{{Short description|Cell surface receptor found in humans}} {{cs1 config |name-list-style=vanc |display-authors=6}} {{Infobox_gene}} '''Toll-like receptor 4''' (TLR4), also designated as '''CD284''' (cluster of differentiation 284), is a key activator of the innate immune response and plays a central role in the fight against bacterial infections. TLR4 is a transmembrane protein of approximately 95 kDa that is encoded by the ''TLR4'' gene.
'''TLR4''' belongs to the toll-like receptor family which is representative of the pattern recognition receptors (PRR), so named for their ability to recognize evolutionarily conserved components of microorganisms (bacteria, viruses, fungi and parasites) called pathogen-associated molecular patterns (PAMPs). The recognition of a PAMP by a PRR triggers rapid activation of the innate immunity essential to fight infectious diseases.<ref name="pmid25071777">{{cite journal | vauthors = Vaure C, Liu Y | title = A comparative review of toll-like receptor 4 expression and functionality in different animal species | journal = Frontiers in Immunology | volume = 5 | page = 316 | date = 2014 | pmid = 25071777 | pmc = 4090903 | doi = 10.3389/fimmu.2014.00316 | doi-access = free }}</ref>
TLR4 is expressed in immune cells mainly of myeloid origin, including monocytes, macrophages and dendritic cells (DC).<ref name="pmid25071777"/> It is also expressed at a lower level on some non-immune cells, including epithelium, endothelium, placental cells and beta cells in Langerhans islets. Most myeloid cells express also high amounts of plasma membrane-anchored CD14, which facilitates the activation of TLR4 by LPS and controls the subsequent internalization of the LPS-activated TLR4 important for receptor signaling and degradation.<ref>{{cite book | vauthors = Mahnke K, Becher E, Ricciardi-Castagnoli P, Luger TA, Schwarz T, Grabbe S | chapter = CD14 is Expressed by Subsets of Murine Dendritic Cells and Upregulated by Lipopolysaccharide |date=1997 | title =Dendritic Cells in Fundamental and Clinical Immunology | series = Advances in Experimental Medicine and Biology |volume=417 |pages=145–159 | veditors = Ricciardi-Castagnoli P |place=Boston, MA |publisher=Springer US |doi=10.1007/978-1-4757-9966-8_25 | pmid = 9286353 |isbn=978-1-4757-9968-2 }}</ref><ref>{{cite journal | vauthors = Sabroe I, Jones EC, Usher LR, Whyte MK, Dower SK | title = Toll-like receptor (TLR)2 and TLR4 in human peripheral blood granulocytes: a critical role for monocytes in leukocyte lipopolysaccharide responses | journal = Journal of Immunology | volume = 168 | issue = 9 | pages = 4701–4710 | date = May 2002 | pmid = 11971020 | doi = 10.4049/jimmunol.168.9.4701 }}</ref>
The main ligands for TLR4 are lipopolysaccharides (LPS), the major components of the outer membrane of Gram-negative bacteria and some Gram-positive bacteria. TLR4 can also be activated by endogenous compounds called damage-associated molecular patterns (DAMPs), including high mobility group box protein 1 (HMGB1), S100 proteins, or histones. These compounds are released during tissue injury and by dying or necrotic cells.<ref name="pmid25559892">{{cite journal | vauthors = Yang H, Wang H, Ju Z, Ragab AA, Lundbäck P, Long W, Valdes-Ferrer SI, He M, Pribis JP, Li J, Lu B, Gero D, Szabo C, Antoine DJ, Harris HE, Golenbock DT, Meng J, Roth J, Chavan SS, Andersson U, Billiar TR, Tracey KJ, Al-Abed Y | title = MD-2 is required for disulfide HMGB1-dependent TLR4 signaling | journal = The Journal of Experimental Medicine | volume = 212 | issue = 1 | pages = 5–14 | date = January 2015 | pmid = 25559892 | pmc = 4291531 | doi = 10.1084/jem.20141318 }}</ref><ref name=":0">{{cite journal | vauthors = Jiang D, Liang J, Fan J, Yu S, Chen S, Luo Y, Prestwich GD, Mascarenhas MM, Garg HG, Quinn DA, Homer RJ, Goldstein DR, Bucala R, Lee PJ, Medzhitov R, Noble PW | title = Regulation of lung injury and repair by Toll-like receptors and hyaluronan | journal = Nature Medicine | volume = 11 | issue = 11 | pages = 1173–1179 | date = November 2005 | pmid = 16244651 | doi = 10.1038/nm1315 | s2cid = 11765495 }}</ref><ref name=":1">{{cite journal | vauthors = Fang H, Ang B, Xu X, Huang X, Wu Y, Sun Y, Wang W, Li N, Cao X, Wan T | title = TLR4 is essential for dendritic cell activation and anti-tumor T-cell response enhancement by DAMPs released from chemically stressed cancer cells | journal = Cellular & Molecular Immunology | volume = 11 | issue = 2 | pages = 150–159 | date = March 2014 | pmid = 24362470 | pmc = 4003380 | doi = 10.1038/cmi.2013.59 }}</ref><ref name=":2">{{cite journal | vauthors = Hernandez C, Huebener P, Schwabe RF | title = Damage-associated molecular patterns in cancer: a double-edged sword | journal = Oncogene | volume = 35 | issue = 46 | pages = 5931–5941 | date = November 2016 | pmid = 27086930 | pmc = 5119456 | doi = 10.1038/onc.2016.104 }}</ref><ref name=":3">{{cite journal | vauthors = Jang GY, Lee JW, Kim YS, Lee SE, Han HD, Hong KJ, Kang TH, Park YM | title = Interactions between tumor-derived proteins and Toll-like receptors | journal = Experimental & Molecular Medicine | volume = 52 | issue = 12 | pages = 1926–1935 | date = December 2020 | pmid = 33299138 | pmc = 8080774 | doi = 10.1038/s12276-020-00540-4 }}</ref>
== Function ==
The first function described for TLR4 was the recognition of exogenous molecules from pathogens (PAMPs), in particular LPS molecules from gram-negative bacteria.<ref name="auto1">{{cite journal | vauthors = Molteni M, Gemma S, Rossetti C | title = The Role of Toll-Like Receptor 4 in Infectious and Noninfectious Inflammation | journal = Mediators of Inflammation | volume = 2016 | article-number = 6978936 | date = 2016 | pmid = 27293318 | pmc = 4887650 | doi = 10.1155/2016/6978936 | doi-access = free }}</ref> As pattern recognition receptor, TLR4 plays a fundamental role in pathogen recognition and activation of innate immunity which is the first line of defense against invading micro-organisms. During infection, TLR4 responds to the LPS present in tissues and the bloodstream and triggers pro-inflammatory reactions facilitating eradication of the invading bacteria.<ref name="auto1"/>
TLR4 is also involved in the recognition of endogenous DAMP molecules leading to different signaling outcomes than PAMPs, both quantitatively and qualitatively.<ref name="auto">{{cite journal | vauthors = Roh JS, Sohn DH | title = Damage-Associated Molecular Patterns in Inflammatory Diseases | journal = Immune Network | volume = 18 | issue = 4 | article-number = e27 | date = August 2018 | pmid = 30181915 | pmc = 6117512 | doi = 10.4110/in.2018.18.e27 }}</ref><ref name=":3"/> DAMPs can activate TLR4 in non-infectious conditions to induce tissue repair and the activation of mainly proinflammatory responses.<ref name="pmid25559892" /><ref name=":0" /><ref name=":1" /><ref name=":2" /><ref name=":3" /> Generally, inflammation has a protective role. It is a complex and coordinated process followed by the induction of resolution pathways that restore tissue integrity and function. However, in some cases, an excessive and/or poorly regulated inflammatory response to DAMPs can be detrimental to the organism, accelerating the development or progression of pathologies such as a number of cancers and neurodegenerative diseases (as discussed below).
TLR4 binds LPS with the help of LPS-binding protein (LBP) and CD14, and an indispensable contribution of the MD-2 protein stably associated with the extracellular fragment of the receptor.<ref>{{cite journal | vauthors = Tsukamoto H, Takeuchi S, Kubota K, Kobayashi Y, Kozakai S, Ukai I, Shichiku A, Okubo M, Numasaki M, Kanemitsu Y, Matsumoto Y, Nochi T, Watanabe K, Aso H, Tomioka Y | title = Lipopolysaccharide (LPS)-binding protein stimulates CD14-dependent Toll-like receptor 4 internalization and LPS-induced TBK1-IKKϵ-IRF3 axis activation | journal = The Journal of Biological Chemistry | volume = 293 | issue = 26 | pages = 10186–10201 | date = June 2018 | pmid = 29760187 | pmc = 6028956 | doi = 10.1074/jbc.M117.796631 | doi-access = free }}</ref> TLR4 signaling responds to signals by forming a complex using an extracellular leucine-rich repeat domain (LRR) and an intracellular toll/interleukin-1 receptor (TIR) domain. LPS stimulation induces a series of interactions with several accessory proteins which form the TLR4 complex on the cell surface. LPS recognition is initiated by an LPS binding to an LBP. This LPS-LBP complex transfers the LPS to CD14 which is a glycosylphosphatidylinositol-anchored membrane protein that binds the LPS-LBP complex and facilitates the transfer of LPS to MD-2 protein, which is associated with the extracellular domain of TLR4. LPS binding promotes the dimerization of TLR4/MD-2 complex. The conformational changes of the TLR4 induce the recruitment of intracellular adaptor proteins containing the TIR domain which is necessary to activate the downstream signaling pathway.
