Radioiodinated inhibitors of cathepsin B as tumor imaging probes

Objectives Various proteases have been implicated in their roles in degrading the extracellular matrix, which is necessary for tumor invasion and metastasis. The protease cathepsin B (CTSB) is overexpressed in various cancers and its expression correlates with the aggressiveness of the disease, making it an attractive target for molecular imaging. Two radioactive probes 125I-CaBInIV-1 (1) and 125I-CaBInIV-3 (2) were developed and assessed for their ability to differentiate tumor types based on CTSB expression.

Methods The radioiodinated inhibitors were prepared by treating the fluorous tin precursors with [125I]NaI and iodogen. The probes were obtained in high purity and specific activity through fluorous solid phase extraction, eliminating the need to use HPLC purification. Probes were tested in vitro for protease binding in cell and tumor lysates derived from the breast cancer cell line MDA-MB231, as well as other cell lines that express different levels of CTSB. Biodistribution studies were preformed using MDA-MB231 tumor bearing mice.

Results The labeled inhibitors showed binding with pure CTSB and with the CTSB expressed in the cells as determined by SDS-PAGE. The binding of (2) was stronger than its lipophilic derivative (1), indicative of its improved binding affinity (181 nM vs 4 μM). In vivo, (1) exhibited rapid blood clearance and distributed primarily in the gall bladder and liver. The polar derivative (2) circulated longer, had lower liver uptake and was taken up into the gall bladder also, but to a much lesser extent. Thyroid uptake did increase over time for both compounds indicative of in vivo metabolism resulting in low tumor uptake (<1.1%).

Conclusions Radioiodinated inhibitors can be used to visualize CTSB in vitro. Structural modifications of the inhibitor resulted in enhanced blood circulation time and reduce liver accumulation in vivo. Higher tumor uptake can be expected with more aggressive cell lines and a more robust iodine containing prosthetic group. Challenges and strategies for advancing this class of probes will be presented.

Research Support Canadian Institutes of Health Research, Canadian Cancer Society, Ontario Institute for Cancer Research, Natural Sciences and Engineering Research Council of Canada