Cy5.5-labeled Affibody molecule for near-infrared fluorescent optical imaging of epidermal growth factor receptor positive tumors.
Affibody protein is an engineered protein scaffold with a three-helical bundle structure. Affibody molecules of small size (7 kD) have great potential for targeting overexpressed cancer biomarkers in vivo. To develop an Affibody-based molecular probe for in vivo optical imaging of epidermal growth factor receptor (EGFR) positive tumors, an anti-EGFR Affibody molecule, Ac-Cys-Z(EGFR:1907) (7 kD), is site-specifically conjugated with a near-IR fluorescence dye, Cy5.5-mono-maleimide. Using fluorescent microscopy, the binding specificity of the probe Cy5.5-Z(EGFR:1907) is checked by a high-EGFR-expressing A431 cell and low-EGFR-expressing MCF7 cells. The binding affinity of Cy5.5-Z(EGFR:1907) (K(D)) to EGFR is 43.6+/-8.4 nM, as determined by flow cytometry. For an in vivo imaging study, the probe shows fast tumor targeting and good tumor contrast as early as 0.5 h postinjection (p.i.) for A431 tumors, while MCF7 tumors are barely visible. An ex vivo imaging study also demonstrates that Cy5.5-Z(EGFR:1907) has high tumor, liver, and kidney uptakes at 24 h p.i.. In conclusion, Cy5.5-Z(EGFR:1907) shows good affinity and high specificity to the EGFR. There is rapid achievement of good tumor-to-normal-tissue contrasts of Cy5.5-Z(EGFR:1907), thus demonstrating its potential for EGFR-targeted molecular imaging of cancers.
Miao
Zheng
Z
Stanford University, Stanford Cancer Center, Molecular Imaging Program at Stanford, Department of Radiology and Bio-X Program, 1201 Welch Road, Lucas Expansion, P095, Stanford, California 94305-5344, USA.
Ren
Gang
G
Liu
Hongguang
H
Jiang
Lei
L
Cheng
Zhen
Z
D000818:Animals; D000915:Antibody Affinity; D014408:Biomarkers, Tumor / Q000378:metabolism*; D002232:Carbocyanines / Q000737:chemistry* / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism*; D005260:Female; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009369:Neoplasms; D055095:Optics and Photonics / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence / Q000379:methods*; D019265:Spectroscopy, Near-Infrared / Q000379:methods*; D014183:Transplantation, Heterologous; D051598:Whole Body Imaging
D014408:Biomarkers, Tumor; C098793:CY5.5 cyanine dye; D002232:Carbocyanines; D011993:Recombinant Fusion Proteins; D066246:ErbB Receptors