The Mevalonate Pathway Is a Druggable Target for Vaccine Adjuvant Discovery.

Motivated by the clinical observation that interruption of the mevalonate pathway stimulates immune responses, we hypothesized that this pathway may function as a druggable target for vaccine adjuvant discovery. We found that lipophilic statin drugs and rationally designed bisphosphonates that target three distinct enzymes in the mevalonate pathway have potent adjuvant activities in mice and cynomolgus monkeys. These inhibitors function independently of conventional "danger sensing." Instead, they inhibit the geranylgeranylation of small GTPases, including Rab5 in antigen-presenting cells, resulting in arrested endosomal maturation, prolonged antigen retention, enhanced antigen presentation, and T cell activation. Additionally, inhibiting the mevalonate pathway enhances antigen-specific anti-tumor immunity, inducing both Th1 and cytolytic T cell responses. As demonstrated in multiple mouse cancer models, the mevalonate pathway inhibitors are robust for cancer vaccinations and synergize with anti-PD-1 antibodies. Our research thus defines the mevalonate pathway as a druggable target for vaccine adjuvants and cancer immunotherapies.

The E3 ligases Itch and WWP2 cooperate to limit TH2 differentiation by enhancing signaling through the TCR.

The mechanisms by which the sensitivity of naive CD4+ T cells to stimulation by the cognate antigen via the T cell antigen receptor (TCR) determines their differentiation into distinct helper T cell subsets remain elusive. Here we demonstrate functional collaboration of the ubiquitin E3 ligases Itch and WWP2 in regulating the strength of the TCR signal. Mice lacking both Itch and WWP2 in T cells showed spontaneous autoimmunity and lung inflammation. CD4+ T cells deficient in Itch and WWP2 exhibited hypo-responsiveness to TCR stimulation and a bias toward differentiation into the TH2 subset of helper T cells. Itch and WWP2 formed a complex and cooperated to enhance TCR-proximal signaling by catalyzing the conjugation of atypical ubiquitin chains to the phosphatase SHP-1 and reducing the association of SHP-1 with the tyrosine kinase Lck. These findings indicate that targeted ubiquitination regulates the strength of the TCR signal and differentiation toward the TH2 lineage.

E3 Ligase VHL Promotes Group 2 Innate Lymphoid Cell Maturation and Function via Glycolysis Inhibition and Induction of Interleukin-33 Receptor.

Group 2 innate lymphoid cells (ILC2s) are a specialized subset of lymphoid effector cells that are critically involved in allergic responses; however, the mechanisms of their regulation remain unclear. We report that conditional deletion of the E3 ubiquitin ligase VHL in innate lymphoid progenitors minimally affected early-stage bone marrow ILC2s but caused a selective and intrinsic decrease in mature ILC2 numbers in peripheral non-lymphoid tissues, resulting in reduced type 2 immune responses. VHL deficiency caused the accumulation of hypoxia-inducible factor 1α (HIF1α) and attenuated interleukin-33 (IL-33) receptor ST2 expression, which was rectified by HIF1α ablation or inhibition. HIF1α-driven expression of the glycolytic enzyme pyruvate kinase M2 downmodulated ST2 expression via epigenetic modification and inhibited IL-33-induced ILC2 development. Our study indicates that the VHL-HIF-glycolysis axis is essential for the late-stage maturation and function of ILC2s via targeting IL-33-ST2 pathway.

Critical role for thymic CD19+CD5+CD1dhiIL-10+ regulatory B cells in immune homeostasis.

This study tested the hypothesis that besides the spleen, LNs, peripheral blood, and thymus contain a regulatory IL-10-producing CD19(+)CD5(+)CD1d(high) B cell subset that may play a critical role in the maintenance of immune homeostasis. Indeed, this population was identified in the murine thymus, and furthermore, when cocultured with CD4(+) T cells, this population of B cells supported the maintenance of CD4(+)Foxp3(+) Tregs in vitro, in part, via the CD5-CD72 interaction. Mice homozygous for Cd19(Cre) (CD19(-/-)) express B cells with impaired signaling and humoral responses. Strikingly, CD19(-/-) mice produce fewer CD4(+)Foxp3(+) Tregs and a greater percentage of CD4(+)CD8(-) and CD4(-)CD8(+) T cells. Consistent with these results, transfer of thymic CD19(+)CD5(+)CD1d(hi) B cells into CD19(-/-) mice resulted in significantly up-regulated numbers of CD4(+)Foxp3(+) Tregs with a concomitant reduction in CD4(+)CD8(-) and CD4(-)CD8(+) T cell populations in the thymus, spleen, and LNs but not in the BM of recipient mice. In addition, thymic CD19(+)CD5(+)CD1d(hi) B cells significantly suppressed autoimmune responses in lupus-like mice via up-regulation of CD4(+)Foxp3(+) Tregs and IL-10-producing Bregs. This study suggests that thymic CD19(+)CD5(+)CD1d(hi)IL-10(+) Bregs play a critical role in the maintenance of immune homeostasis.

