Abstract

In vitro drug sensitivity of leukaemic cells might be influenced by the contamination of such a sample with non-malignant cells and the sample source. To study this, sensitivity of normal peripheral blood (PB) lymphocytes to a number of cytostatic drugs was assessed with the MTT assay. We compared this sensitivity with the drug sensitivity of leukaemic cells of 38 children with acute lymphoblastic leukaemia. We also studied a possible differential sensitivity of leukaemic cells from bone marrow (BM) and PB. The following drugs were used: Prednisolone, dexamethasone, 6-mercaptopurine, 6-thioguanine, cytosine arabinoside, vincristine, vindesine, daunorubicin, doxorubicin, mafosfamide (Maf), 4-hydroperoxy-ifosfamide, teniposide, mitoxantrone, L-asparaginase, methotrexate and mustine. Normal PB lymphocytes were significantly more resistant to all drugs tested, except to Maf. Leukaemic BM and PB cells from 38 patients (unpaired samples) showed no significant differences in sensitivity to any of the drugs. Moreover, in 11 of 12 children with acute leukaemia of whom we investigated simultaneously obtained BM and PB (paired samples), their leukaemic BM and PB cells showed comparable drug sensitivity profiles. In one patient the BM cells were more sensitive to most drugs than those from the PB, but the actual differences in sensitivity were small. We conclude that the contamination of a leukaemic sample with normal PB lymphocytes will influence the results of the MTT assay. The source of the leukaemic sample, BM or PB, does not significantly influence the assay results.

Highlights

  • MethodsPrednisolone disodiumphosphate (PRD), dexamethasone disodiumphosphate (DXM), daunorubicin (DNR), L-asparaginase (L-Asp), mustine hydrochloride (Must), cytosine arabinoside (Ara-C), vindesine (VDS), vincristine (VCR), mitoxantrone (Mitox), methotrexate (MTX), and teniposide (Teni) were obtained from our hospital pharmacy, together with acidified (0.04 N HCI) isopropanol; 6-thioguanine (6TG), 6-mercaptopurine (6-MP), and doxorubicin (Dox) from Sigma; mafosfamide (Maf, 4-hydroxycyclofosfamide) and 4hydroperoxy-ifosfamide (4-HI), active derivatives of cyclofosfamide (CFM) and ifosfamide (IFM) respectively, were kindly provided by ASTA Pharma AG

  • In one patient the bone marrow (BM) cells were more sensitive to most drugs than those from the peripheral blood (PB), but the actual differences in sensitivity were small

  • Cells were suspended in RPMI 1640 (Gibco, Dutch modification), containing 20% foetal calf serum, 2 mM L-glutamine, 1OOIUml-' penicillin, 100l gml-' streptomycin, 0.125 lg ml-' fungizone, 200 igmlml gentamycin, all obtained from Flow Laboratories, and 5 g ml1' insulin, 5 ig ml' transferrin, and 5 ng ml' sodium selenite from Sigma

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Summary

Methods

Prednisolone disodiumphosphate (PRD), dexamethasone disodiumphosphate (DXM), daunorubicin (DNR), L-asparaginase (L-Asp), mustine hydrochloride (Must), cytosine arabinoside (Ara-C), vindesine (VDS), vincristine (VCR), mitoxantrone (Mitox), methotrexate (MTX), and teniposide (Teni) were obtained from our hospital pharmacy, together with acidified (0.04 N HCI) isopropanol; 6-thioguanine (6TG), 6-mercaptopurine (6-MP), and doxorubicin (Dox) from Sigma; mafosfamide (Maf, 4-hydroxycyclofosfamide) and 4hydroperoxy-ifosfamide (4-HI), active derivatives of cyclofosfamide (CFM) and ifosfamide (IFM) respectively, were kindly provided by ASTA Pharma AG PRD (of which 75% corresponds to pure prednisolone) was dissolved in saline. DNR, L-Asp, Must, VDS, and Dox were dissolved in distilled water, 6-MP and 6-TG in 0.1 N NaOH, Maf in PBS, and 4-HI in DMSO/distilled water (1:1). DXM, Ara-C, VCR, Mitox, MTX and Teni were obtained in soluble form.

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