THE ANALYSIS OF THE DIFFERENCE IN ANTI-CANCER DRUG SENSITIVITY OF 3 CLONES SEPARATED FROM BLADDER CANCER CELL LINE

Abstract

One of the major problems with cancer chemotherapy is the development of drug resistance during treatment. Two mechanisms are considered as the cause of drug resistance, natural and acquired. It is now considered that cancers can be composed of multiple clonal subpopulations of cancer cells. In this study, we separated three clones (C1, C3 and C8) from NBT-2 (human bladder cancer cell line) by limiting dilution. We examined the growth rate and the transplantability to nude mice and performed chromosomal analysis of three clones. The doubling time of C1 is 22 hours, and those of C3 and C8 were 25 and 36 hours, respectively. Each clone was transplantable to nude mice, but we could not find out any histological difference among them. The chromosome numbers of C1 was 66, and those of C3 and C8 were 68 and 63, respectively. We could also find out karyotypic difference among them. We could therfore consider that these three clones had different biological features and studied the difference in drug sensitivity among these three clones and the parent cell line. Cells (1×10⁴/well) were incubated in microplates with ten different chemotherapeutic agents for 72hurs. Then ³H-thymidine (1μCi/ml) was added to each. After 24 hours, cells were harvested and the uptake of ³H thymidine was counted with a liquid scintillation counter. According to the reaction pattern, these chemotherapeutic agents were divided into three groups. 1. The radio isotope uptake of three clones and parent cell line was proportionally inhibited by increasing the drug concentration (carboplatin, (glycolato-o, o-) diammine platinum (II), ifosfamide). 2. Clones were inhibited at lower drug concentrations, but parent cell line was first inhibited at higher concentrations of drugs (adriamycine, vinblastine, peplomycin, bleomycin). 3. There was an extensive difference in sensitivity to methotrexate among three clones. Our results demonstrate the presence of clonal heterogeneity of sensitivity to chemotherapeutic agents and it seemes that C1 may be a clone which is naturally resistant to methotrexate.