Immunomodulatory effects of dsRNA and its potential as vaccine adjuvant

Abstract

dsRNA can be detected by pattern recognition receptors, for example, TLR3, MDA-5, NLRP3 to induce proinflammatory cytokines responsible for innate/adaptive immunity. Recognized by endosomal TLR3 in myeloid DCs (mDCs), dsRNA can activate mDCs into mature antigen presenting cells (mAPCs) which in turn present antigen epitopes with MHC-I molecules to naïve T cells. Coadministration of protein and synthetic dsRNA analogues can elicit an antigen-specific Th1-polarized immune response which stimulates the CD8+ CTL response and possibly dampen Th17 response. Synthetic dsRNA analogues have been tested as vaccine adjuvant against viral infections in animal models. However, a dsRNA receptor, TLR3 can be expressed in tumor cells while other members of TLR family, for example, TLR4 and TLR2 have been shown to promote tumor progression, metastasis, and chemoresistance. Thus, the promising potential of dsRNA analogues as a tumor therapeutic vaccine adjuvant should be evaluated cautiously.

Figure 1

dsRNA induced signal pathways. (A) dsRNA signaling through TLR3. dsRNA that is internalized into endosome binds to TLR3. The mechanism of dsRNA internalization is unknown, CD14 may play a role. TLR3 possesses two dsRNA binding sites near the N-terminus and C-terminus. When combined with dsRNA, four dsRNA binding sites from two TLR3 molecules that are linked by the sole dsRNA molecule in an “m” shape, and the B-B loop of the TLR3 TIR domain combines with the TIR domain of TRIF. The interaction of TRIF with RIP1 or TRAF6 results in polyubiquitination of RIP1, the latter binds ubiquitin receptors TAB2 and TAB3 which activates TAK1. Activated TAK1 induces phosphorylation of IKKα and IKKβ leading to phosphorylation and degradation of IκB. The release and translocation of NF-κB to cell nucleus then occurs which results in the specific gene promoter A20 being activated. TAK1 also starts an activation sequence beginning with JNK and p38, which activates c-JUN and ATF2 and this activates the AP-1 transcription factors family. TRIF activates TBK1 and IKKε through NAP1 and this results in phosphorylation and nuclear translocaton of IRF3 resulting in IFN-β production. TRAF3 also binds with the TBK1/IKKε complex inducing IRF3 activation. Combination of TRIF results in phosphorylation of Tyr⁷⁵⁹ and Tyr⁸⁵⁸ in the TLR3 TIR domain which subsequently induces the phosphorylation of, and degradation of IκB leading to NF-κB release. Phosphorylated Tyr⁷⁵⁹ recruits PI3K and activates kinase Akt for full phosphorylation and activation of IRF3 in nucleus. Tyrosine kinase c-Src also plays a role in Akt activation. TRIF interacts with FADD through RIP1 and activates procaspase-8 to initiate cell apoptosis. (B) dsRNA signaling through MDA-5. dsRNA is recognized by MDA-5 in cytosol. MDA-5 interacts with MAVS located on the outer mitochondrial membrane via CARD-CARD interaction leading to the dimerization of the MAVS N-terminal CARD domains which subsequently binds TRAF3 directly through the interaction between the TRAF domain of TRAF3 and the TRAF-interacting motif in the PRR of MAVS. Then the RING domain of TRAF3 forms a scaffold to assemble the noncanonical IKKs signal complex composed of TANK, TBK1, IKKε, NAP1 and NEMO. This complex activates the signal-dependent phosphorylation of IRF3 and IRF7 to form a functional homodimer or heterodimer which translocate to the nucleus to induce expression of type I IFN. TRAF2 and TRAF6 also associates with MAVS and is involved in activation of P38 MAPK and promotion of IL-12 and type I IFN production or activation of NF-κB, JNK and P38 signaling. In addition, MAVS recruits TRADD and interacts with TRAF3, TANK, FADD and RIP1 to activate both IRF3 and NF-κB signaling. (C) dsRNA signaling through NLRP3 inflammasome. NLRP3 recognizes dsRNA by a series of LRRs at the C-terminus. The NACHT domain at the middle of NLRP3 is related to self-oligomerization and the formation of inflammasome. Studies have suggested dsRNA may activate the NLRP3 inflammasome. Activation of NLRP3 results in the interaction of NLRP3 PYD with ASC PYD, and in turn ASC CARD associates with pro-caspase-1 CARD and thereby assembles the NLRP3 inflammasome. Once incorporated into NLRP3 inflammasome, pro-caspase-1 is activated by proteolytic cleavages to remove the CARD prodomain. The active caspase-1 in turn cleaves the IL-1β, IL-18, IL-33 and IL-1F7 precursors into their active forms and these active cytokines are secreted extracellularly.