Nanoparticles based on PLGA:poloxamer blends for the delivery of proangiogenic growth factors.

New blood vessel formation is a critical requirement for treating many vascular and ischemia related diseases, as well as for many tissue engineering applications. Angiogenesis and vasculogenesis, in fact, represent crucial processes for the functional regeneration of complex tissues through tissue engineering strategies. Several growth factors (GFs) and signaling molecules involved in blood vessels formation have been identified, but their application to the clinical setting is still strongly limited by their extremely short half-life in the body. To overcome these limitations, we have developed a new injectable controlled release device based on polymeric nanoparticles for the delivery of two natural proangiogenic GFs: platelet derived growth factor (PDGF-BB) and fibroblast growth factor (FGF-2). The nanoparticle system was prepared by a modified solvent diffusion technique, encapsulating the GF both in presence and in the absence of two stabilizing agents: bovine serum albumin (BSA) and heparin sodium salt (Hp). The developed nanocarriers were characterized for morphology, size, encapsulation efficiency, release kinetics in vitro and GF activity in cell cultures. The results have indicated that the coencapsulation of stabilizing agents can preserve the GF active structure and, in addition, increase their encapsulation efficiency into nanoparticles. Through this optimization process, we were able to raise the encapsulation efficiency of FGF-2 to 63%, and that of PDGF-BB to 87%. These PLGA:poloxamer blend nanoparticles loaded with GFs were able to release PDGF-BB and FGF-2 in a sustained fashion for more than a month. This work also confirms other positive features of PLGA:poloxamer nanoparticles. Namely, they are able to maintain their stability in simulated biological medium, and they are also nontoxic to cell culture models. Incubation of nanoparticles loaded with FGF-2 or PDGF-BB with endothelial cell culture models has confirmed that GFs are released in a bioactive form. Altogether, these results underline the interest of PLGA:poloxamer nanoparticles for the controlled delivery of GFs and substantiate their potential for the treatment of ischemic diseases and for tissue engineering applications.

Improved delivery of angiogenesis inhibitors from PLGA:poloxamer blend micro- and nanoparticles.

Clinical studies have demonstrated the efficacy of new strategies in cancer therapy, such as chemotherapy and radiotherapy, associated to the administration of tumour vascularization inhibitors. A critical limitation for the clinical application of angiogenesis inhibitors relies in their instability in biological environment and high-dose requirements. This work has attempted to overcome this limitation by designing an adequate delivery vehicle consisting of PLGA:poloxamer blend micro- and nanoparticles. The potential of this delivery system was investigated for a new synthetic angiogenesis inhibitor named polyaminoacid JS-2892b. PLGA:poloxamer (ratio 10 : 1) blend microparticles were prepared by the oil-in-oil emulsion technique, while PLGA:poloxamer (ratio 1 : 1) blend nanoparticles were obtained by a modified solvent diffusion technique. The results showed that, by adjusting the formulation conditions, it was possible to efficiently encapsulate the polyaminoacid JS-2892b within PLGA:poloxamer micro- (particle size of 20 microm and encapsulation efficiency higher than 90%) and nanoparticles (particle size of less than 280 nm and encapsulation efficiency of 52%). In addition, the delivery of the polyaminoacid JS-2892b from the particles could be controlled, without altering its stability, for extended periods of time (from a few days to over a month). The release of the encapsulated compound was significantly affected by the particle size and the way the drug is dispersed into the polymeric matrix. Therefore, this study provides information about the formulation conditions and potential of biodegradable particles for the controlled release of polyaminoacid JS-2892b.

Hyaluronic acid/Chitosan nanoparticles as delivery vehicles for VEGF and PDGF-BB.

The development of a vascular network in tissue-engineered constructs is a fundamental bottleneck of bioregenerative medicine, particularly when the size of the implant exceeds a certain limit given by diffusion lengths and/or if the host tissue shows a very active metabolism. One of the approaches to achieve the vascularization of tissue constructs is generating a sustained release of proangiogenic factors from the ischemic site. This work describes the formation and characterization of hyaluronic acid-chitosan (HA/CS) nanoparticles for the delivery of two pro-angiogenic growth factors: vascular endothelial growth factor (VEGF) and platelet derived growth factor (PDGF-BB). These nanoparticles were prepared by an ionic gelification technique, and different formulations were developed by encapsulating the growth factors in association with two stabilizing agents: bovine serum albumin or heparin sodium salt. These carriers were characterized with regard to their physicochemical properties, their stability in biological media, and their cytotoxicity in the C3a hepatoma cell line. The results show that nanoparticles around 200 nm can be prepared by this method. HA/CS nanoparticles were stable when incubated in EMEM cell culture medium or in water at 37°C for 24 h. Cell culture tests confirmed that HA/CS nanoparticles are not cytotoxic within the concentration range used for growth factor delivery. Moreover, HA/CS nanoparticles were able to entrap efficiently both growth factors, reaching association values of 94% and 54% for VEGF and PDGF, respectively. In vitro release studies confirm that PDGF-BB is released from HA/CS nanoparticles in a sustained manner over approximately 1 week. On the other hand, VEGF is completely released within the first 24 h.

PLGA:poloxamer blend micro- and nanoparticles as controlled release systems for synthetic proangiogenic factors.

Tissue engineering is one of the most promising research areas in bioregenerative medicine. However, the restoration of biological functionalities by implanting bioartificially engineered tissues is still highly limited because of their lack of vascular networks. The use of proangiogenic molecules delivered from a controlled release device is a promising strategy to induce tissue vascularization. Indeed, the controlled release system can enhance the therapeutic effect in vivo of many short half-life drugs, while circumventing the need for repeated administrations. In this work, PLGA:poloxamer blend based micro- and nanoparticles have been developed for the sustained delivery of a recently developed synthetic proangiogenic compound: SHA-2-22. Drug-loaded PLGA:poloxamer blend microparticles were prepared by an oil-in-oil solvent extraction/evaporation technique. Drug-loaded PLGA:poloxamer nanoparticles were prepared by a modified solvent diffusion technique. These drug carriers were characterized with regard to their physicochemical properties, morphology, drug encapsulation efficiency and release kinetics in vitro. The results show that by adjusting the formulation conditions, it is possible to obtain PLGA:poloxamer micro- and nanoparticles with very high drug loadings, and with the capacity to release the active compound in a controlled way for up to one month. In vitro cell assays performed in an endothelial cell model confirmed the bioactivity of SHA-22-2 encapsulated in PLGA:poloxamer microparticles.

Effect of bridging ligand structure on the thermal stability and DNA binding properties of iron(II) triple helicates.

Triple helical di-iron complexes, readily prepared through interaction of iron(II) ions with imine-based ligands, are cylinder-shaped tetracations comparable in size and shape to common protein DNA recognition units. They have a strong tendency to coil DNA, and have recently been found to induce formation of three-way junctions in palindromic oligonucleotides. To introduce potential H bond acceptor units onto the surface of triple-helicates, new iron(II) complexes have been synthesized in which the central linking unit in the bridging ligand is S or O, instead of CH(2). The DNA binding properties of these new metallo-helicates were studied using UV-vis spectroscopy and circular and linear dichroism. Results show that the three iron triple helicates bind the DNA in a similar way but that the stability of the triple helicate unit is decreased with the O linked bridging ligand.