The binding of an LPS molecule to the TLR4/MD-2 complex involves acyl chains and phosphate groups of lipid A, the conserved part of LPS and the main inducer of pro-inflammatory responses to LPS.<ref name="auto4">{{cite journal | vauthors = Park BS, Song DH, Kim HM, Choi BS, Lee H, Lee JO | title = The structural basis of lipopolysaccharide recognition by the TLR4-MD-2 complex | journal = Nature | volume = 458 | issue = 7242 | pages = 1191–1195 | date = April 2009 | pmid = 19252480 | doi = 10.1038/nature07830 | s2cid = 4396446 | bibcode = 2009Natur.458.1191P }}</ref><ref>{{cite journal | vauthors = Park BS, Lee JO | title = Recognition of lipopolysaccharide pattern by TLR4 complexes | journal = Experimental & Molecular Medicine | volume = 45 | issue = 12 | pages = e66 | date = December 2013 | pmid = 24310172 | pmc = 3880462 | doi = 10.1038/emm.2013.97 }}</ref>
TLR4 activation and response to LPS is greatly influenced by the polysaccharide domain and the molecular structure of Lipid A moiety of the LPS molecules. Hexa-acylated and diphosphorylated LPS, like Escherichia coli LPS (O111:B4), is one of the most potent agonists of TLR4 whereas under-acylated LPS and dephosphorylated LPS species have a weaker pro-inflammatory activity especially in human cells.<ref>{{cite journal | vauthors = Steimle A, Autenrieth IB, Frick JS | title = Structure and function: Lipid A modifications in commensals and pathogens | journal = International Journal of Medical Microbiology | volume = 306 | issue = 5 | pages = 290–301 | date = August 2016 | pmid = 27009633 | doi = 10.1016/j.ijmm.2016.03.001 | doi-access = free }}</ref> Structural determinants of this phenomenon are found in the TLR4/MD-2 complex and also in CD14 protein.<ref name="auto4"/><ref>{{cite journal | vauthors = Kelley SL, Lukk T, Nair SK, Tapping RI | title = The crystal structure of human soluble CD14 reveals a bent solenoid with a hydrophobic amino-terminal pocket | journal = Journal of Immunology | volume = 190 | issue = 3 | pages = 1304–1311 | date = February 2013 | pmid = 23264655 | pmc = 3552104 | doi = 10.4049/jimmunol.1202446 }}</ref> The polysaccharide portion covalently bound to lipid A also plays an indispensable role in TLR4 activation through CD14/TLR4/MD-2.<ref>{{cite journal | vauthors = Muroi M, Tanamoto K | title = The polysaccharide portion plays an indispensable role in Salmonella lipopolysaccharide-induced activation of NF-kappaB through human toll-like receptor 4 | journal = Infection and Immunity | volume = 70 | issue = 11 | pages = 6043–6047 | date = November 2002 | pmid = 12379680 | pmc = 130318 | doi = 10.1128/IAI.70.11.6043-6047.2002 }}</ref> However, in addition to the lipid A domain, the polysaccharide moiety plays an important role in the binding and activation of the LPS molecules as the lipid A moiety alone was demonstrated to be significantly less active than the full LPS molecule.<ref>{{cite journal | vauthors = Cavaillon JM, Fitting C, Caroff M, Haeffner-Cavaillon N | title = Dissociation of cell-associated interleukin-1 (IL-1) and IL-1 release induced by lipopolysaccharide and lipid A | journal = Infection and Immunity | volume = 57 | issue = 3 | pages = 791–797 | date = March 1989 | pmid = 2537258 | pmc = 313178 | doi = 10.1128/iai.57.3.791-797.1989 }}</ref>
== Signaling ==
Unlike all the other TLRs, TLR4 stimulation triggers two signaling pathways called the MyD88-dependent and the TRIF-dependent one after the adaptor proteins involved in their induction.<ref name="auto9">{{cite journal | vauthors = Shen H, Tesar BM, Walker WE, Goldstein DR | title = Dual signaling of MyD88 and TRIF is critical for maximal TLR4-induced dendritic cell maturation | journal = Journal of Immunology | volume = 181 | issue = 3 | pages = 1849–1858 | date = August 2008 | pmid = 18641322 | pmc = 2507878 | doi = 10.4049/jimmunol.181.3.1849 }}</ref> The MyD88-dependent signaling is triggered by TLR4 localized to the plasma membrane, while the TRIF-dependent one by the TLR4 internalized in endosomes.
These signaling pathways lead to the production of two sets of cytokines. The MyD88-dependent pathway induces the production of pro-inflammatory cytokines while TRIF-dependent pathway induces the production of type I interferons and chemokines.<ref name="auto9"/><ref>{{cite journal | vauthors = Nakayama M, Niki Y, Kawasaki T, Takeda Y, Ikegami H, Toyama Y, Miyamoto T | title = IL-32-PAR2 axis is an innate immunity sensor providing alternative signaling for LPS-TRIF axis | journal = Scientific Reports | volume = 3 | issue = 1 | article-number = 2960 | date = October 2013 | pmid = 24129891 | pmc = 3797434 | doi = 10.1038/srep02960 | bibcode = 2013NatSR...3.2960N }}</ref> The molecular structure of TLR4 ligands (in particular LPS), as well as their complexation with proteins or lipids, greatly influence the action of these TLR4-related signaling pathways, leading to different cytokine balances.<ref>{{cite journal | vauthors = Pridmore AC, Jarvis GA, John CM, Jack DL, Dower SK, Read RC | title = Activation of toll-like receptor 2 (TLR2) and TLR4/MD2 by Neisseria is independent of capsule and lipooligosaccharide (LOS) sialylation but varies widely among LOS from different strains | journal = Infection and Immunity | volume = 71 | issue = 7 | pages = 3901–3908 | date = July 2003 | pmid = 12819075 | pmc = 161978 | doi = 10.1128/IAI.71.7.3901-3908.2003 }}</ref><ref>{{cite journal | vauthors = Stephenson HN, John CM, Naz N, Gundogdu O, Dorrell N, Wren BW, Jarvis GA, Bajaj-Elliott M | title = Campylobacter jejuni lipooligosaccharide sialylation, phosphorylation, and amide/ester linkage modifications fine-tune human Toll-like receptor 4 activation | journal = The Journal of Biological Chemistry | volume = 288 | issue = 27 | pages = 19661–19672 | date = July 2013 | pmid = 23629657 | pmc = 3707672 | doi = 10.1074/jbc.M113.468298 | doi-access = free }}</ref><ref>{{cite journal | vauthors = Alexander-Floyd J, Bass AR, Harberts EM, Grubaugh D, Buxbaum JD, Brodsky IE, Ernst RK, Shin S | title = Lipid A Variants Activate Human TLR4 and the Noncanonical Inflammasome Differently and Require the Core Oligosaccharide for Inflammasome Activation | journal = Infection and Immunity | volume = 90 | issue = 8 | pages = e0020822 | date = August 2022 | pmid = 35862709 | pmc = 9387229 | doi = 10.1128/iai.00208-22 | veditors = Bäumler AJ | article-number = e00208-22 }}</ref><ref>{{cite journal | vauthors = Bonhomme D, Santecchia I, Vernel-Pauillac F, Caroff M, Germon P, Murray G, Adler B, Boneca IG, Werts C | title = Leptospiral LPS escapes mouse TLR4 internalization and TRIF‑associated antimicrobial responses through O antigen and associated lipoproteins | journal = PLOS Pathogens | volume = 16 | issue = 8 | article-number = e1008639 | date = August 2020 | pmid = 32790743 | pmc = 7447051 | doi = 10.1371/journal.ppat.1008639 | doi-access = free }}</ref> none|thumb|320x320px|MyD88 and TRIF dependent signaling pathway of TLR4.
=== MyD88 – dependent pathway ===
The MyD88-dependent pathway is regulated by two adaptor-associated proteins: Myeloid Differentiation Primary Response Gene 88 (MyD88) and TIR Domain-Containing Adaptor Protein (TIRAP). It also involves the activation of IL-1 Receptor-Associated Kinases (IRAKs) and the adaptor molecules TNF Receptor-Associated Factor 6 (TRAF6). TRAF6 induces the activation of TAK1 (Transforming growth factor-β-Activated Kinase 1) that leads to the activation of MAPK cascades (Mitogen-Activated Protein Kinase) and the IκB Kinases (IKK), called IKKα and IKKβ.<ref name="Pålsson-McDermott_2004">{{cite journal | vauthors = Pålsson-McDermott EM, O'Neill LA | title = Signal transduction by the lipopolysaccharide receptor, Toll-like receptor-4 | journal = Immunology | volume = 113 | issue = 2 | pages = 153–162 | date = October 2004 | pmid = 15379975 | pmc = 1782563 | doi = 10.1111/j.1365-2567.2004.01976.x }}</ref> IKKs' signaling pathway leads to the induction of the transcription factor NF-κB, while activation of MAPK cascades lead to the activation of another transcription factor AP-1.<ref name="Pålsson-McDermott_2004" /><ref name="Lu_2008">{{cite journal | vauthors = Lu YC, Yeh WC, Ohashi PS | title = LPS/TLR4 signal transduction pathway | journal = Cytokine | volume = 42 | issue = 2 | pages = 145–151 | date = May 2008 | pmid = 18304834 | doi = 10.1016/j.cyto.2008.01.006 }}</ref> These two transcription factors induce the expression of genes encoding pro-inflammatory mediators, such as tumor necrosis factor α (TNF-α), interleukin (IL)-6, and type III interferons (IFNλ1/2).<ref name="auto7">{{cite journal | vauthors = Meissner F, Scheltema RA, Mollenkopf HJ, Mann M | title = Direct proteomic quantification of the secretome of activated immune cells | journal = Science | volume = 340 | issue = 6131 | pages = 475–478 | date = April 2013 | pmid = 23620052 | doi = 10.1126/science.1232578 | bibcode = 2013Sci...340..475M | s2cid = 40513139 }}</ref><ref name="auto2">{{cite journal | vauthors = Kawai T, Takeuchi O, Fujita T, Inoue J, Mühlradt PF, Sato S, Hoshino K, Akira S | title = Lipopolysaccharide stimulates the MyD88-independent pathway and results in activation of IFN-regulatory factor 3 and the expression of a subset of lipopolysaccharide-inducible genes | journal = Journal of Immunology | volume = 167 | issue = 10 | pages = 5887–5894 | date = November 2001 | pmid = 11698465 | doi = 10.4049/jimmunol.167.10.5887 }}</ref><ref name="auto6">{{cite journal | vauthors = Chanteux H, Guisset AC, Pilette C, Sibille Y | title = LPS induces IL-10 production by human alveolar macrophages via MAPKinases- and Sp1-dependent mechanisms | journal = Respiratory Research | volume = 8 | issue = 1 | article-number = 71 | date = October 2007 | pmid = 17916230 | pmc = 2080632 | doi = 10.1186/1465-9921-8-71 | doi-access = free }}</ref>
=== TRIF – dependent pathway ===
