Inactivation of encapsulated cells and their therapeutic effects by means of TGL triple-fusion reporter/biosafety gene

• Published in: Biomaterials 2013-01, Vol.34 (4), p.1442-1451
• Main Authors: Santos, Edorta, Larzabal, Leyre, Calvo, Alfonso, Orive, Gorka, Pedraz, José Luis, Hernández, Rosa M
• Format: Article
• Language: English
• Subjects: Advanced Basic Science; Animals; Biomedical materials; Biosafety; Cell Line; Cell Survival - drug effects; Dentistry; Drug delivery; Encapsulation; Ganciclovir - therapeutic use; Genes; Genes, Reporter; Green Fluorescent Proteins; Herpes simplex virus 1; Herpesvirus 1, Human - enzymology; HSV-TK; Inactivation; Luciferases, Firefly; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Microcapsule; Myoblasts - drug effects; Myoblasts - physiology; Recombinant Fusion Proteins; Searching; Secretions; Strategy; Surgical implants; TGL; Thymidine Kinase; Treatment Outcome
• ISSN: 0142-9612; 1878-5905
• DOI: 10.1016/j.biomaterials.2012.10.076

Abstract The immobilization of cells within alginate-poly- l -lysine-alginate (APA) microcapsules has been demonstrated to be an effective technology design for long term delivery of therapeutic products. Despite promising advances, biosafety aspects still remain to be improved. Here, we describe a complete characterization of the strategy based on TGL triple-fusion reporter gene – which codifies for Herpes Simplex virus type 1 thymidine-kinase (HSV1-TK), green fluorescent protein (GFP) and Firefly Luciferase – (SFGNES TGL) to inactivate encapsulated cells and their therapeutic effects. Myoblasts genetically engineered to secrete erythropoietin (EPO) were retroviraly transduced with the SFGNES TGL plasmid to further characterize their ganciclovir (GCV)-mediated inactivation process. GCV sensitivity of encapsulated cells was 100-fold lower when compared to cells plated onto 2D surfaces. However, the number of cells per capsule and EPO secretion decayed to less than 15% at the same time that proliferation was arrested after 14 days of GCV treatment in vitro. In vivo, ten days of GCV treatment was enough to restore the increased hematocrit levels of mice implanted with encapsulated TGL-expressing and EPO-secreting cells. Altogether, these results show that TGL triple-fusion reporter gene may be a good starting point in the search of a suitable biosafety strategy to inactivate encapsulated cells and control their therapeutic effects.