PLoS ONE (Jan 2017)

Engineering of monobody conjugates for human EphA2-specific optical imaging.

  • Min-A Kim,
  • Hee Seung Yoon,
  • Seung-Hwan Park,
  • Dong-Yeon Kim,
  • Ayoung Pyo,
  • Hyeon Sik Kim,
  • Jung-Joon Min,
  • Yeongjin Hong

DOI
https://doi.org/10.1371/journal.pone.0180786
Journal volume & issue
Vol. 12, no. 7
p. e0180786

Abstract

Read online

In a previous study, we developed an E1 monobody specific for the tumor biomarker hEphA2 [PLoS ONE (2015) 10(7): e0132976]. E1 showed potential as a molecular probe for in vitro and in vivo targeting of cancers overexpressing hEphA2. In the present study, we constructed expression vectors for E1 conjugated to optical reporters such as Renilla luciferase variant 8 (Rluc8) or enhanced green fluorescent protein (EGFP) and purified such recombinant proteins by affinity chromatography in E. coli. E1-Rluc8 and E1-EGFP specifically bound to hEphA2 in human prostate cancer PC3 cells but not in human cervical cancer HeLa cells, which express hEphA2 at high and low levels, respectively. These recombinant proteins maintained >40% activity in mouse serum at 24 h. In vivo optical imaging for 24 h did not detect E1-EGFP signals, whereas E1-Rluc8 showed tumor-specific luminescence signals in PC3 but not in HeLa xenograft mice. E1-Rluc8 signals were detected at 4 h, peaked at 12 h, and were undetectable at 24 h. These results suggest the potential of E1-Rluc8 as an EphA2-specific optical imaging agent.