To generate a dsDNA break in the genomic regions flanking the PiZ mutation, 2 rAAVs (AAV2/8, containing AAV2 ITRs and AAV8 capsid) were generated to express ZFNs from a hepatocyte-specific promoter consisting of the apolipoprotein E locus control region, human AAT promoter, a portion of intron-A of human coagulation factor IX, 3′-untranslated region, and a bovine growth hormone polyadenylation signal.21 ZFNs were designed with Fok1 nuclease monomer fused at the carboxy terminals of the zinc-finger proteins. When the 2 ZFN molecules home at their respective DNA sequence recognition sites on the 2 complementary DNA strands, the Fok1 monomers dimerize, generating an active nuclease (Figure 1A). For homology-directed correction of the pathogenic mutation on exon-7 of SA1-ATZ, rAAV-TI was designed by replacing the AAV2 genome with a segment of the wild-type human SERPINA1 gene consisting of left and right homology arms, ~1 kb each, flanking the mutation site, between the AAV2 ITRs. Two silent mutations were inserted into the homologous recombination donor to prevent recleavage by ZFN.