Author + information
- Received December 9, 2004
- Revision received January 22, 2005
- Accepted January 25, 2005
- Published online May 17, 2005.
- Jiro Aoki, MD⁎,
- Patrick W. Serruys, MD, PhD, FACC⁎,⁎ (, )
- Heleen van Beusekom, MD, PhD⁎,
- Andrew T.L. Ong, MBBS, FRACP⁎,
- Eugene P. McFadden, MBChB, MD, FRCPI, FACC⁎,
- Georgios Sianos, MD, PhD⁎,
- Willem J. van der Giessen, MD, PhD⁎,
- Evelyn Regar, MD, PhD⁎,
- Pim J. de Feyter, MD, PhD, FACC⁎,
- H. Richard Davis, MSc†,
- Stephen Rowland, PhD† and
- Michael J.B. Kutryk, MD, PhD‡
- ↵⁎Reprint requests and correspondence:
Dr. Patrick W. Serruys, Thoraxcenter, Bd 406, Erasmus Medical Center, Dr. Molewaterplein 40, 3015 GD Rotterdam, the Netherlands
Objectives This study was designed to evaluate whether rapid endothelialization of stainless steel stents with a functional endothelium prevents stent thrombosis and reduces the restenotic process.
Background A “pro-healing” approach for prevention of post-stenting restenosis is theoretically favored over the use of cytotoxic or cytostatic local pharmacologic therapies. It is believed that the central role of the vascular endothelium is to maintain quiescence of the underlying media and adventitia.
Methods Sixteen patients with de novo coronary artery disease were successfully treated with implantation of endothelial progenitor cell (EPC) capture stents.
Results Complete procedural and angiographic success was achieved in all 16 patients. The nine-month composite major adverse cardiac and cerebrovascular events (MACCE) rate was 6.3% as a result of a symptom-driven target vessel revascularization in a single patient. There were no other MACCE despite only one month of clopidogrel treatment. At six-month follow-up, mean angiographic late luminal loss was 0.63 ± 0.52 mm, and percent stent volume obstruction by intravascular ultrasound analysis was 27.2 ± 20.9%.
Conclusions This first human clinical investigation of this technology demonstrates that the EPC capture coronary stent is safe and feasible for the treatment of de novo coronary artery disease. Further developments in this technology are warranted to evaluate the efficacy of this device for the treatment of coronary artery disease.
The emergence of drug-eluting stents has dramatically reduced the incidence of in-stent restenosis (1,2). This therapy interferes with the natural healing response by preventing or significantly delaying the formation of a functional endothelial lining over the stent (3).
Recently, the existence of circulating endothelial progenitor cells (EPCs) has been identified as a key factor for re-endothelialization (4). The early establishment of a functional endothelial layer after vascular injury has been shown to assist in the prevention of neointimal proliferation and thrombus formation (5,6). The EPC capture stents have been developed using immobilized antibodies targeted at EPC surface antigens. The HEALING-FIM (Healthy Endothelial Accelerated Lining Inhibits Neointimal Growth-First In Man) registry is the first clinical investigation using this technology.
The HEALING-FIM registry is a single-center, prospective, non-randomized registry trial. Patients were eligible if they were between ages 18 and 85 years and had a diagnosis of stable or unstable angina or silent ischemia. Additional eligibility criteria were the presence of a single primary target lesion in a native coronary artery that was 2.5 to 3.5 mm in diameter that could be covered by a single trial stent (13 mm or 18 mm length), a stenosis of 51% to 99% of the luminal diameter as estimated visually, and a flow rate of grade 1 or higher according to the classification of the Thrombolysis In Myocardial Infarction (TIMI) trial. Patients were not eligible for enrollment if they had an evolving myocardial infarction; stenosis of the left main coronary artery; a lesion located at an ostial location; a calcified lesion that could not be completely dilated before stenting; angiographically visible thrombus within the target lesion; a left ventricular ejection fraction of <30%; or an intolerance of aspirin, clopidogrel, ticlopidine, heparin, stainless steel, or contrast material. The trial was reviewed and approved by the ethics review committee, and written informed consent was obtained from all patients.
