Establishment and characterization of an in vitro model of acquired resistance to cisplatin in a human testicular nonseminomatous germ cell line.
Clinically, human testicular nonseminomatous germ cell tumors exhibit remarkable sensitivity to platinum-based chemotherapy. To define better the mechanistic basis for this unusual sensitivity, the biochemical determinants of platinum-induced cytotoxicity have been investigated in a human testicular tumor cell line (GCT27) established from a previously untreated patient and in an in vitro derived 5.6-fold cisplatin-resistant stable variant (GCT27cisR). Compared to 12 ovarian and 5 cervical human tumor cell lines, the parent GCT27 line was among the most sensitive to the cytotoxic effects of both cisplatin (dosage producing 50% inhibition, 0.2 microM) and carboplatin (dosage producing 50% inhibition, 2.9 microM), thus reflecting clinical data. A 4-day exposure sulforhodamine B-staining assay was used to determine that GCT27cisR was cross-resistant to carboplatin and iproplatin and the classical bifunctional alkylating agents melphalan and chlorambucil. Partial cross-resistance was observed to tetraplatin, methotrexate, and mitomycin C. No cross-resistance was observed to Adriamycin, etoposide, vinblastine, bleomycin, 1-beta-D-arabinofuranosylcytosine, and 5-fluorouracil. Intracellular cisplatin accumulation across the dose range 2.5-100 microM (for 2 h) was 1.6 +/- 0.39-fold (mean +/- SD) greater for the parent line. There was no significant difference in glutathione levels between the two lines. The acquired resistance line was 1.9-fold more resistant than the parent line to the cytotoxic effects of cadmium chloride. There was no significant difference between the two lines, however, in the total amounts of platinum bound to DNA after cisplatin exposure (25, 50, or 100 microM for 2 h). The removal of total platinum adducts from DNA was significantly faster for GCT27cisR compared to the parent line (half-times of removal, 32 and 67 h, respectively). These data suggest that the abnormal sensitivity of the parent testicular tumor cell line to platinum-containing anticancer drugs may be due predominantly to an inherent defect in the ability of these cells to remove platinum from their DNA. This defect is apparently lost in the acquired resistance counterpart. Reduced intracellular accumulation and increased cytoplasmic concentrations of metallothionein may also contribute, in part, to the acquisition of cisplatin resistance in this model.
Kelland
L R
LR
Drug Development Section, Institute of Cancer Research, Belmont, Sutton, Survey, United Kingdom.
Mistry
P
P
Abel
G
G
Freidlos
F
F
Loh
S Y
SY
Roberts
J J
JJ
Harrap
K R
KR
D000970:Antineoplastic Agents / Q000494:pharmacology*; D002104:Cadmium / Q000494:pharmacology; D019256:Cadmium Chloride; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects*; D002945:Cisplatin / Q000494:pharmacology*; D004273:DNA, Neoplasm / Q000378:metabolism; D004351:Drug Resistance / Q000502:physiology*; D005260:Female; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D008297:Male; D009373:Neoplasms, Germ Cell and Embryonal; D010051:Ovarian Neoplasms; D013736:Testicular Neoplasms; D002583:Uterine Cervical Neoplasms
D000970:Antineoplastic Agents; D004273:DNA, Neoplasm; D002104:Cadmium; D005978:Glutathione; D019256:Cadmium Chloride; D002945:Cisplatin