Homodimeric peptide radiotracer [⁶⁸Ga]Ga-NOTA-(TMVP1)₂ for VEGFR-3 imaging of cervical cancer patients.

Abstract

Vascular endothelial growth factor receptor 3 (VEGFR-3) plays a critical role in tumor lymphangiogenesis and metastasis, holding promise as a promising therapeutic target for solid tumors. TMVP1 (LARGR) is a 5-amino acid peptide previously identified in our laboratory from bacterial peptide display system that specifically targets VEGFR-3. Radiolabeled TMVP1 can be used for non-invasive imaging of VEGFR-3 expressing tumors. Homodimeric peptides have better targeting ability than monomeric peptides, and it is worth exploring whether homodimers of TMVP1 ((TMVP1)₂) can achieve better imaging effects. This study aimed to explore the peptide properties and tumor assessment value of [⁶⁸Ga]Ga-labeled (TMVP1)₂. In this study, we developed a TMVP1 homodimer that was conjugated with 1,4,7-triazacyclononane-N, N', N″-triacetic acid (NOTA) via tetraethyleneglycol (PEG₄) and triglyicine (Gly₃) spacer, and labeled with ⁶⁸Ga, to construct [⁶⁸Ga]Ga-NOTA-(TMVP1)₂. Binding of VEGFR-3 by TMVP1 and (TMVP1)₂, respectively, was modeled by molecular docking. The affinity of [⁶⁸Ga]Ga-NOTA-(TMVP1)₂ for VEGFR-3 and its ability to bind to cells were evaluated. MicroPET imaging and biodistribution studies of [⁶⁸Ga]Ga-NOTA-(TMVP1)₂ were performed in subcutaneous C33A cervical cancer xenografts. Five healthy volunteers and eight patients with cervical cancer underwent whole-body PET/CT acquisition 30-45 min after intravenous injection of [⁶⁸Ga]Ga-NOTA-(TMVP1)₂. Both molecular docking and cellular experiments showed that homodimeric TMVP1 had a higher affinity for VEGFR-3 than monomeric TMVP1. [⁶⁸Ga]Ga-NOTA-(TMVP1)₂ was excreted mainly through the renal route and partly through the liver route. In mice bearing C33A xenografts, [⁶⁸Ga]Ga-NOTA-(TMVP1)₂ specifically localized in the tumor (2.32 ± 0.10% ID/g). Pretreatment of C33A xenograft mice with the unlabeled peptide NOTA-(TMVP1)₂ reduced the enrichment of [⁶⁸Ga]Ga-NOTA-(TMVP1)₂ in tumors (0.58 ± 0.01% ID/g). [⁶⁸Ga]Ga-NOTA-(TMVP1)₂ proved to be safe in all healthy volunteers and recruited patients, with no side effects or allergies noted. In cervical cancer patients, a majority of the [¹⁸F]-FDG identified lesions (18/22, 81.8%) showed moderate to high signal intensity on [⁶⁸Ga]Ga-NOTA-(TMVP1)_2. SUVmax and SUVmean were 2.32 ± 0.77 and 1.61 ± 0.48, respectively. With normal muscle (gluteus maximus) as background, tumor-to-background ratios were 3.49 ± 1.32 and 3.95 ± 1.64 based on SUVmax and SUVmean, respectively. The favorable characterizations of [⁶⁸Ga]Ga-NOTA-(TMVP1)₂ such as convenient synthesis, high specific activity, and high tumor uptake enable the evaluation of VEGFR-3 in cervical cancer patients and warrant further clinical studies. ChiCTR-DOD-17012458. Registered August 23, 2017 (retrospectively registered).