PSMA binding of carbamate-containing peptidomimetics influenced by side chain length and stereocenter configuration in preclinical prostate cancer model.

Abstract

To explore how core backbone modifications influence prostate-specific membrane antigen (PSMA) targeting, we synthesized a series of novel peptidomimetics replacing the classic urea linkage of established radiotracers with a carbamate functionality. This panel systematically varied side chain length (aspartic vs. aminoadipic acid), stereocenter configuration (S/S vs. S/R diastereomers), and the radiometal chelator (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid [DOTA] vs. N,N'-bis[2-hydroxy-5-(carboxyethyl)benzyl]ethylenediamine-N,N'-diacetic acid [HBED-CC]). In vitro binding assays in LNCaP cells revealed that the carbamate backbone severely compromised PSMA affinity (Ki) for short-chain aspartic acid derivatives, whereas long-chain aminoadipic acid conjugates successfully retained low-nanomolar potencies. Across all variations, S/S configurations displayed superior affinities over S/R counterparts, demonstrating that natural stereochemistry is essential for optimal binding pocket interactions. Although both conjugates showed high in vitro potency, high-temperature radiolabeling of the DOTA derivative with gallium-68 caused thermal degradation via hydrolytic cleavage and intramolecular cyclization of the carbamate backbone. Conversely, the companion HBED-CC conjugate, 37(S/S), was successfully radiolabeled at ambient temperature without structural compromise to yield the intact radiotracer, 40(S/S). In vivo positron emission tomography (PET) imaging and biodistribution studies of 40(S/S) in LNCaP tumor-bearing mice demonstrated high tumor uptake (∼12%ID/g) at 1 h post-injection, rapid renal clearance, and excellent tumor-to-background ratios (tumor-to-muscle: ∼23; tumor-to-bone: ∼36). These findings demonstrate that carbamate backbones can successfully mimic urea linkages in PSMA ligands, provided they are paired with precise side chain lengths and mild radiolabeling conditions.