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Association between acquired resistance to PLX4032 (vemurafenib) and ATP-binding cassette transporter expression.

Michaelis M, Rothweiler F, Nerreter T, van Rikxoort M, Zehner R, Dirks WG, Wiese M, Cinatl J - BMC Res Notes (2014)

Bottom Line: Various kinase inhibitors are known to be ATP-binding cassette (ABC) transporter substrates and resistance acquisition to kinase inhibitors has been associated to increased ABC transporter expression.PLX4032 and PLX4720 affected ABCC1- and ABCG2-mediated drug transport in a similar fashion.Since ABC transporters confer multi-drug resistance, this is of relevance for the design of next-line therapies.

View Article: PubMed Central - PubMed

Affiliation: Institut für Medizinische Virologie, Klinikum der Goethe-Universität, Paul Ehrlich-Str, 40, Frankfurt am Main 60596, Germany. Cinatl@em.uni-frankfurt.de.

ABSTRACT

Background: Various kinase inhibitors are known to be ATP-binding cassette (ABC) transporter substrates and resistance acquisition to kinase inhibitors has been associated to increased ABC transporter expression. Here, we investigated the role of the ABC transporters ABCB1, ABCC1, and ABCG2 during melanoma cell resistance acquisition to the V600-mutant BRAF inhibitors PLX4032 (vemurafenib) and PLX4720. PLX4032 had previously been shown to interfere with ABCB1 and ABCG2. PLX4720 had been demonstrated to interact with ABCB1 but to a lower extent than PLX4032.

Findings: PLX4032 and PLX4720 affected ABCC1- and ABCG2-mediated drug transport in a similar fashion. In a panel of 16 V600E BRAF-mutated melanoma cell lines consisting of four parental cell lines and their sub-lines with acquired resistance to PLX4032, PLX4720, vincristine (cytotoxic ABCB1 and ABCC1 substrate), or mitoxantrone (cytotoxic ABCG2 substrate), we detected enhanced ABC transporter expression in 4/4 cytotoxic ABC transporter substrate-resistant, 3/4 PLX4720-resistant, and 1/4 PLX4032-resistant melanoma cell lines.

Conclusion: PLX4032 has the potential to induce ABC transporter expression but this potential is lower than that of PLX4720 or cytotoxic ABC transporter substrates. Since ABC transporters confer multi-drug resistance, this is of relevance for the design of next-line therapies.

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Effect of PLX4032 and PLX4720 on ABCG2 activity. A) Influence of PLX4032 or PLX4720 on BODIPY-prazosine (1 μM) fluorescence in UKF-NB-3ABCG2 cells, B) time kinetics of BODIPY-prazosine (1 μM) fluorescence in UKF-NB-3ABCG2 cells in the presence of PLX4032 or PLX4720 after a 60 min pre-incubation period with subsequent wash-out of extracellular BODIPY-prazosine and PLX4032 or PLX4720 (control = BODIPY-prazosine incubation in the absence of drugs). C) ABCG2 ATPase activity in isolated membranes in the presence of PLX4032 or PLX4720 (control = activity in the absence of drugs). Sulfasalazine, a known ABCG2 substrate, was used for comparison. *P < 0.05 relative to non-treated controls.
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Fig1: Effect of PLX4032 and PLX4720 on ABCG2 activity. A) Influence of PLX4032 or PLX4720 on BODIPY-prazosine (1 μM) fluorescence in UKF-NB-3ABCG2 cells, B) time kinetics of BODIPY-prazosine (1 μM) fluorescence in UKF-NB-3ABCG2 cells in the presence of PLX4032 or PLX4720 after a 60 min pre-incubation period with subsequent wash-out of extracellular BODIPY-prazosine and PLX4032 or PLX4720 (control = BODIPY-prazosine incubation in the absence of drugs). C) ABCG2 ATPase activity in isolated membranes in the presence of PLX4032 or PLX4720 (control = activity in the absence of drugs). Sulfasalazine, a known ABCG2 substrate, was used for comparison. *P < 0.05 relative to non-treated controls.

Mentions: To study the effects of PLX4032 and PLX4720 on ABCG2, an ABCG2-expressing sub-line of the BRAF wild-type neuroblastoma cell line UKF-NB-3 (UKF-NB-3ABCG2) was used that had been established by lentiviral transduction with LeGO vectors (http://www.lentigo-vectors.de) as described previously [21, 22]. All experimental procedures were performed as described previously [22]. PLX4032 and PLX4720 treatment of UKF-NB-3ABCG2 cells (but not of UKF-NB-3 cells or UKF-NB-3 cells transduced with a control vector) resulted in a similar dose-dependent increase in the cellular accumulation of the fluorescent ABCG2 substrate BODIPY-prazosine (Figure 1A) without affecting ABCG2 expression (Additional file 1: Figure S1).Figure 1


Association between acquired resistance to PLX4032 (vemurafenib) and ATP-binding cassette transporter expression.

