Multi-responsive chitosan-based hydrogels for controlled release of vincristine

Cancer is a leading cause of death globally. Chemotherapy, while effective, faces challenges including poor drug penetration into tumor cells, toxicity to healthy cells, and drug instability. Vincristine (VCR) is a highly effective anticancer drug, but its use is limited due to neurotoxicity and other side effects. Controlled release formulations are desirable to mitigate these issues and improve therapeutic efficacy. Various drug delivery platforms, such as liposomes, nanoparticles, and microspheres, have been explored to enhance VCR delivery. Hydrogels, known for their biodegradability, biocompatibility, and responsiveness, have emerged as promising biomaterials for drug delivery. Chitosan (CS) and its derivative carboxymethyl chitosan (CMCS) are biocompatible, non-toxic, and capable of forming gels, making them suitable candidates for hydrogel-based drug delivery systems. This study aims to synthesize and characterize novel chitosan-based hydrogels (CS-g-gly and CMCS-g-gly) for controlled release of vincristine, investigating their in vitro release profiles and cytotoxicity.

Numerous studies have explored alternative VCR formulations to enhance efficacy and reduce side effects. Liu et al. developed VCR-Au conjugated nanoparticles entrapped in liposomes, showing improved antitumor efficiency in mice. Aboutaleb and Dinarvand used VCR-dextran-loaded solid lipid nanoparticles for brain drug delivery, achieving increased plasma and brain drug levels. Salar et al. synthesized vincristine-loaded folic acid-chitosan nanoparticles, demonstrating sustained drug release. Chen et al. created VCR-loaded poly(lactide-co-glycolide) microspheres in a collagen and chitosan swelling solution, achieving sustained release. These studies highlight the ongoing efforts to improve VCR delivery, with hydrogels gaining prominence due to their advantageous properties.

Carboxymethyl chitosan (CMCS) was synthesized by carboxymethylation of chitosan. CS-g-gly and CMCS-g-gly hydrogels were prepared by dissolving chitosan or CMCS in acetic acid, activating with formaldehyde, and crosslinking with glycerol. Ammonia solution was added to induce gelation. The hydrogels were characterized using FTIR, 1H NMR, FESEM, AFM, TGA, DLS, and zeta potential measurements. Vincristine was loaded into the hydrogels, and encapsulation efficiency was determined. In vitro drug release studies were conducted in PBS at pH 5 and 7.4. Cell viability and apoptosis induction were assessed in MCF-7 (breast cancer) and MCF-10 (normal breast) cells using MTT assay and flow cytometry.

FTIR and 1H NMR confirmed the successful synthesis of CS-g-gly and CMCS-g-gly hydrogels. FESEM showed spherical hydrogel structures with diameters increasing after vincristine loading. DLS analysis revealed average particle sizes of approximately 84.5 nm (CS-g-gly) and 87.6 nm (CMCS-g-gly), increasing after drug loading. Zeta potential measurements indicated positive surface charges, enhancing stability and cellular uptake. TGA analysis showed thermal stability of the hydrogels. Both hydrogels displayed high swelling ratios, particularly in acidic pH (pH 5), simulating the tumor microenvironment. Vincristine encapsulation efficiency was high, ranging from 72.28% to 89.97% for CS-g-gly and 56.97% to 71.91% for CMCS-g-gly. VCR/CS-g-gly showed sustained release, reaching 82% release after 120 hours at pH 5. The MTT assay demonstrated that drug-free hydrogels exhibited no significant cytotoxicity. Vincristine-loaded hydrogels significantly inhibited MCF-7 cell viability in a concentration-dependent manner, with IC50 values of 22.49 and 24.43 for VCR/CS-g-gly and VCR/CMCS-g-gly, respectively. Flow cytometry confirmed that vincristine-loaded hydrogels induced apoptosis in MCF-7 cells. The hydrogels showed minimal cytotoxicity against MCF-10 normal cells.

The results demonstrate the successful development of pH-sensitive, biodegradable, and biocompatible chitosan-based hydrogels for controlled vincristine release. The sustained release profile at pH 5, mimicking the tumor microenvironment, suggests potential for enhanced therapeutic efficacy by maximizing drug concentration at the tumor site while minimizing systemic toxicity. The minimal cytotoxicity of drug-free hydrogels underscores their biocompatibility. The significant anti-cancer activity against MCF-7 cells, coupled with the minimal impact on normal cells (MCF-10), highlights the potential selectivity of this drug delivery system. Further optimization may be explored to fine-tune the release kinetics and enhance tumor targeting.

This study successfully synthesized and characterized novel chitosan-based hydrogels for controlled vincristine release. The hydrogels exhibited sustained release properties, particularly at acidic pH, and showed minimal cytotoxicity. These findings suggest potential for improved cancer therapy by reducing side effects and enhancing drug efficacy. Future work could focus on in vivo studies to evaluate the therapeutic potential in animal models and explore further modifications for improved targeted delivery.

The study was limited to in vitro experiments. In vivo studies are necessary to confirm the efficacy and safety of the hydrogels. The long-term stability of the hydrogels in physiological conditions should also be investigated. Further optimization of the hydrogel formulation may be needed to achieve even better control over drug release kinetics and further improve tumor targeting.