مجال التميز | تميز دراسي و بحثي |
البحوث المنشورة |
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البحث (1): | |
عنوان البحث: |
Structural characterization of a novel GPVI-nanobody complex reveals a biologically active domain-swapped GPVI dimer |
رابط إلى البحث: | |
تاريخ النشر: |
27/01/2021 |
موجز عن البحث: |
Glycoprotein VI (GPVI) is the major signaling receptor for collagen on platelets. We have raised 54 nanobodies (Nb), grouped into 33 structural classes based on their complementary determining region 3 loops, against recombinant GPVI-Fc (dimeric GPVI) and have characterized their ability to bind recombinant GPVI, resting and activated platelets, and to inhibit platelet activation by collagen. Nbs from 6 different binding classes showed the strongest binding to recombinant GPVI-Fc, suggesting that there was not a single dominant class. The most potent 3, Nb2, 21, and 35, inhibited collagen-induced platelet aggregation with nanomolar half maximal inhibitory concentration (IC50) values and inhibited platelet aggregation under flow. The binding KD of the most potent Nb, Nb2, against recombinant monomeric and dimeric GPVI was 0.6 and 0.7 nM, respectively. The crystal structure of monomeric GPVI in complex with Nb2 revealed a binding epitope adjacent to the collagen-related peptide (CRP) binding groove within the D1 domain. In addition, a novel conformation of GPVI involving a domain swap between the D2 domains was observed. The domain swap is facilitated by the outward extension of the C-C′ loop, which forms the domain swap hinge. The functional significance of this conformation was tested by truncating the hinge region so that the domain swap cannot occur. Nb2 was still able to displace collagen and CRP binding to the mutant, but signaling was abolished in a cell-based NFAT reporter assay. This demonstrates that the C-C′ loop region is important for GPVI signaling but not ligand binding and suggests the domain-swapped structure may represent an active GPVI conformation. |
البحث (2): | |
عنوان البحث: |
Novel antiplatelet targets in the treatment of acute coronary syndromes |
رابط إلى البحث: |
https://www.tandfonline.com/doi/full/10.1080/09537104.2020.1763731 |
تاريخ النشر: |
12/06/2020 |
موجز عن البحث: |
Acute coronary syndromes (ACS) are a global cause of mortality and morbidity that affect millions of lives worldwide. Following atherosclerotic plaque rupture, platelet activation and aggregation are the two major elements that initiate thrombus formation inside a coronary artery, which can obstruct blood flow and cause myocardial ischemia; ergo, antiplatelet therapy forms a major part of the treatment strategy for ACS. Patients with ACS routinely receive dual antiplatelet therapy (DAPT), which consists of aspirin and a platelet P2Y12 inhibitor to both treat and prevent atherothrombosis. Use of platelet glycoprotein (GP) IIb/IIIa inhibitors is now limited due to the risk of severe bleeding and thrombocytopenia. Thus, administration of GPIIb/IIIa inhibitors is generally restricted to bail out thrombotic events associated with PCI. Furthermore, current antiplatelet medications mainly rely on thromboxane A2 and P2Y12 inhibition, which have broad-acting effects on platelets and are known to cause bleeding, which especially limits the long-term use of these agents. In addition, not all ACS patients treated with current antiplatelet treatments are protected from recurrence of arterial thrombosis, since many platelet mechanisms and activation pathways remain uninhibited by current antiplatelet therapy. Pharmacological antagonism of novel targets involved in platelet function could shape future antiplatelet therapies that could ultimately lead to more effective or safer therapeutic approaches. In this article, we focus on inhibitors of promising targets that have not yet been introduced into clinical practice, including inhibitors of GPVI, protease-activated receptor (PAR)-4, GPIb, 5-hydroxytryptamine receptor subtype 2A (5-HT2A), protein disulfide isomerase, P-selectin and phosphoinositide 3-kinase β. |
البحث (3): | |
عنوان البحث: |
SMIFH2 inhibition of platelets demonstrates a critical role for formin proteins in platelet cytoskeletal dynamics |
رابط إلى البحث: | |
تاريخ النشر: |
13/01/2020 |
موجز عن البحث: |
