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ABSTRACT: In this study, we evaluated the potential of the distamycin derivative MEN 10716 as a telomerase inhibitor. Exposure of human melanoma cell extracts to MEN 10716 induced a dose-dependent inhibition of telomerase activity, with an IC50 of 24+/-3 microM. When intact JR8 melanoma cells were chronically exposed to the drug (200 microM every other day for 50 days), a marked inhibition (>80%) of the enzyme's catalytic activity was consistently observed starting from day 1. At later points in time, MEN 10716 inhibited melanoma cell proliferation and induced apoptosis. Cells surviving MEN 10716 exposure were characterised by a higher melanin content and a greater expression of p16(INK4A) protein than control cells. The effects of MEN 10716 were subsequently evaluated in different tumour cell systems. In particular, even in the H460 non-small cell lung cancer cell line, chronic exposure of the cells to the drug (100 microM every other day for 50 days) induced a consistent inhibition (>85%) of telomerase activity, a reduction of cell proliferation potential, and apoptosis. Conversely, MEN 10716 treatment did not appreciably inhibit cell proliferation in the U2-OS telomerase-negative human osteogenic sarcoma cell line. Interestingly, no variation in the mean telomere length was observed in MEN 10716-treated JR8 melanoma cells, whereas an appreciable increase in the mean telomere length was found in H460 lung cancer cells after drug exposure. Overall, the results of the study indicate that MEN 10716 is a possible telomerase inhibitor and suggest that abrogation of telomerase activity can affect cell proliferation even through pathways that are not dependent on telomere erosion.
European Journal of Cancer 09/2002; 38(13):1792-801. · 5.54 Impact Factor
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M Bigioni,
C Salvatore,
A Bullo, D Bellarosa,
E Iafrate,
F Animati,
G Capranico,
C Goso,
C A Maggi,
G Pratesi,
F Zunino,
S Manzini
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ABSTRACT: MEN 10755 is a disaccharide anthracycline endowed with a broader spectrum of antitumour activity than doxorubicin (DOX). To investigate the cellular and molecular basis of its action, cytotoxic activity, drug uptake, subcellular localisation, induction of DNA damage, and apoptosis were assessed in the human A2780 ovarian carcinoma cell line. Experiments with radiolabelled anthracyclines indicated that MEN 10755 exhibited reduced cellular accumulation and a different subcellular distribution (higher cytoplasmic/nuclear ratio) than DOX. In spite of the lower nuclear concentration, MEN 10755 was as potent as DOX in eliciting DNA single- and double-strand breaks, G2/M cell arrest, and apoptosis. Sequencing of drug-induced topoisomerase II cleavage sites showed a common DNA cleavage pattern for MEN 10755 and DOX. Cleavage sites were always characterised by the presence of adenine in -1 position. However, the extent of DNA cleavage stimulation induced by MEN 10755 was greater than that produced by DOX. Reversibility studies showed that MEN 10755-stimulated DNA cleavage sites were more persistent than those induced by DOX, thus suggesting a more stable interaction of the drug in the ternary complex. As a whole, the study indicated that the cellular pharmacokinetics of MEN 10755 substantially differs from that of DOX, showing a lower uptake and a different subcellular disposition. In spite of the apparently unfavourable cellular pharmacokinetics, MEN 10755 was still as potent as DOX in inducing topoisomerase-mediated DNA damage. Although the extent and persistence of protein-associated DNA breaks may contribute to the cytotoxic effects, the drug's efficacy as apoptosis inducer and antitumour agent could not be adequately explained on the basis of DNA damage mediated by the known target (i.e. topoisomerase II), thus supporting additional cellular effects that may be relevant in cellular response.
