Small molecule compounds which interact with the tubulin-microtubule system are recognized as one of the major clinical options for cancer treatment such taxol and vincristine. The stilbene Combretastatin A-4 (CA-4) demonstrates potent anticancer activity in human cancer cells together with inhibition of tubulin polymerisation and antivascular effects. We report the synthesis of a panel of nineteen conformationally restricted 1,4-diaryl-2-azetidinones containing the 3,5-dimethoxyphenyl substituent at N-1 replacing the characteristic 3,4,5-trimethoxyphenyl Ring A of CA-4, together with phenyl, chloro, hydroxyl, phenoxy substituents at C-3. X-Ray crystallographic studies determined the specific structural features of selected compounds together with the role of the novel 3,5-dimethoxyphenyl Ring A interactions with the colchicine tubulin binding site residues.

The effect of these compounds on cancer cell proliferation was determined in vitro with the 3-hydroxyl, 3-phenoxy and 3-unsubstituted compounds identified as the most potent examples, having IC50 values of 3-25 nM in MCF-7 human breast cancer and 8-25 nM in chemoresistant HT-29 colon cancer cells which compare favourably with control CA-4. Low levels of in vitro cytotoxicity was confirmed for the novel β-lactams in MCF-7 cells using the lactate dehydrogenase (LDH) assay (< 3.5% cell death observed). The interactions of the methoxy groups at C-3 and C-5 of Ring A with key hydrophobic tubulin binding site residues Valβ318, Cysβ241, Alaβ354, Alaβ316, Alaβ317, Leuβ242, Valβ238 and Ileβ378 are explored to provide binding stabilisation. The structural study of these compounds will facilitate further design of more effective and diverse β-lactams for potential development in breast and chemoresistant colon cancer applications.