Neuronal precursor-restricted transduction via in utero CNS gene delivery of a novel bipartite HSV amplicon/transposase hybrid vector.

The ability to modify genetically in utero the precursors of neuronal lineage contributing to multiple postmitotic cell types in the adult central nervous system would provide a means to evaluate strategies to ameliorate conditions affecting cellular patterning, metabolism, or survival. The herpes simplex virus (HSV)-derived amplicon, a vector devoid of viral genes and with the largest known payload capacity, normally exists episomally within nuclei of transduced cells, thus precluding conveyance during mitosis. Herein, we modify the Tc1-like Sleeping Beauty (SB) transposon system to create an integrating amplicon vector platform wherein provision of transposase in trans effectively catalyzes integration of a transgenomic segment. Cotransduction with a Rous sarcoma virus promoter-driven beta-galactosidase-neomycin (betageo) fusion flanked by SB terminal repeats (HSVT-betageo) and a second expressing the SB transposase gene under HSV immediate-early 4/5 gene promoter control (HSVsb) resulted in integration and extension of expression duration. Most notably, in utero intraventricular application led to extensive transgene expression within neuronal precursors and their derivatives without attendant adverse consequences, suggesting this new platform could be used to evaluate prenatally the function of gene products in neuronal lineages and evaluate therapeutic strategies for correction of genetic abnormalities affecting the developing CNS.