Modeling of the time-dependency of in vitro drug cytotoxicity and resistance

For potential clinical extrapolation of in vitro findings, it is of interes t to relate the measured effect of an anticancer agent to concentration and exposure time. The Hill model (A, V, Hill, J, Physiol., 40: iv-vii, 1910) is commonly used to describe pharmacodynamic (PD) effects, including drug-i nduced growth inhibition of cancer cells irt vitro, The ICXn x T = k relati onship, in which IC, is the concentration of agent required to reduce cell growth by X%, T is the exposure time, and II and k are estimable parameters , was first applied to bacterial disinfectant action and then was successfu lly used to model anticancer drug potency as a function of exposure time (D , J, Adams, Cancer Res., 49: 6615-6620, 1989), Our goal was to create a new global PD modeling paradigm to facilitate the quantitative assessment of t he growth-inhibitory effect of anticancer agents as a function of concentra tion and exposure time. Wild-type human ovarian A2780 and ileocecal HCT-8 c arcinoma cells and sublines that were resistant to cisplatin (A2780/CP3), d oxorubicin (A2780/DX5B), and raltitrexed (RTX) (HCT-8/DW2) were exposed to various anticancer agents, cisplatin, doxorubicin, paclitaxel, trimetrexate , RTX, methotrexate, and AG2034, for periods ranging from 1 to 96 h. Cell g rowth inhibition was measured with the sulforhodamine B protein dye assay. Patterns of time-dependency of drug potency, slope of the concentration-eff ect curves, and relative degree of resistance were characterized. Empirical mathematical expressions were built into a global concentration-time-effec t model. The global PD model was then fit to the concentration-time-effect data with iteratively reweighted nonlinear regression. Under specific treat ment conditions, the examination of the slope and the shape of the concentr ation-effect curves revealed a large heterogeneity in drug response, e.g., shallow concentration-effect curve or double or triple Hill "roller coaster " concentration-effect curve. These patterns, which were observed at interm ediate exposure times in parental and resistant cells for paclitaxel and tr imetrexate or only in resistant HCT-8/DW2 cells for RTX, methotrexate, and AG2034, revealed mechanistic insights for the former cases but possible met hodological artifacts for the latter cases. The comprehensive PD modeling o f the cytotoxic effect of anticancer agents showed that it was possible to modulate drug effect, response heterogeneity, and drug resistance by alteri ng the time of exposure to the agents. This approach will be useful for: (a ) describing complex concentration-time-effect surfaces; (b) refining biolo gical interpretations of data; (c) providing insights on mechanisms of drug action and resistance; and (d) generating leads for clinical use of antica ncer drugs.