title: A Novel Class of Small Molecule Agonists with Preference for Human over Mouse TLR4 Activation.
abstract: The best-characterized Toll-like receptor 4 (TLR4) ligands are lipopolysaccharide (LPS) and its chemically modified and detoxified variant, monophosphoryl lipid A (MPL). Although both molecules are active for human TLR4, they demonstrate a potency preference for mouse TLR4 based on data from transfected cell lines and primary cells of both species. After a high throughput screening process of small molecule libraries, we have discovered a new class of TLR4 agonist with a species preference profile differing from MPL. Products of the 4-component Ugi synthesis reaction were demonstrated to potently trigger human TLR4-transfected HEK cells but not mouse TLR4, although inclusion of the human MD2 with mTLR4 was able to partially recover activity. Co-expression of CD14 was not required for optimal activity of Ugi compounds on transfected cells, as it is for LPS. The species preference profile for the panel of Ugi compounds was found to be strongly active for human and cynomolgus monkey primary cells, with reduced but still substantial activity for most Ugi compounds on guinea pig cells. Mouse, rat, rabbit, ferret, and cotton rat cells displayed little or no activity when exposed to Ugi compounds. However, engineering the human versions of TLR4 and MD2 to be expressed in mTLR4/MD2 deficient mice allowed for robust activity by Ugi compounds both in vitro and in vivo. These findings extend the range of compounds available for development as agonists of TLR4 and identify novel molecules which reverse the TLR4 triggering preference of MPL for mouse TLR4 over human TLR4. Such compounds may be amenable to formulation as more potent human-specific TLR4L-based adjuvants than typical MPL-based adjuvants.
authors.affiliation: Vaccine Platform Group, MedImmune, Gaithersburg, Maryland, United States of America.
authors.forename: Jason D
authors.identifier: http://orcid.org/0000-0002-7864-803X
authors.initials: JD
authors.lastname: Marshall
authors.affiliation: Translational Biology Group, MedImmune, Mountain View, California, United States of America.
authors.forename: Darren S
authors.initials: DS
authors.lastname: Heeke
authors.affiliation: Translational Biology Group, MedImmune, Mountain View, California, United States of America.
authors.forename: Eileen
authors.initials: E
authors.lastname: Rao
authors.affiliation: Vaccine Platform Group, MedImmune, Gaithersburg, Maryland, United States of America.
authors.forename: Sean K
authors.initials: SK
authors.lastname: Maynard
authors.affiliation: Department of Cardiovascular and Metabolic Diseases, MedImmune, Cambridge, United Kingdom.
authors.forename: David
authors.initials: D
authors.lastname: Hornigold
authors.affiliation: Translational Science, Respiratory, Inflammation and Autoimmunity Innovative Medicines, AstraZeneca R&D, Mölndal, Sweden.
authors.forename: Christopher
authors.initials: C
authors.lastname: McCrae
authors.affiliation: Biology Department, AstraZeneca R&D, Charnwood, United Kingdom.
authors.forename: Neil
authors.initials: N
authors.lastname: Fraser
authors.affiliation: Research Bioinformatics, MedImmune, Gaithersburg, Maryland, United States of America.
authors.forename: Andrey
authors.initials: A
authors.lastname: Tovchigrechko
authors.affiliation: Statistical Sciences, MedImmune, Gaithersburg, Maryland, United States of America.
authors.forename: Li
authors.initials: L
authors.lastname: Yu
authors.affiliation: Biology Department, AstraZeneca R&D, Charnwood, United Kingdom.
authors.forename: Nicola
authors.initials: N
authors.lastname: Williams
authors.affiliation: Medicinal Chemistry Department, AstraZeneca R&D, Charnwood, United Kingdom.
authors.forename: Sarah
authors.initials: S
authors.lastname: King
authors.affiliation: Translational Science, Respiratory, Inflammation and Autoimmunity Innovative Medicines, AstraZeneca R&D, Mölndal, Sweden.
authors.forename: Martin E
authors.initials: ME
authors.lastname: Cooper
authors.affiliation: Department of Comparative Medicine, University of Washington, Seattle, Washington, United States of America.
authors.forename: Adeline M
authors.initials: AM
authors.lastname: Hajjar
authors.affiliation: Translational Biology Group, MedImmune, Mountain View, California, United States of America.
authors.forename: Jennifer C
authors.initials: JC
authors.lastname: Woo
mesh_terms: D000577:Amides / Q000138:chemical synthesis / Q000737:chemistry; D000818:Animals; D001665:Binding Sites; D001854:Bone Marrow Cells / Q000166:cytology; D002478:Cells, Cultured; D018925:Chemokines / Q000378:metabolism; D016207:Cytokines / Q000378:metabolism; D003713:Dendritic Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D017930:Genes, Reporter / Q000235:genetics; D006168:Guinea Pigs; D057809:HEK293 Cells; D006801:Humans; D018950:Lipopolysaccharide Receptors / Q000235:genetics; D008070:Lipopolysaccharides / Q000633:toxicity; D051218:Lymphocyte Antigen 96 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008822:Mice, Transgenic; D062105:Molecular Docking Simulation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011817:Rabbits; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects; D054852:Small Molecule Libraries / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D013154:Spleen / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000819:agonists / Q000235:genetics / Q000378:metabolism*
chemical_list: D000577:Amides; D018925:Chemokines; D016207:Cytokines; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D051218:Lymphocyte Antigen 96; D054852:Small Molecule Libraries; D051197:Toll-Like Receptor 4