A new biological scaffold was produced by replacing the 6π-electron phenyl ring-B of a natural flavone skeleton with a 10π-electron benzothiophene (BT). Since aromatic rings are important for ligand protein interactions, this expansion of the π-electron system of ring-B might change the bioactivity profile. One of the resulting novel natural product-inspired compounds, 2-(benzo[b]thiophen-3-yl)-5-hydroxy-7-isopropoxy-6-methoxyflavone (6), effectively arrested the cell cycle at the G2/M phase and displayed significant antiproliferative effects with IC<sub>50</sub> values of 0.05-0.08 μM against multiple human tumor cell lines, including a multidrug resistant line. A structure-activity relationship study revealed that a 10π-electron system with high aromaticity, juxtaposed 4-oxo and 5-hydroxy groups, and 7-alkoxy groups were important for potent antimitotic activity. Interestingly, two BT-flavonols (3-hydroxyflavone), 16 and 20, with 3-hydroxy and 5-alkoxy groups, induced distinct biological profiles affecting the cell cycle at the G1/S phase by inhibition of DNA replication through an interaction with topoisomerase I.