Purpose AAZTA (1,4-bis (carboxymethyl)-6-[bis (carboxymethyl)]amino-6-methylperhydro-1,4-diazepine) based chelators were initially developed in the context of magnetic resonance imaging. First radiochemical studies showed the capability of AAZTA to form stable complexes with radiolanthanides and moderately stable complexes with 68 Ga. For a systematic comparison of the labelling capabilities with current diagnostic and therapeutic trivalent radiometals, AAZTA 5 (1,4-bis (carboxymethyl)-6-[bis (carboxymethyl)]amino-6-[pentanoic-acid]perhydro-1,4-diazepine) was synthesized representing a bifunctional version with a pentanoic acid at the carbon-6 atom. To evaluate the effect of adding a targeting vector (TV) to the bifunctional chelator on the complex formation, AAZTA 5 -TOC was synthesized, radiolabeled and tested in comparison to the uncoupled AAZTA 5 . Methods AAZTA 5 was synthesized in a 5-step synthesis. It was coupled to the cyclic peptide TOC (Phe 1 -Tyr 3 octreotide) via amide bound formation. AAZTA and AAZTA 5 -TOC complex formations with 68 Ga, 44 Sc and 177 Lu were investigated at different pH, temperature and precursor amounts. Stability studies against human serum, PBS buffer, EDTA and DTPA were performed. Results AAZTA 5 and AAZTA 5 -TOC achieved quantitative labelling (> 95%) at room temperature in less than 5 min with all three nuclides at pH ranges from 4 to 5.5 with low precursor amounts of 1 to 10 nmol. [ 44 Sc]Sc-AAZTA 5 complexes as well as [ 44 Sc]Sc-AAZTA 5 -TOC were completely stable. The 177 Lu complexes of AAZTA 5 and AAZTA 5 -TOC showed high stability comparable to the 44 Sc complexes. In contrast, the [ 68 Ga]Ga-AAZTA 5 complex stability was rather low, but interestingly, [ 68 Ga]Ga-AAZTA 5 -TOC was completely stable. Conclusion AAZTA 5 appears to be a promising bifunctional chelator for 68 Ga, 44 Sc and 177 Lu with outstanding labelling capabilities at room temperature. Complex stabilities are high in the case of 44 Sc and 177 Lu. While [ 68 Ga]Ga-AAZTA complexes alone lacking stability, [ 68 Ga]Ga-AAZTA ...

Background Pretargeted imaging allows the use of short-lived radionuclides when imaging the accumulation of slow clearing targeting agents such as antibodies. The biotin-(strept)avidin and the bispecific antibody-hapten interactions have been applied in clinical pretargeting studies; unfortunately, these systems led to immunogenic responses in patients. The inverse electron demand Diels-Alder (IEDDA) reaction between a radiolabelled tetrazine (Tz) and a trans -cyclooctene (TCO)-functionalized targeting vector is a promising alternative for clinical pretargeted imaging due to its fast reaction kinetics. This strategy was first applied in nuclear medicine using an 111 In-labelled Tz to image TCO-functionalized antibodies in tumour-bearing mice. Since then, the IEDDA has been used extensively in pretargeted nuclear imaging and radiotherapy; however, these studies have only been performed in mice. Herein, we report the 44 Sc labelling of a Tz and evaluate it in pretargeted imaging in Wistar rats. Results 44 Sc was obtained from an in house 44 Ti/ 44 Sc generator. A 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-functionalized tetrazine was radiolabelled with 44 Sc resulting in radiochemical yields of 85–95%, a radiochemical purity > 99% at an apparent molar activity of 1 GBq/mmol. The 44 Sc-labelled Tz maintained stability in solution for up to 24 h. A TCO-functionalized bisphosphonate, which accumulates in skeletal tissue, was used as a targeting vector to evaluate the 44 Sc-labelled Tz. Biodistribution data of the 44 Sc-labelled Tz showed specific uptake (0.9 ± 0.3% ID/g) in the bones (humerus and femur) of rats pre-treated with the TCO-functionalized bisphosphonate. This uptake was not present in rats not receiving pre-treatment (< 0.03% ID/g). Conclusions We have prepared a 44 Sc-labelled Tz and used it in pretargeted PET imaging with rats treated with TCO-functionalized bisphosponates. This allowed for the evaluation of the IEDDA reaction in animals larger than a typical mouse. Non-target accumulation was low, and there was a 30-fold higher bone uptake in the pre-treated rats compared to the non-treated controls. Given its convenient half-life and the ability to perform positron emission tomography with a previously studied DOTA-functionalized Tz, scandium-44 ( t 1/2 = 3.97 h) proved to be a suitable radioisotope for this study. Electronic supplementary material The online version of this article (10.1186/s13550-019-0520-y) contains supplementary material, which is available to authorized users.

