4. vWF-related antithrombotic treatment strategies

Despite recent major advances in the vascular biology of thrombosis, we face unmet treatment needs which warrant search for novel antithrombotic medication. Inhibition of vWF secretion constitutes an attractive therapeutic strategy to counteract thrombus initiation and propagation as the plasma concentration of vWF increases in high risk populations and predicts cardiovascular disease outcome. In this section, we review the drugs with the potential of vWF antagonism, illustrating how they might become an option to overcome current limitations of antithrombotic therapy.

activation is sepsis-induced elevation of plasma vWF levels [121, 129, 163, 179]. Sepsis can promote DIC, which is common in critically ill patients. When unfolded, highly multimeric sepsis-induced vWF interacts with platelets [26] and generates small clots in the microvasculature [45, 124]. Microvascular thrombosis appears to be a major pathological mechanism in sepsis pathology resulting in multi-organ dysfunction syndrome (MODS) [124]. Hence, sepsisinduced vWF secretion in excess in DIC and MODS are inevitably linked. In addition, sepsisinduced ultra-large high molecular weight multimeric vWF permits complement activation,

In murine model of sepsis, mice subjected to cecal ligation and puncture (CLP) have increased circulating levels of vWF that promote a procoagulant phenotype and poor outcome [29]. vWF-deficient mice subjected to the CLP model of sepsis exhibit increased survival [180]. For the purpose of testing whether Gα12 modulates vWF secretion in sepsis pathological setting, we subjected Gα12/ mice and WT control mice to the CLP model of polymicrobial sepsis and monitored survival [181]. Our preliminary results indicate that 80% of the Gα12/ mice survived, while all WT mice succumbed in the first 96 h [182]. To further determine whether decreased vWF was responsible for the protective effect of G12 deficiency, we restored circulating vWF level in Gα12/ mice, by intravenous administration of purified vWF [10], to the level observed in WT mice. This procedure resulted in the loss of the protective effect of G12 deletion [182]. Furthermore, we employed a pharmacological G12 inhibition approach to prove G12 involvement in sepsis-induced vWF secretion [183]. We hypothesized that a synthetic peptide derived from Gα12 would inhibit α-SNAP-dependent WPB priming and fusion with the plasma membrane [89, 182], thereby inhibiting vWF secretion from activated ECs,

Transient elevations of plasma vWF level were also observed after epinephrine infusion [184], and chronic elevation of circulating vWF (2–3-fold) was reported in hyperthyroidism, which is thought to be due to β-adrenergic receptor stimulation. In contrast, hypothyroidism is associated with a 15% decrease in vWF level, although this is reversible with appropriate treatment [185]. vWF secretion from ECs might play a role in sickle cell disease pathogenesis [186, 187]. Erythrocytes bind specifically to vWF [186] and stasis, or the reduction of blood flow and shear, promotes binding of erythrocytes to vWF which form fibrin-rich regions in venous thrombi [186].

Despite recent major advances in the vascular biology of thrombosis, we face unmet treatment needs which warrant search for novel antithrombotic medication. Inhibition of vWF secretion constitutes an attractive therapeutic strategy to counteract thrombus initiation and propagation as the plasma concentration of vWF increases in high risk populations and predicts cardiovascular disease outcome. In this section, we review the drugs with the potential of

[121] leading to a positive feedback cycle of inflammation and thrombosis.

and reducing the risk of thrombotic microangiopathy during sepsis [182].

4. vWF-related antithrombotic treatment strategies

3.5. vWF role in other inflammatory diseases

160 Endothelial Dysfunction - Old Concepts and New Challenges

Most of the studies involving anti-vWF antibodies were conducted in animal models. Monoclonal antibodies have been widely tested in murine, monkey and rabbit models of cardiovascular disease, including coronary thrombosis, stroke and in-stent stenosis: GPG-290, 6B4-Fab, AJW200, 82D6A6 and SZ-123 [188, 189]. These are all agents with antithrombotic and antiinflammatory effects which do not prolong bleeding time and do not provoke thrombocytopenia. However, to date, none of these have made it into clinical trials. Hillgruber and colleagues recently published a study that found a massive accumulation of vWF in skin biopsies of patients suffering from leukocytoclastic vasculitis, an immune complex (IC)-mediated vasculitis (ICV) frequently encountered in dermatology that is caused by IC precipitation in the vessel wall followed by recruitment of neutrophils [28]. These results were confirmed in a murine model of vasculitis [28], and importantly, they identified ICs as possible vWF secretagogues. Of note, polyclonal anti-vWF blocking antibodies had an anti-inflammatory effect, reducing leukocyte recruitment and edema formation in a murine model of vascular inflammation [28]. Although the physiological implications of elevated vWF levels (increasing up to 400% of the normal level in various conditions [190, 191]) require additional investigation, it is currently believed that plasma vWF level is a surrogate marker of increased risk of vascular complications in septic patients and in those with vascular disease, which makes vWF antagonism a promising therapeutic target in thrombosis and inflammation.

Aptamers and nanobodies have been demonstrated to directly interfere with the vWF pathway, suggesting their powerful antithrombotic properties and acceptable level of safety may prove to be useful strategies. Aptamers are oligonucleotides that have similar characteristics with monoclonal antibodies and with small molecules. ARC1779 is an aptamer that binds to the A1 domain of vWF, blocking its interaction with GpIb and thereby inhibiting vWFdependent platelet function. Clinical trials with ARC1779 were conducted and completed, but until truly meaningful clinical endpoints can be obtained that indicate effective inhibition of thrombosis without increasing bleeding, the utility of aptamers will remain unclear. Before the clinical trial was prematurely closed by the sponsor, the clinical and laboratory data after 14 days of dosing in nine patients with TTP aged 18–75 were very promising, and the authors optimistically proposed the study continuation (www.clinicaltrials.com; [192]) suggesting further development may be in order.

Nanobodies have the advantage of being highly specific therapeutic agents. Caplacizumab (ALX-0081) is an anti-vWF humanized nanobody that selectively targets vWF A1 domain, locking platelets launching on VWF A1 domain via interaction with their receptor GPIb. TITAN phase II trial concluded that caplacizumab administration in 75 patients with acquired TTP lead to a rapid resolution of the acute TTP episode, but it was adversely seconded by mild bleeding when compared to placebo group [193–195].

Unlike the vWF aptamer or antibodies, a pharmacological inhibitor of vWF secretion would have the advantage of acting from the inside of ECs, thereby limiting the amount of vWF available to support binding of platelets and thrombus formation. Studies thus far from our group suggest inhibition of Gα12-dependent activation of vWF secretion from WPBs may in fact be one such option [89, 90, 181, 182, 196]. Ongoing studies are demonstrating that inhibition of vWF secretion from ECs, using a novel peptide inhibitor to block Gα12/α-SNAP-dependent WPB exocytosis [89], reduces DIC and mortality in septic mice [89, 90, 182, 183, 196].

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