Background: Reorganization of the actin cytoskeleton is required for proper functioning of platelets following activation in response to vascular damage. Formins are a family of proteins that regulate actin polymerization and cytoskeletal organization via a number of domains including the FH2 domain. However, the role of formins in platelet spreading has not been studied in detail. Objectives: Several formin proteins are expressed in platelets so we used an inhibitor of FH2 domains (SMIFH2) to uncover the role of these proteins in platelet spreading and in maintenance of resting platelet shape. Methods: Washed human and mouse platelets were treated with various concentrations of SMIFH2 and the effects on platelet spreading, platelet size, platelet cytoskeletal dynamics, and organization were analyzed using fluorescence and electron microscopy. Results: Pretreatment with SMIFH2 completely blocks platelet spreading in both mouse and human platelets through effects on the organization and dynamics of actin and microtubules. However, platelet aggregation and secretion are unaffected. SMIFH2 also caused a decrease in resting platelet size and disrupted the balance of tubulin post-translational modification. Conclusions: These data therefore demonstrated an important role for formin-mediated actin polymerization in platelet spreading and highlighted the importance of formins in cross-talk between the actin and tubulin cytoskeletons. |
المؤتمرات العلمية |
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المؤتمر (1): | |
عنوان المؤتمر: |
1st Platelet Society Annual Meeting |
تاريخ الإنعقاد: |
12-13/09/2019 |
مكان الإنعقاد: |
Cambridge, UK |
طبيعة المشاركة: |
Oral presentation |
عنوان المشاركة: |
Synergistic antiplatelet effects of a novel GPVI inhibitor ACT017 when used in combination with aspirin and ticagrelor |
ملخص المشاركة: |
Background: Aspirin and a P2Y12 inhibitor, such as ticagrelor, are routine treatments for myocardial infarction (MI). However, these drugs are not always sufficient for heavy coronary thrombus burden during ST-elevation MI (STEMI). More potent antiplatelet drugs (GPIIb/IIIa-inhibitors) may help in this setting but are limited by excessive bleeding. Aims: Since GPVI has major roles in thrombosis but is much less important than GPIIb/IIIa for haemostasis, we aimed to investigate whether a novel platelet GPVI inhibitor, ACT017 (Acticor Biotech), provides additional antithrombotic effects when combined with aspirin and ticagrelor. Methods: Blood was sampled from healthy volunteers and ACT017 (a clinicalgrade Fab fragment of a monoclonal antibody) was added to blood with or without aspirin (30μM) or ticagrelor (1μM) and compared to eptifibatide (9μM) ex-vivo. Platelet aggregation was assessed using light transmission aggregometry (PAP8) and multiple electrode aggregometry (Multiplate). Platelet thrombin generation in response to tissue factor was assessed using a calibrated automated thrombogram. Results: ACT017 (50μg/ml) potently inhibited collagen- and fibrin-induced platelet aggregation. Addition of ACT017 to aspirin and ticagrelor provided significantly greater inhibition of platelet aggregation induced by collagen. A 10-fold lower concentration of ACT017 (5μg/ml) could also maximally inhibit collagen-induced platelet aggregation in the presence of aspirin and ticagrelor. ACT017 and eptifibatide (a GPIIb/IIIa antagonist) both provided similar inhibition of collagen-induced platelet aggregation when used in combination with aspirin and ticagrelor. ACT017 also inhibited peak platelet thrombin generation and did not cause any off-target inhibition of non-GPVI-mediated pathways. Conclusion: This is the first study to show that the addition of a novel GPVI inhibitor, ACT017, to aspirin and ticagrelor provides greater inhibition of multiple critical mechanisms of arterial thrombosis. This strategy is appealing for further development for the treatment of STEMI due to the minimal role of GPVI in haemostasis. |
الرابط: | |
المؤتمر (2): | |
عنوان المؤتمر: |
BSHT Annual Scientific Meeting |
تاريخ الإنعقاد: |
23-24 /01/2020 |
مكان الإنعقاد: |
Birmingham. UK |
طبيعة المشاركة: |
Oral presentation |
عنوان المشاركة: |
AMPLIFIED ANTITHROMBOTIC EFFECTS OF A NOVEL GPVI INHIBITOR ACT017 WHEN USED IN COMBINATION WITH ASPIRIN AND TICAGRELOR |
ملخص المشاركة: |
Aspirin and a P2Y12 inhibitor, such as ticagrelor, are routine treatments for myocardial infarction (MI). However, these drugs are not always sufficient for heavy coronary thrombus burden during ST-elevation MI (STEMI). More potent antiplatelet drugs (GPIIb/IIIa-inhibitors) may help in this setting, but are limited by excessive bleeding. Aims: Since GPVI has major roles in thrombosis but is much less important than GPIIb/IIIa for haemostasis, we aimed to investigate whether a novel platelet GPVI inhibitor, ACT017 (Acticor Biotech), provides additional antithrombotic effects when combined with aspirin and ticagrelor. Methods: Blood was sampled from healthy volunteers and ACT017 (a clinical-grade Fab fragment of a monoclonal antibody) was added to blood with or without aspirin (30µM) or ticagrelor (1µM) and compared to eptifibatide (9µM) ex-vivo. Platelet aggregation was assessed using light transmission aggregometry (PAP8) and multiple electrode aggregometry (Multiplate). Platelet thrombin generation in response to tissue factor was assessed using a calibrated automated thrombogram. Results: ACT017 (50µg/ml) potently inhibited collagen- and