68Ga-bisphosphonates for the imaging of extraosseous calcification by positron emission tomography

Abstract

Radiolabelled bisphosphonates (BPs) and [18F]NaF (18F-fluoride) are the two types of radiotracers available to image calcium mineral (e.g. bone), yet only [18F]NaF has been widely explored for the non-invasive molecular imaging of extraosseous calcification (EC) using positron emission tomography (PET) imaging. These two radiotracers bind calcium mineral deposits via different mechanisms, with BPs chelating to calcium ions and thus being non-selective, and [18F]NaF being selective for hydroxyapatite (HAp) which is the main component of bone mineral. Considering that the composition of EC has been reported to include a diverse range of non-HAp calcium minerals, we hypothesised that BPs may be more sensitive for imaging EC due to their ability to bind to both HAp and non-HAp deposits. We report a comparison between the 68Ga-labelled BP tracer [68Ga]Ga-THP-Pam and [18F]NaF for PET imaging in a rat model of EC that develops macro- and microcalcifications in several organs. Macrocalcifications were identified using preclinical computed tomography (CT) and microcalcifications were identified using µCT-based 3D X-ray histology (XRH) on isolated organs ex vivo. The morphological and mineral analysis of individual calcified deposits was performed using scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDX). PET imaging and ex vivo analysis results demonstrated that while both radiotracers behave similarly for bone imaging, the BP-based radiotracer [68Ga]Ga-THP-Pam was able to detect EC more sensitively in several organs in which the mineral composition departs from that of HAp. Our results strongly suggest that BP-based PET radiotracers such as [68Ga]Ga-THP-Pam may have a particular advantage for the sensitive imaging and early detection of EC by being able to detect a wider array of relevant calcium minerals in vivo than [18F]NaF, and should be evaluated clinically for this purpose.

[68Ga]Ga-THP-tetrazine for bioorthogonal click radiolabelling: pretargeted PET imaging of liposomal nanomedicines

“…The nature of this insoluble impurity that binds irreversibly to glass surfaces remains unknown despite our efforts to characterise it. It is worth noting that it has not been observed with other THP-based small molecule radiotracers with thiourea linkers, 29,31,32,34,35 strongly suggesting it is specific to this tetrazine conjugate. The purified [ 68 Ga]Ga-THP-Tz (RCY = 45 ± 5%, Apparent A m ≈ 2.2 GBq μmol −1 ) eluted in pure methanol was subsequently dried under nitrogen and resuspended in 5% DMSO/water used for further in vitro and in vivo validation.…”

“…30 The resulting thiourea linker has been successfully used in other THP-based radiotracers, and leads to stable compounds in vivo, based on several clinical and preclinical studies. 29,[31][32][33][34][35] The conjugation was performed at room temperature using DIPEA as a base to achieve a pH of 9-10 to keep the target amine on the 6-methyl tetrazine deprotonated. The mild conditions and aprotic solvent DMSO minimised the hydrolysis of THP-NCS to amine (m/z = 460) and degradation of tetrazine.…”