Abstract 715: Pseudotyped AAV Vectors in Combination with the Cardiac Troponin-T Promoter Dramatically Accelerate the Onset of Cardiac-Specific Gene Expression After Direct Injection into Adult Murine Myocardium
Introduction: AAV2 suffers from a prolonged lag phase (up to 6 wks) before full expression after injection into adult myocardium. Newly-isolated AAV serotypes 1, 6 & 8 provide early onset expression after IV injection. We hypothesized that packaging AAV2 DNA in capsids from serotypes 1, 6 or 8 would overcome the lag phase seen after injection into the LV wall.
Methods: An AAV2 ITR-based vector carrying firefly luciferase under control of the cardiac troponin-T promoter (AcTnTLuc) was packaged into capsids from AAV serotypes 1, 2, 6 & 8 (AAV2/1, 2/2, 2/6 & 2/8). 5E+ 10 viral particles were injected into the LV wall of adult mice (n = 4/group) and transgene expression was monitored by in vivo bioluminescence (IVIS, Xenogen Corp), in vitro luciferase assays and immunostains for luciferase.
Results: At Day 7 post-injection, in vivo light output from hearts injected with AAV2/2 was no different from background (p = NS, Panel A), whereas it had reached steady state levels in hearts injected with AAV2/1, 2/6 & 2/8. At Day 14, light output was 670x higher in AAV2/1, 1820x higher in AAV2/6 and 3070x higher in AAV2/8 than in AAV2/2-injected mice (p < 0.05 for all). Multi-organ in vitro luciferase assays confirmed that luciferase was largely restricted to the heart. Luciferase immunostains of AAV2/8 hearts indicated high efficiency transduction (Panel B).
Conclusions: Capsids from AAV serotypes 1, 6 or 8 are far more efficient for cardiac gene delivery than AAV2, as evidenced by early onset and high-level gene expression. The use of tissue-specific promoters in combination with highly efficient AAV capsids further enhances the potential of AAV vectors for animal experiments and human gene therapy protocols.