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. Author manuscript; available in PMC: 2016 Aug 11.
Published in final edited form as: Cell Rep. 2015 Jul 30;12(6):1056–1068. doi: 10.1016/j.celrep.2015.07.019

Figure 4. Anc80L65 in vivo transduction biology.

Figure 4

A, top panel: Mouse liver transduction and lacZ transgene expression comparison of AAV2, AAV8 and Anc80L65.TBG.nLacZ in liver 28 days after intraperitoneal delivery at a dose of 3.9 × 1010 GC (C57Bl/6, n=3). A, middle panel: AAV2, AAV8 and Anc80L65 muscle tropism in mouse 28 days following an intramuscular delivery at a dose of 1010 GC to the rear-right thigh (gastrocnemius) (n=5). See also Figure S1. A, lower panel: Comparison of eGFP transgene expression between AAV2, AAV8, and Anc80L65 in the murine retina after subretinal delivery at a dose of 2×109 GC. AAV2 shows high affinity for RPE cells while both RPE and photoreceptors are targeted using AAV8 and Anc80L65 vectors with Anc80L65 showing higher transduction efficiency compared to the AAV2 and 8 (C57Bl/6, n=4 eyes). B. Qualitative dose response hepatic eGFP-expression analysis following dosing of 1011 (top panel), 1010 (middle panel), and 109 (bottom panel) GC comparing AAV-8 and Anc80L65 by retro-orbital intravenous injection in mouse. Both AAV8 and Anc80L65 lead to comparable eGFP expression at equal dose throughout the dose ranging. (C57Bl/6, n=3) C. Quantitative AAV dose response analysis measuring mouse serum levels of recombinant human alpha 1-antitrypsin (hA1AT) transgene expression from AAV-8 (black symbols: square-1011 GC, circle-1010 GC, and four-square-109 GC) and Anc80L65 (grey symbols: diamond-1011 GC, square-1010 GC, and triangle-109 GC). (C57Bl/6, n=5). See also Table S1, S3, S4 and S5. D. Rhesus macaque liver gene transfer of AAV-8 and Anc80L65 expressing Rhesus chorionic-gonadotropin (rhCG) following saphenous vein injection of a dose of 1 × 1012 GC/kg. Genomic DNA was harvested from macaque liver-lobes and viral genome (vg) per diploid genome (dpg) was measured by qPCR assay. One AAV8 and all three Anc80L65 animals successfully received ~1–3 vg per diploid cell of the caudal liver lobe, while 2 AAV8 animals likely had low level NAB resulting in vector neutralization and limited liver gene transfer. See also Table S2, S6 and S7. E. Transgene mRNA expression of AAV8 and Anc80L65 in NHP caudal, right, left and middle liver-lobes by TaqMan probe-specific, quantitative reverse-transcriptase PCR (qRT-PCR). Protein levels are not available due to lacking antibody and rhCG standards. Quantitation of rhCG transcript is normalized with endogenous GAPDH mRNA levels.