title: Heterologous protein production capacity of mammalian cells cultivated as monolayers and microtissues.
abstract: A precise understanding of processes managing heterologous protein production in vitro and in vivo is essential for the manufacture of sophisticated biopharmaceuticals as well as for future gene therapy and tissue engineering initiatives. Capitalizing on the gravity-enforced self-assembly of monodispersed cells into coherent (multicellular) microtissues we studied heterologous protein production of microtissues and monolayers derived from cell lines and primary cells engineered/transduced for (i) constitutive, (ii) proliferation-controlled, (iii) macrolide-, or (iv) gas-inducible expression of the human placental secreted alkaline phosphatase (SEAP) and of the Bacillus stearothermophilus-derived secreted alpha-amylase (SAMY). Specific productivity of cells assembled in microtissues was up to 20-fold higher than isogenic monolayer cultures. Diffusion across microtissues could be further increased by HUVEC-mediated vascularization. As well as higher specific protein productivities, microtissues were also more efficient than monolayer cultures in assembling transgenic lentiviral particles. Our results showed that mammalian cells embedded in a tissue-like three-dimensional (3D) microenvironment exhibit increased production capacity. This observation should be considered for gene therapy and tissue engineering scenarios as well as for biopharmaceutical manufacturing.
authors.affiliation: Institute for Chemical and Bio-Engineering, ICB, Swiss Federal Institute of Technology, ETH Hoenggerberg, HCI F115, Wolfgang-Pauli-Strasse 10, CH-8093 Zurich, Switzerland.
authors.forename: Carlota Diaz
authors.initials: CD
authors.lastname: Sanchez-Bustamante
authors.forename: Jens M
authors.initials: JM
authors.lastname: Kelm
authors.forename: Barbara
authors.initials: B
authors.lastname: Mitta
authors.forename: Martin
authors.initials: M
authors.lastname: Fussenegger
mesh_terms: D000469:Alkaline Phosphatase / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D018929:Cell Culture Techniques; D002460:Cell Line; D006224:Cricetinae; D005260:Female; D001411:Geobacillus stearothermophilus / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D011257:Pregnancy Proteins / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D046509:Tissue Culture Techniques
chemical_list: D001426:Bacterial Proteins; D011257:Pregnancy Proteins; D011994:Recombinant Proteins; D000469:Alkaline Phosphatase