Hans Brightbill

IgE⁺ memory B cells and plasma cells generated through a germinal-center pathway.

Immunoglobulin E (IgE) antibodies are pathogenic in asthma and allergic diseases, but the in vivo biology of IgE-producing (IgE+) cells is poorly understood. A model of the differentiation of IgE+ B cells proposes that IgE+ cells develop through a germinal-center IgG1+ intermediate and that IgE memory resides in the compartment of IgG1+ memory B cells. Here we have used a reporter mouse expressing green fluorescent protein associated with membrane IgE transcripts (IgE-GFP) to assess in vivo IgE responses. In contrast to the IgG1-centered model of IgE switching and memory, we found that IgE+ cells developed through a germinal-center IgE+ intermediate to form IgE+ memory B cells and plasma cells. Our studies delineate a new model for the in vivo biology of IgE switching and memory.

A microRNA upregulated in asthma airway T cells promotes TH2 cytokine production

MicroRNAs (miRNAs) exert powerful effects on immunological function by tuning networks of target genes that orchestrate cell activity. We sought to identify miRNAs and miRNA-regulated pathways that control the type 2 helper T cell (TH2 cell) responses that drive pathogenic inflammation in asthma. Profiling miRNA expression in human airway-infiltrating T cells revealed elevated expression of the miRNA miR-19a in asthma. Modulating miR-19 activity altered TH2 cytokine production in both human and mouse T cells, and TH2 cell responses were markedly impaired in cells lacking the entire miR-17∼92 cluster. miR-19 promoted TH2 cytokine production and amplified inflammatory signaling by direct targeting of the inositol phosphatase PTEN, the signaling inhibitor SOCS1 and the deubiquitinase A20. Thus, upregulation of miR-19a in asthma may be an indicator and a cause of increased TH2 cytokine production in the airways.

Conditional Deletion of NF-κB–Inducing Kinase (NIK) in Adult Mice Disrupts Mature B Cell Survival and Activation

NF-κB–inducing kinase (NIK) is a primary regulator of the noncanonical NF-κB signaling pathway, which plays a vital role downstream of BAFF, CD40L, lymphotoxin, and other inflammatory mediators. Germline deletion or inactivation of NIK in mice results in the defective development of B cells and secondary lymphoid organs, but the role of NIK in adult animals has not been studied. To address this, we generated mice containing a conditional allele of NIK. Deletion of NIK in adult mice results in decreases in B cell populations in lymph nodes and spleen, similar to what is observed upon blockade of BAFF. Consistent with this, B cells from mice in which NIK is acutely deleted fail to respond to BAFF stimulation in vitro and in vivo. In addition, mice with induced NIK deletion exhibit a significant decrease in germinal center B cells and serum IgA, which is indicative of roles for NIK in additional pathways beyond BAFF signaling. Our conditional NIK-knockout mice may be broadly useful for assessing the postdevelopmental and cell-specific roles of NIK and the noncanonical NF-κB pathway in mice.

Addendum: IgE + memory B cells and plasma cells generated through a germinal-center pathway

Addendum: IgE + memory B cells and plasma cells generated through a germinal-center pathway

Dendritic cells require NIK for CD40-dependent cross-priming of CD8+ T cells.

Significance The noncanonical NF-κB signaling pathway via the serine kinase NIK (NF-κB–inducing kinase) is essential for normal immune system development and has been implicated in tumor cell survival and growth. Because NIK is under investigation as a therapeutic target, it is important to understand NIK’s role in the context of a fully developed immune system, particularly in regard to mounting adaptive T-cell responses. We have generated and characterized transgenic mice with conditionally deleted NIK in CD11c+ dendritic cells and observe impaired antigen cross-priming of a naive CD8 T-cell response. This defect results from defective antigen cross-presentation by CD8+ dendritic cells and also is associated with their reduced ability to secrete IL-12p40, a cytokine known to promote cross-priming in vivo. Dendritic cells (DCs) link innate and adaptive immunity and use a host of innate immune and inflammatory receptors to respond to pathogens and inflammatory stimuli. Although DC maturation via canonical NF-κB signaling is critical for many of these functions, the role of noncanonical NF-κB signaling via the serine/threonine kinase NIK (NF-κB–inducing kinase) remains unclear. Because NIK-deficient mice lack secondary lymphoid organs, we generated transgenic mice with targeted NIK deletion in CD11c+ cells. Although these mice exhibited normal lymphoid organs, they were defective in cross-priming naive CD8+ T cells following vaccination, even in the presence of anti-CD40 or polyinosinic:polycytidylic acid to induce DC maturation. This impairment reflected two intrinsic defects observed in splenic CD8+ DCs in vitro, namely antigen cross-presentation to CD8+ T cells and secretion of IL-12p40, a cytokine known to promote cross-priming in vivo. In contrast, antigen presentation to CD4+ T cells was not affected. These findings reveal that NIK, and thus probably the noncanonical NF-κB pathway, is critical to allow DCs to acquire the capacity to cross-present antigen and prime CD8 T cells after exposure to licensing stimuli, such as an agonistic anti-CD40 antibody or Toll-like receptor 3 ligand.

