TY - JOUR
T1 - Quantum Dot-Based Screening System for Discovery of G Protein-Coupled Receptor Agonists
AU - Lee, Junghan
AU - Kwon, Yong Jun
AU - Choi, Youngseon
AU - Kim, Hi Chul
AU - Kim, Keumhyun
AU - Kim, Jinyeop
AU - Park, Sun
AU - Song, Rita
PY - 2012/7/9
Y1 - 2012/7/9
N2 - Cellular imaging has emerged as an important tool to unravel biological complexity and to accelerate the drug-discovery process, including cell-based screening, target identification, and mechanism of action studies. Recently, semiconductor nanoparticles known as quantum dots (QDs) have attracted great interest in cellular imaging applications due to their unique photophysical properties such as size, tunable optical property, multiplexing capability, and photostability. Herein, we show that QDs can also be applied to assay development and eventually to high-throughput/content screening (HTS/HCS) for drug discovery. We have synthesized QDs modified with PEG and primary antibodies to be used as fluorescent probes for a cell-based HTS system. The G protein-coupled receptor (GPCR) family is known to be involved in most major diseases. We therefore constructed human osteosarcoma (U2OS) cells that specifically overexpress two types of differently tagged GPCRs: influenza hemagglutinin (HA) peptide-tagged κ-opioid receptors (κ-ORs) and GFP-tagged A3 adenosine receptors (A3AR). In this study, we have demonstrated that 1) anti-HA antibody-conjugated QDs could specifically label HA-tagged κ-ORs, 2) subsequent treatment of QD-tagged GPCR agonists allowed agonist-induced translocation to be monitored in real time, 3) excellent emission spectral properties of QD permitted the simultaneous detection of two GPCRs in one cell, and 4) the robust imaging capabilities of the QD-antibody conjugates could lead to reproducible quantitative data from high-content cellular images. These results suggest that the present QD-based GPCR inhibitor screening system can be a promising platform for further drug screening applications.
AB - Cellular imaging has emerged as an important tool to unravel biological complexity and to accelerate the drug-discovery process, including cell-based screening, target identification, and mechanism of action studies. Recently, semiconductor nanoparticles known as quantum dots (QDs) have attracted great interest in cellular imaging applications due to their unique photophysical properties such as size, tunable optical property, multiplexing capability, and photostability. Herein, we show that QDs can also be applied to assay development and eventually to high-throughput/content screening (HTS/HCS) for drug discovery. We have synthesized QDs modified with PEG and primary antibodies to be used as fluorescent probes for a cell-based HTS system. The G protein-coupled receptor (GPCR) family is known to be involved in most major diseases. We therefore constructed human osteosarcoma (U2OS) cells that specifically overexpress two types of differently tagged GPCRs: influenza hemagglutinin (HA) peptide-tagged κ-opioid receptors (κ-ORs) and GFP-tagged A3 adenosine receptors (A3AR). In this study, we have demonstrated that 1) anti-HA antibody-conjugated QDs could specifically label HA-tagged κ-ORs, 2) subsequent treatment of QD-tagged GPCR agonists allowed agonist-induced translocation to be monitored in real time, 3) excellent emission spectral properties of QD permitted the simultaneous detection of two GPCRs in one cell, and 4) the robust imaging capabilities of the QD-antibody conjugates could lead to reproducible quantitative data from high-content cellular images. These results suggest that the present QD-based GPCR inhibitor screening system can be a promising platform for further drug screening applications.
KW - Agonists
KW - G protein-coupled receptors
KW - High-throughput screening
KW - Kappa-opioid receptor
KW - Quantum dots
UR - http://www.scopus.com/inward/record.url?scp=84863447547&partnerID=8YFLogxK
UR - https://pubmed.ncbi.nlm.nih.gov/22692979
U2 - 10.1002/cbic.201200128
DO - 10.1002/cbic.201200128
M3 - Article
C2 - 22692979
AN - SCOPUS:84863447547
SN - 1439-4227
VL - 13
SP - 1503
EP - 1508
JO - ChemBioChem
JF - ChemBioChem
IS - 10
ER -