A short hairpin RNA-based adjuvant targeting NF-kappa B repressor I kappa B alpha promotes migration of dermal dendritic cells to draining lymph nodes and antitumor CTL responses induced by DNA vaccination

Abstract

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-kappa B. To this end, NF-kappa B repressor I kappa B alpha needs to be degraded, allowing NF-kappa B to translocate to the nucleus and transcribe proinflammatory target genes, as well as its repressor I kappa B alpha. Therefore, NF-kappa B activation is self-limited by de novo synthesis of I kappa B alpha, which sequesters NF-kappa B in the cytosol. Hence, we tested whether co-delivering a shRNA-based adjuvant able to silence I kappa B alpha expression would further promote DNA-induced NF kappa B activation, DC activation and tumor-protective CTL responses induced by DNA vaccination in a preclinical model. First, an I kappa B alpha-targeting shRNA plasmid (shI kappa B alpha) was shown to reduce I kappa B alpha expression and promote NP kappa B-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, shI kappa B alpha further promoted the migration of mature skin migratory dendritic cells, in particular dDC, which are specialized in antigen cross-presentation and activation of CDS+ T cells. Consistently, mice vaccinated with a plasmid encoding the melanoma-associated antigen tyrosinase-related protein 2 (TRP2) in combination with shI kappa B alpha 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 shI kappa B alpha delayed the growth of B16F10 melanoma subcutaneous tumors. Our data suggest that adjuvants promoting NF-kappa B activation represent an attractive strategy to boost DC activation and promote the generation of tumor-protective CTL responses elicited by DNA vaccines.