Background/Objectives: This study aimed to further enhance the properties of paclitaxel (PTX) and honokiol (HNK) through encapsulation in planetary ball-milled nanoparticles (PBM NPs) and specific targeting of breast cancer (BrCa) cells via MUC1 targeting using an aptamer (S2.2) coating. Methods: Tissue microarray (TMA) analysis was utilized to measure MUC1 expression in stages 1, 2, 3, and 4 BrCa tissue samples. Pharmacokinetic simulations were performed to explore the potential advantages of using PTX and HNK in combination while targeting MUC1 for BrCa treatment. To investigate the efficacy of the PBM NPs for MUC1 targeting, we synthesized the aptamer-conjugated PTX and HNK PBM NPs (PTX-S2.2-PBM NPs, HNK-S2.2-PBM NPs) using N-hydroxysuccinimide (NHS) coupling. Dynamic light scattering (DLS), Fourier-transform infrared (FTIR), and high-performance liquid chromatography (HPLC) were utilized to characterize the NPs. MTT and live/dead cell assays were used to evaluate the cytotoxicity of the NPs. Results: TMA sample analysis confirmed the upregulation of MUC1 in BrCa tissues, which increased with the stage of BrCa. DLS analysis revealed that the PTX-S2.2 and HNK-S2.2 NPs have a desirable size (83.4 nm and 163 nm, respectively) and zeta potential (−9.74 mV and −7.16 mV, respectively), which are suitable for systemic circulation and improved therapeutic outcomes. FTIR and HPLC analysis suggest proper conjugation was achieved, and an encapsulation efficiency of PTXS2.2 and HNKS2.2 NPs at 77% and 84%, respectively, was achieved. Cell viability assays demonstrated that PTX-S2.2-PBM and HNK-S2.2-PBM NPs exhibit cytotoxicity comparable to or greater than free PTX and HNK, respectively. Conclusions: These findings support the belief that using PTX-S2.2 and HNK-S2.2 PBM NPs could be a promising treatment option for BrCa.
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