Inhibition of the redox function of APE1/Ref-1 in myeloid leukemia cell lines results in a hypersensitive response to retinoic acid-induced differentiation and apoptosis.
OBJECTIVE
The standard of care for promyelocytic leukemia includes use of the differentiating agent all-trans retinoic acid (RA) and chemotherapy. RA induces cell differentiation through retinoic acid receptor (RAR) transcription factors. Because redox mechanisms influence how readily transcription factors bind to DNA response elements (RARE), the impact of small molecule (E3330) inhibition of the redox regulatory protein, apurinic-apyrimidinic endonuclease/redox effector factor (APE1/Ref-1) on RAR DNA binding and function in RA-induced myeloid leukemia cell differentiation and apoptosis was investigated.


METHODS
The redox function of APE1 was studied using the small molecule inhibitor E3330 in HL-60 and PLB acute myeloid leukemia cells. Electrophoretic mobility shift assays were employed to determine effect of inhibitor on APE1/Ref-1 redox signaling function. Trypan blue assays, Annexin-V/propidium iodide and CD11b staining, and real-time polymerase chain reaction analyses were employed to determine survival, apoptosis, and differentiation status of cells in culture.


RESULTS
RARα binds to its RARE in a redox-dependent manner mediated by APE1/Ref-1 redox regulation. Redox-dependent RAR-RARE binding is blocked by E3330, a small molecule redox inhibitor of APE1/Ref-1. Combination treatment of RA + E3330 results in a profound hypersensitivity of myeloid leukemia cells to RA-induced differentiation and apoptosis. Additionally, redox inhibition by E3330 results in enhanced RAR target gene, BLR-1, expression in myeloid leukemia cells.


CONCLUSIONS
The redox function of APE1/Ref-1 regulates RAR binding to its DNA RAREs influencing the response of myeloid leukemia cells to RA-induced differentiation. Targeting of APE1/Ref-1 redox function may allow manipulation of the retinoid response with therapeutic implications.
Fishel
Melissa L
ML
Department of Pediatrics (Section of Hematology/Oncology), Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, 980 W. Walnut, Indianapolis, IN 46202, USA. mfishel@iupui.edu
Colvin
E Scott
ES
Luo
Meihua
M
Kelley
Mark R
MR
Robertson
Kent A
KA
D017209:Apoptosis / Q000187:drug effects*; D016227:Benzoquinones / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000037:antagonists & inhibitors* / Q000502:physiology; D006801:Humans; D007951:Leukemia, Myeloid / Q000188:drug therapy* / Q000473:pathology; D010084:Oxidation-Reduction; D011422:Propionates / Q000494:pharmacology; D054380:Receptors, CXCR5 / Q000032:analysis; D018168:Receptors, Retinoic Acid / Q000378:metabolism; D020218:Response Elements; D000072482:Retinoic Acid Receptor alpha; D014212:Tretinoin / Q000494:pharmacology*
D016227:Benzoquinones; C095085:CXCR5 protein, human; D011422:Propionates; C000607071:RARA protein, human; D054380:Receptors, CXCR5; D018168:Receptors, Retinoic Acid; D000072482:Retinoic Acid Receptor alpha; C075569:E 3330; D014212:Tretinoin; C085108:APEX1 protein, human; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase
Oncogene. 2005 Nov 3;24(48):7145-55
16044154
Semin Hematol. 2001 Jan;38(1):13-25
11172536
J Immunol. 1998 Jan 15;160(2):810-9
9551916
Antioxid Redox Signal. 2009 Mar;11(3):601-20
18976116
Gene. 1994 Aug 5;145(2):197-203
8056331
Blood. 2009 Feb 26;113(9):1875-91
18812465
Circulation. 2005 Feb 15;111(6):782-90
15699255
Mol Cancer Ther. 2003 Jul;2(7):633-40
12883036
Cell. 2001 Mar 9;104(5):719-30
11257226
Blood. 2003 Jul 1;102(1):237-45
12586626
Best Pract Res Clin Haematol. 2007 Mar;20(1):57-65
17336255
Gene. 2004 Mar 17;328:1-16
15019979
FEBS Lett. 2008 Jul 23;582(17):2508-14
18571505
J Mol Med (Berl). 2009 Aug;87(8):785-91
19424677
Endocrinology. 2001 Jun;142(6):2600-5
11356710
Antioxid Redox Signal. 2010 Jun 1;12(11):1247-69
19764832
Cancer Res. 2007 Aug 15;67(16):7765-72
17699781
J Biol Chem. 2000 Apr 28;275(17):12470-4
10777532
Cell Growth Differ. 1997 Apr;8(4):443-9
9101090
Leukemia. 2007 Jul;21(7):1363-77
17443228
Nat Biotechnol. 2000 Aug;18(8):877-81
10932159
Blood. 2009 Jan 22;113(4):775-83
18945964
Blood. 1991 Jan 15;77(2):340-7
1670759
Nat Rev Drug Discov. 2007 Oct;6(10):793-810
17906642
J Biol Chem. 2008 Feb 15;283(7):4375-86
18006504
Mutat Res. 2008 Aug 25;643(1-2):54-63
18579163
Mol Endocrinol. 2007 Feb;21(2):359-75
17138644
Mol Cell Biol. 2007 Jun;27(11):4179-97
17325034
Antioxid Redox Signal. 2008 Nov;10(11):1853-67
18627350
Ann N Y Acad Sci. 2006 Dec;1090:203-8
17384263
Leukemia. 2003 Jan;17(1):98-113
12529666
Genes Dev. 2009 Mar 15;23(6):681-93
19299558
Blood. 2007 Mar 1;109(5):1917-22
17053053
Best Pract Res Clin Haematol. 2001 Mar;14(1):153-74
11355929
Biochem Biophys Res Commun. 2009 Dec 11;390(2):241-6
19799861
Curr Drug Targets Immune Endocr Metabol Disord. 2001 Aug;1(2):153-64
12476796
Biol Pharm Bull. 2006 Sep;29(9):1803-9
16946489
Exp Cell Res. 2000 Feb 1;254(2):287-98
10640427
Eur J Cell Biol. 2001 Jan;80(1):59-67
11211936