The Middle East respiratory syndrome coronavirus (MERS-CoV) is a highly pathogenic and zoonotic virus with a fatality rate in humans of over 35%. Although several vaccine candidates have been developed, there is still no clinically available vaccine for MERS-CoV. In this study, we developed two types of MERS-CoV vaccines: a recombinant adenovirus serotype 5 encoding the MERS-CoV spike gene (Ad5/MERS) and spike protein nanoparticles formulated with aluminum (alum) adjuvant. Next, we tested a heterologous prime-boost vaccine strategy, which compared priming with Ad5/MERS and boosting with spike protein nanoparticles and vice versa, with homologous prime-boost vaccination comprising priming and boosting with either spike protein nanoparticles or Ad5/MERS. Although both types of vaccine could induce specific immunoglobulin G against MERS-CoV, neutralizing antibodies against MERS-CoV were induced only by heterologous prime-boost immunization and homologous immunization with spike protein nanoparticles. Interestingly, Th1 cell activation was induced by immunization schedules including Ad5/MERS, but not by those including only spike protein nanoparticles. Heterologous prime-boost vaccination regimens including Ad5/MERS elicited simultaneous Th1 and Th2 responses, but homologous prime-boost regimens did not. Thus, heterologous prime-boost may induce longer-lasting immune responses against MERS-CoV because of an appropriate balance of Th1/Th2 responses. However, both heterologous prime-boost and homologous spike protein nanoparticles vaccinations could provide protection from MERS-CoV challenge in mice. Our results demonstrate that heterologous immunization by priming with Ad5/MERS and boosting with spike protein nanoparticles could be an efficient prophylactic strategy against MERS-CoV infection.

Inhibitor K562 (IK) protein was first isolated from the culture medium of K562, a leukemia cell line. It is known to be an inhibitory regulator of interferon-γ-induced major histocompatibility complex class (MHC) II expression. Previously, we found that transgenic (Tg) mice constitutively expressing truncated IK (tIK) showed reduced numbers of pathogenic Th1 and Th17 cells, which are known to be involved in the development of rheumatoid arthritis (RA). Here, we investigated whether exogenous tIK protein has a therapeutic effect in arthritis in disease models and analyzed its mechanism. Exogenous tIK protein was produced in an insect expression system and applied to the collagen antibody-induced arthritis (CAIA) mouse disease model. Injection of tIK protein alleviated the symptoms of arthritis in the CAIA model and reduced Th1 and Th17 cell populations. In addition, treatment of cultured T cells with tIK protein induced expression of A20, a negative regulator of nuclear factor-κB (NFκB)-induced inflammation, and reduced expression of several transcription factors related to T cell activation. We conclude that exogenous tIK protein has the potential to act as a new therapeutic agent for RA patients, because it has a different mode of action to biopharmaceutical agents, such as tumor necrosis factor antagonists, that are currently used to treat RA.

The Middle East respiratory syndrome coronavirus (MERS-CoV) is a single-strained RNA virus and is included in Betacoronavirus. While, in the Arabian Peninsula, it has shown an endemic pattern, a sporadic pattern was observed elsewhere. In 2015, there was big outbreak of the MERS-CoV in Korea. However, since we still have no clinically available vaccine to prevent the MERS-CoV infection, such a vaccine is urgently required. In the present study, we developed two types of MERS-CoV vaccines: a spike nanoparticle vaccine produced by an insect culture system and a recombinant adenovirus 5 vaccine expressing spike protein. Both vaccines induced total IgG, IgG1, and IgG2a against spike protein of the MERS-CoV. Thus, both vaccines can trigger Th1 and Th2 immune responses. Interestingly, as compared to spike nanoparticle formulation with Alum without RNA adjuvant, spike nanoparticle formulation with Alum and RNA adjuvant showed higher titers of IgG1 and IgG2a at 10 days after boosting immunization. Moreover, T cell proliferation in splenocytes of immunized mice was well induced after the spike nanoparticle treatment. Thus, RNA adjuvant may be effective to increase vaccine efficacy against the MERS-CoV. Taken together, the results of the present study suggest that the developed two type of MERS-CoV vaccines–the spike nanoparticle vaccine and the recombinant virus vaccine–may be helpful to prevent the spread of the MERS-CoV.

In 2015, there was an outbreak of Middle East respiratory syndrome coronavirus (MERS-CoV) in Korea. Therefore, rapid diagnostic kits for the viral antigen and antibodies against MERS-CoV are urgently required. First, we produced and purified a spike nanoparticle of MERS-CoV by the insect expression system. It was highly soluble and intact for immunization. The spike nanoparticle was used as a coating antigen for enzyme-linked immunosorbent assay (ELISA) to detect MERS-CoV-specific antibodies in the serum. The spike nanoparticle, at least with 1.5 ng/well as a coating antigen, can detect serially diluted standard antibodies purified from rabbits immunized with inactivated MERS-CoV, and from rats immunized with spike nanoparticle. Moreover, it can successfully detect MERS-CoV in patient serum. Next, to develop sandwich ELISA to detect MERS-CoV antigens, we generated 616 hybridoma clones and selected 37 monoclonal antibodies (MAbs), which exclusively reacted against the spike nanoparticle. Among them, three MAbs (S-15, S-16, and S-34) showed about 40% plaque reduction against MERS-CoV. Of the 37 MAbs, we selected the two that showed the optimal combinations to develop sandwich ELISA. This sandwich ELISA also can detect serially diluted MERS-CoV antigen. In this study, we developed two MERS-CoV-specific ELISA kits. One is for detecting MERS antibodies in patient serum and the other is for detecting MERS-CoV itself. Both ELISA kits can be reliably used for diagnosis of MERS-CoV.

The truncated IK (tIK) cytokine is known for its ability to downregulate the major histocompatibility complex class II on antigen-presenting cells and to regulate immune activation in inflammatory diseases by suppressing differentiation of pathogenic T cells and activation of macrophage. Because of these immune regulatory functions, previous research has proposed tIK as a new candidate for autoimmune disease therapy such as that for rheumatoid arthritis (RA). However, its immunological mechanism is still not identified clearly. In this study, we showed that exogenous tIK protein produced by the insect expression system suppressed the differentiation of TH1 and TH17 cells, which are key factors of inflammatory arthritis. We also showed that tIK protein reduced symptoms of the disease in the collagen antibody-induced arthritis (CAIA) mouse model. In addition, to investigate the receptor and signal transducers that directly interact with tIK and/or the intracellular molecules as a signal pathway, we performed a protein–protein interaction array. According to protein array data, FAM104B, CUTA, GSN, YEATS4 and some other proteins might interact with tIK protein. Among them some proteins may serve as the negative regulators of NF-kappaB transcription factor. Therefore, we suggest that tIK protein has the potential to act as a new therapeutic agent for RA patients, because it has a different mode of action compared with the currently-used biologics for RA, such as monoclonal antibody drugs.