Study device: EPC capture stent
The EPC antibody surface consists of a covalently coupled polysaccharide intermediate coating with murine monoclonal anti-human CD34 antibodies, attached to a stainless steel stent (R stent, OrbusNeich, Fort Lauderdale, Florida) (Fig. 1).This antibody specifically targets CD34+ cells (endothelial progenitor cells are CD34 positive) in the vascular circulation. This device was supplied in aqueous sodium azide solution as a preservative to maintain bioactivity and required hand crimping by the operator onto a percutaneous transluminal coronary angioplasty balloon catheter before implantation.
Lesions were treated according to local standard interventional techniques. Specifically, the decision to predilate or direct stent was at the investigator’s discretion, and post-dilation was performed as required to ensure that the residual stenosis was <20% by visual assessment, with a TIMI flow grade rate 3. In case of a dissection or incomplete coverage of the lesion, implantation of additional EPC capture R stents was permitted.
Intravenous boluses of heparin were administered to maintain an activated clotting time >300 s during the implantation. Treatment with aspirin, at a dose of at least 80 mg/day, was initiated at least 12 h before the procedure and continued for one month. In addition, a loading dose of 300 mg of clopidogrel was administered before the procedure, followed by 75 mg daily for 28 days. Glycoprotein IIb/IIIa inhibitors were used at the operator’s discretion. Angiographic success was defined as the successful implantation of the study device, with a stenosis of <20% of the vessel diameter with TIMI flow grade 3.
All patients were scheduled for a clinical follow-up at one, six, and nine months following the implantation procedure to assess the anginal status and the occurrence of major adverse cardiac and cerebrovascular events (MACCE). An electrocardiogram was obtained at each visit, and an angiographic and intravascular ultrasound (IVUS) study was performed at a mean of 185 ± 14 days.
Quantitative angiographic and IVUS analysis
Coronary angiograms were obtained in multiple views after an intracoronary injection of nitrates. Offline quantitative analyses of preprocedural, postprocedural, and six-month follow-up angiographic data were performed. Restenosis was defined as a reduction of 50% or more of the luminal diameter. Late luminal loss was defined as the difference between the minimal luminal diameter after procedure and at six months. The target lesion was defined as the stented segment plus the 5-mm segments proximal and distal to the stented segment.
Intravascular ultrasound was performed with an automated pullback at 0.5 mm/s to examine the target lesion at postprocedure and six-month follow-up. Lumen, stent, and external elastic membrane contours were detected with the use of CUARD QCU analysis software (CUARD BV, Wijk Bij Duurstede, the Netherlands), applying a three-dimensional reconstruction as described elsewhere (7).
Study end points
The primary safety end point of this study was the absence of stent thrombosis up to six months. The second end point was a composite of MACCE, defined as cardiac death, stroke, Q-wave or non-Q-wave myocardial infarction (MI), and target vessel revascularization. Stroke was defined as a focal neurologic deficit resulting from a vascular cause involving the central nervous system. Q-wave MI was defined as development of new abnormal Q waves not present on the patient’s baseline. Non-Q-wave MI was defined as a creatine kinase of more than twice the upper limit of normal with an abnormal level of the MB isoenzyme of creatine kinase. The efficacy end point was late luminal loss as determined by quantitative coronary angiography and % stent volume obstruction by IVUS at six months. Stent thrombosis was angiographically documented as a complete occlusion (TIMI flow grade 0 or 1) or a flow-limiting thrombus (TIMI flow grade 1 or 2) of a previously successfully treated artery.
Detection of human antimurine antibody (HAMA)
Human antimurine antibody testing was not added to the study until several patients had already returned for follow-up. Testing with baseline data was conducted on 4 of 16 patients. The HAMA was determined by a commercially available enzyme-linked immunosorbent assay kit (MEDAC, Hamburg, Germany) (8). A positive assay was defined as ≥10 ng/ml. Significant levels of HAMA were defined as ≥150 ng/ml.
