Nicholas Kipshidze

Intramural coronary delivery of advanced antisense oligonucleotides reduces neointimal formation in the porcine stent restenosis model

OBJECTIVES We evaluated the long-term influence of intramural delivery of advanced c-myc neutrally charged antisense oligonucleotides (Resten-NG) on neointimal hyperplasia after stenting in a pig model. BACKGROUND Neointimal hyperplasia after percutaneous coronary interventions is one of the key components of the restenotic process. The c-myc is a critical cell division cycle protein involved in the formation of neointima. METHODS In short-term experiments, different doses (from 500 microg to 5 mg) of Resten-NG or saline were delivered to the stent implantation site with an infiltrator delivery system (Interventional Technologies, San Diego, California). Animals were euthanized at 2, 6 and 18 h after interventions, and excised vessels were analyzed for c-myc expression by Western blot. In long-term experiments, either saline or a dose of 1, 5 or 10 mg of Resten-NG was delivered in the same fashion, and animals were euthanized at 28 days after the intervention. RESULTS Western blot analysis demonstrated inhibition of c-myc expression and was dose dependent. Morphometry showed that the intimal area was 3.88 +/- 1.04 mm(2) in the control. There was statistically significant reduction of intimal areas in the 5 and 10 mg groups (2.01 +/- 0.66 and 1.95 +/- 0.91, respectively, p < 0.001) but no significant reduction in the 1 mg group (2.81 +/- 0.56, p > 0.5) in comparison with control. CONCLUSIONS This study demonstrated that intramural delivery of advanced c-myc neutrally charged antisense morpholino compound completely inhibits c-myc expression and dramatically reduces neointimal formation in a dose dependent fashion in a porcine coronary stent restenosis model, while allowing for complete vascular healing.

Estrogen-eluting, phosphorylcholine-coated stent implantation is associated with reduced neointimal formation but no delay in vascular repair in a porcine coronary model

Estrogen can inhibit intimal proliferation and accelerate endothelial regeneration after angioplasty. This suggests that estrogen may prevent in‐stent restenosis. Unlike other therapies to prevent restenosis, estrogen may also not delay endothelial regrowth, thereby avoiding the risk of late stent thrombosis. The purpose of this work was to determine the effect of a 17β‐estradiol–eluting stent on neointimal formation in a porcine model. Each artery of six pigs was randomized to either a control, low‐dose, or high‐dose 17β‐estradiol–eluting stent. All animals were sacrificed at 30 days for histopathological analysis. There was a 40% reduction in intimal area in the high‐dose stents compared with control stents (2.54 ± 1.0 vs. 4.13 ± 1.1 mm2, for high dose vs. control, respectively; P < 0.05). There was complete endothelial regeneration at 30 days and similar inflammatory response to stenting on histopathology in all the stent groups. This is the first study to show that 17β‐estradiol–eluting stents are associated with reduced neointimal formation without affecting endothelial regeneration in the pig model of in‐stent restenosis. Estrogen‐coated stents may have a potential benefit in the prevention and treatment of in‐stent restenosis. Cathet Cardiovasc Intervent 2002;57:266–271.

Transplantation of autologous endothelial cells induces angiogenesis.

CHEKANOV, V., et al.: Transplantation of Autologous Endothelial Cells Induces Angiogenesis. This study examined the feasibility and efficacy of autologous endothelial cell (EC) transplantation using a fibrin matrix in the ischemic myocardium of sheep. Four weeks after placing an ameroid constrictor in the circumflex artery of 12 adult sheep, four animals (EC group) were subjected to EC transplantation. In four others (saline [SAL] group) saline with added inactivated cells was injected and four animals served as controls. Eight weeks after treatment the animals were sacrificed to assess histology and ultrastructure. Eight weeks after injection, ventricular function was markedly improved in the EC transplant group, but had deteriorated in the SAL and control groups. Myocardial blood flow was also increased in the EC group. Histology and electron microscopy revealed extensive neovascularization after EC transplantation and improved myocardial appearance. Heterotopic transplantation of EC within a fibrin matrix enhances neovascularization, increases myocardial blood flow, and improves left ventricular function. (PACE 2003; 26[Pt. II]:496–499)

