Characterization of anti-enteroviral activity of heparan sulphate mimetic compounds

Enterovirus 71 (EV71) infection remains a public health problem at a global level, particularly in the Asia-pacific region. The infection normally manifests as hand-foot-mouth disease (HFMD); however, it is capable of developing into potentially fatal neurological complications. There is currently no approved vaccine or antiviral substance available for prevention or treatment of EV71 infection. There has been no research undertaken to examine the possible antiviral activity of heparan sulphate (HS) mimetics against the Human Enterovirus A (HEV-A), in particular EV71. Therefore, this PhD research began with the aim of investigating the antiviral potencies of HS mimetics against EV71 infection. Initially, a colorimetric-based method was developed for the titration of EV71 strains, whereby the viral titre was quantified more precisely. Three soluble HS mimetics including heparin (Hep), HS, and pentosan polysulphate (PPS) were then shown to substantially inhibit a cloned strain of EV71 from infecting in Vero cells. Further investigations revealed that the compounds most likely exerted their antiviral action through interference with the EV71 attachment. The role of cell surface HS in mediating viral infection was then studied for several clinical isolates of HEV-A and HEV-B in Vero cells as well as a human neuroblastoma cell line, SK-N-SH. The findings revealed that the clinical isolate of EV71 utilizes low sulphated domains of cellular HS to bind to Vero cells, in contrast to Coxsackievirus B4 (CVB4), Coxsackievirus A16 (CVA16), and the cloned EV71. In the neural cells, Hep and PPS significantly prevented both clinical EV71 and clinical CVA16 binding and infections, although it could not be confirmed whether the viruses utilize cellular HS to bind to the cells. Finally, an Affymetrix DNA microarray was performed to gain insight into the mechanisms of action of Hep against the clinical EV71 infection in the neural cells. The results showed many genes with significant down- or up-regulation across the undermentioned conditions: negative control cells, compound control (cells treated with Hep only), virus control (cells treated with virus only), and treatment control (EV71-infected cells treated with Hep). Several genes were finally found to be targets for the anti-EV71 activity of Hep in SK-N-SH cells using a strict multi-level selection procedure. In parallel, the results revealed the significant induction of more than 1000 genes by EV71 infection with expression fold changes ranged from +46.5 to -10.7. The findings of this research may suggest new directions for studies of designing molecular drug targets against EV71 infection.