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An important advantage of the microencapsulated cell approach is that the therapeutic protein can reach therapeutic concentrations locally in the affected tissue, while maintaining a low systemic concentration, thus avoiding systemic side effects, such as systemic immunosuppression. Production of protein therapeutics by implanted microencapsulated cells also eliminates the need for repeated invasive administrations of therapeutics, which are typically applied by injections. Repeated application of therapeutics into the liver, brain or other complex tissues, where local delivery could be beneficial, is quite difficult. The usual obvious solution would be gene therapy, which has, despite its early promises, not delivered many successful clinical applications because of the problems of viral delivery and variable effectiveness caused by the random site of integration. We therefore aimed to develop an implantable cellular device for the local, regulated and safe delivery of therapeutic proteins. Long term survival of encapsulated cells beyond months may however be problematic with current technology and their efficiency of therapeutic production may decrease, although efficient therapy of diabetes and hemophilia using microencapsulated cells has been demonstrated in animals for more than 6months. Nevertheless we concluded that we should focus at least initially on the therapy of acute conditions.

The existing genetic switches used in mammalian cells are mainly based on the available prokaryotic transcriptional regulators, whose number is currently limited and whose properties may differ, hindering balanced switching. Our idea was to investigate the development of orthogonal genetic switches based on designed DNA-binding domains, such as TAL effectors that could support simultaneous introduction of several toggle switches into mammalian cells. Although designed transcription factors have been created based on modular DNA binding proteins, such as zinc fingers and TAL effectors, to our best knowledge there have been no reports of genetic switches based on these types of elements. These devices would be extremely valuable since we could produce numerous orthogonal switches allowing development of complex control functions and therapeutic devices.

Safety mechanisms for therapy with microencapsulated cells

For the safe therapeutic implementation of microencapsulated engineered cells in the organism we envisioned that we should be able to cease the therapy without having to physically remove the capsules. We therefore designed safety mechanisms that allow a complete removal of all exogenous material after the therapy has been completed.

Figure 3. Safety mechanisms that we planned to introduce for the application of microencapsulated engineered mammalian cells in therapy.

Microcapsule degradation - Most of the microcapsules used in literature and therapy are composed of polymerized alginate that is biocompatible but cannot be degraded in mammalian tissues. Alginate lyase harvested from a marine microorganism effectively degrades alginate, so secretion of this enzyme from cells should degrade the capsules and allow their resorption by phagocytic cells.

Termination - We should be able to inactivate the implanted cells at the end of the therapy or at any other stage if the therapy has to be terminated, regardless of the cause. We introduced a safety mechanism that would enable controlled induction of apoptosis of the implanted cells that is leak-free until the selected moment. Therefore we could inactivate therapeutic cells and enable their resorption without causing inflammation.

Escape tag – This ensures that any cell that may have escaped from a microcapsule is eliminated by the immune system of the host organism. The escaped cells would most likely be recognized as foreign in any case but in order to provide maximal safety measures we decided to introduce an additional tag that would alert the natural killer cells of the host immune response.

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