Labeling of oxidizable proteins with a photoactivatable analog of the antitumor agent DMXAA: evidence for redox signaling in its mode of action.
The signaling pathway(s) and molecular target(s) for 5,6-dimethylxanthenone-4-acetic acid (DMXAA), a tumor vascular disrupting agent in late stages of clinical development, are still undefined. As an approach toward identifying potential targets for DMXAA, a tritiated azido-analog of DMXAA was used to probe for cellular binding proteins. More than 20 cytosolic proteins from murine splenocytes, RAW 264.7 cells, and the HECPP immortalized endothelial cells were photoaffinity-labeled. Although no protein domain, fold, or binding site for a specific ligand was found to be shared by all the candidate proteins, essentially all were noted to be oxidizable proteins, implicating a role for redox signaling in the action of DMXAA. Consistent with this hypothesis, DMXAA caused an increase in concentrations of reactive oxygen species (ROS) in RAW264.7 cells during the first 2 hours. This increase in ROS was suppressed in the presence of the antioxidant, N-acetyl-L-cysteine, which also suppressed DMXAA-induced cytokine production in the RAW 264.7 cells with no effects on cell viability. Short interfering RNA (siRNA)-mediated knockdown of one of the photoaffinity-labeled proteins, superoxide dismutase 1, an ROS scavenger, resulted in an increase in tumor necrosis factor-alpha production by RAW 264.7 cells in response to DMXAA compared with negative or positive controls transfected with nontargeting or lamin A/C-targeting siRNA molecules, respectively. The results from these lines of study all suggest that redox signaling plays a central role in cytokine induction by DMXAA.
Brauer
Romy
R
Auckland Cancer Society Research Centre, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand.
Wang
Liang-Chuan S
LC
Woon
See-Tarn
ST
Bridewell
David J A
DJ
Henare
Kimiora
K
Malinger
Dieter
D
Palmer
Brian D
BD
Vogel
Stefanie N
SN
Kieda
Claudine
C
Tijono
Sofian M
SM
Ching
Lai-Ming
LM
D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology; D014408:Biomarkers, Tumor / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D002478:Cells, Cultured; D015180:Electrophoresis, Gel, Two-Dimensional; D042783:Endothelial Cells / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009389:Neovascularization, Pathologic / Q000378:metabolism / Q000473:pathology; D010084:Oxidation-Reduction; D019920:Photoaffinity Labels / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D010777:Photochemistry; D011506:Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D013154:Spleen / Q000166:cytology / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D013194:Staining and Labeling / Q000379:methods*; D044004:Xanthones / Q000737:chemistry / Q000494:pharmacology*
D000970:Antineoplastic Agents; D014408:Biomarkers, Tumor; D019920:Photoaffinity Labels; D011506:Proteins; D034741:RNA, Small Interfering; D044004:Xanthones; C066668:vadimezan
Antioxid Redox Signal. 2005 Jul-Aug;7(7-8):964-72
15998251
Neoplasia. 2009 Aug;11(8):793-803
19649209
Oncol Res. 2002;13(2):95-101
12392157
J Biol Chem. 2004 Mar 5;279(10):9298-305
14613939
J Natl Cancer Inst. 1989 Jul 5;81(13):1005-13
2733044
Biochem Biophys Res Commun. 2005 Jul 1;332(2):362-9
15910747
J Med Chem. 1991 Jan;34(1):217-22
1992120
Exp Mol Pathol. 2007 Dec;83(3):347-56
17936751
Mol Pharmacol. 2006 Feb;69(2):501-8
16288082
Free Radic Biol Med. 1993 Oct;15(4):385-94
8225020
Bioorg Med Chem. 2007 Jan 15;15(2):605-15
17070061
Biol Cell. 1993;79(1):27-35
8118408
Br J Cancer. 2002 Aug 12;87(4):465-70
12177785
Nat Rev Cancer. 2005 Jun;5(6):423-35
15928673
Cancer Res. 2005 Dec 15;65(24):11752-61
16357188
J Biol Chem. 2004 Oct 1;279(40):41352-60
15292244
Eur J Cancer. 2001 Oct;37(15):1930-7
11576850
Electrophoresis. 1999 Dec;20(18):3551-67
10612281
Anal Biochem. 1976 May 7;72:248-54
942051
Br J Cancer. 2002 Jun 17;86(12):1937-42
12085190
Oncol Res. 2006;16(1):1-14
16783963
Invest New Drugs. 1986;4(3):207-20
3546183
Biochim Biophys Acta. 1966 Mar 28;117(1):115-33
5914328
Eur J Cancer. 2003 May;39(8):1176-83
12736120
Cancer Res. 1990 Sep 1;50(17):5537-42
2386959
J Exp Med. 2007 Jul 9;204(7):1559-69
17562815
Proteomics. 2003 Jul;3(7):1154-61
12872216
Eur J Cancer Clin Oncol. 1989 Feb;25(2):263-9
2702981
J Med Chem. 2007 Aug 9;50(16):3757-64
17616114
J Interferon Cytokine Res. 2008 Mar;28(3):133-9
18338946
Arch Biochem Biophys. 2002 Oct 15;406(2):229-40
12361711
Curr Med Chem Anticancer Agents. 2005 Sep;5(5):501-6
16178775
Biochem J. 2005 Aug 1;389(Pt 3):785-95
15801906
J Natl Cancer Inst. 1990 Mar 21;82(6):528-9
2313725
J Neurochem. 1994 Sep;63(3):1126-32
8051554
Biochim Biophys Acta. 2004 Dec 6;1742(1-3):191-201
15590070
Hear Res. 2001 Sep;159(1-2):14-22
11520631
J Immunol. 1999 Jun 15;162(12):7335-42
10358184
Cardiovasc Res. 2005 Oct 1;68(1):26-36
16009356
J Natl Cancer Inst. 1990 Jul 4;82(13):1107-12
2359136
J Biol Chem. 2001 Aug 10;276(32):30188-98
11402028
Proc Natl Acad Sci U S A. 1988 Oct;85(19):7187-91
2902625
Biochem Pharmacol. 2006 Nov 30;72(11):1493-505
16723122
Int J Cancer. 2006 Jul 1;119(1):41-8
16450384
Cancer Res. 2007 Jul 15;67(14):7011-9
17638914
Biochem Pharmacol. 2004 Mar 1;67(5):937-45
15104247
Proteomics. 2003 Apr;3(4):479-95
12687615
Cancer Res. 1988 Apr 15;48(8):2179-83
3349488
Proc Natl Acad Sci U S A. 2002 Mar 19;99(6):3505-10
11904414