Glufosfamide is a new agent for cancer chemotherapy. The objective of the study was the comparison of the in vitro drug resistance profile of glufosfamide with other oxazaphosphorines in 106 samples of childhood acute leukemia by means of the MTT assay. The following drugs were tested: glufosfamide, 4-HOO-ifosfamide, 4-HOO-cyclophosphamide, mafosfamide cyclohexylamine salt, prednisolone, vincristine, L-asparaginase, daunorubicin and cytarabine. In the group of initial Acute Lymphoblastic Leukemia (ALL) samples, equivalent cytotoxicity values for glufosfamide, 4-HOO-ifosfamide, 4-HOO-cyclophosphamide and mafosfamide were 5.95, 9.92, 4.60 and 3.90 microg/ml, respectively. In comparison to initial ALL samples, the relative resistance for glufosfamide and 4-HOO-ifosfamide in relapsed ALL samples were 1.9 (p=0.049) and 1.3 (ns), and in initial Acute Myeloblastic Leukemia (AML) samples, respectively, 31 (p<0.001) and 5 (p=0.001). All oxazaphosphorines showed highly significant cross-resistance. In conclusion, in vitro activity of glufosfamide is comparable to ifosfamide. Glufosfamide shows high activity against lymphoblasts both on diagnosis and on relapse, however it cannot circumvent resistance to other oxazaphosphorines.