Synthesis and Mechanism of Action Studies of a Series of Norindenoisoquinoline Topoisomerase I Poisons Reveal an Inhibitor with a Flipped Orientation in the Ternary DNA−Enzyme−Inhibitor Complex As Determined by X-ray Crystallographic Analysis

Abstract

Several norindenoisoquinolines substituted with methoxy or methylenedioxy groups have been prepared and their anticancer properties evaluated in cancer cell cultures and in topoisomerase I inhibition assays. 2,3-Dimethoxy-8,9-methylenedioxy-11H-indeno[1,2-c]isoquinoline hydrochloride (14) is a strong topoisomerase I inhibitor and also displays very high cytotoxicity in the NCI cancer cell culture screen (mean graph midpoint of 50 nM). The X-ray crystal structure of norindenoisoquinoline 14 in complex with topoisomerase I and DNA has been solved, providing insight into the structure-activity relationships within this class of new anticancer agents. The number and position of the norindenoisoquinoline substituents have a significant influence on biological activity and demonstrate that substitution on the nitrogen atom is not an absolute requirement for the antitumor effect of the indenoisoquinolines. Removal of the 11-keto group from the lead compound 1 and replacement of the N-alkyllactam with an unsubstituted pyridine ring causes the indenoisoquinoline ring system to flip over in the DNA-enzyme-inhibitor ternary complex. This allows the nitrogen atom to assume the hydrogen bond acceptor role of the 11-keto group, resulting in hydrogen bonding to Arg364.

Repair of Topoisomerase I‐Mediated DNA Damage

“…Misalignments can be generated by drugs bound at the interface of the enzyme and broken DNA [26,27] and by alterations of the DNA substrate (Table 1 and Camptothecins and non-camptothecin Top1 inhibitors trap Top1 cleavage complexes by binding at the enzyme-DNA interface ( Fig. 3D-E) [22,[28][29][30]. Hence Top1 inhibitors represent a paradigm for "interfacial inhibitors" [26,27].…”

“…[29]] showing "interfacial inhibition" [26,27] of the Top1 cleavage complex by camptothecin. Interfacial inhibition also applies to non-camptothecin Top1 inhibitors shown in panel F [29,30]. E. Same structure as in panel D. The Top1 has been removed except for the catalytic tyrosine (in orange).…”

A novel norindenoisoquinoline structure reveals a common interfacial inhibitor paradigm for ternary trapping of the topoisomerase I-DNA covalent complex

“…E, topoisomerase I is represented in ribbon diagram to allow visualization of the catalytic tyrosine (Y; red) and to show the drug intercalation between the À1 and +1 bp. F, chemical structure of the five drugs cocrystallized with topoisomerase I-DNA complexes (15,16,24).…”

“…This study describes the crystal structure of a potent norindenoisoquinoline (24), topoisomerase I inhibitor (AI-III-52), in a complex with DNA and topoisomerase I. Comparison with the corresponding ternary complexes of the MJ-238 indenoisoquinoline (16) and with the structures of camptothecin (16), topotecan (15), and an indolocarbazole (16) reveals a common molecular mechanism of drug action.…”

“…The PDB codes for the different topoisomerase I crystal structures were 1LT8 for norindenoisoquinoline (24), 1SC7 for MJ-238 (16), 1SEU for indolocarbazole (16), 1K4T for topotecan (15), 1T81 for camptothecin (16), and 1A31 for topoisomerase I in the absence of inhibitor (34). The overall structure the AI-III-52 norindenoisoquinoline in a topoisomerase I cleavage complex is presented in Fig.…”