The TRIF-dependent pathway involves the internalization of TLR4 in endosomes and the recruitment of the adaptor proteins TIR-domain-containing adaptor inducing interferon-β (TRIF) and TRIF-related Adaptor Molecule (TRAM). TRAM-TRIF signals activate the ubiquitin ligase TRAF3 followed by the activation of non-canonical IKK kinases: TANK binding kinase 1 (TBK1) and IKKε. TBK1 phosphorylates the pLxIS consensus motif of TRIF that is necessary to recruit interferon regulatory factor (IRF) 3. IRF3 is also phosphorylated by TBK1 and then dissociates from TRIF, dimerizes and translocates to the nucleus.<ref name="pmid33057840">{{cite journal | vauthors = Ciesielska A, Matyjek M, Kwiatkowska K | title = TLR4 and CD14 trafficking and its influence on LPS-induced pro-inflammatory signaling | journal = Cellular and Molecular Life Sciences | volume = 78 | issue = 4 | pages = 1233–1261 | date = February 2021 | pmid = 33057840 | pmc = 7904555 | doi = 10.1007/s00018-020-03656-y }}</ref> Finally, IRF3 induces the expression of genes encoding type I IFN such as interferon beta (IFN-β), the chemokine CCL5/ RANTES and interferon-regulated genes as that encoding the chemokine CXCL10/IP-10.<ref name="auto7"/><ref name="auto2"/><ref name="auto6"/><ref name="O'Neill _2013">{{cite journal | vauthors = O'Neill LA, Golenbock D, Bowie AG | title = The history of Toll-like receptors - redefining innate immunity | journal = Nature Reviews. Immunology | volume = 13 | issue = 6 | pages = 453–460 | date = June 2013 | pmid = 23681101 | doi = 10.1038/nri3446 | bibcode = 2013NatRI..13..453O | hdl-access = free | s2cid = 205491986 | hdl = 2262/72552 }}</ref> TRIF-dependent signaling pathway of TLR4 is known to play a central role in the stimulation of innate immune cells such as macrophages, the maturation of DCs and the induction and recruitment of Th1 adaptive immune responses.<ref>{{cite journal | vauthors = Watanabe S, Kumazawa Y, Inoue J | title = Liposomal lipopolysaccharide initiates TRIF-dependent signaling pathway independent of CD14 | journal = PLOS ONE | volume = 8 | issue = 4 | article-number = e60078 | date = 2013 | pmid = 23565187 | pmc = 3615118 | doi = 10.1371/journal.pone.0060078 | doi-access = free | bibcode = 2013PLoSO...860078W }}</ref>
== Immune cell activation == TLR4 activation by LPS enables a rapid stimulation of a wide range of innate immune cells such as macrophages and DCs. This leads to the secretion of pro-inflammatory and type I interferons cytokines, chemokines. Production levels of these cytokines/chemokines vary according to the degree of activation of the MyD88 and TRIF signaling pathways by TLR4 agonist molecules. TLR4 activation also induces the stimulation of antigen presentation and upregulation of costimulatory molecules (such as CD40, CD80 and CD86) on innate immune cells which are required for antigen presentation for T lymphocytes.<ref>{{cite journal | vauthors = Lien E, Means TK, Heine H, Yoshimura A, Kusumoto S, Fukase K, Fenton MJ, Oikawa M, Qureshi N, Monks B, Finberg RW, Ingalls RR, Golenbock DT | title = Toll-like receptor 4 imparts ligand-specific recognition of bacterial lipopolysaccharide | journal = The Journal of Clinical Investigation | volume = 105 | issue = 4 | pages = 497–504 | date = February 2000 | pmid = 10683379 | pmc = 289161 | doi = 10.1172/JCI8541 | bibcode = 2000JCliI.105..497L }}</ref><ref name="auto10">{{cite journal | vauthors = Shetab Boushehri MA, Lamprecht A | title = TLR4-Based Immunotherapeutics in Cancer: A Review of the Achievements and Shortcomings | journal = Molecular Pharmaceutics | volume = 15 | issue = 11 | pages = 4777–4800 | date = November 2018 | pmid = 30226786 | doi = 10.1021/acs.molpharmaceut.8b00691 | s2cid = 52297047 }}</ref> This explains why TLR4 activation by LPS is also known to stimulate the generation of effective adaptive immune responses and to induce their recruitment, polarization and maintenance via the panel of cytokines and chemokines produced.<ref name="auto10"/><ref name="auto9"/>
The TRIF and MyD88 signaling pathways have a different but complementary impact on immune cell activation. Macrophages stimulation has been shown to be strictly dependent on TRIF pathway activation whereas DC activation and maturation depend on both the MyD88 and TRIF pathways.<ref>{{cite journal | vauthors = Kaisho T, Takeuchi O, Kawai T, Hoshino K, Akira S | title = Endotoxin-induced maturation of MyD88-deficient dendritic cells | journal = Journal of Immunology | volume = 166 | issue = 9 | pages = 5688–5694 | date = May 2001 | pmid = 11313410 | doi = 10.4049/jimmunol.166.9.5688 }}</ref><ref>{{cite journal | vauthors = Hoebe K, Janssen EM, Kim SO, Alexopoulou L, Flavell RA, Han J, Beutler B | title = Upregulation of costimulatory molecules induced by lipopolysaccharide and double-stranded RNA occurs by Trif-dependent and Trif-independent pathways | journal = Nature Immunology | volume = 4 | issue = 12 | pages = 1223–1229 | date = December 2003 | pmid = 14625548 | doi = 10.1038/ni1010 | s2cid = 8505015 }}</ref><ref>{{cite journal | vauthors = Shen H, Tesar BM, Walker WE, Goldstein DR | title = Dual signaling of MyD88 and TRIF is critical for maximal TLR4-induced dendritic cell maturation | journal = Journal of Immunology | volume = 181 | issue = 3 | pages = 1849–1858 | date = August 2008 | pmid = 18641322 | pmc = 2507878 | doi = 10.4049/jimmunol.181.3.1849 }}</ref><ref>{{cite journal | vauthors = Trombetta ES, Ebersold M, Garrett W, Pypaert M, Mellman I | title = Activation of lysosomal function during dendritic cell maturation | journal = Science | volume = 299 | issue = 5611 | pages = 1400–1403 | date = February 2003 | pmid = 12610307 | doi = 10.1126/science.1080106 | s2cid = 46594244 }}</ref> The increased expression of costimulatory and MHC molecules is a hallmark of DC maturation required for antigen presentation by these cells.<ref>{{cite journal | vauthors = Turley SJ, Inaba K, Garrett WS, Ebersold M, Unternaehrer J, Steinman RM, Mellman I | title = Transport of peptide-MHC class II complexes in developing dendritic cells | journal = Science | volume = 288 | issue = 5465 | pages = 522–527 | date = April 2000 | pmid = 10775112 | doi = 10.1126/science.288.5465.522 }}</ref> However, significant differences were found in the signaling pathways leading to this phenomenon. In macrophages, the upregulation of costimulatory molecules depends strictly on the TRIF-dependent pathway, whereas in DC both the MyD88- and TRIF-dependent ones are involved.<ref>{{cite journal | vauthors = Kaisho T, Takeuchi O, Kawai T, Hoshino K, Akira S | title = Endotoxin-induced maturation of MyD88-deficient dendritic cells | journal = Journal of Immunology | volume = 166 | issue = 9 | pages = 5688–5694 | date = May 2001 | pmid = 11313410 | doi = 10.4049/jimmunol.166.9.5688 }}</ref><ref>{{cite journal | vauthors = Hoebe K, Janssen EM, Kim SO, Alexopoulou L, Flavell RA, Han J, Beutler B | title = Upregulation of costimulatory molecules induced by lipopolysaccharide and double-stranded RNA occurs by Trif-dependent and Trif-independent pathways | journal = Nature Immunology | volume = 4 | issue = 12 | pages = 1223–1229 | date = December 2003 | pmid = 14625548 | doi = 10.1038/ni1010 | s2cid = 8505015 }}</ref><ref name="auto9" /><ref>{{cite journal | vauthors = Trombetta ES, Ebersold M, Garrett W, Pypaert M, Mellman I | title = Activation of lysosomal function during dendritic cell maturation | journal = Science | volume = 299 | issue = 5611 | pages = 1400–1403 | date = February 2003 | pmid = 12610307 | doi = 10.1126/science.1080106 | s2cid = 46594244 }}</ref> The increased cell surface presence of the costimulatory molecules and also of MHC II is a hallmark of DC maturation required for antigen presentation by these cells.<ref>{{cite journal | vauthors = Turley SJ, Inaba K, Garrett WS, Ebersold M, Unternaehrer J, Steinman RM, Mellman I | title = Transport of peptide-MHC class II complexes in developing dendritic cells | journal = Science | volume = 288 | issue = 5465 | pages = 522–527 | date = April 2000 | pmid = 10775112 | doi = 10.1126/science.288.5465.522 }}</ref>
The activation of MyD88 and TRIF signaling pathways were also found to induce Th1 polarization of the T cells responses through DC maturation and the panel of cytokines produced.<ref name=":4">{{cite journal | vauthors = Nair-Gupta P, Baccarini A, Tung N, Seyffer F, Florey O, Huang Y, Banerjee M, Overholtzer M, Roche PA, Tampé R, Brown BD, Amsen D, Whiteheart SW, Blander JM | title = TLR signals induce phagosomal MHC-I delivery from the endosomal recycling compartment to allow cross-presentation | journal = Cell | volume = 158 | issue = 3 | pages = 506–521 | date = July 2014 | pmid = 25083866 | pmc = 4212008 | doi = 10.1016/j.cell.2014.04.054 }}</ref><ref>{{cite journal | vauthors = Han JE, Wui SR, Kim KS, Cho YJ, Cho WJ, Lee NG | title = Characterization of the structure and immunostimulatory activity of a vaccine adjuvant, de-O-acylated lipooligosaccharide | journal = PLOS ONE | volume = 9 | issue = 1 | article-number = e85838 | date = 2014-01-22 | pmid = 24465739 | pmc = 3899070 | doi = 10.1371/journal.pone.0085838 | doi-access = free | bibcode = 2014PLoSO...985838H | veditors = Shin EC }}</ref><ref>{{cite journal | vauthors = Sharif O, Bolshakov VN, Raines S, Newham P, Perkins ND | title = Transcriptional profiling of the LPS induced NF-kappaB response in macrophages | journal = BMC Immunology | volume = 8 | issue = 1 | article-number = 1 | date = January 2007 | pmid = 17222336 | pmc = 1781469 | doi = 10.1186/1471-2172-8-1 | doi-access = free }}</ref> Low activation of MYD88 pathway is however important for effective cytotoxic T-cell differentiation by facilitating fusion of MHC I-bearing recycling endosomes with phagosomes allowing cross-presentation of antigens.<ref name=":4" /> In contrast, robust activation of MYD88 pathway induces excessive production of pro-inflammatory cytokines leading to life-threatening pathological consequences such as cytokine storms.
