Fluorescence resonance energy transfer-based biosensors for monitoring prostate specific antigen

Abstract

Prostate cancer has become the most commonly diagnosed cancer in men in the United States. Clinical diagnostic procedures currently include prostate-specific antigen (PSA) screening, digital rectal exam, and prostatic needle biopsy. However, these methods lack the sensitivity to detect small lesions that occur in the early stages of cancer and metastasis. I propose a molecular imaging modality that provides a biochemical characterization of localized regions of prostate tissue. Using fluorescence resonance energy transfer (FRET), several peptide substrates have been designed to respond to varying concentrations of PSA with a concomitant increase in fluorescence. In the near-infrared wavelength range, these fluorescent substrates can be imaged through thin sections of tissue to allow surface volume imaging of biochemical function, and thus, to provide additional insight into prostate cancer localization and progression. The goal of this study was to develop novel fluorescent substrates for prostate-specific antigen to serve as indicators of prostate cancer progression. PSA is a biomolecular marker that has gained widespread clinical use in prostate cancer detection. Produced primarily by prostate epithelium, PSA is an androgen-regulated serine protease that acts to cleave semenogelins. Several peptide substrates for PSA have been identified and optimized for specific and efficient hydrolysis. Two of these substrates, QFYSSN and SSIYSQTEEQ were modified with fluorescent dye and quencher molecules to suppress fluorescence in the inactivated form. Light absorbed by the fluorescent molecule is dissipated via nonradiative interaction with the quencher molecule.

(cont.) Disruption of dye and quencher interaction, as in substrate proteolysis, results in an increase in fluorescence. I report several promising substrates that generate significant increases in fluorescence upon cleavage by PSA in purified systems as well as with human prostate cancer cell lines. Selected FRET substrates can distinguish between PSA- producing and non-PSA-producing human prostate cancer cells.