Antiherpes virus activity and effect on deoxyribonucleoside triphosphate pools of (E)-5-(2-bromovinyl)-2′-deoxycytidine in combination with deaminase inhibitors

Abstract

The antiviral activity and cytotoxicity of (E)-5-(2-bromovinyl)-2′-deoxycytidine (BrVdCyd) against herpes simplex virus type 1 (HSV-1), singly and in combination with deaminase inhibitors was determined using rabbit kidney (RK-13), HEP-2, BHK-21 and VERO cells. BrVdCyd was a potent inhibitor of HSV-1 replication with ED 50 values of 0.30 to 1.20 μM depending on the cell line used. In the presence of tetrahydrouridine or tetrahydrodeoxyuridine (H 4dUrd), potency of BrVdCyd increased approximately two fold (ED 50: 0.54 μM) in HSV-infected VERO cells. The combination of BrVdCyd and H 4dUrd was also effective in decreasing virus yield. Dihydrodeoxyuridine (H 2dUrd) reversed the activity of BrVdCyd (ED 50: 6 to 7 μM). The effect of (E)-5-(2-bromovinyl)-2′-deoxyuridine (BrVdUrd), BrVdCyd and BrVdCyd in combination with H 4dUrd on deoxyribonucleoside triphosphate (dNTP) pools was assessed in VERO cells infected with a high multiplicity of infection (10 PFU/cell). Significant differences in dNTP poll sizes (pmol/10 6 cell) were observed with different treatments. BrVdUrd and BrVdCyd treatment resulted in marked expansion of the dTTP pool (> 1200 pmol) compared to HSV-infected VERO cells (303 pmol). Exposure to H 4dUrd resulted in a 12-fold expansion of the dCTP pool (326 pmol) and barely detectable levels of dTTP (<1.0 pmol). BrVdCyd plus H 4dUrd treatment resulted in a slight expansion of the dTTP pool (515 pmol). These results indicate: (i) H 4dUrd inhibits de novo dCyd/dCMP deaminase pathway and (ii) exposure to BrVdCyd plus H 4dUrd puts a strain on viral DNA synthesis to such an extent that even though dTTP is being formed from alternative pathways, its eventual utilization as a substrate is reduced and hence it builds up.