The impact of TLR4 activation on the innate and adaptive immune system explains why TLR4 agonists, such as LPS derivatives, have been developed as vaccine adjuvants. Among them is GSK's Monophosphorylated Lipid A (MPL), a detoxified Lipid A derived from Salmonella LPS, which is the first and only natural immunostimulant to have been approved as adjuvant in five human vaccines.<ref>{{cite journal | vauthors = Paavonen J, Jenkins D, Bosch FX, Naud P, Salmerón J, Wheeler CM, Chow SN, Apter DL, Kitchener HC, Castellsague X, de Carvalho NS, Skinner SR, Harper DM, Hedrick JA, Jaisamrarn U, Limson GA, Dionne M, Quint W, Spiessens B, Peeters P, Struyf F, Wieting SL, Lehtinen MO, Dubin G | title = Efficacy of a prophylactic adjuvanted bivalent L1 virus-like-particle vaccine against infection with human papillomavirus types 16 and 18 in young women: an interim analysis of a phase III double-blind, randomised controlled trial | journal = Lancet | volume = 369 | issue = 9580 | pages = 2161–2170 | date = June 2007 | pmid = 17602732 | doi = 10.1016/S0140-6736(07)60946-5 | s2cid = 26318328 }}</ref><ref>{{cite journal | vauthors = Kundi M | title = New hepatitis B vaccine formulated with an improved adjuvant system | journal = Expert Review of Vaccines | volume = 6 | issue = 2 | pages = 133–140 | date = April 2007 | pmid = 17408363 | doi = 10.1586/14760584.6.2.133 | s2cid = 35472093 }}</ref><ref>{{cite journal | vauthors = Garçon N, Di Pasquale A | title = From discovery to licensure, the Adjuvant System story | journal = Human Vaccines & Immunotherapeutics | volume = 13 | issue = 1 | pages = 19–33 | date = January 2017 | pmid = 27636098 | pmc = 5287309 | doi = 10.1080/21645515.2016.1225635 }}</ref>
== Evolutionary history and polymorphism == TLR4 originated when TLR2 and TLR4 diverged about 500 million years ago near the beginning of vertebrate evolution.<ref>{{cite book | vauthors = Beutler B, Rehli M | title = Toll-Like Receptor Family Members and Their Ligands | chapter = Evolution of the TIR, Tolls and TLRS: Functional Inferences from Computational Biology | series = Current Topics in Microbiology and Immunology | volume = 270 | pages = 1–21 | year = 2002 | pmid = 12467241 | doi = 10.1007/978-3-642-59430-4_1 | isbn = 978-3-642-63975-3 }}</ref> Sequence alignments of human and great ape TLR4 exons have demonstrated that not much evolution has occurred in human TLR4 since our divergence from our last common ancestor with chimpanzees; human and chimp TLR4 exons only differ by three substitutions while humans and baboons are 93.5% similar in the extracellular domain.<ref>{{cite journal | vauthors = Smirnova I, Poltorak A, Chan EK, McBride C, Beutler B | title = Phylogenetic variation and polymorphism at the toll-like receptor 4 locus (TLR4) | journal = Genome Biology | volume = 1 | issue = 1 | pages = RESEARCH002 | year = 2000 | article-number = research002.1 | pmid = 11104518 | pmc = 31919 | doi = 10.1186/gb-2000-1-1-research002 | doi-access = free }}</ref> Notably, humans possess a greater number of early stop codons in TLR4 than great apes; in a study of 158 humans worldwide, 0.6% had a nonsense mutation.<ref>{{cite journal | vauthors = Quach H, Wilson D, Laval G, Patin E, Manry J, Guibert J, Barreiro LB, Nerrienet E, Verschoor E, Gessain A, Przeworski M, Quintana-Murci L | title = Different selective pressures shape the evolution of Toll-like receptors in human and African great ape populations | journal = Human Molecular Genetics | volume = 22 | issue = 23 | pages = 4829–4840 | date = December 2013 | pmid = 23851028 | pmc = 3820138 | doi = 10.1093/hmg/ddt335 }}</ref><ref name="Barreiro_2009">{{cite journal | vauthors = Barreiro LB, Ben-Ali M, Quach H, Laval G, Patin E, Pickrell JK, Bouchier C, Tichit M, Neyrolles O, Gicquel B, Kidd JR, Kidd KK, Alcaïs A, Ragimbeau J, Pellegrini S, Abel L, Casanova JL, Quintana-Murci L | title = Evolutionary dynamics of human Toll-like receptors and their different contributions to host defense | journal = PLOS Genetics | volume = 5 | issue = 7 | article-number = e1000562 | date = July 2009 | pmid = 19609346 | pmc = 2702086 | doi = 10.1371/journal.pgen.1000562 | doi-access = free }}</ref> This suggests that there are weaker evolutionary pressures on the human TLR4 than on our primate relatives. The distribution of human TLR4 polymorphisms matches the out-of-Africa migration, and it is likely that the polymorphisms were generated in Africa before migration to other continents.<ref name="Barreiro_2009" /><ref>{{cite journal | vauthors = Plantinga TS, Ioana M, Alonso S, Izagirre N, Hervella M, Joosten LA, van der Meer JW, de la Rúa C, Netea MG | title = The evolutionary history of TLR4 polymorphisms in Europe | journal = Journal of Innate Immunity | volume = 4 | issue = 2 | pages = 168–175 | year = 2012 | pmid = 21968286 | pmc = 6741577 | doi = 10.1159/000329492 }}</ref>
Various single nucleotide polymorphisms (SNPs) of TLR4 have been identified in humans . For some of them, an association with increased susceptibility to Gram-negative bacterial infections or faster progression and a more severe course of sepsis in critically ill patients was reported.However, they are very rare, and their frequency varies according to ethnic origin. The 2 predominant SNPs are Asp299Gly and Thr399Ile, with a frequency of <10% in the Caucasian population and even lower in the Asian population.<ref>{{cite journal | vauthors = Noreen M, Shah MA, Mall SM, Choudhary S, Hussain T, Ahmed I, Jalil SF, Raza MI | title = TLR4 polymorphisms and disease susceptibility | journal = Inflammation Research | volume = 61 | issue = 3 | pages = 177–188 | date = March 2012 | pmid = 22277994 | doi = 10.1007/s00011-011-0427-1 | s2cid = 9500302 }}</ref> These two SNPs are missense mutations, thus associated with a loss of function, which may explain their negative impact on infection control. Studies have indeed shown that TLR4 D299G SNP limits the response to LPS by compromising MyD88 and TRIF recruitment to TLR4, and thus cytokine secretion, but without affecting TLR4 expression <ref>{{cite journal | vauthors = Long H, O'Connor BP, Zemans RL, Zhou X, Yang IV, Schwartz DA | title = The Toll-like receptor 4 polymorphism Asp299Gly but not Thr399Ile influences TLR4 signaling and function | journal = PLOS ONE | volume = 9 | issue = 4 | article-number = e93550 | date = 2014-04-02 | pmid = 24695807 | pmc = 3973565 | doi = 10.1371/journal.pone.0093550 | doi-access = free | bibcode = 2014PLoSO...993550L }}</ref><ref name="pmid22474023">{{cite journal | vauthors = Figueroa L, Xiong Y, Song C, Piao W, Vogel SN, Medvedev AE | title = The Asp299Gly polymorphism alters TLR4 signaling by interfering with recruitment of MyD88 and TRIF | journal = Journal of Immunology | volume = 188 | issue = 9 | pages = 4506–4515 | date = May 2012 | pmid = 22474023 | pmc = 3531971 | doi = 10.4049/jimmunol.1200202 }}</ref> Structural analyses of human TLR4 with SNP D299G suggest that this amino acid change affects van der Waals interaction and hydrogen bonding in leucine-rich repeats, modulating its surface properties which may affect LPS ligand binding to TLR4.<ref>{{cite journal | vauthors = Ohto U, Yamakawa N, Akashi-Takamura S, Miyake K, Shimizu T | title = Structural analyses of human Toll-like receptor 4 polymorphisms D299G and T399I | journal = The Journal of Biological Chemistry | volume = 287 | issue = 48 | pages = 40611–40617 | date = November 2012 | pmid = 23055527 | pmc = 3504774 | doi = 10.1074/jbc.M112.404608 | doi-access = free }}</ref>
== Clinical significance==
TLR4 has been reported to play both friend and foe in a variety of human diseases, such as bacterial infections and cancers. This dual role of TLR4 depends on the intensity, duration and site (surface or endosome) of its activation, its polymorphism and the balance of activation of signaling pathways (MyD88 ''vs''. TRIF).
=== Infectious diseases === TLR4 play a central role in the control of bacterial infections through the recognition of LPS molecules from gram-negative, and some gram-positive, bacteria.<ref>{{cite journal | vauthors = Akira S, Takeda K | title = Toll-like receptor signalling | journal = Nature Reviews. Immunology | volume = 4 | issue = 7 | pages = 499–511 | date = July 2004 | pmid = 15229469 | doi = 10.1038/nri1391 }}</ref> During infections, TLR4s on innate immunity cells are activated by LPS molecules present in tissues and the bloodstream. This activates innate immunity, the first line of defense against invading microorganisms, and triggers pro-inflammatory responses that facilitate the eradication of invading bacteria.<ref name="auto1"/> Generally, inflammation has a protective role. It is a complex and coordinated process followed by the induction of resolution pathways that restore tissue integrity and function. However, in some cases, exaggerated and uncontrolled inflammation triggered by TLR4 during infection can lead to sepsis and septic shock.<ref name="pmid33057840"/> Infections with Gram-negative bacteria such as ''Escherichia coli'' and ''Pseudomonas aeruginosa'' are the prevailing causes of severe sepsis in humans.Some studies have linked TLR4 polymorphisms (Asp299Gly and Thr399Ile SNPs) to an increased susceptibility to sepsis due to gram-negative infection but other studies failed to confirm this.<ref>{{cite journal | vauthors = Netea MG, Wijmenga C, O'Neill LA | title = Genetic variation in Toll-like receptors and disease susceptibility | journal = Nature Immunology | volume = 13 | issue = 6 | pages = 535–542 | date = May 2012 | pmid = 22610250 | doi = 10.1038/ni.2284 | s2cid = 24438756 }}</ref>
=== Cancer ===
The role of the TLR4 in the control of cancer progression and in cancer therapy is well documented.
