title: P-glycoprotein and breast cancer resistance protein in acute myeloid leukaemia cells treated with the aurora-B kinase inhibitor barasertib-hQPA.
abstract: BACKGROUND
  Aurora kinases play an essential role in orchestrating chromosome alignment, segregation and cytokinesis during mitotic progression, with both aurora-A and B frequently over-expressed in a variety of human malignancies. Over-expression of the ABC drug transporter proteins P-glycoprotein (Pgp) and Breast cancer resistance protein (BCRP) is a major obstacle for chemotherapy in many tumour types with Pgp conferring particularly poor prognosis in acute myeloid leukaemia (AML). Barasertib-hQPA is a highly selective inhibitor of aurora-B kinase that has shown tumouricidal activity against a range tumour cell lines including those of leukaemic AML origin.
  
  
  METHODS
  Effect of barasertib-hQPA on the pHH3 biomarker and cell viability was measured in a panel of leukaemic cell lines and 37 primary AML samples by flow cytometry. Pgp status was determined by flow cytometry and BCRP status by flow cytometry and real-time PCR.
  
  
  RESULTS
  In this study we report the creation of the cell line OCI-AML3DNR, which over-expresses Pgp but not BCRP or multidrug resistance-associated protein (MRP), through prolonged treatment of OCI-AML3 cells with daunorubicin. We demonstrate that Pgp (OCI-AML3DNR and KG-1a) and BCRP (OCI-AML6.2) expressing AML cell lines are less sensitive to barasertib-hQPA induced pHH3 inhibition and subsequent loss of viability compared to transporter negative cell lines. We also show that barasertib-hQPA resistance in these cell lines can be reversed using known Pgp and BCRP inhibitors. We report that barasertib-hQPA is not an inhibitor of Pgp or BCRP, but by using 14[C]-barasertib-hQPA that it is effluxed by these transporters. Using phosphoHistone H3 (pHH3) as a biomarker of barasertib-hQPA responsiveness in primary AML blasts we determined that Pgp and BCRP positive primary samples were less sensitive to barasertib-hQPA induced pHH3 inhibition (p = <0.001) than samples without these transporters. However, we demonstrate that IC50 inhibition of pHH3 by barasertib-hQPA was achieved in 94.6% of these samples after 1 hour drug treatment, in contrast to the resistance of the cell lines.
  
  
  CONCLUSIONS
  We conclude that Pgp and BCRP status and pHH3 down-regulation in patients treated with barasertib should be monitored in order to establish whether transporter-mediated efflux is sufficient to adversely impact on the efficacy of the agent.
authors.affiliation: Department of Academic Haematology, The University of Nottingham, Clinical Sciences Building, Hucknall Road, Nottingham, NG5 1PB, UK. martin.grundy@nottingham.ac.uk
authors.forename: Martin
authors.initials: M
authors.lastname: Grundy
authors.forename: Claire
authors.initials: C
authors.lastname: Seedhouse
authors.forename: Nigel H
authors.initials: NH
authors.lastname: Russell
authors.forename: Monica
authors.initials: M
authors.lastname: Pallis
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis*; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000032:analysis* / Q000378:metabolism; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology / Q000627:therapeutic use*; D064107:Aurora Kinase B; D064127:Aurora Kinases; D045744:Cell Line, Tumor / Q000187:drug effects / Q000201:enzymology; D003630:Daunorubicin / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm; D005260:Female; D006657:Histones / Q000378:metabolism; D006801:Humans; D020128:Inhibitory Concentration 50; D015470:Leukemia, Myeloid, Acute / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D008297:Male; D009363:Neoplasm Proteins / Q000032:analysis / Q000037:antagonists & inhibitors* / Q000502:physiology; D010755:Organophosphates / Q000493:pharmacokinetics / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology / Q000627:therapeutic use*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000502:physiology; D011799:Quinazolines / Q000493:pharmacokinetics / Q000494:pharmacology*
chemical_list: C520647:2-((3-((4-((5-(2-((3-fluorophenyl)amino)-2-oxoethyl)-1H-pyrazol-3-yl)amino)quinazolin-7-yl)oxy)propyl)(ethyl)amino)ethyl dihydrogen phosphate; C116614:ABCG2 protein, human; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D000903:Antibiotics, Antineoplastic; D000970:Antineoplastic Agents; D006657:Histones; D009363:Neoplasm Proteins; D010755:Organophosphates; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; C531556:AZD 1152-HQPA; C576921:AURKB protein, human; D064107:Aurora Kinase B; D064127:Aurora Kinases; D017346:Protein Serine-Threonine Kinases; D003630:Daunorubicin