Evaluation of ⁶⁸Ga-labeled iNGR peptide with tumor-penetrating motif for microPET imaging of CD13-positive tumor xenografts

The aim of the study is to evaluate the efficacy of ⁶⁸Ga-labeled iNGR, containing Asn-Gly-Arg (NGR) homing sequence and CendR (R/KXXR/K) penetrating motif, as a new molecular probe for microPET imaging of CD13-positive xenografts. The synthesized iNGR and NGR peptides were conjugated with DOTA and then labeled with ⁶⁸Ga. ⁶⁸Ga-iNGR and ⁶⁸Ga-NGR were compared in the performance of the in vitro stability, partition coefficient, binding affinity, cell uptake analysis, in vivo microPET imaging, and biodistribution studies in CD13-positive HT-1080 and CD13-negative HT-29 cell lines. The in vitro results revealed that both probes exhibited high radiochemical purity and stability, and no significant difference between two probes was observed in terms of the binding affinity to CD13. In vivo microPET/CT imaging showed that the uptake of ⁶⁸Ga-iNGR in HT-1080 tumor was significantly higher than that of ⁶⁸Ga−NGR. Moreover, tumor ⁶⁸Ga-iNGR uptake could be completely blocked by cold NGR and partially blocked by neutralizing NRP-1 antibody. We concluded that ⁶⁸Ga-iNGR has a higher tumor uptake and better tumor retention than ⁶⁸Ga-NGR through NRP-1, indicating that CendR motif modification is a promising method for improving NGR peptide performance.