Publications
98 results found
South K, Salles-Crawley I, Crawley JT, et al., 2015, The importance of conformational activation of ADAMTS13 for control of platelet deposition under flow, Publisher: WILEY-BLACKWELL, Pages: 766-767, ISSN: 1538-7933
Thomas M, Groot RD, Crawley J, et al., 2015, Pathogenicity of anti-ADAMTS13 autoantibodies in acquired TTP, Publisher: WILEY-BLACKWELL, Pages: 473-473, ISSN: 1538-7933
Salles-Crawley I, Monkman JH, Lane DA, et al., 2015, Endothelial BAMBI (BMP and activin membrane bound inhibitor) is important for fibrin generation and thrombus stability, Publisher: WILEY-BLACKWELL, Pages: 110-110, ISSN: 1538-7933
Reglinska-Matveyev N, Crawley JT, Camire R, et al., 2015, FV enhances protein S cofactor function for TFPI in the inhibition of FXa, Publisher: WILEY-BLACKWELL, Pages: 54-54, ISSN: 1538-7933
Gierula M, Salles-Crawley II, Crawley JTB, et al., 2015, Factor VA in synergy with protein s enhances activated protein C binding to phospholipids, Publisher: WILEY-BLACKWELL, Pages: 313-313, ISSN: 1538-7933
Andreou AP, Efthymiou M, Yu Y, et al., 2015, Protective Effects of Non-Anticoagulant Activated Protein C Variant (D36A/L38D/A39V) in a Murine Model of Ischaemic Stroke, PLOS One, Vol: 10, ISSN: 1932-6203
Ischaemic stroke is caused by occlusive thrombi in the cerebral vasculature. Although tissue-plasminogenactivator (tPA) can be administered as thrombolytic therapy, it has majorlimitations, which include disruption of the blood-brain barrier and an increased risk ofbleeding. Treatments that prevent or limit such deleterious effects could be of major clinicalimportance. Activated protein C (APC) is a natural anticoagulant that regulates thrombingeneration, but also confers endothelial cytoprotective effects and improved endothelialbarrier function mediated through its cell signalling properties. In murine models of stroke,although APC can limit the deleterious effects of tPA due to its cell signalling function, its anticoagulantactions can further elevate the risk of bleeding. Thus, APC variants such asAPC(5A), APC(Ca-ins) and APC(36-39) with reduced anticoagulant, but normal signallingfunction may have therapeutic benefit. Human and murine protein C (5A), (Ca-ins) and (36-39) variants were expressed and characterised. All protein C variants were secreted normally,but 5-20% of the protein C (Ca-ins) variants were secreted as disulphide-linked dimers.Thrombin generation assays suggested reductions in anticoagulant function of 50- to57-fold for APC(36-39), 22- to 27-fold for APC(Ca-ins) and 14- to 17-fold for APC(5A). Interestingly,whereas human wt APC, APC(36-39) and APC(Ca-ins) were inhibited similarly byprotein C inhibitor (t½ - 33 to 39 mins), APC(5A) was inactivated ~9-fold faster (t½ - 4 mins).Using the murine middle cerebral artery occlusion ischaemia/repurfusion injury model, incombination with tPA, APC(36-39), which cannot be enhanced by its cofactor protein S, significantlyimproved neurological scores, reduced cerebral infarct area by ~50% and reducedoedema ratio. APC(36-39) also significantly reduced bleeding in the brain induced by administrationof tPA, whereas wt APC did not. If our data can be extrapolated to clinical settings,then APC(36-39
de Groot R, Lane DA, Crawley JTB, 2015, The role of the ADAMTS13 cysteine-rich domain in VWF binding and proteolysis, Blood, Vol: 125, Pages: 1968-1975, ISSN: 0006-4971
ADAMTS13 proteolytically regulates the platelet-tethering function of von Willebrand factor (VWF). ADAMTS13 function is dependent upon multiple exosites that specifically bind the unraveled VWF A2 domain and enable proteolysis. We carried out a comprehensive functional analysis of the ADAMTS13 cysteine-rich (Cys-rich) domain using engineered glycans, sequence swaps, and single point mutations in this domain. Mutagenesis of Cys-rich domain–charged residues had no major effect on ADAMTS13 function, and 5 out of 6 engineered glycans on the Cys-rich domain also had no effect on ADAMTS13 function. However, a glycan attached at position 476 appreciably reduced both VWF binding and proteolysis. Substitution of Cys-rich sequences for the corresponding regions in ADAMTS1 identified a hydrophobic pocket involving residues Gly471-Val474 as being of critical importance for both VWF binding and proteolysis. Substitution of hydrophobic VWF A2 domain residues to serine in a region (residues 1642-1659) previously postulated to interact with the Cys-rich domain revealed the functional importance of VWF residues Ile1642, Trp1644, Ile1649, Leu1650, and Ile1651. Furthermore, the functional deficit of the ADAMTS13 Cys-rich Gly471-Val474 variant was dependent on these same hydrophobic VWF residues, suggesting that these regions form complementary binding sites that directly interact to enhance the efficiency of the proteolytic reaction.
