Base sequence determinants of amonafide stimulation of topoisomerase II DNA cleavage.
A number of antitumor drugs including naphthalimides, a new class of intercalating agents, interfere with the DNA breakage-reunion activity of mammalian DNA topoisomerase II resulting in DNA cleavage stimulation. In this work, the sequence specificity of a lead compound of this series, amonafide, in stimulating DNA cleavage by murine topoisomerase II has been studied. Amonafide-stimulated cleavage intensity patterns were markedly different from those of other antitumor drugs by using pBR322 and SV40 DNAs. This drug had an unusually high site selectivity since about 60% of DNA cleavage was observed at only one site in pBR322 DNA, and at two sites in SV40 DNA. A total of ninety-four drug-stimulated sites were collected, and a statistical analysis of their sequences showed that amonafide highly prefers a cytosine, and excludes guanines and thymines instead, at position -1. A lower preference for an adenine at position +1 was also noted. In agreement with the statistical analysis, the DNA sequences of the three sites stimulated by amonafide at exceptionally high levels showed that the drug requirements of a cytosine (-1) and adenine (+1) were present in both the two strands. In addition, a particular feature of these prominent cleavage sites was the presence of an inverted repeat from position -3 to +7. Comparison of amonafide stimulation of DNA cleavage in oligonucleotides bearing base mutations at positions -2, -3 and/or +6, +7 suggested that DNA sequence, and not a putative cruciform structure, was critical for drug action. Moreover, the results showed that, for strong cleavage stimulation, the primary drug requirements at -1 and +1 positions were not sufficient and that the sequence 5'-WRC decreases A-3' (W, A or T; R, A or G) is required from -3 to +1 positions at both strands. The results suggest that the exceptionally high sequence specificity of amonafide is the result of optimal drug interactions with both the two enzyme subunits.
De Isabella
P
P
Division of Experimental Oncology B, Istituto Nazionale per lo Studio e la Cura dei Tumori, Milan, Italy.
Zunino
F
F
Capranico
G
G
D000225:Adenine; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism; D007094:Imides / Q000494:pharmacology*; D007546:Isoquinolines / Q000494:pharmacology*; D007941:Leukemia P388; D051379:Mice; D008969:Molecular Sequence Data; D053644:Naphthalimides; D063065:Organophosphonates; D017422:Sequence Analysis, DNA; D013539:Simian virus 40 / Q000235:genetics
D000970:Antineoplastic Agents; D004279:DNA, Viral; D007094:Imides; D007546:Isoquinolines; D053644:Naphthalimides; D063065:Organophosphonates; C037020:amonafide; D004247:DNA; D004250:DNA Topoisomerases, Type II; D000225:Adenine