P0117DOSE ESCALATING TOXICOLOGY STUDY OF ELP-VEGF, A NOVEL BIOLOGIC FOR RENAL THERAPEUTIC ANGIOGENESIS

Abstract

Abstract Background and Aims Chronic renal disease, irrespective of the etiology, is hallmarked by renal microvascular rarefaction. This rarefaction is associated with a reduced bioavailability of the pro-angiogenic cytokine vascular endothelial growth factor (VEGF). Recently, we developed a therapeutic intervention for ischemic renal conditions using therapeutic angiogenesis – supplementing renal VEGF levels. This was achieved by administration of a biopolymer-stabilized form of VEGF using a fusion protein between human VEGF-A121 and an elastin-like polypeptide carrier protein (ELP-VEGF). ELP-VEGF induced microvascular remodeling and increased microvascular density when administered intra-renally in swine models of unilateral renovascular disease (RVD) and chronic kidney disease (CKD), which was associated with improved renal function, reduced renal inflammation, and reduced renal fibrosis. Also, ELP-VEGF targeted the kidney when administered systemically and induced similar beneficial effects on renal function and renal vascular density. The therapeutic dose of ELP-VEGF in the swine models was 0.1 mg/kg when administered intra-renally and 1.0 mg/kg when administered intravenously. The aims of the present study were to determine the maximum tolerated dose of ELP-VEGF and assess its dose-related toxicity. We hypothesize that ELP-VEGF will not exhibit toxicity at therapeutic doses. Method A dose escalating toxicology experiment was performed in Sprague Dawley. Female rats were instrumented with either carotid catheters or carotid artery telemeters for blood pressure monitoring. After a recovery period, a baseline 24-hour urine sample was collected, and baseline glomerular filtration rate (GFR) was measured by monitoring FITC-sinistrin clearance via transdermal fluorescence. A single injection of saline control or ELP-VEGF (0.1, 1, 10, 100, or 200 mg/kg) was administered, and blood pressure was monitored by telemetry or by direct carotid arterial pressure measurements. Body weights were monitored daily throughout the study, and 24-hour urine collections and GFR measurements were repeated 7 and 14 days after protein injection. On day 14 after injection, blood was collected, one kidney was collected and fixed for histological examination, and the second kidney was perfused with Microfil vascular contrast agent for quantification of renal vascular density by micro-CT. Results ELP-VEGF caused no significant changes in body weight at any dose. At the highest doses there was an acute drop in blood pressure (20 – 30 mm Hg at 100 and 200 mg/kg dose, respectively) for approximately twenty minutes post-injection which then normalized. GFR was unchanged by ELP-VEGF at doses up to 100 mg/kg. However, GFR was significantly increased 14 days after treatment with 200 mg/kg ELP-VEGF (1.5 +/- 0.3 versus 2.7 +/- 0.3 mL/min/100 g body weight, p=0.0021). Plasma toxicology revealed no changes in markers of liver or kidney toxicity (AST, ALT, BUN, creatinine, LDH, bilirubin) at any dose or time point. Renal vascular density was unaffected by ELP-VEGF at doses up to 10 mg/kg. However, at 200 mg/kg ELP-VEGF, renal vascular density was significantly decreased for small (0 – 100 micron, p=0.0007) and intermediate (100 – 200 micron, p=0.028) sized vessels Conclusion ELP-VEGF has a proven therapeutic potential for treatment of RVD and CKD at therapeutic doses ranging from 0.1 – 1.0 mg/kg in swine models. Dose escalating toxicity assessment in rats found no effect of ELP-VEGF on body weight, blood pressure, plasma markers of liver and renal toxicity, GFR, or renal vascular density at these therapeutic doses. ELP-VEGF doses of 100 – 200 mg/kg caused transient hypotension immediately after the injection and were associated with increased GFR and loss of renal vascular density, possibly indicating renal damage at ultra-high doses. We conclude that the therapeutic window for ELP-VEGF is 100 – 1,000 times the effective therapeutic dose.