BackgroundThe gastrin-releasing peptide receptor (GRPr) is a molecular target for the visualization of prostate cancer. Bombesin (BN) analogs are short peptides with a high affinity for GRPr. RM2 is a bombesin-based antagonist. It has been demonstrated that RM2 have superior in vivo biodistribution and targeting properties than high-affinity receptor agonists. This study developed new RM2-like antagonists by introducing the novel bifunctional chelators AAZTA5and DATA5mto RM2.ResultsThe effects of different macrocyclic chelating groups on drug targeting properties and the possibility of preparing68Ga-radiopharmaceuticals in a kit-based protocol were investigated using68Ga-labeled entities. Both new RM2 variants were labelled with68Ga3+resulting in high yields, stability, and low molarity of the ligand. DATA5m-RM2 and AAZTA5-RM2 incorporated68Ga3+nearly quantitatively at room temperature within 3–5 min, and the labelling yield for68Ga-DOTA-RM2 was approximately 10% under the same conditions.68Ga-AAZTA5-RM2 showed stronger hydrophilicity according to partition coefficient. Although the maximal cellular uptake values of the three compounds were similar,68Ga-AAZTA5-RM2 and68Ga-DATA5m-RM2 peaked more rapidly. Biodistribution studies showed high and specific tumor uptake, with a maximum of 9.12 ± 0.81 percentage injected activity per gram of tissue (%ID/g) for68Ga-DATA5m-RM2 and 7.82 ± 0.61%ID/g for68Ga-AAZTA5-RM2 at 30 min after injection.ConclusionsThe conditions for complexation of DATA5m-RM2 and AAZTA5-RM2 with gallium-68 are milder, faster and require less amount of precursors than DOTA-RM2. Chelators had an evident influence on the pharmacokinetics and targeting properties of68Ga-X-RM2 derivatives. Positively charged68Ga-DATA5m-RM2 provided a high tumor uptake, high image contrast and good capability of targeting GRPr.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13550-023-01005-1.