Considerations for the use of targeted fluorescence contrast agents to detect circulating cancer cell populations with diffuse

Significance: Metastasis is a leading cause of cancer-related deaths. Disseminated circulating tumor cells (CTCs) through the bloodstream seed metastatic tumors at distant sites. Most methods for enumerating CTCs in humans clinically rely on drawing and analyzing small blood samples, but these may yield inaccurate estimates of CTC burden and cannot measure CTC changes over time. Identification and enumeration of CTCs for experimental or clinical purposes largely rely on marker-driven analyses by flow cytometry. Aim: In principle, noninvasive fluorescence enumeration of CTCs directly Approach: We evaluated a clinical small-molecule folate receptor-targeted molecular imaging agent (OTL38, pafolacianine), a fluorogenic pan-cathepsin imaging agent (VGT-309, abenacianine), and a set of custom-designed, small-molecule prostate-specific membrane antigen (PSMA)-targeted fluorophores. We tested these contrast agents using Results: All tested contrast agents showed not only high uptake and labeling by target cell lines but also small but significant labeling of non-cancer blood cells. Contrast agents that exhibited rapid clearance from circulation and the fluorogenic approach resulted in significantly reduced non-specific interfering background fluorescence signals. Conclusions: Although all of the tested targeted fluorescence contrast agents have properties useful for labeling of CTCs, thus far, none has exhibited the required high specificity. This resulted in some labeling of non-cancer blood cells, which presented false-positive CTC counts. Improved contrast agent design and multiplexed use of more than one contrast agent may improve this specificity.