Multiple Immediate-Early Gene-Deficient Herpes Simplex Virus Vectors Allowing Efficient Gene Delivery to Neurons in Culture and Widespread Gene Delivery to the Central Nervous System In Vivo

Abstract

Herpes simplex virus (HSV) has several potential advantages as a vector for delivering genes to the nervous system. The virus naturally infects and remains latent in neurons and has evolved the ability of highly efficient retrograde transport from the site of infection at the periphery to the site of latency in the spinal ganglia. HSV is a large virus, potentially allowing the insertion of multiple or very large transgenes. Furthermore, HSV does not integrate into the host chromosome, removing any potential for insertional activation or inactivation of cellular genes. However, the development of HSV vectors for the central nervous system that exploit these properties has been problematical. This has mainly been due to either vector toxicity or an inability to maintain transgene expression. Here we report the development of highly disabled versions of HSV-1 deleted for ICP27, ICP4, and ICP34.5/open reading frame P and with an inactivating mutation in VP16. These viruses express only minimal levels of any of the immediate-early genes in noncomplementing cells. Transgene expression is maintained for extended periods with promoter systems containing elements from the HSV latency-associated transcript promoter (J. A. Palmer et al., J. Virol. 74:5604-5618, 2000). Unlike less-disabled viruses, these vectors allow highly effective gene delivery both to neurons in culture and to the central nervous system in vivo. Gene delivery in vivo is further enhanced by the retrograde transport capabilities of HSV. Here the vector is efficiently transported from the site of inoculation to connected sites within the nervous system. This is demonstrated by gene delivery to both the striatum and substantia nigra following striatal inoculation; to the spinal cord, spinal ganglia, and brainstem following injection into the spinal cord; and to retinal ganglion neurons following injection into the superior colliculus and thalamus.

HSV trafficking and development of gene therapy vectors with applications in the nervous system

“…Inoculation at the striatum, spinal cord, and superior colliculus was accompanied by expression of transgene at the SNc, retinal ganglion cells, DRG, brainstem, hind-, and mid-brain. 147 These results demon-HSV vector trafficking AR Frampton et al strated that an HSV vector could be efficiently delivered to, and utilize the retrograde transport machinery of, CNS neurons without a decrease in transgene expression, and suggest that HSV gene therapy may be extended to applications involving CNS disorders.…”

“…The development of vectors that are deleted for multiple IE genes has allowed the successful use of this system in the CNS with little observed toxicity and vectors for applications in neurodegenerative disease are currently under development. [145][146][147] In one study, reporter gene expression at CNS sites remote from the site of initial inoculation was observed. Inoculation at the striatum, spinal cord, and superior colliculus was accompanied by expression of transgene at the SNc, retinal ganglion cells, DRG, brainstem, hind-, and mid-brain.…”

Innocuous full-length botulinum neurotoxin targets and promotes the expression of lentiviral vectors in central and autonomic neurons

“…First, no retro-axonal transport has been demonstrated conclusively for lentiviral vectors, although this has been seen with other vector systems. [36][37][38] Second, the transduction mechanisms of retroviruses and their relatively large size (4100 nm) would strongly disfavour their entrapment within syt-II bearing small synaptic vesicles for retro-axonal trafficking. 39 The viability of CS-BoTIM/B as a moiety for guiding viral vectors to neurons has been established through transduction of functional MTR-S25.…”

Infection of mature dendritic cells with herpes simplex virus type 1 dramatically reduces lymphoid chemokine-mediated migration

“…The HSV-1 pR20.5/vhs strain contains a cassette expressing enhanced green fluorescent protein (EGFP) and LacZ. This cassette has been inserted into the single NruI site in the HSV-1 UL41 region, which leads to a deletion of the vhs gene as described previously (Lilley et al, 2001). HSV-1 wt 'ang' represents an HSV-1 wild-type laboratory strain.…”