AN ANALYSIS OF THE CYTOTOXIC EFFECTS OF ALKYLATING AGENTS IN CELL CULTURE

MARY EILEEN DOLAN, Purdue University

Abstract

The effect of chloroethylnitrosoureas, methylnitrosoureas methyltriazenes, and 1-alkyl-3-nitro-1-nitrosoguanidines on the growth rate of cultured P388, L1210 and 1,3-Bis(2-chloroethyl)-1-nitrosourea (BCNU) resistant P388 cells was studied. Dose functions have been established for these alkylating agents. These functions are used to analyse observed cytotoxic activity in relation to the amount of active alkylating species formed during the period of exposure to the agent. Dose function equations consider terms for the rate of partitioning of agent into the cell, the rate and yield of active intermediate formation and the duration of exposure period. The cytotoxic activity of alkylating agents under a variety of conditions including different initial drug concentration, length of exposure time, incubation temperature or different extracellular medium may be compared in terms of the concentration of intracellular active intermediate formed. Using this approach, it may be shown that the cytotoxicity of these agents is not dependent on initial dose, duration of exposure or extracellular medium, but on the number of alkylation events that take place in the cell. Furthermore, the effects caused by a variation of the temperature at which alkylnitrosoureas are incubated with P388 cells was found to be related to changes in the rate of conversion to active intermediate and not to a change in the sensitivity of cells to these drugs for the temperature range of 27 to 42(DEGREES)C. Structurally different compounds, such as the chloroethylnitrosoureas, which decompose to the same active intermediate are shown to have identical activity when the dose functions are applied. This was also found for the methylating agents that have different modes of activation. The cytotoxic activity of BCNU is inversely proportional to the intracellular thiol content. The decomposition of alkylnitronitrosoguanidines to active intermediate is accelerated in the presence of thiol containing compounds. A population of P388 cells found to have a 26% higher thiol content than normal cells were 2 to 3 fold less sensitive to the cytotoxic effects of 1-methyl-1-nitrosourea (MNU) and BCNU, respectively. This same population of cells was fully sensitive to the cytotoxic effects of 1-chloroethyl-3-nitro-1-nitrosoguanidine (ClENNG) and 1-methyl-3-nitro-1-nitrosoguanidine (MNNG).

Degree

Ph.D.

Subject Area

Pharmacology

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