Structure-Based Design of Tricyclic NF-κB Inducing Kinase (NIK) Inhibitors That Have High Selectivity over Phosphoinositide-3-kinase (PI3K)

We report here structure-guided optimization of a novel series of NF-κB inducing kinase (NIK) inhibitors. Starting from a modestly potent, low molecular weight lead, activity was improved by designing a type 11/2 binding mode that accessed a back pocket past the methionine-471 gatekeeper. Divergent binding modes in NIK and PI3K were exploited to dampen PI3K inhibition while maintaining NIK inhibition within these series. Potent compounds were discovered that selectively inhibit the nuclear translocation of NF-κB2 (p52/REL-B) but not canonical NF-κB1 (REL-A/p50).

Cell intrinsic role of NF-κB-inducing kinase in regulating T cell-mediated immune and autoimmune responses

NF-κB inducing kinase (NIK) is a central component of the noncanonical NF-κB signaling pathway. Although NIK has been extensively studied for its function in the regulation of lymphoid organ development and B-cell maturation, the role of NIK in regulating T cell functions remains unclear and controversial. Using T cell-conditional NIK knockout mice, we here demonstrate that although NIK is dispensable for thymocyte development, it has a cell-intrinsic role in regulating the homeostasis and function of peripheral T cells. T cell-specific NIK ablation reduced the frequency of effector/memory-like T cells and impaired T cell responses to bacterial infection. The T cell-conditional NIK knockout mice were also defective in generation of inflammatory T cells and refractory to the induction of a T cell-dependent autoimmune disease, experimental autoimmune encephalomyelitis. Our data suggest a crucial role for NIK in mediating the generation of effector T cells and their recall responses to antigens. Together, these findings establish NIK as a cell-intrinsic mediator of T cell functions in both immune and autoimmune responses.

Scaffold-Hopping Approach To Discover Potent, Selective, and Efficacious Inhibitors of NF-κB Inducing Kinase

NF-κB-inducing kinase (NIK) is a protein kinase central to the noncanonical NF-κB pathway downstream from multiple TNF receptor family members, including BAFF, which has been associated with B cell survival and maturation, dendritic cell activation, secondary lymphoid organ development, and bone metabolism. We report herein the discovery of lead chemical series of NIK inhibitors that were identified through a scaffold-hopping strategy using structure-based design. Electronic and steric properties of lead compounds were modified to address glutathione conjugation and amide hydrolysis. These highly potent compounds exhibited selective inhibition of LTβR-dependent p52 translocation and transcription of NF-κB2 related genes. Compound 4f is shown to have a favorable pharmacokinetic profile across species and to inhibit BAFF-induced B cell survival in vitro and reduce splenic marginal zone B cells in vivo.

NIK signaling axis regulates dendritic cell function in intestinal immunity and homeostasis

Dendritic cells (DCs) play an integral role in regulating mucosal immunity and homeostasis, but the signaling network mediating this function of DCs is poorly defined. We identified the noncanonical NF-κB-inducing kinase (NIK) as a crucial mediator of mucosal DC function. DC-specific NIK deletion impaired intestinal immunoglobulin A (IgA) secretion and microbiota homeostasis, rendering mice sensitive to an intestinal pathogen, Citrobacter rodentium. DC-specific NIK was required for expression of the IgA transporter polymeric immunoglobulin receptor (pIgR) in intestinal epithelial cells, which in turn relied on the cytokine IL-17 produced by TH17 cells and innate lymphoid cells (ILCs). NIK-activated noncanonical NF-κB induced expression of IL-23 in DCs, contributing to the maintenance of TH17 cells and type 3 ILCs. Consistent with the dual functions of IL-23 and IL-17 in mucosal immunity and inflammation, NIK deficiency also ameliorated colitis induction. Thus, our data suggest a pivotal role for the NIK signaling axis in regulating DC functions in intestinal immunity and homeostasis.The kinase NIK activates a noncanonical NF-κB2 signaling pathway. Shao-Cong Sun and colleagues show that conditional loss of NIK in dendritic cells alters gut microbiome composition and TH17 cell responses, as a result of reduced expression of IL-23, pIgR and fecal IgA transcytosis.