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They act through binding to the DNA minor groove and forming covalent adducts with guanine residues. However, the limited availability of structural data restricts mechanistic insights into their mode of action and hampers the discovery of novel compounds. We report crystal structures of duplex DNA adducts with first-, second-, and third-generation ecteinascidins. The structures show that trabectedin, lurbinectedin, and PM54 bind through their shared A- and B-subunits, forming a covalent bond with the N2 atom of guanine and an extensive network of noncovalent interactions, leading to significant minor groove widening. In contrast, their C-subunit, which differs across the compounds, establishes distinct contacts with the modified strand that affect binding strength and sequence specificity. These structural findings, supported by F\u00f6rster resonance energy transfer and biochemical assays, reveal the molecular determinants underlying differential sequence selectivity and reactivity. Our results provide a mechanistic framework for the anticancer activity of ecteinascidins and a structural basis to guide the design of next-generation analogues with improved therapeutic potential.<\/jats:p>","DOI":"10.1093\/nar\/gkag749","type":"journal-article","created":{"date-parts":[[2026,7,17]],"date-time":"2026-07-17T11:44:56Z","timestamp":1784288696000},"source":"Crossref","is-referenced-by-count":0,"title":["The structures of ecteinascidin anticancer agents bound to DNA shed light on their mechanism of action"],"prefix":"10.1093","volume":"54","author":[{"given":"Tommy","family":"Darri\u00e8re","sequence":"first","affiliation":[{"name":"Centro de Investigaciones Biol\u00f3gicas Margarita Salas, CSIC , Madrid 28040 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Federico M","family":"Ruiz","sequence":"additional","affiliation":[{"name":"Centro de Investigaciones Biol\u00f3gicas Margarita Salas, CSIC , Madrid 28040 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marta","family":"Mart\u00ednez-D\u00edez","sequence":"additional","affiliation":[{"name":"PharmaMar S.A. , Colmenar Viejo 28770 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marcelo Lima","family":"Ribeiro","sequence":"additional","affiliation":[{"name":"PharmaMar S.A. , Colmenar Viejo 28770 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carmen","family":"Cuevas","sequence":"additional","affiliation":[{"name":"PharmaMar S.A. , Colmenar Viejo 28770 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5097-731X","authenticated-orcid":false,"given":"Carlos","family":"Fern\u00e1ndez-Tornero","sequence":"additional","affiliation":[{"name":"Centro de Investigaciones Biol\u00f3gicas Margarita Salas, CSIC , Madrid 28040 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2026,7,29]]},"reference":[{"key":"2026072905572821600_B1","doi-asserted-by":"publisher","first-page":"111624","DOI":"10.1016\/j.jinorgbio.2021.111624","article-title":"Covalent and noncovalent interactions of coordination compounds with DNA: an overview","volume":"225","author":"Andrez\u00e1lov\u00e1","year":"2021","journal-title":"J Inorg Biochem"},{"key":"2026072905572821600_B2","doi-asserted-by":"publisher","first-page":"321","DOI":"10.2174\/0929867023371085","article-title":"DNA-binding of drugs used in medicinal therapies","volume":"9","author":"Bischoff","year":"2002","journal-title":"Curr Med Chem"},{"key":"2026072905572821600_B3","doi-asserted-by":"publisher","first-page":"26","DOI":"10.1039\/C8MD00425K","article-title":"Recent developments in compounds acting in the DNA minor groove","volume":"10","author":"Rahman","year":"2018","journal-title":"Med Chem Commun"},{"key":"2026072905572821600_B4","doi-asserted-by":"publisher","first-page":"409","DOI":"10.2174\/1568011054222355","article-title":"Target practice: aiming at satellite repeats with DNA minor groove binders","volume":"5","author":"Susbielle","year":"2005","journal-title":"Curr Med Chem Anticancer Agents"},{"key":"2026072905572821600_B5","doi-asserted-by":"publisher","first-page":"1213","DOI":"10.1351\/pac199365061213","article-title":"DNA\u2013drug interactions","volume":"65","author":"Kennard","year":"1993","journal-title":"Pure Appl Chem"},{"key":"2026072905572821600_B6","doi-asserted-by":"publisher","first-page":"328","DOI":"10.1039\/C4NP00051J","article-title":"Ecteinascidins. 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