Replication and homologous recombination repair regulate DNA double-strand break formation by the antitumor alkylator ecteinascidin 743.
Adducts induced by the antitumor alkylator ecteinascidin 743 (ET-743, Yondelis, trabectedin) represent a unique challenge to the DNA repair machinery because no pathway examined to date is able to remove the ET adducts, whereas cells deficient in nucleotide excision repair show increased resistance. We here describe the processing of the initial ET adducts into cytotoxic lesions and characterize the influence of cellular repair pathways on this process. Our findings show that exposure of proliferating mammalian cells to pharmacologically relevant concentrations of ET-743 is accompanied by rapid formation of DNA double-strand breaks (DSBs), as shown by the neutral comet assay and induction of focalized phosphorylated H2AX. The ET adducts are stable and can be converted into DSBs hours after the drug has been removed. Loss of homologous recombination repair has no influence on the initial levels of DSBs but is associated with the persistence of unrepaired DSBs after ET-743 is removed, resulting in extensive chromosomal abnormalities and pronounced sensitivity to the drug. In comparison, loss of nonhomologous end-joining had only modest effect on the sensitivity. The identification of DSB formation as a key step in the processing of ET-743 lesions represents a novel mechanism of action for the drug that is in agreement with its unusual potency. Because loss of repair proteins is common in human tumors, expression levels of selected repair factors may be useful in identifying patients particularly likely to benefit, or not, from treatment with ET-743.
Soares
Daniele Grazziotin
DG
Group of Cancer Biology and Therapeutics, Institut National de la Santé et de la Recherche Médicale, Unité 673, and Université Pierre et Marie Curie, Hôpital Saint-Antoine, 75571 Paris Cedex 12, France.
Escargueil
Alexandre E
AE
Poindessous
Virginie
V
Sarasin
Alain
A
de Gramont
Aimery
A
Bonatto
Diego
D
Henriques
João Antonio Pêgas
JA
Larsen
Annette K
AK
D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D002453:Cell Cycle / Q000187:drug effects; D002869:Chromosome Aberrations; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects*; D004260:DNA Repair; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000502:physiology; D004149:Dioxoles / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D006367:HeLa Cells; D006801:Humans; D011995:Recombination, Genetic; D044005:Tetrahydroisoquinolines / Q000494:pharmacology*; D000077606:Trabectedin
D018906:Antineoplastic Agents, Alkylating; D004268:DNA-Binding Proteins; D004149:Dioxoles; D044005:Tetrahydroisoquinolines; C110901:X-ray repair cross complementing protein 3; D000077606:Trabectedin