Interpreting cell cycle effects of drugs: the case of melphalan.
Multiple effects usually occur in the cell cycle, during and after the exposure to a drug, while treated cells flowing through the cycle encounter G1, S and G2M checkpoints. We developed a simulation tool connecting the microscopic level of the cellular response in G1, S and G2M with the experimental data of growth inhibition and flow cytometry. We found that multiple-often not intuitive-combinations of cytostatic and cytotoxic effects can be in keeping with the observations. This multiplicity of interpretation can be strongly reduced by considering together data with different methods, ideally reaching a reconstruction of the underlying cell cycle perturbations. Here, we propose an experimental plan including a time course of DNA flow cytometry and absolute cell count measurements with several drug concentrations and a limited number of flow cytometric DNA-Bromodeoxyuridine and TUNEL analyses, coupled with computer simulation. We showed its use in the attempt to define the complete time course of the effects of melphalan on three cancer cell lines. After drug treatment, each subset of cells experienced blocks and lethality in all phases of the cell cycle, but the dynamics was different, the differences being strongly dose-dependent. Our approach allows a better appreciation of the complexity of the cell cycle phenomena associated with drug treatment. It is expected that such level of understanding of the time- and dose-dependence of the cytostatic and cytotoxic effects of a drug might support rational therapeutic design.
Lupi
Monica
M
Biophysics Unit, Laboratory of Cancer Pharmacology, Department of Oncology, Istituto di Ricerche Farmacologiche Mario Negri, Via Eritrea 62, 20157, Milano, Italy. ubezio@marionegri.it
Cappella
Paolo
P
Matera
Giada
G
Natoli
Claudia
C
Ubezio
Paolo
P
D000964:Antimetabolites, Antineoplastic / Q000378:metabolism; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D001973:Bromodeoxyuridine / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects*; D016923:Cell Death / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D003198:Computer Simulation; D004273:DNA, Neoplasm / Q000737:chemistry / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate; D016193:G1 Phase / Q000187:drug effects; D016195:G2 Phase / Q000187:drug effects; D006801:Humans; D020287:In Situ Nick-End Labeling; D007700:Kinetics; D008558:Melphalan / Q000494:pharmacology*; D016196:S Phase / Q000187:drug effects; D013997:Time Factors
D000964:Antimetabolites, Antineoplastic; D018906:Antineoplastic Agents, Alkylating; D004273:DNA, Neoplasm; D001973:Bromodeoxyuridine; D016650:Fluorescein-5-isothiocyanate; D008558:Melphalan