[⁶⁸Ga]Ga-THP-Pam: A Bisphosphonate PET Tracer with Facile Radiolabeling and Broad Calcium Mineral Affinity

Abstract

Calcium minerals such as hydroxyapatite (HAp) can be detected non-invasively in vivo using nuclear imaging agents such as [ 18 F]NaF (available from cyclotrons), for positron emission tomography (PET) and 99m Tc-radiolabelled bisphosphonates (BP; available from 99m Tc generators for single photon emission computed tomography (SPECT) or scintigraphy). These two types of imaging agents allow detection of bone metastases (based on the presence of HAp) and vascular calcification lesions (that contain HAp and other calcium minerals). With the aim of developing a cyclotron-independent PET radiotracer for these lesions, with broad calcium mineral affinity and simple one-step radiolabelling, we developed [ 68 Ga]Ga-THP-Pam. Radiolabelling with 68 Ga is achieved using a mild single-step kit (5 min, room temperature, pH 7) to high radiochemical yield and purity (>95%). NMR studies demonstrate that Ga binds via the THP chelator, leaving the BP free to bind to its biological target. [ 68 Ga]Ga-THP-Pam shows high stability in human serum. The calcium mineral binding of [ 68 Ga]Ga-THP-Pam was compared in vitro to two other 68 Ga-BPs which have been successfully evaluated in humans, [ 68 Ga]Ga-NO2AP BP and [ 68 Ga]Ga-BPAMD, as well as [ 18 F]NaF. Interestingly, we found that all 68 Ga-BPs have a high affinity for a broad range of calcium minerals implicated in vascular calcification disease, while [ 18 F]NaF is selective for HAp. Using healthy young mice as a model of metabolically active growing calcium mineral in vivo, we compared the pharmacokinetics and biodistribution of [ 68 Ga]Ga-THP-Pam with [ 18 F]NaF as well as [ 68 Ga]NO2AP BP. These studies revealed that [ 68 Ga]Ga-THP-Pam has high in vivo affinity for bone tissue (high bone/muscle and bone/blood ratios) and fast blood clearance (t1/2 <10 min) comparable to both [ 68 Ga]NO2AP BP and [ 18 F]NaF. Overall, [ 68 Ga]Ga-THP-Pam shows high potential for clinical translation as a cyclotron-independent calcium mineral PET radiotracer, with simple and efficient radiochemistry that can be easily implemented in any radiopharmacy.

Towards Facile Radiolabeling and Preparation of Gallium-68-/Bismuth-213-DOTA-[Thi8, Met(O2)11]-Substance P for Future Clinical Application: First Experiences

“…Other DOTA-conjugated peptides are also able to incorporate radionuclides, such as 68 Ga, at moderate acidity of the eluate [ 42 ] although, from a pH above 4.5, colloidal- 68 Ga formation has been reported [ 41 ]. Alternate chelators such as 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA) and tris(hydroxypyridinone) (THP) [ 47 , 48 ] though more favorable for complexation of 68 Ga, are not as compatible with 213 Bi when considering a theranostic approach. For radionuclides that are dissolved in acidic solutions such as hydrochloric acid (HCl), fast reaction kinetics are supported using sodium acetate buffer to stabilize the pH during the radiolabeling reaction [ 49 ].…”

NHS-Functionalized THP Derivative for Efficient Synthesis of Kit-Based Precursors for 68Ga Labeled PET Probes

“…Its performance has previously been evaluated against a range of common chelators, showing better properties for radiolabeling under mild conditions. THP has also been functionalized for conjugation to peptides, proteins, and other targeting molecules such as bisphosphonates while retaining the required mild radiolabeling and in vivo targeting properties [ 15 , 16 , 17 , 18 ]. Moreover, peptide-THP-based PET tracers have been already tested in clinical trials with promising results [ 19 ].…”

“…When technetium-99m ( 99m Tc) is used as radioactive source the tracer is labeled with speed, simplicity, and reproducibility from commercially available "cold kits" compatible with good manufacturing practice (GMP) [2]. With the growth of 18 F and 11 C-based PET imaging tracers, more complex and costly infrastructures have been built to support PET because a kit model is not compatible with those tracers and due to the need for an on-site cyclotron and more complex synthetic chemistry procedures involved in radiolabeling. Modern 68 Ga generators are compatible with GMP and have the potential to be used in the manufacturing of kit-based radiotracers if a simple chelation step can be achieved [3,4], making 68 Ga tracers widely available without the costly infrastructures related with 18 F and 11 C production.…”

“…With the growth of 18 F and 11 C-based PET imaging tracers, more complex and costly infrastructures have been built to support PET because a kit model is not compatible with those tracers and due to the need for an on-site cyclotron and more complex synthetic chemistry procedures involved in radiolabeling. Modern 68 Ga generators are compatible with GMP and have the potential to be used in the manufacturing of kit-based radiotracers if a simple chelation step can be achieved [3,4], making 68 Ga tracers widely available without the costly infrastructures related with 18 F and 11 C production. Despite several efforts for a kit-based 68 Ga tracer production [5], an ideal one-step procedure for gallium radiolabeling, with the same simplicity of the long-established technetium labeling procedures (addition of generator eluate to the kit vial), has not yet been widely achieved [6,7].…”

HAP-multitag, a PET and positive MRI contrast nanotracer for the longitudinal characterization of vascular calcifications in atherosclerosis

“…4 On the other hand, the [ 18 F]FNa only binds to HAP, while bisphosphonate-based (BP) tracers or nanoparticles cover a broader spectrum of calcium salts recognition, relevant in several diseases. 5–8 The mechanism of accumulation in calcifications is different for both types of tracers: in using [ 18 F]FNa, 18 F substitutes one hydroxyl group in the HAP matrix, forming fluorapatite, while when using BP-based probes, the bisphosphonate moiety coordinates with the Ca atom. When using [ 18 F]FNa or BP-based tracers, one of the main drawbacks is the high uptake they show in bone, increasing the off-target signal, often complicating vasculature differentiation.…”