RNAi-mediated silencing of p190Bcr-Abl inactivates Stat5 and cooperates with imatinib mesylate and 17-allylamino-17-demetoxygeldanamycin in selective killing of p190Bcr-Abl-expressing leukemia cells.
The 190 kD (p190) and 210 kD (p210) Bcr-Abl proteins are responsible for the pathophysiology of Philadelphia chromosome (Ph)(+) leukemia. We applied RNA interference (RNAi) to specific killing of p190(+) cells, and determined the optimal sequences for gene silencing in the BCR, junctional and ABL regions of p190, respectively. Then, p190(+) and p210(+) cells were infected with lentiviral vectors encoding these shRNAs, resulting in efficient killing of p190(+) cells, while p210(+) cells were only sensitive to shBCR and shABL. In p190-transformed Ba/F3 cells, silencing of p190 specifically inhibited tyrosine phospohorylation of Stat5 prior to their death, but did not affect phosphorylation of Jak2, Akt or MEK1/2. In contrast, downregulation of p190 by their treatment with 17-allylamino-17-demetoxygeldanamycin (17-AAG) was associated with reduced protein levels of Jak2, Akt and MEK1/2. shRNA targeting p190 collaborated additively with imatinib and 17-AAG in growth inhibition of Ba/F3-p190wt and imatinib-resistant Ba/F3-p190Y253 H cells. Collectively, RNAi-mediated silencing of p190 is a promising option both for delineating signal transduction and for therapeutic application in 190(+) leukemia.
Futami
M
M
Division of Molecular Therapy, Advanced Clinical Research Center, Institute of Medical Science, University of Tokyo, Tokyo, Japan.
Hatano
T
T
Soda
Y
Y
Kobayashi
S
S
Miyagishi
M
M
Tojo
A
A
D000818:Animals; D001549:Benzamides; D016227:Benzoquinones / Q000494:pharmacology; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D003114:Colony-Forming Units Assay; D016044:Fusion Proteins, bcr-abl / Q000235:genetics* / Q000378:metabolism; D006410:Hematopoiesis / Q000187:drug effects / Q000502:physiology; D000068877:Imatinib Mesylate; D047029:Lactams, Macrocyclic / Q000494:pharmacology; D016086:Lentivirus / Q000235:genetics; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000473:pathology*; D051379:Mice; D010879:Piperazines / Q000494:pharmacology; D054448:Precursor Cells, B-Lymphoid / Q000378:metabolism / Q000473:pathology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D011743:Pyrimidines / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000494:pharmacology; D050799:STAT5 Transcription Factor / Q000378:metabolism*; D015398:Signal Transduction
D001549:Benzamides; D016227:Benzoquinones; D047029:Lactams, Macrocyclic; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D034741:RNA, Small Interfering; D050799:STAT5 Transcription Factor; C112765:tanespimycin; D000068877:Imatinib Mesylate; D011505:Protein-Tyrosine Kinases; D016044:Fusion Proteins, bcr-abl