Discovery of Novel Azetidine Amides as Potent Small-Molecule STAT3 Inhibitors

We optimized our previously reported proline-based STAT3 inhibitors into an exciting new series of (R)-azetidine-2-carboxamide analogues that have sub-micromolar potencies. (5a) under bar, (5o) under bar, and (8i) under bar have STAT3-inhibitory potencies (IC50) of 0.55, 0.38, and 0.34 mu M, respectively, compared to potencies greater than 18 mu M against STAT1 or STATS activity. Further modifications derived analogues, including 7 (e) under bar, (7f) under bar, (7g) under bar, and (9k) under bar, that addressed cell membrane permeability and other physicochemical issues. Isothermal titration calorimetry analysis confirmed high-affinity binding to STAT3, with K-D of 880 nM ((7g) under bar) and 960 nM ((9k) under bar). (7g) under bar and (9k) under bar inhibited constitutive STAT3 phosphorylation and DNA-binding activity in human breast cancer, MDA-MB-231 or MDA-MB-468 cells. Furthermore, treatment of breast cancer cells with (7e) under bar, 7 (f) under bar , (7g) under bar, or (9k) under bar inhibited viable cells, with an EC50 of 0.9-1.9 mu M, cell growth, and colony survival, and induced apoptosis while having relatively weaker effects on normal breast epithelial, MCF-10A or breast cancer, MCF-7 cells that do not harbor constitutively active STAT3.