Ultrasound-Mediated Delivery of Nonviral Col4a5 Gene Therapy Vector in X-Linked Alport Syndrome (XLAS) Disease Model Mice and Nonhuman Primates

Background: XLAS is a hereditary disorder marked by progressive kidney dysfunction, hearing loss, and ocular abnormalities. The renal pathology results from structural defects in the glomerular basement membrane caused by mutations in the COL4A5 gene. Lack of delivery technology allowing efficient delivery of a therapeutic transgene expressing full-length Col4a5 in podocytes prevents the development of genetic medicines for XLAS patients. Ultrasound-mediated gene delivery (UMGD) offers a noninvasive, non-viral and targeted approach for transgene delivery to renal cells, presenting a promising strategy for COL4A5 gene replacement therapy in XLAS Methods: UMGD parameters were optimized for efficient transgene delivery to podocytes in both mice and non-human primates (NHPs). A series of non-viral gene therapy constructs encoding codon-optimized COL4A5 sequences under the control of podocyte-specific promoters were engineered. The lead vector was delivered to the kidneys of an XLAS mouse model (G5X) and NHP using UMGD. Safety of the gene delivery approach was assessed using established methods. Results: To evaluate the safety and efficacy of UMGD for renal targeting, a reporter transgene was delivered to mice, resulting in kidney-specific and durable expression that was further enhanced with repeat dosing. Podocyte-specific construct was developed which demonstrated robust full-length COL4A5 expression in primary human podocytes. The vector was successfully delivered to the kidneys of G5X mice using UMGD, resulting in strong transgene expression within the glomeruli. Further refinement of UMGD conditions enabled efficient delivery of the COL4A5 transgene to NHP kidneys, with substantial delivery observed in podocytes. Safety and tolerability assessments confirmed a favorable safety profile. Conclusion: The efficient delivery of a nonviral COL4A5 vector to podocytes in the kidneys of both the XLAS disease model and NHP, combined with the favorable safety profile of UMGD, supports the translational potential of this approach for clinical development in the treatment of XLAS.