The tetraamine chelator outperforms HYNIC in a new technetium-99m-labelled somatostatin receptor 2 antagonist

Somatostatin receptor targeting radiopeptides are successfully being used to image, stage, and monitor patients with neuroendocrine tumours. They are exclusively agonists that internalise upon binding to the relevant receptor. According to recent reports, antagonists may be preferable to agonists. To date, ^99mTc-labelled somatostatin receptor antagonists have attracted little attention. Here, we report on a new somatostatin receptor subtype 2 (sst2) antagonist, SS-01 (p-Cl-Phe-cyclo(D-Cys-Tyr-D-Trp-Lys-Thr-Cys)D-Tyr-NH₂), with the aim of developing ^99mTc-labelled ligands for SPECT/CT imaging. SS-01 was prepared using Fmoc solid-phase synthesis and subsequently coupled to the chelators 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), 6-carboxy-1,4,8,11-tetraazaundecane (N4), and 6-hydrazinonicotinic acid (HYNIC) to form the corresponding peptide-chelator conjugates SS-03, SS-04, and SS-05, respectively. SS-04 and SS-05 were radiolabelled with ^99mTc and SS-03 with ¹⁷⁷Lu. Binding affinity and antagonistic properties were determined using autoradiography and immunofluorescence microscopy. Biodistribution and small animal SPECT/CT studies were performed on mice bearing HEK293-rsst2 xenografts.

The conjugates showed low nanomolar sst2 affinity and antagonistic properties. ¹⁷⁷Lu-DOTA-SS-01 (¹⁷⁷Lu-SS-03) and ^99mTc-N4-SS-01 (^99mTc-SS-04) demonstrated high cell binding and low internalisation, whereas ^99mTc-HYNIC/edda-SS-01 (^99mTc-SS-05) showed practically no cellular uptake in vitro. The ^99mTc-SS-04 demonstrated impressive tumour uptake at early time points, with 47% injected activity per gram tumour (%IA/g) at 1 h post-injection. The tumour uptake persisted after 4 h and was 32.5 %IA/g at 24 h. The uptake in all other organs decreased much more rapidly leading to high tumour-to-normal organ ratios, which was reflected in high-contrast SPECT/CT images.

These data indicate a very promising ^99mTc-labelled sst2-targeting antagonist. The results demonstrate high sensitivity of the ^99mTc-labelling strategy, which was shown to strongly influence the receptor affinity, contrary to corresponding agonists. ^99mTc-SS-04 exhibits excellent pharmacokinetics and imaging properties and appears to be a suitable candidate for SPECT/CT clinical translation.