Högskolan i Skövde

Human rhinovirus (HRV) is one of the leading causes of upper respiratory tract infections in humans. This infection is usually mild and self-limiting in the immunocompetent host, but in some cases HRV infection can be associated with bronchiolitis in infants, pneumonia in immune suppressed, and the deterioration of pre-existing respiratory diseases such as asthma. There is currently no vaccine or drug for clinical prophylactic/therapeutic use against HRV infection, mostly because of the fact that this virus exists in over 150 serotypes which are sufficiently different and thereby difficult to target by a single vaccine or antiviral compound.

Here, a HeLa cell-based assay was developed to be used for screening the cellular kinase inhibitor library (KIL) for anti-HRV activity. The KIL library comprise 1200 of kinase inhibitors, and the screening identified 22 hits that completely protected HeLa cells against the development of HRV-induced cytopathic effect (CPE) at concentrations of 1 μM and 10 μM. Thereafter, these hits were assessed in a dose-dependent manner to establish their concentration that inhibited the viral CPE in 50% (EC₅₀). Of these hits, two kinase inhibitors, i.e., a phosphatidylinositol 3-kinase (PI3Ka) inhibitor and an epidermal growth factor receptor tyrosine kinase (EGFR-TK) inhibitor were, investigated in more detail. In particular, it was found that the PI3K and EGFR inhibitors blocked HRV infection in HeLa cell cultures at EC₅₀ of 0.64 μM and 0.49 μM, respectively. Furthermore, these two compounds exhibited 50% cytotoxicity (CC₅₀) for HeLa cells at >100 μM and 13 μM, respectively, which resulted in a selectivity index (SI; CC₅₀/EC₅₀) of >156 for the PI3K inhibitor and 27 for the EGFR-TK inhibitor. These data indicated that the anti-HRV activity of tested of the non-toxic PI3K, and EGFR inhibitors was virus selective and occurred at comparatively low concentrations. Attempts to identify the mode of antiviral activity showed that both inhibitors lacked a direct virus-inactivating (virucidal) activity. Instead, the time-of-addition assay revealed that the PI3Ka inhibitor exhibited antiviral activity even when added as late as 10 h after HRV infection of HeLa cells while the EGFR-TK inhibitor was active when added at 10-12 h post infection. This suggests that both these inhibitors affected the late stage of HRV life cycle, most likely by abrogating release of progeny HRV particles from infected cells.

In conclusion, these results indicate that the cellular kinases represent an important host-related target for anti-HRV intervention. A further identification and characterization of kinase inhibitors that block the virus infection of cells, and their responsible mechanisms of action, may help in the development of potential anti-HRV drugs.