In vivo detection of phospholipase C by enzyme-activated near-infrared probes.

Abstract

In this article, the characterization of the first near-infrared (NIR) phospholipase-activated molecular beacon is reported, and its utility for in vivo cancer imaging is demonstrated. The probe consists of three elements: a phospholipid (PL) backbone to which the NIR fluorophore, pyropheophorbide a (Pyro), and the NIR Black Hole Quencher 3 (BHQ) were conjugated. Because of the close proximity of BHQ to Pyro, the Pyro-PtdEtn-BHQ probe is self-quenched until enzyme hydrolysis releases the fluorophore. The Pyro-PtdEtn-BHQ probe is highly specific to one isoform of phospholipase C, phosphatidylcholine-specific phospholipase C (PC-PLC), responsible for catabolizing phosphatidylcholine directly to phosphocholine. Incubation of Pyro-PtdEtn-BHQ in vitro with PC-PLC demonstrated a 150-fold increase in fluorescence that could be inhibited by the specific PC-PLC inhibitor tricyclodecan-9-yl xanthogenate (D609) with an IC(50) of 34 ± 8 μM. Since elevations in phosphocholine have been consistently observed by magnetic resonance spectroscopy in a wide array of cancer cells and solid tumors, we assessed the utility of Pyro-PtdEtn-BHQ as a probe for targeted tumor imaging. Injection of Pyro-PtdEtn-BHQ into mice bearing DU145 human prostate tumor xenografts followed by in vivo NIR imaging resulted in a 4-fold increase in tumor radiance over background and a 2 fold increase in the tumor/muscle ratio. Tumor fluorescence enhancement was inhibited with the administration of D609. The ability to image PC-PLC activity in vivo provides a unique and sensitive method of monitoring one of the critical phospholipase signaling pathways activated in cancer, as well as the phospholipase activities that are altered in response to cancer treatment.

• Authors: Theresa M Mawn, Anatoliy V Popov, Nancy J Beardsley, Klara Stefflova, Matthew Milkevitch, Gang Zheng, E James Delikatny
• Journal: Bioconjugate chemistry (Bioconjug Chem) Vol. 22 Issue 12 Pg. 2434-43 (Dec 21 2011) ISSN: 1520-4812 [Electronic] United States
• PMID: 22034913 (Publication Type: Journal Article, Research Support, N.I.H., Extramural)

Chemical References

• Fluorescent Dyes
• Phospholipids
• Type C Phospholipases

Topics

• Animals
• Cell Line, Tumor
• Enzyme Activation
• Fluorescent Dyes (chemistry)
• Humans
• Male
• Mice
• Mice, Nude
• Microscopy, Confocal
• Phospholipids (chemistry)
• Prostatic Neoplasms (enzymology)
• Type C Phospholipases (analysis, metabolism)