In vivo imaging of tumors with protease-activated near-infrared fluorescent probes

Abstract

We have developed a method to image tumor-associated lysosomal protease activity in a xenograft mouse model in vivo using autoquenched near-infrared fluorescence (NIRF) probes. NIRF probes were bound to a long circulating graft copolymer consisting of poly-L-lysine and methoxypolyethylene glycol succinate. Following intravenous injection, the NIRF probe carrier accumulated in solid tumors due to its long circulation time and leakage through tumor neovasculature. Intratumoral NIRF signal was generated by lysosomal proteases in tumor cells that cleave the macromolecule, thereby releasing previously quenched fluorochrome. In vivo imaging showed a 12-fold increase in NIRF signal, allowing the detection of tumors with submillimeter-sized diameters. This strategy can be used to detect such early stage tumors in vivo and to probe for specific enzyme activity.

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Figure 1: (A) Schematic diagram of probe activation.
Figure 2: Fluorescence microscopy of LX-1 cells incubated with C-PGC (magnification 250×).
Figure 3: LX-1 tumor implanted into the mammary fat pad of a nude mouse.
Figure 4: NIRF histology of tumor excised from animal shown in Figure 3 (magnification 200×).

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Acknowledgements

We thank Anna Moore for providing all cell cultures and Lee Josephson and Michael Feld for valuable discussion. This study was supported, in part, by RO1 CA74424-01.

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Correspondence to Ralph Weissleder.

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Weissleder, R., Tung, C., Mahmood, U. et al. In vivo imaging of tumors with protease-activated near-infrared fluorescent probes. Nat Biotechnol 17, 375–378 (1999). https://doi.org/10.1038/7933

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