Assessment of Novel Anti-thrombotic Fusion Proteins for Inhibition of Stenosis in a Porcine Model of Arteriovenous Graft.

BACKGROUND: Hemodialysis arteriovenous synthetic grafts (AVG) provide high volumetric blood flow rates shortly after surgical placement. However, stenosis often develops at the vein-graft anastomosis contributing to thrombosis and early graft failure. Two novel fusion proteins, ANV-6L15 and TAP-ANV, inhibit the tissue factor/factor VIIa coagulation complex and the factor Xa/factor Va complex, respectively. Each inhibitor domain is fused to an annexin V domain that targets the inhibitor activity to sites of vascular injury to locally inhibit thrombosis. This study's objective was to determine if these antithrombotic proteins are safe and effective in inhibiting AVG stenosis. METHODS: A bolus of either TAP-ANV or ANV-6L15 fusion protein was administered intravenously immediately prior to surgical placement of a synthetic graft between the external jugular vein and common carotid artery in a porcine model. At surgery, the vein and artery were irrigated with the anti-thrombotic fusion protein. Control animals received intravenous heparin. At 4 weeks, MRI was performed to evaluate graft patency, the pigs were then euthanized and grafts and attached vessels were explanted for histomorphometric assessment of neointimal hyperplasia at the vein-graft anastomosis. Blood was collected at surgery, immediately after surgery and at euthanasia for serum metabolic panels and coagulation chemistries. RESULTS: No acute thrombosis occurred in the control group or in either experimental group. No abnormal serum chemistries, activated clotting times or PT, PTT values were observed after treatment in experimental or control animals. However, at the vein-graft anastomosis, there was no difference between the control and experimental groups in cross-sectional lumen areas, as measured on MRI, and no difference in hyperplasia areas as determined by histomorphometry. These results suggest that local irrigation of TAP-ANV or ANV-6L15 intra-operatively was as effective in inhibiting acute graft thrombosis as intravenous administration of heparin, but failed to inhibit hyperplasia development and stenosis in AVG.

In stent restenosis and thrombosis assessment after EP224283 injection in a rat model.

OBJECTIVE: After stent implantation, platelet aggregation and thrombus formation are thought to play a key role in the early phase of in-stent restenosis (ISR). Drug-eluting stents have reduced ISR, but are associated with healing-related issues or hypersensitivity reactions, leading to an increased risk of late acute stent thrombosis. EP224283 is a new dual-action antithrombotic molecule combining a GPIIbIIIa antagonist and a factor Xa inhibitor. We investigated its efficacy on restenosis in a rat model of ISR and on platelet adhesion. METHODS AND RESULTS: Rat aortas were stented and the animals received either EP224283 or vehicle subcutaneously every 48 h. At day 7 and day 28 after surgery, the stented aortas were removed and processed for morphometric analysis or protein analysis. At day 28, EP224283 significantly reduced neointima growth (in the range of 20%). Protein analysis revealed that EP224283 reduced cell proliferation pathways: ERK1/2 and Akt were down-regulated and p38 up-regulated. Expression of Ki67 was also reduced. In vitro assessment depicted a reduction of platelet activation and platelet adhesion among treated rats. CONCLUSION: These results show a beneficial effect of EP224283 on in-stent restenosis and on stent thrombogenicity that may improve results after stent implantation. Further investigations are required to assess the efficacy of a local delivery of EP224283 on both acute thrombosis and ISR.

"Pharmacologic" distal protection using prophylactic, intragraft nicardipine to prevent no-reflow and non-Q-wave myocardial infarction during elective saphenous vein graft intervention.

