic molecules (e.g., proteins) may also be extended by conjugating them to Fc fragments, thus decreasing the frequency of patient injections and allowing the administration of low and potentially nontoxic concentrations of the therapeutics. To investigate the possibility for further increasing the half-life of Fc, a pair of quantitative methods is presented to complement combinatorial screening and in vivo testing. Specifically, a simple molecular modeling procedure was developed to predict relative Gibbs free energies of binding values (Delta Delta G(bind)) between Fc and FcRn across different mutants and species. This procedure was found to reasonably reproduce experimental Delta Delta G(bind) values from our experiments and the literature, and may be used as an initial screen to explore Fc sequence space more fully prior to experimental testing. In addition, a mathematical model of Fc trafficking was formulated and combined with a cell-level pulse-chase assay to obtain a quantitative recycling parameter in human T84 cells. This Fc recycling parameter was found to be correlated with binding affinity, but captures the pH dependent nature of the interaction between Fc and FcRn and may serve as an additional screen following combinatorial experiments. (c) 2005 Wiley Periodicals, Inc.