Significant reduction of activity retention in the kidneys via optimized linker sequences in radiohybrid-based minigastrin analogs.

BACKGROUND

We recently introduced radiohybrid (rh)-based minigastrin analogs e.g., DOTA-rhCCK-18 (DOTA-D-Dap(p-SiFA)-(D-γ-Glu)-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH), that revealed substantially increased activity retention in the tumor. However, one major drawback of these first generation rh-based cholecystokinin-2 receptor (CCK-2R) ligands is their elevated activity levels in the kidneys, especially at later time points (24 h p.i.). Therefore, this study aimed to reduce kidney retention with regard to a therapeutic use via substitution of negatively charged D-glutamic acid moieties by hydrophilic uncharged polyethylene glycol (PEG) linkers of various length ((PEG) to (PEG)). Furthermore, the influence of differently charged silicon-based fluoride acceptor (SiFA)-moieties (p-SiFA: neutral, SiFA-ipa: negatively charged, and SiFAlin: positively charged) on in vitro properties of minigastrin analogs was evaluated. Out of all compounds evaluated in vitro, the two most promising minigastrin analogs were further investigated in vivo.

RESULTS

CCK-2R affinity of most compounds evaluated was found to be in a range of 8-20 nM (by means of apparent IC), while ligands containing a SiFA-ipa moiety displayed elevated IC values. Lipophilicity was noticeably lower for compounds containing a D-γ-glutamate (D-γ-Glu) moiety next to the D-Dap(SiFA) unit as compared to their counterparts lacking the additional negative charge. Within this study, combining the most favorable CCK-2R affinity and lipophilicity, [Lu]Lu-DOTA-rhCCK-70 (DOTA-D-Dap(p-SiFA)-D-γ-Glu-(PEG)-D-γ-Glu-(PEG)-Trp-(N-Me)Nle-Asp-1-Nal-NH; IC: 12.6 ± 2.0 nM; logD: - 1.67 ± 0.08) and [Lu]Lu-DOTA-rhCCK-91 (DOTA-D-Dap(SiFAlin)-D-γ-Glu-(PEG)-D-γ-Glu-(PEG)-Trp-(N-Me)Nle-Asp-1-Nal-NH; IC: 8.6 ± 0.7 nM; logD =  - 1.66 ± 0.07) were further evaluated in vivo. Biodistribution data of both compounds revealed significantly reduced (p < 0.0001) activity accumulation in the kidneys compared to [Lu]Lu-DOTA-rhCCK-18 at 24 h p.i., leading to enhanced tumor-to-kidney ratios despite lower tumor uptake. However, overall tumor-to-background ratios of the novel compounds were lower than those of [Lu]Lu-DOTA-rhCCK-18.

CONCLUSION

We could show that the reduction of negative charges within the linker section of radiohybrid-based minigastrin analogs led to decreased activity levels in the kidneys at 24 h p.i., while maintaining a good tumor uptake. Thus, favorable tumor-to-kidney ratios were accomplished in vivo. However, further optimization has to be done in order to improve tumor retention and general biodistribution profile.