Purpose The widespread use of 68 Ga for positron emission tomography (PET) relies on the development of radiopharmaceutical precursors that can be radiolabelled and dispensed in a simple, quick, and convenient manner. The DATA (6-amino-1,4-diazapine-triacetate) scaffold represents a novel hybrid chelator architecture possessing both cyclic and acyclic character that may allow for facile access to 68 Ga-labelled tracers in the clinic. We report the first bifunctional DATA chelator conjugated to [Tyr 3 ]octreotide (TOC), a somatostatin subtype 2 receptor (SST 2 )-targeting vector for imaging and functional characterisation of SSTR 2 expressing tumours. Methods The radiopharmaceutical precursor, DATA-TOC, was synthesised as previously described and used to complex nat Ga(III) and 68 Ga(III). Competition binding assays of [ nat Ga]Ga-DATA-TOC or [ nat Ga]Ga-DOTA-TOC against [ 125 I-Tyr 25 ]LTT-SS28 were conducted in membranes of HEK293 cells transfected to stably express one of the hSST 2,3,5 receptor subtypes (HEK293-hSST 2/3/5 cells). First in vivo studies were performed in female NMRI-nude mice bearing SST 2 -positive mouse phaeochromocytoma mCherry (MPC-mCherry) tumours to compare the in vivo SST 2 -specific tumour-targeting of [ 68 Ga]Ga-DATA-TOC and its overall pharmacokinetics versus the [ 68 Ga]Ga-DOTA-TOC reference. A direct comparison of [ 68 Ga]Ga-DATA-TOC with the well-established PET radiotracer [ 68 Ga]Ga-DOTA-TOC was additionally performed in a 46-year-old male patient with a well-differentiated NET (neuroendocrine tumour), representing the first in human administration of [ 68 Ga]Ga-DATA-TOC. Results DATA-TOC was labelled with 68 Ga with a radiolabelling efficiency of > 95% in less than 10 min at ambient temperature. A molar activity up to 35 MBq/nmol was achieved. The hSST 2 -affinities of [ nat Ga]Ga-DATA-TOC and [ nat Ga]Ga-DOTA-TOC were found similar with only sub-nanomolar differences in the respective IC 50 values. In mice, [ 68 Ga]Ga-DATA-TOC was able to visualise the tumour lesions, showing standardised uptake values (SUVs) similar to [ 68 Ga]Ga-DOTA-TOC. Direct comparison of the two PET tracers in a NET patient revealed very similar tumour uptake for the two 68 Ga-radiotracer...

The L‐lysine urea‐L‐glutamate (KuE) represents a key motif in recent diagnostic and therapeutic radiopharmaceuticals targeting the prostate specific membrane antigen (PSMA). Using a squaric acid moiety for coupling of KuE with a radioactive label, the squaric acid as a linker in the PSMA ligand seems to mimic the aromatic structure of the naphthylalanine unit on PSMA‐617. In this work, we investigate the influence of squaric acid moiety on the biological activity of the compound carrying a KuE motif and three typical chelates. The derivatives TRAM.SA.KuE, DOTAGA.SA.KuE and NODAGA.SA.KuE were all synthesized in straightforward organic reactions and purified by HPLC afterward. Different amounts of tracer were labeled at different temperatures with 68Ga. PET examinations were performed on NMRInu/nu nude mice with an LNCaP tumor on the right hind leg including ex vivo investigations of the organs. For comparison, 68Ga‐derivatives of PSMA‐11 and PSMA‐617, the derivatives most commonly used in clinics, were investigated in the same animal model.

Background The AAZTA chelator and in particular its bifunctional derivative AAZTA5 was recently investigated to demonstrate unique capabilities to complex diagnostic and therapeutic trivalent radiometals under mild conditions. This study presents a comparison of 68Ga, 44Sc and 177Lu-labeled AAZTA5-PSMA-617 with DOTA-PSMA-617 analogues. We evaluated the radiolabeling characteristics, in vitro stability of the radiolabeled compounds and evaluated their binding affinity and internalization behavior on LNCaP tumor cells in direct comparison to the radiolabeled DOTA-conjugated PSMA-617 analogs. Results AAZTA5 was synthesized in a five-step synthesis and coupled to the PSMA-617 backbone on solid phase. Radiochemical evaluation of AAZTA5-PSMA-617 with 68Ga, 44Sc and 177Lu achieved quantitative radiolabeling of > 99% after less than 5 min at room temperature. Stabilities against human serum, PBS buffer and EDTA and DTPA solutions were analyzed. While there was a small degradation of the 68Ga complex over 2 h in human serum, PBS and EDTA/DTPA, the 44Sc and 177Lu complexes were stable at 2 h and remained stable over 8 h and 1 day. For all three compounds, i.e. [natGa]Ga-AAZTA5-PSMA-617, [natSc]Sc-AAZTA5-PSMA-617 and [natLu]Lu-AAZTA5-PSMA-617, in vitro studies on PSMA-positive LNCaP cells were performed in direct comparison to radiolabeled DOTA-PSMA-617 yielding the corresponding inhibition constants (Ki). Ki values were in the range of 8–31 nM values which correspond with those of [natGa]Ga-DOTA-PSMA-617, [natSc]Sc-DOTA-PSMA-617 and [natLu]Lu-DOTA-PSMA-617, i.e. 5–7 nM, respectively. Internalization studies demonstrated cellular membrane to internalization ratios for the radiolabeled 68Ga, 44Sc and 177Lu-AAZTA5-PSMA-617 tracers (13–20%IA/106 cells) in the same range as the ones of the three radiolabeled DOTA-PSMA-617 tracers (17–20%IA/106 cells) in the same assay. Conclusions The AAZTA5-PSMA-617 structure proved fast and quantitative radiolabeling with all three radiometal complexes at room temperature, excellent stability with 44Sc, very high stability with 177Lu and medium stability with 68Ga in human serum, PBS and EDTA/DTPA solutions. All three AAZTA5-PSMA-617 tracers showed binding affinities and internalization ratios in LNCaP cells comparable with that of radiolabeled DOTA-PSMA-617 analogues. Therefore, the exchange of the chelator DOTA with AAZTA5 within the PSMA-617 binding motif has no negative influence on in vitro LNCaP cell binding characteristics. In combination with the faster and milder radiolabeling features, AAZTA5-PSMA-617 thus demonstrates promising potential for in vivo application for theranostics of prostate cancer.