One specimen was retrieved by directional atherectomy (Flexi-cut, Guidant Europe SA, Diegem, Belgium), fixed in 4% buffered formaldehyde with metal stent fragments removed and embedded in paraffin.
Sections were stained with hematoxylin eosin and an elastin stain (Resorcin Fuchsin) for general assessment of the tissue. Cellular characterization was performed with immunocytochemistry, using antibodies against smooth muscle (specific alpha-actin), leukocytes (CD45), macrophages (mac 387), proliferation cells (Mib 1), and EPC (CD34). All antibodies except anti-CD34 were obtained from DAKO (DakoCytomation, Produktionsvej, Denmark).
Because of the size of the patient population in this nonrandomized registry, no formal statistical analysis was conducted to determine the efficacy of the device. Continuous variables are expressed as mean ± SD. Comparisons between postprocedure and six-month follow-up values were performed with a two-tailed paired ttest. A p value <0.05 was considered statistically significant.
Baseline characteristics and procedural outcome
Overall, 16 patients were enrolled in the HEALING-FIM registry between May and November 2003. Table 1presents the baseline characteristics and procedural outcomes for this patient population. A second overlapping stent was implanted in two patients to treat a dissection following implantation of the first stent.
Table 2provides an overview of the MACCE at one and nine months. There was no subacute thrombosis or MACCE in the first month after implantation. During the first nine months, MACCE occurred in a patient with insulin-dependent diabetes with diabetic nephropathy. This patient (#12) was admitted with a non-ST-segment elevation MI (maximum creatinine kinase level was 524 U/l) at six months after the index procedure and underwent recatheterization that demonstrated focal in-stent restenosis with TIMI flow grade 3 and no visible contrast detect, which could have been caused by stent thrombosis. No MACCE was reported in other patients.
Six-month angiographic and IVUS results in stented segment
Scheduled coronary angiography at six months was performed in 15 patients (93.8%). The patient who refused angiography was asymptomatic. At six months, mean late luminal loss was 0.63 mm on quantitative coronary angiography and percent stent volume obstruction was 27.2% on IVUS (Table 3).Binary restenosis occurred in two patients (13.3%). Restenosis was focal in one patient and diffuse in the other. In the case of the patient with diffuse restenosis (case #4), the patient had no symptoms and a percent diameter stenosis was 58% at six months. This patient did not undergo repeat revascularization. Focal restenosis occurred in an insulin-dependent patient (case #12). This patient was subsequently treated with directional coronary atherectomy and implantation of a paclitaxel-eluting stent. Intravascular ultrasound analysis revealed incomplete stent apposition in one patient (6.3%), which had resolved at six months. Furthermore, no late-acquired incomplete stent apposition was observed for the entire patient population.
Histology of the atherectomy specimen
Histologic analysis showed tissue consisting of a mixture of myxoid and fibrous material with variable cell density. Some of the fibrous pieces showed a paucity of cells. The myxoid fragments consisted of smooth muscle cells (SMCs) within variable amounts of proteoglycan (Figs. 2Ato 2C). Some thrombus remnants were seen and assessed to be several days old (Figs. 2D and 2E, no SMCs, only some nuclei), presumably related to the small MI. The atherectomy specimens included metal stent fragments. That indicates that the plaques behind the stent struts were included in the specimens. There were large areas of calcification in both the fibrous and myxoid tissue (Fig. 2F), presumably old plaque, as these were also observed with IVUS. Immunocytochemistry (Figs. 2C and 2E) shows that most cells were SMC-alpha-actin positive. Mib-1 (proliferation), Mac387 (macrophages), and CD34 were negative. There were only a few CD45-positive cells (leukocytes).
HAMA test results
The HAMA testing was conducted on a subset of the population (last four consecutive patients). A positive assay due to the treatment with the study device was not observed in these patients.
