Novel mechanism of resistance to paclitaxel (Taxol) in human K562 leukemia cells by combined selection with PSC 833.
A paclitaxel-resistant cell line, KPTA5, was established by co-selecting the parental erythroleukemic cell line K562 with stepwise increased concentrations of paclitaxel (Taxol) in the presence of the cyclosporin D analogue PSC 833 (2 microM), a potent modulator of the multidrug resistance phenotype. KPTA5 cells are 9-fold resistant to paclitaxel and taxotere, but do not exhibit significant resistance to Vinca alkaloids, etoposide, anthracyclines, antimetabolites, or alkylating agents. Doubling time and morphology were similar to the parental K562 cells. Reverse transcriptase-polymerase chain reaction (rt-PCR) analysis revealed no alterations in the expression of the mdr1 and MRP genes. Cellular paclitaxel accumulation was unchanged. Cell cycle analyses showed that at 20 nM there was a significantly higher proportion of K562 cells blocked in G2/M, in comparison with KPTA5 cells. In both cases, disruption of the mitotic spindles and the presence of multiple mitotic asters were comparable but occurred at lower paclitaxel concentrations in K562 cells than in KPTA5 cells. There was no difference in total tubulin content between K562 and KPTA5 cells as analyzed by immunoblotting with an anti-beta-tubulin monoclonal antibody. However, we found that KPTA5 cells presented a 2-fold increase both in 5 beta-tubulin mRNA expression and in the corresponding tubulin protein Class IV isotype content, as evaluated by rt-PCR and immunostaining. In conclusion, the KPTA5 cell line displays a novel mechanism of resistance to paclitaxel which does not involve altered cellular drug accumulation. The data presented suggest that alterations in expression of the 5 beta-tubulin gene may be involved in paclitaxel resistance.
Jaffrézou
J P
JP
Department of Medicine, Stanford University Medical Center, CA 94305, USA.
Dumontet
C
C
Derry
W B
WB
Durán
G
G
Chen
K G
KG
Tsuchiya
E
E
Wilson
L
L
Jordan
M A
MA
Sikic
B I
BI
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000096:biosynthesis; D018528:ATP-Binding Cassette Transporters / Q000032:analysis; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000493:pharmacokinetics / Q000494:pharmacology*; D001483:Base Sequence; D002453:Cell Cycle / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects; D003524:Cyclosporins / Q000008:administration & dosage; D018432:Drug Resistance, Multiple / Q000502:physiology*; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D008938:Mitosis / Q000187:drug effects; D008969:Molecular Sequence Data; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D017239:Paclitaxel / Q000008:administration & dosage / Q000493:pharmacokinetics; D010641:Phenotype; D016133:Polymerase Chain Reaction; D008941:Spindle Apparatus / Q000187:drug effects; D014316:Tritium; D014404:Tubulin / Q000032:analysis / Q000378:metabolism; D014407:Tumor Cells, Cultured
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D018528:ATP-Binding Cassette Transporters; D003524:Cyclosporins; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D017239:Paclitaxel; D014316:Tritium; D014404:Tubulin; C070272:valspodar