Intramyocardial Gene Therapy with Naked DNA Encoding Vascular Endothelial Growth Factor Improves Collateral Flow to Ischemic Myocardium

Abstract

Both VEGF protein and VEGF DNA in combination with an adenoviral vector have been shown to enhance collateral formation in a porcine model of chronic myocardial ischemia. We sought to determine whether direct intramyocardial injection of naked DNA encoding for VEGF could similarly improve myocardial perfusion. Initially, 23 nonischemic pigs received either 200 microg of plasmid DNA encoding beta-galactosidase (pCMVbeta, n = 11) or 500 microg of phVEGF165 (n = 12) into four separate sites in the myocardium via a small anterolateral thoracotomy incision in the fourth intercostal space. Two additional groups of pigs received an intramyocardial injection of either phVEGF165 (n = 6) or pCMVbeta (n = 7) 3 to 4 weeks after implantation of an ameroid constrictor around the left circumflex coronary artery. The injections caused no change in heart rate or blood pressure, and no ventricular arrhythmias or histologic evidence of inflammation. VEGF protein was detected by Western blot in VEGF-treated animals, with the strongest bands closest to the injection site. Plasma VEGF concentration (ELISA) increased from 3+/-2 to 27+/-13 pg/ml (p = 0.035) by day 4 after treatment. No increase in VEGF protein was noted in pCMVbeta-treated animals whereas these did stain positive for beta-Gal. Resting myocardial blood flow (colored microspheres) was significantly reduced in the ischemic versus nonischemic territory in control animals (1.07+/-0.05 versus 1.32+/-0.05; p < 0.05) but not VEGF-treated pigs (1.32+/-0.24 versus 1.13+/-0.12; p = NS). Maximal vasodilatation with adenosine significantly increased flow to the ischemic region in VEGF-treated pigs (2.16+/-0.57 versus 1.32+/-0.24; p < 0.05) but not controls (1.31+/-0.05 versus 1.17+/-0.06;p = NS). Collateral filling of the occluded circumflex artery improved in five of six VEGF-treated pigs (mean change in Rentrop score, +1.5). We conclude that direct intramyocardial transfection phVEGF165 is safe and capable of producing sufficient VEGF protein to enhance collateral formation and myocardial perfusion. This approach may offer an alternative therapy for patients with intractable myocardial ischemia not amenable to PTCA or CABG.

Effects of intramyocardial pVEGF165 delivery on regional myocardial blood flow: evidence for a spatial ‘delivery–efficacy’ mismatch

“…As suggested by previous studies, the latter could only be established at the 3 (rather than 6) week time point by tissue, but not serum VEGF protein levels. 5,7 Further, RT-PCR showed evidence for gene transcription in the target zone. CAT levels were measurable 3 weeks after gene delivery into the ischaemic myocardium, suggesting that this reporter may be useful to address questions such as optimal delivery devices in this model.…”

“…9 Thus, the significance of improvements in collateralization after administration of VEGF protein or VEGF gene therapy suggested by other studies is unclear. 5,6,10,11 Effects of pVEGF 165 gene transfer on regional myocardial blood flow: spatial 'delivery-efficacy' mismatch…”

“…In turn, therefore, the ischaemic target zone in our study should be the focus for the effects of the administered VEGF. However, VEGF is a secreted protein, and has been demonstrated in myocardial sections taken up to 3 cm from the site of injection 5 Capillary proliferation in healing wounds or bordering areas of myocardial infarction originate at the border zones and progress towards the centre. Therefore, one explanation for the observed delivery-efficacy mismatch is that 3 weeks following delivery, the majority of angiogenic activity is occuring at the border zone.…”

“…Previous preclinical studies using microsphere techniques may have underestimated the angiogenic efficacy when tissue samples were selectively taken from the treated myocardial segments. 5,6 Sensitive noninvasive imaging techniques providing regional assessment of blood flow such as positron emission tomography and MRI may overcome this problem in preclinical studies, and are already being evaluated in clinical angiogenesis trials. [14][15][16] In conclusion, plasmid VEGF 165 delivery by direct intramyocardial injection to the pig myocardium improves regional perfusion but only in areas adjacent to the site of administration into a region of ischaemia.…”

Cardiac gene therapy in large animals: bridge from bench to bedside

“…VEGF 165 has been the most frequently tested growth factor in large animal gene transfer studies. 14,[42][43][44][45][46][47][48][49][50][51][52][53][54] This form enhances collateral formation and myocardial perfusion in ameroid constrictor chronic ischemia models 46,48,50,54 as well as in coronary artery ligation ischemia models. 14,51,52 VEGF 121 gene transfer was evaluated in the chronic ischemia model, 20,55 and the pacing induced heart failure model, 26 showing improved myocardial perfusion and preserved cardiac function.…”