Identification of a CEACAM5 targeted nanobody for positron emission tomography imaging and near-infrared fluorescence imaging of colorectal cancer

Yitai Xiao; Chaoming Mei; Duo Xu; Fan Yang; Meilin Yang; Lei Bi; Junjie Mao; Pengfei Pang; Dan Li

doi:10.1007/s00259-023-06183-7

Identification of a CEACAM5 targeted nanobody for positron emission tomography imaging and near-infrared fluorescence imaging of colorectal cancer

Eur J Nucl Med Mol Imaging. 2023 Jul;50(8):2305-2318. doi: 10.1007/s00259-023-06183-7. Epub 2023 Mar 14.

Authors

Yitai Xiao^#^{1

2}, Chaoming Mei^#^{1

2}, Duo Xu^{1

2}, Fan Yang^{1

2}, Meilin Yang^{1

2}, Lei Bi², Junjie Mao³, Pengfei Pang⁴, Dan Li^{5

6}

Affiliations

¹ Department of Nuclear Medicine, The Fifth Affiliated Hospital, Sun Yat-Sen University, Guangdong Province, 519000, China.
² Guangdong Provincial Key Laboratory of Biomedical Imaging and Guangdong Provincial Engineering Research Center of Molecular Imaging, The Fifth Affiliated Hospital, Sun Yat-Sen University, Guangdong Province, 519000, China.
³ Center for Interventional Medicine, The Fifth Affiliated Hospital, Sun Yat-Sen University, Guangdong Province, 519000, China. maojunj@mail.sysu.edu.cn.
⁴ Center for Interventional Medicine, The Fifth Affiliated Hospital, Sun Yat-Sen University, Guangdong Province, 519000, China. pangpf@mail.sysu.edu.cn.
⁵ Department of Nuclear Medicine, The Fifth Affiliated Hospital, Sun Yat-Sen University, Guangdong Province, 519000, China. lidan25@mail.sysu.edu.cn.
⁶ Guangdong Provincial Key Laboratory of Biomedical Imaging and Guangdong Provincial Engineering Research Center of Molecular Imaging, The Fifth Affiliated Hospital, Sun Yat-Sen University, Guangdong Province, 519000, China. lidan25@mail.sysu.edu.cn.

^# Contributed equally.

PMID: 36914753
DOI: 10.1007/s00259-023-06183-7

Abstract

Purpose: Here, we aim to identify a CEACAM5-targeted nanobody and demonstrate its application in positron emission tomography (PET) imaging and near-infrared (NIR) fluorescence imaging in colorectal cancer (CRC).

Methods: Immunohistochemistry was applied to verify CEACAM5 expression in CRC and metastatic lymph nodes (mLNs). CEACAM5-targeted nanobodies were obtained by immunization of human CEACAM5 protein in a dromedary, followed by several rounds of phage screenings. Immunofluorescence staining and flow cytometry was carried out to determine the binding affinity of the nanobodies. The nanobodies were radiolabeled by coupling ¹⁸F-SFB for PET imaging of CRC subcutaneous xenografts and lymph node metastasis (LNM). IRDye800CW (IR800) were conjugated to form NIR probes for NIR imaging in CRC subcutaneous models.

Results: CEACAM5 was overexpressed in either human CRC tissues or mLNs. A CEACAM5 targeted nanobody, Nb41 was successfully generated, with excellent in vitro binding properties. Incorporation of albumin binding domain (ABD) did not affect the affinity of Nb41. In vivo imaging showed that both ¹⁸F-FB-Nb41 and ¹⁸F-FB-Nb41-ABD showed obvious accumulation in the tumor. Due to the longer retention in the blood, ¹⁸F-FB-Nb41-ABD enrichment in tumors was significantly delayed but higher compared to ¹⁸F-FB-Nb41. Both ¹⁸F-FB-Nb41 and ¹⁸F-FB-Nb41-ABD showed prominent LNM enrichment. Similarly, the IR800-conjugated nanobodies Nb41-IR800 and Nb41-ABD-IR800 exhibited superior imaging effects in subcutaneous models, while Nb41-ABD-IR800 exhibited higher fluorescence intensity in the tumor accompanied with a remarkedly delay compared to Nb41-IR800.

Conclusion: Collectively, we presented the identification and in vivo validation of a CEACAM5-targeted nanobody and a fused nanobody with an ABD, which enabled to the non-invasive visualization of malignancy of CRC using PET imaging and NIR imaging in subcutaneous models as well as LNM models.

Keywords: CEACAM5; Colorectal cancer; Nanobody; Near-infrared fluorescence imaging; Positron emission tomography imaging.

MeSH terms

Carcinoembryonic Antigen
Cell Line, Tumor
Colorectal Neoplasms* / diagnostic imaging
GPI-Linked Proteins
Humans
Optical Imaging
Positron-Emission Tomography
Single-Domain Antibodies* / metabolism

Substances

Single-Domain Antibodies
CEACAM5 protein, human
Carcinoembryonic Antigen
GPI-Linked Proteins

Grants and funding

2018B030322006/The Department of Science and Technology of Guangdong Province to the Guangdong Provincial Key Laboratory of Biomedical Imaging