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Sasha Bakhru — 2007-08 Fellow

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Stem cells hold enormous potential for applications in regenerative medicine. However, reliable mechanisms for control of stem cell fate are needed to develop effective technologies in support of stem cell therapies. Our general objective is to establish stem cell-containing, biofunctional, polymeric microcapsule systems serving the dual purpose of advanced in-vitro 3D stem cell culture system as well as vehicle for in-vivo delivery of therapeutic stem cells.

Polymeric microcapsules afford opportunity to engineer and tune the microenvironment of stem cells in unprecedented ways by modular incorporation of natural and artificial niche constituents in order to elicit desired, specific cellular responses. In combination with genetic engineering and cued expression of genes in transplanted stem cells, external control will be gained over stem cell fate and function. In particular, adult neural stem cells (NSCs) are genetically engineered for enzyme replacement in the brain, as the basis for treatment of Mucopolysaccharidosis Type II (MPSII, Hunter’s Syndrome). NSCs are microencapsulated in methylated collagen (polycationic)/alginate (polyanionic)-based microcapsules formed by complex coacervation for expansion and subsequent delivery to the brain.

MPSII is a lethal, inborn metabolic disorder affecting roughly 1 in 100,000 to 160,000 live births worldwide. Inherited as an X-linked, recessive trait, it affects primarily young boys. Hunter patients produce insufficient quantities or dysfunctional copies of the lysosomal enzyme iduronate-2-sulfatase (IDS) in all cells of the body. Cells of all tissues of such patients become more and more dysfunctional through build-up of mucopolysaccharides (or glycosaminoglycans, GAGs), and untreated patients usually do not survive past their teen years. The only currently commercially available enzyme replacement therapy for MPSII, Elaprase® (Shire, PA, USA), effects nearly all cells of the body, however not the brain, as IDS is size-excluded from passage across the blood-brain barrier. The proposed microcapsule-based approach, optimized for treatment of the brain specifically, can complement systemic Elaprase® administration. Microcapsules consist of an alginate/methylated collagen-based material system. Microcapsules are to be optimized for rapid self-renewal of NSCs in vitro, so that NSCs that are isolated from biopsied patient brain tissues samples can be expanded for autologous treatments.

Microcapsules also serve as delivery vehicles in order to ensure NSC survival and functional integration after implantation. NSCs are genetically modified so that they can express the enzyme poly-β-D-mannuronate lyase (alginase) under control of the drug tetracycline to cause microcapsule digestion and their own cued release. NSCs are also genetically modified to inducibly express the enzyme IDS, so that they can provide adjacent cells with the lacking enzyme in the patient brain. Expression levels of both genes are controllable through dosed oral administration of the corresponding transgene activators. Pre-clinical testing in an animal model of Hunter Syndrome, the C57BL/6 IDS knock-out mouse, will follow.