Thymidine kinase-negative herpes simplex virus mutants establish latency in mouse trigeminal ganglia but do not reactivate.

Abstract

Herpes simplex virus infection of mammalian hosts involves lytic replication at a primary site, such as the cornea, translocation by axonal transport to sensory ganglia and replication, and latent infection at a secondary site, ganglionic neurons. The virus-encoded thymidine kinase, which is a target for antiviral drugs such as acyclovir, is not essential for lytic replication yet evidently is required at the secondary site for replication and some phase of latent infection. To determine the specific stage in viral pathogenesis at which this enzyme is required, we constructed virus deletion mutants that were acyclovir resistant and exhibited no detectable thymidine kinase activity. After corneal inoculation of mice, the mutants replicated to high titers in the eye but were severely impaired for acute replication in trigeminal ganglia and failed to reactivate from ganglia upon cocultivation with permissive cells. Nevertheless, latency-associated transcripts were expressed in neuronal nuclei of ganglia from mutantinfected mice and superinfection of the ganglia with a second virus rescued the latent mutant virus. Thus, contrary to a widely accepted hypothesis, the thymidine kinase-negative mutants established latent infections, implying that neither thymidine kinase activity nor ganglionic replication is necessary for establishment of latency. Rather, thymidine kinase appears to be necessary for reactivation from latency. These results suggest that acyclovir-resistant viruses could establish latent infections in clinical settings and have implications for the use of genetically engineered herpesviruses to deliver foreign genes to neurons.

High efficiency gene transfer to the central nervous system of rodents and primates using herpes virus vectors lacking functional ICP27 and ICP34.5

“…Although a number of viral genes are necessary for neurovirulence the deletion of some of these such as those encoding ICP6 or thymidine kinase does not provide an absolute block to replication in the brain and they are thus not suitable for production of viral vectors. 12,13 However, the ICP34.5 protein has been shown to be absolutely required for a neurovirulent phenotype. Thus ICP34.5 deletion mutants have an LD 50 of Ͼ10 6 p.f.u.…”

Highly Reliable Heterologous System for Evaluating Resistance of Clinical Herpes Simplex Virus Isolates to Nucleoside Analogues

“…The former mechanism is most frequently seen in the clinic (8,16,17,29), probably because TK is not essential for viral replication in most tissues and culture cells (36). However, several reports have demonstrated that TK activity facilitates HSV reactivation from latency in neural ganglia (11,13,44,46).Changes in the TK gene can result in viruses producing no or partial amounts of TK or with an altered substrate specificity (4, 23). Darby et al have proposed a preliminary model for the active center of the HSV type 1 (HSV-1) TK enzyme including three distinct regions: an ATP-binding site (amino acids 51 to 63), a nucleoside-binding site (amino acids 168 to 176), and amino acid 336 (12).…”

Selective in vitro replication of herpes simplex virus type 1 (HSV-1) ICP34.5 null mutants in primary human CNS tumours - evaluation of a potentially effective clinical therapy

“…One of the earliest studies using a thymidine kinase (TK)-negative HSV mutant dlsptk (Coen et al, 1989) demonstrated a dose-dependent improvement in survival of nude mice bearing intracranial tumours following intratumoral therapy (Martuza et al, 1991). However, subsequent studies demonstrated that this mutant is unsuitable for therapeutic use in humans as it can cause encephalitis.…”