Stimulation of TLR4 by natural derivatives and LPS is well known to induce potent antitumor activity. This anti-tumor activity is linked to the ability of LPS to stimulate innate immunity via TLR4, resulting in the production of pro-inflammatory cytokines and type 1 interferons, and the indirect generation of adaptive anti-tumor responses.<ref>{{cite journal | vauthors = Chettab K, Fitzsimmons C, Novikov A, Denis M, Phelip C, Mathé D, Choffour PA, Beaumel S, Fourmaux E, Norca P, Kryza D, Evesque A, Jordheim LP, Perrial E, Matera EL, Caroff M, Kerzerho J, Dumontet C | title = A systemically administered detoxified TLR4 agonist displays potent antitumor activity and an acceptable tolerance profile in preclinical models | journal = Frontiers in Immunology | volume = 14 | article-number = 1066402 | date = 2023 | pmid = 37223101 | pmc = 10200957 | doi = 10.3389/fimmu.2023.1066402 | doi-access = free }}</ref><ref>{{cite journal | vauthors = Richert I, Berchard P, Abbes L, Novikov A, Chettab K, Vandermoeten A, Dumontet C, Karanian M, Kerzerho J, Caroff M, Blay JY, Dutour A | title = A TLR4 Agonist Induces Osteosarcoma Regression by Inducing an Antitumor Immune Response and Reprogramming M2 Macrophages to M1 Macrophages | journal = Cancers | volume = 15 | issue = 18 | page = 4635 | date = September 2023 | pmid = 37760603 | pmc = 10526955 | doi = 10.3390/cancers15184635 | doi-access = free }}</ref>
The first clues about the efficacy of TLR4 agonists like LPS in cancer immunotherapy was found in the 19th centuries, when bacterial infections were found to induce tumor regressions.<ref>{{cite journal | vauthors = Maruyama K, Selmani Z, Ishii H, Yamaguchi K | title = Innate immunity and cancer therapy | journal = International Immunopharmacology | volume = 11 | issue = 3 | pages = 350–357 | date = March 2011 | pmid = 20955832 | doi = 10.1016/j.intimp.2010.09.012 }}</ref> Later, Dr William Coley showed the therapeutic efficacy of a mixed bacterial vaccine, so-called "Coley's toxin", to human cancer.<ref>{{cite journal | vauthors = Starnes CO | title = Coley's toxins in perspective | journal = Nature | volume = 357 | issue = 6373 | pages = 11–12 | date = May 1992 | pmid = 1574121 | doi = 10.1038/357011a0 | s2cid = 4265230 | bibcode = 1992Natur.357...11S }}</ref> Since then, a number of developments have been made in the treatment or prevention of cancer using bacterial mixtures strongly activating TLR4 due to LPS content. The antituberculosis vaccine Bacillus Calmette–Guérin (BCG) was approved by the Federal Drug Administration (FDA) in 1990 for the local treatment of superficial bladder cancer. BCG promotes dendritic cell maturation, and this effect is TLR4 (as well as TLR2) dependent.<ref>{{cite journal | vauthors = Tsuji S, Matsumoto M, Takeuchi O, Akira S, Azuma I, Hayashi A, Toyoshima K, Seya T | title = Maturation of human dendritic cells by cell wall skeleton of Mycobacterium bovis bacillus Calmette-Guérin: involvement of toll-like receptors | journal = Infection and Immunity | volume = 68 | issue = 12 | pages = 6883–6890 | date = December 2000 | pmid = 11083809 | pmc = 97794 | doi = 10.1128/IAI.68.12.6883-6890.2000 | veditors = Kaufmann SH }}</ref> There are also reports on the treatment of oral squamous cell carcinoma, gastric , Head-and-neck and cervical cancers with lyophilized streptococcal preparation OK-432 (Picibanil).<ref>{{cite journal | vauthors = Ryoma Y, Moriya Y, Okamoto M, Kanaya I, Saito M, Sato M | title = Biological effect of OK-432 (picibanil) and possible application to dendritic cell therapy | journal = Anticancer Research | volume = 24 | issue = 5C | pages = 3295–3301 | date = 2004-09-01 | pmid = 15515424 | url = https://ar.iiarjournals.org/content/24/5C/3295 }}</ref> The mechanism of action of OK-432 involves TLR4 activation, since OKA-432 does not inhibit tumor growth on TLR4 knockouts as it does on wild-type mice.<ref name="Okamoto_2004">{{cite journal | vauthors = Okamoto M, Oshikawa T, Tano T, Ohe G, Furuichi S, Nishikawa H, Ahmed SU, Akashi S, Miyake K, Takeuchi O, Akira S, Moriya Y, Matsubara S, Ryoma Y, Saito M, Sato M | title = Involvement of Toll-like receptor 4 signaling in interferon-gamma production and antitumor effect by streptococcal agent OK-432 | journal = Journal of the National Cancer Institute | volume = 95 | issue = 4 | pages = 316–326 | date = February 2003 | pmid = 12591988 | doi = 10.1093/jnci/95.4.316 }}</ref>
Purified LPS also showed potent anti-tumor efficacy as systemic therapeutic agents in several tumor models.<ref>{{cite journal | vauthors = Shear MB, Perrault M |title=Chemical Treatment of Tumors. IX. Reactions of Mice with Primary Subcutaneous Tumors to Injection of a Hemorrhage-Producing Bacterial Polysaccharide1 |journal=JNCI: Journal of the National Cancer Institute |date=April 1944 | volume = 4 | issue = 5 | pages = 461–476 |doi=10.1093/jnci/4.5.461}}</ref><ref>{{cite journal | vauthors = Berendt MJ, North RJ, Kirstein DP | title = The immunological basis of endotoxin-induced tumor regression. Requirement for a pre-existing state of concomitant anti-tumor immunity | journal = The Journal of Experimental Medicine | volume = 148 | issue = 6 | pages = 1560–1569 | date = December 1978 | pmid = 309922 | pmc = 2185097 | doi = 10.1084/jem.148.6.1560 }}</ref> In the 90's, clinical trials evaluating the intravenous administration of LPS to patients with cancer provided positive results including several cases of disease stabilization and partial responses. However, limiting toxicities at doses in the ng/kg range has been reported which are too low to obtain significant antitumor effects.<ref>{{cite journal | vauthors = Engelhardt R, Mackensen A, Galanos C | title = Phase I trial of intravenously administered endotoxin (Salmonella abortus equi) in cancer patients | journal = Cancer Research | volume = 51 | issue = 10 | pages = 2524–2530 | date = May 1991 | pmid = 2021932 }}</ref>
Subsequently, detoxified TLR4 agonists (LPS derivatives) have been produced and evaluated in the clinic. This includes the MPL, a chemically modified LPS which was the first TLR4 agonist to be approved and commercialized by GSK in 5 human vaccines (HPV, Zoster, Hepatitis B, Malaria, RSV). MPL was investigated as an adjuvant for curative anti-tumor vaccines, with the approval of Melacine in Canada for the treatment of patients with melanoma.<ref>{{cite journal | vauthors = Koller KM, Wang W, Schell TD, Cozza EM, Kokolus KM, Neves RI, Mackley HB, Pameijer C, Leung A, Anderson B, Mallon CA, Robertson G, Drabick JJ | title = Malignant melanoma-The cradle of anti-neoplastic immunotherapy | journal = Critical Reviews in Oncology/Hematology | volume = 106 | pages = 25–54 | date = October 2016 | pmid = 27637351 | doi = 10.1016/j.critrevonc.2016.04.010 }}</ref> Synthetic LPS derivatives based on dephosphorylated lipid A moiety structures were also developed and confirmed potent adjuvant and antitumor activities as therapeutic agents. In particular, the intratumoral administration of Glucopyranosyl Lipid Adjuvant (GLA-SE/G100), a synthetic detoxified analog of lipid A formulated in a stable emulsion, showed anti-tumor immune responses and tumor regression in patients with Merkel cell carcinoma,<ref>{{cite journal | vauthors = Bhatia S, Miller NJ, Lu H, Longino NV, Ibrani D, Shinohara MM, Byrd DR, Parvathaneni U, Kulikauskas R, Ter Meulen J, Hsu FJ, Koelle DM, Nghiem P | title = Intratumoral G100, a TLR4 Agonist, Induces Antitumor Immune Responses and Tumor Regression in Patients with Merkel Cell Carcinoma | journal = Clinical Cancer Research | volume = 25 | issue = 4 | pages = 1185–1195 | date = February 2019 | pmid = 30093453 | pmc = 6368904 | doi = 10.1158/1078-0432.CCR-18-0469 }}</ref> and potent adjuvant activity in phase 2 trials in combination with pembrolizumab in patients with follicular lymphoma.<ref>{{cite journal | vauthors = Halwani AS, Panizo C, Isufi I, Herrera AF, Okada CY, Cull EH, Kis B, Chaves JM, Bartlett NL, Ai W, de la Cruz-Merino L, Bryan LJ, Houot R, Linton K, Briones J, Chau I, von Keudell GR, Lu H, Yakovich A, Chen M, Meulen Jh T, Yurasov S, Hsu FJ, Flowers CR | title = Phase 1/2 study of intratumoral G100 (TLR4 agonist) with or without pembrolizumab in follicular lymphoma | journal = Leukemia & Lymphoma | volume = 63 | issue = 4 | pages = 821–833 | date = April 2022 | pmid = 34865586 | doi = 10.1080/10428194.2021.2010057 | s2cid = 244943266 }}</ref><ref name="auto5">{{Cite journal | vauthors = Flowers C, Panizo C, Isufi I, Herrera AF, Okada C, Cull EH, Kis B, Chaves JM, Bartlett NL, Ai W, de la Cruz-Merino L |date=2017-12-08 |title=Intratumoral G100 Induces Systemic Immunity and Abscopal Tumor Regression in Patients with Follicular Lymphoma: Results of a Phase 1/ 2 Study Examining G100 Alone and in Combination with Pembrolizumab |url=https://ashpublications.org/blood/article/130/Supplement%201/2771/80455 |journal=Blood |volume=130 |page=2771 |doi=10.1182/blood.V130.Suppl_1.2771.2771 |doi-broken-date=12 July 2025 }}</ref>
Besides the recognized anti-tumor efficacy of TLR4 activation by LPS, some studies suggest that TLR4 may also contribute to the development of some cancers, (prostate, liver, breast and lung cancers) and may contribute to resistance to paclitaxel chemotherapy in breast cancer.<ref>{{cite journal | vauthors = Rajput S, Volk-Draper LD, Ran S | title = TLR4 is a novel determinant of the response to paclitaxel in breast cancer | journal = Molecular Cancer Therapeutics | volume = 12 | issue = 8 | pages = 1676–1687 | date = August 2013 | pmid = 23720768 | pmc = 3742631 | doi = 10.1158/1535-7163.MCT-12-1019 }}</ref> Some clinical studies also suggested a potential correlation between TLR4 expression on tumor cells and tumor progression. However, no such effect was reported in the numerous clinical studies conducted with natural LPS or LPS derivatives. On the contrary, in phase 2 studies with GLA, a positive association between baseline TLR4 expression in tumors and the increase of overall response rates has been reported.<ref name="auto5"/>
