Proteomic (antibody microarray) exploration of the molecular mechanism of action of the specific COX-2 inhibitor DuP 697

Our plain-English summary of Proteomic (antibody microarray) exploration of the molecular mechanism of action of the specific COX-2 inhibitor DuP 697, by V. Agarwal, V.C. Hodgkinson, G.L. Eagle, et al (International Journal of Oncology, 2013).

The study

• Authors: V. Agarwal, V.C. Hodgkinson, G.L. Eagle, L. Scaife, M.J. Lind, L. Cawkwell
• Journal: International Journal of Oncology
• Published: 2013;42(3):1088-92
• Cited by: 3 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

We have previously shown that specific COX-2 inhibitors, including DuP 697, have anti-proliferative effects on mesothelioma cells and potentiate the cytotoxicity of pemetrexed. Here, we used a novel proteomic approach to explore the mechanism of action of this agent. COX-2-positive cell lines MSTO-211H (mesothelioma) and A549 (lung cancer) were exposed to DuP 697 for 72 h. Drug carrier only was added to control cells. Extracted proteins from treated and control cells were analysed using a comparative proteomic platform. Differentially expressed proteins, identified by the Panorama Xpress Profiler725 antibody microarray were submitted to Ingenuity Pathway Analysis. A total of 32 unique differentially expressed proteins were identified with a significant (>1.8-fold) difference in expression between treated and untreated cells in at least one cell line. Five molecules, BCL2L1 (Bcl-xL), BID, CHUK (IKK), FASLG and RAF1, were mapped to the Apoptosis Signaling pathway following Ingenuity Pathway Analysis. BCL2L1 (Bcl-xL) and BID were analysed using immuno-blotting and differential expression was confirmed. Proteomic (antibody microarray) analysis suggests that the mechanism of action of DuP 697 may be exerted via the induction of apoptosis. The antibody microarray platform can be utilised to explore the molecular mechanism of action of novel anticancer agents.

In plain English

This was a laboratory study on cancer cells grown in the lab. It did not study people or asbestos exposure. The authors treated two cell lines, MSTO-211H (mesothelioma) and A549 (lung cancer), with DuP 697, a specific COX-2 inhibitor, for 72 hours. Control cells received only the drug carrier (DMSO). The authors compared proteins in treated and control cells using an antibody microarray. They found 32 proteins whose levels differed by at least 1.8-fold in at least one cell line, and 20 of these differed in both. Computer pathway analysis linked five of these proteins to an apoptosis (programmed cell death) signalling pathway. In follow-up immunoblotting, the protein Bcl-xL fell 2.48-fold in the mesothelioma cells and could not be detected in the lung cancer cells. Full-length BID fell 10.16-fold in the mesothelioma cells and 14.52-fold in the lung cancer cells. At 72 hours, no differences in caspases or in COX-2 itself were seen. The authors concluded that DuP 697 may act by inducing apoptosis, and said further studies are required.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) cell-line study using antibody microarray proteomics, computer pathway analysis and confirmatory immunoblotting
• Who was studied: Two human cancer cell lines: MSTO-211H (COX-2 positive mesothelioma) and A549 (lung cancer); no human participants
• Exposure: The drug DuP 697 (a specific COX-2 inhibitor): 31.7 µM for MSTO-211H cells and 50 µM for A549 cells, for 72 hours. Control cells received drug carrier (DMSO) only. No asbestos exposure was studied.
• Main results:
  • 32 unique proteins differed in expression by at least 1.8-fold between DuP 697-treated and control cells in at least one cell line
  • 20 of the 32 proteins differed by at least 1.8-fold in both cell lines
  • Pathway analysis identified 'Apoptosis Signaling' as the top pathway, involving 5 of the proteins: BCL2L1 (Bcl-xL), BID, CHUK (IKK), FASLG and RAF1
  • In immunoblotting, BCL2L1 (Bcl-xL) was down-regulated 2.48-fold in the MSTO-211H mesothelioma cells after DuP 697 treatment
  • BCL2L1 (Bcl-xL) could not be detected in the A549 lung cancer cells by immunoblotting
  • In immunoblotting, full-length BID was down-regulated 10.16-fold in MSTO-211H cells and 14.52-fold in A549 cells after DuP 697 treatment
  • At 72 hours, no differential expression of caspases 3 to 13 or pro-caspase 8 was found in either cell line
  • COX-2 expression itself did not change in either cell line after 72 hours of DuP 697 treatment
• The authors' conclusion: The authors concluded that their proteomic analysis suggests the anti-proliferative effect of DuP 697 in mesothelioma cell lines may be exerted via induction of apoptosis. They also said further studies are required to test whether COX-2 inhibitors may act as apoptosis sensitisers with chemotherapy.
• Limitations the authors state:
  • The antibody against truncated BID (tBID) used on the microarray was unreliable in immunoblotting, so tBID could not be confirmed
  • Only one time point (72 hours) was examined; caspase changes may have been missed because the proteins may have returned to basal levels by then
  • Some proteins (such as Zyxin) are repeatedly identified on this platform, so the choice of proteins for further analysis must be made carefully

Topics

The National Library of Medicine indexes this study under subjects our site covers: Lung cancer, Mesothelioma.

Medical terms in this study, explained in our Lexicon: Mesothelioma, Pemetrexed (Alimta), Lung Cancer, Apoptosis.

Related studies

• COX-2 specific inhibitors enhance the cytotoxic effects of pemetrexed in mesothelioma cell lines — S.L. O'Kane, G.L. Eagle, et al. (2010)
• Manumycin A induces apoptosis in malignant pleural mesothelioma through regulation of Sp1 and activation of the mitochondria-related apoptotic pathway — K.H. Kim, J.I. Chae, et al. (2016)
• Novel ATP-competitive Akt inhibitor afuresertib suppresses the proliferation of malignant pleural mesothelioma cells — M. Yamaji, A. Ota, et al. (2017)
• The flavonoid resveratrol suppresses growth of human malignant pleural mesothelioma cells through direct inhibition of specificity protein 1 — K.A. Lee, Y.J. Lee, et al. (2012)
• Antitumor effect and antiangiogenic potential of the mTOR inhibitor temsirolimus against malignant pleural mesothelioma — M. Moriya, T. Yamada, et al. (2014)
• Mechanisms of cisplatin-induced cell death in malignant mesothelioma cells: role of inhibitor of apoptosis proteins (IAPs) and caspases — I.L. Cregan, A.M. Dharmarajan, et al. (2013)

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