Overexpression of P-glycoprotein and alterations in topoisomerase II in P388 mouse leukemia cells selected in vivo for resistance to mitoxantrone.
The overexpression of P-glycoprotein (PGP) and alterations in DNA topoisomerase II (TOPO II) were evaluated in mouse leukemia P388 cells selected in vivo for mitoxantrone (MTT) resistance (P388/MTT) and compared to doxorubicin (DOX) resistant (P388/DOX) or vincristine (VCR) resistant (P388/VCR) models. Among a panel of TOPO II inhibitors which included etoposide (VP-16), DOX, MTT and 4'-[(9-acridinyl)-amino]methanesulfon-m-anisidide (m-AMSA), the relative resistance compared to parental sensitive P388/S cells was: P388/DOX greater than P388/MTT greater than P388/VCR. All the resistant sublines exhibited minimal cell kill (less than 20%) at vincristine concentrations greater than 100-fold the IC50 for P388/S cells. In a soft-agar colony-forming assay, the modulation of cytotoxicity in P388/MTT cells by the calmodulin inhibitor trifluoperazine following a 3-hr drug treatment demonstrated a marked potentiation in cell kill with MTT, VP-16, DOX and m-AMSA but not VCR. Immunoblotting data revealed that while PGP was not detectable in P388/S cells, the overexpression of PGP was apparent in P388/MTT cells and the relative expression between the resistant sublines was: P388/DOX greater than P388/MTT greater than P388/VCR. Although the amount and DNA cleavage activity of TOPO II in nuclear extracts from P388/VCR cells were comparable to those in P388/S cells, they were markedly lower in both P388/DOX and P388/MTT cells. However, decatenation activity of TOPO II in nuclear extracts was comparable between the sensitive (P388/S) and resistant sublines (P388/MTT, P388/DOX, and P388/VCR). Results from the present study demonstrated that P388 cells selected for resistance to mitoxantrone exhibit changes in TOPO II and overexpression of PGP similar to P388/DOX cells, while vincristine resistant cells only overexpress PGP. Since therapeutic strategies are primarily designed to interfere with PGP-mediated drug efflux, the choice of agents for modulating resistance in tumors which overexpress PGP versus tumors which overexpress PGP with altered TOPO II could be different.
Kamath
N
N
Research Institute, Cleveland Clinic Foundation, OH 44195.
Grabowski
D
D
Ford
J
J
Kerrigan
D
D
Pommier
Y
Y
Ganapathi
R
R
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000818:Animals; D016923:Cell Death; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004317:Doxorubicin / Q000627:therapeutic use; D004351:Drug Resistance; D007941:Leukemia P388 / Q000188:drug therapy* / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D051379:Mice; D008942:Mitoxantrone / Q000627:therapeutic use*; D014407:Tumor Cells, Cultured; D014750:Vincristine / Q000627:therapeutic use
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D008562:Membrane Glycoproteins; D014750:Vincristine; D004317:Doxorubicin; D008942:Mitoxantrone; D004250:DNA Topoisomerases, Type II