Preclinical evaluation of a dual-receptor targeted tracer [ 68 Ga]Ga-HX01 in 10 different subcutaneous and orthotopic tumor models

Purpose The integrin α v β 3 and aminopeptidase N (APN/CD13) play vital roles in the tumor angiogenesis process. They are highly expressed in a variety of tumor cells and proliferating endothelial cells during angiogenesis, which have been considered as highly promising targets for tumor imaging. Arginine-glycine-aspartic (RGD) and asparagine-glycine-arginine (NGR) are two peptides specifically binding to the integrin α v β 3 and CD13, respectively. In this study, we optimized our previously developed probe and preclinically evaluated the new integrin α v β 3 and CD13 dual-targeted probe, NOTA-RGD-NGR (denoted as HX01) radiolabeled with 68 Ga, in 10 different subcutaneous and orthotopic tumor models. Methods The specific activity and radiochemical purity of [ 68 Ga]Ga-HX01 were identified. The dual-receptor targeting ability was confirmed by a series of blocking studies and partly muted tracers using BxPC-3 xenograft model. The dynamic imaging study and dose escalation study were explored to determine the optimal imaging time point and dosage in the BxPC-3 xenograft model. Next, we established a variety of subcutaneous and orthotopic tumor models including pancreas (BxPC-3), breast (MCF-7), gallbladder (NOZ), lung (HCC827), ovary (SK-OV-3), colorectal (HCT-8), liver (HuH-7), stomach (NUGC-4), and glioma (U87) cancers. All models underwent [ 68 Ga]Ga-HX01 PET/CT imaging about 2 weeks post-inoculation, with a subset of them undergoing [ 18 F]FDG PET/CT scan performed concurrently, and their results were compared. In addition, ex vivo biodistribution studies were also performed for verifying the semi-quantitative results of the non-invasive PET images. Results [ 68 Ga]Ga-HX01 significantly outperformed single target probes in the BxPC-3 xenograft model. All blocking and single target groups exhibited significantly descending tumor uptake. The high tumor uptakes were found in BxPC-3, MCF-7, and NOZ subcutaneous tumors (%ID/g > 1.1), while middle uptakes were observed in HCC827, SK-OV-3, HCT-8, and HuH-7 subcutaneous tumor (%ID/g 0.7–1.0). Due to the low background, the tumor-to-muscle and tumor-to-blood ratios of [ 68 Ga]Ga-HX01 were higher than that of [ 18 F]FDG. Conclusions [ 68 Ga]Ga-HX01, as a dual target imaging agent, exhibited superior in vivo performance in different subcutaneous and orthotopic mice models of human tumors over [ 18 F]FDG and its respectively mono-receptor targeted agents, which warrants the future clinical translation for tumor imaging.