Principal Investigator: Tracy Stokol

DESCRIPTION (provided by applicant): 

Equine herpes virus type 1 (EHV-1) is a highly contagious pathogen, infecting horses worldwide. The virus, which is acquired by inhalation, causes pneumonia and neonatal deaths. In addition, the virus can spread systemically, resulting in abortion and a devastating neurologic syndrome, equine herpes myeloencephalopathy (EHM), in adult horses. Diagnosis of EHV-1 at a training facility or racetrack has a huge negative economic impact due to event cancellations and implementation of biosecurity procedures to limit virus transmission. Beyond financial burden, owners experience emotional toll from their horses’ suffering and need for euthanasia. Due to short-lived immunity, incomplete vaccine protection, and latency, outbreaks of disease continue to occur. The Equine Disease Communication Center reported 21 outbreaks of EHV-1 in the USA to date in 2016. In many states, EHV-1 is a reportable disease. The devastating clinical consequences of abortion and EHM are unpredictable and veterinarians have no effective means of preventing their occurrence in EHV-1-exposed horses. One of the major contributing factors to abortion and EHM is ischemic necrosis in the placenta and spinal cord. The ischemia is caused by thrombosis and vasculitis in placental and spinal blood vessels. We propose that treating horses with anticoagulant drugs will reduce the risk of abortion and EHM in EHV-1-exposed horses. How EHV-1 infection results in thrombosis is unknown. Thrombosis requires activation of the coagulation cascade to generate thrombin, which forms the fibrin clot, and activation of platelets, which amplify thrombin generation and form part of the clot. EHV-1 infection causes a hypercoagulable or prothrombotic state, characterized by excess thrombin generation. This hypercoagulability occurs during viremia, which coincides with the timing of abortion and EHM. We have preliminary data showing that platelets are also activated in horses during viremia. Together, this data supports the role of thrombin and activated platelets in pathologic thrombus formation that manifests as abortion and EHM. The mechanism of EHV-1 induced thrombosis is complex. We have previously shown that EHV-1 upregulates tissue factor on monocytes. Tissue factor activates coagulation, which culminates in fibrin formation. We have also shown that EHV-1 associates with and activates platelets. Thrombin, the key terminal enzyme of the coagulation cascade, is central to both of these prothrombotic EHV-1 actions. Here we show with preliminary data that the anticoagulant drugs, unfractionated heparin (UFH), low molecular weight heparin (LMWH), and apixaban (a direct inhibitor of activated factor X [Xa], which is upstream of thrombin), abolish EHV-1- induced platelet activation when spiked into equine platelet-rich plasma (PRP) ex vivo. Importantly, in a previous Grayson-funded study, we found that anti-platelet drugs, including aspirin and clopidogrel, were ineffective in blocking EHV-1-induced platelet activation. Our first Aim will be to confirm that the observed ex vivo inhibitory effects of UFH or LMWH can be reproduced with in vivo administration. If we find that these parenteral drugs are effective, they would be immediately available for thromboprophylaxis in EHV-1-infected horses. However, these heparin products have substantial disadvantages, such as the need for parenteral administration and injection site reactions. Therefore, an oral alternative would be advantageous. In Aim 2, by defining the pharmacokinetics of a newly approved human oral anticoagulant, apixaban, in horses, this study will provide necessary data for future drug trials of this orally active drug.