Cucurbituril–Ferrocene: Host–Guest Based Pretargeted Positron Emission Tomography in a Xenograft Model

Abstract

Pretargeted positron emission tomography is a macromolecule-driven nuclear medicine technique that involves targeting a preadministered antigen target-bound macromolecule with a radioligand in vivo, aiming to minimize the overall radiation dose. This study investigates the use of antibody based host−guest chemistry methodology for pretargeted positron emission tomography. We hypothesize that the novel pretargeting approach reported here overcomes the challenges the current pretargeting platforms have with the in vivo stability and modularity of the pretargeting components. A cucurbit [7]uril host molecule modified, anti-carcinoembryonic antigen antibody (M5A; CB7-M5A) and a 68 Ga-radiolabeled ferrocene guest radioligand ([ 68 Ga]Ga-NOTA-PEG 3 -NMe 2 -Fc) were studied as potential host−guest chemistry pretargeting agents for positron emission tomography in BxPC3 xenografted nude mice. The viability of the platform was studied via in vivo biodistribution and positron emission tomography. Tumor uptake of [ 68 Ga]Ga-NOTA-PEG 3 -NMe 2 -Fc was significantly higher in mice which received CB7-M5A prior to the radioligand injection (pretargeted) (3.3 ± 0.7%ID/g) compared to mice which only received the radioligand (nonpretargeted) (0.2 ± 0.1%ID/g).

Investigation of Copper-64-Based Host–Guest Chemistry Pretargeted Positron Emission Tomography

“…These studies expand upon our previous work which first reported CB7–Fc, host–guest chemistry-based pretargeting platform . That study explored CB7–Fc pretargeting using a gallium-68 radiolabeled Fc-radioligand, [ 68 Ga]­Ga-NOTA-PEG 3 -NMe 2 -Fc.…”

“…As previously reported, each M5A antibody molecule was determined to have 0.8 ± 0.0 CB7 moieties on average. Additionally, the immunoreactivity of CB7-M5A was studied to be 95.7 ± 0.7%.…”

“…The primary pretargeting agent, CB7-M5A, was synthesized and characterized as previously reported by our laboratory . In short, the antibody (3.0 mg; 20 nmol; 1 equiv) was first modified with dibenzocyclooctyne (DBCO) using DBCO NHS-ester (0.017 mg, 39 nmol, 2 equiv) in PBS (2 mL, pH 8.7) over 60 min at room temperature.…”

“…[ 64 Cu]­Cu-NOTA-PEG 3 -Fc ( 1 ) and [ 64 Cu]­Cu-NOTA-PEG 7 -Fc ( 2 ) (Figure A) were synthesized using NOTA-PEG 3 -Fc ( 4 ) and NOTA-PEG 7 -Fc ( 3 ) as the radiolabeling precursors. 4 was synthesized following the synthesis scheme reported previously by our laboratory Figure S1): Ferrocenecarboxaldehyde (133 mg; 0.621 mmol) and t -Boc- N -amido-PEG7-amine (73 mg; 0.156 mmol) were dissolved in methanol (40 mL) followed by addition of triethylamine (7.26 mg; 0.0717 mmol).…”

“…The number of CB7-moieties per M5A mAb was studied by incubating the CB7-modified mAb with high excess of adamantane-fluorescein CB7 guest molecule (FL-Adma), purifying the FL-Adma-CB7-M5A and measuring its degree of labeling with a UV/Vis spectrophotometer. The characterization of FL-Adma and the method used to study the number of CB7 moieties per mAb have been previously reported by our laboratory …”