Novel IL-6-secreting γδT cells increased in patients with atherosclerotic cerebral infarction.

Mounting evidence has suggested that inflammation associated with interleukin (IL)­6 and T­helper (Th)17 cells, has a role in the development of atherosclerotic cerebral infarction (ACI). However, it remains unclear which population of cells determines the levels of IL­6, and the role of IL­6­secreting cells in inducing Th17 cell production. In the present study, IL­6 levels were determined in patients with ACI, by ELISA. The percentage of CD3+T, CD4+T, CD8+T, CD11c+ dendritic cells and γδT cells were determined by flow cytometry, and the correlation between cytokine IL­6 and γδT cells was determined by statistical analysis. An in vitro culture assay was used to determine whether γδT cells secreted high levels of IL­6, and induced production of Th17 cells. The patients with ACI had significantly higher levels of IL­6 and γδT cells. Furthermore, γδT cells were associated with the secretion of a high level of IL­6 in patients with ACI. These results indicate that γδT cells are novel IL­6­secreting cells, which from then on were known as γδT6 cells. In addition, the novel γδT6 cells induced Th17­cell production, and this induction was dependent on IL­6. Novel γδT6 cells increased the induction of Th17­cell production in patients with ACI. The results of the present study suggest that novel γδT6 cells may be a target for strategic therapies of ACI.

Interaction of CD5 and CD72 is involved in regulatory T and B cell homeostasis.

Regulatory IL-10-producing CD1d(high)CD5(+)CD19(+) B cells and CD4(+)CD25(+)Foxp3(+) T cells have been found to modulate immune responses in autoimmunity, infection, and cancer, but the interaction between these two cell subsets remains unclear. Through cell culture and flow cytometry (FACS), we analyzed the interaction of regulatory T cells (Tregs) and regulatory B cells (Bregs). A neutralizing antibody was used to determine the role of CD5 and CD72 in maintaining regulatory T and B cell homeostasis. We found that CD19(+)CD5(+)CD1d(hi) Bregs induced expansion of CD4(+)Foxp3(+) Tregs, and CD4(+)CD25(+) Tregs also induced expansion of IL-10-expressing Bregs. Once CD72 or CD5 was blocked, both IL-10-expressing Bregs and CD4(+)Foxp3(+)Tregs were reduced in the different cultures. Finally, FACS analysis demonstrated that Foxp3(+)CD4(+)Treg cells were reduced in CD19(Cre) mice defective of CD5 on the surface of B cells. The study suggests that the interaction of CD5 and CD72 plays a critical role in maintaining regulatory T and B cell homeostasis.

Combination of TACI-IgG and anti-IL-15 treats murine lupus by reducing mature and memory B cells.

Clinical trials suggest that BAFF inhibitors such as atacicept (TACI-IgG) and belimumab (anti-BAFF antibody) could not reduce memory B-cell numbers, although they reduced the numbers of CD20(+) naïve B cells and activated B cells. In the present study, we explored the way to reduce memory B-cell numbers. First, we used TACI-IgG to treat murine lupus. We found that TACI-IgG was effective in reducing mature B cell numbers. Accordingly it controlled the level of the anti-dsDNA antibody in lupus-like mice. In addition, TACI-IgG up-regulated memory B cells in murine lupus. Furthermore, we found that TACI-IgG up-regulated IL-15 expression in lupus-like mice. Thus, the combination of TACI-IgG and anti-IL-15 antibodies was explored to understand their effects on the treatment of murine lupus. Compared to treatments with TACI-IgG or anti-IL-15 alone, the combination of TACI-IgG and anti-IL-15 antibodies efficiently ameliorated murine lupus phenotypes. The study provides hints for the clinical application of BAFF- and IL-15-specific therapeutic agents.