Lonidamine, a dichlorinated derivative of indazole-3-carboxylic acid, was shown to play a significant role in reversing or overcoming multidrug resistance. Here, we show that exposure to 50 g/ml of lonidamine induces apoptosis in adriamycin and nitrosourea-resistant cells (MCF-7 ADR R human breast cancer cell line, and LB9 glioblastoma multiform cell line), as demonstrated by sub-G1 peaks in DNA content histograms, condensation of nuclear chromatin, and internucleosomal DNA fragmentation. Moreover, we find that apoptosis is preceded by accumulation of the cells in the G0/G1 phase of the cell cycle. Interestingly, lonidamine fails to activate the apoptotic program in the corresponding sensitive parental cell lines (ADR-sensitive MCF-7 WT, and nitrosourea-sensitive LI cells) even after long exposure times. The evaluation of bcl-2 protein expression suggests that this different effect of lonidamine treatment in drug-resistant and -sensitive cell lines might not simply be due to dissimilar expression levels of bcl-2 protein.To determine whether the lonidamine-induced apoptosis is mediated by p53 protein, we used cells lacking endogenous p53 and overexpressing either wild-type p53 or dominant-negative p53 mutant. We find that apoptosis by lonidamine is independent of the p53 gene. ( J. Clin. Invest. 1996.

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Two continuous human glioma derived cell lines, LI and DF, were established in our laboratory. Both cell lines showed cytological features and in vitro behavior similar to those of the respective original neoplasms. These two lines were characterized for their main biological properties including in vitro and in vivo growth rate, clonogenic ability and tumorigenicity in nude mice. The plating efficiencies were generally high both during exponential and stationary growth phases and a high tumorigenicity was observed. All injected nude mice developed tumors. The two lines were tested for chemosensitivity to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and cis-Diamminedichloroplatinum II (DDP). Heterogeneity in biological features and in drug sensitivity was observed. Exposure of the two lines to BCNU and DDP showed that the glioblastoma (LI) was less sensitive than the anaplastic astrocytoma (DF). For both lines BCNU was more effective on cells in plateau than in exponential phase, while the killing effect of DDP was not phase-dependent.

Summary In HT29 cells 5-fluorouracil (5FU) cytotoxicity is enhanced by subsequent incubation of cells in medium containing 1% N-methylformamide (NMF). This enhancement does not appear to be related to differences in the repair of 5FU-induced DNA damage. It is proposed that the inhibition of DNA synthesis by NMF (that is reversible and does not result in any detectable toxicity) becomes a lethal event in a cell in which DNA synthesis has already been altered by 5FU exposure. The synergism is sequence dependent (i.e. it does not occur when NMF is given before 5FU) and specific for some cell types as shown by the fact that no synergism was found in L1210 mouse leukaemia cells. In nude mice transplanted s.c. with HT29 cells daily 5FU treatment (for 5 days) followed by daily NMF treatment (for 10 days) caused much greater inhibition of tumour growth than either drug alone or the same combination given in the opposite order (NMF then 5FU). These results, if confirmed on other human colon tumours, could be of clinical interest as a means of increasing the therapeutic efficacy of 5FU in patients with colon cancer.The polar solvent N-methylformamide (NMF) has recently been reported to induce the expression of a better differentiated or less malignant phenotype in both rodent and human tumour cells (Langdon & Hickman, 1987). Cellular alterations associated with NMF-mediated induction of differentiation include changes in morphology, clonogenicity, tumorigenicity and cell culture doubling time (Zupi et al., 1988), which ultimately result in a lower tumour aggressiveness. NMF-treated cells revert to the original phenotype upon removal of the inducer, but the maturational treatment has been associated with increased tumour cell sensitivity to certain cytotoxic agents, such as X-rays, cisplatinum and bleomycin (Dexter et al., 1984;Harpur et al., 1986;Langdon et al., 1985;Leith et al., 1985Leith et al., , 1986. It has been proposed that the sequential use of cytotoxic and differentiating agents might improve anti-cancer therapy by preventing or slowing tumour cell proliferation and phenotypic diversification (Lotan & Nicolson, 1988).In the human colon cancer cell line HT29 we have already studied the lethal effects of the combination of NMF and 5-fluorouracil (5FU) (Zupi et al., 1988). 5FU cytotoxicity was potentiated by subsequent exposure of cells to a nontoxic dose of NMF (1%), but when the two compounds were given in the opposite sequence there was no synergism. The enhanced cytotoxicity of the sequence 5FU to NMF could be related to the NMF-treated cells being less able to recover from the sublethal damage produced by the anti-metabolite.In order to shed some light on the mechanism of this potentiation we designed a study to assess the DNA damage and alteration of macromolecule synthesis caused by 5FU alone or SFU followed by NMF on HT29 cells. Materials and methods Cell culturesThe HT29 colon adenocarcinoma cell line was maintained as monolayer culture in RPMI 1640 medium supplemented with 10% FCS, L-glutamine and antibi...

Six cell lines were derived from pleural effusions of two lung cancer patients and established in vitro in our laboratory. Cell line AE1 was obtained from a small cell lung cancer (SCLC) before the patient had received any chemotherapy; the other lines (AE2 and AE3) were established from tumour recurrences in the same patient after therapy. Cell lines DG1 and DG2 were derived from specimens of an untreated non-small cell lung cancer (NSCLC), while cell line DG3 originated from pleural effusions recurring in the same patient after therapy. The results of the present study show that: (a) the SCLC lines AE1, AE2 and AE3 are heterogeneous in their biological characteristics and in their chemosensitivity patterns. In particular lines AE2 and AE3 are less responsive to cis-Platinum (DDP) and Adriamycin (ADM) than line AE1, so that they may reflect resistant subpopulations existing within the original tumour, selected following therapy with these drugs. In contrast, however, line AE1 proved more resistant to Vepesid (VP16) than lines AE2 and AE3. (b) The three NSCLC lines are similar in various biological features as well as in their chemosensitivity to DDP and Vinblastine (VBL).