Amide-to-triazole switch vs. in vivo NEP-inhibition approaches to promote radiopeptide targeting of GRPR-positive tumors

Abstract

Introduction: Radiolabeled bombesin (BBN)-analogs have been proposed for diagnosis and therapy of gastrin-releasing peptide receptor (GRPR)-expressing tumors, such as prostate, breast and lung cancer. Metabolic stability represents a crucial factor for the success of this approach by ensuring sufficient delivery of circulating radioligand to tumor sites. The amide-to-triazole switch on the backbone of DOTA-PEG₄-[Nle¹⁴]BBN(7-14) (1) was reported to improve the in vitro stability of resulting ¹⁷⁷Lu-radioligands. On the other hand, in-situ inhibition of neutral endopeptidase (NEP) by coinjection of phosphoramidon (PA) was shown to significantly improve the in vivo stability and tumor uptake of biodegradable radiopeptides. We herein compare the impact of the two methods on the bioavailability and localization of ¹⁷⁷Lu-DOTA-PEG₄-[Nle¹⁴]BBN(7-14) analogs in GRPR-positive tumors in mice.

Methods: The 1,4-disubstituted [1-3]-triazole was used to replace one (2: Gly¹¹-His¹²; 3: Ala⁹-Val¹⁰) or two (4: Ala⁹-Val¹⁰ and Gly¹¹-His¹²) peptide bonds in 1 (reference) and all compounds were labeled with ¹⁷⁷Lu. Each of [¹⁷⁷Lu]1-[¹⁷⁷Lu]4 was injected without (control) or with PA in healthy mice. Blood samples collected 5min post-injection (pi) were analyzed by HPLC. Biodistribution of [¹⁷⁷Lu]1-[¹⁷⁷Lu]4 was conducted in SCID mice bearing human prostate adenocarcinoma PC-3 xenografts at 4h pi. Groups of 4 animals were injected with radioligand, alone (controls), or with coinjection of PA, or of a mixture of PA and excess and [Tyr⁴]BBN to determine GRPR-specificity of uptake (Block).

Results: The in vivo stability of the radioligands was: [¹⁷⁷Lu]1 (25% intact), [¹⁷⁷Lu]2 (45% intact), [¹⁷⁷Lu]3 (30% intact) and [¹⁷⁷Lu]4 (40% intact). By PA-coinjection these values notably increased to 90%-93%. Moreover, treatment with PA induced an impressive and GRPR-specific uptake of all radioligands in the PC-3 xenografts at 4h pi: [¹⁷⁷Lu]1: 4.7±0.4 to 24.8±4.9%ID/g; [¹⁷⁷Lu]2: 8.3±1.2 to 26.0±1.1%ID/g; [¹⁷⁷Lu]3: 6.6±0.4 to 21.3±4.4%ID/g; and [¹⁷⁷Lu]4: 4.8±1.6 to 13.7±3.8%ID/g.

Conclusions: This study has shown that amide-to-triazole substitutions in ¹⁷⁷Lu-DOTA-PEG₄-[Nle¹⁴]BBN(7-14) induced minor effects on bioavailability and tumor uptake in mice models, whereas in-situ NEP-inhibition(s) by PA impressively improved in vivo profiles.

Keywords: (177)Lu-bombesin; GRPR-radioligand; NEP-inhibition; Triazolyl-bombesin; Tumor targeting.