The potential impact of TLR4 on the progression of some cancers was associated with the excessive production of pro-inflammatory cytokines via activation of the TLR4-MyD88/NF-kB signaling pathway.<ref>{{cite journal | vauthors = Zhang R, Zhao J, Xu J, Jiao DX, Wang J, Gong ZQ, Jia JH | title = Andrographolide suppresses proliferation of human colon cancer SW620 cells through the TLR4/NF-κB/MMP-9 signaling pathway | journal = Oncology Letters | volume = 14 | issue = 4 | pages = 4305–4310 | date = October 2017 | pmid = 28943944 | pmc = 5604146 | doi = 10.3892/ol.2017.6669 }}</ref><ref>{{cite journal | vauthors = Wang CH, Wang PJ, Hsieh YC, Lo S, Lee YC, Chen YC, Tsai CH, Chiu WC, Chu-Sung Hu S, Lu CW, Yang YF, Chiu CC, Ou-Yang F, Wang YM, Hou MF, Yuan SS | title = Resistin facilitates breast cancer progression via TLR4-mediated induction of mesenchymal phenotypes and stemness properties | journal = Oncogene | volume = 37 | issue = 5 | pages = 589–600 | date = February 2018 | pmid = 28991224 | doi = 10.1038/onc.2017.357 | s2cid = 24926622 }}</ref><ref>{{cite journal | vauthors = Kelly MG, Alvero AB, Chen R, Silasi DA, Abrahams VM, Chan S, Visintin I, Rutherford T, Mor G | title = TLR-4 signaling promotes tumor growth and paclitaxel chemoresistance in ovarian cancer | journal = Cancer Research | volume = 66 | issue = 7 | pages = 3859–3868 | date = April 2006 | pmid = 16585214 | doi = 10.1158/0008-5472.CAN-05-3948 }}</ref> Several studies showed that this is mediated by the misuse of DAMP signaling by tumor cells.<ref name=":3"/><ref>{{cite journal | vauthors = Khademalhosseini M, Arababadi MK | title = Toll-like receptor 4 and breast cancer: an updated systematic review | journal = Breast Cancer | volume = 26 | issue = 3 | pages = 265–271 | date = May 2019 | pmid = 30543015 | doi = 10.1007/s12282-018-00935-2 | s2cid = 56143069 }}</ref><ref name="auto"/>
Many DAMPs are released by dying or necrotic tumor cells and present during cancer progression. DAMPs released from tumor cells can directly activate tumor-expressed TLR4 that induce chemoresistance, migration, invasion, and metastasis. Furthermore, DAMP-induced chronic inflammation in the tumor microenvironment causes an increase in immunosuppressive populations, such as M2 macrophages, myeloid-derived suppressor cells (MDSCs), and regulatory T cells (Tregs).<ref name=":3"/> DAMPs, such as HMGB1, S100 proteins, and heat shock proteins (HSPs), were found to strongly activate inflammatory pathways and release IL-1, IL-6, LT-β, IFN-γ, TNF, and transforming growth factor (TGF)-β promoting inflammation, immunosuppression, angiogenesis, and tumor cell proliferation.<ref name=":2"/>
Several studies have evaluated the potential association of this TLR4 polymorphism with cancer risk, but the data are highly conflicting. However, some meta-analyses suggest an association of SNP D299G with gastric, viral-induced and female-specific cancers (cervix, ovary).<ref>{{cite journal | vauthors = Zhu L, Yuan H, Jiang T, Wang R, Ma H, Zhang S | title = Association of TLR2 and TLR4 polymorphisms with risk of cancer: a meta-analysis | journal = PLOS ONE | volume = 8 | issue = 12 | article-number = e82858 | date = 2013-12-20 | pmid = 24376595 | pmc = 3869723 | doi = 10.1371/journal.pone.0082858 | doi-access = free | bibcode = 2013PLoSO...882858Z }}</ref>
=== Neurogenerative diseases === Growing evidence suggests an implication of TLR4 in the development and progression of neurogenerative disorders such as Alzheimer's disease, Parkinson's disease, and Huntington's disease. In the brain, TLR4 is expressed by neurons as well as the non-neuronal glial cells, which include microglia, astrocytes, and oligodendrocytes. TLR4 is expressed primarily by microglia, and to a lesser extent by astrocytes, oligodendrocytes, and neurons.<ref name="pmid25071777"/> Microglia are representatives of the mononuclear phagocyte system in the brain, and TLR4 activation regulates some of their functions, such as phagocytic activity.<ref>{{cite journal | vauthors = Wardill HR, Van Sebille YZ, Mander KA, Gibson RJ, Logan RM, Bowen JM, Sonis ST | title = Toll-like receptor 4 signaling: a common biological mechanism of regimen-related toxicities: an emerging hypothesis for neuropathy and gastrointestinal toxicity | journal = Cancer Treatment Reviews | volume = 41 | issue = 2 | pages = 122–128 | date = February 2015 | pmid = 25512119 | doi = 10.1016/j.ctrv.2014.11.005 }}</ref><ref name="auto1"/>
Activation of microglial TLR4 has been suggested to protect against or slow the development of neurodegenerative diseases, notably by enhancing the clearance of neurotoxic proteins such as Aβ and its aggregates, thanks to increased phagocytic and autophagic activity.<ref>{{cite journal | vauthors = Tahara K, Kim HD, Jin JJ, Maxwell JA, Li L, Fukuchi K | title = Role of toll-like receptor signalling in Abeta uptake and clearance | journal = Brain | volume = 129 | issue = Pt 11 | pages = 3006–3019 | date = November 2006 | pmid = 16984903 | pmc = 2445613 | doi = 10.1093/brain/awl249 }}</ref>
However, chronic TLR4 activation is believed to be associated with glia-mediated neuronal death due to excessive secretion of pro-inflammatory cytotoxins leading to neuroinflammation, a key factor in the development of many neurodegenerative diseases.<ref name="auto3">{{cite journal | vauthors = Buchanan MM, Hutchinson M, Watkins LR, Yin H | title = Toll-like receptor 4 in CNS pathologies | journal = Journal of Neurochemistry | volume = 114 | issue = 1 | pages = 13–27 | date = July 2010 | pmid = 20402965 | pmc = 2909662 | doi = 10.1111/j.1471-4159.2010.06736.x }}</ref><ref>{{cite journal | vauthors = Qin Y, Liu Y, Hao W, Decker Y, Tomic I, Menger MD, Liu C, Fassbender K | title = Stimulation of TLR4 Attenuates Alzheimer's Disease-Related Symptoms and Pathology in Tau-Transgenic Mice | journal = Journal of Immunology | volume = 197 | issue = 8 | pages = 3281–3292 | date = October 2016 | pmid = 27605009 | doi = 10.4049/jimmunol.1600873 }}</ref> In the brain, TLR4 can be activated by various endogenous DAMPs in addition to pathology-associated proteins such as aggregates of amyloid-βpeptides (Aβ) or α-synuclein.<ref>{{cite journal | vauthors = Gambuzza M, Licata N, Palella E, Celi D, Foti Cuzzola V, Italiano D, Marino S, Bramanti P | title = Targeting Toll-like receptors: emerging therapeutics for multiple sclerosis management | journal = Journal of Neuroimmunology | volume = 239 | issue = 1–2 | pages = 1–12 | date = October 2011 | pmid = 21889214 | doi = 10.1016/j.jneuroim.2011.08.010 | s2cid = 3277551 }}</ref> All these structures bind TLR4 and activate downstream signaling pathways in glia, inducing secretion of reactive oxygen species (ROS) and proinflammatory cytokines such as IL-1β and TNF-α, which can lead to damage and death of neurons.<ref name="auto3"/><ref>{{cite journal | vauthors = Rannikko EH, Weber SS, Kahle PJ | title = Exogenous α-synuclein induces toll-like receptor 4 dependent inflammatory responses in astrocytes | journal = BMC Neuroscience | volume = 16 | article-number = 57 | date = September 2015 | pmid = 26346361 | pmc = 4562100 | doi = 10.1186/s12868-015-0192-0 | doi-access = free }}</ref><ref>{{cite journal | vauthors = Walter S, Letiembre M, Liu Y, Heine H, Penke B, Hao W, Bode B, Manietta N, Walter J, Schulz-Schuffer W, Fassbender K | title = Role of the toll-like receptor 4 in neuroinflammation in Alzheimer's disease | journal = Cellular Physiology and Biochemistry | volume = 20 | issue = 6 | pages = 947–956 | date = 2007 | pmid = 17982277 | doi = 10.1159/000110455 | s2cid = 6752610 }}</ref> Neuronal death is accompanied by the release of DAMPs into the extracellular space, which can then further activate TLR4, aggravating neuroinflammation.<ref>{{cite journal | vauthors = Land WG | title = The Role of Damage-Associated Molecular Patterns in Human Diseases: Part I - Promoting inflammation and immunity | journal = Sultan Qaboos University Medical Journal | volume = 15 | issue = 1 | pages = e9–e21 | date = February 2015 | pmid = 25685392 | pmc = 4318613 }}</ref> In patients with Alzheimer's disease (AD), the levels of circulating DAMPs like HMGB1 and soluble RAGE, are significantly elevated, which was correlated with the levels of amyloid beta.<ref>{{cite journal | vauthors = Festoff BW, Sajja RK, van Dreden P, Cucullo L | title = HMGB1 and thrombin mediate the blood-brain barrier dysfunction acting as biomarkers of neuroinflammation and progression to neurodegeneration in Alzheimer's disease | journal = Journal of Neuroinflammation | volume = 13 | issue = 1 | article-number = 194 | date = August 2016 | pmid = 27553758 | pmc = 4995775 | doi = 10.1186/s12974-016-0670-z | doi-access = free }}</ref> In AD patients, the serum levels of S100B are also intimately related to the severity of the disease.<ref>{{cite journal | vauthors = Chaves ML, Camozzato AL, Ferreira ED, Piazenski I, Kochhann R, Dall'Igna O, Mazzini GS, Souza DO, Portela LV | title = Serum levels of S100B and NSE proteins in Alzheimer's disease patients | journal = Journal of Neuroinflammation | volume = 7 | page = 6 | date = January 2010 | pmid = 20105309 | pmc = 2832635 | doi = 10.1186/1742-2094-7-6 | doi-access = free }}</ref> The role of the HMGB1-TLR4 axis is very important in the pathogenesis of Parkinson's disease (PD). The serum HMGB1 and TLR4 protein levels were significantly elevated in PD patients and correlated with the PD stages.<ref>{{cite journal | vauthors = Yang Y, Han C, Guo L, Guan Q | title = High expression of the HMGB1-TLR4 axis and its downstream signaling factors in patients with Parkinson's disease and the relationship of pathological staging | journal = Brain and Behavior | volume = 8 | issue = 4 | article-number = e00948 | date = April 2018 | pmid = 29670828 | pmc = 5893335 | doi = 10.1002/brb3.948 }}</ref>
