A quantitative chemical proteomics approach to profile the specific cellular targets of andrographolide, a promising anticancer agent that suppresses tumor metastasis.
Drug target identification is a critical step toward understanding the mechanism of action of a drug, which can help one improve the drug's current therapeutic regime and expand the drug's therapeutic potential. However, current in vitro affinity-chromatography-based and in vivo activity-based protein profiling approaches generally face difficulties in discriminating specific drug targets from nonspecific ones. Here we describe a novel approach combining isobaric tags for relative and absolute quantitation with clickable activity-based protein profiling to specifically and comprehensively identify the protein targets of andrographolide (Andro), a natural product with known anti-inflammation and anti-cancer effects, in live cancer cells. We identified a spectrum of specific targets of Andro, which furthered our understanding of the mechanism of action of the drug. Our findings, validated through cell migration and invasion assays, showed that Andro has a potential novel application as a tumor metastasis inhibitor. Moreover, we have unveiled the target binding mechanism of Andro with a combination of drug analog synthesis, protein engineering, and mass-spectrometry-based approaches and determined the drug-binding sites of two protein targets, NF-κB and actin.
Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore, 117543;
Jigang
J
Wang
Xing Fei
XF
Tan
Van Sang
VS
Nguyen
Peng
P
Yang
Jing
J
Zhou
Mingming
M
Gao
Zhengjun
Z
Li
Teck Kwang
TK
Lim
Yingke
Y
He
Chye Sun
CS
Ong
Yifei
Y
Lay
Jianbin
J
Zhang
Guili
G
Zhu
Siew-Li
SL
Lai
Dipanjana
D
Ghosh
Yu Keung
YK
Mok
Han-Ming
HM
Shen
Qingsong
Q
Lin
D000970:Antineoplastic Agents / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology / Q000627:therapeutic use*; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003545:Cysteine / Q000378:metabolism; D004224:Diterpenes / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D006801:Humans; D015335:Molecular Probes / Q000138:chemical synthesis / Q000737:chemistry; D016328:NF-kappa B / Q000378:metabolism; D009361:Neoplasm Invasiveness; D009362:Neoplasm Metastasis / Q000188:drug therapy*; D011485:Protein Binding / Q000187:drug effects; D020543:Proteome / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D015398:Signal Transduction / Q000187:drug effects
D000970:Antineoplastic Agents; D004224:Diterpenes; D015335:Molecular Probes; D016328:NF-kappa B; D020543:Proteome; C030419:andrographolide; D003545:Cysteine