Safety and efficacy of local periadventitial delivery of sirolimus for improving hemodialysis graft patency: first human experience with a sirolimus-eluting collagen membrane (Coll-R)

BACKGROUND Neointimal hyperplasia causes a high rate of hemodialysis synthetic graft failure. Thus, therapies that inhibit neointimal hyperplasia are urgently needed. The Coll-R is a sirolimus-eluting collagen matrix designed for intra-operative perivascular implantation around the graft-venous anastomosis. Sirolimus is an anti-proliferative drug that has proven clinical utility in suppressing neointimal tissue growth in coronary artery disease when delivered locally to the vascular wall by an endovascular drug eluting stent. METHODS A cohort of 12 chronic hemodialysis patients underwent surgical placement of 13 polytetrafluoroethylene grafts + Coll-R and were followed for up to 24 months. The primary endpoint was safety (freedom from device related adverse events). Secondary endpoints were pharmacokinetics of sirolimus release, success of Coll-R implantation and primary unassisted graft patency. RESULTS There were no technical failures, infections, vascular anastomotic or wound-healing problems. Whole blood sirolimus levels rose to a mean peak of 4.8 ng/mL at 6 h and fell to <1 ng/mL at 1 week (n = 5). Twelve and 24-month primary unassisted patencies were 76 and 38%, respectively, and the thrombosis rate was 0.37/patient-year. CONCLUSIONS Perivascular implantation of the Coll-R during graft surgery safely delivered sirolimus to the vascular wall. Systemic sirolimus levels were sub-therapeutic for immunosuppression. This small first-in-human study supports the concept that the Coll-R can safely deliver sirolimus to the graft-venous anastomosis. Safety and patency in this small study were sufficiently encouraging to justify randomized controlled trials to further test the efficacy of the Coll-R.

Advanced c-myc antisense (AVI-4126)-eluting phosphorylcholine-coated stent implantation is associated with complete vascular healing and reduced neointimal formation in the porcine coronary restenosis model

An advanced six‐ring morpholino backbone c‐myc antisense (AVI‐4126) was shown to inhibit c‐myc expression and intimal hyperplasia after local catheter delivery in a porcine balloon injury model. The purpose of this study was to investigate the effects of an AVI‐4126‐eluting phosphorylcholine‐coated (PC) stent on c‐myc expression restenosis and vascular healing after stent implantation in porcine coronary arteries. PC stents were loaded with AVI‐4126 using soak trap. Nine pigs underwent AVI‐4126 PC coronary stent implantation (two stents/animal). Two to six hours postprocedure, three pigs were sacrificed and stented segments were analyzed by Western blot for c‐myc expression. In chronic experiments, six pigs (12 stent sites) were sacrificed at 28 days following intervention and vessels were perfusion‐fixed. High‐performance liquid chromatography analysis of plasma samples showed minimal presence of the antisense. Western blot analysis of the stented vessels demonstrated inhibition of c‐myc expression at 2 and 6 hr after procedure. Quantitative histologic morphometry showed that the neointimal area was significantly reduced (by 40%) in the antisense‐coated group compared with control (2.3 ± 0.7 vs. 3.9 ± 0.8 mm2, respectively; P = 0.0077). Immunostaining and electron microscopy demonstrated complete endothelialization, without fibrin deposition, thrombosis, or necrosis in all implanted stents. In the porcine coronary model, an advanced c‐myc‐eluting PC stent blocked c‐myc expression and significantly inhibited myointimal hyperplasia and allowed complete reendothelialization and healing response. Catheter Cardiovasc Interv 2004;61:518–527.

