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Screening Bioactive Nanoparticles in Phagocytic Immune Cells for Inhibitors of Toll-like Receptor Signaling
Published on: July 26, 2017
Distinct indirect pathways govern human NK-cell activation by TLR-7 and TLR-8 agonists
Kevin S Gorski1, Emily L Waller, Jacqueline Bjornton-Severson
13M Pharmaceuticals, Department of Pharmacology, 3M Center, 270-2S-06, St Paul, MN 55144, USA. kgorski@mmm.com
Abstract:
NK cells limit the emergence of cancers and viral infections by surveillance of 'missing-self' and 'induced-self' ligands, and by direct recognition of pathogen-associated molecules. We examined individual roles for Toll-like receptors (TLRs)-7 and -8 in human NK-cell activation using synthetic, small molecule agonists of either TLR-7 (imiquimod and 3M-001), TLR-8 (3M-002) or both TLR-7/8 (3M-003 and R-848) for comparison with known ligands of TLR-2 to -9. Tracking cytokine production in PBMC initially revealed that a subset of TLR agonists including polyinosinic-polycytidylic acid (poly I:C), 3M-002, 3M-003, R-848 and single-stranded RNA trigger relatively high levels of IFN-gamma expression by NK cells. Isolated NK cells did not express TLR-7 or TLR-8. Unlike MALP-2 and poly I:C, 3M-001-3 did not induce expression of either CD69 or IFN-gamma by purified NK cells suggesting indirect activation. IL-18 and IL-12p70 were primarily required for induction of IFN-gamma by both synthetic and natural TLR-8 ligands, while type I IFN was required for induction of CD69 on NK cells by the TLR-7 agonist 3M-001. In addition to expression of IFN-gamma and CD69, relative induction of NK-cell cytotoxicity by TLR-7 and TLR-8 agonists was compared. Immune response modifiers (IRMs) with a TLR-8 agonist component (3M-002 and 3M-003) stimulated greater levels of K562 cytolysis than achieved with 3M-001 or IL-2 (1000 units ml(-1)). In vivo NK-cell cytotoxicity was also enhanced by R-848, but not in type I IFNR-deficient mice. We conclude that IRMs can modulate NK-cell function both in vitro and in vivo and that distinct indirect pathways control human NK-cell activation by TLR-7 and TLR-8 agonists.
Insights
Toll-like receptors (TLRs) 7 and 8 modulate natural killer (NK) cell functions. Immune response modifiers (IRMs) activate NK cells indirectly, influencing cytokine production and cytotoxicity, with TLR-8 agonists showing enhanced effects.
Area of Science:
- Immunology
- Cellular Biology
- Infectious Disease Research
Background:
- Natural killer (NK) cells are crucial for immune surveillance against cancer and viral infections.
- Toll-like receptors (TLRs) are key components of the innate immune system, recognizing pathogen-associated molecules.
- The specific roles of TLR-7 and TLR-8 in human NK-cell activation remain incompletely understood.
Purpose of the Study:
- To investigate the individual contributions of TLR-7 and TLR-8 to human NK-cell activation.
- To compare the effects of synthetic TLR-7 and TLR-8 agonists on NK-cell cytokine production and cytotoxicity.
- To elucidate the distinct indirect pathways mediating NK-cell responses to TLR-7 and TLR-8 agonists.
Main Methods:
- Utilized synthetic small molecule agonists for TLR-7 (imiquimod, 3M-001), TLR-8 (3M-002), and dual TLR-7/8 (3M-003, R-848).
- Analyzed cytokine production (IFN-gamma, IL-18, IL-12p70, type I IFN) and surface marker expression (CD69) in peripheral blood mononuclear cells (PBMCs) and isolated NK cells.
- Assessed NK-cell cytotoxicity against K562 target cells in vitro and in vivo, including experiments with type I IFNR-deficient mice.
Main Results:
- Specific TLR agonists (poly I:C, 3M-002, 3M-003, R-848, ssRNA) induced significant IFN-gamma production by NK cells.
- Isolated NK cells lacked TLR-7 and TLR-8 expression, indicating indirect activation pathways.
- TLR-8 agonists (3M-002, 3M-003) significantly enhanced NK-cell cytotoxicity compared to TLR-7 agonists or IL-2.
Conclusions:
- Immune response modifiers (IRMs) effectively modulate human NK-cell function both in vitro and in vivo.
- Distinct indirect signaling pathways mediate NK-cell activation by TLR-7 and TLR-8 agonists.
- TLR-8 agonists demonstrate potent induction of NK-cell cytotoxicity, suggesting therapeutic potential.
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