BACKGROUND: Coronary saphenous vein bypass graft (SVG) stenting has been associated with up to a 30% rate of no-reflow or myocardial infarction (MI) when performed without distal protection. METHODS: We evaluated the technique using prophylactic pharmacologic arteriolar vasodilatation with intracoronary nicardipine followed by immediate direct stenting for the treatment of degenerated coronary SVGs without mechanical distal protection. Data were collected from 83 consecutive elective SVG interventions in 68 patients. Quantitative coronary angiographic measurements were performed by the Borgess angiographic core lab. Electrocardiograms (ECGs), CPKs, and CPK-MBs were obtained preprocedure and at 12 to 18 hours after the intervention. Follow-up data at 30 days were obtained in 67/68 (98%) patients. RESULTS: The average graft age was 11.9 +/- 6.6 years with thrombus in 26/83 vessels (31%). The primary adverse endpoint of total CPK >3 times the upper limit of normal (ULN), or CPK-MB >3 times the ULN were seen in 1/68 (1.5%) and 3/68 (4.4%) patients, respectively. No-/slow-reflow was observed transiently in 2/83 SVG interventions (2.4%). Of the patients, 1/68 had persistent, minor ECG changes after stenting (1.4%). No patient had a Q-wave MI. Inhospital major adverse cardiac events (MACE) (death, MI, repeat TLR) were observed in only 3/68 patients (CPK-MB elevation). There were no additional MACE events (0/68) from hospital discharge to 30 days. CONCLUSIONS: (1) Prophylactic vasodilatation with intragraft nicardipine followed by direct stenting appears to be a safe and effective means of performing elective SVG revascularization; (2) this approach may provide a simple and time- and cost-effective alternative or adjunct to mechanical distal protection for elective SVG interventions.

Dialysis graft declotting with very low dose urokinase: is it feasible to use "less and wait?".

PURPOSE: "Lyse and wait" dialysis graft declotting is simple and effective, but the minimum necessary dose of urokinase is unknown. The efficacy of the technique with very low dose urokinase is evaluated. MATERIALS AND METHODS: Twenty-one grafts in 17 patients were declotted with use of the lyse and wait technique, but with 5,000-15,000 U of urokinase initially. Graft angiography was performed when an interventional suite was available. Declotting was completed in the manner chosen by the individual operator. Angiograms, interventional radiology records, and dialysis records were reviewed. RESULTS: Technical and clinical success were achieved in 95% of cases. Mean initial urokinase dose was 6,667 U. Initial angiography was performed at a mean 86 minutes. Two cases required second 5,000-U boluses to achieve complete graft thrombolysis. In all other cases, complete or near complete graft thrombolysis was observed with the initial very low dose. No bleeding, arterial embolic, or pulmonary embolic complications were observed. CONCLUSIONS: Doses of urokinase as low as 5,000 U are effective for lyse and wait declotting. A substantial reduction in drug costs can be expected with the "less and wait" modification. Bleeding risk may also be reduced.

Limitations of intra-arterial thrombolysis.

The purpose of this study was to assess the efficacy, costs and complications of acutely ischaemic limbs initially treated with urokinase-induced thrombolysis, and to compare the subsequent patencies of occluded native arteries, vein grafts and prosthetic grafts. Data from 45 consecutive episodes of thrombolysis in 37 patients (37 limbs) were reviewed retrospectively. An initial bolus dose of urokinase was used in all limbs, with pulse-spray in nine 30 limbs received additional infusions of urokinase including six who received initial treatment with pulse-spray. Percutaneous transluminal angioplasty or surgical intervention was undertaken at any stage when considered appropriate. There were 14 occluded native arteries (31%), 12 vein grafts (27%), 16 prosthetic polytetrafluoroethylene grafts (35%), and three composite grafts (7%). Thrombolysis was initially successful in 33 episodes (73%). Of these, no additional procedure was required in three, percutaneous transluminal angioplasty in 23, other surgery in 11 and bypass surgery in seven with more than one additional procedure being undertaken in 10. Of the 12 episodes with initial failure, major surgery was performed in eight and no procedure in four. Lysis-related complications occurred in 19 treatment episodes (42%) with major bleeding in seven (16%), minor bleeding in 10 (22%), distal embolization in 11 (24%) and arterial dissection in two (4%). One amputation (2%) and one death (2%), both directly attributable to thrombolysis, occurred within 1 month of treatment. Complications occurred in 10 of 19 (53%) prosthetic or composite grafts, five of 12 (42%) vein grafts, and four of 14 (29%) native artery occlusions (P = 0.38). The median follow-up of patients remaining alive was 24 months. The overall mean (s.e.m.)patency rate at 6 months of lysed vessels or replacement bypass grafts was 44% (8%). Presentation with a native artery and or graft occlusion was associated with superior long-term patency (any subsequent conduit) compared with presentation with an occluded prosthetic graft (log rank chi2[1] = 7.00, P = 0.008) and reflects the inferior run-off of the latter patients. The average cost of thrombolysis in radiological procedures and disposables alone excluding intensive care and complications was AUD$2440.

