Abstract 3506: Rules of DNA adduct formation for pyrrolobenzodiazepine (PBD) dimers

Abstract

Abstract Pyrrolo[2,1-c][1,4]benzodiazepine (PBD) dimers are synthetic sequence-selective DNA minor-groove cross-linking agents developed from the tricyclic anthramycin family of naturally-occurring antitumor agents. The “lead” agent, SJG-136, is about to enter Phase II clinical trials as a “first-in-class” sequence-selective DNA minor-groove binding agent. PBDs possess electrophilic imine or carbinolamine moieties that form covalent aminal linkages with guanine C2-NH2 functionalities within the DNA minor groove. Previous studies have shown that PBD dimers with odd numbers of methylenes (e.g., n = 3 or 5) in the -O-(CH2)n-O- linker connecting the two PBD units through their C8/C8′-positions are the most potent interstrand cross-linking agents. We have developed a reversed-phase ion pair HPLC/MS method as a tool to evaluate the interaction of DNA-binding molecules with oligonucleotides of varying length and sequence. Based on this methodology, we recently reported that SJG-136 (n = 3) can form three different types of adducts: inter- and intrastrand cross-linked duplex adducts, and mono-alkylated adducts) (J. Am. Chem. Soc., 2009, 131, 13756-13766). Extending these studies to PBD dimers with longer central linkers (e.g., ELB-21 [n = 5] and DRG-16 [n = 5]), we have found that the type and distribution of the various adducts appear to depend on (1) the positioning of the two reactive guanine bases on the same or opposite strands, (2) their separation (typically the number of base pairs, usually ATs, between them) and (3) the length of the C8/C8′-linker connecting the two PBD units in the dimers. Based on these data, a set of rules have emerged that can be used to predict the binding behaviour of a particular PBD dimer towards a given DNA sequence. For example, for PBD dimers containing an -O-CH2CH2CH2-O- linker (i.e., n = 3) between the C8/C8′-positions the following rules apply: Rule 1 - For binding sequences with two guanines separated by two A/T base pairs (ideal sequence length for “n = 3” dimers according to previous modeling studies), there is a preference for interstrand rather than intrastrand cross-link formation; Rule 2 - For shorter binding sequences (e.g., two guanines separated by a single A/T base pair), interstrand cross-linking is preferred over intrastrand, although the rate of reaction is significantly slower; Rule 3 - For longer sequences (e.g., two guanines separated by three A/T base pairs), intrastrand cross-link formation is preferred and the rate of formation is faster. Similar rules have been found to apply to “n = 5” PBD dimers (e.g., ELB-21 and DRG-16) in relation to sequences with two guanines separated by 2, 3 or 4 base pairs. These results, and the simple rules that appear to explain them, suggest that it may be possible to design PBD dimers to target specific DNA sequences. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3506.