Gene Therapy for Attenuating Cardiac Allograft Arteriopathy Using Ex Vivo E2F Decoy Transfection by HVJ-AVE–Liposome Method in Mice and Nonhuman Primates

Abstract

Abstract-Cardiac allograft arteriopathy, which limits the long-term survival of recipients, is characterized by diffuse intimal thickening composed of proliferative smooth muscle cells. The transcription factor E2F plays a pivotal role in the coordinated transcription of cell-cycle regulatory genes. To test the hypothesis that double-stranded DNA with specific affinity for E2F (E2F decoy) is effective in preventing intimal hyperplasia, we performed ex vivo single intraluminal delivery of E2F decoy into cardiac allografts of mice and Japanese monkeys using the hemagglutinating virus of Japan (HVJ) artificial viral envelope-liposome method. In murine models, antisense cyclin-dependent kinase 2 (cdk2) kinase oligodeoxynucleotide (ODN) and no transfers were performed to compare the effects. Severe intimal thickening was observed, and multiple cell-cycle regulatory genes were enhanced in untreated allografts. E2F decoy prevented neointimal formation and suppressed these genes for up to 8 weeks, whereas antisense cdk2 kinase ODN had limited effects. In primate models, E2F decoy dramatically prevented neointimal thickening and suppressed multiple cell-cycle regulatory genes, whereas intimal thickening developed in the nontransfected or mismatch decoy-transfected allografts. Gel mobility shift assay proved the specific effects of E2F decoy, and reverse transcriptase-polymerase chain reaction documented that neither complication nor dissemination of HVJ into other organs was observed. We demonstrate that ex vivo gene delivery to allografts is a potent strategy to modify allograft gene expression, resulting in prevention of graft arteriopathy without systemic adverse effects. Key Words: primate Ⅲ heart transplantation Ⅲ transcription factor Ⅲ arteriosclerosis Ⅲ gene therapy C ardiac transplantation often results in accelerated graft coronary disease in long-term survivors. 1 Neointimal hyperplasia of the coronary arteries results from vascular smooth muscle cell (SMC) proliferation; this proliferation is dependent on the coordinated actions of cell-cycle regulatory genes. 2 Recently, we reported that antisense cyclin-dependent kinase (cdk) 2 kinase oligodeoxynucleotide (ODN) prevents neointimal formation in murine cardiac allografts. 3 The transcription factor E2F regulates multiple cell-cycle regulatory genes, which are critical to the process of cell growth and proliferation. 4,5 Double-stranded DNA with high affinity for E2F acting as a decoy (E2F decoy) inhibits cell-cycle regulatory gene expression and SMC proliferation in rat carotid injury models. 6 However, the effect of E2F decoy in preventing graft coronary arterial neointimal formation has not been investigated. Several gene therapy trials have been performed in search of methods to prevent and treat several diseases using the hemagglutinating virus of Japan (HVJ)-liposome method. 7 Recently, HVJ artificial viral envelope (AVE) liposome, which is a modified method of HVJliposome delivery, has been demonstrated to increase efficiency of cellular uptake of ODN. 8...

Ring-Sp1 decoy oligonucleotide effectively suppresses extracellular matrix gene expression and fibrosis of rat kidney induced by unilateral ureteral obstruction

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Transcription Factor Decoy Molecules Based on a Peptide Nucleic Acid (PNA)-DNA Chimera Mimicking Sp1 Binding Sites

“…In vitro transfection of cis elements that decoy against nuclear factors leads to alteration of gene expression and was recently proposed in molecular medicine as a novel tool for the possible therapy of several disorders (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12). One of the most effective decoy approaches so far described involves nuclear proteins belonging to the NF-B 1 superfamily (7)(8)(9)(13)(14)(15)(16)(17)(18)(19)(20).…”

Characteristics of Chronic Rejection in Heart Transplantation: Important Elements of Pathogenesis and Future Treatments

“…The transcription factor E2F regulates multiple cell-cycle regulatory genes, which are critical to the process of cell growth and proliferation. 70 To clarify the effects of E2F decoy ODN on CAV, we used both murine and monkey cardiac transplant models, which showed that E2F decoy ODN transfer into the allografts specifically abolishes E2F activity and inhibites CAV 71 (Figure 3). Another critical transcription factor, nuclear factor-κB (NF-κB), plays a pivotal role in the coordinated transcription of multiple inflammatory genes.…”