Synthesis and evaluation of novel gonadotropin-releasing hormone receptor-targeting peptides

The purpose of this study was to develop novel radiolabeled gonadotropin-releasing hormone (GnRH) receptor-targeting peptides for breast cancer imaging. Three novel 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-conjugated GnRH peptides were designed and synthesized. The radiometal chelator DOTA was conjugated to the epsilon or alpha amino group of d-lysine, or the epsilon amino group of l-lysine via an Ahx {aminohexanoic acid} linker to generate DOTA-Ahx-(d-Lys⁶-GnRH1), DOTA-Ahx-(d-Lys ⁶-GnRH2) and DOTA-Ahx-(l-Lys⁶-GnRH3), respectively. The conjugation of the DOTA to the epsilon amino group of d-lysine (rather than alpha amino group of d-lysine nor epsilon amino group of l-lysine) maintained the nanomolar GnRH receptor binding affinity. The IC₅₀ values of DOTA-Ahx-(d-Lys⁶-GnRH1), DOTA-Ahx-(d-Lys⁶-GnRH2) and DOTA-Ahx-(l-Lys⁶-GnRH3) were 36.1 nM, 10.6 mM and 4.3 mM, respectively. Since only DOTA-Ahx-(d-Lys⁶-GnRH1) displayed nanomolar receptor binding affinity, the specific GnRH receptor binding of ¹¹¹In-DOTA-Ahx-(d-Lys⁶-GnRH1) was determined in human GnRH receptor membrane preparations. Furthermore, the biodistribution and tumor imaging properties of ¹¹¹In-DOTA-Ahx-(d-Lys⁶-GnRH1) were examined in MDA-MB-231 human breast cancer-xenografted nude mice. ¹¹¹In-DOTA-Ahx-(d-Lys⁶-GnRH1) exhibited specific GnRH receptor binding and rapid tumor uptake (1.76 ± 0.58% ID/g at 0.5 h postinjection) coupled with fast whole-body clearance through the urinary system. The MDA-MB-231 human breast cancer-xenografted tumor lesions were clearly visualized by single photon emission computed tomography (SPECT)/CT at 1 h postinjection of ¹¹¹In-DOTA-Ahx-(d-Lys⁶-GnRH1). The profound impact of DOTA position on the binding affinity of the GnRH peptide provided a new insight into the design of novel radiolabeled GnRH peptides. The successful imaging of MDA-MB-231 human breast cancer-xenografted tumor lesions using ¹¹¹In-DOTA-Ahx-(d-Lys⁶-GnRH1) suggested its potential as a novel imaging probe for human breast cancer imaging.