Efficient and safe lung gene delivery using AAV6.2FF in neonatal pigs demonstrates pediatric translational potential

While adeno-associated virus (AAV) vectors have demonstrated efficacy in multiple organ systems, gene delivery to the lung remains an unmet therapeutic target. To treat a monogenic lung disease like surfactant protein B deficiency, a gene delivery system must overcome pulmonary barriers and be validated in models that closely resemble the human respiratory tract. Here, we optimized the endotracheal delivery of AAV6.2FF, an engineered capsid with enhanced pulmonary tropism, in neonatal piglets with lung features comparable to those of human newborns. We optimized delivery parameters that improved pulmonary distribution and applied them to administer AAV6.2FF expressing secreted alkaline phosphatase (SEAP) at two different doses: 5 × 10¹² vg/kg and 1.73 × 10¹³ vg/kg to assess transgene expression, biodistribution, and the safety and tolerability of this AAV platform. Transgene expression was dose-dependent, with the higher dose producing more robust and widespread SEAP expression across the cranial, middle, accessory, and upper caudal lobe regions. Both doses were well tolerated, with low anti-AAV6.2FF IgG titers and no hematology and biochemistry markers showing trends indicative of systemic toxicity. This study establishes the neonatal pig as a clinically relevant model for optimizing lung-directed AAV dosing and delivery, while also highlighting the need for further optimization to maximize this platform's therapeutic potential.