Toll-Like Receptor 4 Activation in Cancer Progression and Therapy

Cancer immunotherapy has been the focus of intense research since the late 19th century when Coley observed that bacterial components can contribute to cancer regression by eliciting an antitumor immune response. Successful activation and maturation of tumor-specific immune cells is now known to be mediated by bacterial endotoxin, which activates Toll-like receptor 4 (TLR4). TLR4 is expressed on a variety of immune as well as tumor cells, but its activation can have opposing effects. While TLR4 activation can promote antitumor immunity, it can also result in increased tumor growth and immunosuppression. Nevertheless, TLR4 engagement by endotoxin as well as by endogenous ligands represents notable contribution to the outcome of different cancer treatments, such as radiation or chemotherapy. Further research of the role and mechanisms of TLR4 activation in cancer may provide novel antitumor vaccine adjuvants as well as TLR4 inhibitors that could prevent inflammation-induced carcinogenesis

Zwiespältige Bilanz für Afrika : die Milleniumsziele der UN haben keine Gleichberechtigung in der Entwicklungszusammenarbeit gebracht

An increasing number of pathologies have been linked to Toll-like receptor 4 (TLR4) activation and signaling, therefore new hit and lead compounds targeting this receptor activation process are urgently needed. We report on the synthesis and biological properties of glycolipids based on glucose and trehalose scaffolds which potently inhibit TLR4 activation and signaling in vitro and in vivo. Structure–activity relationship studies on these compounds indicate that the presence of fatty ester chains in the molecule is a primary prerequisite for biological activity and point to facial amphiphilicity as a preferred architecture for TLR4 antagonism. The cationic glycolipids here presented can be considered as new lead compounds for the development of drugs targeting TLR4 activation and signaling in infectious, inflammatory, and autoimmune diseases. Interestingly, the biological activity of the best drug candidate was retained after adsorption at the surface of colloidal gold nanoparticles, broadening the options for clinical development.Italian, Ministry of University and Research (MIUR), PRIN 2010 - 11Ministerio de Economía y Competitividad AF2013-44021- R, CTQ2010 - 15848Junta de Andalucía FQM 1467Slovenian Research Agency P4-176, J1 - 417

Species-Specific Activation of TLR4 by Hypoacylated Endotoxins Governed by Residues 82 and 122 of MD-2

<div><p>The Toll-like receptor 4/MD-2 receptor complex recognizes endotoxin, a Gram-negative bacterial cell envelope component. Recognition of the most potent hexaacylated form of endotoxin is mediated by the sixth acyl chain that protrudes from the MD-2 hydrophobic pocket and bridges TLR4/MD-2 to the neighboring TLR4 ectodomain, driving receptor dimerization via hydrophobic interactions. In hypoacylated endotoxins all acyl chains could be accommodated within the binding pocket of the human hMD-2. Nevertheless, tetra- and pentaacylated endotoxins activate the TLR4/MD-2 receptor of several species. We observed that amino acid residues 82 and 122, located at the entrance to the endotoxin binding site of MD-2, have major influence on the species-specific endotoxin recognition. We show that substitution of hMD-2 residue V82 with an amino acid residue with a bulkier hydrophobic side chain enables activation of TLR4/MD-2 by pentaacylated and tetraacylated endotoxins. Interaction of the lipid A phosphate group with the amino acid residue 122 of MD-2 facilitates the appropriate positioning of the hypoacylated endotoxin. Moreover, mouse TLR4 contributes to the agonistic effect of pentaacylated msbB endotoxin. We propose a molecular model that explains how the molecular differences between the murine or equine MD-2, which both have sufficiently large hydrophobic pockets to accommodate all five or four acyl chains, influence the positioning of endotoxin so that one of the acyl chains remains outside the pocket and enables hydrophobic interactions with TLR4, leading to receptor activation.</p></div

MD-2 determinants of nickel and cobalt-mediated activation of human TLR4.

Recent findings unexpectedly revealed that human TLR4 can be directly activated by nickel ions. This activation is due to the coordination of nickel by a cluster of histidine residues on the ectodomain of human TLR4, which is absent in most other species. We aimed to elucidate the role of MD-2 in the molecular mechanism of TLR4/MD-2 activation by nickel, as nickel binding site on TLR4 is remote from MD-2, which directly binds the endotoxin as the main pathological activator of TLR4. We identified MD-2 and TLR4 mutants which abolished TLR4/MD-2 receptor activation by endotoxin but could nevertheless be significantly activated by nickel, which acts in synergy with LPS. Human TLR4/MD-2 was also activated by cobalt ions, while copper and cadmium were toxic in the tested concentration range. Activation of TLR4 by cobalt required MD-2 and was abolished by human TLR4 mutations of histidine residues at positions 456 and 458. We demonstrated that activation of TLR4 by nickel and cobalt ions can trigger both the MyD88-dependent and the -independent pathway. Based on our results we propose that predominantly hydrophobic interactions between MD-2 and TLR4 contribute to the stabilization of the TLR4/MD-2/metal ion complex in a conformation that enables activation

Responsiveness to different types of endotoxins by combinations of MD-2 and TLR4.

<p>Responsiveness to different types of endotoxins by combinations of MD-2 and TLR4.</p

Mutations of murine MD-2 residues E122 and L125 diminish responsiveness to tetraacylated endotoxin while preserving activation by hexaacylated endotoxin.

<p>(A) Murine MD-2 mutants mE122K and mE122K L125K gain the ability to activate the human TLR4 with hexaacylated endotoxin. (B) Murine MD-2 mutants mE122K and mE122K L125K in combination with murine TLR4 exhibit reduced activation by tetraacylated lipid IVa.</p

The structure of the human MD-2 with bound endotoxin.

<p>(A) Human MD-2 (pdb id 3FXI, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0107520#pone.0107520-Park1" target="_blank">[27]</a>) is shown in grey ribbon. The side chains of amino acid residues at positions 82 (valine), 122 (lysine) and 125 (lysine) are shown in stick representation. These amino acid residues are positioned at the entrance to the binding pocket and come in close proximity to the ligand (here the hexaacylated endotoxin, shown in yellow). Figure was prepared with the USCF Chimera package <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0107520#pone.0107520-Pettersen1" target="_blank">[26]</a>. (B) Amino acid alignment of residues at positions 82, 122 and 125 of human, murine and equine MD-2 (red – acidic amino acid; blue – basic amino acid; green – nonpolar/hydrophobic amino acid). (C) The structure of lipid A. The arrows indicate acyl chains that are absent in the structure of the msbB endotoxin and in lipid IVa.</p