Targeting TLR4 with agonists or antagonists, or modulating its downstream signaling pathways, may have a therapeutic potential in treating neurodegenerative diseases.<ref>{{cite journal | vauthors = Wu L, Xian X, Xu G, Tan Z, Dong F, Zhang M, Zhang F | title = Toll-Like Receptor 4: A Promising Therapeutic Target for Alzheimer's Disease | journal = Mediators of Inflammation | volume = 2022 | article-number = 7924199 | date = 2022-08-21 | pmid = 36046763 | pmc = 9420645 | doi = 10.1155/2022/7924199 | doi-access = free }}</ref> TLR4-specific antagonists could suppress neuroinflammation by reducing overproduction of inflammatory mediators and cytotoxins by glia. However, TLR4 antagonists could have adverse CNS effects by inhibiting phagocytosis by glia, reducing protein clearance, and interfering with myelination.<ref name="auto8">{{cite journal | vauthors = Leitner GR, Wenzel TJ, Marshall N, Gates EJ, Klegeris A | title = Targeting toll-like receptor 4 to modulate neuroinflammation in central nervous system disorders | journal = Expert Opinion on Therapeutic Targets | volume = 23 | issue = 10 | pages = 865–882 | date = October 2019 | pmid = 31580163 | doi = 10.1080/14728222.2019.1676416 | s2cid = 203652175 }}</ref> Some studies showed that selective TLR4 agonists could be beneficial by upregulating the phagocytic activity of microglia, leading to enhanced clearance of damaged tissue and abnormal protein aggregates associated with several different CNS diseases. Repeated injections of MPL, at doses that are nonpyrogenic, were found to significantly improved AD-related pathology mice.<ref>{{cite journal | vauthors = Michaud JP, Hallé M, Lampron A, Thériault P, Préfontaine P, Filali M, Tribout-Jover P, Lanteigne AM, Jodoin R, Cluff C, Brichard V, Palmantier R, Pilorget A, Larocque D, Rivest S | title = Toll-like receptor 4 stimulation with the detoxified ligand monophosphoryl lipid A improves Alzheimer's disease-related pathology | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 110 | issue = 5 | pages = 1941–1946 | date = January 2013 | pmid = 23322736 | pmc = 3562771 | doi = 10.1073/pnas.1215165110 | doi-access = free | bibcode = 2013PNAS..110.1941M }}</ref> MPL led to a significant reduction in Aβ load in the brain, as well as enhanced cognitive function. MPL induced a potent phagocytic response by microglia while triggering a moderate inflammatory reaction. However, adverse effects can be caused by TLR 4 agonists inducing secretion of inflammatory mediators. Studies therefore suggested that TLR4 agonists that selectively activate the TRIF signaling pathway could be highly beneficial in the treatment of neurodegenerative disorders by increasing glial cell phagocytic activity without significantly increasing glial cytokines and cytotoxins.<ref name="auto8"/>
== Drugs targeting TLR4 ==
=== Opioids === TLR4 has been shown to be important for the long-term side-effects of opioid analgesic drugs. Various μ-opioid receptor ligands have been tested and found to also possess action as agonists or antagonists of TLR4, with opioid agonists such as (+)-morphine being TLR4 agonists, while opioid antagonists such as naloxone were found to be TLR4 antagonists. Activation of TLR4 leads to downstream release of inflammatory modulators including TNF-α and Interleukin-1, and constant low-level release of these modulators is thought to reduce the efficacy of opioid drug treatment with time, and be involved in both the development of tolerance to opioid analgesic drugs,<ref name="pmid15836969">{{cite journal | vauthors = Shavit Y, Wolf G, Goshen I, Livshits D, Yirmiya R | title = Interleukin-1 antagonizes morphine analgesia and underlies morphine tolerance | journal = Pain | volume = 115 | issue = 1–2 | pages = 50–59 | date = May 2005 | pmid = 15836969 | doi = 10.1016/j.pain.2005.02.003 | s2cid = 7286123 }}</ref><ref name="pmid20615556">{{cite journal | vauthors = Mohan S, Davis RL, DeSilva U, Stevens CW | title = Dual regulation of mu opioid receptors in SK-N-SH neuroblastoma cells by morphine and interleukin-1β: evidence for opioid-immune crosstalk | journal = Journal of Neuroimmunology | volume = 227 | issue = 1–2 | pages = 26–34 | date = October 2010 | pmid = 20615556 | pmc = 2942958 | doi = 10.1016/j.jneuroim.2010.06.007 }}</ref> and in the emergence of side-effects such as hyperalgesia and allodynia that can become a problem following extended use of opioid drugs.<ref name="pmid19607972">{{cite book | vauthors = Komatsu T, Sakurada S, Katsuyama S, Sanai K, Sakurada T | title = Mechanism of allodynia evoked by intrathecal morphine-3-glucuronide in mice | volume = 85 | pages = 207–19 | year = 2009 | pmid = 19607972 | doi = 10.1016/S0074-7742(09)85016-2 | isbn = 978-0-12-374893-5 | series = International Review of Neurobiology }}</ref><ref name="pmid19833175">{{cite journal | vauthors = Lewis SS, Hutchinson MR, Rezvani N, Loram LC, Zhang Y, Maier SF, Rice KC, Watkins LR | title = Evidence that intrathecal morphine-3-glucuronide may cause pain enhancement via toll-like receptor 4/MD-2 and interleukin-1beta | journal = Neuroscience | volume = 165 | issue = 2 | pages = 569–583 | date = January 2010 | pmid = 19833175 | pmc = 2795035 | doi = 10.1016/j.neuroscience.2009.10.011 }}</ref> Drugs that block the action of TNF-α or IL-1β have been shown to increase the analgesic effects of opioids and reduce the development of tolerance and other side-effects,<ref name="pmid21081778">{{cite journal | vauthors = Shen CH, Tsai RY, Shih MS, Lin SL, Tai YH, Chien CC, Wong CS | title = Etanercept restores the antinociceptive effect of morphine and suppresses spinal neuroinflammation in morphine-tolerant rats | journal = Anesthesia and Analgesia | volume = 112 | issue = 2 | pages = 454–459 | date = February 2011 | pmid = 21081778 | doi = 10.1213/ANE.0b013e3182025b15 | s2cid = 12295407 | doi-access = free }}</ref><ref name="pmid20974246">{{cite journal | vauthors = Hook MA, Washburn SN, Moreno G, Woller SA, Puga D, Lee KH, Grau JW | title = An IL-1 receptor antagonist blocks a morphine-induced attenuation of locomotor recovery after spinal cord injury | journal = Brain, Behavior, and Immunity | volume = 25 | issue = 2 | pages = 349–359 | date = February 2011 | pmid = 20974246 | pmc = 3025088 | doi = 10.1016/j.bbi.2010.10.018 }}</ref> and this has also been demonstrated with drugs that block TLR4 itself.
The response of TLR4 to opioid drugs has been found to be enantiomer-independent, so the "unnatural" enantiomers of opioid drugs such as (-)-morphine and (+)-naloxone, which lack affinity for opioid receptors, still produce the same activity at TLR4 as their "normal" enantiomers.<ref name="pmid19762094">{{cite journal | vauthors = Watkins LR, Hutchinson MR, Rice KC, Maier SF | title = The "toll" of opioid-induced glial activation: improving the clinical efficacy of opioids by targeting glia | journal = Trends in Pharmacological Sciences | volume = 30 | issue = 11 | pages = 581–591 | date = November 2009 | pmid = 19762094 | pmc = 2783351 | doi = 10.1016/j.tips.2009.08.002 }}</ref><ref name="pmid18662331">{{cite journal | vauthors = Hutchinson MR, Zhang Y, Brown K, Coats BD, Shridhar M, Sholar PW, Patel SJ, Crysdale NY, Harrison JA, Maier SF, Rice KC, Watkins LR | title = Non-stereoselective reversal of neuropathic pain by naloxone and naltrexone: involvement of toll-like receptor 4 (TLR4) | journal = The European Journal of Neuroscience | volume = 28 | issue = 1 | pages = 20–29 | date = July 2008 | pmid = 18662331 | pmc = 2588470 | doi = 10.1111/j.1460-9568.2008.06321.x }}</ref> This means that the unnatural enantiomers of opioid antagonists, such as (+)-naloxone, can be used to block the TLR4 activity of opioid analgesic drugs, while leaving the μ-opioid receptor mediated analgesic activity unaffected.<ref name="pmid18599265">{{cite journal | vauthors = Hutchinson MR, Coats BD, Lewis SS, Zhang Y, Sprunger DB, Rezvani N, Baker EM, Jekich BM, Wieseler JL, Somogyi AA, Martin D, Poole S, Judd CM, Maier SF, Watkins LR | title = Proinflammatory cytokines oppose opioid-induced acute and chronic analgesia | journal = Brain, Behavior, and Immunity | volume = 22 | issue = 8 | pages = 1178–1189 | date = November 2008 | pmid = 18599265 | pmc = 2783238 | doi = 10.1016/j.bbi.2008.05.004 }}</ref><ref name="pmid18662331" /><ref name="pmid20178837">{{cite journal | vauthors = Hutchinson MR, Lewis SS, Coats BD, Rezvani N, Zhang Y, Wieseler JL, Somogyi AA, Yin H, Maier SF, Rice KC, Watkins LR | title = Possible involvement of toll-like receptor 4/myeloid differentiation factor-2 activity of opioid inactive isomers causes spinal proinflammation and related behavioral consequences | journal = Neuroscience | volume = 167 | issue = 3 | pages = 880–893 | date = May 2010 | pmid = 20178837 | pmc = 2854318 | doi = 10.1016/j.neuroscience.2010.02.011 }}</ref> This may also be the mechanism behind the beneficial effect of ultra-low dose naltrexone on opioid analgesia.<ref name="pmid19799935">{{cite journal | vauthors = Lin SL, Tsai RY, Tai YH, Cherng CH, Wu CT, Yeh CC, Wong CS | title = Ultra-low dose naloxone upregulates interleukin-10 expression and suppresses neuroinflammation in morphine-tolerant rat spinal cords | journal = Behavioural Brain Research | volume = 207 | issue = 1 | pages = 30–36 | date = February 2010 | pmid = 19799935 | doi = 10.1016/j.bbr.2009.09.034 | s2cid = 5128970 }}</ref>
