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ABSTRACT: Therapies based on RNA interference, using agents such as siRNA, are limited by the absence of safe, efficient vehicles for targeted delivery in vivo. The barriers to siRNA delivery are well known and can be individually overcome by addition of functional modules, such as conjugation of moieties for cell penetration or targeting. But, so far, it has been impossible to engineer multiple modules into a single unit. Here, we describe the synthesis of degradable nanoparticles that carry eight synergistic functions: 1) polymer matrix for stabilization/controlled release; 2) siRNA for gene knockdown; 3) agent to enhance endosomal escape; 4) agent to enhance siRNA potency; 5) surface-bound PEG for enhancing circulatory time; and surface-bound peptides for 6) cell penetration; 7) endosomal escape; and 8) tumor targeting. Further, we demonstrate that this approach can provide prolonged knockdown of PLK1 and control of tumor growth in vivo. Importantly, all elements in these octa-functional nanoparticles are known to be safe for human use and each function can be individually controlled, giving this approach to synthetic RNA-loaded nanoparticles potential in a variety of clinical applications.
Biomaterials 01/2012; 33(2):583-91. · 7.40 Impact Factor
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ABSTRACT: Inhibition of the epidermal growth factor receptor (EGFR) reduces tumour growth and metastases and promotes axon regeneration in the central nervous system. Current EGFR inhibition strategies include the administration of reversible small-molecule tyrosine kinase inhibitors (TKIs). However, to be effective in vivo sustained delivery is required. This study explored the feasibility of encapsulating the tyrphostin 4-(3-chloroanilino)-6,7-dimethoxyquinazoline (AG1478) in poly(lactic-co-glycolic acid) (PLGA) microspheres using three different emulsion methods: solid-in-oil-in-water, oil-in-water and oil-in-water with co-solvent. Addition of a co-solvent increased loading and release of AG1478 and significantly (p < 0.001) decreased microsphere size. Co-solvent addition also prolonged AG1478 release from 6 months to over 9 months. Once released AG1478 remained bioactive and inhibited EGFR in immortalized rat fibroblasts and EGFR-amplified human carcinoma cells. These results demonstrate that AG1478 can be encapsulated in PLGA with sustained release and retain bioactivity; thereby providing a new platform for controlled administration of EGFR TKIs.
Journal of Microencapsulation 05/2010; 27(3):263-71. · 1.55 Impact Factor
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ABSTRACT: Blood loss is the major cause of death in both civilian and battlefield traumas. Methods to staunch bleeding include pressure dressings and absorbent materials. For example, QuikClot effectively halts bleeding by absorbing large quantities of fluid and concentrating platelets to augment clotting, but these treatments are limited to compressible and exposed wounds. An ideal treatment would halt bleeding only at the injury site, be stable at room temperature, be administered easily, and work effectively for internal injuries. We have developed synthetic platelets based on Arg-Gly-Asp functionalized nanoparticles, which halve bleeding time after intravenous administration in a rat model of major trauma. The effects of these synthetic platelets surpass other treatments, including recombinant factor VIIa, which is used clinically for uncontrolled bleeding. Synthetic platelets were cleared within 24 hours at a dose of 20 mg/ml, and no complications were seen out to 7 days after infusion, the longest time point studied. These synthetic platelets may be useful for early intervention in trauma and demonstrate the role that nanotechnology can have in addressing unmet medical needs.
