Defective HSV-1 Vector Expressing BDNF in Auditory Ganglia Elicits Neurite Outgrowth: Model for Treatment of Neuron Loss Following Cochlear Degeneration

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Abstract
The neurotrophins are a family of growth factors that play an important role in the development and maintenance of the nervous system. Brain-derived neurotrophic factor (BDNF) is a member of the neurotrophin family that appears to participate in the maturation and function of mammalian auditory neurons. Forms of deafness due to varied injurious stimuli that are amenable to treatment with implantable prosthetic devices require the survival of these BDNF-responsive auditory neurons for effective outcome. To evaluate the feasibility of developing a gene therapy for deafness that may be used in conjunction with a prosthetic device, we constructed replication-defective herpes simplex virus (HSV) amplicon vectors that carry the human BDNF cDNA. Using these vectors, HSVbdnf and HSVbdnflac (expresses BDNF and Escherichia coli β-galactosidase), we evaluated the expression and biological activity in established cell lines and explant cultures prepared from spiral ganglia of the murine ear. Gene transfer with HSVbdnf resulted in the efficient expression of human BDNF mRNA in murine fibroblasts. Using two BDNF-responsive cell lines, PC12trkB and MG87trkB, we demonstrate efficient secretion of biologically active BDNF. Finally, transduction of explanted spiral ganglia with HSVbdnflac elicited robust neuritic process outgrowth comparable to exogenously added BDNF. Overall, these data demonstrate that HSV vectors can efficiently transfer and express the BDNF gene in many cell types, including auditory neurons. Moreover, they suggest that similar vectors may be used to express the neurotrophin in auditory neurons in vivo and perhaps as adjunctive gene therapy for deafness. Effective therapy for some forms of deafness will likely require methods to promote survival of auditory neurons that can then be stimulated by an implantable prosthetic device. Auditory neurons require brain-derived neurotrophic factor (BDNF) for maximal survival during development and appear to continue to utilize the factor to maintain function in the mature ear. To develop a gene transfer method to deliver human BDNF, we constructed two replication-defective HSV viral vectors, HSVbdnf and HSVbdnflac. Gene transfer studies with these vectors performed in cell culture demonstrated efficient expression of the BDNF mRNA and biologically active factor. Direct gene transfer into spiral ganglion explants from the ear resulted in pronounced biological activity manifest by elaboration of extensive neural processes from the BDNF-responsive auditory neurons. These data demonstrate that the HSV vectors are efficient vehicles for the expression of the BDNF gene in auditory neurons affected in some forms of deafness.