Pharmacological evaluation of uPAR expression by 68Ga-PET

Objectives The urokinase plasminogen activator receptor (uPAR) has been implicated in tumor growth, invasion, and metastasis, making it an attractive target for early detection of primary and metastatic tumors. Previously, cyclotron-produced radiometals have been used to labeled uPAR-targeting peptides but require delayed imaging time points to allow for blood clearance and attain optimal contrast. Since 68Ga-labeled peptides have shown rapid clearance from circulation, this study focused on radiolabeling of a high-affinity uPAR-targeting peptide with 68Ga, and pharmacological evaluation of the agent in a mouse melanoma model.

Methods DOTA was conjugated to a linear uPAR-targeting peptide (AE105) by solid phase peptide synthesis to yield U1. 68Ga labeling was performed in sodium acetate (pH 4) using the 2 ml peak fraction and heated at 95oC for 10 mins. In vitro studies in B16F10 melanoma cells were conducted to evaluate binding kinetics and tracer specificity. Nude mice bearing B16F10 tumors were injected with 68Ga-U1 and imaged by PET/CT.

Results Routine radiolabeling resulted in >99% radiochemical purity as determined by radio-HPLC. In vitro assays demonstrated rapid tracer association with the cells as early as 30 minutes with moderate increase over time. Blocking studies showed a decreased uptake of 68Ga-U1 in response to increasing concentrations of competitive inhibitor. Imaging studies were performed and the optimal imaging time point selected was 1 hr post-injection due to high renal clearance and low background signal. 68Ga-U1 uptake was evident in all tumors and tumor-to-background ratios ranged from 3-4.

Conclusions A 68Ga-radiopeptide was developed and evaluated for imaging uPAR-expressing tumors. Highly-selective targeting was observed and further evaluation in human melanoma cells is underway to determine potential usefulness of the agent for detecting primary and metastatic tumors