This study is the first clinical experience with implantation of a bioengineered stent. The results of the HEALING-FIM registry show that the EPC capture stent is safe and feasible: with no stent thrombosis (30 days or 6 months), and MACCE occurred in only one patient (6.3%), despite only 30 days of clopidogrel therapy.
This clinical registry was preceded by several experimental studies. In an in vivo study, at 1 h after deployment the EPC capture stent showed a >90% cell coverage, while the bare stainless steel stents were almost completely devoid of cells. Histologic analysis at 31 days showed that percent area luminal stenosis was significantly reduced with the EPC capture stents compared with stainless steel stents (15.49 ± 4.54% vs. 23.96 ± 7.70%, p = 0.01) (9).
These preclinical and preliminary clinical results have to be interpreted carefully, considering the recent emergence of new technologies such as drug-eluting stents. Drug-eluting stents inhibit the inflammatory and proliferative process of the normal healing response, including the formation of a confluent endothelial layer on the stent (3). The EPC capture stents induce the rapid establishment of a functional endothelial layer early in the healing response. In this registry, the atherectomy specimen indicated a well-healed artery with minimal inflammation.
Of note, the neointimal hyperplasia in the EPC capture coating stents was not significantly reduced when compared with the usual late loss seen after conventional bare metal stent implantation. It has been argued that EPC capture coating covers only the stent struts, and theoretically no early functional endothelial lining can be expected in the interstrut space, although the interstrut area in the animal model (healthy coronary artery undergoing direct stenting) was covered with functional endothelium within 48 h; this situation differs substantially from a human pathologic atherosclerosis vessel after balloon injury. In addition, the bioactivity of these prototype EPC capture stents used in HEALING-FIM registry was unstable and was easily reduced by sterilization with gamma irradiation. It was subsequently discovered through the use of a bioassay that gamma irradiation lessened the immunoaffinity of EPC capture prototype stents used in this trial. Therefore, it is likely that the reduced bioactivity of EPC capture stents in the HEALING-FIM registry may not have been enough to inhibit neointimal hyperplasia after stent implantation.
The technology behind the creation of an EPC affinity surface is achieved by attaching murine monoclonal antihuman CD 34 antibodies to the stent. An immunoreaction against the murine monoclonal antibody may occur in patients who have human ant-murine antibody (HAMA). In addition, production of HAMA may result in the neutralization of the EPC capture surface. In this registry, increased HAMA levels were not observed in the four patients who underwent serial HAMA testing, and no patients exhibited systemic symptoms of immunoreaction. This safety issue was in agreement with other trials, evaluating immunologic treatment using murine antibody for ovarian cancer (10).
Because of the small sample size and single-center enrollment, the present study did not fully evaluate the efficacy of this device. However, further developments in this technology, such a providing a dry stent premounted on a delivery system and maintaining good activity post-sterilization, are currently being evaluated in the HEALING-II registry (Table 4).The technology for preserving antibody structure and bioactivity has advanced, resulting in a higher capture of EPCs (Fig. 3).As a result of these improvements, further clinical investigation of this technology is warranted to evaluate the efficacy of this device for the treatment of coronary artery disease.
The present study demonstrates that the EPC capture stent is safe and feasible. Further study and development of this promising technology are needed to confirm the clinical efficacy of this bio-engineered stent.
This study was supported by OrbusNeich, Fort Lauderdale, Florida. Mr. Davis and Dr. Rowland are full-time employees of OrbusNeich, and Dr. Kutryk is a consultant of Orbus Medical Technologies.
- Abbreviations and acronyms
- endothelial progenitor cell
- human antimurine antibody
- Healthy Endothelial Accelerated Lining Inhibits Neointimal Growth-First In Man
- intravascular ultrasound
- major adverse cardiac and cerebrovascular events
- myocardial infarction
- smooth muscle cell
- Thrombolysis In Myocardial Infarction
- Received December 9, 2004.
- Revision received January 22, 2005.
- Accepted January 25, 2005.
- American College of Cardiology Foundation
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