INHIBITION OF HUMAN CYTOMEGALOVIRUS IMMEDIATE-EARLY GENE-EXPRESSION BY AN ANTISENSE OLIGONUCLEOTIDE COMPLEMENTARY TO IMMEDIATE-EARLY RNA

ISIS 2922 is a phosphorothioate oligonucleotide that is complementary to human cytomegalovirus (CMV) immediate-early (IE) RNA and that exhib its potent and specific antiviral activity against CMV in cell culture assays, Specific assay systems were developed to separately character ize the antisense and nonantisense components of the antiviral activit y mediated by ISIS 2922, In U373 cells transformed with cDNA encoding the CMV IE 55-kDa (IE55) protein, expression was inhibited at nanomola r concentrations comparable to effective concentrations in antiviral a ssays. The specificity of inhibition was demonstrated by using control oligonucleotides incorporating progressive base changes to destabiliz e oligonucleotide-RNA base pairing and by showing a lack of inhibition of the CMV IE72 product expressed from the same promoter, Inhibition of IE55 protein expression correlated with a reduction in mRNA levels consistent with an RNase II-mediated termination event, Studies with v irus-infected cells demonstrated that antisense and nonantisense mecha nisms contribute to the antiviral activity of ISIS 2922, Base compleme ntarity to target RNA was important for optimal activity in antiviral assays, but base changes affecting parameters other than hybridization affinity also influenced antiviral activity, Sequence-independent inh ibition of virus adsorption to host cells by phosphorothioate oligonuc leotides was also observed at high concentrations. Therefore, at least three different mechanisms may contribute to the antiviral activity o f ISIS 2922 in cell culture: antisense-mediated inhibition of target g ene expression; nonantisense, sequence-dependent inhibition of virus r eplication; and sequence-independent inhibition of virus adsorption to host cells.