Pharmacoeconomic considerations in peripheral arterial thrombolytic therapy.

Intraarterial thrombolytic therapy with urokinase (UK) offers documented advantages to alternatives for treating both subacute and the initial presentation of acute peripheral arterial occlusion (PAO), including reduced morbidity and mortality. Treatment with intraarterial UK does not increase overall health care costs; hospital length of stay is either similar to that with other therapies (acute PAO) or is shortened (subacute PAO). Total hospital charges associated with use of intraarterial UK are also not significantly elevated. Thus, thrombolysis with UK offers both a clinically superior and a cost-beneficial way to treat PAO.

Differential mechanisms of failure of autogenous and non-autogenous bypass conduits: an assessment following successful graft thrombolysis.

The advent of graft thrombolysis has provided an objective means for evaluating the etiology of graft occlusion. Over a 10-year period, intra-arterial urokinase (102 cases) or streptokinase (seven cases) was used in 109 infrainguinal conduits (30 autogenous and 79 non-autogenous) that failed 30 days or more after implantation. Thrombolysis was not achieved in 19 additional graft occlusions; these cases were excluded from study because of an inability to define the mechanism of failure. Non-invasive laboratory data were available within 6 months of graft occlusion in 82 (75%) of the cases, with Doppler segmental studies in 80 cases (73%) and duplex ultrasonography studies in 39 cases (36%). Pre-failure non-invasive laboratory abnormalities were detected more frequently in autogenous grafts (21 of 24 patients, 88%), while non-autogenous grafts usually occluded without prior hemodynamic change (11 of 58 patients had abnormalities, 19%) (P < 0.001). Thrombolysis uncovered anatomic defects responsible for thrombosis in 27 (90%) of 30 autogenous grafts compared with only 32 (41%) of non-autogenous conduits (P < 0.001). The most common lesions underlying autogenous graft failure comprised stenoses within the body of the graft (11 cases, 37%), while the most common lesions in failed non-autogenous grafts appeared to be stenoses at an anastomosis (21 cases, 27%). Thus, the mechanisms underlying the late failure of autogenous and non-autogenous grafts differ markedly; autogenous grafts most commonly fail as a result of the gradual development of lesions intrinsic to the graft, while non-autogenous grafts fail precipitously, presumably as a result of some non-anatomic mechanism.

Acute myocardial infarction complicating urokinase infusion for total saphenous vein graft occlusion.

Saphenous vein graft occlusions have been successfully treated with extended urokinase infusions. We report a case of myocardial infarction complicating this treatment. A review of reported cases suggests that this complication may not be uncommon. The optional drug, dose, and infusion technique for intra-graft lytic therapy has not been determined. The costs, risks, and difficulty of this technique may limit its application.

Cost-effectiveness of intra-arterial thrombolytic therapy.

We reviewed the clinical course of 23 patients who received 24 intra-arterial infusions of either streptokinase or urokinase to treat 14 arteries and ten arterial grafts that were occluded due to primary thrombosis (22) or artery-artery embolism (two). Time from symptom onset to treatment was one to 28 days (mean, 11 days). Five infusions (21%) were completely successful since symptoms were eliminated without subsequent operation. Seven infusions (29%) were partially successful since thrombolysis aided, limited, or postponed subsequent surgery. Six infusions (25%) were failures since thrombolysis or clinical improvement did not occur and surgery was required. Six infusions (25%) were associated with thrombolytic complications that required urgent operation (less severe complications occurred in an additional 17% of cases [4/24]). Of the 19 patients without complete success after thrombolytic therapy, 16 underwent surgery during the same admission, two were not operable due to distal disease, and one declined operation. Of the 16 operations, 15 (94%) were successful in restoring graft or artery patency and achieving limb salvage, whereas one failed. In the 12 patients with failure or major complications of thrombolytic treatment, all had successful surgical outcome without morbidity. The actual mean cost of thrombolytic treatment was $8200 per patient and was comparable with the actual mean cost of subsequent surgical treatment in the 16 patients who required operation ($8900 per patient). The effective cost of thrombolytic and surgical treatment was calculated by dividing the actual costs by the proportion of successful cases. The effective cost of thrombolytic therapy per complete success was $39,200 and per complete or partial success was $16,500. This was significantly more than the effective cost of $9400 per complete success of surgical therapy.