Preclinical Evaluation of Molecular-Targeted Ultrasound Contrast Agents in a Rabbit Model

Abstract

The localization of prostate cancer by ultrasound remains limited by the lack of B-mode conspicuity and the confinement of clinically approved microbubbles (MBs) to the vasculature. This precludes the differentiation of viable tumor tissue, necrotic tissue, and margin-associated disease. We investigate the use of prostate-specific membrane antigen (PSMA)-targeted lipid-shelled perfluorocarbon nanobubbles (PSMA-NBs) in an orthotopic rabbit model using a clinical CEUS system. We implanted PSMA-positive PC3pip-GFP tumors into the prostates of immunosuppressed New Zealand White rabbits and performed transabdominal imaging with PSMA-NBs, MBs, and untargeted nanobubbles (Plain-NB) using the same imaging and analysis protocols. To address tumor heterogeneity and ultrasound boundary ambiguity, ROIs were defined from baseline B-mode images and segmented into the tumor core, rim, and a surrounding peritumoral area. Pixel-wise parametric and decorrelation time (DT) maps were generated and correlated with whole-slide histology (H&E) and PSMA IHC. Compared to MBs, PSMA-NBs exhibited higher peak intensities in the tumor core and rim (1.60-fold and 1.50-fold, respectively) and improved retention (mean transit time: 4.20 to 4.50-fold higher) for up to 10 minutes in the tumor and peritumoral areas. PSMA-NB kinetic and DT parameters also correlated with histology-defined tumor viability and PSMA expression. Compared to Plain-NBs, PSMA-NBs also exhibited improved retention (MTT +21%; AUCwo) in the rim and peritumoral areas. This study demonstrates the capability of PSMA-NBs to characterize prostate cancer by molecular CEUS beyond what is possible with conventional MBs.