Thiolated chitosan nanoparticles as a delivery system for antisense therapy: evaluation against EGFR in T47D breast cancer cells

Abstract

Thiolated chitosan has high transfection and mucoadhesive properties. We investigated the potential of two recently synthesized polymers: NAC-C (N-acetyl cysteine-chitosan) and NAP-C (N-acetyl penicillamine-chitosan) in anticancer drug delivery targeting epidermal growth factor receptor (EGFR). Doxorubicin (DOX) and antisense oligonucleotide (ASOND)-loaded polymer nanoparticles were prepared in water by a gelation process. Particle characterization, drug loading, and drug release were evaluated. To verify drug delivery efficiency in vitro experiments on a breast cancer cell line (T47D) were performed. EGFR gene and protein expression was analyzed by real time quantitative polymerase chain reaction and Western blotting, respectively. A loading percentage of 63% ± 5% for ASOND and 70% ± 5% for DOX was achieved. Drug release data after 15 hours showed that ASOND and DOX were completely released from chitosan-based particles while a lower and more sustained release of only 22% ± 8% was measured for thiolated particles. In a cytosol simulated release medium/reducing environment, such as found intracellularly, polymer-based nanoparticles dissociated, liberating approximately 50% of both active substances within 7 hours. ASOND-loaded polymer nanoparticles had higher stability and high mucoadhesive properties. The ASOND-loaded thiolated particles significantly suppressed EGFR gene expression in T47D cells compared with ASOND-loaded chitosan particles and downregulated EGFR protein expression in cells. This study could facilitate future investigations into the functionality of NAP-C and NAC-C polymers as an efficient ASOND delivery system in vitro and in vivo.

Thiolated-Polymer-Based Nanoparticles as an Avant-Garde Approach for Anticancer Therapies—Reviewing Thiomers from Chitosan and Hyaluronic Acid

“…Thus, CTS-based NPs were formulated via gelation with sodium sulfate as nanocarriers for doxorubicin (DOX) and antisense oligonucleotide (ASOND). They displayed a spherical, uniform shape with a mean particle size of 150-300 nm [114]. Another option for CTS-based particles is ionic gelation with TPP, feasible for both CTS and thiolated CTS-GSH [29].…”

“…Results showed that the corneal status significantly improved in more than 60% of patients, with most of them starting to show beneficial results as early as 10 min after a single instillation of CTS-NAC [162]. This polymer has also been tested, along with CTS-NAP (CTS-N-acetylpenicillamine), as a platform to deliver doxorubicin (DOX) and antisense oligonucleotides (ASOND) targeting EGFR, which is a transmembrane receptor overexpressed in most of the cases of breast cancer with poor prognosis [114]. When evaluated on T47D cells (human breast epithelial tumor cells), these NPs showed similar properties as the ones stated before, such as sustained release, good encapsulation efficiency, and stability (especially those carrying ASOND), so the clinical potential of this formulation is obvious.…”

Thiolated Chitosans: A Multi-talented Class of Polymers for Various Applications

“…For the analysis of the amount of immobilized −SH groups of these thiolated chitosans, Ellman’s reagent optionally with previous reduction of disulfide bonds with borohydride or iodometric titration can be used . Phadungcharoen et al, for instance, have conducted the Ellman’s assay via smartphone, providing an easy, fast, and reliable analysis of thiolated polymers without access to sophisticated equipment. …”

Drug Delivery Systems, CNS Protection, and the Blood Brain Barrier

“…Moreover, 5-aminolevulinic acid-incorporated nanoparticles of methoxy poly(ethylene glycol)-chitosan copolymer are used in photodynamic therapy [ 191 ], while, FVIII-chitosan nanoparticles [ 192 ], cyclosporin A-loaded, PEGylated chitosan-modified, lipid-based nanoparticles [ 193 , 194 ], and chitosan and poly(lactic- co -glycolic acid) incorporated nanoparticles (heparin) are also prepared for quick CNS therapeutics [ 195 ]. Similarly, thiolated chitosan nanoparticles are also prepared for drug delivery system for antisense therapy [ 196 ] ( Table 2 ).…”