Gálvez-Cancino Felipe, Roco Jonathan, Rojas-Colonelli Nicole, Flores Camila, Murgas Paola, Cruz-Gómez Sebastián, Oyarce César, Varas-Godoy Manuel, Sauma Daniela, Lladser Alvaro
Laboratorio de Inmunoterapia Génica, Fundación Ciencia & Vida, Av. Zañartu 1482, Santiago 7780272, Chile.
Centro de Investigación Biomédica, Facultad de Medicina, Universidad de los Andes, Santiago, Chile.
Vaccine. 2017 Jul 24;35(33):4148-4154. doi: 10.1016/j.vaccine.2017.06.041. Epub 2017 Jun 27.
DNA vaccination is an attractive approach to elicit tumor-specific cytotoxic CD8 T lymphocytes (CTL), which can mediate protective immunity against tumors. To initiate CTL responses, antigen-encoding plasmids employed for DNA vaccination need to activate dendritic cells (DC) through the stimulation of DNA-sensing innate immune receptors that converge in the activation of the master transcription factor NF-κB. To this end, NF-κB repressor IκBα needs to be degraded, allowing NF-κB to translocate to the nucleus and transcribe proinflammatory target genes, as well as its repressor IκBα. Therefore, NF-κB activation is self-limited by de novo synthesis of IκBa, which sequesters NF-κB in the cytosol. Hence, we tested whether co-delivering a shRNA-based adjuvant able to silence IκBα expression would further promote DNA-induced NFκB activation, DC activation and tumor-protective CTL responses induced by DNA vaccination in a preclinical model. First, an IκBα-targeting shRNA plasmid (shIκBα) was shown to reduce IκBα expression and promote NFκB-driven transcription in vitro, as well as up-regulate inflammatory target genes in vivo. Then, we showed that intradermal DNA electroporation induced the migration of skin migratory dendritic cells to draining lymph nodes and maturation of dermal dendritic cells (dDC). Interestingly, shIκBα further promoted the migration of mature skin migratory dendritic cells, in particular dDC, which are specialized in antigen cross-presentation and activation of CD8 T cells. Consistently, mice vaccinated with a plasmid encoding the melanoma-associated antigen tyrosinase-related protein 2 (TRP2) in combination with shIκBα enhanced TRP2-specific CTL responses and reduced the number of lung melanoma foci in mice challenged with intravenous injection of B16F10 cells. Moreover, therapeutic vaccination with pTRP2 and shIκBα delayed the growth of B16F10 melanoma subcutaneous tumors. Our data suggest that adjuvants promoting NF-κB activation represent an attractive strategy to boost DC activation and promote the generation of tumor-protective CTL responses elicited by DNA vaccines.
DNA疫苗接种是一种诱导肿瘤特异性细胞毒性CD8 T淋巴细胞（CTL）的有吸引力的方法，CTL可介导针对肿瘤的保护性免疫。为启动CTL反应，用于DNA疫苗接种的编码抗原的质粒需要通过刺激DNA传感固有免疫受体来激活树突状细胞（DC），这些受体在主转录因子NF-κB的激活中汇聚。为此，NF-κB抑制因子IκBα需要被降解，使NF-κB易位至细胞核并转录促炎靶基因及其抑制因子IκBα。因此，NF-κB的激活受到IκBa从头合成的自我限制，IκBa将NF-κB隔离在细胞质中。因此，我们测试了在临床前模型中，共同递送一种能够沉默IκBα表达的基于短发夹RNA（shRNA）的佐剂是否会进一步促进DNA诱导的NFκB激活、DC激活以及DNA疫苗接种诱导的肿瘤保护性CTL反应。首先，一种靶向IκBα的shRNA质粒（shIκBα）在体外显示可降低IκBα表达并促进NFκB驱动的转录，在体内还可上调炎症靶基因。然后，我们表明皮内DNA电穿孔可诱导皮肤迁移树突状细胞向引流淋巴结迁移以及真皮树突状细胞（dDC）成熟。有趣的是，shIκBα进一步促进了成熟皮肤迁移树突状细胞的迁移，特别是dDC，dDC专门负责抗原交叉呈递和CD8 T细胞的激活。一致地，用编码黑色素瘤相关抗原酪氨酸酶相关蛋白2（TRP2）的质粒与shIκBα联合接种的小鼠增强了TRP2特异性CTL反应，并减少了静脉注射B16F10细胞攻击的小鼠肺部黑色素瘤病灶数量。此外，用pTRP2和shIκBα进行治疗性疫苗接种可延缓B16F10黑色素瘤皮下肿瘤的生长。我们的数据表明，促进NF-κB激活的佐剂是增强DC激活并促进DNA疫苗引发的肿瘤保护性CTL反应产生的一种有吸引力的策略。
