A high throughput drug screen based on fluorescence resonance energy transfer (FRET) for anticancer activity of compounds from herbal medicine.
OBJECTIVE
We report the development of a very efficient cell-based high throughput screening (HTS) method, which utilizes a novel bio-sensor that selectively detects apoptosis based on the fluorescence resonance energy transfer (FRET) technique.


METHODS
We generated a stable HeLa cell line expressing a FRET-based bio-sensor protein. When cells undergo apoptosis, they activate a protease called 'caspase-3'. Activation of this enzyme will cleave our sensor protein and cause its fluorescence emission to shift from a wavelength of 535 nm (green) to 486 nm (blue). A decrease in the green/blue emission ratio thus gives a direct indication of apoptosis. The sensor cells are grown in 96-well plates. After addition of different chemical compounds to each well, a fluorescence profile can be measured at various time-points using a fluorescent plate reader. Compounds that can trigger apoptosis are potential candidates as anti-cancer drugs.


RESULTS
This novel cell-based HTS method is highly effective in identifying anti-cancer compounds. It was very sensitive in detecting apoptosis induced by various known anti-cancer drugs. Further, this system detects apoptosis, but not necrosis, and is thus more useful than the conventional cell viability assays, such as those using MTT. Finally, we used this system to screen compounds, isolated from two plants used in Chinese medicine, and identified several effective compounds for inducing apoptosis.


CONCLUSIONS
This FRET-based HTS method is a powerful tool for identifying anti-cancer compounds and can serve as a highly efficient platform for drug discovery.
Tian
H
H
Bioengineering Graduate Program, Department of Chemical Engineering, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong.
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Luo
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Chang
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DC
Luo
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D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D015374:Biosensing Techniques / Q000379:methods; D004354:Drug Screening Assays, Antitumor / Q000379:methods*; D004365:Drugs, Chinese Herbal / Q000494:pharmacology*; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D006367:HeLa Cells; D006801:Humans; D011035:Podophyllum; D027561:Salvia miltiorrhiza
D000970:Antineoplastic Agents; D004365:Drugs, Chinese Herbal