Biological properties of photocrosslinked alginate microcapsules.

An alternative form of gene therapy using recombinant cell lines delivering therapeutic products encapsulated in alginate hydrogel has proven effective in treating many murine models. The lack of long-term capsule stability has led to a new strategy to reinforce the microcapsules with a photopolymerized interpenetrating covalent network of N-vinylpyrrolidone (NVP) and sodium acrylate. Here the properties for potential application in gene therapy are reported. In assessing potential toxicity of the unpolymerized residues, HPLC showed that even after 1 week of washing, no toxic monomers could be detected. Their ability to sustain cell growth was monitored with growth of the encapsulated cells in vitro and in vivo. Although the initial photopolymerization caused significant cell damage, the cells were able to recover normal growth rates thereafter. After implanting into mice, the NVP-modified capsules showed a high level of biocompatibility as measured by hematological and biochemical functional tests. There was also no difference in the amount and type of plasma proteins adsorbing to the NVP-modified and the classical alginate capsules, thus indicating their similar biological compatibility. Both in vitro and in vivo tests confirmed that the NVP-modified capsules were more resistant to osmotic stress than the alginate microcapsules. Furthermore, when applied to the treatment of a murine model of human cancer by delivering encapsulated cells secreting angiostatin, the NVP-modified microcapsules suppressed tumor growth as successfully as the regular alginate microcapsules. In conclusion, the covalently modified microcapsules have shown a high level of biocompatibility, safety, increase in stability, and clinical efficacy for use as immunoisolation devices in gene therapy.

Quantitative chemical mapping of sodium acrylate- and N-vinylpyrrolidone-enhanced alginate microcapsules.

Alginate microcapsules enclosing recombinant cells secreting therapeutic products have been used successfully to treat several murine models of human diseases. The mechanical and chemical properties of these alginate capsules can be improved by the addition and in situ photo-polymerization of sodium acrylate and N-vinylpyrrolidone in the alginate capsule. The purpose of this modification was to form additional covalent cross-links. In this work we have used scanning transmission X-ray microscopy (STXM) to probe the nature and location of the chemical modifications in the modified capsules by comparison with unmodified capsules. Analysis of X-ray image sequences and selected area spectra has been used to map the calcium gradient in capsules, to identify the presence of polyacrylate throughout the capsules and the localization of poly-N-vinylpyrrolidone in the outer regions of the alginate capsules. The differences in the spatial distributions of these species have led to better understanding of the chemical modifications that provide a mechanically more stable capsule structure.

A novel method to enhance the stability of alginate-poly-L-lysine-alginate microcapsules.

Implantation of microencapsulated recombinant cells is an alternative approach to gene therapy. These genetically-engineered cells enclosed in microcapsules to deliver therapeutic recombinant products have been effective in treating several murine models of human diseases. However, the most commonly used microcapsules fabricated from alginate ionically cross-linked with calcium suffer from loss of long-term mechanical stability. We now report on a method to improve their stability by introducing additional polymers to provide covalent linkages via photopolymerization. Vinyl monomers and a photoinitiator were allowed to diffuse into the initially formed calcium-alginate microcapsules. In situ photopolymerization in the presence of sodium acrylate and N-vinylpyrrolidone substantially enhanced their mechanical strength. After four months of storage in saline, > 70% of these capsules remained intact in the osmotic pressure test, while the un-modified alginate microcapsules totally disintegrated. Tests of their permeability to polyethylene glycol of different molecular weight and their ability to support cell survival showed that these properties remained unaffected by the photopolymerization. Hence, these microcapsules modified by adding a network of vinyl polymers are promising candidates to use for long-term delivery of recombinant gene products in this cell-based method of gene therapy.