Introduction: In this study two In-111-labeled dimeric cyclic RGD peptides, In-111(DOTA-Galacto-RGD(2)) and In-111(DOTA-Galacto-RGD(2)), were evaluated as radiotracers for breast tumor imaging. The objective was to evaluate the impact of SAA, PEG(2) and 1,2,3-triazole linkers as compare to PEG(4) on the tumor uptake and excretion kinetics of In-111 radiotracers. Methods: DOTA-Galacto-RGD(2) was prepared by conjugation of Galacto-RGD(2) with DOTA-OSu in the presence of diisopropylethylamine. Its integrin alpha(v)beta(3) binding affinity was determined using a whole-cell displacement assay against I-125-echistatin bound to U87MG glioma cells, and was compared with those of c(RGDfK), DOTA-3P-RGD(2) and DOTA-3P-RGK(2) (a nonsense peptide conjugate with "scrambled" RGK sequences). In-111(DOTA-Galacto-RGD(2)) and In-111(DOTA-Galacto-RGD(2)) were prepared and evaluated for their tumor-targeting capability and biodistribution properties in athymic nude mice bearing MDA-MB-435 breast tumor xenografts. Planar imaging studies were performed to demonstrate the utility of In-111(DOTA-Galacto-RGD(2)) and In-111(DOTA-Galacto-RGD(2)) for breast tumor imaging. Results: IC50 values of DOTA-Galacto-RGD(2), DOTA-3P-RGD(2), and DOTA-3P-RGK(2) were calculated to be 27 +/- 2, 29 +/- 4, 596 +/- 48 nM, respectively. The tumor uptake values of In-111(DOTA-Galacto-RGD(2)) (6.79 +/- 0.98, 6.56 +/- 0.56, 4.17 +/- 0.61 and 1.09 +/- 0.13 %ID/g at 1, 4, 24 and 72 hours p.i., respectively) were almost identical to those of In-111(DOTA-Galacto-RGD(2)) (6.17 +/- 1.65, 5.94 +/- 0.84, 3.40 +/- 0.50 and 0.99 +/- 0.20 %ID/g, respectively). In-111(DOTA-Galacto-RGD(2)) had a faster clearance from blood and muscle than In-111(DOTA-Galacto-RGD(2)), leading to higher tumor/blood and tumor/muscle ratios. In-111(DOTA-Galacto-RGD(2)) had lower liver uptake and better tumor/liver ratios than In-111(DOTA-Galacto-RGD(2)). The tumor uptake of In-111(DOTA-Galacto-RGD(2)) and In-111(DOTA-Galacto-RGD(2)) was both integrin alpha(v)beta(3) and RGD-specific. Imaging data suggest that In-111(DOTA-Galacto-RGD(2)) and In-111(DOTA-Galacto-RGD(2)) are useful as radiotracers for imaging integrin alpha(v)beta(3)-positive breast tumors. Conclusion: The results from this study suggest that replacing PEG(4) linkers between two RGD moieties with a pair of SAA, PEG(2) and 1,2,3-triazole groups has little impact on integrin alpha(v)beta(3) binding affinity and tumor uptake of In-111-labeled dimeric cyclic RGD peptides. Despite the subtle differences in their excretion kinetics from non-cancerous tissues, In-111(DOTA-Galacto-RGD(2)) and In-111(DOTA-Galacto-RGD(2)) are useful radiotracers for imaging integrin alpha(v)beta(3)-positive breast tumors. (C) 2014 Elsevier Inc. All rights reserved.