Efficient Transfer of Genes into Murine Cardiac Grafts by Starburst Polyamidoamine Dendrimers

Abstract

Starburst dendrimer, a structurally defined, spherical macromolecule composed of repeating polyamidoamino subunits, was investigated to augment plasmid-mediated gene transfer efficiency in a murine cardiac transplantation model. The grafts were directly injected with naked pCH110, a plasmid encoding β-galactosidase (β-Gal), or pCH110-dendrimer complex, and reporter gene expression determined by X-Gal staining. The grafts injected with pCH110-dendrimer demonstrated widespread and extended β-Gal expression in both myocytes and the graft infiltrating cells from 7 to 28 days, compared to the grafts injected with naked pCH110 that expressed β-Gal only in myocytes for less than 14 days. pαMHC-vIL-10, as plasmid encoding viral interleukin-10 (vIL-10) under the control of α-myosin heavy chain promoter, was able to prolong allograft survival from 13.9 ± 0.9 days to 21.4 ± 2.3 days (p < 0.005). When dendrimer G5EDA was used with pαMHC-vIL-10, 60-fold less DNA resulted in significant prolongation of graft survival to 38.6 ± 4.7 days (p < 0.0005). The dose of DNA, the charge ratio of DNA to dendrimer, and the size generation of the dendrimers were all determined to be critical variables for prolongation of allograft survival in this model system. Thus, the use of the Starburst dendrimer dramatically increased the efficiency of plasmid-mediated gene transfer and expression. Production of immunosuppressive cytokines at higher amounts for longer periods of time in a greater expanse of tissue enhanced the immunosuppressive effect and prolonged graft survival further. Plasmid-mediated gene therapy has been used to deliver immunosuppressive molecules into allografts to prolong graft survival. However, direct injection of naked plasmid DNA is inefficient because transgene expression is low and transient. This study investigated the ability of Starburst dendrimers to augment plasmid-mediated gene transfer efficiency in a murine cardiac transplantation model. The results demonstrate that dendrimers increased the efficiency of transfer and expression of exogenous DNA in cardiac grafts. Improved expression of an immunosuppressive cytokine viral interleukin-10 (vIL-10) by dendrimers significantly prolonged allograft survival. The dose of DNA, the charge ratio of DNA to dendrimer, and the size generation of the dendrimers were all critical for prolongation of allograft survival. Thus, the use of the Starburst dendrimer as a carrier molecule for plasmid-mediated gene transfer improved the efficiency of transfer and expression, providing further therapeutic value for treatment of cardiac allograft rejection.