Synthesis of a Bifunctional Peptide Inhibitor-IgG1 Fc Fusion That Suppresses Experimental Autoimmune Encephalomyelitis.
Synthesis of a Bifunctional Peptide Inhibitor-IgG1 Fc Fusion That Suppresses Experimental Autoimmune Encephalomyelitis.
复制标题
抑制实验性自身免疫性脑脊髓炎的双功能肽抑制剂-IgG1 Fc 融合体的合成。
DOI：
10.1021/acs.bioconjchem.7b00175
复制
发表时间：
2017
影响因子：
4.7
通讯作者：
Tolbert,ThomasJ
中科院分区：
文献类型：
--
作者：
White,DerekR;Khedri,Zahra;Kiptoo,Paul;Siahaan,TerunaJ;Tolbert,ThomasJ
多发性硬化症（MS）是一种神经退行性疾病，估计影响全球超过230万人。这种疾病的确切原因尚不清楚，但涉及免疫系统攻击和中枢神经系统神经元周围髓鞘蛋白的破坏。选择性地防止参与导致髓鞘破坏的途径的免疫细胞的活化的一类有前途的化合物是双功能肽抑制剂（BPIs）。治疗与BPI减少神经退行性症状的实验性自身免疫性脑脊髓炎（EAE），MS的小鼠模型。在这项工作中，作为一项努力，以进一步提高BPI的生物活性，BPI肽共轭的N-和C-末端的片段结晶（Fc）区的人IgG 1抗体。最初，使用重组DNA技术将两种肽缀合至IgG 1 Fc。然而，在酵母中的表达导致低产量和被严重蛋白水解的肽之一。为了避免这个问题，相反，通过固相肽合成产生表达差的肽，并使用分选酶介导的连接酶促缀合。分选酶介导的方法显示出在小规模反应中通过SDS-PAGE和质谱法观察到的接近完全的缀合产率。将该方法按比例放大，以获得足够的量用于在EAE诱导的小鼠中检测BPI-Fc融合。与PBS处理的对照相比，用BPI-Fc融合体处理的小鼠显示出显著减轻的疾病症状，没有经历体重减轻，并且显示出减少的脱髓鞘作用。这些结果表明，当以这种方式与大Fc支架缀合时，BPI肽在抑制EAE方面具有高度活性。
Multiple sclerosis (MS) is a neurodegenerative disease that is estimated to affect over 2.3 million people worldwide. The exact cause for this disease is unknown but involves immune system attack and destruction of the myelin protein surrounding the neurons in the central nervous system. One promising class of compounds that selectively prevent the activation of immune cells involved in the pathway leading to myelin destruction are bifunctional peptide inhibitors (BPIs). Treatment with BPIs reduces neurodegenerative symptoms in experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. In this work, as an effort to further improve the bioactivity of BPIs, BPI peptides were conjugated to the N- and C-termini of the fragment crystallizable (Fc) region of the human IgG1 antibody. Initially, the two peptides were conjugated to IgG1 Fc using recombinant DNA technology. However, expression in yeast resulted in low yields and one of the peptides being heavily proteolyzed. To circumvent this problem, the poorly expressed peptide was instead produced by solid phase peptide synthesis and conjugated enzymatically using a sortase-mediated ligation. The sortase-mediated method showed near-complete conjugation yield as observed by SDS-PAGE and mass spectrometry in small-scale reactions. This method was scaled up to obtain sufficient quantities for testing the BPI-Fc fusion in mice induced with EAE. Compared to the PBS-treated control, mice treated with the BPI-Fc fusion showed significantly reduced disease symptoms, did not experience weight loss, and showed reduced de-myelination. These results demonstrate that the BPI peptides were highly active at suppressing EAE when conjugated to the large Fc scaffold in this manner.