title: Threshold levels of ABL tyrosine kinase inhibitors retained in chronic myeloid leukemia cells determine their commitment to apoptosis.
abstract: The imatinib paradigm in chronic myelogenous leukemia (CML) established continuous BCR-ABL inhibition as a design principle for ABL tyrosine kinase inhibitors (TKI). However, clinical responses seen in patients treated with the ABL TKI dasatinib despite its much shorter plasma half-life and the apparent rapid restoration of BCR-ABL signaling activity following once-daily dosing suggested acute, potent inhibition of kinase activity may be sufficient to irrevocably commit CML cells to apoptosis. To determine the specific requirements for ABL TKI-induced CML cell death for a panel of clinically important ABL TKIs (imatinib, nilotinib, dasatinib, ponatinib, and DCC-2036), we interrogated response of CML cell lines and primary CML cells following acute drug exposure using intracellular fluorescence-activated cell sorting and immunoblot analyses of BCR-ABL signaling, apoptosis measurements, liquid chromatography/tandem mass spectrometry of intracellular drug levels, and biochemical TKI dissociation studies. Importantly, significant intracellular TKI stores were detected following drug washout, levels of which tracked with onset of apoptosis and incomplete return of BCR-ABL signaling, particularly pSTAT5, to baseline. Among TKIs tested, ponatinib showed the most robust capacity for apoptotic commitment showing sustained suppression of BCR-ABL signaling even at low intracellular levels following extensive washout, consistent with high-affinity binding and slow dissociation from ABL kinase. Together, our findings suggest commitment of CML cells to apoptosis requires protracted incomplete restoration of BCR-ABL signaling mediated by intracellular retention of TKIs above a quantifiable threshold. These studies refine our understanding of apoptotic commitment in CML cells and highlight parameters important to design of therapeutic kinase inhibitors for CML and other malignancies.
authors.affiliation: Division of Hematology and Hematologic Malignancies, University of Utah, Huntsman Cancer Institute, Salt Lake City, Utah, USA. Thomas.OHare@hci.utah.edu
authors.forename: Thomas
authors.initials: T
authors.lastname: O'Hare
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Eide
authors.forename: Anupriya
authors.initials: A
authors.lastname: Agarwal
authors.forename: Lauren T
authors.initials: LT
authors.lastname: Adrian
authors.forename: Matthew S
authors.initials: MS
authors.lastname: Zabriskie
authors.forename: Ryan J
authors.initials: RJ
authors.lastname: Mackenzie
authors.forename: Dorian H
authors.initials: DH
authors.lastname: Latocha
authors.forename: Kara J
authors.initials: KJ
authors.lastname: Johnson
authors.forename: Huihong
authors.initials: H
authors.lastname: You
authors.forename: Jenny
authors.initials: J
authors.lastname: Luo
authors.forename: Steven M
authors.initials: SM
authors.lastname: Riddle
authors.forename: Bryan D
authors.initials: BD
authors.lastname: Marks
authors.forename: Kurt W
authors.initials: KW
authors.lastname: Vogel
authors.forename: Dennis R
authors.initials: DR
authors.lastname: Koop
authors.forename: John
authors.initials: J
authors.lastname: Apgar
authors.forename: Jeffrey W
authors.initials: JW
authors.lastname: Tyner
authors.forename: Michael W
authors.initials: MW
authors.lastname: Deininger
authors.forename: Brian J
authors.initials: BJ
authors.lastname: Druker
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D001549:Benzamides / Q000493:pharmacokinetics / Q000494:pharmacology; D016044:Fusion Proteins, bcr-abl / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D000068877:Imatinib Mesylate; D020014:K562 Cells; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000188:drug therapy* / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D010879:Piperazines / Q000493:pharmacokinetics / Q000494:pharmacology; D047428:Protein Kinase Inhibitors / Q000493:pharmacokinetics* / Q000494:pharmacology; D011743:Pyrimidines / Q000493:pharmacokinetics / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D001549:Benzamides; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D000068877:Imatinib Mesylate; D016044:Fusion Proteins, bcr-abl