Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity, The University of Melbourne, Melbourne, 3000, Australia.
Research Center for Zoonosis Control, Hokkaido University, Sapporo, 001-0020, Japan; Global Institution for Collaborative Research and Education, Hokkaido University, Sapporo, 001-0020, Japan.
Biomaterials. 2017 Aug;137:61-72. doi: 10.1016/j.biomaterials.2017.05.018. Epub 2017 May 11.
The lipopeptide RPamCys is an agonist for toll-like receptor-2 (TLR2), a key pathogen-associated molecular pattern receptor expressed on many antigen-presenting cells such as dendritic cells (DCs). Electrostatic association of RPamCys with soluble protein antigens significantly enhances their immunogenicity and there is evidence to suggest that reducing the size of suitably adjuvanted-antigen complexes in solution may further improve their immunostimulatory capabilities. In this study, we investigated how incorporation of polyethylene glycol (PEG) into RPamCys affects the size, activity and efficacy of formed antigen-lipopeptide complexes. The presence of PEG was shown to increase solubility with a concomitant reduction in the particle size of vaccine formulations that was dependent on the length of PEG used. When compared to non-PEGylated RPamCys, vaccination of animals with antigen-complexed PEGylated RPamCys resulted not only in improvements in antibody production but significantly higher antigen-specific CD8 T cell responses. Both lipopeptides exhibited similar in vitro capabilities to induce DC maturation, facilitate antigen uptake and presentation to T cells. Moreover, analyses of the transcriptomes obtained from DCs treated with either lipopeptide revealed a large number of commonly induced genes with similar transcript expression levels, suggesting that common signalling pathways and processes were engaged following activation by either lipopeptide. In vivo analysis however revealed that vaccination with antigen-complexed PEGylated RPamCys resulted in improved antigen presentation to T cells. These heightened responses were not attributed to prolonged antigen persistence but rather due to more rapid transportation of antigen from the injection site into the draining lymph nodes over a short period of time. Our results indicate that reducing the size of formed antigen-TLR2-agonist complexes by PEGylation does not compromise the activity of the agonist but in fact enhances its trafficking in vivo ultimately leading to improved humoral and cell-mediated immune responses.
脂肽 RPamCys 是 Toll 样受体 2（TLR2）的激动剂，TLR2 是一种表达在许多抗原呈递细胞（如树突状细胞（DC））上的关键病原体相关分子模式受体。RPamCys 与可溶性蛋白抗原的静电结合显著增强了它们的免疫原性，有证据表明，降低溶液中适当佐剂化抗原复合物的大小可能进一步提高它们的免疫刺激能力。在这项研究中，我们研究了将聚乙二醇（PEG）掺入 RPamCys 中如何影响形成的抗原-脂肽复合物的大小、活性和功效。结果表明，PEG 的存在增加了溶解度，同时疫苗制剂的粒径减小，这取决于使用的 PEG 的长度。与非 PEG 化的 RPamCys 相比，用抗原结合的 PEG 化 RPamCys 免疫动物不仅导致抗体产生的改善，而且还显著提高了抗原特异性 CD8 T 细胞反应。两种脂肽在体外均具有诱导 DC 成熟、促进抗原摄取和呈递给 T 细胞的能力。此外，用两种脂肽处理的 DC 的转录组分析显示，大量共同诱导的基因具有相似的转录表达水平，这表明在被两种脂肽激活后，共同的信号通路和过程被激活。然而，体内分析表明，用抗原结合的 PEG 化 RPamCys 免疫导致抗原向 T 细胞的呈递得到改善。这些增强的反应不是由于抗原持续存在时间延长，而是由于在短时间内从注射部位向引流淋巴结更快地运输抗原。我们的结果表明，通过 PEG 化降低形成的抗原-TLR2-激动剂复合物的大小不会损害激动剂的活性，但实际上增强了其在体内的运输，最终导致更好的体液和细胞介导的免疫反应。
