SYNERGISTIC ANTIVIRAL ACTIVITIES OF OLIGONUCLEOSIDE METHYLPHOSPHONATES COMPLEMENTARY TO HERPES SIMPLES VIRUS TYPE-1 IMMEDIATE-EARLY MESSENGER RNA-4, RNA-5, AND RNA-1

An oligonucleoside methylphosphonate (ONMP) complementary to the splic e acceptor site of immediate-early (IE) pre-mRNAs 4 and 5 (IE4,5SA) in hibits herpes simplex virus type 1 (HSV-1) growth in vitro and in infe cted animals. The antiviral effect appears to be due to inhibition of IE pre-mRNA 4 and 5 splicing and/or IE4 gene expression (M. Kulka, M. Wachsman, S. Miura, R. Fishelevich, P. S. Miller, P. 0. P. Ts'o, and L . Aurelian, Antiviral Res. 20:115-130, 1993). We describe the potentia tion of antiviral activity when we targeted two IE genes with differen t ONMPs. A psoralen derivative of an ONMP complementary to the IE mRNA 1 (IE1) translation initiation site (IE1TI) covalently bound a 2.8-kb transcript that hybridized with a 20-base oligonucleotide complementa ry to the 5' leader sequence of IE1 but not a 20-base oligonucleotide complementary to the first intron of IE1. IE1TI inhibited IE1 gene exp ression and virus replication in cells infected with HSV-1 in vitro. I nhibition was specific because it was not observed with oligomers muta ted in two (IE1TImu1) or four (IE1TImu2) central residues or in cells infected with an IE1 deletion mutant (HSV-1 dl1403). IE1TI potentiated the antiviral activity of IE4,5SA (synergistic effect), while potenti ation was not observed when IE4,5SA was mixed with IE1TImu1. A similar synergistic effect was seen when IE1TI was mixed with an ONMP complem entary to the translation initiation site of IE mRNA 4 but not with an ONMP complementary to the translation initiation site of IE mRNA 5. T hese findings suggest that synergistic antiviral activity is mediated by targeting at least two IE genes (IE1 and IE4).