Engineering herpes simplex virus vectors for CNS applications

We have engineered highly defective HSV genomic vectors (i) deleted for multiple IE gene functions which fail to initiate lytic viral replication, (ii) that express few viral functions, (iii) that display reduced toxicity even for primary neurons in culture, (iv) that are able to incorporate multiple transgenes or single large genes, (v) that are able to efficiently establish latency in neurons and serve as a platform for long-term gene expression using the latency promoter system, (vi) that cannot reactivate from latency or spread to other nerves of tissues, (vii) that can enhance or regulate transgene expression, and (viii) that can be targeted to specific cell types using their normal receptor-recognition ligands modified to contain novel attachment functions. These vectors may prove useful in applications in which short-term gene expression and multiple gene products are required to achieve a therapeutic outcome such as tumor-cell killing and vaccination. Expression of these therapeutic genes may be coordinately regulated or controlled by drug-responsive transactivators that will enable regulation of the level and duration of therapeutic gene expression. Applications that require long-term expression of transgene in PNS neurons, such as diabetic neuropathy, may employ the LAP system for expression of therapeutic genes, such as NGF. Although we have been able to redirect virus binding using novel ligands introduced into the viral glycoproteins, significant improvements will be required to more efficiently bind the virus to the target cell without perturbing normal cellular pathways and still enable the virus to enter the target cell by a mechanism similar to that naturally employed. Future experiments will be designed using vectors that are capable of regulation of therapeutic gene expression using their own native promoters, thus responding to the natural stimuli involved in their production. Further work will be necessary to modify the vector for non- nervous-system approaches to treat diseases of other tissues such as arthritis, muscular dystrophy, or autoimmune disorders.