Objective    To investigate the effect of combining use of rapamycin (RAP) and ACNU (nimuatine) on human glioma cell line U87-MG and their interaction.     Methods    Modified Karber’s method was used to test the lethal dose 50% (IC

50

) of RAP or ACNU to U87-MG cells. Then the cells were divided into control group

RAP groups (0.01

0.1

1

10 or 100 μmol/L)

ACNU groups (0.

1

1

10

100 or 1 000 μg/ml)

and combining groups (ACNU at above doses combined with RAP 1/4

1/2 or 3/4 IC

50

). Cell vitality of different groups was determined by MTT assay. The combined effect of RAP plus ACNU in inhibition of U87-MG cells was analyzed by isobolographic analysis. Cell cycle and apoptosis of U87-MG cells were evaluated by flow cytometry. Acridine orange and MDC staining were conducted to detect the autophagy of U87-MG cells. Ultrastructure of the cells was observed by transmission electron microscopy.     Results    As MTT assay showed

the tumor cell survival rate was inhibited significantly afer treatment of rapamycin alone or plus ACNU. IC

50

was 0.726±0.063 μmol/L in the RAP group

and 26.9±2.3 μg/ml in the ACNU group. Isobolographic analysis demonstrated that RAP and ACNU in combination had a synergistic effect during treatment. Although ACNU significantly induced apoptosis in U87-MG cells when compared with control group

no significant apoptosis was found in RAP groups. In RAP group

cells were arrested in G

1

phase. More apoptosis cells and cells arrested in G

1

phase were detected in combining RAP and ACNU group. Staining of acridine orange and MDC demonstrated that RAP induced autophagy alone

which was enhanced by ACNU treatment. Autophagosomes and apoptosis were found in combining groups by transmission electron microscopy.     Conclusion    RAP and ACNU had synergistic interactions in suppressing the growth and proliferation of U87-MG cells.