Exploring the impact of trifluoromethyl (-CF 3 ) functional group on the anti-cancer activity of isoxazole-based molecules: design, synthesis, biological evaluation and molecular docking analysis.

Herein we report the design and synthesis of a series of fully-substituted 4-(trifluoromethyl)isoxazoles and evaluation of their anti-cancer activities against MCF-7, 4T1 and PC-3 cell lines as a proof of concept study. 4-(Trifluoromethyl)isoxazole is a synthetically challenging class of molecules and very few synthetic methods have been developed so far and all of them suffered from several serious limitations. Recently we developed a novel, metal-free, and general synthetic strategy to access synthetically challenging 4-(trifluoromethyl)isoxazoles starting from readily available chalcones using cheap CF 3 SO 2 Na as the source of the -CF 3 group and multitasking t BuONO as an oxidant as well as the source of N and O and thus we have overcome the limitations of the previous methods. Based on the structure of an isoxazole-based anti-cancer agent, 3-(3,4-dimethoxyphenyl)-5-(thiophen-2-yl)isoxazole 14, we designed a set of 4-(trifluoromethyl)isoxazoles for synthesis and further anti-cancer evaluation. Among various molecules, 3-(3,4-dimethoxyphenyl)-5-(thiophen-2-yl)-4-(trifluoromethyl)isoxazole 2g (IC 50 = 2.63 μM) and 3-(thiophen-2-yl)-5-(4-(thiophen-2-yl)-1 H -pyrrol-3-yl)-4-(trifluoromethyl)isoxazole 5 (IC 50 = 3.09 μM) exhibited the best anti-cancer activity against the human breast cancer cell-lines (MCF-7), 2g being the lead molecule among all. Interestingly, 2g is found to be almost 8 times more active compared to its non-trifluoromethylated analogue, i.e. , 3-(3,4-dimethoxyphenyl)-5-(thiophen-2-yl)isoxazole 14 (IC 50 = 19.72 μM) which revealed the importance of a 'CF 3 ' moiety in enhancing the anti-cancer activity of 14. Further studies such as apoptosis induction, cell cycle analysis, and nuclear staining revealed an apoptotic cell death mechanism. The in silico molecular docking, induced fit analysis, and ADME studies further supported the effect of a -CF 3 moiety on the enhancement of anti-cancer activity of isoxazole-based anti-cancer molecules. Further exploration of the biodistribution and therapeutic efficacy of lead 2g in vivo holds significant promise, positioning it as a potential candidate for anticancer therapy.