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Abstract Firstly, the study investigates the feasibility of hexosomes (HEX) as a competent platform for fluoxetine hydrochloride (FH) repurposing against HepG2 hepatocellular carcinoma. Different FH-loaded HEX formulations were prepared and optimized by the hot emulsification method. The HEX features as particle size, zeta potential, and drug entrapment efficiency (EE %) can be tailored by tuning HEX components and fabrication conditions. The composition of the optimized FH hexosomes (OFH-HEX) was 3.1, 1.4, 0.5, 0.2, and 94.8% for glyceryl monooleate, oleic acid, pluronic F127, FH, and deionized water respectively. The anionic OFH-HEXwith a particle size of 145.5±2.5 nm and drug EE % of 45.4±1.2% was able to prolong in vitro FH release where only 19.5±2.3% released in PBS pH7.4 after 24 h. Contrarily, HEX rapidly released FH in acetate buffer pH5.5 and achieved a 90.5±4.7% release after 24 h which facilitates the passive targeting of FH to the cancer cells. Moreover, the obtained HEX showed an improved cellular internalization in a time-dependent manner and enhanced cytotoxicity (2-fold higher than FH solution). Consequently, this chapter suggests the potential of FH-HEX as a possible anticancer agent against hepatocellular carcinoma |