Development of a conditional in vivo model to evaluate the efficacy of small molecule inhibitors for the treatment of Raf-transformed hematopoietic cells.
Conditionally active forms of the Raf proteins (Raf-1, B-Raf, and A-Raf) were created by ligating NH2-terminal truncated activated forms (Delta) to the estrogen receptor (ER) hormone-binding domain resulting in estradiol-regulated constructs (DeltaRaf:ER). These different Raf:ER oncoproteins were introduced into the murine FDC-P1 hematopoietic cell line, and cells that grew in response to the three DeltaRaf:ER oncoproteins were isolated. The ability of FDC-P1, DeltaRaf-1:ER, DeltaA-Raf:ER, and DeltaB-Raf:ER cells to form tumors in severe combined immunodeficient mice was compared. Mice injected with DeltaRaf:ER cells were implanted with beta-estradiol pellets to induce the DeltaRaf:ER oncoprotein. Cytokine-dependent parental cell lines did not form tumors. Implantation of beta-estradiol pellets into mice injected with DeltaRaf:ER cells significantly accelerated tumor onset and tumor size. The recovered DeltaRaf:ER cells displayed induction of extracellular signal-regulated kinase (ERK) in response to beta-estradiol stimulation, indicating that they had retained conditional activation of ERK even when passed through a severe combined immunodeficient mouse. The DeltaRaf:ER cells were very sensitive to induction of apoptosis by the mitogen-activated protein/ERK kinase (MEK) 1 inhibitor CI1040 whereas parental cells were much less affected, demonstrating that the MEK1 may be useful in eliminating Ras/Raf/MEK-transformed cells. Furthermore, the effects of in vivo administration of the MEK1 inhibitor were evaluated and this inhibitor was observed to suppress the tumorigenicity of the injected cells. This DeltaRaf:ER system can serve as a preclinical model to evaluate the effects of signal transduction inhibitors which target the Raf and MEK proteins.
Konopleva
Marina
M
Department of Blood and Marrow Transplantation, The University of Texas M.D. Anderson Cancer Center, Houston, Texas 77030, USA.
Shi
Yuexi
Y
Steelman
Linda S
LS
Shelton
John G
JG
Munsell
Mark
M
Marini
Frank
F
McQueen
Teresa
T
Contractor
Rooha
R
McCubrey
James A
JA
Andreeff
Michael
M
D000818:Animals; D001549:Benzamides / Q000494:pharmacology; D002471:Cell Transformation, Neoplastic / Q000187:drug effects / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D004354:Drug Screening Assays, Antitumor / Q000379:methods; D004958:Estradiol / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D007938:Leukemia / Q000139:chemically induced / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D051379:Mice; D016513:Mice, SCID; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism; D011960:Receptors, Estrogen / Q000096:biosynthesis; D014183:Transplantation, Heterologous; D048490:raf Kinases / Q000096:biosynthesis*
C120227:2-(2-chloro-4-iodophenylamino)-N-cyclopropylmethoxy-3,4-difluorobenzamide; D001549:Benzamides; D011960:Receptors, Estrogen; D004958:Estradiol; D048490:raf Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3