IN-VIVO COMPLEMENTATION STUDIES OF A GLYCOPROTEIN H-DELETED HERPES-SIMPLEX VIRUS-BASED VECTOR

The utilization of herpes simplex virus (HSV) as a vector for gene del ivery to the nervous system or as a live vaccine delivery system is de pendent on the construction and characterization of disabled virus mut ants which are unable to cause disease, Under certain circumstances, h owever, replication-defective vectors may carry a potential risk if th ey can be efficiently complemented by a co-infecting wild-type virus. Stocks of defective vectors should, therefore, be free from replicatio n-competent virus, and helper cell lines should be incapable of genera ting replication-competent virus by recombination between the vector a nd the complementary gene, We describe a glycoprotein H-negative (gH(- )) virus/helper cell line combination which generates helper-free defe ctive virus stocks containing replication-competent virus at a frequen cy no higher than 1 in 10(9) p.f.u, This virus/helper cell system prov ides a suitable background for the construction of safe replication-de fective gene delivery vectors, In vivo studies demonstrate that gH(-) virus is unable to initiate disease in mice and establishes latency at low efficiency compared to wild-type HSV. To determine whether gH(-) virus can be complemented by wild-type virus in vivo, mice were infect ed with a variety of mixtures of these viruses, Complementation was ob served in a minority of animals infected with more than 10(6) p,f,u. O f both wild-type and defective virus but the most common observation w as that the presence of defective virus suppressed entry of wild-type virus into the nervous system.