Structural simplification of bioactive natural products with multicomponent synthesis. 3. Fused uracil-containing heterocycles as novel topoisomerase-targeting agents

Abstract

After the initial discovery of antiproliferative and apoptosis-inducing properties of a camptothecin-inspired pentacycle based on a 1H-indeno[2',1':5,6]dihydropyrido[2,3-d]pyrimidine scaffold, a library of its analogues as well as their oxidized planar counterparts were prepared utilizing a practical multicomponent synthetic protocol. The synthesized compounds exhibited submicromolar to low micromolar antiproliferative potencies toward a panel of human cancer cell lines. Biochemical experiments are consistent with the dihydropyridine library members undergoing intracellular oxidation to the corresponding planar pyridines, which then inhibit topoisomerase II activity, leading to inhibition of proliferation and cell death. Because of facile synthetic preparation and promising antitopoisomerase activity, both the dihydropyridine and planar pyridine-based compounds represent a convenient starting point for anticancer drug discovery.

Figure 1

Structures of camptothecin (1), topotecan (2), silatecan (3), luotonin A (4), batracylin (5) and indenopyridine 6.

Figure 2

(a) MCR used for the preparation of hetero-fused indenopyridines 7 and (b) preparation of the uracil-fused pentacycle 8h in the dihydropyridine form.

Figure 3

Preparation of the dihydro- and indenopyridine libraries.

Figure 4

Mean IC₅₀ values for camptothecin (1) and the best analogues from the MCR library determined on 4 distinct highly sensitive (HS) versus 4 distinct weakly sensitive (WS) human cancer cell lines to camptothecin. HS cancer cell lines display mean IC₅₀ values for camptothecin < 0.1 µM (Jurkat, LoVo, U373 and SKMEL), while WS cancer cell lines display mean IC₅₀ values for camptothecin > 0.1 µM (HeLa, MCF-7, A-549 and PC-3).

Figure 5

Quantitative videomicroscopy analyses carried out on human U373 glioblastoma cells revealed that camptothecin (1) manifests cytostatic features, while compounds 13h, 13o, 18h and 18o also display cytotoxic effects in addition to the cytostatic ones. The cytotoxic effects relate to refringent and rounded cells that correspond to dying or dead cells (see the white arrows). Each compound was assayed at its GI₅₀ concentration as revealed by the MTT colorimetric assay on U373 glioblastoma cells (Table 1), i.e. 0.03 µM for 1 (camptothecin), 0.4 µM for 13h, 0.5 µM for 13o, 0.1 µM for 18h and 0.1 µM for 18o.

Figure 6

Effects of “h” and “o” pentacycles on relaxation of supercoiled pGEM1 plasmid DNA by topoisomerase I. Camptothecin was used as reference at 100 µM concentration. DNA samples were separated by gel electrophoresis.

Figure 7

Effects of “h” and “o” pentacycles on decatination of kinetoplast DNA (KDNA) by topoisomerase II. Doxorubicin (DOX) was used as reference at 5 µM concentration. DNA samples were separated by gel electrophoresis.