Research Papers:

Enzastaurin inhibits ABCB1-mediated drug efflux independently of effects on protein kinase C signalling and the cellular p53 status

Martin Michaelis _, Florian Rothweiler, Nadine Löschmann, Mohsen Sharifi, Taravat Ghafourian and Jindrich Cinatl Jr.

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Oncotarget. 2015; 6:17605-17620. https://doi.org/10.18632/oncotarget.2889

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Martin Michaelis1,2, Florian Rothweiler1, Nadine Löschmann1, Mohsen Sharifi3, Taravat Ghafourian3, Jindrich Cinatl Jr.1

1Institut für Medizinische Virologie, Klinikum der Goethe-Universität, Frankfurt am Main, Germany

2Centre for Molecular Processing and School of Biosciences, University of Kent, Canterbury, UK

3Medway School of Pharmacy, Universities of Kent and Greenwich, Chatham Maritime, Kent, UK

Correspondence to:

Jindrich Cinatl Jr., e-mail: [email protected]

Keywords: ONC201, cancer drug, TIC10, NSC350625

Abbreviations: ABC transporter, ATP-binding cassette transporter; MARCKS, myristoylated alanine-rich C-kinase substrate; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; RCCL collection, Resistant Cancer Cell Line collection.

Received: September 09, 2014     Accepted: December 09, 2014     Published: February 13, 2015


The PKCβ inhibitor enzastaurin was tested in parental neuroblastoma and rhabdomyosarcoma cell lines, their vincristine-resistant sub-lines, primary neuroblastoma cells, ABCB1-transduced, ABCG2-transduced, and p53-depleted cells. Enzastaurin IC50s ranged from 3.3 to 9.5 μM in cell lines and primary cells independently of the ABCB1, ABCG2, or p53 status. Enzastaurin 0.3125 μM interfered with ABCB1-mediated drug transport. PKCα and PKCβ may phosphorylate and activate ABCB1 under the control of p53. However, enzastaurin exerted similar effects on ABCB1 in the presence or absence of functional p53. Also, enzastaurin inhibited PKC signalling only in concentrations ≥ 1.25 μM. The investigated cell lines did not express PKCβ. PKCα depletion reduced PKC signalling but did not affect ABCB1 activity. Intracellular levels of the fluorescent ABCB1 substrate rhodamine 123 rapidly decreased after wash-out of extracellular enzastaurin, and enzastaurin induced ABCB1 ATPase activity resembling the ABCB1 substrate verapamil. Computational docking experiments detected a direct interaction of enzastaurin and ABCB1. These data suggest that enzastaurin directly interferes with ABCB1 function. Enzastaurin further inhibited ABCG2-mediated drug transport but by a different mechanism since it reduced ABCG2 ATPase activity. These findings are important for the further development of therapies combining enzastaurin with ABC transporter substrates.

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