Scott C. Berta

The complete sequence of adeno-associated virus type 1 (AAV-1) was defined. Its genome of 4,718 nucleotides demonstrates high homology with those of other AAV serotypes, including AAV-6, which appears to have arisen from homologous recombination between AAV-1 and AAV-2. Analysis of sera from nonhuman and human primates for neutralizing antibodies (NAB) against AAV-1 and AAV-2 revealed the following. (i) NAB to AAV-1 are more common than NAB to AAV-2 in nonhuman primates, while the reverse is true in humans; and (ii) sera from 36% of nonhuman primates neutralized AAV-1 but not AAV-2, while sera from 8% of humans neutralized AAV-2 but not AAV-1. An infectious clone of AAV-1 was isolated from a replicated monomer form, and vectors were created with AAV-2 inverted terminal repeats and AAV-1 Rep and Cap functions. Both AAV-1- and AAV-2-based vectors transduced murine liver and muscle in vivo; AAV-1 was more efficient for muscle, while AAV-2 transduced liver more efficiently. Strong NAB responses were detected for each vector administered to murine skeletal muscle; these responses prevented readministration of the same serotype but did not substantially cross-neutralize the other serotype. Similar results were observed in the context of liver-directed gene transfer, except for a significant, but incomplete, neutralization of AAV-1 from a previous treatment with AAV-2. Vectors based on AAV-1 may be preferred in some applications of human gene therapy.

Although autogenous iliac bone is frequently used for bone graft, many well-documented complications are associated with this procedure-including chronic pain; nerve, arterial, and ureteral injury; herniation of abdominal contents; sacroiliac joint instability; pelvic fractures; hematoma; and infection. An understanding of the morbidities associated with bone graft harvesting and of the strategies for avoiding them is imperative for surgeons using this grafting source. In addition, although synthetic grafting materials are considered relatively expensive compared with autogenous sources, the majority of physicians are unaware of the actual direct and indirect costs associated with autogenous bone graft harvesting. Contemporary allograft and synthetic grafting composites are being developed to optimize and surpass the native qualities of autogenous sources (ie, osteogenesis, osteoinductivity, osteoconductivity). Careful comparison of the cost of these alternative sources with the physical and monetary costs of autogenous bone graft will undoubtedly make allograft, recombinant, synthetic graft composites the logical choice in the very near future.

Factors relevant to the successful application of adeno-associated virus (AAV) vectors for liver-directed gene therapy were evaluated. Vectors with different promoters driving expression of human alpha-1-antitrypsin (alpha-1AT) were injected into the portal circulation of immunodeficient mice. alpha-1AT expression was stable but dependent on the promoter. Southern analysis of liver DNA revealed approximately 0.1 to 2.0 provirus copies/diploid genome in presumed head-to-tail concatamers. In situ hybridization and immunohistochemical analysis revealed expression in approximately 5% of hepatocytes clustered in the pericentral region. These results support the use of AAV as a vector for diseases treatable by targeting of hepatocytes.