Gregory D. Chapman

Direct in vivo gene transfer into the coronary and peripheral vasculatures of the intact dog.

Gene therapy approaches have been suggested for the treatment of cardiovascular disease. Recently, direct transfer of the gene encoding beta-galactosidase into peripheral arteries of the pig has been demonstrated. To determine whether this approach is applicable to other arterial beds and to other species, we first evaluated the use of beta-galactosidase as a marker protein in the canine model. We demonstrate that variable but substantial endogenous beta-galactosidase-like activity is induced by manipulation of canine peripheral arteries, which precludes the use of this marker protein in evaluating the efficiency of gene transfer in this model. A marker gene encoding firefly luciferase was then evaluated, and background luciferase activity was found to be low in the dog even after arterial manipulation. Using the luciferase gene, we then demonstrated lipid-mediated gene transfer directly into both coronary and peripheral arteries of the intact dog. These results indicate the feasibility of in vivo gene transfer into coronary arteries and demonstrate the use of the luciferase marker protein in quantifying recombinant protein expression following gene transfer in canine models. This simple and effective method for direct in vivo gene transfer into coronary and peripheral arteries may be applicable to the localized production of therapeutically important proteins for the treatment of cardiovascular diseases.

Gene transfer into coronary arteries of intact animals with a percutaneous balloon catheter.

Genetic manipulation of the vasculature may offer insights into the pathogenesis of coronary artery disease and may lead to gene therapy for disorders such as restenosis after percutaneous coronary angioplasty. The goal of this study was to develop a percutaneous method for gene transfer into coronary arteries of intact animals. Liposomes were used to facilitate transfection in coronary arteries with a plasmid containing the cDNA encoding luciferase. This reporter was chosen since it is not expressed in mammalian cells, and it can be quantified using a sensitive assay (light production). Mongrel dogs were catheterized, and DNA was delivered to coronary arteries via a porous perfusion balloon system. Luciferase expression was measured 3-5 days after the procedure, when the dogs were killed. Luciferase activity in control arteries (n = 12) was no higher than average background activity. Eight of 12 transfected arteries exhibited gene expression, averaging 4.3 +/- 2.1 pg luciferase (p less than 0.01, transfected versus control arteries). In addition, the ability to transfect DNA into femoral arteries without a transfection vehicle was tested. Five dogs were subjected to surgical transfection attempts in their femoral arteries with either DNA alone or DNA plus liposomes. Luciferase was expressed in all 10 femoral arteries; those treated with DNA alone expressed 35.6 +/- 8 pg luciferase, and those treated with DNA plus liposomes expressed 42.3 +/- 14 pg luciferase (p = 0.70). These results demonstrate the use of a percutaneous catheter to achieve gene transfer and expression in coronary arteries of intact dogs and suggest that the efficiency of intra-arterial gene transfer may be similar whether or not a transfection vehicle is used.

Minimizing the risk of inappropriately administering thrombolytic therapy (Thrombolysis and Angioplasty in Myocardial Infarction [TAMI] study group)☆

Despite the proven benefits of thrombolytic therapy in acute myocardial infarction, concern for its complications, especially in patients misdiagnosed with myocardial infarction, has led to hesitancy in its use. Historical, clinical and electrocardiographic criteria were developed for enrolling patients with suspected acute myocardial infarction into thrombolytic trials by noncardiovascular specialists. The incidence of misdiagnosis of myocardial infarction and the clinical outcomes when these criteria were used were evaluated for 1,387 consecutive patients given thrombolytic therapy. Twenty-five community hospitals and 7 interventional centers were the sites of enrollment. Most patients (63%) were enrolled from community hospitals. Criteria for thrombolytic therapy included: symptoms of acute myocardial infarction < 6 hours but > 20 minutes, and not relieved by nitroglycerin; and ST-segment elevation > or = 1 mm in 2 contiguous leads or ST-segment depression of posterior myocardial infarction. Exclusion criteria reflecting increased risk of bleeding were used. A final diagnosis of myocardial infarction was based on creatinine kinase-MB, electrocardiographic and ventriculographic evaluation. Acute myocardial infarction was misdiagnosed in 20 patients (1.4%; 95% confidence interval 0.8-2.0%). These patients were demographically similar to those with acute myocardial infarction. All misdiagnosed patients survived; no significant adverse events occurred. Thus, in several clinical settings, a simple algorithm with specific criteria was used for diagnosing acute myocardial infarction and administering thrombolytic therapy. The inclusion criteria used in this study led to a low rate of misdiagnosis.

Early and late outcome following deployment of a new flexible tantalum intracoronary stent in dogs

A new radiopaque, highly flexible balloon-expandable tantalum stent was tested. Thirty-six of 40 stents were successfully deployed percutaneously in the coronary arteries of 31 dogs. The dogs were given aspirin before, intravenous heparin during, and aspirin alone after the procedure. One dog died at 24 hours because of coronary occlusion following traumatic implantation. Four dogs were put to death early, revealing re-endothelialization by 9 days. Eleven dogs were put to death from 2 weeks to 9 months during long-term follow-up, showing all vessels widely patent with the stent uniformly embedded within a stable neointimal layer. Follow-up arteriography showed patency in all remaining stents up to 1 year, with no perforation or aneurysm formation. Four stents were placed into canine peripheral arteries and were removed percutaneously after deployment. Pathology revealed no significant trauma to involved vessels. This tantalum stent exhibits feasibility of percutaneous deployment, early neointimal formation, low thrombogenicity on long-term aspirin therapy alone, and patency up to 1 year in this canine model.