Development of novel chimeric transmembrane proteins for multimodality imaging of cancer cells.
Tracking the migration of cancer cells is essential to understanding the metastatic process. In order to facilitate the tracking of metastatic progression, we have generated transgenic cancer cell lines that express novel chimeric proteins composed of truncated human type II transmembrane proteins fused in-frame to a red fluorescence protein. These chimeric proteins have been engineered to display the fluorescence domain on the surface of cells. The three novel chimeric proteins exhibited high transient expression in several cancer cell lines. Membrane expression was well characterized in MCF-7 cell lines that stably express the chimeric proteins. Indirect immunocytochemistry of non-premeablized cells demonstrated co-localization of endogenous red and green fluorescence labeled secondary antibody bound on the cell surfaces. Immunoblots of total protein prepared from membrane, cytosolic and nuclear fractions indicated that the chimeric proteins were mainly associated with the membrane fraction. Further evidence of the membrane expression of these proteins was confirmed by confocal microscopy. Moreover, the chimeric protein was detected on the cell surface by T2-weighted magnetic resonance imaging using anti-red fluorescence protein antibody and superparamagnetic iron oxide particles in vitro and by optical imaging in vivo. The development of this non-mammalian cell surface marker, that can be detected by multi-modality imaging, will find utility in non-invasive longitudinal tracking of biological processes including metastatic progression, solid tumor treatment regimes and the fate of cells used in cell therapies such as islet or stem cells.
Department of Radiology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Paul T
PT
Winnard
Jessica B
JB
Kluth
Yoshinori
Y
Kato
Dmitri
D
Artemov
Venu
V
Raman
D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000032:analysis* / Q000235:genetics; D015415:Biomarkers; D045744:Cell Line, Tumor / Q000737:chemistry / Q000648:ultrastructure*; D004578:Electron Spin Resonance Spectroscopy; D053222:Fas Ligand Protein / Q000032:analysis* / Q000235:genetics; D019084:Fluorescent Antibody Technique, Indirect; D005456:Fluorescent Dyes / Q000032:analysis*; D006801:Humans; D008279:Magnetic Resonance Imaging; D008565:Membrane Proteins / Q000032:analysis* / Q000235:genetics; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000379:methods*; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000737:chemistry / Q000648:ultrastructure*; D015202:Protein Engineering; D011990:Receptors, Transferrin / Q000032:analysis* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000032:analysis* / Q000235:genetics; D023041:Xenograft Model Antitumor Assays / Q000379:methods
D015703:Antigens, CD; D015415:Biomarkers; C068126:CD71 antigen; D053222:Fas Ligand Protein; D005456:Fluorescent Dyes; D008565:Membrane Proteins; D011990:Receptors, Transferrin; D011993:Recombinant Fusion Proteins