Kannika Khantasup1,2, Pairash Saiviroonporn3, Suwatchai Jarussophon4, Warangkana Chantima5, Tararaj Dharakul6,7. 1. Research Division, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand. 2. NANOTEC-Mahidol University Center of Excellence in Nanotechnology for Cancer Diagnosis and Treatment, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand. 3. Department of Radiology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand. 4. National Nanotechnology Center, National Science and Technology Development Agency, Pathum Thani, Thailand. 5. Graduate Program in Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand. 6. NANOTEC-Mahidol University Center of Excellence in Nanotechnology for Cancer Diagnosis and Treatment, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand. tararaj.dha@mahidol.ac.th. 7. Department of Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 17000, Thailand. tararaj.dha@mahidol.ac.th.
Abstract
OBJECTIVES: The development of targeted contrast agents for magnetic resonance imaging (MRI) facilitates enhanced cancer imaging and more accurate diagnosis. In the present study, a novel contrast agent was developed by conjugating anti-EpCAM humanized scFv with gadolinium chelate to achieve target specificity. MATERIALS AND METHODS: The material design strategy involved site-specific conjugation of the chelating agent to scFv. The scFv monomer was linked to maleimide-DTPA via unpaired cysteine at the scFv C-terminus, followed by chelation with gadolinium (Gd). Successful scFv-DTPA conjugation was achieved at 1:10 molar ratio of scFv to maleimide-DTPA at pH 6.5. The developed anti-EpCAM-Gd-DTPA MRI contrast agent was evaluated for cell targeting ability, in vitro serum stability, cell cytotoxicity, relaxivity, and MR contrast enhancement. RESULTS: A high level of targeting efficacy of anti-EpCAM-Gd-DTPA to an EpCAM-overexpressing HT29 colorectal cell was demonstrated by confocal microscopy. Good stability of the contrast agent was obtained and no cytotoxicity was observed in HT29 cells after 48 h incubation with 25-100 µM of Gd. Favorable imaging was obtained using anti-EpCAM-Gd-DTPA, including 1.8-fold enhanced relaxivity compared with Gd-DTPA, and MR contrast enhancement observed after binding to HT29. CONCLUSION: The potential benefit of this contrast agent for in vivo MR imaging of colorectal cancer, as well as other EpCAM positive cancers, is suggested and warrants further investigation.
OBJECTIVES: The development of targeted contrast agents for magnetic resonance imaging (MRI) facilitates enhanced cancer imaging and more accurate diagnosis. In the present study, a novel contrast agent was developed by conjugating anti-EpCAM humanized scFv with gadolinium chelate to achieve target specificity. MATERIALS AND METHODS: The material design strategy involved site-specific conjugation of the chelating agent to scFv. The scFv monomer was linked to maleimide-DTPA via unpaired cysteine at the scFv C-terminus, followed by chelation with gadolinium (Gd). Successful scFv-DTPA conjugation was achieved at 1:10 molar ratio of scFv to maleimide-DTPA at pH 6.5. The developed anti-EpCAM-Gd-DTPA MRI contrast agent was evaluated for cell targeting ability, in vitro serum stability, cell cytotoxicity, relaxivity, and MR contrast enhancement. RESULTS: A high level of targeting efficacy of anti-EpCAM-Gd-DTPA to an EpCAM-overexpressing HT29 colorectal cell was demonstrated by confocal microscopy. Good stability of the contrast agent was obtained and no cytotoxicity was observed in HT29 cells after 48 h incubation with 25-100 µM of Gd. Favorable imaging was obtained using anti-EpCAM-Gd-DTPA, including 1.8-fold enhanced relaxivity compared with Gd-DTPA, and MR contrast enhancement observed after binding to HT29. CONCLUSION: The potential benefit of this contrast agent for in vivo MR imaging of colorectal cancer, as well as other EpCAM positive cancers, is suggested and warrants further investigation.
Entities:
Keywords:
Gadolinium-diethylenetriamine pentaacetic acid (Gd-DTPA); Human epithelial cell adhesion molecule (EpCAM); Humanized scFv; Magnetic resonance imaging (MRI)
Authors: Elyse D Bernard; Michael A Beking; Karunanithi Rajamanickam; Eve C Tsai; Maria C Derosa Journal: J Biol Inorg Chem Date: 2012-08-19 Impact factor: 3.358