Our long term goal is to elucidate the molecular mechanism of HMGB1 A box (A box), a DNA binding domain of the molecule, as an antagonist of HMGB1. Severe sepsis is the 3rd leading cause of death nationwide, and does not currently have sufficient medical treatment, finding relevant therapeutic targets and understanding the mechanisms underlying this pathology are highly significant. HMGB1 is a necessary and sufficient mediator of severe sepsis and has been proven to be involved in both sepsis progression, organ damage and in sterile injury. In structure-function analyses, we observed that A box (amino acids 1-85 on HMGB1 protein), has an antagonistic effect to HMGB1. A box specifically inhibits HMGB1-induced cytokine release in macrophage cultures. Administration of A box rescues mice from lethal endotoxemia or sepsis caused by cecal ligation and puncture, a clinically relevant sepsis model. Following our first publication in 2004, several reports confirmed that A box acts as an antagonist of HMGB1 and extended our observations by showing beneficial effects of A box in several models of infection and sterile injury where excessive amount of HMGB1 release was observed. But the mechanisms underlying A box-mediated antagonistic effects of HMGB1 remain unknown.
In aim 1 of this proposal, we will synthesize segments of A box (over-lapping 20-mer peptides) and make point mutation mutants (cysteines at positions of 23 and 45 replaced by serine) of A box and assess the inhibition of these segments/mutants on HMGB1-mediated cytokine responses in macrophages;to start to identify the minimum sequence and/or critical region of A box required as an antagonist of HMGB1 in aim 2, we will investigate mechanism of A box action by examining the interaction of A box, and derivatives identified from above, with HMGB1 and with HMGB1 receptor TLR4/MD2;
in aim 3, we will examine the therapeutic efficacy of these effective A box derivatives in clinical relevant animal models of endotoxemia and sepsis induced by cecal perforation. We hypothesize that A box segment(s) binds to HMGB1 and inhibits HMGB1 further binding to MD2 in the TLR4/MD2 complex, consequently inhibits HMGB1-induced toxicity in sepsis. We further hypothesize that cysteines on positions 23 and 45 of A box are critical for the antagonistic activity of A box to HMGB1. Results of this research will contribute both to the scientific effort of elucidating this important mechanism for A box action, as well as to yield new directions in developing therapeutics for sepsis, by using HMGB1 antagonists as drug targets.

Public Health Relevance

Sepsis is a leading cause of death in intensive care units and HMGB1 has been shown as a critical mediator in sepsis. A box, a DNA binding domain of the molecule, has been shown as a specific antagonist of HMGB1. The overall aim is to elucidate the molecular mechanism of A box action and to determine the critical region and/or minimum sequence of A box as an antagonist of HMGB1.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM098446-01
Application #
8159694
Study Section
Special Emphasis Panel (ZRG1-SBIB-Q (02))
Program Officer
Dunsmore, Sarah
Project Start
2011-09-15
Project End
2015-07-31
Budget Start
2011-09-15
Budget End
2012-07-31
Support Year
1
Fiscal Year
2011
Total Cost
$282,200
Indirect Cost
Name
Feinstein Institute for Medical Research
Department
Type
DUNS #
110565913
City
Manhasset
State
NY
Country
United States
Zip Code
11030
Yang, H; Wang, H; Wang, Y et al. (2017) The haptoglobin beta subunit sequesters HMGB1 toxicity in sterile and infectious inflammation. J Intern Med 282:76-93
Olofsson, Peder S; Steinberg, Benjamin E; Sobbi, Roozbeh et al. (2016) Blood pressure regulation by CD4(+) lymphocytes expressing choline acetyltransferase. Nat Biotechnol 34:1066-1071
Yang, Huan; Wang, Haichao; Levine, Yaakov A et al. (2016) Identification of CD163 as an antiinflammatory receptor for HMGB1-haptoglobin complexes. JCI Insight 1:
Yang, Huan; Wang, Haichao; Chavan, Sangeeta S et al. (2015) High Mobility Group Box Protein 1 (HMGB1): The Prototypical Endogenous Danger Molecule. Mol Med 21 Suppl 1:S6-S12
Pribis, John P; Al-Abed, Yousef; Yang, Huan et al. (2015) The HIV Protease Inhibitor Saquinavir Inhibits HMGB1-Driven Inflammation by Targeting the Interaction of Cathepsin V with TLR4/MyD88. Mol Med 21:749-757
Yang, Huan; Wang, Haichao; Ju, Zhongliang et al. (2015) MD-2 is required for disulfide HMGB1-dependent TLR4 signaling. J Exp Med 212:5-14
Lu, Ben; Antoine, Daniel J; Kwan, Kevin et al. (2014) JAK/STAT1 signaling promotes HMGB1 hyperacetylation and nuclear translocation. Proc Natl Acad Sci U S A 111:3068-73
Parker, Katherine H; Sinha, Pratima; Horn, Lucas A et al. (2014) HMGB1 enhances immune suppression by facilitating the differentiation and suppressive activity of myeloid-derived suppressor cells. Cancer Res 74:5723-33
Ju, Zhongliang; Chavan, Sangeeta S; Antoine, Daniel J et al. (2014) Sequestering HMGB1 via DNA-conjugated beads ameliorates murine colitis. PLoS One 9:e103992
Yang, Huan; Antoine, Daniel J; Andersson, Ulf et al. (2013) The many faces of HMGB1: molecular structure-functional activity in inflammation, apoptosis, and chemotaxis. J Leukoc Biol 93:865-73

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