Upon neutrophil activation, peptidylarginine deiminase 4 (PAD4) may translocate to the nucleus to citrullinate histones. This decondenses chromatin which is released as neutrophil extracellular traps (NETs). NETs trap microbes but also promote inflammation. Relevant to this application, PAD4/NETs play an important role in pathological thrombosis and their formation can be stimulated by cancer. The central hypotheses of this proposal are: NETs are involved in the formation of a stable organized and vascularized thrombus and breaking up NETs is necessary for thrombolysis. Thrombosis promotes NET deposition in the adjacent vessel wall and in distant organs leading to post-thrombotic syndrome (PTS) and an increased systemic pro-coagulant and pro-inflammatory state. Finally, PAD4/NETs elevate coagulation in cancer and promote tumor angiogenesis and growth. To address the hypotheses, we will study several models of deep vein thrombosis (DVT) and breast cancer in wild-type (WT) and genetically engineered mice. The special knockouts to be used include PAD4-/- and newly generated DNase 1-/- mice. Novel technologies will be applied to extract 3D models of the neovasculature to generate vasometrics from the thrombi. Also, shear-activated nanotherapeutics (SA-NTs) will be tested to induce thrombolysis. The proposal has two specific aims:
Aim1 : to study NETs in venous thrombosis and thrombolysis. In this aim, we propose to investigate the role of NETs in various stages of DVT formation and in resulting fibrosis of the vessel wall. We will evaluate the role of extracellular chromatin and platelets in thrombus neo- vascularization and how chromatin and neovascularization affect thrombolysis.
Aim 2 : to study NETs/PAD4 in thrombosis-induced systemic inflammation and cancer-induced thrombosis. In this aim, we will examine what systemic effects and susceptibilities a mouse with DVT experiences and to what extent NETs contribute to pro-coagulant and pro-angiogenic activities of cancer. Thrombosis, including DVT/pulmonary embolism, is now the biggest killer in United States. We hope that our studies will broaden understanding of the roles NETs play in thrombosis and help to guide new treatments and prevention of pathological thrombosis, PTS and cancer-induced thrombosis.

Public Health Relevance

Thrombotic events are now the biggest killer in United States. Thrombi are formed by platelets sticking together but white blood cells influence the process. They actively release their DNA forming a mesh together with plasma proteins cementing the thrombus together. We observed that cancer stimulates white cells to release a lot of DNA promoting thrombosis. We propose that this release of DNA could be inhibited and that thrombi could be dissolved by breaking both the protein and DNA scaffold-a new anti-thrombotic approach.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL102101-07
Application #
9043165
Study Section
Hemostasis and Thrombosis Study Section (HT)
Program Officer
Kindzelski, Andrei L
Project Start
2010-04-01
Project End
2019-05-31
Budget Start
2016-06-01
Budget End
2017-05-31
Support Year
7
Fiscal Year
2016
Total Cost
Indirect Cost
Name
Children's Hospital Boston
Department
Type
DUNS #
076593722
City
Boston
State
MA
Country
United States
Zip Code
Gavillet, Mathilde; Martinod, Kimberly; Renella, Raffaele et al. (2018) A key role for Rac and Pak signaling in neutrophil extracellular traps (NETs) formation defines a new potential therapeutic target. Am J Hematol 93:269-276
Zitomersky, Naamah L; Demers, Melanie; Martinod, Kimberly et al. (2017) ADAMTS13 Deficiency Worsens Colitis and Exogenous ADAMTS13 Administration Decreases Colitis Severity in Mice. TH Open 1:e11-e23
Hayashi, Hideki; Cherpokova, Deya; Martinod, Kimberly et al. (2017) Sirt3 deficiency does not affect venous thrombosis or NETosis despite mild elevation of intracellular ROS in platelets and neutrophils in mice. PLoS One 12:e0188341
Martinod, Kimberly; Witsch, Thilo; Erpenbeck, Luise et al. (2017) Peptidylarginine deiminase 4 promotes age-related organ fibrosis. J Exp Med 214:439-458
Erpenbeck, Luise; Chowdhury, Chanchal Sur; Zsengellér, Zsuzsanna K et al. (2016) PAD4 Deficiency Decreases Inflammation and Susceptibility to Pregnancy Loss in a Mouse Model. Biol Reprod 95:132
Shaw, Maureen A; Kombrinck, Keith W; McElhinney, Kathryn E et al. (2016) Limiting prothrombin activation to meizothrombin is compatible with survival but significantly alters hemostasis in mice. Blood 128:721-31
Demers, Mélanie; Wong, Siu Ling; Martinod, Kimberly et al. (2016) Priming of neutrophils toward NETosis promotes tumor growth. Oncoimmunology 5:e1134073
Pillai, Padmini S; Molony, Ryan D; Martinod, Kimberly et al. (2016) Mx1 reveals innate pathways to antiviral resistance and lethal influenza disease. Science 352:463-6
Martinod, K; Witsch, T; Farley, K et al. (2016) Neutrophil elastase-deficient mice form neutrophil extracellular traps in an experimental model of deep vein thrombosis. J Thromb Haemost 14:551-8
Erpenbeck, Luise; Demers, Melanie; Zsengellér, Zsuzsanna K et al. (2016) ADAMTS13 Endopeptidase Protects against Vascular Endothelial Growth Factor Inhibitor-Induced Thrombotic Microangiopathy. J Am Soc Nephrol 27:120-31

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