Michaelis M, Rothweiler F, Nerreter T, van Rikxoort M, Zehner R, Dirks WG, Wiese M, Cinatl J - BMC Res Notes (2014)

Effect of PLX4032 and PLX4720 on ABCG2 activity. A) Influence of PLX4032 or PLX4720 on BODIPY-prazosine (1 μM) fluorescence in UKF-NB-3ABCG2 cells, B) time kinetics of BODIPY-prazosine (1 μM) fluorescence in UKF-NB-3ABCG2 cells in the presence of PLX4032 or PLX4720 after a 60 min pre-incubation period with subsequent wash-out of extracellular BODIPY-prazosine and PLX4032 or PLX4720 (control = BODIPY-prazosine incubation in the absence of drugs). C) ABCG2 ATPase activity in isolated membranes in the presence of PLX4032 or PLX4720 (control = activity in the absence of drugs). Sulfasalazine, a known ABCG2 substrate, was used for comparison. *P < 0.05 relative to non-treated controls.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4197243&req=5

Fig1: Effect of PLX4032 and PLX4720 on ABCG2 activity. A) Influence of PLX4032 or PLX4720 on BODIPY-prazosine (1 μM) fluorescence in UKF-NB-3ABCG2 cells, B) time kinetics of BODIPY-prazosine (1 μM) fluorescence in UKF-NB-3ABCG2 cells in the presence of PLX4032 or PLX4720 after a 60 min pre-incubation period with subsequent wash-out of extracellular BODIPY-prazosine and PLX4032 or PLX4720 (control = BODIPY-prazosine incubation in the absence of drugs). C) ABCG2 ATPase activity in isolated membranes in the presence of PLX4032 or PLX4720 (control = activity in the absence of drugs). Sulfasalazine, a known ABCG2 substrate, was used for comparison. *P < 0.05 relative to non-treated controls.
Mentions: To study the effects of PLX4032 and PLX4720 on ABCG2, an ABCG2-expressing sub-line of the BRAF wild-type neuroblastoma cell line UKF-NB-3 (UKF-NB-3ABCG2) was used that had been established by lentiviral transduction with LeGO vectors (http://www.lentigo-vectors.de) as described previously [21, 22]. All experimental procedures were performed as described previously [22]. PLX4032 and PLX4720 treatment of UKF-NB-3ABCG2 cells (but not of UKF-NB-3 cells or UKF-NB-3 cells transduced with a control vector) resulted in a similar dose-dependent increase in the cellular accumulation of the fluorescent ABCG2 substrate BODIPY-prazosine (Figure 1A) without affecting ABCG2 expression (Additional file 1: Figure S1).Figure 1

Bottom Line: Various kinase inhibitors are known to be ATP-binding cassette (ABC) transporter substrates and resistance acquisition to kinase inhibitors has been associated to increased ABC transporter expression.PLX4032 and PLX4720 affected ABCC1- and ABCG2-mediated drug transport in a similar fashion.Since ABC transporters confer multi-drug resistance, this is of relevance for the design of next-line therapies.

View Article: PubMed Central - PubMed

Affiliation: Institut für Medizinische Virologie, Klinikum der Goethe-Universität, Paul Ehrlich-Str, 40, Frankfurt am Main 60596, Germany. Cinatl@em.uni-frankfurt.de.

ABSTRACT

Background: Various kinase inhibitors are known to be ATP-binding cassette (ABC) transporter substrates and resistance acquisition to kinase inhibitors has been associated to increased ABC transporter expression. Here, we investigated the role of the ABC transporters ABCB1, ABCC1, and ABCG2 during melanoma cell resistance acquisition to the V600-mutant BRAF inhibitors PLX4032 (vemurafenib) and PLX4720. PLX4032 had previously been shown to interfere with ABCB1 and ABCG2. PLX4720 had been demonstrated to interact with ABCB1 but to a lower extent than PLX4032.

Findings: PLX4032 and PLX4720 affected ABCC1- and ABCG2-mediated drug transport in a similar fashion. In a panel of 16 V600E BRAF-mutated melanoma cell lines consisting of four parental cell lines and their sub-lines with acquired resistance to PLX4032, PLX4720, vincristine (cytotoxic ABCB1 and ABCC1 substrate), or mitoxantrone (cytotoxic ABCG2 substrate), we detected enhanced ABC transporter expression in 4/4 cytotoxic ABC transporter substrate-resistant, 3/4 PLX4720-resistant, and 1/4 PLX4032-resistant melanoma cell lines.

Conclusion: PLX4032 has the potential to induce ABC transporter expression but this potential is lower than that of PLX4720 or cytotoxic ABC transporter substrates. Since ABC transporters confer multi-drug resistance, this is of relevance for the design of next-line therapies.

Show MeSH
Related in: MedlinePlus