Selection of genetically modified cell population using hapten-specific antibody/receptor chimera.
Efficient selection of the genetically modified cell population is a critical step to obtain the cells with desired properties. In this study, we propose an antigen-mediated genetically modified cell amplification (AMEGA) system employing an antibody/receptor chimera that triggers a growth signal in response to a non-toxic hapten dimer. An anti-fluorescein single-chain Fv fused to the extracellular D2 domain of erythropoietin receptor and transmembrane/intracellular domains of gp130 was expressed together with a model transgene, enhanced green fluorescent protein (EGFP) downstream of IRES sequence, by retroviral infection to IL-3-dependent Ba/F3 cells. Addition of fluorescein dimers connected by various oligo-DNA linkers induced selective growth of transfectants, thus leading to efficient expansion of EGFP-positive cell population. Also, digestion of the oligonucleotides by specific restriction endonuclease completely suppressed cell growth. Because these hapten dimers are not harmful for normal cells, the approach will be especially useful for reversible in vitro or in vivo expansion of genetically modified cell population employed for cell therapy and tissue engineering.
Kawahara
Masahiro
M
Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan. kawahara@bio.t.u-tokyo.ac.jp
Kimura
Hiroko
H
Ueda
Hiroshi
H
Nagamune
Teruyuki
T
D000818:Animals; D000906:Antibodies / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D000918:Antibody Specificity; D000941:Antigens / Q000276:immunology; D001483:Base Sequence; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D015245:Deoxyribonuclease BamHI / Q000378:metabolism; D019281:Dimerization; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins; D006241:Haptens / Q000737:chemistry / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000276:immunology; D008164:Luminescent Proteins / Q000737:chemistry / Q000235:genetics; D051379:Mice; D009841:Oligonucleotides / Q000737:chemistry; D017467:Receptors, Erythropoietin / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D014162:Transfection / Q000379:methods*; D019076:Transgenes
D000906:Antibodies; D000941:Antigens; D006241:Haptens; D007135:Immunoglobulin Variable Region; D008164:Luminescent Proteins; D009841:Oligonucleotides; D017467:Receptors, Erythropoietin; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D015245:Deoxyribonuclease BamHI