Significantly Increased Expression of β-Glucuronidase in the Central Nervous System of Mucopolysaccharidosis Type VII Mice from the Latency-Associated Transcript Promoter in a Nonpathogenic Herpes Simplex Virus Type 1 Vector

Abstract

Herpes simplex virus (HSV) has the ability to establish life-long latent infections in postmitotic neurons and to remain transcriptionally active, continuously expressing latency-associated transcripts (LAT) while producing minimal disease. These properties have made HSV an excellent candidate for neuronal gene transfer. Previously, we have shown that in mucopolysaccharidosis type VII mice (MPS VII, beta-glucuronidase deficiency) the LAT promoter is capable of expressing beta-glucuronidase (GUSB) in the trigeminal ganglion and the brainstem after latency is established. However, the number of neurons expressing GUSB is much lower than the number expressing 2-kb LAT following a wild-type virus infection. In this study, we have evaluated the effect of the position of the coding sequence relative to the LAT promoter on beta-glucuronidase gene expression in the central nervous system (CNS). Non-neurovirulent (ICP-34.5-deleted HSV-1) vectors were used, allowing direct intracranial injection. Significantly more GUSB activity was detected in brains of MPS VII mice inoculated with a recombinant virus (HSV-LAT-GUSB-JS) in which the GUSB cDNA was inserted near the LAT promoter, compared to viruses where it was inserted farther downstream in either the LAT exon 1 or overlapping exon 1 and the 2-kb LAT intron. This vector produced more than 100 times the number of positive cells than the other constructs. During acute infection, the distribution of viral replication differed from the distribution of GUSB enzyme expression. Viral antigen was predominately present in cells around the site of injection in the caudate putamen and in ependymal cells lining the ventricles. In contrast, GUSB expression was present mainly in cells of the thalamus and hypothalamus, which did not exhibit viral antigen, suggesting that GUSB enzyme activity was expressed from latently but not acutely infected neuronal cells. This vector design should be useful for high-level expression of various genes in the CNS.

Deletion of the Virion Host Shut-off Gene Enhances Neuronal-Selective Transgene Expression from an HSV Vector Lacking Functional IE Genes

“…HSV genomes persisting in neurons do not integrate and can serve as a platform for therapeutic gene expression. Replication-defective HSV vectors have been tested for gene therapy approaches in the CNS1, 2, 3 and peripheral nervous system 4, 5, 6, 7. However, CNS applications have been plagued by residual neuronal toxicity and progressive loss of transgene expression 8, 9, 10.…”

Critical Issues in Gene Therapy for Neurologic Disease

“…HSV, Ad, AAV, and lentivirus vectors have all been used for direct gene transfer of lysosomal enzymes to the CNS (Wolfe et al, 1992a;Ghodsi et al, 1998;Skorupa et al, 1999;Bosch et al, 2000a,b;Sferra et al, 2000;Zhu et al, 2000;Shen et al, 2001). Although both HSV and Ad recombinant vectors are capable of mediating strong gene expression in many cell types in the CNS, long-term gene expression at significant levels has been elusive.…”

“…Recombinant HSV vectors (capacity, ,50 kb) retain the ability to enter an episomal state of latency in nondividing host cells, which is benign to the cell as essentially no viral genes are expressed with the exception of latency-associated transcripts (LATs) (Fink et al, 1996). LAT promoter elements can maintain expression of transgenes for extended periods of time in vivo (Lokensgard et al, 1994;Coffin et al, 1998;Zhu et al, 2000). Multiple mutations in the viral genome can reduce toxicity to virtually null levels and prevent reactivation from latency.…”

“…Reactivation of promoters has been demonstrated in the brain (Starr et al, 1996). Reports suggest that the strength of the promoter and its ability to avoid inactivation are key to achieving prolonged transgene expression (Gerdes et al, 2000;Zhu et al, 2000). Although in many conditions even low levels of a transgene product can suffice for normal function, expression in nontargeted cells or overexpression may prove pathologic.…”

Long-term normalization in the central nervous system, ocular manifestations, and skeletal deformities by a single systemic adenovirus injection into neonatal mice with mucopolysaccharidosis VII

“…As an alternative, gene therapy for MPSVII has been explored using several viral vectors such as retroviruses, [10][11][12] herpes viruses, 13,14 adenoviruses, [15][16][17][18][19][20] lentiviruses, 21,22 and adeno-associated viruses. [23][24][25][26] Both in vivo and ex vivo gene transduction methods have been tested experimentally; however, only limited therapeutic success has been reported, when studies were carried out using adult B6/MPSVII.…”