Science translational medicine 12/2009; 1(11):11ra22. · 7.80 Impact Factor
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ABSTRACT: Poly(lactic-co-glycolic acid) (PLGA) is one of the more widely used polymers for biomedical applications. Nonetheless, PLGA lacks chemical moieties that facilitate cellular interactions and surface chemistries. Furthermore, incorporation of hydrophilic molecules is often problematic. The integration of polymer functionalities would afford the opportunity to alter device characteristics, thereby enabling control over drug interactions, conjugations and cellular phenomena. In an effort to introduce amine functionalities and improve polymer versatility, we synthesized two block copolymers (PLGA-PLL 502H and PLGA-PLL 503H) composed of PLGA and poly(epsilon-carbobenzoxy-l-lysine) utilizing dicyclohexyl carbodiimide coupling. PLGA-PLL microspheres encapsulated approximately sixfold (502H) and threefold (503H) more vascular endothelial growth factor, and 41% (503H) more ciliary neurotrophic factor than their PLGA counterparts. While the amine functionalities were amenable to the delivery of large molecules and surface conjugations, they did not compromise polymer biocompatibility. With the versatile combination of properties, biocompatibility and ease of synthesis, these block copolymers have the potential for diverse utility in the fields of drug delivery and tissue engineering.
Acta biomaterialia 06/2009; 5(8):2860-71. · 3.98 Impact Factor
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ABSTRACT: Transplantation of Bcl-2-transduced human umbilical vein endothelial cells (ECs) in protein gels into the gastrocnemius muscle improves local reperfusion in immunodeficient mouse hosts with induced hind limb ischemia. We tested the hypothesis that incorporation of local, sustained growth factor delivery could enhance and accelerate this effect. Tissue engineering scaffolds often use synthetic polymers to enable controlled release of proteins, but most synthetic delivery systems have major limitations, most notably hydrophobicity and inefficient protein loading. Here, we report the development of a novel alginate-based delivery system for vascular endothelial growth factor-A(165) (VEGF) that exhibits superior loading efficiency and physical properties to previous systems in vitro. In vivo, VEGF released from alginate microparticles within protein gels was biologically active and, when combined with EC transplantation, led to increased survival of transplanted cells at 28 days. The composite graft described also improved early (14 days) tissue perfusion and late (28 days) muscle myoglobin expression, a sign of recovery from ischemia, compared with EC transplantation and VEGF delivery separately. We conclude that our improved approach to sustained VEGF delivery in tissue engineering is useful in vivo and that the integration of high efficiency protein delivery enhances the therapeutic effect of protein gel-based EC transplantation.
The FASEB Journal 06/2008; 22(8):2949-56. · 5.71 Impact Factor
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ABSTRACT: Neural stem cells (NSCs) have the potential to replace the major cell types of the central nervous system (CNS) and may be important in therapies for injuries to and diseases of the CNS. However, for such treatments to be safe and successful, NSCs must survive and differentiate appropriately following transplantation. A number of polymer scaffolds have shown promise in improving the survival and promoting the differentiation of NSCs. To capitalize on the interaction between scaffolds and NSCs, we need to determine the fundamental material properties that influence NSC behavior. To investigate the role of material properties on NSCs, we synthesized a library of 52 hydrogels composed of poly(ethylene glycol) and poly(L-lysine) (PLL). This library of hydrogels allows independent variation of chemical and mechanical properties across a wide range of values. By culturing NSCs on this library, we have identified a subset of gels that promotes NSC migration and a further subset that promotes NSC differentiation. By combining the material properties of these subsets with the cell behavior, we determined that mechanical properties play a critical role in NSC behavior with elastic moduli promoting NSC migration and neuronal differentiation. Amine concentration is less critical, but PLL molecular weight also plays a role in NSC differentiation.