South K, Luken BM, Crawley JTB, et al., 2014, Conformational activation of ADAMTS13, Proceedings of the National Academy of Sciences of the United States of America, Vol: 111, Pages: 18578-18583, ISSN: 0027-8424
A disintegrin and metalloprotease with thrombospondin motifs 13 (ADAMTS13) is a metalloprotease that regulates von Willebrand factor (VWF) function. ADAMTS13-mediated proteolysis is determined by conformational changes in VWF, but also may depend on its own conformational activation. Kinetic analysis of WT ADAMTS13 revealed ∼2.5-fold reduced activity compared with ADAMTS13 lacking its C-terminal tail (MDTCS) or its CUB1-2 domains (WTΔCUB1-2), suggesting that the CUB domains naturally limit ADAMTS13 function. Consistent with this suggestion, WT ADAMTS13 activity was enhanced ∼2.5-fold by preincubation with either an anti-CUB mAb (20E9) or VWF D4CK (the natural binding partner for the CUB domains). Furthermore, the isolated CUB1-2 domains not only bound MDTCS, but also inhibited activity by up to 2.5-fold. Interestingly, a gain-of-function (GoF) ADAMTS13 spacer domain variant (R568K/F592Y/R660K/Y661F/Y665F) was ∼2.5-fold more active than WT ADAMTS13, but could not be further activated by 20E9 mAb or VWF D4CK and was unable to bind or to be inhibited by the CUB1-2 domains, suggesting that the inhibitory effects of the CUB domains involve an interaction with the spacer domain that is disrupted in GoF ADAMTS13. Electron microscopy demonstrated a “closed” conformation of WT ADAMTS13 and suggested a more “open” conformation for GoF ADAMTS13. The cryptic spacer domain epitope revealed by conformational unfolding also represents the core antigenic target for autoantibodies in thrombotic thrombocytopenic purpura. We propose that ADAMTS13 circulates in a closed conformation, which is maintained by a CUB–spacer domain binding interaction. ADAMTS13 becomes conformationally activated on demand through interaction of its C-terminal CUB domains with VWF, making it susceptible to immune recognition.
Thomas MR, Crawley JT, Machin SJ, et al., 2014, Domain Specificity and Mechanism of Action of Anti-ADAMTS13 Antibodies at Presentation and Relapse in Acquired Thrombotic Thrombocytopenic Purpura (TTP), 56th Annual Meeting of the American-Society-of-Hematology, Publisher: AMER SOC HEMATOLOGY, ISSN: 0006-4971
Reglinska-Matveyev N, Andersson HM, Rezende SM, et al., 2014, TFPI cofactor function of protein S: essential role of the protein S SHBG-like domain, Blood, Vol: 123, Pages: 3979-3987, ISSN: 0006-4971
Protein S is a cofactor for tissue factor pathway inhibitor (TFPI), accelerating the inhibition of activated factor X (FXa). TFPI Kunitz domain 3 residue Glu226 is essential for enhancement of TFPI by protein S. To investigate the complementary functional interaction site on protein S, we screened 44 protein S point, composite or domain swap variants spanning the whole protein S molecule for their TFPI cofactor function using a thrombin generation assay. Of these variants, two protein S/growth arrest–specific 6 chimeras, with either the whole sex hormone–binding globulin (SHBG)-like domain (Val243-Ser635; chimera III) or the SHBG laminin G-type 1 subunit (Ser283-Val459; chimera I), respectively, substituted by the corresponding domain in growth arrest–specific 6, were unable to enhance TFPI. The importance of the protein S SHBG-like domain (and its laminin G-type 1 subunit) for binding and enhancement of TFPI was confirmed in FXa inhibition assays and using surface plasmon resonance. In addition, protein S bound to C4b binding protein showed greatly reduced enhancement of TFPI-mediated inhibition of FXa compared with free protein S. We show that binding of TFPI to the protein S SHBG-like domain enables TFPI to interact optimally with FXa on a phospholipid membrane.
