Toll-Like Receptor 7 Agonist RG7854 Mediates Therapeutic Efficacy and Seroconversion in Woodchucks With Chronic Hepatitis B.

Conventional treatment of chronic hepatitis B (CHB) is rarely curative due to the immunotolerant status of patients. RG7854 is an oral double prodrug of a toll-like receptor 7 (TLR7) agonist that is developed for the treatment of CHB. The therapeutic efficacy, host immune response, and safety of RG7854 were evaluated in the woodchuck model of CHB. Monotreatment with the two highest RG7854 doses and combination treatment with the highest RG7854 dose and entecavir (ETV) suppressed viral replication, led to loss of viral antigens, and induced seroconversion in responder woodchucks. Since viral suppression and high-titer antibodies persisted after treatment ended, this suggested that a sustained antiviral response (SVR) was induced by RG7854 in a subset of animals. The SVR rate, however, was comparable between both treatment regimens, suggesting that the addition of ETV did not enhance the therapeutic efficacy of RG7854 although it augmented the proliferation of blood cells in response to viral antigens and magnitude of antibody titers. The induction of interferon-stimulated genes in blood by RG7854/ETV combination treatment demonstrated on-target activation of TLR7. Together with the virus-specific blood cell proliferation and the transient elevations in liver enzymes and inflammation, this suggested that cytokine-mediated non-cytolytic and T-cell mediated cytolytic mechanisms contributed to the SVR, in addition to the virus-neutralizing effects by antibody-producing plasma cells. Both RG7854 regimens were not associated with treatment-limiting adverse effects but accompanied by dose-dependent, transient neutropenia and thrombocytopenia. The study concluded that finite, oral RG7854 treatment can induce a SVR in woodchucks that is based on the retrieval of antiviral innate and adaptive immune responses. This supports future investigation of the TLR7 agonist as an immunotherapeutic approach for achieving functional cure in patients with CHB.

Gasdermin D in macrophages restrains colitis by controlling cGAS-mediated inflammation.

The functional relevance and mechanistic basis of the effects of the pyroptosis executioner Gasdermin D (GSDMD) on colitis remain unclear. In this study, we observed that GSDMD protein was activated during intestinal inflammation in a model of chemically induced colitis. GSDMD deficiency exacerbated experimental colitis independent of changes in the microbiota and without affecting the production of antimicrobial peptides. GSDMD deficiency in macrophages, but not epithelial cells, was sufficient to drive this exacerbated experimental colitis. We further demonstrate that GSDMD functions in macrophages as a negative regulator to control cyclic GMP-AMP synthase (cGAS)-dependent inflammation, thereby protecting against colitis. Moreover, the administration of cGAS inhibitor can rescue the colitogenic phenotype in GSDMD-deficient mice. Collectively, these findings provide the first demonstration of GSDMD's role in controlling colitis and a detailed delineation of the underlying mechanism.

Evaluating the potential of IL-27 as a novel therapeutic agent in HIV-1 infection.

Interleukin 27 (IL-27) is an immunomodulatory cytokine with important roles in both the innate and adaptive immune systems. In the last five years, the addition of exogenous IL-27 to primary cell cultures has been demonstrated to decrease HIV-1 replication in a number of cell types including peripheral blood mononuclear cells (PBMCs), CD4+ T cells, macrophages and dendritic cells. These in vitro findings suggest that IL-27 may have therapeutic value in the setting of HIV-1 infection. In this review, we describe the current knowledge of the biology of IL-27, its effects primarily on HIV-1 replication but also in other viral infections and explore its potential role as a therapeutic cytokine for the treatment of patients with HIV-1 infection.

Interleukin-2 inhibits HIV-1 replication in some human T cell lymphotrophic virus-1-infected cell lines via the induction and incorporation of APOBEC3G into the virion.

IL-2 has been used in culture of primary T cells to maintain cell proliferation. We have previously reported that IL-27 inhibits HIV-1 replication in primary T cells in the presence of IL-2. To gain a better understanding of the mechanisms involved in this inhibitory effect, we attempted to investigate in detail the effects of IL-27 and IL-2 using several cell lines. Unexpectedly, IL-27 did not inhibit HIV-1 in T cell lines, whereas IL-2 inhibited HIV-1 replication in the human T cell lymphotrophic virus (HTLV)-1-transformed T cell lines, MT-2, MT-4, SLB-1, and ATL-2. No effects were seen in HTLV-1-negative cell lines. Utilizing MT-2 cells, we demonstrated that IL-2 treatment inhibited HIV-1 syncytia-inducing ability and dose-dependently decreased supernatant p24 antigen levels by >90%. Using real time PCR and Western blot analysis, we observed that IL-2 treatment induced the host restriction factor, APOBEC3G with accumulation into the lower molecular mass active form as characterized by FPLC. Further analysis revealed that the virus recovered from IL-2-treated MT-2 cells had impaired replication competency. This was found to be due to incorporation of APOBEC3G into the virion despite the presence of Vif. These findings demonstrate a novel role for IL-2 in regulating production of infectious HIV-1 virions in HTLV-1-infected cells through the induction of APOBEC3G.

