Abstract. Postconfluent cultures of HT29 cells form a heterogeneous multilayer of which >95% of the cells are undifferentiated . In contrast, when stably adapted to normally lethal concentrations of methotrexate (10~-10-5 M), they form a monolayer of gobletlike cells (Lesuffieur et al ., 1990) which secrete large quantities of mucins and display a discrete brush border with the presence of villin, dipeptidylpeptidase-IV, and carcinoembryonic antigen . When adapted to even higher concentrations of methotrexate (10-4 and
The HT-29 cell line contains a small proportion of differentiated polarized, enterocytic and mucus-secreting cell types (less than 95%) which can be selected under various pressure conditions, e.g., glucose deprivation or methotrexate. The purpose of the present work was to investigate whether this also applies to 5-fluorouracil (FUra). Stepwise adaptation of exponentially growing cells to 1, 5, 10 and 20 microM FUra results, after a phase of high mortality, in the emergence of adapted sub-populations with stable growth rates and curves, and IC50 6, 18, 37, and 110 times higher than in untreated cells respectively. FUra-adapted cells are all differentiated, according to 2 phenotypes: (I) polarized dome-forming cells which express carcinoembryonic antigen at their apical surface and (2) goblet cells which secrete a mucus of colonic immunoreactivity. These phenotypes are present in the parental population and are different from those selected e.g., by glucose deprivation or methotrexate. This differentiation pattern is maintained when the cells are subcultured in drug-free medium. Resistance to FUra is acquired through gene amplification as substantiated by a 4- to 6-fold increase of thymidylate synthase gene copies in cells stably adapted to the drug. Whether the same mechanism or others are responsible for the first steps of resistance to FUra remains to be elucidated. Altogether, these results support the hypothesis that some of the cells which are present in the parental line and are committed to differentiation possess advantages which allow them to immediately resist and secondarily adapt to FUra. Comparison of the differentiation characteristics of FUra-adapted cells with those from cells selected under other pressure conditions suggests that resistance and adaptation to either type of pressure may depend on the differentiated phenotype to which the cells are committed.
Adaptation of the heterogeneous human colon carcinoma cell line HT-29 to lethal concentrations of methotrexate (MTX) and 5-fluorouracil (FUra) was shown to result in the emergence of sub-populations of cells all stably committed to differentiation. It was postulated that these populations result from selection of a few cells present in the parental line which possess, associated with their ability to differentiate, particular advantages allowing them to adapt to adverse conditions such as MTX or FUra. The purpose of the present study was to further verify this hypothesis by investigating whether HT-29 sub-populations selected for the commitment of all cells to differentiation would spontaneously be more resistant and adaptable than the parental cells to MTX and FUra. This study included a mucus-secreting clone (HT29-16E), a transporting clone (HT29-19A), and an enterocytic population selected by glucose deprivation (HT29-Glc-/+). Although all 3 populations show only a slight increase in their spontaneous resistance to both drugs, as substantiated by the values of IC50 which are only less than 2-fold higher than in parental cells, they are more adaptable as judged by growth curves, over a 50-day culture period, under exposure to 1 microM FUra and 0.1 microM MTX. In sharp contrast to parental cells, which, at these concentrations, show a high rate of mortality, all 3 populations, although growing slowly, reach densities more or less close, depending on the drug and population concerned, to that of control untreated cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Steroid sulfatase (STS) inhibition may have a therapeutic role in suppression of endocrine-responsive breast cancer. This study aimed to determine the optimal biological dose and recommended dose (RD) of the STS inhibitor irosustat. A three-part, open-label, multicenter, dose escalation study of irosustat in estrogen receptor-positive breast cancer patients involved administration of a single dose of irosustat with a 7-day observation period; followed by a daily oral dose of irosustat for 28 days; and an extension phase, in which the daily oral dose of irosustat was continued at the discretion of the investigator and as long as the patient was benefitting from the treatment. Five doses of irosustat were tested (1, 5, 20, 40, and 80 mg) in 50 patients. After 28 days of daily administration of irosustat, all the evaluated patients in the 5, 20, 40, and 80 mg cohorts achieved ≥95 % STS inhibition in peripheral blood mononuclear cells and corresponding endocrine suppression. The maximum tolerated dose was not reached, and the 40 mg dose was established as the RD. The median time to disease progression in the 40 mg cohort was 11.2 weeks. Disease stabilization was achieved in 10 % of patients potentially indicative of drug activity. Dry skin was the most frequent adverse event. The RD of irosustat is 40 mg. Disease stabilization occurred in 10 % of this heavily pretreated patient population. A larger study is required to define an accurate response rate to irosustat as a single agent and whether co-administration with an aromatase inhibitor is needed.
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