TY - JOUR
T1 - Development of a time-resolved fluorescence probe for evaluation of competitive binding to the cholecystokinin 2 receptor
AU - Elshan, N. G.R.Dayan
AU - Jayasundera, Thanuja
AU - Weber, Craig S.
AU - Lynch, Ronald M.
AU - Mash, Eugene A.
N1 - Funding Information:
The authors thank Renata Patek and Professor Josef Vagner for assistance and useful discussions. The HEK-293 cell line overexpressing hCCK2R and hMC4R was the generous gift of Professor Robert J. Gillies, Professor David L. Morse, and Dr. Liping Xu of the H. Lee Moffitt Cancer Center and Research Institute, Tampa, FL, USA. This work was supported by grants R33 CA95944 , RO1 CA97360 , RO1 CA123547 , and P30 CA23074 from the National Cancer Institute .
Publisher Copyright:
© 2015 Elsevier Ltd.All rights reserved.
PY - 2015/4/15
Y1 - 2015/4/15
N2 - The synthesis, characterization, and use of Eu-DTPA-PEGO-Trp-Nle-Asp-Phe-NH2 (Eu-DTPA-PEGO-CCK4), a luminescent probe targeted to cholecystokinin 2 receptor (CCK2R, aka CCKBR), are described. The probe was prepared by solid phase synthesis. A Kd value of 17 ± 2 nM was determined by means of saturation binding assays using HEK-293 cells that overexpress CCK2R. The probe was then used in competitive binding assays against Ac-CCK4 and three new trivalent CCK4 compounds. Repeatable and reproducible binding assay results were obtained. Given its ease of synthesis, purification, receptor binding properties, and utility in competitive binding assays, Eu-DTPA-PEGO-CCK4 could become a standard tool for high-throughput screening of compounds in development targeted to cholecystokinin receptors.
AB - The synthesis, characterization, and use of Eu-DTPA-PEGO-Trp-Nle-Asp-Phe-NH2 (Eu-DTPA-PEGO-CCK4), a luminescent probe targeted to cholecystokinin 2 receptor (CCK2R, aka CCKBR), are described. The probe was prepared by solid phase synthesis. A Kd value of 17 ± 2 nM was determined by means of saturation binding assays using HEK-293 cells that overexpress CCK2R. The probe was then used in competitive binding assays against Ac-CCK4 and three new trivalent CCK4 compounds. Repeatable and reproducible binding assay results were obtained. Given its ease of synthesis, purification, receptor binding properties, and utility in competitive binding assays, Eu-DTPA-PEGO-CCK4 could become a standard tool for high-throughput screening of compounds in development targeted to cholecystokinin receptors.
KW - Cholecystokinin 2 receptor
KW - Dissociation-enhanced lanthanide
KW - Fluoroimmunoassay
KW - High-throughput screening
KW - Multivalent binding
KW - Time-resolved fluorescence
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U2 - 10.1016/j.bmc.2015.02.028
DO - 10.1016/j.bmc.2015.02.028
M3 - Article
C2 - 25769518
AN - SCOPUS:84925682601
SN - 0968-0896
VL - 23
SP - 1841
EP - 1848
JO - Bioorganic and Medicinal Chemistry
JF - Bioorganic and Medicinal Chemistry
IS - 8
ER -