Screening, Construction, and Preliminary Evaluation of CLDN18.2-Specific Peptides for Noninvasive Molecular Imaging

Zilei Wang; Chuanke Zhao; Jin Ding; Yan Chen; Jiayue Liu; Xingguo Hou; XiangXing Kong; Bin Dong; Zhi Yang; Hua Zhu

doi:10.1021/acsptsci.3c00165

Screening, Construction, and Preliminary Evaluation of CLDN18.2-Specific Peptides for Noninvasive Molecular Imaging

ACS Pharmacol Transl Sci. 2023 Nov 21;6(12):1829-1840. doi: 10.1021/acsptsci.3c00165. eCollection 2023 Dec 8.

Authors

Zilei Wang¹, Chuanke Zhao², Jin Ding¹, Yan Chen¹, Jiayue Liu¹, Xingguo Hou¹, XiangXing Kong¹, Bin Dong³, Zhi Yang¹, Hua Zhu¹

Affiliations

¹ State Key Laboratory of Holistic Integrative Management of Gastrointestinal Cancers, Beijing Key Laboratory of Carcinogenesis and Translational Research, NMPA Key Laboratory for Research and Evaluation of Radiopharmaceuticals (National Medical Products Administration), Department of Nuclear Medicine, Peking University Cancer Hospital & Institute, Beijing 100142, China.
² Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Biochemistry and Molecular Biology, Peking University Cancer Hospital & Institute, Beijing 100142, China.
³ Central Laboratory, Peking University Cancer Hospital & Institute, Beijing 100142, China.

PMID: 38093841
PMCID: PMC10714438 (available on 2024-11-21)
DOI: 10.1021/acsptsci.3c00165

Abstract

Recent global clinical trials have shown that CLDN18.2 is an ideal target for the treatment of gastric cancer and that patients with high CLDN18.2 expression can benefit from targeted therapy. Therefore, accurate and comprehensive detection of CLDN18.2 expression is important for patient screening and guidance in anti-CLDN18.2 therapy. Phage display technology was used to screen CLDN18.2-specific peptides from 100 billion libraries. 293T^CLDN18.1 cells were used to exclude nonspecific binding and CLDN18.1 binding sequences, while 293T^CLDN18.2 cells were used to screen CLDN18.2-specific binding peptides. The monoclonal clones obtained from phage screening were sequenced, and peptides were synthesized based on the sequencing results. Binding specificity and affinity were assessed with a fluorescein isothiocyanate (FITC)-conjugated peptide. A 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-conjugated peptide was also synthesized for ⁶⁸Ga radiolabeling. The in vitro and in vivo stability, partition coefficients, in vivo molecular imaging, and biodistribution were also characterized. Overall, 54 monoclonal clones were selected after phage display screening. Subsequently, based on the cell ELISA results, CLDN18.2 preference monoclonal clones were selected for deoxyribonucleic acid (DNA) sequencing, and four 7-peptide sequences were obtained after sequence comparison; among them, a peptide named T37 was further validated in vitro and in vivo. The T37 peptide specifically recognized CLDN18.2 but not CLDN18.1 and bound strongly to CLDN18.2-positive cell membranes. The ⁶⁸Ga-DOTA-T37 probe exhibits good in vitro properties and high stability as a hydrophilic probe; it has high biological safety, and positron emission tomography/computed tomography (PET/CT) studies have shown that it can specifically target CLDN18.2 protein and CLDN18.2-positive tumors in mice. ⁶⁸Ga-DOTA-T37 demonstrated the superiority and feasibility of using a CLDN18.2-specific probe in PCT/CT imaging, which deserves further development and exploitation.