IL-27 inhibits HIV-1 infection in human macrophages by down-regulating host factor SPTBN1 during monocyte to macrophage differentiation.

The susceptibility of macrophages to HIV-1 infection is modulated during monocyte differentiation. IL-27 is an anti-HIV cytokine that also modulates monocyte activation. In this study, we present new evidence that IL-27 promotes monocyte differentiation into macrophages that are nonpermissive for HIV-1 infection. Although IL-27 treatment does not affect expression of macrophage differentiation markers or macrophage biological functions, it confers HIV resistance by down-regulating spectrin ß nonerythrocyte 1 (SPTBN1), a required host factor for HIV-1 infection. IL-27 down-regulates SPTBN1 through a TAK-1-mediated MAPK signaling pathway. Knockdown of SPTBN1 strongly inhibits HIV-1 infection of macrophages; conversely, overexpression of SPTBN1 markedly increases HIV susceptibility of IL-27-treated macrophages. Moreover, we demonstrate that SPTBN1 associates with HIV-1 gag proteins. Collectively, our results underscore the ability of IL-27 to protect macrophages from HIV-1 infection by down-regulating SPTBN1, thus indicating that SPTBN1 is an important host target to reduce HIV-1 replication in one major element of the viral reservoir.

Interleukin-27 is a potent inhibitor of cis HIV-1 replication in monocyte-derived dendritic cells via a type I interferon-independent pathway.

IL-27, a member of the IL-12 family of cytokines, plays an important and diverse role in the function of the immune system. Whilst generally recognized as an anti-inflammatory cytokine, in addition IL-27 has been found to have broad anti-viral effects. Recently, IL-27 has been shown to be a potent inhibitor of HIV-1 infection in CD4+ T cells and macrophages. The main objective of this study was to see whether IL-27 has a similar inhibitory effect on HIV-1 replication in dendritic cells (DCs). Monocytes were differentiated into immature DCs (iDCs) and mature DCs (mDCs) with standard techniques using a combination of GM-CSF, IL-4 and LPS. Following differentiation, iDCs were infected with HIV-1 and co-cultured in the presence or absence of IL-27. IL-27 treated DCs were shown to be highly potent inhibitors of cis HIV-1, particularly of CCR5 tropic strains. Of note, other IL-12 family members (IL-12, IL-23 and IL-35) had no effect on HIV-1 replication. Microarray studies of IL-27 treated DCs showed no up-regulation of Type I (IFN) gene expression. Neutralization of the Type-I IFN receptor had no impact on the HIV inhibition. Lastly, IL-27 mediated inhibition was shown to act post-viral entry and prior to completion of reverse transcription. These results show for the first time that IL-27 is a potent inhibitor of cis HIV-1 infection in DCs by a Type I IFN independent mechanism. IL-27 has previously been reported to inhibit HIV-1 replication in CD4+ T cells and macrophages, thus taken together, this cytokine is a potent anti-HIV agent against all major cell types targeted by the HIV-1 virus and may have a therapeutic role in the future.

A novel motif in HIV-1 Nef that regulates MIP-1beta chemokine release in macrophages.

All primate lentiviruses encode Nef, an accessory protein that is important for viral pathogenicity in vivo. Lentiviral Nef proteins regulate the release of chemokines (MIP-1 alpha/beta) from infected macrophages, thereby enhancing virus dissemination (S. Swingler, A. Mann, J. Jacque, B. Brichacek, V. G. Sasseville, K. Williams, A. A. Lackner, E. N. Janoff, R. Wang, D. Fisher, and M. Stevenson, Nat. Med. 5:997-1003, 1999). In the current study, we have identified a novel domain within Nef (K(92)EK) that is required for Nef-dependent MIP-1beta production by infected macrophages. Mutations in this domain abrogated MIP-1beta induction but did not affect other Nef-ascribed activities, such as CD4 or major histocompatibility complex (MHC) class Iota downregulation. This further underscores Nef as a modular protein with genetically separable activities that may contribute to its role in viral replication and pathogenicity.