Preparation and bioevaluation of [Tc-99m]Tc-labeled A7R and (D)A7R for SPECT imaging of triple-negative breast cancer

Novel strategies for diagnosing triple-negative breast cancer (TNBC) are essential for effective clinical treatment. Vascular endothelial growth factor receptor 2 (VEGFR2) and neuropilin-1 (NRP-1) are potential targets for tumor imaging agents. We designed and synthesized [Tc-99m]Tc-labeled heptapeptide (A7R) and its D-type peptide ((D)A7R) with high affinity and specificity for VEGFR2 and NRP-1 as novel single-photon emission computed tomography (SPECT) probes for TNBC imaging. The specificities of A7R and (D)A7R were first evaluated in vitro using flow cytometry and confocal microscopy and ex vivo using fluorescence imaging. Subsequently, A7R and (D)A7R were labeled with [Tc-99m]Tc through 6-hydrazino nicotinamide (HYNIC), and their radiochemical purities (RCPs) and stability in vitro were assessed. The imaging performance and biodistribution of [Tc-99m]Tc-HYNIC-A7R and [Tc-99m]Tc-HYNIC-(D)A7R were evaluated in TNBC mouse models. A7R and (D)A7R exhibited good TNBC cell-targeting abilities and were readily labeled with [Tc-99m]Tc through HYNIC. Both [Tc-99m]Tc-HYNIC-A7R and [Tc-99m]Tc-HYNIC-(D)A7R had high RCPs and stability in vitro, and their accumulation in tumor tissues in the TNBC models was evident, with fast blood clearance and favorable biodistribution. More importantly, [Tc-99m]Tc-HYNIC-(D)A7R showed better tumor-to-muscle ratios and lower renal uptake than [Tc-99m]Tc-HYNIC-A7R. These results suggest that both [Tc-99m]Tc-HYNIC-(D)A7R and [Tc-99m]Tc-HYNIC-A7R have substantial potential as probes for targeted SPECT imaging of TNBC, and the former may be considered for future clinical translation owing to its superior tumor imaging performance.

Automated summary

Novel heptapeptides, [Tc-99m]Tc-HYNIC-A7R and [Tc-99m]Tc-HYNIC-(D)A7R, show great potential as probes for targeted SPECT imaging of TNBC, with the latter demonstrating superior tumor imaging performance compared to the former.