Multidrug resistance-modifying components in human plasma with potential clinical significance.
P-Glycoprotein (P-gp) and multidrug resistance protein (MRP) are plasma membrane associated proteins which can confer multidrug resistance (MDR) to cancer cells by lowering the intracellular amount of drug. Although clinical trials with MDR-reverting agents have been initiated, not much attention has been paid to blood components which may modulate the activity of P-gp or MRP. The present investigation was performed to identify and characterize blood components which may influence the drug content and the drug cytotoxicity of MDR cells. Human plasma, from healthy volunteers, was tested for its effects on the daunorubicin (DNR) accumulation and cytotoxicity in the MDR cell lines SW-1573/2R160 (2R160) and GLC4/ADR containing P-gp and MRP, respectively. The data were compared to the effects observed in wild-type cells. MDR-modifying plasma components were isolated by extraction procedures and characterized using ultrafiltration, high-performance liquid chromatography (HPLC) and mass spectrometry. An increase in the proportion of plasma in the culture medium led to a reduction of the ratio between the DNR content of wild-type and corresponding MDR cells. At 100% plasma we observed an increase in the cellular DNR content of 2R160 cells, which was 10-30% (median 18%) of the maximum possible increase induced by well-known MDR-reverting agents, such as verapamil (for GLC4/ADR cells: 10-20%, median 15%). The DNR cytotoxicity in MDR cells also increased with an increasing amount of plasma included in the culture media. There was neither an increase in the cellular DNR content nor an effect on the DNR cytotoxicity in wild-type cells. Plasma extract analysis by HPLC showed a major peak which increased the DNR content of MDR cells. The HPLC column retention time of this fraction was identical to that of a standard of cortisol and it was further confirmed to be cortisol using mass spectrometry. Moreover, inclusion of a standard of cortisol in culture media induced a similar effect. We analyzed the data for one of the plasma pools and found that blood cortisol was responsible for the MDR-modulating effect only for 35% of the effect of 100% plasma. Other plasma components were responsible for the remaining modulation effect on MDR cells. In conclusion, the DNR pumping activity of P-gp and MRP is inhibited by human plasma, resulting in 10-30% of the maximum possible increase in cellular drug content. Based on cellular pharmacokinetic calculations this percentage will most likely increase at clinical levels of drug resistance (reaching 40-50%). In one sample blood cortisol accounted for 35% of the effect of plasma on the DNR content in MDR 2R160 cells. These data show the need for additional studies to test plasma samples for their MDR modulating effects before the administration of MDR-reverting agents in chemotherapy. The data suggest that the effectiveness of chemotherapeutic drugs may be enhanced when administered in accordance with the circadian peak of endogenous corticoids.
Mülder
H S
HS
Department of Medical Oncology, University Hospital Vriije Universiteit, Amsterdam, The Netherlands.
Pinedo
H M
HM
Timmer
A T
AT
Rao
B R
BR
Lankelma
J
J
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000097:blood*; D018528:ATP-Binding Cassette Transporters / Q000097:blood*; D000328:Adult; D000903:Antibiotics, Antineoplastic / Q000378:metabolism / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D003630:Daunorubicin / Q000378:metabolism / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005260:Female; D006801:Humans; D006854:Hydrocortisone / Q000097:blood; D006863:Hydrogen-Ion Concentration; D008297:Male; D013058:Mass Spectrometry; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009363:Neoplasm Proteins / Q000097:blood*; D014407:Tumor Cells, Cultured
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D018528:ATP-Binding Cassette Transporters; D000903:Antibiotics, Antineoplastic; D003630:Daunorubicin; D006854:Hydrocortisone; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009363:Neoplasm Proteins