Investigating endothelial cell transduction and hexon:PF4 binding of ChAdOx1 in the context of VITT

Abstract

BackgroundVaccines against SARS-CoV2 have been essential in controlling COVID-19 related mortality and have saved millions of lives. Adenoviral (Ad) based vaccines have played an integral part in this vaccine campaign, with licensed vaccines based on the simian Y25 isolate (Vaxzevria, Astrazeneca) and human Ad type 26 (Jcovden, Janssen) widely adopted. As part of the largest global vaccination programme ever undertaken, ultrarare thromboembolic events have been described in approximately 1:200,000 vaccinees administered with Ad based SARS-CoV2 vaccines. ObjectivesThe mechanism underpinning these adverse events remain to be completely delineated, but is characterised by elevated autoantibodies against PF4 which, in complex with PF4, cluster, bind FC{gamma}RIIa on platelets and induce thrombus formation. Here we investigated the ability of ChAdOx1 to transduce and activate endothelial cells. MethodsUsing protein sequence alignment tools and in vitro transduction assays, the ability of ChAdOx1 to infect endothelial cells was assessed. Furthermore, the ability of ChAdOx1 infection to activate endothelial cells was determined. Finally, using surface plasmon resonance we assessed the electrostatic interactions between the ChAdOx1 hexon and PF4. Results and ConclusionsDespite lacking the primary cell entry receptor, Coxsackie and Adenovirus Receptor (CAR), ChAdOx1 efficiently transduced endothelial cells in a CAR-independent manner. This transduction did not result in endothelial cell activation. Purified hexon protein from ChAdOx1 preps did, however, bind PF4 with a similar affinity to that previously reported for the whole ChAdOx1 capsid. These data confirm the need to develop non-PF4 binding adenoviral capsids to reduce the potential adverse events associated with VITT.