Hyaluronic acid lipid-conjugate coated liposomes for targeted delivery of drugs to triple-negative breast cancer : in vitro evaluation on cellular models
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Date
2025-01-08
Authors
Hajimolaali, Mohammad
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Abstract
Triple-negative breast cancer (TNBC) is one of the most aggressive and deadly types of breast cancer. There are limited treatment options for TNBC; conventional treatments include surgery, radiotherapy, and chemotherapy. A promising approach to increase efficacy and reduce side effects involves using targeted drug delivery systems that selectively target tumor cells.
In this study, targeted liposomes containing hyaluronic acid (HA) loaded with doxorubicin (DOX) were developed. The purpose of this study is to investigate the stability of targeted liposomes, to evaluate the reduction of cytotoxicity compared to non-targeted liposomes, and to investigate the cellular uptake of three different breast cancer cell lines, MDA-MB-231, MCF-7and 4T1.
Liposomes were prepared through thin film hydration. Hyaluronic acid was bound to the liposome surface by a strong covalent bond. The stability of liposomes in PBS and FBS buffers was evaluated and the percentage of calcein retention was reported. Acceptable encapsulation efficiency of 97.77 ± 1.36%, particle size of 151.33 ± 0.96 nm, zeta potential of -8.75 ± 0.30 mV, PDI 0.229, and the percentage of release of DOX 25.77 ± 0.03%, in 48 hours were measured.
In the cytotoxicity study on three triple-negative breast cancer cell lines, MDA-MB-231, MCF-7 and 4T1, the targeted and non-targeted liposomes were evaluated and the percentage of cell viability and cytotoxicity were measured. The cellular uptake percentages of the targeted liposomes-labeled with the fluorescent dyes FITC and Rhodamine were also recorded and found to be significantly higher in all cell lines studied, compared to non-targeted liposomes.
The results of the cellular uptake studies, conducted using two methods—flow cytometry and fluorescence spectrophotometry with the fluorescent dyes Rhodamine and FITC—revealed that the flow cytometry device exhibited higher sensitivity and recorded a greater percentage of cellular uptake. According to the findings, the MDA-MB-231 and 4T1 cancer cells demonstrated a higher sensitivity to fluorescent light. The sensitivity of Rhodamine compared to FITC in flow cytometry for the MDA-MB-231, 4T1 and MCF-7 cells was reported as 20.1±2.9%, 31.5±3.2% and 2.3±0.90% respectively, for the formulation containing 0.02 mol hyaluronic acid. Additionally, the cellular uptake percentage for the MCF-7 cells at the same concentration of HA was reported to be 2.3±0.90%.
By comparing the cellular uptake percentages from both devices, it can be concluded that the 0.02 mol concentration of hyaluronic acid in the liposomal formulation with the Rhodamine fluorescent dye shows higher uptake compared to other formulations.
The results of cell uptake studies showed that the presence of HA on the surface of liposome increases their antitumor activity, especially towards MDA-MB-231 cells that overexpress the CD44 receptor.
The results showed that HA-coated liposomes exhibit enhanced antitumor activity of encapsulated DOX, especially in cancer cells overexpressing the CD44 receptor. In addition, future studies are expected to investigate CD44 overexpression and the high affinity of HA for these receptors.
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Keywords
Targeted delivery, Liposomes, Triple-negative breast cancer, Hyaluronic acid