title: RNAi screening of the kinome with cytarabine in leukemias.
abstract: To identify rational therapeutic combinations with cytarabine (Ara-C), we developed a high-throughput, small-interference RNA (siRNA) platform for myeloid leukemia cells. Of 572 kinases individually silenced in combination with Ara-C, silencing of 10 (1.7%) and 8 (1.4%) kinases strongly increased Ara-C activity in TF-1 and THP-1 cells, respectively. The strongest molecular concepts emerged around kinases involved in cell-cycle checkpoints and DNA-damage repair. In confirmatory siRNA assays, inhibition of WEE1 resulted in more potent and universal sensitization across myeloid cell lines than siRNA inhibition of PKMYT1, CHEK1, or ATR. Treatment of 8 acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), and chronic myeloid leukemia (CML) cell lines with commercial and the first-in-class clinical WEE1 kinase inhibitor MK1775 confirmed sensitization to Ara-C up to 97-fold. Ex vivo, adding MK1775 substantially reduced viability in 13 of 14 AML, CML, and myelodysplastic syndrome patient samples compared with Ara-C alone. Maximum sensitization occurred at lower to moderate concentrations of both drugs. Induction of apoptosis was increased using a combination of Ara-C and MK1775 compared with using either drug alone. WEE1 is expressed in primary AML, ALL, and CML specimens. Data from this first siRNA-kinome sensitizer screen suggests that inhibiting WEE1 in combination with Ara-C is a rational combination for the treatment of myeloid and lymphoid leukemias.
authors.affiliation: Division of Hematology and Oncology, Mayo Clinic, Scottsdale, AZ 85259, USA. tibes.raoul@mayo.edu
authors.forename: Raoul
authors.initials: R
authors.lastname: Tibes
authors.forename: James M
authors.initials: JM
authors.lastname: Bogenberger
authors.forename: Leena
authors.initials: L
authors.lastname: Chaudhuri
authors.forename: R Tanner
authors.initials: RT
authors.lastname: Hagelstrom
authors.forename: Donald
authors.initials: D
authors.lastname: Chow
authors.forename: Megan E
authors.initials: ME
authors.lastname: Buechel
authors.forename: Irma M
authors.initials: IM
authors.lastname: Gonzales
authors.forename: Tim
authors.initials: T
authors.lastname: Demuth
authors.forename: James
authors.initials: J
authors.lastname: Slack
authors.forename: Ruben A
authors.initials: RA
authors.lastname: Mesa
authors.forename: Esteban
authors.initials: E
authors.lastname: Braggio
authors.forename: Hongwei H
authors.initials: HH
authors.lastname: Yin
authors.forename: Shilpi
authors.initials: S
authors.lastname: Arora
authors.forename: David O
authors.initials: DO
authors.lastname: Azorsa
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D003561:Cytarabine / Q000494:pharmacology*; D004354:Drug Screening Assays, Antitumor; D057166:High-Throughput Screening Assays; D006801:Humans; D007951:Leukemia, Myeloid / Q000201:enzymology*; D009687:Nuclear Proteins / Q000378:metabolism*; D010770:Phosphotransferases / Q000032:analysis* / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D034741:RNA, Small Interfering; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D000964:Antimetabolites, Antineoplastic; D018797:Cell Cycle Proteins; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D003561:Cytarabine; D010770:Phosphotransferases; D011505:Protein-Tyrosine Kinases; C499349:WEE1 protein, human