Response to BCNU of spheroids grown from mixtures of drug-sensitive and drug-resistant cells.

Multicellular spheroids were grown from mixtures of rat brain tumor cells sensitive (9L) and resistant (R3) to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). As shown previously, after treatment with 3 microM BCNU, percentages of each cell subpopulation in mixed-cell spheroids were estimated with the sister chromatid exchange (SCE) assay and found to be approximately the same as percentages used to initiate spheroids. The sensitivity of 9L cells in mixed-cell spheroids treated with BCNU, estimated by changes in the number of SCEs induced by treatment, decreased as the percentage of R3 cells increased. When spheroids were disaggregated into single cells before treatment, however, the number of SCEs induced in the 9L population did not decrease but remained at levels similar to those found for spheroids grown from 9L cells only. These data suggest that the cell-cell interactions that influence BCNU sensitivity in mixed cell spheroids depend on three-dimensional intercellular contact. The response of purely 9L, purely R3, and mixed-cell spheroids to BCNU was also determined using the cell survival and spheroid growth delay assays. The surviving fractions of individual spheroids treated with 40 microM BCNU were slightly greater than expected; growth delays found for mixed-cell spheroids were 2-3 days less than expected. These findings suggest that cells in mixed-cell spheroids are more resistant to BCNU than would be predicted from the sensitivities of purely 9L and R3 spheroids.