Thomas M, Crawley J, Machin S, et al., 2014, DOMAIN SPECIFICITY AND INHIBITORY POTENTIAL OF ANTI-ADAMTS13 ANTIBODIES AT PRESENTATION AND RELAPSE IN THROMBOTIC THROMBOCYTOPENIC PURPURA (TTP), 19th Congress of the European-Hematology-Association, Publisher: FERRATA STORTI FOUNDATION, Pages: 538-538, ISSN: 0390-6078
Salles-Crawley II, Monkman JH, Ahnstroem J, et al., 2014, Vessel wall BAMBI contributes to hemostasis and thrombus stability., Blood, Vol: 123, Pages: 2873-2881, ISSN: 0006-4971
Bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI) is a transmembrane protein related to the transforming growth factor-β superfamily, and is highly expressed in platelets and endothelial cells. We previously demonstrated its positive role in thrombus formation using a zebrafish thrombosis model. In the present study, we used Bambi-deficient mice and radiation chimeras to evaluate the function of this receptor in the regulation of both hemostasis and thrombosis. We show that Bambi−/− and Bambi+/− mice exhibit mildly prolonged bleeding times compared with Bambi+/+ littermates. In addition, using 2 in vivo thrombosis models in mesenterium or cremaster muscle arterioles, we demonstrate that Bambi-deficient mice form unstable thrombi compared with Bambi+/+ mice. No defects in thrombin generation in Bambi−/− mouse plasma could be detected ex vivo. Moreover, the absence of BAMBI had no effect on platelet counts, platelet activation, aggregation, or platelet procoagulant function. Similar to Bambi−/− mice, Bambi−/− transplanted with Bambi+/+ bone marrow formed unstable thrombi in the laser-induced thrombosis model that receded more rapidly than thrombi that formed in Bambi+/+ mice receiving Bambi−/− bone marrow transplants. Taken together, these results provide strong evidence for an important role of endothelium rather than platelet BAMBI as a positive regulator of both thrombus formation and stability.