Biochemical Pharmacology 08/2001; 62(1):63-70. · 4.70 Impact Factor
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ABSTRACT: Cytotoxic drugs commonly used in cancer therapy promote tumor cell death by inducing apoptosis, but the cell death pathway(s) is likely dependent on the mechanism of drug action. In the present study, we investigated the mechanisms of cell death induced by doxorubicin (DXR) and the novel disaccharide anthracycline MEN 10755, in a human ovarian cancer cell line (A2780). Exposure to either anthracycline induced the up-regulation of several genes known to promote cell cycle arrest and DNA repair (WAF1/p21, GADD45) or apoptosis (bax, Fas). Although the expression of Fas was increased, an antagonistic anti-Fas antibody ZB4 did not inhibit anthracycline-induced apoptosis, suggesting that the stimulation of the Fas receptor did not play a critical role in the induction of apoptosis in this cell line. We also observed that neither MEN 10755 nor DXR were able to induce apoptosis in A2780 cells deprived of the nucleus but retaining an intact mitochondrial function (cytoplasts) and that apoptosis induced by either anthracycline was inhibited by cycloheximide, indicating that it is an active process requiring new protein synthesis. Both the caspases inhibitors, ZVAD-fmk and DEVD-cho, inhibited at similar extent apoptosis induced by either DXR or MEN 10755, suggesting an involvement of caspase-3 in this response. We conclude that, in a tumor cell line of epithelial origin, the apoptosis following exposure to anthracyclines is an active process requiring protein synthesis and drug interaction with nuclear structures. The pathway was Fas-independent but likely involved bax and caspase-3 as effectors of the cascade culminating in apoptosis.
Journal of Pharmacology and Experimental Therapeutics 03/2001; 296(2):276-83. · 3.83 Impact Factor
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ABSTRACT: On human endothelial cells from umbilical cord (HUVEC) are present, in addition to E- and P-selectins, their cognate ligands. Differently from selectins, the ligand expression is constitutive and not modulated by interleukin-1beta. Such ligands appear to be different from the ones present in promyelocytic cells in order to promote cell adhesion to immobilized selectins. The expression of selectin-ligands on HUVEC cells suggest that selectins can participate in endothelial signalling besides their role as adhesion molecules for circulating blood cells. However, despite their role in chemotaxis, selectins do not contribute to HUVEC tube formation in Matrigel.
Mediators of Inflammation 02/1998; 7(3):211-5. · 3.26 Impact Factor
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D Bellarosa,
G Antonelli,
F Bambacioni,
D Giannotti,
G Viti,
R Nannicini,
A Giachetti,
F Dianzani,
M Witvrouw,
R Pauwels,
J Desmyter,
E De Clercq
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ABSTRACT: A series of pyridobenzothiodiazepindioxides such as the 11-ethyl-6,8,9-trimethyl-6,11-dihydro-pyrido[2,3-f] [2,1,5]benzothiodiazepine-5,5-dioxide and arylpiridodiazepines such as the 6,7-dihydro-7-methyl-12-ethyl-pyrido[2,3-b] pyrido(2',3'-4,5]furo[2,3-f][1,4]diazepin-6(12H)-thio and the 6,7-dihydro-7-methyl-12-ethyl-pyrido[2,3-b]pyrido- [2,3-4,5]thieno[2,3-f][1,4] diazepin-6(12H)-thione were found to inhibit human immunodeficiency virus type 1 [HIV-1(IIIB)] replication at a concentration of 0.003-0.04 microM without being cytotoxic at a 3,000- to 15,000-fold higher concentration. These compounds proved effective against a variety of HIV-1 strains, including those that are resistant to 3'-azido-3' deoxythymidine (AZT), but not against HIV-2, simian immunodeficiency virus or herpes simplex virus. An HIV-1 strain containing the 188 Tyr-->His mutation in the reverse transcriptase displayed severely reduced sensitivity to the compounds. The specificity of these compounds is due to an interaction with the reverse transcription process. The 6,7-dihydro-7-methyl-12-ethyl-pyrido[2,3-b]pyrido [2,3-4,5]thieno[2,3-f][1,4]diazepin-6(12H)-thione (MEN 10979) enhanced the anti-HIV-1 activity of AZT and dideoxyinosine (ddI) in a synergistic manner. The new arylpyrido-diazepine and -thiodiazepine derivatives appear to be drug candidates for the treatment of HIV-1 infection.
Antiviral Research 05/1996; 30(2-3):109-24. · 4.30 Impact Factor
