NMR-Based Model of an Ecteinascidin 743−DNA Adduct

Moore, Bob M.; Seaman, Fred C.; Hurley, Laurence H.

doi:10.1021/ja9704500

Cited by 78 publications

(66 citation statements)

References 8 publications

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“…Indeed, it has been shown to bind in the minor groove of DNA and to alkylate the N2 position of guanine with some degree of specificity. 6,7 Such ET-743 minor-groove alkylation induces a bend in the DNA helix toward the major groove, a finding not common to the other minor groove-alkylating agents, which bend DNA into the minor groove. 8 No single-strand breaks, double-strand breaks or DNAprotein cross-links could be found by alkaline elution in cells exposed to IC 50 concentrations of ET-743.…”

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confidence: 98%

Unique pattern of ET-743 activity in different cellular systems with defined deficiencies in DNA-repair pathways

et al. 2001

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The cytotoxic activity of ecteinascidin 743 (ET-743), a natural product derived from the marine tunicate Ecteinascidia turbinata that exhibits potent anti-tumor activity in pre-clinical systems and promising activity in phase I and II clinical trials, was investigated in a number of cell systems with well-defined deficiencies in DNA-repair mechanisms. ET-743 binds to N2 of guanine in the minor groove, but its activity does not appear to be related to DNA-topoisomerase I poisoning as the drug is equally active in wild-type yeast and in yeast with a deletion in the DNA-topoisomerase I gene. Defects in the mismatch repair pathway, usually associated with increased resistance to methylating agents and cisplatin, did not affect the cytotoxic activity of ET-743. However, ET-743 did show decreased activity (from 2-to 8-fold) in nucleotide excision repair ( Ecteinascidin 743 (ET-743) is a natural product derived from the marine tunicate Ecteinascidia turbinata, selected for its significant anti-tumor activity in different in vitro and in vivo models. [1][2][3] Currently, ET-743 is undergoing phase II clinical trials after having shown activity in phase I clinical studies with responses observed in different human tumors, including soft tissue sarcoma, osteosarcoma, melanoma and breast cancer. 4,5 The mechanism of action of ET-743 has yet to be fully defined, but DNA appears to be the primary target. Indeed, it has been shown to bind in the minor groove of DNA and to alkylate the N2 position of guanine with some degree of specificity. 6,7 Such ET-743 minor-groove alkylation induces a bend in the DNA helix toward the major groove, a finding not common to the other minor groove-alkylating agents, which bend DNA into the minor groove. 8 No single-strand breaks, double-strand breaks or DNAprotein cross-links could be found by alkaline elution in cells exposed to IC 50 concentrations of ET-743. 9 However, Takebayashi et al. 10 showed that ET-743 was able to induce DNA-topoisomerase I cross-linking but that high concentrations were required to produce the effect.To provide some insight into the mechanism of action of ET-743, we have evaluated its cytotoxic activity using a panel of different cellular systems characterized by well-defined deficiencies in different mammalian repair pathways. Our data indicate that ET-743 has a unique mechanism of interaction with DNA. MATERIAL AND METHODS Cells and drugsYeast strains and the isogenic derivatives CY2823 (⌬rad52) and CY2278 (⌬topI) (kindly provided by Dr. M. Lopes, Instituto F.I.R.C. di Oncologia Molecolare, Milan, Italy) were grown in YPD plates (1% yeast extract, 2% bacto-peptone, 2% glucose) with or without the drugs.The Chinese hamster ovary (CHO) parental cell line CHO-AA8 and the UV-sensitive DNA repair-deficient mutant cell lines CHO-UV23, CHO-UV61 and CHO-UV96 (hereafter named UV23, UV61 and UV96) 11 were kindly provided by Dr. M. Stefanini (CNR IGBE, Pavia, Italy). The UV96 cell line was transfected by the calcium phosphate technique using 10 g of a CMV-ERCC1 plasmid enc...

