Summary 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.

Glucocorticoids (GC) are being used in the treatment of childhood leukemia for several decades, most successfully in newly diagnosed acute lymphoblastic leukemia (ALL). However, GC resistance is seen in 10-30% of untreated ALL patients, and is much more frequent in relapsed ALL and in acute nonlymphoblastic leukemia (ANLL). Sensitivity or resistance to GC can be measured using a cell culture drug resistance assay. For this purpose, we use the colorimetric methyl-thiazol-tetrazolium (MTT) assay. We have shown that GC resistance in childhood leukemia is related to clinical and cell biological features, and to the clinical outcome after multi-drug chemotherapy. These results are summarized in this review. In addition, we describe the apoptotic 'cell-lysis pathway' by which GC exert their antileukemic activity. This description provides a model to discuss the mechanisms of GC resistance, and to summarize the relevant literature. Possible levels of resistance relate to the diffusion of GC through the cell membrane, binding to the GC receptor (GCR), activation of the GC-GCR complex, translocation of the complex into the nucleus, binding to DNA, endonuclease-mediated DNA fragmentation, and DNA repair. A low number of GCR has been shown to be the cause of resistance in some children with ALL. However, GC resistance is likely to be caused at the post-receptor level in most leukemias. Unfortunately, there is still a lack of knowledge relating to the clinical relevance of mechanisms of GC resistance at the post-receptor level. Studies on the mechanisms of GC resistance other than those directly related to the GCR should be initiated, especially if patient material is used, as the results might indicate ways to circumvent or modulate GC resistance. A further increase in our knowledge regarding the relation between GC resistance and patient and cell biological features, the clinical relevance of GC resistance, and the mechanisms of GC resistance in leukemia patients, may contribute to further improvement in the results of GC therapy in leukemia.

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Gonadal function was evaluated in 23 men (aged 14.8\p=n-\28.8 years) treated in childhood with cytotoxic drugs for a solid tumour. Group 1 (N = 14) had been treated with non-alkylating drugs only, while group 2 (N=9) received the alkylating drug cyclophosphamide in addition (range 3.8\p=n-\19.5 g/m2).Median age at the start of treatment was 4.6 years (range 0.6\p=n-\16.1) in group 1 and 13.9 years (range 3.7\p=n-\16.9) in group 2. Data of the patients were compared with a reference group consisting of 14 normal men. Almost all patients of both groups showed normal development of puberty; 13 of the 14 men in group 1 showed normal hormonal values. In group 2, basal LH and FSH as well as the LH and FSH responses to GnRH showed higher levels compared to those of a reference group (p<0.001). Correlation analysis showed an evident correlation between the total dose of received cyclophosphamide and the basal FSH level (r=0.78; p=0.002), the FSH response to GnRH (r=0.73; p=0.002) and the LH response to GnRH (r=0.67; p=0.002). There was no correlation between the hormonal

DUP 785 (NSC 368390; Brequinar sodium) is a new inhibitor of pyrimidine de novo biosynthesis with antitumor activity against several experimental tumors. DUP 785 inhibits the mitochondrial enzyme dihydroorotate dehydrogenase, blocking the conversion of dihydroorotate to orotate. We examined the influence of exposure time to DUP 785 on its growth-inhibitory effects in L1210 murine leukemia and WiDR human adenocarcinoma cells and the effects of pyrimidine (deoxy) nucleosides on reversal of growth-inhibition. The results were correlated with changes in intracellular pyrimidine nucleotide pools and cell cycle distribution. In L1210 cells, a continuous exposure to 25 microM DUP 785 up to 96 hr caused complete growth inhibition. A 2 hr exposure of cells to the drug did not affect growth. In WiDR cells, exposure to the drug for 1-24 hr, followed by cultivation in drug-free medium resulted in recovery of growth. However, cells exposed to the drug for 48 hr or longer were not able to resume growth when recultured in drug-free medium. Reversal studies were performed to know whether selective depletion of one of the pyrimidine (deoxy) nucleotides might be related to the growth-inhibitory effects of DUP 785. Neither thymidine, deoxycytidine alone, deoxycytidine plus tetrahydrouridine; nor cytidine plus tetrahydrouridine added after 24 hr were able to reverse cell growth inhibition induced by 25 microM DUP 785. However, uridine and cytidine alone reversed growth inhibition. UTP and CTP pools in L1210 cells decreased to about 30-40% of control levels after 4 hr of drug exposure, while dTTP and dCTP pools decreased to about 30% of control levels. There were no significant changes in purine nucleotide pools. In WiDR cells, UTP and CTP pools decreased rapidly after drug exposure and were substantially depleted after 24 hr. Reculture of cells in drug-free medium resulted in a significant recovery of UTP and CTP levels only for cells exposed to DUP 785 for 1-24 hr. For cells exposed to the drug for 48 and 72 hr recovery of nucleotide pools was minimal. In L1210 cells, a 12-hr exposure to the drug caused an accumulation of cells in the early S-phase. In WiDR cells, there was a clear accumulation of cells in the S-phase of the cell cycle after 24 hr drug exposure.(ABSTRACT TRUNCATED AT 400 WORDS)