Factors influencing the cellular accumulation of SN-38 and camptothecin.
OBJECTIVE
The influence of biophysical factors (drug metabolism, transport proteins, and chemical stability) on the cellular accumulation of camptothecin (CPT) and SN-38 was examined.


METHODS
Drug transporter RNA transcript levels were measured by real-time reverse transcriptase polymerase chain reaction (RT-PCR). Intracellular drug accumulation, metabolism, and drug stability studies were all performed by HPLC.


RESULTS
A panel of three human cell lines exhibiting different drug resistant phenotypes was investigated. HT29 colon cells glucuronidated SN-38 but did not express P-gp or MRP1 or 2. HCT116 colon cells expressed P-gp and MRP2 but did not catalyse conjugation. A2780 ovarian cells neither catalysed drug metabolism nor contained these drug transporters. In all lines, SN-38 lactone was rapidly taken up achieving peak concentrations at the earliest time point studied (5 min, 3.3-4.1 ng/10(6) cells). Subsequently, a fall in intracellular lactone concentration occurred, stabilising after 4 h at 0.48-1.18 ng/10(6) cells. No significant differences in intracellular levels of lactone were observed between the three cell lines with one exception: a twofold increase in HCT116 cells at 24 h. Stability studies in culture medium revealed that SN-38 lactone concentrations disappeared at the same rate regardless of whether cells were present, initially falling to reach equilibrium with the hydroxy acid by 4 h. Indeed, changes in intracellular lactone concentrations followed closely chemical stability profiles in media. Similar patterns of cellular retention and chemical degradation were observed with CPT.


CONCLUSIONS
The major determinant of drug accumulation in three diverse cell line phenotypes was lactone chemical stability in culture medium.
Cummings
Jeffrey
J
Imperial Cancer Research Fund, Medical Oncology Unit, Western General Hospital, Edinburgh, EH4 2XU, UK. j.cummings@icrf.icnet.uk
Boyd
Gary
G
Macpherson
Janet S
JS
Wolf
Helga
H
Smith
Gillian
G
Smyth
John F
JF
Jodrell
Duncan I
DI
D000230:Adenocarcinoma; D001692:Biological Transport; D002166:Camptothecin / Q000031:analogs & derivatives / Q000493:pharmacokinetics*; D003110:Colonic Neoplasms; D018432:Drug Resistance, Multiple; D004791:Enzyme Inhibitors / Q000493:pharmacokinetics; D005260:Female; D019450:Genes, MDR; D006801:Humans; D000077146:Irinotecan; D007700:Kinetics; D026901:Membrane Transport Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics; D010051:Ovarian Neoplasms; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D059004:Topoisomerase I Inhibitors; D014407:Tumor Cells, Cultured
D002166:Camptothecin; D004791:Enzyme Inhibitors; D000077146:Irinotecan; D026901:Membrane Transport Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D059004:Topoisomerase I Inhibitors; C000715087:ABCC2 protein, human; C421109:multidrug resistance-associated protein 1