 |
|
Optimizing Cardiovascular Gene Therapy
Increased Vascular Gene Transfer With Modified Adenoviral Vectors
Melina R. Kibbe, MD;
Alan Murdock, MD;
Thomas Wickham, PhD;
Alena Lizonova, PhD;
Imre Kovesdi, PhD;
Suhua Nie, MD;
Larry Shears, MD;
Timothy R. Billiar, MD;
Edith Tzeng, MD
Arch Surg. 2000;135:191-197.
Background Adenovirus is widely used as a vector for gene transfer to the vasculature. However, the efficiency of these vectors can be limited by ineffective viral-target cell interactions. Viral attachment, which largely determines adenoviral tropism, is mediated through binding of the adenoviral fiber coat protein to the Coxsackievirus and adenovirus receptor, while internalization follows binding of the adenoviral RGD motif to  v-integrin receptors. Modifications of the fiber coat protein sequence have been successful for targeting the adenovirus to more prevalent receptors in the vasculature, including heparan sulfate–containing receptors and v-integrin receptors.
Hypothesis Modified adenoviral vectors targeted to receptors more prevalent in the vasculature result in an increased transfer efficiency of the virus in vitro and in vivo even in the presence of clinically relevant doses of heparin.
Design We tested 2 modified E1- and E3-deleted Ad5 type adenoviral vectors containing the  -galactosidase gene. AdZ.F(pK7) contains multiple positively charged lysines in the fiber coat protein that target the adenovirus to heparan sulfate receptors, while AdZ.F(RGD) contains an RGD integrin-binding sequence in the fiber coat protein that allows binding to v-integrin receptors. The gene transfer efficiency of these modified viruses was compared in rat aortic smooth muscle cells in vitro and in an in vivo porcine model of balloon-induced arterial injury. Because of the use of heparin during most vascular surgical procedures and the concern that heparin might interfere with the binding of AdZ.F(pK7) to heparan sulfate receptors, the effect of heparin on the in vitro and in vivo transfer efficiency of these 2 modified adenoviruses was evaluated.
Results In vitro infection of rat aortic smooth muscle cells with AdZ.F(pK7) and AdZ.F(RGD) resulted in significantly higher levels of -galactosidase expression compared with the unmodified adenovirus (mean ± SEM, 1766.3 ± 89.1 and 44.8 ± 3.4 vs 10.1 ± 0.7 mU per milligram of protein; P<.001). Following heparin administration, the gene transfer efficiency achieved with AdZ.F(pK7) diminished slightly in a concentration-dependent manner. However, the transfer efficiency was still greater than with the unmodified virus (mean ± SEM, 1342.3 ± 101.8 vs 4.8 ± 0.4 mU per milligram of protein; P<.001). In vivo, following injury to the pig iliac artery with a 4F Fogarty balloon catheter, we found that AdZ.F(pK7) transduced the artery approximately 35-fold more efficiently than AdZ.F and 3-fold more efficiently than AdZ.F(RGD) following the administration of intravenous heparin, 100 U/kg body weight, and heparinized saline irrigation.
Conclusions Modifications of the adenovirus that lead to receptor targeting resulted in significantly improved gene transfer efficiencies. These improvements in transfer efficiencies observed with the modified vectors decreased slightly in the presence of heparin. However, AdZ.F(pK7) was still superior to AdZ.F(RGD) and AdZ.F despite heparin administration. These data demonstrate that modifications of adenoviral vectors that enhance binding to heparan sulfate receptors significantly improve gene transfer efficiency even in the presence of heparin and suggest an approach to optimize gene transfer into blood vessels.
From the Department of Surgery, University of Pittsburgh, Pittsburgh, Pa (Drs Kibbe, Murdock, Nie, Shears, Billiar, and Tzeng); and GenVec Incorporation, Rockville, Md (Drs Wickham, Lizonova, and Kovesdi).
THIS ARTICLE HAS BEEN CITED BY OTHER ARTICLES
Vascular Gene Therapy: A Reality of the 21st Century
Zuckerbraun and Tzeng
Arch Surg 2002;137:854-861.
ABSTRACT
| FULL TEXT
Gene Therapy of Erectile Dysfunction in the Rat with Penile Neuronal Nitric Oxide Synthase
Magee et al.
Biol. Reprod. 2002;67:20-28.
ABSTRACT
| FULL TEXT
|
