Fabrication of DNA-antibody–apatite composite layers for cell-targeted gene transfer

Abstract

Surface-mediated gene transfer systems using apatite (Ap)-based composite layers have received increased attention in tissue engineering applications owing to their safety, biocompatibility and relatively high efficiency. In this study, DNA-antibody-apatite composite layers (DA-Ap layers), in which DNA and antibody molecules are immobilized within a matrix of apatite nanocrystals, were fabricated using a biomimetic coating process. They were then assayed for their gene transfer capability for application in a specific cell-targeted gene transfer. A DA-Ap layer that was fabricated with an anti-CD49f antibody showed a higher gene transfer capability to the CD49f-positive CHO-K1 cells than a DNA-apatite composite layer (D-Ap layer). The antibody facilitated the gene transfer capability of the DA-Ap layer only to the specific cells that were expressing corresponding antigens. When the DA-Ap layer was fabricated with an anti-N-cadherin antibody, a higher gene transfer capability compared with the D-Ap layer was found in the N-cadherin-positive P19CL6 cells, but not in the N-cadherin-negative UV♀2 cells or in the P19CL6 cells that were pre-blocked with anti-N-cadherin. Therefore, the antigen-antibody binding that takes place at the cell-layer interface should be responsible for the higher gene transfer capability of the DA-Ap than D-Ap layer. These results suggest that the DA-Ap layer works as a mediator in a specific cell-targeted gene transfer system.

Delivery Systems for Nucleic Acids and Proteins: Barriers, Cell Capture Pathways and Nanocarriers

“…Yazaki et al [ 156 ] co-immobilized DNA and antibodies within a hydroxyapatite matrix and assessed the obtained materials’ cell-specific gene transfer. They observed the transfection of CD49f-positive CHO-K1 cells in the presence of an anti-CD49f antibody.…”

Overview of Safety and Efficacy of Non-Viral Gene Transfer in Cartilage Tissue Engineering from the Worldview of Islam

“…After conducting countless numbers of gene transfer studies, researchers identified the pros and cons of both viral and non-viral approaches. The latter is favoured by some researchers because of several advantages [18][19][20] as this approach somehow triggers low immune response compared with viral approach which makes it suitable for human therapy. 21 It was also observed that the non-viral approach caused low toxicity to cells.…”

Spontaneous assembly of DNA–amorphous calcium phosphate nanocomposite spheres for surface-mediated gene transfer

“…Over the past decade, various approaches to increase the gene transfer efficiency of the DNA-CaP nanocomposite layer have been proposed. 1,[4][5][6][7] Previous approaches were based on the co-immobilization of biofunctional molecules such as cell adhesion molecules (laminin, fibronectin), 4,5 lipids, 6 and antibodies 7 within the DNA-CaP nanocomposite layer to enhance cell-layer interactions. The co-immobilization of these biofunctional molecules increased the gene transfer efficiency of the nanocomposite layer by 1-4 orders of magnitude.…”

“…The co-immobilization of these biofunctional molecules increased the gene transfer efficiency of the nanocomposite layer by 1-4 orders of magnitude. 1,[4][5][6][7] Here we discovered another approach without biofunctional molecules that is based on kinetic control of CaP nucleation in the fabrication process of the DNA-CaP nanocomposite layer to increase its gene transfer efficiency. Nucleation of CaPs includes amorphous phase nucleation as well as crystalline phase nucleation.…”

“…We hypothesized that the micro-and crystal structure of the nanocomposite layer should be altered considerably by changing the nucleation kinetics of CaPs in the nanocomposite fabrication process. Previously, DNA-CaP nanocomposite layers were fabricated using metastable supersaturated CaP solutions (e.g., a so-called CP solution 1,4,5,7 ) that remain stable without inducing homogeneous nucleation for several days to weeks. In such metastable solutions, DNA-CaP nanocomposite layers were fabricated via heterogeneous CaP nucleation at the substrate surface followed by relatively faster crystallization (amorphous to crystalline phase) and/or CaP crystal growth 8 with simultaneous immobilization of DNA molecules on the crystal surface and interstices.…”