Anti-thrombotic efficacy of S007-867: Pre-clinical evaluation in experimental models of thrombosis in vivo and in vitro

Abstract

Pharmacological inhibition of platelet collagen interaction is a promising therapeutic strategy to treat intra-vascular thrombosis. S007-867 is a novel synthetic inhibitor of collagen-induced platelet aggregation. It has shown better antithrombotic protection than aspirin and clopidogrel with minimal bleeding tendency in mice. The present study is aimed to systematically investigate the antithrombotic efficacy of S007-867 in comparison to aspirin and clopidogrel in vivo and to delineate its mechanism of action in vitro. Aspirin, clopidogrel, and S007-867 significantly reduced thrombus weight in arterio-venous (AV) shunt model in rats. In mice, following ferric chloride induced thrombosis in either carotid or mesenteric artery; S007-867 significantly prolonged the vessel occlusion time (1.2-fold) and maintained a sustained blood flow velocity for >30 min. Comparatively, clopidogrel showed significant prolongation in TTO (1.3-fold) while aspirin remained ineffective. Both S007-867 and aspirin did not alter bleeding time in either kidney or spleen injury models, and thus maintained hemostasis, while clopidogrel showed significant increase in spleen bleeding time (1.7-fold). The coagulation parameters namely thrombin time, prothrombin time or activated partial thromboplastin time remained unaffected even at high concentration of S007-867 (300 µM), thus implying its antithrombotic effect to be primarily platelet mediated. S007-867 significantly inhibited collagen-mediated platelet adhesion and aggregation in mice ex-vivo. Moreover, when blood was perfused over a highly thrombogenic combination of collagen mimicking peptides like CRP-GFOGER-VWF-III, S007-867 significantly reduced total thrombus volume or ZV (53.4 ± 5.7%). Mechanistically, S007-867 (10-300 μM) inhibited collagen-induced ATP release, thromboxane A (TxA) generation, intra-platelet [Ca] flux and global tyrosine phosphorylation including PLCγ2. Collectively the present study highlights that S007-867 is a novel synthetic inhibitor of collagen induced platelet activation, that effectively maintains blood flow velocity and delays vascular occlusion. It inhibits thrombogenesis without compromising hemostasis. Therefore, S007-867 may be further developed for the treatment of thrombotic disorders in clinical settings.

Hsp47 Inhibitor Col003 Attenuates Collagen-Induced Platelet Activation and Cerebral Ischemic–Reperfusion Injury in Rats

“…A total of 250 μl of washed platelet (2 × 10 7 /ml concentration, 1x Tyrodes buffer) containing 1 mM of CaCl 2 was incubated with Col003 (25 or 50 μM) or vehicle for 5 minutes and subsequently added to the cover slips (coated with 5 μg/ml of collagen) for 1 hour. Finally, the fixed platelets were stained with 1 μg/ml of FITC-labeled phalloidin for 15 min and protected from light, as previously described ( Misra et al, 2018 ). Images from at least three different microscope fields were captured with an Olympus fluorescence microscope (Olympus Corp., Tokyo, Japan) and analyzed with the ImageJ software (United States National Institutes of Health, Bethesda, MD, United States).…”

Oral glucose feeding enhances adherence of quiescent lymphocytes to fibronectin via non-canonical insulin signalling

“…Mesenteric artery of the recipient mice was exposed through an abdominal wall incision and a Whatmann filter paper (2mm x 0.5mm) saturated with 5% FeCl 3 (w/v) solution was applied topically for 3 minutes for inducing the endothelial/vessel injury. Adherence of labelled donor PBMCs to the exposed basement membrane of the mesenteric artery of the recipient mice was monitored under intra-vital microscope [Olympus FV1200 (BX61Wi), Japan] coupled to a CCD camera [28, 29]. The images were captured and quantified using FV1000 (Olympus) and image J software.…”

Feeding enhances fibronectin adherence of quiescent lymphocytes through non‐canonical insulin signalling

“…Mesenteric artery of the recipient mice was exposed through an abdominal wall incision and a Whatmann filter paper (2 mm  0.5 mm) saturated with 5% FeCl 3 (w/v) solution was applied topically for 3 min for inducing vessel injury. Adherence of labelled donor PBMCs to the exposed basement membrane of the mesenteric artery of the recipient mice was monitored under intravital microscope [Olympus FV1200 (BX61Wi), Japan] coupled to a CCD camera [11,12]. The images were captured and quantified using FV1000 (Olympus) and ImageJ software.…”