CRAF inhibition induces apoptosis in melanoma cells with non-V600E BRAF mutations.
Here, we identify a panel of melanoma lines with non-V600E mutations in BRAF. These G469E- and D594G-mutated melanomas were found to exhibit constitutive levels of phospho-extracellular signal-regulated kinase (pERK) and low levels of phospho-mitogen-activated protein kinase/ERK kinase (pMEK) and were resistant to MEK inhibition. Upon treatment with the CRAF inhibitor sorafenib, these lines underwent apoptosis and associated with mitochondrial depolarization and relocalization of apoptosis-inducing factor, whereas the BRAF-V600E-mutated melanomas did not. Studies have shown low-activity mutants of BRAF (G469E/D594G) instead signal through CRAF. Unlike BRAF, CRAF directly regulates apoptosis through mitochondrial localization where it binds to Bcl-2 and phosphorylates BAD. The CRAF inhibitor sorafenib was found to induce a time-dependent reduction in both BAD phosphorylation and Bcl-2 expression in the D594G/G469E lines only. Knockdown of CRAF using a lentiviral shRNA suppressed both Bcl-2 expression and induced apoptosis in the D594G melanoma line but not in a V600E-mutated line. Finally, we showed in a series of xenograft studies that sorafenib was more potent at reducing the growth of tumors with the D594G mutation than those with the V600E mutation. In summary, we have identified a group of melanomas with low-activity BRAF mutations that are reliant upon CRAF-mediated survival activity.
Smalley
K S M
KS
The Wistar Institute, Philadelphia, PA, USA. k.smalley@mac.com
Xiao
M
M
Villanueva
J
J
Nguyen
T K
TK
Flaherty
K T
KT
Letrero
R
R
Van Belle
P
P
Elder
D E
DE
Wang
Y
Y
Nathanson
K L
KL
Herlyn
M
M
D017209:Apoptosis / Q000235:genetics; D001557:Benzenesulfonates / Q000494:pharmacology; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D055785:Gene Knockdown Techniques; D018698:Glutamic Acid / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008545:Melanoma / Q000201:enzymology* / Q000235:genetics; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D009154:Mutation; D009536:Niacinamide / Q000031:analogs & derivatives; D010671:Phenylurea Compounds; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*; D019908:Proto-Oncogene Proteins c-raf / Q000037:antagonists & inhibitors* / Q000235:genetics; D011725:Pyridines / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000235:genetics; D015398:Signal Transduction; D000077157:Sorafenib; D014633:Valine / Q000235:genetics / Q000378:metabolism
D001557:Benzenesulfonates; D010671:Phenylurea Compounds; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D034741:RNA, Small Interfering; D009536:Niacinamide; D018698:Glutamic Acid; D000077157:Sorafenib; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D019908:Proto-Oncogene Proteins c-raf; D020929:Mitogen-Activated Protein Kinase Kinases; D014633:Valine
Cancer Res. 2006 Oct 1;66(19):9483-91
17018604
Mol Cell Biol. 2001 Apr;21(7):2324-36
11259582
Cell. 1996 Nov 15;87(4):629-38
8929532
J Mol Diagn. 2007 Sep;9(4):464-71
17690212
Proc Natl Acad Sci U S A. 2001 Jul 3;98(14):7783-8
11427728
Proc Natl Acad Sci U S A. 1996 Jul 9;93(14):7063-8
8692945
Cell. 2004 Mar 19;116(6):855-67
15035987
Science. 2004 Dec 24;306(5705):2267-70
15618521
Br J Cancer. 2007 Feb 12;96(3):445-9
17245336
Curr Opin Genet Dev. 2007 Feb;17(1):31-9
17208430
Nature. 2007 Feb 22;445(7130):851-7
17314971
Mol Cancer Ther. 2006 May;5(5):1136-44
16731745
Nature. 2006 Jan 19;439(7074):358-62
16273091
Cell. 1996 Nov 15;87(4):619-28
8929531
Int J Cancer. 2003 May 1;104(5):527-32
12594806
Biochim Biophys Acta. 2004 Mar 1;1644(2-3):149-58
14996499
Clin Cancer Res. 2008 Jan 1;14(1):230-9
18172275
J Biol Chem. 2005 Jul 1;280(26):24698-705
15849194
Br J Cancer. 2006 Sep 4;95(5):581-6
16880785
Am J Pathol. 2005 May;166(5):1541-54
15855653
Nature. 2002 Jun 27;417(6892):949-54
12068308
Nature. 1992 Jul 30;358(6385):417-21
1322500
Mol Cell. 2005 Dec 22;20(6):963-9
16364920
J Invest Dermatol. 2008 Aug;128(8):2003-12
18323787
Cancer Res. 2007 Jan 1;67(1):209-17
17210701
Cancer Res. 2006 Feb 1;66(3):1611-9
16452220
Cancer Res. 2004 Oct 1;64(19):7099-109
15466206