Anti-CD44 and EGFR Dual-Targeted Solid Lipid Nanoparticles for Delivery of Doxorubicin to Triple-Negative Breast Cancer Cell Line: Preparation, Statistical Optimization, and In Vitro Characterization

Abstract

Background. Despite being more aggressive than other types of breast cancer, there is no suitable treatment for triple-negative breast cancer (TNBC). Here, we designed doxorubicin-containing solid lipid nanoparticles (SLNs) decorated with anti-EGFR/CD44 dual-RNA aptamers, which are overexpressed in TNBC. For more efficiency in the nuclear delivery of doxorubicin, dexamethasone (Dexa) was chemically attached to the surface of nanoparticles. Methods. To prepare the cationic SLNs, 6-lauroxyhexyl BOC-ornithine (LHON) was synthesized and was chemically attached to dexamethasone to form Dexa-LHON complexes. The doxorubicin-containing SLNs were prepared via double emulsification ( $w/o/w$ ) and the solvent evaporation technique. The preparation of SLNs was statistically optimized using the central composite response surface methodology. Independent factors were the GMS/lecithin concentration ratio and the amount of Tween 80, while responses considered were particle size, polydispersity index, and entrapment efficiency of the nanoparticles. The optimized nanoparticles were studied morphologically using transmission electron microscopy, and in vitro release of doxorubicin from nanoparticles was studied in phosphate-buffered saline. Then, the designated aptamers were attached to the surface of nanoparticles using electrostatic interactions, and their cytotoxicity was assessed in vitro. Results. The size, PDI, zeta potential, EE%, and LE% of the prepared nanoparticles were $101\pm 12.6\text{nm}$ , $0.341\pm 0.005$ , $+13.6\pm 1.83\text{mV}$ , $69.98\pm 7.54\%$ , and $10.2\pm 1.06\%$ , respectively. TEM images revealed spherical nanoparticles with no sign of aggregation. In vitro release study exhibited that $96.1\pm 1.97\%$ of doxorubicin was released within 48 h of incubation. The electrostatic attachment of the designated aptamers to the nanoparticles’ surface was confirmed by reducing the zeta potential to $-15.6\pm 2.07\text{mV}$ . The in vitro experiments revealed that the SLNs/DOX/Dexa/CD44 or EGFR aptamers were substantially more successful than SLNs/DOX/Dexa at inhibiting cell proliferation. Using the MDA-MB-468 cell line, we discovered that SLN/DOX/Dexa/CD44/EGFR aptamers were more effective than other constructs in inhibiting cell proliferation ( $p<0.001$ ). The reduction of cell viability using this construct suggests that targeting numerous proliferation pathways is effective. Conclusion. Overall, the finding of this investigation suggested that SLNs/DOX/Dexa/CD44/EGFR could be a promising new enhanced anticancer delivery system and deserved further preclinical consideration.