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Unique pattern of ET-743 activity in different cellular systems with defined deficiencies in DNA-repair pathways

et al. 2001

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“…ET-743 has been shown to alkylate DNA, preferentially at GC-rich sequences (8,9). Previous reports have shown that ET-743 inhibits at the M range (Ͼ10 M) the activity of various DNA-binding factors, including TBP, E2F, SRF, and NF-Y (11).…”

Section: Discussionmentioning

confidence: 99%

“…Two recent studies reported that relatively low concentrations (10 -50 nM) inhibited the transcriptional activation of cellular genes regulated by the NF-Y transcription factor, including MDR1, and possibly by the Sp1 transcription factor (12,43). However, although ET-743 interaction with the DNA seems to be well characterized (8,9), the molecular mechanisms by which such an interaction leads to growth arrest and cytotoxicity are not well understood. The data reported here, showing that pharmacologically relevant concentrations (nM) of ET-743 induce a potent apoptotic response, indicate that triggering of apoptosis can be the major action in ET-743 cytotoxicity.…”

Section: Discussionmentioning

confidence: 99%

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Differential Cytostatic and Apoptotic Effects of Ecteinascidin-743 in Cancer Cells

Gajate

Mollinedo

2002

Journal of Biological Chemistry

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We have found that ecteinascidin-743 (ET-743) inhibited cell proliferation at 1-10 ng/ml, leading to S and G 2 /M arrest and subsequent apoptosis, and induced early apoptosis without previous cell cycle arrest at 10 -100 ng/ml in cancer cells. ET-743-mediated apoptosis, did not involve Fas/CD95. ET-743 induced c-Jun NH 2 -terminal kinase (JNK) and caspase-3 activation, and JNK and caspase inhibition prevented ET-743-induced apoptosis. ET-743 failed to promote apoptosis in caspase-3-deficient MCF-7 cells, further implicating caspase-3 in its proapoptotic action. Overexpression of bcl-2 by gene transfer abrogated ET-743-induced apoptosis, but cells underwent cell cycle arrest. ET-743 triggered cytochrome c release from mitochondria that was inhibited by Bcl-2 overexpression. Inhibition of transcription or protein synthesis did not prevent ET-743-induced apoptosis, but abrogated ET-743-induced cell cycle arrest. Microarray analyses revealed changes in the expression of a small number of cell cycle-related genes (p21, GADD45A, cyclin G2, MCM5, and histones) that suggested their putative involvement in ET-743-induced cell cycle arrest. These data indicate that ET-743 is a very potent anticancer drug showing dose-dependent cytostatic and proapoptotic effects through activation of two different signaling pathways, namely a transcriptiondependent pathway leading to cell cycle arrest and a transcription-independent route leading to rapid apoptosis that involves mitochondria, JNK, and caspase-3. Ecteinascidin-743 (ET-743)1 is a marine-derived compound isolated from the marine tunicate Ecteinascidia turbinata (1, 2), with a potent cytotoxic activity against a variety of tumors in vitro and in vivo (3-5). The preclinical in vivo experiments with ET-743 showed cytotoxic activity of the drug when administered at g/m 2 dosages, yielding nanomolar plasma concentrations (6, 7). Current phase II clinical trials in Europe and the United States indicate that ET-743 represents a highly promising antitumor agent. However, the mechanism by which ET-743 exerts its anticancer activity remains to be elucidated. ET-743 has been reported to bind to the minor groove of DNA (8, 9), bending DNA toward the major groove (10). DNA-bound ET-743 appeared to modify the interaction between DNA and several transcription factors (11,12). Also, at high concentrations ET-743 and the related synthetic drug phthalascidin, have been reported to target topoisomerase I (13, 14). However, the relevance of these actions for the antitumor activity of ET-743 can be questioned as they are evidenced at drug concentrations much higher than those required for achieving its antitumor effect. On the other hand, ET-743-treated cells have been reported to accumulate in S and G 2 /M phases (15-18).To elucidate the mechanism underlying the anticancer effect of ET-743, we investigated the putative role of apoptosis in ET-743 action as an explanation for its cytotoxic effect. In this work we have found evidence for the induction of c-Jun NH 2 -terminal kinase (JNK)-, mitoch...