Journal of Biomedical Materials Research Part A 05/2008; 89(2):499-509. · 2.63 Impact Factor
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ABSTRACT: A microvascular network is critical for the survival and function of most tissues. We have investigated the potential of neural progenitor cells to augment the formation and stabilization of microvascular networks in a previously uncharacterized three-dimensional macroporous hydrogel and the ability of this engineered system to develop a functional microcirculation in vivo. The hydrogel is synthesized by cross-linking polyethylene glycol with polylysine around a salt-leached polylactic-co-glycolic acid scaffold that is degraded in a sodium hydroxide solution. An open macroporous network is formed that supports the efficient formation of tubular structures by brain endothelial cells. After subcutaneous implantation of hydrogel cocultures in mice, blood flow in new microvessels was apparent at 2 weeks with perfused networks established on the surface of implants at 6 weeks. Compared to endothelial cells cultured alone, cocultures of endothelial cells and neural progenitor cells had a significantly greater density of tubular structures positive for platelet endothelial cell adhesion molecule-1 at the 6-week time point. In implant cross sections, the presence of red blood cells in vessel lumens confirmed a functional microcirculation. These findings indicate that neural progenitor cells promote the formation of endothelial cell tubes in coculture and the development of a functional microcirculation in vivo. We demonstrate a previously undescribed strategy for creating stable microvascular networks to support engineered tissues of desired parenchymal cell origin.
Proceedings of the National Academy of Sciences 03/2006; 103(8):2512-7. · 9.68 Impact Factor
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ABSTRACT: A microvascular network is critical for the survival and function of most tissues. We have investigated the potential of neural
progenitor cells to augment the formation and stabilization of microvascular networks in a previously uncharacterized three-dimensional
macroporous hydrogel and the ability of this engineered system to develop a functional microcirculation in vivo. The hydrogel is synthesized by cross-linking polyethylene glycol with polylysine around a salt-leached polylactic-co-glycolic
acid scaffold that is degraded in a sodium hydroxide solution. An open macroporous network is formed that supports the efficient
formation of tubular structures by brain endothelial cells. After subcutaneous implantation of hydrogel cocultures in mice,
blood flow in new microvessels was apparent at 2 weeks with perfused networks established on the surface of implants at 6
weeks. Compared to endothelial cells cultured alone, cocultures of endothelial cells and neural progenitor cells had a significantly
greater density of tubular structures positive for platelet endothelial cell adhesion molecule-1 at the 6-week time point.
In implant cross sections, the presence of red blood cells in vessel lumens confirmed a functional microcirculation. These
findings indicate that neural progenitor cells promote the formation of endothelial cell tubes in coculture and the development
of a functional microcirculation in vivo. We demonstrate a previously undescribed strategy for creating stable microvascular networks to support engineered tissues
of desired parenchymal cell origin.
Proceedings of the National Academy of Sciences 02/2006; 103(8):2512-2517. · 9.68 Impact Factor
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[show abstract]
[hide abstract]
ABSTRACT: A microvascular network is critical for the survival and function of
most tissues. We have investigated the potential of neural progenitor
cells to augment the formation and stabilization of microvascular
networks in a previously uncharacterized three-dimensional macroporous
hydrogel and the ability of this engineered system to develop a
functional microcirculation in vivo. The hydrogel is synthesized by
cross-linking polyethylene glycol with polylysine around a salt-leached
polylactic-co-glycolic acid scaffold that is degraded in a sodium
hydroxide solution. An open macroporous network is formed that supports
the efficient formation of tubular structures by brain endothelial
cells. After subcutaneous implantation of hydrogel cocultures in mice,
blood flow in new microvessels was apparent at 2 weeks with perfused
networks established on the surface of implants at 6 weeks. Compared to
endothelial cells cultured alone, cocultures of endothelial cells and
neural progenitor cells had a significantly greater density of tubular
structures positive for platelet endothelial cell adhesion molecule-1 at
the 6-week time point. In implant cross sections, the presence of red
blood cells in vessel lumens confirmed a functional microcirculation.
These findings indicate that neural progenitor cells promote the
formation of endothelial cell tubes in coculture and the development of
a functional microcirculation in vivo. We demonstrate a previously
undescribed strategy for creating stable microvascular networks to
support engineered tissues of desired parenchymal cell origin.
microvasculature | neural stem cells | polymer | scaffold
Proceedings of the National Academy of Sciences 01/2006; 103:2512-2517. · 9.68 Impact Factor
