Adenoassociated Viral Vectors

Adeno-associated virus (AAV) is a defective human parvovirus that is not able to replicate unless a helper virus, such as adenovirus, is also present.19 There are several features that make AAV an attractive vector. The virus can be prepared at high titer, is not pathogenic in humans, and infects a broad range of cell lines.20 Wild-type AAV integrates in a site-specific manner in a 7-kb region of human chromosomes.20,21 The AAV genome is a single-stranded, linear, 5-kb DNA molecule.

The genome is flanked by two 145-bp inverted terminal repeats (ITRs) that contain the sequences required for packing, integration, and DNA replication. The coding region contains two open reading frames (ORFs). Either of these ORFs can be replaced with the transgene and regulatory elements to construct an AAV vector. The ORF can only accept a transgene of 4 to 5 kb, thus limiting the size of the transgene insert. Propagation of AAV vectors requires AAV Rep and Cap proteins and five adenoviral proteins: E1A, E1B, E2A, E4, and VA. These are complex packaging requirements, and thus it has been difficult to construct a packaging cell line. Instead, AAV vectors are propagated by cotransfection of the AAV vector with a plasmid wild-type or mutant helper adenovirus. AAV vectors have been used very successfully when injected into skeletal muscle to produce proteins secreted into the circulation, such as factor IX for hemophilia B.22 Cardiovascular applications include cardiac myocytes, where Svensson et al. have demonstrated stable expression.23 Whether AAV vectors can be adapted to efficiently transduce vascular endothelial cells and smooth muscle cells is not known.

Nonviral Vectors

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