Augmentation of the expression of proangiogenic genes in cardiomyocytes with low dose laser irradiation In vitro

BACKGROUND AND OBJECTIVE Several reports suggest that low power red laser light (LPRLL) is capable of affecting cellular processes in the absence of significant thermal effect. The objective of the present study was to determine the effect of LPRLL on proliferation of fetal cardiomyocytes in vitro and on the expression of proangiogenic genes, transforming growth factor-beta (TGF-beta), and vascular endothelial growth factor (VEGF). STUDY DESIGN/MATERIALS AND METHODS All cell cultures were irradiated with single-dose LPRLL using a He-Ne continuous wave laser (632 nm) with different doses. The effect of LPRLL on new DNA synthesis was studied by 3H thymidine-incorporation assay. VEGF and TGF-beta expression by cardiomyocytes was studied by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS We observed that a dose-dependent increase in cardiomyocytes proliferation can be obtained with LPRLL and that there is a significant increase in VEGF and TGF-beta mRNA expression by cardiomyocytes. CONCLUSIONS These data may have significant importance leading to the establishment of new methods for myocardial photoangiogenesis and photoregeneration as well as in vitro proliferation of cardiac myocytes.

Endoluminal reconstruction of the arterial wall with endothelial cell/glue matrix reduces restenosis in an atherosclerotic rabbit.

OBJECTIVES The objectives of this study were 1) to improve the attachment of reimplanted endothelial cells (EC) using a fibrin glue, and 2) to assess the impact of endothelial reseeding on restenosis eight weeks after balloon angioplasty. BACKGROUND A possible mechanism contributing to restenosis after balloon angioplasty is the loss of the EC lining. Previous attempts to reseed EC had little effect due to rapid loss of the seeded cells. METHODS Twelve atherosclerotic rabbits were subjected to angioplasty of iliac arteries and reseeding procedure. One iliac artery was subjected to EC/glue reconstruction and a contralateral site to EC seeding without glue. The animals were sacrificed after 4 h. In another series 12 rabbits were treated in the same fashion and were restudied at eight weeks. Additionally, in 10 animals one iliac was subjected to glue treatment, and another served as control. RESULTS Histological examination demonstrated the ability of this method to reattach the EC/glue matrix circumferentially to 68.0 +/- 6.7% of the arterial wall in comparison with 13.5 +/- 3.9% reattachment after EC seeding. Morphometry at eight weeks showed that the lumen area was significantly greater in the EC/glue group (1.23 +/- 0.35 mm2) than in the EC seeding alone (0.65 +/- 0.02 mm2) and 0.72 +/- 0.41 mm2 in the glue group. This was principally accounted for by the statistically significant differences in the intimal area (0.76 +/- 0.18 mm vs. 1.25 +/-0.26 mm2 and 1.01 +/- 0.53 mm2, respectively). CONCLUSIONS The attachment of EC after angioplasty can be greatly improved with fibrin glue matrix. The near 70% endothelial coverage achieved by this method resulted in a significant reduction of restenosis in atherosclerotic rabbit.

Update on drug-eluting coronary stents

The introduction of drug-eluting stents (DES) to interventional cardiology practice has resulted in a significant improvement in the long-term efficacy of percutaneous coronary interventions. DES successfully combine mechanical benefits of bare-metal stents and stabilizing the lumen, with direct delivery and the controlled elution of a pharmacologic agent to the injured vessel wall to suppress further neointimal proliferation. The dramatic reduction in restenosis has resulted in the implementation of DES in clinical practice, and has rapidly expanded the whole spectrum of successfully treatable coronary conditions, particularly in high-risk patients and complex lesions. In this review the authors present current data on DES. Currently, two types of DES are available in the USA: sirolimus-eluting stents (SES) CYPHER™ (Cordis Corp., FL, USA) and paclitaxel-eluting stents (PES) TAXUS™(Boston Scientific, MA, USA), and many more are on the way to approval. In addition to sirolimus and paclitaxel, several other drugs have been successfully used in DES. Everolimus and ABT-578 are both analogs of sirolimus that also have immunosuppressive and antiproliferative properties. Another approach in the development of DES is to use drugs that can accelerate re-endothelialization and restore normal endothelial function following vascular injury. Recent advances in vascular gene transfer have also demonstrated potential new treatment modalities for cardiovascular disease, particularly in the treatment of vascular restenosis.

