Therapeutic modulation of inflammasome pathways

Abstract

Inflammasomes are macromolecular complexes formed in response to pathogen-associated molecular patterns (PAMPs) and danger-associated molecular patterns (DAMPs) that drive maturation of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18, and cleave gasdermin D (GSDMD) for induction of pyroptosis. Inflammasomes are highly important in protecting the host from various microbial pathogens and sterile insults. Inflammasome pathways are strictly regulated at both transcriptional and post-translational checkpoints. When these checkpoints are not properly imposed, undue inflammasome activation may promote inflammatory, metabolic and oncogenic processes that give rise to autoinflammatory, autoimmune, metabolic and malignant diseases. In addition to clinically approved IL-1-targeted biologics, upstream targeting of inflammasome pathways recently gained interest as a novel pharmacological strategy for selectively modulating inflammasome activation in pathological conditions.

Keywords: GSDMD; IL-1; NLRP3; disease; immunotherapy; inflammasome; inflammation; pyroptosis.

FIGURE 1

Overview of different inflammasome complexes: (From left to right) Engagement of different TLR receptors leads to activation of IKKs, which in turn results in activation of NF‐κB signaling. Consequently, NF‐κB upregulates different pro‐inflammatory cytokines such as TNF, IL‐6, and pro‐IL‐1β. In addition, NF‐κB signaling upregulates NLRP3 expression (priming step). In the activation step of the NLRP3 inflammasome, diverse stimuli such as nigericin (ionophore), extracellular ATP, and medically relevant crystals (MSU, cholesterol) are sensed by NLRP3 and trigger its activation. Given the diversity of stimuli that activate NLRP3, K⁺ efflux has been proposed as a common mechanism for NLRP3 inflammasome activation, although K⁺ efflux‐independent pathways also exist. NEK7 protein is an essential mediator for NLRP3 activation that acts independently of its kinase activity. NLRP3 recruits the adapter protein ASC and the protease caspase‐1, which drives maturation of pro‐IL‐1β and pro‐IL‐18 into their respective active forms. At the same time, caspase‐1 cleaves GSDMD to release GSDMD‐N that forms GSDMD pores into the plasma membrane and drives pyroptosis, causing the release of various DAMPs such as IL‐1β, IL‐18, IL‐1α, and HMGB1. In the non‐canonical NLRP3 pathway, intracellular sensing of LPS activates caspase‐4/11 (human/mouse) that cleaves GSDMD, which induces pyroptosis, consequently activating the NLRP3 inflammasome and IL‐1β maturation through K⁺‐efflux. NLRP1 undergoes auto‐catalytic processing in its FIIND domain. Proteasomal degradation of its auto‐inhibitory N‐terminus is considered a primary requirement of NLRP1 inflammasome activation. In rodents, cleavage on the N‐terminal part of the NLRP1b protein by anthrax lethal toxin or IpaH7.8 allows the C‐terminal CARD domain to engage caspase‐1 for activation. In addition, mouse NLRP1b and human NLRP1 are activated by DPP8/DPP9 inhibitors. Activation of the NLRC4 inflammasome requires NAIPs that recognize T3SS inner rod proteins, needle proteins, or flagellin. In case of NLRP1b and NLRC4, caspase‐1 can directly be recruited independently of the adapter ASC. dsDNA of microbial or host origin in the cytosolic compartment activates AIM2, which requires ASC to activate caspase‐1. Clostridium difficile toxins (TcdA/B) inhibit RhoA GTPases activity, which inhibits PKN1/2‐dependent phosphorylation of Pyrin. Consequently, 14‐3‐3 disengages from Pyrin, allowing the assembly of the Pyrin inflammasome

IGURE 2

F IL‐1 and IL‐18 signaling and its modulation by therapeutic agents: (Left) Engagement of IL‐1 and IL‐18 receptors by IL‐1α, IL1β, and IL‐18, respectively, leads to the cytosolic recruitment of the TIR domain‐containing adapter MyD88. Subsequently, MyD88 binds to the IRAK kinases IRAK1/2/4, which associate with the E3‐ubiquitin ligase TRAF6. This allows dissociation of IRAK kinases and TRAF6 from the receptor complex and results in the polyubiquitination of the kinase TAK‐1, which leads to the phosphorylation of IKKs. In turn, IKKs phosphorylate and degrade IκB, subsequently inducing the upregulation of NF‐κB target genes and the release of pro‐inflammatory cytokines. To maintain the balance of IL‐1 signaling, extracellular IL‐1Ra competitively inhibits the binding of active IL‐1β and IL‐1α to IL‐1R1. Furthermore, the decoy receptor IL‐1R2 binds to IL‐1β and IL‐1α and inhibits IL‐1 signaling. Similarly, IL‐18 signaling is regulated by the inhibitory activity of IL‐18BP that binds IL‐18 and prevents its interaction with its cell surface receptors. (Right) Three biologic agents targeting IL‐1 signaling have been approved by the FDA. As a recombinant form of the IL‐1Ra, anakinra competes with IL‐1β and IL‐1α for binding to IL‐1R1. The human anti‐IL‐1β monoclonal antibody canakinumab specifically binds and neutralizes IL‐1β to prevent its interaction with IL‐1R1. Rilonacept is a soluble decoy receptor, which consists of the extracellular domains of IL‐1R1 and IL‐1RAcP, and binds and neutralizes both IL‐1β and IL‐1α. Similar to IL‐1 blocking agents, the recombinant IL‐18BP, tadekinig alfa, and the human anti‐IL‐18 monoclonal antibody, GSK1070806, bind to IL‐18 and inhibit the interaction of IL‐18 to its cell surface receptors. Both IL‐18 blockers are currently under investigation in clinical trials

FIGURE 3

Pharmacological targeting of inflammasomes. NLRP3 inflammasome activation requires a two‐step mechanism. The priming step involves NF‐κB‐dependent transcriptional upregulation of NLRP3, which is activated by the engagement of TLRs, CLRs, and cytokine receptors. Small molecule compounds such as Bay 11‐7082, parthenolide, and disulfiram were shown to inhibit inflammasome signaling at several levels, including the priming step. These compounds inhibit NF‐κB signaling, consequently blocking the expression of pro‐inflammatory cytokines such as pro‐IL‐1β, TNF, and IL‐6, as well as the synthesis of new NLRP3 protein. Small molecules were also shown to inhibit NLRP3 activation at the post‐translational level. The HAT inhibitor, NU9056, was reported to block NLRP3 acetylation, and the JNK kinase inhibitor, SP600125, was shown to inhibit NLRP3 phosphorylation. Other small molecules directly inhibit NLRP3 inflammasome activation. MCC950/CRID3, which is structurally related to the sulfonylurea compound glyburide, is currently the most potent and selective inhibitor of NLRP3 that directly targets its NACHT domain. The MCC950/CRID3‐related compounds IZD334 and Inzomelid are currently under investigation for the treatment of CAPS. CY‐09, MNS, OLT1177, Bay 11‐7082, and parthenolide were shown to bind directly to NLRP3 and inhibit its ATPase activity. Tranilast was shown to inhibit NLRP3 oligomerization via an ATPase‐independent manner. The natural product oridonin prevents the direct interaction between NLRP3 and NEK7. In addition to various inhibitors, the NLRP3 activator, BMS‐986299, is currently being investigated in a clinical trial for cancer treatment. VX‐740 and VX‐765 are specific inhibitors of caspase‐1. Blocking inflammasome signaling by targeting GSDMD is an alternative strategy currently being explored with small molecule inhibitors, such as LDC7559, NSA, Disulfiram, and Bay 11‐7082