fibrin-induced platelet aggregation. Addition of ACT017 to aspirin and ticagrelor provided significantly greater inhibition of platelet aggregation induced by collagen. A 10-fold lower concentration of ACT017 (5µg/ml) could also maximally inhibit collagen-induced platelet aggregation in the presence of aspirin and ticagrelor. ACT017 and eptifibatide (a GPIIb/IIIa antagonist) both provided similar inhibition of collagen-induced platelet aggregation when used in combination with aspirin and ticagrelor (Figure 1C). ACT017 also inhibited peak platelet thrombin generation and did not cause any off-target inhibition of non-GPVI-mediated pathways (not shown). Conclusion: This is the first study to show that the addition of a novel GPVI inhibitor, ACT017, to aspirin and ticagrelor provides greater inhibition of multiple critical mechanisms of arterial thrombosis. This strategy is appealing for further development for the treatment of STEMI due to the minimal role of GPVI in haemostasis. |
الرابط: |
https://bsht.org.uk/wp-content/uploads/2020/02/BSHT-2020-programme-book-for-website.pdf |
المؤتمر (3): | |
عنوان المؤتمر: |
ESC Congress 2021 – The Digital Experience |
تاريخ الإنعقاد: |
28/08/2021 |
مكان الإنعقاد: |
Paris, France |
طبيعة المشاركة: |
Oral presentation in young investigator award session in thrombosis |
عنوان المشاركة: |
GPVI inhibition by glenzocimab synergistically inhibits atherosclerotic plaque-induced platelet activation when combined with conventional dual antiplatelet therapy |
ملخص المشاركة: |
Introduction: Aspirin and a potent platelet P2Y12 inhibitor, such as prasugrel or ticagrelor, are not always sufficient to prevent thrombus formation in patients with ST-elevation MI (STEMI), leading to “slow flow” or “no reflow” effects after stenting. GPIIb/IIIa inhibitors, such as eptifibatide, may help in this setting, but are not used routinely due to their bleeding risk. GPVI has critical roles in thrombosis and a minimal role in haemostasis. Here we tested whether depletion of GPVI has effects on thrombus formation after MI in an animal model and investigated the effects of a novel platelet GPVI inhibitor, glenzocimab (a Fab fragment of a monoclonal antibody), on platelet activation and thrombus formation when combined with aspirin and ticagrelor. Methods: We used intravital microscopy in a murine model of ST-elevation myocardial infarction and ischaemia-reperfusion injury to investigate microvascular thrombosis. We investigated the antithrombotic effects of adding glenzocimab (previously known as ACT017) to blood from healthy donors and 20 patients with ACS treated with aspirin and ticagrelor. We compared the effect of glenzocimab with the GPIIb/IIIa inhibitor eptifibatide ex-vivo. We stimulated platelets with collagen and atherosclerotic plaque material that was sourced from patients undergoing carotid endarterectomy. We investigated effects on platelet aggregation, spreading, signalling, adhesion, thrombin generation, thrombus formation and clot stability ex vivo. Results: Genetic depletion of GPVI in an animal model of myocardial infarction reduced microvascular thrombosis. Ex vivo, aspirin and ticagrelor partially inhibited atherosclerotic plaque-induced platelet aggregation (assessed by multiple electrode aggregometry) by 48% compared to control (34±3 vs. 65±4 U; P < 0.001; Figure 1). Atherosclerotic plaque-induced platelet aggregation, adhesion, secretion and activation were critically dependent on platelet GPVI activation and were potently inhibited by glenzocimab. Glenzocimab alone reduced atherosclerotic plaque-induced platelet aggregation by 75% compared to control (16±4 vs. 65±4 U; P < 0.001; Figure 1) and by over 95% when combined with aspirin and ticagrelor (3±1 vs 65±4 U; P <0.001; Figure 1). Furthermore, glenzocimab provided multiple synergistic antithrombotic effects when added to the blood of aspirin and ticagrelor-treated patients with ACS ex vivo. Glenzocimab and the GPIIb/IIIa inhibitor, eptifibatide, had many similar antithrombotic effects but glenzocimab had less effect on mechanisms of general haemostasis compared to eptifibatide, as assessed by ROTEM (Figure 2). Conclusions: The addition of glenzocimab to aspirin and ticagrelor provides synergistic inhibition of multiple critical mechanisms of atherothrombosis. Glenzocimab and the GPIIb/IIIa inhibitor, eptifibatide, share many similar antithrombotic effects, although glenzocimab has less impact on mechanisms involved in haemostasis compared to eptifibatide. |
الرابط: | |
أخرى |
One of the Top Posters in the ISTH 2019 Congress For the poster entitled: – Amplified antithrombotic effects of a novel GPVI inhibitor ACT017 when used in combination with aspirin and ticagrelor. |
الرابط: |
https://2019.isthcongressdaily.org/top-posters-for-sunday-july-7/ |
فواز عبيدالله مطيران العنزي
دكتوراه
الطب والخدمات الصحية
University of Birmingham