The pretargeting strategy has recently emerged in order to overcome the limitations of direct targeting, mainly in the field of radioimmunotherapy (RIT). This strategy is directly dependent on chemical reactions, namely bioorthogonal reactions, which have been developed for their ability to occur under physiological conditions. The Staudinger ligation, the copper catalyzed azide-alkyne cycloaddition (CuAAC) and the strain-promoted [3 + 2] azide–alkyne cycloaddition (SPAAC) were the first bioorthogonal reactions introduced in the literature. However, due to their incomplete biocompatibility and slow kinetics, the inverse-electron demand Diels-Alder (IEDDA) reaction was advanced in 2008 by Blackman et al. as an optimal bioorthogonal reaction. The IEDDA is the fastest bioorthogonal reaction known so far. Its biocompatibility and ideal kinetics are very appealing for pretargeting applications. The use of a trans-cyclooctene (TCO) and a tetrazine (Tz) in the reaction encouraged researchers to study them deeply. It was found that both reagents are sensitive to acidic or basic conditions. Furthermore, TCO is photosensitive and can be isomerized to its cis-conformation via a radical catalyzed reaction. Unfortunately, the cis-conformer is significantly less reactive toward tetrazine than the trans-conformation. Therefore, extensive research has been carried out to optimize both click reagents and to employ the IEDDA bioorthogonal reaction in biomedical applications.

Nuclear and optical dual-modality probes can be of great assistance in prostate cancer localization, providing the means for both preoperative nuclear imaging and intraoperative surgical guidance. We developed a series of probes based on the backbone of the established GRPR-targeting radiotracer NeoB. The inverse electron demand of the Diels–Alder reaction was used to integrate the sulfo-cyanine 5 dye. Indium-111 radiolabeling, stability studies and a competition binding assay were carried out. Pilot biodistribution and imaging studies were performed in PC-3 tumor-bearing mice, using the best two dual-labeled probes. The dual-modality probes were radiolabeled with a high yield (> 92%), were proven to be hydrophilic and demonstrated high stability in mouse serum (> 94% intact labeled ligand at 4 h). The binding affinity for the GRPR was in the nanomolar range (21.9–118.7 nM). SPECT/CT images at 2 h p.i. clearly visualized the tumor xenograft and biodistribution studies, after scanning confirmed the high tumor uptake (8.47 ± 0.46%ID/g and 6.90 ± 0.81%ID/g for probe [111In]In-12 and [111In]In-15, respectively). Receptor specificity was illustrated with blocking studies, and co-localization of the radioactive and fluorescent signal was verified by ex vivo fluorescent imaging. Although optimal tumor-to-blood and tumor-to-kidney ratios might not yet have been reached due to the prolonged blood circulation, our probes are promising candidates for the preoperative and intraoperative visualization of GRPR-positive prostate cancer.

Background The [177Lu]Lu-DOTA-TATE mediated peptide receptor radionuclide therapy (PRRT) of neuroendocrine tumors (NETs) is sometimes leading to treatment resistance and disease recurrence. An interesting alternative could be the somatostatin antagonist, [177Lu]Lu-DOTA-JR11, that demonstrated better biodistribution profile and higher tumor uptake than [177Lu]Lu-DOTA-TATE. Furthermore, treatment with alpha emitters showed improvement of the therapeutic index of PRRT due to the high LET offered by the alpha particles compared to beta emitters. Therefore, [225Ac]Ac-DOTA-JR11 can be a potential candidate to improve the treatment of NETs (Graphical abstract). DOTA-JR11 was radiolabeled with [225Ac]Ac(NO3)3 and [177Lu]LuCl3. Stability studies were performed in phosphate buffered saline (PBS) and mouse serum. In vitro competitive binding assay has been carried out in U2OS-SSTR2 + cells for natLa-DOTA-JR11, natLu-DOTA-JR11 and DOTA-JR11. Ex vivo biodistribution studies were performed in mice inoculated with H69 cells at 4, 24, 48 and 72 h after injection of [225Ac]Ac-DOTA-JR11. A blocking group was included to verify uptake specificity. Dosimetry of selected organs was determined for [225Ac]Ac-DOTA-JR11 and [177Lu]Lu-DOTA-JR11. Results [225Ac]Ac-DOTA-JR11 has been successfully prepared and obtained in high radiochemical yield (RCY; 95%) and radiochemical purity (RCP; 94%). [225Ac]Ac-DOTA-JR11 showed reasonably good stability in PBS (77% intact radiopeptide at 24 h after incubation) and in mouse serum (~ 81% intact radiopeptide 24 h after incubation). [177Lu]Lu-DOTA-JR11 demonstrated excellent stability in both media (> 93%) up to 24 h post incubation. Competitive binding assay revealed that complexation of DOTA-JR11 with natLa and natLu did not affect its binding affinity to SSTR2. Similar biodistribution profiles were observed for both radiopeptides, however, higher uptake was noticed in the kidneys, liver and bone for [225Ac]Ac-DOTA-JR11 than [177Lu]Lu-DOTA-JR11. Conclusion [225Ac]Ac-DOTA-JR11 showed a higher absorbed dose in the kidneys compared to [177Lu]Lu-DOTA-JR11, which may limit further studies with this radiopeptide. However, several strategies can be explored to reduce nephrotoxicity and offer opportunities for future clinical investigations with [225Ac]Ac-DOTA-JR11.

Somatostatin receptor subtype 2 (SSTR2) has become an essential target for radionuclide therapy of neuroendocrine tumors (NETs). JR11 was introduced as a promising antagonist peptide to target SSTR2. However, due to its rapid blood clearance, a better pharmacokinetic profile is necessary for more effective treatment. Therefore, two JR11 analogs (8a and 8b), each carrying an albumin binding domain, were designed to prolong the blood residence time of JR11. Both compounds were labeled with lutetium-177 and evaluated via in vitro assays, followed by in vivo SPECT/CT imaging and ex vivo biodistribution studies. [177Lu]Lu-8a and [177Lu]Lu-8b were obtained with high radiochemical purity (>97%) and demonstrated excellent stability in PBS and mouse serum (>95%). [177Lu]Lu-8a showed better affinity towards human albumin compared to [177Lu]Lu-8b. Further, 8a and 8b exhibited binding affinities 30- and 48-fold lower, respectively, than that of the parent peptide JR11, along with high cell uptake and low internalization rate. SPECT/CT imaging verified high tumor accumulation for [177Lu]Lu-8a and [177Lu]Lu-JR11 at 4, 24, 48, and 72 h post-injection, but no tumor uptake was observed for [177Lu]Lu-8b. Ex vivo biodistribution studies revealed high and increasing tumor uptake for [177Lu]Lu-8a. However, its extended blood circulation led to an unfavorable biodistribution profile for radionuclide therapy.