A facile hybrid ‘flow and batch’ access to substituted 3,4-dihydro-2H-benzo[b][1,4]oxazinones

• Andrew J. S. Lin^a
• Cecilia C. Russell^a
• Jennifer R. Baker^a
• Shelby L. Frailey^ab
• Jennette A. Sakoff^c
• Adam McCluskey^*a

• ^* Corresponding authors
• ^a Chemistry, Centre for Chemical Biology, School of Environmental & Life Sciences, University of Newcastle, University Drive, Callaghan, Australia
• ^b Chemical Engineering, Trine University, Angola, 46703 USA
• ^c Department of Medical Oncology, Calvary Mater Newcastle Hospital, Waratah, Australia

We describe a simple flow chemistry approach to libraries of ethyl 3-oxo-2-(substituted-phenylamino)-3,4-dihydro-2H-benzo[b][1,4]oxazine-6-carboxylates (12a–l) and N-ethyl-3-oxo-2-(substituted-phenylamino)-3,4-dihydro-2H-benzo[b][1,4]oxazine-6-carboxamides (13a–l) in 38–87% yields. This scaffold is poorly described in the chemical literature. Screening against a panel of 11 cancer and one normal cell line showed that the amide linked library 13a–l was devoid of toxicity. Whereas the ester linked analogues 12b, 12c, 12g, 12j and 12l were highly cytotoxic with growth inhibition (GI50) values from 0.34 to >50 μM across all cell lines, with the 2-OH-Ph substituted 12l analogue presenting with sub-micromolar potency against the A2780 (ovarian; 0.34 ± 0.04 μM), BEC-2 (glioblastoma; 0.35 ± 0.06 μM), MIA (pancreas; 0.91 ± 0.054 μM) and SMA (murine glioblastoma; 0.77 ± 0.029 μM) carcinoma cell lines. Interestingly, the U87 glioblastoma cell line showed inherent resistance to growth inhibition by all analogues (GI50 32 to >50 μM) while the A2780 cells were highly sensitive (GI50 3.8–0.34 μM), suggesting that the analogues developed herein may be valuable lead compounds for the development of ovarian carcinoma specific cytotoxic agents. The differences in amide versus ester cytotoxicity was consitent with esterase cleaveage to release the cytotoxic warhead.

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