Laurence Hurley

PMID: 3078076;Abstract:

DNA is the presumed target for a number of clinically useful anticancer drugs. In this review, Laurence Hurley and Leslie Boyd discuss the appropriateness of the term 'receptor' for DNA and outline the forseeable problems in designing drugs that will produce a defined pharmacological response through interaction with DNA. They describe the structural features which present DNA as an attractive target for drug design, and the possible characteristics of drugs that react with DNA to produce a predetermined biochemical response. Finally, they outline modern approaches to elucidating the structural and biological consequences of drug modification. © 1998.

PMID: 10924124;Abstract:

N,N'-Bis[2-(1-piperidino)ethyl]-3,4,9,10-perylenetetracarboxylic diimide (PIPER), a perylene derivative, is a very potent and selective G-quadruplex DNA-interactive agent. It has been shown to inhibit DNA polymerase and telomerase by stacking externally to the G-tetrads in the G-quadruplex structures. Recently, we have demonstrated that this small molecule greatly accelerates the assembly of G-quadruplex structures in a cell-free system. In this report, we present data demonstrating that PIPER prevents the unwinding of G-quadruplex structures by yeast Sgsl helicase. Sgsl belongs to the RecQ DNA helicase family whose members include other G-quadruplex DNA unwinding helicases, such as human Bloom's syndrome and human Werner's syndrome helicases. PIPER specifically prevents the unwinding of G-quadruplex DNA but not duplex DNA by Sgsl. Competition experiments indicate that this inhibitory activity is due to the interaction of PIPER with G-quadruplex structures rather than the helicase itself. These results combined with previous studies suggest a possible mechanism of action for these G-quadruplex-interactive agents inside cells: they might induce G-quadruplex formation in G-rich regions on genomic DNA, stabilize these structures, and prevent them from being cleared by enzymes such as helicases. The G-quadruplex structures may, in turn, disrupt some critical cellular events such as DNA replication, transcription regulation, and telomere maintenance.

Abstract:

Gel electrophoresis analysis of CPI-I bisalkylation of a 21-mer duplex containing 5′-TAA²TTA¹-3′ (the palindromic preferred cross-linking sequence of the (+)-CC-1065 analog Bizelesin) shows same-strand (strand one) alkylation of first A¹ and then A² instead of the anticipated symmetrical A¹ alkylation of strands one and two. Two-dimensional NMR analyses (NOESY and COSY) confirm the head-to-tail minor groove orientation of the same-strand-bound drugs. CPI-I contrasts sharply with Bizelesin (two CPI-I units linked tail-to-tail by a ureadiyl "linker"), which symmetrically cross-links this sequence at A¹ (strands one and two), but only by first rearranging the duplex structure and consequently removing the duplex distortion stemming from monoadduct formation. CPI-I induces no such major DNA rearrangement prior to or during bisalkylation. Why does CPI-I react with the adenines of only a single strand? Two possible causes for the unexpected strand one A² alkylation are, first, retardation of strand two A' site's reactivity by focusing of monoadduct conformational distortion on this site and, second, elevation of A² reactivity above other competing adenine sites due to unusual monoadduct strand one A²T-step conformational properties. The relative importance of these two nonmutually exclusive factors was investigated using gel electrophoresis experiments: Time-course CPI-I bisalkylation studies were conducted on the AT-step sequence 5′-TAA²TTA1-3′ and an A-tract sequence, 5′-TAA²AAA¹-3′, to see if the former sequence's AT-step flexibility, high base-pair opening rate, and unwinding capability (traits not shared by the latter sequence) controlled selection of the second target site. The observed parallel AT-step and A-tract sequence A¹ and A² bisalkylation patterns suggest that AT-step properties play at best a secondary role (compared to 5′-end TA-step behavior) in directing the second alkylation reaction to the AT-step site. rMD (solvated) simulations of the AT-step and A-tract monoadducts display distortion that is focused on this 5′-end TA-step site. While two-dimensional ¹H NMR spectra of the final bisadduct reveal no significant TA-step conformational distortion, they demonstrate that conformational adjustment at the A² ligand attachment site diminishes head-to-tail steric clash of the two drugs. These results suggest that the CPI-I monoadduct propagates bending distortion (to the 5′-side) through five base pairs toward the TA-step junction site. In the AT-step and A-tract sequences, neither adenine straddling this TA-step junction site is alkylated by CPI-I, suggesting that the base pairs forming the junction site are distorted away from a suitable orientation or are unable to assume a conformation suitable for alkylation. Consequently, the second alkylation occurs at a site (AT-step) that requires a modest displacement of the second ligand away from the already attached drug. The results and analysis of the data included in this paper provide important lessons for the design of inter- and intrastrand DNA-DNA cross-linkers.

MYC is deregulated in most tumour types, but an effective means to selectively target its aberrant expression is not yet available. Supercoiling that is induced by transcription has been demonstrated to have dynamic effects on DNA in the MYC promoter element: it converts duplex DNA to non-duplex DNA structures, even at considerable distances from the transcriptional start site. These non-duplex DNA structures, which control both turning on and off of transcription and the rate of transcription firing, are amenable to small-molecule targeting. This dynamic system provides a unique opportunity for the treatment of tumours in which MYC is an important oncogene.