Overexpression of the multidrug resistance genes mdr1, mdr3, and mrp in L1210 leukemia cells resistant to inhibitors of ribonucleotide reductase.
L1210 MQ-580 is a murine leukemia cell line resistant to the cytotoxic activity of the alpha-(N)-heterocyclic carboxaldehyde thiosemicarbazone class of inhibitors of ribonucleotide reductase. The line is cross-resistant to etoposide, daunomycin, and vinblastine. L1210 MQ-580 cells expressed 8-fold resistance to 3-aminopyridine-2-carboxaldehyde thiosemicarbazone (3-AP), a relatively newly developed inhibitor of ribonucleotide reductase. The accumulation of [14C]3-AP by L1210 MQ-580 cells was 5- to 6-fold less than by parental L1210 cells. An increased rate of efflux of 3-AP was responsible for the lower steady-state concentration of 3-AP in resistant cells. In reverse transcription-polymerase chain reaction assays, L1210 MQ-580 cells were found to overexpress the multidrug resistance genes mdr1, mdr3, and mrp, but not the mdr2 gene, compared with parental L1210 cells. Measurement of the steady-state concentration of doxorubicin, a potential substrate for both the mdr and mrp gene products, demonstrated that L1210 MQ-580 cells accumulated 4-fold less anthracycline than parental cells. These findings indicate that drug efflux is a major determinant of the pattern of cross-resistance of L1210 MQ-580 cells. To extrapolate these observations to the human homologues of the mdr1, mdr3, and mrp murine genes, the effects of 3-AP were measured in L1210/VMDRC0.06 and NIH3T3 36-8-32 cells transfected with human MDR1 and MRP cDNAs, respectively. The transfectants were 2- to 3-fold resistant to the cytotoxic effects of 3-AP and accumulated less [14C]3-AP than their parental mock-transfected counterparts. Moreover, the cytotoxic activity of 3-AP was significantly greater in two double mrp gene knockout cell lines than in parental W 9.5 embryonic stem cells. Thus, the results suggest that 3-AP is a substrate for both the P-glycoprotein and MRP and that baseline MRP expression has the capacity to exert a protective role against the toxicity of this agent.
Rappa
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Department of Pharmacology and Developmental Therapeutics Program, Cancer Center, Yale University School of Medicine, New Haven, CT, 06520, U.S.A.
Lorico
A
A
Liu
M C
MC
Kruh
G D
GD
Cory
A H
AH
Cory
J G
JG
Sartorelli
A C
AC
D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics*; D018528:ATP-Binding Cassette Transporters / Q000235:genetics*; D000818:Animals; D018432:Drug Resistance, Multiple / Q000235:genetics; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D007939:Leukemia L1210 / Q000235:genetics* / Q000473:pathology; D051379:Mice; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D012264:Ribonucleotide Reductases / Q000037:antagonists & inhibitors*; D013882:Thiosemicarbazones / Q000494:pharmacology
D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D018528:ATP-Binding Cassette Transporters; D004791:Enzyme Inhibitors; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D012264:Ribonucleotide Reductases; D013882:Thiosemicarbazones; C095122:multidrug resistance protein 3