A promising design paradigm for small-molecule inhibitors of tubulin polymerization that bind to the colchicine site draws structural inspiration from the natural products colchicine and combretastatin A-4 (CA4). Our previous studies with benzocycloalkenyl and heteroaromatic ring systems yielded promising inhibitors with dihydronaphthalene and benzosuberene analogues featuring phenolic (KGP03 and KGP18) and aniline (KGP05 and KGP156) congeners emerging as lead agents. These molecules demonstrated dual mechanism of action, functioning both as potent vascular disrupting agents (VDAs) and as highly cytotoxic anticancer agents. A further series of analogues was designed to extend functional group diversity and investigate regioisomeric tolerance. Ten new molecules were effective inhibitors of tubulin polymerization (IC50 < 5 μM) with seven of these exhibiting highly potent activity comparable to CA4, KGP18, and KGP03. For one of the most effective agents, dose-dependent vascular shutdown was demonstrated using dynamic bioluminescence imaging in a human prostate tumor xenograft growing in a rat.