Cytostatic and Cytotoxic Effects of Topotecan Decoded by a Novel Mathematical Simulation Approach

Abstract

Topotecan (TPT) is a topoisomerase I inhibitor, and like the other drugs of this family, it is believed to act in a specific way on cells in S phase at the time of treatment. Exploiting a new method, coupling a particular experimental plan with computer simulation, a complete quantitative study of the time dependence and dose dependence of the activity of cell cycle controls has become feasible, and the overall scenario of events after treatment can be reconstructed in detail. We were able to demonstrate that the response of an ovarian cancer cell line to 1 h of treatment with TPT is not limited to inhibition of DNA synthesis, leading to cell death, but involves G 1 and G 2 -M checkpoints. G 1 and G 2 -M block, recycling, and death follow specific dose-dependent kinetics, lasting no less than 3 days after treatment. We also found that cells treated outside S phase contribute significantly to the overall activity. The utility of this analysis was demonstrated by reproducing more complex treatment schemes in which low TPT concentrations were applied for 1 h three times at 24-h intervals. In this case, the simulation clarified the origin of the auto-potentiation observed with repeated 0.2 M treatments, in which the cytotoxicity, particularly against S-phase cells, was higher than the cytotoxicity in cells treated with 10 M only once. We believe that this approach will help us to understand the complexity and heterogeneity of the response of a cell population to a drug challenge and could help us to establish the rationale for drug scheduling or drug combinations.

The Contribution of p53 in the Dynamics of Cell Cycle Response to DNA Damage Interpreted by a Mathematical Model

“…At different times after treatment the cells were harvested and prepared for biparametric DNA-BrdUrd flow cytometric analysis as previously described. 16 The protocol for two-parameter flow cytometry analysis of DNA content and p53 level was previously described by Erba. 20 It was not possible to establish a clear-cut separation between cells with a raised p53 level and the non-specific fluorescence.…”

“…Biparametric DNA-dUTP analysis was made as previously described. 16 Monoparametric DNA histograms were analyzed as described. 21 Computer simulation of cell population growth and drug effects.…”

“…An important tool for quantitatively combining all the experimental data is the computer program already described in detail elsewhere. [14][15][16] Starting from the cell cycle distribution at a given time, this model predicts cell cycle flux. It reproduces the unperturbed growth of a cell population and its response to several drug concentrations, constructing a complete kinetic scenario based on a quantitative estimate of the time-and dose-dependence of the probabilities of cell arrest and killing.…”

“…The use of a wide range of drug concentrations, with a method permitting a distinction between cytostatic and cytotoxic effects should help clarify matters. We used an approach previously applied to study cell cycle effects induced by short treatment with cisplatin, 14 taxol, 15 topotecan 16 or melphalan 17 on a population of ovarian cancer cells. A large amount of experimental data (flow cytometric percentages and absolute cell numbers) were analyzed using a mathematical model that allows their interpretation in terms of the underlying phenomena of inhibition of DNA synthesis, G 1 and G 2 M block, death or recycling, as they superimpose on the "physiological" cell cycle progression.…”

Optimisation of Cancer Drug Treatments Using Cell Population Dynamics

“…Ubezio and co-workers [94,95,104,135] based themselves on an age and phase structured PDE model to develop a discrete age-structured model of cell cycle describing the time evolution of the number of cells of age a at time t in the phases G 1 , S and G 2 /M of the cell cycle. This model also takes into account the inter-cell variability in phase duration.…”