Morphine causes inflammation by binding to the protein lymphocyte antigen 96, which, in turn, causes the protein to bind to Toll-like receptor 4 (TLR4).<ref>{{cite journal | title = Neuroscience: Making morphine work better | journal = Nature | volume = 484 | issue = 7395 | page = 419 | date = 26 April 2012 | doi=10.1038/484419a | bibcode = 2012Natur.484Q.419. | s2cid = 52805136 | doi-access = free }}</ref> The morphine-induced TLR4 activation attenuates pain suppression by opioids and enhances the development of opioid tolerance and addiction, drug abuse, and other negative side effects such as respiratory depression and hyperalgesia. Drug candidates that target TLR4 may improve opioid-based pain management therapies.<ref name="urlcen.acs.org">{{cite web | url = http://cen.acs.org/articles/90/web/2012/08/Small-Molecules-Target-Toll-Like.html | title = Small Molecules Target Toll-Like Receptors | work = Chemical & Engineering News | author = Drahl C | date = 22 August 2012 }}</ref>
=== Agonists === Apart from LPS and its derivatives, up to 30 natural TLR4 agonists with diverse chemical structures have been postulated. Most of them are PAMPs or DAMPs.<ref name="pmid25559892"/> However, besides LPS (and lipid A), paclitaxel, and Ni<sup>2+</sup>, the others have not demonstrated to be direct activators of TLR4 and could instead act as chaperones for TLR4 or as promoters of LPS internalization.<ref>{{cite journal | vauthors = Kim HM, Kim YM | title = HMGB1: LPS Delivery Vehicle for Caspase-11-Mediated Pyroptosis | journal = Immunity | volume = 49 | issue = 4 | pages = 582–584 | date = October 2018 | pmid = 30332623 | doi = 10.1016/j.immuni.2018.09.021 | doi-access = free }}</ref> Further experiments should try to test whether the proposed agonists satisfy three postulates: that the activation depends on the existence of TLR4 and MD-2, that the test environment is not contaminated by other agonists such as LPS, and that the substance in question forms a novel molecular interaction with TLR4/MD-2 as the "ultimate proof".<ref name=Manchek15>{{cite journal | vauthors = Manček-Keber M, Jerala R | title = Postulates for validating TLR4 agonists | journal = European Journal of Immunology | volume = 45 | issue = 2 | pages = 356–370 | date = February 2015 | pmid = 25476977 | doi = 10.1002/eji.201444462 | s2cid = 32029412 }}</ref> TLR4/MD-2 from different species may also interact differently with proposed ligands. For example, paclitaxel is a confirmed direct TLR4/MD-2 agonist in mice, but acts as an antagonist in humans. Ni<sup>2+</sup> causes confirmed activation in humans (an explanation of nickle-associated contact dermatitis) but not in mice.<ref name=Manchek15/>
It is unclear whether opioids act as true agonists. As of 2021, molecular docking computations suggest a possible interaction at the LPS binding site of MD-2, but the way opioids modify LPS-induced interaction suggest instead a noncompetitve action at a different site.<ref>{{cite journal | vauthors = Gabr MM, Saeed I, Miles JA, Ross BP, Shaw PN, Hollmann MW, Parat MO | title = Interaction of Opioids with TLR4-Mechanisms and Ramifications | journal = Cancers | volume = 13 | issue = 21 | date = October 2021 | pmid = 34771442 | pmc = 8582379 | doi = 10.3390/cancers13215274 | doi-access = free }}</ref>
=== Antagonists === As of 2020, there were no specific TLR4 antagonists approved as drugs.<ref>{{cite journal | vauthors = Romerio A, Peri F | title = Increasing the Chemical Variety of Small-Molecule-Based TLR4 Modulators: An Overview | journal = Frontiers in Immunology | volume = 11 | article-number = 1210 | date = 2020 | pmid = 32765484 | pmc = 7381287 | doi = 10.3389/fimmu.2020.01210 | doi-access = free }}</ref> {{div col|colwidth=20em}} * Amitriptyline<ref name="pmid20381591">{{cite journal | vauthors = Hutchinson MR, Loram LC, Zhang Y, Shridhar M, Rezvani N, Berkelhammer D, Phipps S, Foster PS, Landgraf K, Falke JJ, Rice KC, Maier SF, Yin H, Watkins LR | title = Evidence that tricyclic small molecules may possess toll-like receptor and myeloid differentiation protein 2 activity | journal = Neuroscience | volume = 168 | issue = 2 | pages = 551–563 | date = June 2010 | pmid = 20381591 | pmc = 2872682 | doi = 10.1016/j.neuroscience.2010.03.067 }}</ref> * Cyclobenzaprine<ref name="pmid20381591"/> * CyRL-QN15<ref>{{cite journal | vauthors = Ru ZQ, Wu YT, Yang CY, Yang YT, Li YJ, Liu M, Peng Y, Yang YL, Wang JY, Jia QY, Li YS, Fu Z, Yang MF, Tang J, Fan Y, Liu CX, Su WR, Liu NX, He L, Wang Y, Yang XW | title = Ultra-short cyclic peptide Cy <sub>RL-QN15</sub> acts as a TLR4 antagonist to expedite oral ulcer healing | journal = Zoological Research | volume = 46 | issue = 5 | pages = 1187–1202 | date = September 2025 | pmid = 41017403 | pmc = 12780494 | doi = 10.24272/j.issn.2095-8137.2025.211 }}</ref> * Eritoran<ref>{{cite journal | vauthors = Chen F, Zou L, Williams B, Chao W | title = Targeting Toll-Like Receptors in Sepsis: From Bench to Clinical Trials | journal = Antioxidants & Redox Signaling | volume = 35 | issue = 15 | pages = 1324–1339 | date = November 2021 | pmid = 33588628 | pmc = 8817700 | doi = 10.1089/ars.2021.0005 }}</ref> * Ketotifen<ref name="pmid20381591" /> * Imipramine<ref name="pmid20381591"/> * Mianserin<ref name="pmid20381591"/> * Ibudilast<ref name="pmid22776604">{{cite journal | vauthors = Jia ZJ, Wu FX, Huang QH, Liu JM | title = [Toll-like receptor 4: the potential therapeutic target for neuropathic pain] | journal = Zhongguo Yi Xue Ke Xue Yuan Xue Bao. Acta Academiae Medicinae Sinicae | volume = 34 | issue = 2 | pages = 168–173 | date = April 2012 | pmid = 22776604 | doi = 10.3881/j.issn.1000-503X.2012.02.013 }}</ref> * Pinocembrin<ref name="pmid28007523">{{cite journal | vauthors = Lan X, Han X, Li Q, Li Q, Gao Y, Cheng T, Wan J, Zhu W, Wang J | title = Pinocembrin protects hemorrhagic brain primarily by inhibiting toll-like receptor 4 and reducing M1 phenotype microglia | journal = Brain, Behavior, and Immunity | volume = 61 | pages = 326–339 | date = March 2017 | pmid = 28007523 | pmc = 5453178 | doi = 10.1016/j.bbi.2016.12.012 }}</ref> * Resatorvid<ref name="pmid31697454">{{cite journal | vauthors = Kaieda A, Takahashi M, Fukuda H, Okamoto R, Morimoto S, Gotoh M, Miyazaki T, Hori Y, Unno S, Kawamoto T, Tanaka T, Itono S, Takagi T, Sugimoto H, Okada K, Lane W, Sang BC, Saikatendu K, Matsunaga S, Miwatashi S | title = Structure-Based Design, Synthesis, and Biological Evaluation of Imidazo[4,5-b]Pyridin-2-one-Based p38 MAP Kinase Inhibitors: Part 2 | journal = ChemMedChem | volume = 14 | issue = 24 | pages = 2093–2101 | date = December 2019 | pmid = 31697454 | doi = 10.1002/cmdc.201900373 | s2cid = 207951964 }}</ref> * M62812 * Naloxone<ref name="pmid19679181">{{cite journal | vauthors = Hutchinson MR, Zhang Y, Shridhar M, Evans JH, Buchanan MM, Zhao TX, Slivka PF, Coats BD, Rezvani N, Wieseler J, Hughes TS, Landgraf KE, Chan S, Fong S, Phipps S, Falke JJ, Leinwand LA, Maier SF, Yin H, Rice KC, Watkins LR | title = Evidence that opioids may have toll-like receptor 4 and MD-2 effects | journal = Brain, Behavior, and Immunity | volume = 24 | issue = 1 | pages = 83–95 | date = January 2010 | pmid = 19679181 | pmc = 2788078 | doi = 10.1016/j.bbi.2009.08.004 }}</ref> * (+)-Naloxone ("unnatural" isomer, lacks opioid receptor affinity so selective for TLR4 inhibition)<ref name="pmid18662331" /> * Naltrexone<ref name="pmid19679181"/> * (+)-Naltrexone<ref name="pmid19679181"/> * LPS-RS<ref name="pmid19679181"/> * Propentofylline{{citation needed|date=December 2012}} * Pentoxifylline<ref>{{cite journal | vauthors = Speer EM, Dowling DJ, Ozog LS, Xu J, Yang J, Kennady G, Levy O | title = Pentoxifylline inhibits TLR- and inflammasome-mediated in vitro inflammatory cytokine production in human blood with greater efficacy and potency in newborns | journal = Pediatric Research | volume = 81 | issue = 5 | pages = 806–816 | date = May 2017 | pmid = 28072760 | doi = 10.1038/pr.2017.6 | s2cid = 47210724 | doi-access = free }}</ref> (and downregulate TLR4 expression<ref>{{cite journal | vauthors = Schüller SS, Wisgrill L, Herndl E, Spittler A, Förster-Waldl E, Sadeghi K, Kramer BW, Berger A | title = Pentoxifylline modulates LPS-induced hyperinflammation in monocytes of preterm infants in vitro | journal = Pediatric Research | volume = 82 | issue = 2 | pages = 215–225 | date = August 2017 | pmid = 28288151 | doi = 10.1038/pr.2017.41 | s2cid = 24897100 | doi-access = free }}</ref>) * Tapentadol (mixed agonist/antagonist) * TLR4-IN-C34<ref name="pmid23776545">{{cite journal | vauthors = Neal MD, Jia H, Eyer B, Good M, Guerriero CJ, Sodhi CP, Afrazi A, Prindle T, Ma C, Branca M, Ozolek J, Brodsky JL, Wipf P, Hackam DJ | title = Discovery and validation of a new class of small molecule Toll-like receptor 4 (TLR4) inhibitors | journal = PLOS ONE | volume = 8 | issue = 6 | article-number = e65779 | date = 2013 | pmid = 23776545 | pmc = 3680486 | doi = 10.1371/journal.pone.0065779 | doi-access = free | bibcode = 2013PLoSO...865779N }}</ref> * Palmitoylethanolamide<ref>{{cite journal | vauthors = Impellizzeri D, Campolo M, Di Paola R, Bruschetta G, de Stefano D, Esposito E, Cuzzocrea S |doi = 10.1177/1721727X15575869|title = Ultramicronized palmitoylethanolamide reduces inflammation an[sic] a Th1-mediated model of colitis|journal = European Journal of Inflammation|volume = 13|pages = 14–31|year = 2015 |s2cid = 79398556|doi-access = free}}</ref> {{Div col end}}
== References == {{Reflist|33em}}
== External links == * {{MeshName|Toll-Like+Receptor+4}} * {{PDBe-KB2|O00206|Toll-like receptor 4}}
{{Clusters of differentiation}} {{TLR signaling pathway}}
Category:Clusters of differentiation 4 Category:LRR proteins