Molecular imaging of drug-modulated protein-protein interactions in living subjects

被引:87
作者
Paulmurugan, R
Massoud, TF
Huang, J
Gambhir, SS
机构
[1] Stanford Univ, Sch Med, Dept Radiol, Palo Alto, CA USA
[2] Stanford Univ, Sch Med, BioX Program, Palo Alto, CA USA
[3] Univ Calif Los Angeles, David Geffen Sch Med, Crump Inst Mol Imaging, Los Angeles, CA USA
[4] Univ Calif Los Angeles, David Geffen Sch Med, Dept Mol & Med Pharmacol, Los Angeles, CA USA
[5] Med Coll Wisconsin, Dept Radiol, Milwaukee, WI 53226 USA
[6] Univ Cambridge, Sch Clin Med, Dept Radiol, Cambridge, England
[7] Univ Cambridge, Sch Clin Med, Dept Oncol, Cambridge, England
关键词
D O I
10.1158/0008-5472.CAN-03-2972
中图分类号
R73 [肿瘤学];
学科分类号
100214 ;
摘要
Networks of protein interactions mediate cellular responses to environmental stimuli and direct the execution of many different cellular functional pathways. Small molecules synthesized within cells or recruited from the external environment mediate many protein interactions. The study of small molecule-mediated interactions of proteins is important to understand abnormal signal transduction pathways in cancer and in drug development and validation. In this study, we used split synthetic renilla luciferase (hRLUC) protein fragment-assisted complementation to evaluate heterodimerization of the human proteins FRB and FKBP12 mediated by the small molecule rapamycin. The concentration of rapamycin required for efficient dimerization and that of its competitive binder ascomycin required for dimerization inhibition were studied in cell lines. The system was dually modulated in cell culture at the transcription level, by controlling nuclear factor kappaB promoter/enhancer elements using tumor necrosis factor a, and at the interaction level, by controlling the concentration of the dimerizer rapamycin. The rapamycin-mediated dimerization of FRB and FKBP12 also was studied in living mice by locating, quantifying, and timing the hRLUC complementation-based bioluminescence imaging signal using a cooled charged coupled device camera. This split reporter system can be used to efficiently screen small molecule drugs that modulate protein-protein interactions and also to assess drugs in living animals. Both are essential steps in the preclinical evaluation of candidate pharmaceutical agents targeting protein-protein interactions, including signaling pathways in cancer cells.
引用
收藏
页码:2113 / 2119
页数:7
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