A Nanoparticle-Based Strategy to Stabilize 5-Azacytidine and Preserve DNA Demethylation Activity in Human Cardiac...

Area of Science:

• Pharmacology
• Biotechnology
• Epigenetics

Background:

• 5-Azacytidine (5-Aza) is a DNA methyltransferase (DNMT) inhibitor with potential in cardiac remodeling via epigenetic reprogramming of human cardiac fibroblasts (HCFs).
• The clinical use of 5-Aza is limited by its rapid hydrolytic degradation.
• Nanoparticle (NP) encapsulation is explored to improve the stability of labile drugs like 5-Aza.

Purpose of the Study:

• To evaluate the physical and chemical stability of free 5-Aza and 5-Aza-loaded lipid nanoparticles (5-Aza-NP).
• To assess the impact of NP encapsulation on the stability and biological activity of 5-Aza.
• To examine the effects of stored 5-Aza and 5-Aza-NP on DNA methylation-related gene expression in HCFs.

Main Methods:

• Lipid nanoparticles (NPs) were formulated using a hyaluronic acid-stabilized lipid system via solvent displacement.
• Physical stability (size, PDI, zeta potential) was monitored over 96 hours at -20 °C, 4 °C, and 30 °C.
• Chemical stability was assessed using HPLC and kinetic modeling. Functional activity was determined by measuring DNMT1, DNMT3A, and DNMT3B gene expression in HCFs post-treatment with stored formulations.

Main Results:

• 5-Aza-NP exhibited physical stability at 4 °C, while aggregation occurred at -20 °C and thermal variability at 30 °C.
• Free 5-Aza degraded rapidly at 30 °C (6.56% remaining at 72h), whereas 5-Aza-NP showed significantly improved stability (11.54% remaining).
• 5-Aza-NP maintained its ability to suppress DNMT1 expression after 96h storage at 4 °C, unlike free 5-Aza, while DNMT3A/B levels remained unaffected.

Conclusions:

• Nanoparticle encapsulation significantly enhances the physicochemical stability of 5-Azacytidine.
• The 5-Aza-NP formulation preserves the drug's DNMT1-inhibitory activity after storage.
• NP-based delivery represents a promising strategy for stabilizing labile epigenetic drugs like 5-Aza for therapeutic applications.