Effects of cationic adjuvant formulation particle type, fluidity and immunomodulators on delivery and immunogenicity of saRNA
Effects of cationic adjuvant formulation particle type, fluidity and immunomodulators on delivery and immunogenicity of saRNA
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DOI：
10.1016/j.jconrel.2019.04.043
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发表时间：
2019-06-28
影响因子：
10.8
通讯作者：
Shattock, Robin J.
中科院分区：
文献类型：
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作者：
Blakney, Anna K.;McKay, Paul F.;Shattock, Robin J.
自扩增RNA（saRNA）非常适合作为针对衣原体的疫苗平台，因为它相对经济实惠且可扩展，已显示可诱导针对多价抗原的免疫力，并且可导致蛋白质表达长达60天。阳离子佐剂制剂（CAFs）以前已被研究作为蛋白质亚单位疫苗的佐剂，在这里，我们优化CAFs在体内传递的saRNA，并观察在细胞和体液免疫的背景下，对沙眼衣原体的主要外膜蛋白（MOMP）的免疫原性。我们测试了具有固相和液相脂质的基于脂质体和乳液的CAF，有或没有TLR激动剂R848和3 M-052，用于体外转染效率和细胞毒性。然后，我们使用编码萤火虫荧光素酶（fLuc）的saRNA作为体内报告蛋白，优化了用于体内递送的RNA/递送系统比率。我们观察到，虽然流体相脂质体制剂显示出最高的体外转染效率，但流体和固相脂质体在体内具有等同的荧光素酶表达。未观察到将R848或3 M-052掺入制剂中影响saRNA的体外或体内递送效率。与CAF复合的编码MOMP的saRNA导致MOMP特异性细胞和体液免疫，并且虽然当R848掺入制剂中时IFN-γ（+）T细胞应答略有增强，但RNA的自佐剂效应似乎主导免疫应答。这些研究确定CAF是saRNA的有效递送载体，用于体外转染和体内免疫原性，并产生与蛋白质表达成比例的细胞和体液应答。
Self-amplifying RNA (saRNA) is well suited as a vaccine platform against chlamydia, as it is relatively affordable and scalable, has been shown to induce immunity against multivalent antigens, and can result in protein expression for up to 60 days. Cationic adjuvant formulations (CAFs) have been previously investigated as an adjuvant for protein subunit vaccines; here we optimize the CAFs for delivery of saRNA in vivo and observe the immunogenicity profile in the context of both cellular and humoral immunity against the major outer membrane protein (MOMP) of Chlamydia trachomatis. We tested both liposomal and emulsion based CAFs with solid and fluid phase lipids, with or without the TLR agonists R848 and 3M-052, for in vitro transfection efficiency and cytotoxicity. We then optimized the RNA/delivery system ratio for in vivo delivery using saRNA coding for firefly luciferase (fLuc) as a reporter protein in vivo. We observed that while the fluid phase liposome formulations showed the highest in vitro transfection efficiency, the fluid and solid phase liposomes had equivalent luciferase expression in vivo. Incorporation of R848 or 3M-052 into the formulation was not observed to affect the delivery efficiency of saRNA either in vitro or in vivo. MOMP-encoding saRNA complexed with CAFs resulted in both MOMP-specific cellular and humoral immunity, and while there was a slight enhancement of IFN-gamma(+) T-cell responses when R848 was incorporated into the formulation, the self-adjuvanting effects of RNA appeared to dominate the immune response. These studies establish that CAFs are efficient delivery vehicles for saRNA both for in vitro transfections and in vivo immunogenicity and generate cellular and humoral responses that are proportionate to protein expression.