title: Isolation and Characterization of a Monobody with a Fibronectin Domain III Scaffold That Specifically Binds EphA2.
abstract: Monobodies are binding scaffold proteins originating from a human fibronectin domain III (Fn3) scaffold that can be easily engineered with specificity and affinity. Human EphA2 (hEphA2) is an early detection marker protein for various tumors including lung, breast, and colon cancer. In this study, we isolated two hEphA2-specific monobodies (E1 and E10) by screening a yeast surface display library. They showed the same amino acid sequence except in the DE loop and had high affinity (~2 nM Kd) against hEphA2. E1 bound only hEphA2 and mEphA2, although it bound hEphA2 with an affinity 2-fold higher than that of mEphA2. However, E10 also bound the mEphA6 and mEphA8 homologs as well as hEphA2 and mEphA2. Thus, E1 but not E10 was highly specific for hEphA2. E1 specifically bound human cells and xenograft tumor tissues expressing hEphA on the cell surface. In vivo optical imaging showed strong targeting of Cy5.5-labeled E1 to mouse tumor tissue induced by PC3 cells, a human prostate cancer cell line that expresses a high level of hEphA2. In conclusion, the highly specific monobody E1 is useful as a hEphA2 probe candidate for in vivo diagnosis and therapy.
authors.affiliation: Department of Nuclear Medicine, Chonnam National University Medical School, Gwangju, Republic of Korea.
authors.forename: Seung-Hwan
authors.initials: SH
authors.lastname: Park
authors.affiliation: Department of Microbiology, Chonnam National University Medical School, Gwangju, Republic of Korea.
authors.forename: Sukho
authors.initials: S
authors.lastname: Park
authors.affiliation: Department of Nuclear Medicine, Chonnam National University Medical School, Gwangju, Republic of Korea.
authors.forename: Dong-Yeon
authors.initials: DY
authors.lastname: Kim
authors.affiliation: Department of Nuclear Medicine, Chonnam National University Medical School, Gwangju, Republic of Korea.
authors.forename: Ayoung
authors.initials: A
authors.lastname: Pyo
authors.affiliation: Molecular Imaging Program at Stanford, Department of Radiology, Bio-X Program, Stanford University, Palo Alto, CA, United States of America.
authors.forename: Richard H
authors.initials: RH
authors.lastname: Kimura
authors.affiliation: Molecular Imaging Program at Stanford, Department of Radiology, Bio-X Program, Stanford University, Palo Alto, CA, United States of America.
authors.forename: Ataya
authors.initials: A
authors.lastname: Sathirachinda
authors.affiliation: Department of Microbiology, Chonnam National University Medical School, Gwangju, Republic of Korea.
authors.forename: Hyon E
authors.initials: HE
authors.lastname: Choy
authors.affiliation: Department of Nuclear Medicine, Chonnam National University Medical School, Gwangju, Republic of Korea.
authors.forename: Jung-Joon
authors.initials: JJ
authors.lastname: Min
authors.affiliation: Molecular Imaging Program at Stanford, Department of Radiology, Bio-X Program, Stanford University, Palo Alto, CA, United States of America.
authors.forename: Sanjiv Sam
authors.initials: SS
authors.lastname: Gambhir
authors.affiliation: Department of Microbiology, Chonnam National University Medical School, Gwangju, Republic of Korea.
authors.forename: Yeongjin
authors.initials: Y
authors.lastname: Hong
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D002478:Cells, Cultured; D005353:Fibronectins / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D009369:Neoplasms / Q000175:diagnosis* / Q000378:metabolism / Q000473:pathology; D019151:Peptide Library; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D036104:Receptor, EphA2 / Q000378:metabolism*; D012441:Saccharomyces cerevisiae; D013379:Substrate Specificity; D023041:Xenograft Model Antitumor Assays
chemical_list: D000906:Antibodies; D005353:Fibronectins; D019151:Peptide Library; D036104:Receptor, EphA2