The differences in immunoadjuvant mechanisms of TLR3 and TLR4 agonists on the level of antigen-presenting cells during immunization with recombinant adenovirus vector

Ekaterina Lebedeva, Alexander Bagaev, Alexey Pichugin, Marina Chulkina, Andrei Lysenko, Irina Tutykhina, Maxim Shmarov, Denis Logunov, Boris Naroditsky, Ravshan Ataullakhanov

Agonists of TLR3 and TLR4 are effective immunoadjuvants for different types of vaccines. The mechanisms of their immunostimulatory action differ significantly; these differences are particularly critical for immunization with non-replicating adenovirus vectors (rAds) based vaccines. Unlike traditional vaccines, rAd based vaccines are not designed to capture vaccine antigens from the external environment by antigen presenting cells (APCs), but rather they are targeted to the de novo synthesis of vaccine antigens in APCs transfected with rAd. To date, there is no clear understanding about approaches to improve the efficacy of rAd vaccinations with immunoadjuvants. In this study, we investigated the immunoadjuvant effect of TLR3 and TLR4 agonists on the level of activation of APCs during vaccination with rAds. We demonstrated that TLR3 and TLR4 agonists confer different effects on the molecular processes in APCs that determine the efficacy of antigen delivery and activation of antigen-specific CD4+ and CD8+ T cells. APCs activated with agonists of TLR4 were characterized by up-regulated production of target antigen mRNA and protein encoded in rAd, as well as enhanced expression of the co-activation receptors CD80, CD86 and CD40, and pro-inflammatory cytokines TNF-α, IL6 and IL12. These effects of TLR4 agonists have provided a significant increase in the number of antigen-specific CD4+ and CD8+ T cells. TLR3 agonist, on the contrary, inhibited transcription and synthesis of rAd-encoded antigens, but improved expression of CD40 and IFN-β in APCs. The cumulative effect of TLR3 agonist have resulted in only a slight improvement in the activation of antigen-specific T cells. Also, we demonstrated that IFN-β and TNF-α, secreted by APCs in response to TLR3 and TLR4 agonists, respectively, have an opposite effect on the transcription of the targeted gene encoded in rAd. Specifically, IFN-β inhibited, and TNF-α stimulated the expression of target vaccine antigens in APCs. Our data demonstrate that agonists of TLR4 but not TLR3 merit further study as adjuvants for development of vaccines based on recombinant adenoviral vectors.