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“…It binds to DNA in a sequence-specific manner and forms a covalent adduct with the N-2 amino group of guanine bases in the minor groove, which bends the double helix toward the major groove [13,14]. Recognition of these drug-associated lesions by the transcription-coupled nucleotide-excision repair system, resulting in the generation of lethal single-strand breaks in DNA, appears to be the principal mechanism of cell death [15].…”

Section: Introductionmentioning

confidence: 99%

A Phase II and Pharmacokinetic Study of Ecteinascidin 743 in Patients with Gastrointestinal Stromal Tumors

et al. 2002

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Purpose. To assess the efficacy, tolerability, and pharmacokinetics of ecteinascidin 743 (ET-743) in patients with advanced gastrointestinal stromal tumors (GISTs).Patients and Methods. The study was confined to adult patients with radiographically measurable GISTs. ET-743 was administered as a 24-hour continuous i.v. infusion at a dose of 1.5 mg/m 2 repeated every 3 weeks. Pharmacokinetic blood sampling was performed during the first cycle of therapy. Tumors were restaged after every second cycle of therapy.Results. A total of 20 patients was enrolled in the study, 19 of whom were treated with 47 cycles of ET-743 (median 2, range 1-10). Severe toxicities were limited to reversible grade 3 transaminitis in 10 patients and grade 3 fatigue in one patient. There were no objective responses, and disease stabilization occurred in two patients lasting for periods of 4 and 10 months. The 1-year survival rate was 71.1%. Mean ± standard deviation values of the maximum plasma concentration and total plasma clearance were 1.1 ± 0.4 ng/ml and 44 ± 16 l/h/m 2 , respectively, for 19 of the 20 patients.Conclusion. This study is the first report of a prospective phase II trial to evaluate a cytotoxic agent in patients with GISTs. This study underscores the primary resistance of GISTS to chemotherapy and stands in stark contrast to the encouraging results recently achieved with STI571. The lack of response may be associated with a therapeutically ineffective exposure to the drug based upon the lower incidence of severe toxicities and greater clearance than described in phase I and II trials of ET-743. The Oncologist 2002;7:531-538 The Oncologist 2002;7:531-538 www.TheOncologist.com Correspondence: David P. Ryan, M.D., MGH Cancer Center, 100 Blossom Street, Cox 640, Boston, Massachusetts 02114, USA. Telephone: 617-724-4000; Fax: 617-724-3166; e-mail: dpryan@partners.org Received August 6, 2002; accepted for publication October 17, 2002. ©AlphaMed Press 1083-7159/2002 The Oncologist ® LEARNING OBJECTIVESAfter completing this course, the reader will be able to:1. Describe the mechanism of action, administration, and pharmacokinetics of ecteinascidin 743.2. Recognize the classification and pathophysiology of gastrointestinal stromal tumors.3. Appreciate that conventional cytotoxic chemotherapy has no efficacy in patients with gastrointestinal stromal tumors and that STI571 has dramatic efficacy.Access and take the CME test online and receive one hour of AMA PRA category 1 credit at CME.TheOncologist.com CME CME

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NMR-Based Model of an Ecteinascidin 743−DNA Adduct

Cited by 78 publications

References 8 publications

Unique pattern of ET-743 activity in different cellular systems with defined deficiencies in DNA-repair pathways

Unique pattern of ET-743 activity in different cellular systems with defined deficiencies in DNA-repair pathways

Differential Cytostatic and Apoptotic Effects of Ecteinascidin-743 in Cancer Cells

A Phase II and Pharmacokinetic Study of Ecteinascidin 743 in Patients with Gastrointestinal Stromal Tumors

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