Thomas MR, Crawley J, Machin S, et al., 2014, Domain specificity of anti-ADAMTS13 antibodies at presentation and relapse in thrombotic thrombocytopenic purpura (TTP), 54th Annual Scientific Meeting of the British-Society-for-Haematology, Publisher: WILEY-BLACKWELL, Pages: 3-4, ISSN: 0007-1048
Crawley JTB, Scully MA, 2013, Thrombotic thrombocytopenic purpura: basic pathophysiology and therapeutic strategies, HEMATOLOGY-AMERICAN SOCIETY OF HEMATOLOGY EDUCATION PROGRAM, Pages: 292-299, ISSN: 1520-4391
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- Citations: 49
Onubogu IK, Cooper N, Naresh K, et al., 2013, Alternative Methods Of Thrombocytopenia: Patients With ITP Have Increased Megakaryocyte/T Cell Interactions In The Bone Marrow and Higher Serum TRAIL Levels, 55th Annual Meeting of the American-Society-of-Hematology, Publisher: AMER SOC HEMATOLOGY, ISSN: 0006-4971
Reglinska NB, Andersson HMH, Rezende SM, et al., 2013, The search for functionally important residues in protein S required for its enhancement of TFPI, JOURNAL OF THROMBOSIS AND HAEMOSTASIS, Vol: 11, Pages: 328-328, ISSN: 1538-7933
Salles I, Monkman JH, Ahnstroem J, et al., 2013, BMP and activin membrane bound inhibitor (BAMBI): A novel regulator of thrombus formation, Publisher: WILEY-BLACKWELL, Pages: 274-275, ISSN: 1538-7933
Crawley JTB, Efthymiou M, Yu Y, et al., 2013, Protective effects of non-anticoagulant activated protein C variant (D36A/L38D/A39V) in a murine model of ischaemic stroke, JOURNAL OF THROMBOSIS AND HAEMOSTASIS, Vol: 11, Pages: 64-64, ISSN: 1538-7933
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- Citations: 1
Ahnstroem J, Andersson HM, Hockey V, et al., 2012, Identification of functionally important residues in TFPI Kunitz domain 3 required for the enhancement of its activity by protein S, BLOOD, Vol: 120, Pages: 5059-5062, ISSN: 0006-4971
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- Citations: 40
Godfrey CL, Terrinoni I, Laffan M, et al., 2012, Elevated Plasma Von Willebrand Factor and Decreased ADAMTS13 Antigen Levels in Patients with Immune Thrombocytopenia (ITP), 54th Annual Meeting and Exposition of the American-Society-of-Hematology (ASH), Publisher: AMER SOC HEMATOLOGY, ISSN: 0006-4971
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- Citations: 5
Salles II, Crawley JTB, 2012, Blocking VWF platelet binding to treat TTP, BLOOD, Vol: 120, Pages: 3390-3392, ISSN: 0006-4971
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- Citations: 2
Crawley JTB, de Groot R, 2012, Cardiovascular string theory, BLOOD, Vol: 119, Pages: 2181-2182, ISSN: 0006-4971
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- Citations: 1
Andersson HM, Siegerink B, Luken BM, et al., 2012, High VWF, low ADAMTS13, and oral contraceptives increase the risk of ischemic stroke and myocardial infarction in young women, BLOOD, Vol: 119, Pages: 1555-1560, ISSN: 0006-4971
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- Citations: 109
Andreou AP, Crawley JTB, 2011, Thrombomodulin analogues for the treatment of ischemic stroke, Journal of Thrombosis and Haemostasis, Vol: 9, Pages: 1171-1173, ISSN: 1538-7933
Crawley JTB, de Groot R, Xiang Y, et al., 2011, Unraveling the scissile bond: how ADAMTS13 recognizes and cleaves von Willebrand factor, BLOOD, Vol: 118, Pages: 3212-3221, ISSN: 0006-4971
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- Citations: 218
Xiang Y, de Groot R, Crawley JTB, et al., 2011, Mechanism of von Willebrand factor scissile bond cleavage by a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13), PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, Vol: 108, Pages: 11602-11607, ISSN: 0027-8424
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- Citations: 41
Yu Y, Crawley JTB, Mason JC, et al., 2011, Protein kinase C isoforms in the divergent endothelial signalling exerted by thrombin- and activated protein c-dependent activation of PAR-1, Publisher: WILEY-BLACKWELL, Pages: 239-239, ISSN: 1538-7933
Siegerink B, Andersson HM, Luken BM, et al., 2011, VWF and ADAMTS13 levels and the risk of myocardial infarction and ischaemic stroke in young women: results from the ratio case-control study, Publisher: WILEY-BLACKWELL, Pages: 209-209, ISSN: 1538-7933
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- Citations: 1
Ahnstrom J, Andersson HM, Canis K, et al., 2011, Activated protein S cofactor function of protein S: a novel function for a γ-carboxyglutamic acid residue, Publisher: WILEY-BLACKWELL, Pages: 14-14, ISSN: 1538-7933
Xiang Y, de Groot R, Crawley JTB, et al., 2011, Essential role of the P3 residue (Leu1603) of von Willebrand factor in scissile bond cleavage by ADAMTS13, Publisher: WILEY-BLACKWELL, Pages: 909-909, ISSN: 1538-7933
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