IinQ attenuates systemic inflammatory responses via selectively impairing the Myddosome complex formation upon TLR4 ligation.
A specific small-molecule inhibitor of the TLR4 signaling complex upstream of the IKK would likely provide therapeutic benefit for NF-κB-mediated inflammatory disease. We previously identified brazilin as a selective upstream IKK inhibitor targeting the Myddosome complex. In this study, using a cell-based ubiquitination assay for IRAK1 and a chemical library comprising a series of structural analogues of brazilin, a novel small molecule, 2-hydroxy-5,6-dihydroisoindolo[1,2-a]isoquinoline-3,8-dione (IinQ), was identified as a selective and potent inhibitor of IRAK1-dependent NF-κB activation upon TLR4 ligation. In RAW264.7 macrophages, IinQ drastically suppressed activation of upstream IKK signaling events including membrane-bound IRAK1 ubiquitination and IKK phosphorylation by the TLR4 ligand, resulting in reduced expression of proinflammatory mediators including IL-6, TNF-α, and nitric oxide. Interestingly, IinQ did not suppress NF-κB activation via the TLR3 ligand, DNA damaging agents, or a protein kinase C activator, indicating IinQ is specific for TLR4 signaling. Analysis of upstream signaling events further confirmed that IinQ disrupts the MyD88-IRAK1-TRAF6 complex formation induced by LPS treatment, without affecting TLR4 oligomerization. Moreover, intravenous administration of IinQ significantly reduced lethality and attenuated systemic inflammatory responses in an in vivo mouse model of endotoxin shock following LPS challenge. Thus, IinQ represents a novel class of brazilin analogues with improved potency and specificity toward disruption of Myddosome complex formation in TLR4 signaling, indicating that IinQ may be a promising therapeutic candidate for the treatment of systemic inflammatory diseases.
Department of Pharmacology, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea; Department of Medical Science, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea.
Kidong
K
Kang
Department of Pharmacology, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea.
Minho
M
Won
Department of Medical Science, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea; Department of Infection Biology, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea.
Jae-Min
JM
Yuk
Eco-Friendly New Materials Research Center, Korea Research Institute of Chemical Technology, 141 Gajeong-ro, Yusung-gu, Daejeon 34114, Republic of Korea.
Chan-Yong
CY
Park
Department of Pharmacology, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea.
Hee Sun
HS
Byun
Department of Pharmacology, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea.
Kyeong Ah
KA
Park
Department of Pharmacology, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea; Department of Medical Science, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea.
So-Ra
SR
Lee
Eco-Friendly New Materials Research Center, Korea Research Institute of Chemical Technology, 141 Gajeong-ro, Yusung-gu, Daejeon 34114, Republic of Korea.
Young-Goo
YG
Kang
Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
Han-Ming
HM
Shen
Eco-Friendly New Materials Research Center, Korea Research Institute of Chemical Technology, 141 Gajeong-ro, Yusung-gu, Daejeon 34114, Republic of Korea. Electronic address: iylee@krict.re.kr.
Ill Young
IY
Lee
Department of Pharmacology, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea; Department of Medical Science, College of Medicine, Chungnam National University, 266 Munhwa-ro, Daejeon 35015, Republic of Korea. Electronic address: gmhur@cnu.ac.kr.
Gang Min
GM
Hur
D000818:Animals; D002460:Cell Line; D005260:Female; D051550:I-kappa B Kinase / Q000037:antagonists & inhibitors*; D053592:Interleukin-1 Receptor-Associated Kinases / Q000378:metabolism*; D054833:Isoindoles / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D007546:Isoquinolines / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000633:toxicity; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D053594:Myeloid Differentiation Factor 88 / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D018746:Systemic Inflammatory Response Syndrome / Q000188:drug therapy* / Q000378:metabolism; D048029:TNF Receptor-Associated Factor 6 / Q000378:metabolism*; D051197:Toll-Like Receptor 4 / Q000378:metabolism*; D054875:Ubiquitination / Q000187:drug effects
C000619252:2-hydroxy-5,6-dihydroisoindolo(1,2-a)isoquinoline-3,8-diol; D054833:Isoindoles; D007546:Isoquinolines; D008070:Lipopolysaccharides; C507780:Myd88 protein, mouse; D053594:Myeloid Differentiation Factor 88; D048029:TNF Receptor-Associated Factor 6; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; C433788:lipopolysaccharide, E. coli O26-B6; D053592:Interleukin-1 Receptor-Associated Kinases; C507769:Irak1 protein, mouse; D051550:I-kappa B Kinase
IRAK1 ubiquitination; IinQ; Inflammatory response; Nuclear factor-κB; TLR4