Abstract
Previously, we successfully developed a pretargeted imaging strategy (atezolizumab-TCO/[99mTc]HYNIC-PEG11-Tz) for evaluating programmed cell death ligand-1 (PD-L1) expression in xenograft mice. However, the surplus unclicked [99mTc]HYNIC-PEG11-Tz is cleared somewhat sluggishly through the intestines, which is not ideal for colorectal cancer (CRC) imaging. To shift the excretion of the Tz-radioligand to the renal system, we developed a novel Tz-radioligand by adding a polypeptide linker between HYNIC and PEG11. Pretargeted molecular probes [99mTc]HYNIC-polypeptide-PEG11-Tz and cetuximab-TCO were synthesized. [99mTc]HYNIC-polypeptide-PEG11-Tz was evaluated for in vitro stability and in vivo blood pharmacokinetics. In vitro ligation reactivity of [99mTc]HYNIC-polypeptide-PEG11-Tz towards cetuximab-TCO was also tested. Biodistribution assay and imaging of [99mTc]HYNIC-polypeptide-PEG11-Tz were performed to observe its excretion pathway. Pretargeted biodistribution was measured at three different accumulation intervals to determine the optimal pretargeted interval time. Pretargeted (cetuximab-TCO 48h/[99mTc]HYNIC-PEG11-Tz 6h) and (cetuximab-TCO 48h/[99mTc]HYNIC-Polypeptide-PEG11-Tz 6h) imagings were compared to examine the effect of the excretion pathway on tumor imaging. [99mTc]HYNIC-polypeptide-PEG11-Tz showed favorable in vitro stability and rapid blood clearance in mice. SEC-HPLC revealed almost complete reaction between cetuximab-TCO and [99mTc]HYNIC-polypeptide-PEG11-Tz in vitro, with the 8:1 Tz-to-mAb reaction providing a conversion yield of 87.83 ± 3.27%. Biodistribution and imaging analyses showed that the Tz-radioligand was cleared through the kidneys. After 24, 48, and 72h of accumulation in HCT116 tumor, the tumor-to-blood ratio of cetuximab-TCO was 0.83 ± 0.13, 1.40 ± 0.31, and 1.15 ± 0.21, respectively. Both pretargeted (cetuximab-TCO 48h/[99mTc]HYNIC-PEG11-Tz 6h) and (cetuximab-TCO 48h/[99mTc]HYNIC-polypeptide-PEG11-Tz 6h) clearly delineated HCT116 tumor. Pretargeted imaging strategy using cetuximab-TCO/[99mTc]HYNIC-polypeptide-PEG11-Tz could be used for diagnosing CRC, as the surplus unclicked [99mTc]HYNIC-polypeptide-PEG11-Tz was cleared through the urinary system, leading to low abdominal uptake background. Our novel pretargeted imaging strategy (cetuximab-TCO/[99mTc]HYNIC-polypeptide-PEG11-Tz) was useful for imaging CRC, broadening the application scope of pretargeted imaging strategy. The pretargeted imaging strategy clearly delineated HCT116 tumor, showing that its use could be extended to selection of internalizing antibodies.
Highlights
Colorectal cancer (CRC) is a common malignant tumor of the gastrointestinal tract with its morbidity ranking 3th among all malignant tumors and its mortality ranking 4th in the world, which poses a serious threat to human health [1]
We developed a pretargeted single photon emission computed tomography (SPECT) imaging strategy for evaluating immune checkpoint ligand PD-L1 expression in tumor based on bioorthogonal Diels-Alder click chemistry [17]
After the reaction of purified Cetuximab-TCO with 20-fold molar equivalents of HYNIC-Polypeptide-PEG11-Tz was completed, the consumption of HYNIC-Polypeptide-PEG11-Tz was used for calculating the average number of TCO conjugated to each Cetuximab
Summary
Colorectal cancer (CRC) is a common malignant tumor of the gastrointestinal tract with its morbidity ranking 3th among all malignant tumors and its mortality ranking 4th in the world, which poses a serious threat to human health [1]. Diagnosis and treatment will greatly improve the prognosis of CRC, most of which could be cured in the early stage with a 5-year survival rate of 90%, while only 12.5% for advanced CRC [2] It is greatly significant for the diagnosis of CRC, especially for the early diagnosis, to research and develop the specific targeted molecular probe for CRC. The clinical diagnosis for CRC mainly relies on endoscopy and endoscopy-guided biopsy, imaging methods such as computed tomography (CT), magnetic resonance imaging (MRI), and 2-deoxy–2-[18F]fluoro-D-glucose (18F-FDG) positron emission tomography/computed tomography (PET/CT). It is still difficult to detect surrounding tissue invasion and distant metastasis and achieve targeted imaging of CRC for these endoscopic methods. The most common positron radioactive tracer, 18F-FDG, is still not a specific targeting molecular probe for CRC
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