Inhibition of foot-and-mouth disease virus in cell cultures with antisense Morpholino oligomers

Foot-and-mouth disease (FMD) is a highly contagious viral disease of cloven-hoofed ungulates that can lead to severe losses in the livestock production and export industries. Although vaccines have been extensively used to control FMD, there is no antiviral therapy available to treat ongoing infections with FMD virus (FMDV). Six peptide-conjugated morpholino oligomers (PPMO), with sequences complementary to various 21 nucleotide segments of the 5' and 3'untranslated regions (UTRs) of the FMDV genome (strain A24 Cruzeiro/Brazil 1955 (A24Cru), were evaluated in cell cultures. Three of the PPMO, targeting domain 5 of the IRES (5D PPMO), and the two translation start codon regions (AUG1 and AUG2 PPMO), showed high anti-FMDV activity. A dose-dependent and sequence-specific reduction in viral titer of greater than 5 log10, with a concomitant reduction of viral protein and RNA expression, was achieved at low micromolar concentrations. Under identical conditions, three other PPMO, targeting the 5'-terminal region of the genome, the cis-acting replication element in the 5' UTR, and the 3' 'ab' stem-loop, showed less dramatic titer reductions of 1.5 to 2 log10. Treatment with 5D PPMO reduced the titer of FMDV strains representing 5 different serotypes by 2-4 log10, compared to controls. A24Cru-infected BHK-21 cells treated repeatedly with 5D or AUG2 PPMO generated resistant virus for which phenotypic and genotypic properties were defined. Notably, three passages with low concentrations of the AUG1 PPMO extinguished all traces of detectable virus. The results indicate that PPMO have potential for treating FMDV infection, and also represent useful tools for study of picornaviral translation and evolution.