Abstract
The cytotoxic activity and cross-resistance pattern of the novel topoisomerase I inhibitor topotecan (Topo) were investigated in ten cell lines, representing different mechanisms of cytotoxic drug resistance, and in 218 fresh human tumour samples using the fluorometric microculture cytotoxicity assay (FMCA). Resistance to Topo in the cell lines was associated with expression of the multidrug resistance-associated protein (MRP), whereas the cell lines with P-glycoprotein (P-gp), topoisomerase II and glutathione-associated resistance did not show decreased sensitivity to the drug. Topo was more active in haematological than in solid tumour samples, but substantial activity was observed in carcinomas of the ovary and breast, sarcoma and childhood solid tumours. Cross-resistance to standard drugs representing different mechanisms of action was generally low in patient cells. The effect of Topo was better after longer exposure, but this time-dependent effect was largely abolished when adjustment for in vitro exposure was made. Topo showed activity both in proliferative and non-proliferative cell systems. The results indicate that Topo is insensitive to major mechanisms of resistance except for MRP. Proliferation does not seem to be necessary for the effect of Topo, and no superiority for protracted dosing schedules was observed. The results also suggest that, for example, leukaemias, lymphomas, sarcomas and childhood solid tumours may be suitable targets for future phase II trials.
Highlights
The H69AR cell line expresses a multidrug-resistant (MDR) phenotype proposed to be mediated by a multidrug resistance-associated protein (MRP) and the CEMNVM-l expresses an atypical MDR, which is associated with altered topoisomerase II activity
The cell lines of haematological origin were more and sensitive, except for the myeloma cell line RPMI 8226/S with its sublines 8226/Dox40 and 8226/LR-5, which were of intermediate sensitivity
The greatest difference in sensitivity between a cell line and its subline was seen for National Cancer Institute (NCI)-H69 and its multidrug resistanceassociated protein (MRP)-expressing resistant subline H69AR, where the resistance factor was 77 (Table 1)
Summary
To evaluate the resistance pattern of Topo, a human cell line panel of four sensitive parental cell lines, five drug resistant sublines, representing different mechanisms of resistance, and one cell line with primary resistance was used. A complete medium consisting of culture medium RPMI- 1640 (HyClone) supplemented with 10% inactivated FCS, 2 mm glutamine, 50 ,ug ml-' streptomycin and 60 jg ml-' penicillin was used throughout for both cell lines and patient samples Both in the cell lines and in the patient samples the activity of Topo and the standard drugs cisplatin, cytarabine, doxorubicin, etoposide, melphalan and taxol was determined. To determine the dose-response relationship for Topo in patient samples, five different drug concentrations were used, obtained by a fivefold serial dilution of the drug from 12.5 to 0.02 jig ml-'. The cells were exposed to different Topo concentrations for 8 h, followed by washing with PBS, addition of new culture medium and continuation of incubation up to 72 h, before performing FMCA. Cells were exposed for the doubled concentrations for 4 h, and for four times the original concentrations for 2 h, before washing the Topo solution away
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