Sustained release formulations of rhVEGF165 produce a durable response in a murine model of peripheral angiogenesis |
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Authors: | Ann L DaughertyLinda K Rangell Renee EckertJose Zavala-Solorio Frank PealeRandall J Mrsny |
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Institution: | Genentech Inc., S. San Francisco, CA 94080, USA |
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Abstract: | Local delivery of therapeutic angiogenic agents that stimulate blood vessel formation represents a promising strategy for the treatment of peripheral vascular disease (PVD). At present, requirements for temporal and spatial parameters for localized delivery are unclear, with a variety of sustained delivery approaches being examined. Two polymer-based sustained formulations containing the 165 amino acid isoform of human recombinant vascular endothelial growth factor-A (rhVEGF165) were evaluated for their potential application in the treatment of PVD following intramuscular injection. Microspheres prepared from a 50:50 ratio of polylactic-co-glycolic acid (PLGA) and a gel of PLGA polymer solubilized in N-methyl pyrrolidone (PLGA:NMP) were each loaded with rhVEGF165 and tested in vitro and in vivo. PLGA microspheres averaged ∼30 μm in diameter and contained 8.9% (w/w) rhVEGF165, while the PLGA:NMP gel was formulated with varying amounts of spray freeze-dried rhVEGF165 to result in final gel formulations having concentrations of 0.36, 0.72, or 3.6 mg/mL rhVEGF165. In vitro release of rhVEGF165 from PLGA microspheres showed ∼10% cumulative release by day 6, whereas the cumulative release of rhVEGF165 from the PLGA:NMP gel matrices (0.65% w/w loading) was less than 0.25% at this same time point. While the in vitro release characteristics of these two sustained release formulations were broadly different, the plasma rhVEGF165 concentration-time profiles following hind-limb intramuscular (IM) injection of these formulations in non-compromised rats revealed similar in vivo pharmacokinetics. Three-dimensional resin casts of vascular architecture were prepared at days 3, 7, 14, 21, 28, 60, and 75 following a single IM dosing of these sustained release microsphere and gel matrix formulations in the gastrocnemius muscle of immune-compromised mice. Scanning electron microscopic visualization of these vascular casts demonstrated spatial arrangement of capillary sprouts and vessel enlargement consistent with profound vascular changes occurring within 3 days of dosing that persisted for 2 months, approximately 1 month beyond the anticipated completion of rhVEGF165 release from these sustained delivery formulations. Vascular re-modeling events were correlated with histological and immunohistochemical parameters attributed to known biological actions of rhVEGF165 signaling. Together, these pharmacokinetic and pharmacodynamic results support the use of sustained release PLGA-based formulations for the local delivery of rhVEGF165 to achieve a durable vascular re-modeling response. |
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Keywords: | Angiogenesis Vascular endothelial growth factor Peripheral vascular disease PLGA microspheres PLGA:NMP gels Vascular casts |
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