Topical retrovirus gene therapy for Scheie keratopathy

Purpose. Corneal epithelium is a rapidly-regenerating tissue and may be an ideal target for in vivo retrovirus gene therapy for conditions such as Scheie keratopathy, the corneal dystrophy of mucopolysaccharidosis (MPS) type I. In this disorder, progressive corneal clouding is caused by accumulation of undegraded glycosaminoglycans due to deficient α-L-iduronidase (IDUA). Methods. To study the response to IDUA gene transfer, a MoMLV-based vector L1SN was constructed and packaged in PA317 cells yielding concentrated vector (10⁷ cfu/ml) with high IDUA activity (26,500 nmol/mg/hr; 20 - 31 in nontransfected PA317 cells). New Zealand White rabbits were selected as a model for in vivo gene therapy after determining that native IDUA levels were relatively low, and that leporine IDUA coding sequence was highly homologous to human IDUA cDNA. Results. Cultured rabbit fibroblasts and conjunctival epithelium transduced with L1SN showed durable, high-level IDUA expression (80- and 60-fold above control, respectively). Transduction of peripheral blood lymphocytes from a patient with MPS I produced exceedingly high IDUA enzyme activity, increased from negligible levels (< 1 nmol/mg/hr) to > 1,400 (normal range 65 - 150). To evaluate vector transduction in vivo, L1SN was applied topically to the cornea after superficial epithelial keratectomy. This approach resulted in elevated IDUA activity for 3 weeks reflecting the survival of transiently activated cells (TAC). To further refine a means of targeting corneal epithelial stem cells for more durable expression, marker vector MFG-lacZ was applied in the same fashion. Three days after topical administration, small numbers of lacZ+ cells were identifiable. Conclusion. In vivo transduction, albeit currently limited to the survival of TAC, provides the means to map corneal epithelial stem cells and the impetus to seek a more durable treatment.