Evaluation of Two Models for Human Topoisomerase I Interaction with dsDNA and Camptothecin Derivatives

Human topoisomerase I (Top1) relaxes supercoiled DNA during cell division. Camptothecin stabilizes Top1/dsDNA covalent complexes which ultimately results in cell death, and this makes Top1 an anti-cancer target. There are two current models for how camptothecin and derivatives bind to Top1/dsDNA covalent complexes (Staker, et al., 2002, Proc Natl Acad Sci USA 99: 15387–15392; and Laco, et al., 2004, Bioorg Med Chem 12: 5225–5235). The interaction energies between bound camptothecin, and derivatives, and Top1/dsDNA in the two models were calculated. The published structure-activity-relationships for camptothecin and derivatives correlated with the interaction energies for camptothecin and derivatives in the Laco et al. model, however, this was not the case for several camptothecin derivatives in the Stacker et al. model. By defining the binding orientation of camptothecin and derivatives in the Top1/dsDNA active-site these results allow for the rational design of potentially more efficacious camptothecin derivatives

Relative interaction energy in kcal/mol between CPT/hCPT-derivatives and the Top1/dsDNA active-site.

<p>The interaction energy scores for the derivatives of CPT and hCPT were subtracted from the score of the respective parent inhibitor with the resulting difference values plotted on the graph. The CPT and hCPT interaction energy scores were set to zero. A negative kcal/mol score indicates that a derivative bound tighter than the parent inhibitor, while a positive kcal/mol score indicates that it bound weaker than the parent inhibitor. A) Interaction energy values for CPT, hCPT and derivatives when bound in the Rotated +1 Nucleoside model Top1/dsDNA active-site. B) Interaction energy values for CPT, hCPT and derivatives bound in the Intercalated model Top1/dsDNA active-site.</p

Relative <i>in vitro</i> inhibition of Top1 by CPT, hCPT, and derivatives.

<p>Modified A-ring ring positions (10, 11) and E-ring position (20) are indicated for CPT/hCPT derivatives; modifications shown in italics (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone-0024314-g001" target="_blank">Fig. 1</a> for CPT and hCPT structures).</p><p>*As reported by Laco et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone.0024314-Laco1" target="_blank">[15]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone.0024314-Laco2" target="_blank">[16]</a>.</p><p>**As reported by Wang et al. for racemic CPT derivatives <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone.0024314-Wang1" target="_blank">[20]</a>. All Top1 inhibition assays were performed using an end-labeled dsDNA oligonucleotide.</p

Rotated +1 Nucleoside model for Top1 interaction with dsDNA and 10-OH CPT.

<p>A) Rotated +1 Nucleoside model, Top1 shown as blue ribbon, dsDNA in teal with the rotated +1 deoxyguanosine left of 10-OH CPT (CPK rendering; carbon, green; oxygen, red; nitrogen, blue; hydrogen, white). B) Rotated +1 Nucleoside model, close up of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone-0024314-g002" target="_blank">Fig. 2A</a> active-site. 10-OH CPT with E-ring in foreground, bound in the Top1/dsDNA active-site in which the +1 scissile strand G is rotated out of the helix to the left until trapped in a network of H-bonds/electrostatic interactions with Asp533 (center) and Arg488/Arg590 (not shown for clarity, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone-0024314-g004" target="_blank">Fig. 4A</a>). Selected atoms involved in H-bonds/electrostatic interactions are colored: nitrogen, blue; oxygen, red; hydrogen, white. Top1 side-chain carbons are yellow, except for Tyr723 (red) in tyrosyl-phosphate bond (phosphorus, orange) to the -1 scissile strand T. 10-OH CPT interactions: 10-OH CPT D-ring stacks over the -1 scissile strand T; 20-OH H-bonds to -1 scissile strand T carbonyl oxygen; A-ring 10-OH oxygen makes electrostatic interaction with Asn352 nitrogen (3.6 Å); E-ring carbonyl oxygen H-bonds to Lys532 nitrogen; C-ring carbonyl oxygen H-bonds to Asn722 nitrogen. Scissile strand rotated +1 G 5′OH H-bonds to Asp533. Arg364 nitrogens H-bond to +1 non-scissile strand C carbonyl oxygen and -1 non-scissile strand A nitrogen. Scissile strand, ss; non-scissile strand, ns. For flat image see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone-0024314-g004" target="_blank">Fig. 4A</a>.</p

Structure of CPT and hCPT E-ring.

<p>CPT left, hCPT E-ring right; hCPT and derivatives differ from CPT in that they contain an additional E-ring carbon between the 20-OH and the adjacent carbonyl oxygen to give a seven-member E-ring.</p

Flattened images of two models for Top1 interaction with dsDNA and 10-OH CPT.

<p>A) Flat image of Rotated +1 Nucleoside model from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone-0024314-g002" target="_blank">Fig. 2</a>, H-bonds and electrostatic interactions within 3.6 Å are indicated with dashed lines. B) Flat image of Intercalated model from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone-0024314-g003" target="_blank">Fig. 3</a>, H-bonds and electrostatic interactions within 3.6 Å are indicated with dashed lines.</p

Intercalated model for Top1 interaction with dsDNA and 10-OH CPT.

<p>A) Intercalated model; Top1 shown as blue ribbon, dsDNA in teal, 10-OH CPT center (CPK rendering; carbon, bronze; oxygen, red; nitrogen, blue; hydrogen, white). B) Intercalated model, close up of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone-0024314-g003" target="_blank">Fig. 3A</a> active-site. 10-OH CPT (carbons bronze) with E-ring in foreground intercalated between the -1 and +1 base pairs in the Top1/dsDNA active-site. 10-OH CPT interactions: 10-OH CPT stacks in between the +1 and -1 base pairs; A-ring 10-OH H-bonds to Glu356 oxygen; E-ring 20-OH H-bonds to Asp533 oxygen; D-ring carbonyl oxygen makes an electrostatic interaction with Asn722 (4.1 Å). Arg364 H-bonds with the -1 non-scissile strand T, Lys 532 H-bonds to the -1 scissile strand T carbonyl oxygen. Top1 active-site Tyr723 (red, left) is shown making a tyrosyl-phosphate bond to the -1 scissile strand T. Scissile strand, ss; non-scissile strand, ns. For flat image see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0024314#pone-0024314-g004" target="_blank">Fig. 4B</a>.</p