Optimizing uPAR-targeting radiopeptides for improved tissue distribution: progress towards radionuclide therapy

The aim of this study was to develop radiopeptides for targeting the urokinase-type plasminogen activator receptor (uPAR) modified with an albumin-binding moiety to improve their tissue distribution profiles.

uPAR-11, uPAR-12, uPAR-14, uPAR-15, uPAR-17 and uPAR-18 were synthesized based on the AE105 nonapeptide which was modified with the p-tolyl-based albumin binder and variable linker entities and chelators. The ¹⁷⁷Lu-labeled peptides were evaluated in vitro with regard to their stability, albumin-binding properties and uPAR-binding affinity using HEK-uPAR cells. Biodistribution and SPECT/CT imaging studies were performed with HEK-uPAR xenografted nude mice. The acquired data were compared to those obtained with [¹⁷⁷Lu]Lu-DOTA-AE105.

The radiopeptides showed 11‒155-fold higher albumin-binding affinity in human blood plasma than [¹⁷⁷Lu]Lu-DOTA-AE105. The uPAR-binding affinity reached K_D values of 31‒42 nM, similar to the K_D value of 20 ± 1 nM determined for [¹⁷⁷Lu]Lu-DOTA-AE105. Accumulation in the HEK-uPAR xenograft was 6.0-16% IA/g at 4 h p.i., which was 7‒18-fold higher than that of [¹⁷⁷Lu]Lu-DOTA-AE105. PEG spacers next to the albumin binder as exemplified in [¹⁷⁷Lu]Lu-uPAR-11, [¹⁷⁷Lu]Lu-uPAR-15 and [¹⁷⁷Lu]Lu-uPAR-18 led to increased xenograft accumulation while their replacement with an alkane spacer in [¹⁷⁷Lu]Lu-uPAR-12 led to unfavorably high blood retention. A diaminopropionic acid to connect the different entities did not improve the tissue distribution profile of [¹⁷⁷Lu]Lu-uPAR-14 as compared to other peptides which were designed with a lysine residue. The use of a DOTAGA chelator instead of a DOTA resulted in unfavorable kidney retention of [¹⁷⁷Lu]Lu-uPAR-17.

In view of a clinical translation, [¹⁷⁷Lu]Lu-uPAR-11 emerged as the most favorable candidate. Future studies will, thus, focus on the therapeutic potential of [¹⁷⁷Lu]Lu-uPAR-11 in tumor-bearing mice.