Perspectives of drug-eluting stents: the next revolution.

Coronary stent implantation has become a well established therapy in the management of coronary artery disease (CAD). Although the Stent Restenosis Study (STRESS) and Belgium-Netherlands Stent (BENESTENT) trials demonstrated convincingly that stenting is superior to percutaneous transluminal coronary angioplasty with respect to restenosis in de novo lesions, there is, however, still a high incidence (10 to 50%) of restenosis following stent implantation.Improvements in stent design and implantation techniques resulted in an increase in the use of coronary stents and today, in most centers in the US and Europe, stenting has become the predominant form of nonsurgical revascularization accounting for about 80% of all percutaneous coronary intervention procedures. Coronary stents provide luminal scaffolding that virtually eliminates elastic recoil and remodelling. Stents, however, do not decrease neointimal hyperplasia and in fact lead to an increase in the proliferative comportment of restenosis.Agents that inhibit cell-cycle progression indirectly have also been tested as inhibitors of vascular proliferation. When coated onto stents, sirolimus, a macrolide antibiotic with immunosuppressive properties, and paclitaxel and dactinomycin, both chemotherapeutic agents, induced cell-cycle arrest in smooth muscle cells (SMC) and inhibited neointimal formation in animal models.Preliminary clinical studies with drug-eluting stents produced dramatic results eliminating restenosis in large and mid-size arteries. Quantitative coronary angiography and intravascular ultrasound demonstrated virtually complete inhibition of tissue growth at 6 and 12 months after sirolimus-eluting stent implantation. Results are also very encouraging with paclitaxel-coated stents. However, it needs to be proven that current drug-eluting stents will produce similar results in ‘real life’ interventional practice (long lesions, lesions in small vessels, in vein grafts, chronic total occlusions, and bifurcated and ostial lesions). The ongoing randomized, double-blind sirolimus-coated Bx Velocity™ balloon expandable stent in the treatment of patients with de novo coronary artery lesions (SIRIUS) trial may answer some of these concerns.With further improvements, including the expansion of drug-loading capacity, double coatings and coatings with programmable pharmacokinetic capacity using advances in nanotechnology (which may allow for more precise and controlled release of less toxic and improved molecules), we think that in the next few years the practice of interventional cardiology may undergo major changes. A new era of dramatic improvements in the treatment of CAD may have dawned. The prospect of approval of this technology should herald a host of clinical trials to revisit basic assumptions about the place of coronary stenting in the contemporary care of obstructive (and nonobstructive) CAD.

Novel site-specific systemic delivery of Rapamycin with perfluorobutane gas microbubble carrier reduced neointimal formation in a porcine coronary restenosis model

Earlier studies demonstrated that perfluorobutane gas microbubble carrier (PGMC) adheres to injured arteries and enhances the drug uptake specifically into the cells of the denuded vessel segment. The purpose of this study was to investigate the effect of PGMC‐based systemic delivery of Rapamycin on expression of p27 in vascular tissue and restenosis in porcine coronary arteries after stent implantation. Eight pigs underwent coronary stent implantation (three stents per animal). Five pigs were treated with i.v. injection of PGMC with 2 mg of Rapamycin and three animals served as control. Four hours postprocedure, three pigs were sacrificed and stented segments were analyzed by high‐performance liquid chromatography (HPLC) and Western blot. In chronic experiments, five pigs (15 stent sites) were sacrificed at 28 days following intervention and vessels were perfusion‐fixed. HPLC of the treated arteries demonstrated high drug concentration in the vessel tissue, and Western blot analysis showed elevated expression of p27 at 4 hr postprocedure. Histomorphometry revealed significantly reduced (by 40%) neointimal formation in the PGMC/Rapamycin group compared with controls (1.84 ± 0.84 vs. 4.77 ± 1.71 mm2, respectively; P < 0.001). In the porcine coronary model, site‐specific systemic delivery of Rapamycin utilizing PGMC resulted in overexpression of p27 and a significant reduction of neointimal formation within the stented segments. Catheter Cardiovasc Interv 2005;64:389–394.