thioguanine-anhydrous and thiazolyl-blue

The clinical relevance of cellular drug resistance in children with acute lymphoblastic leukaemia (ALL) is unknown. The relation between in-vitro sensitivity to chemotherapeutic drugs at initial diagnosis and long-term clinical outcome was investigated in 44 children with ALL. The short-term MTT assay was used to assess sensitivity to prednisolone, vincristine, colaspase (asparaginase), daunorubicin, and thioguanine (instead of mercaptopurine which is unstable in vitro). For vincristine and colaspase there was no difference in outcome (probability of continuous complete remission) between sensitive and resistant patients. However, the probability of continuous complete remission was significantly lower in patients with resistant cells than in those with sensitive cells for thioguanine (p less than 0.01), daunorubicin (p less than 0.02), and prednisolone (p less than 0.05). For prednisolone there was a significant worsening of the prognosis (p less than 0.05) from the extremely sensitive patients through an intermediate group to the most resistant group. The prognostic significance of cellular drug resistance was independent of white-blood-cell count, age, sex, and hepatosplenomegaly. Leukaemic cells from boys were more resistant to thioguanine than those from girls. Thus, the short-term highly efficient MTT assay can help to predict long-term response to chemotherapy in childhood ALL.

Topics: Antineoplastic Agents; Asparaginase; Chi-Square Distribution; Child; Coloring Agents; Daunorubicin; Drug Resistance; Drug Screening Assays, Antitumor; Female; Humans; Male; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Prednisolone; Prognosis; Remission Induction; Tetrazolium Salts; Thiazoles; Thioguanine; Tumor Cells, Cultured; Vincristine

The knowledge about drug resistance in childhood leukemias and acute lymphoblastic leukemia (ALL) in general is limited. This is because of the lack of a suitable in vitro drug sensitivity assay, which is in part due to low in vitro ALL cell survival. We recently adapted the highly efficient 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) assay to test cells from ALL patients and showed that its results were comparable with those of the DiSC assay, up to now the most valid but laborious assay. In this study, in vitro drug sensitivity was assessed in cells from 82 children with leukemia, 79 of whom had ALL, with the MTT assay. Dose response curves were obtained for 6-mercaptopurine, 6-thioguanine (6-TG), prednisolone (Pred), daunorubicin (DNR), vincristine (VCR), cytosine arabinoside (Ara-C), L-asparaginase (L-Asp), mafosfamide, and mustine. A cytotoxic effect of methotrexate could be detected in only a few cases. Large interindividual differences in drug sensitivity were detected. Compared with leukemia cells from newly diagnosed patients, leukemia cells from relapsed patients were significantly more in vitro resistant to 6-TG, Pred, Ara-C, mafosfamide and mustine but not to DNR, VCR, and L-Asp. Improvements of culture medium and methods to increase MTT reduction were studied. From 10 components tested, addition of insulin and bovine serum albumin to serum-containing medium improved ALL cell survival. Addition of succinate did not increase the amount of MTT reduction. We conclude that the in vitro MTT assay highly facilitates large-scale studies on drug resistance of ALL patients that can lead to rational improvements in existing treatment protocols.

Topics: Asparaginase; Cells, Cultured; Child; Coloring Agents; Culture Media; Cyclophosphamide; Cytarabine; Daunorubicin; Dose-Response Relationship, Drug; Drug Resistance; Drug Screening Assays, Antitumor; Humans; Mechlorethamine; Mercaptopurine; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Prednisolone; Tetrazolium Salts; Thiazoles; Thioguanine; Vincristine