Synthesis of HA-SS-MP: A Prodrug With High Specificity for Cancer Cells

Thiolated hyaluronic acid was synthesized, and a quantification method was established to determine the content of the thiol group. Three thiol compounds, 2-nitro-5-mercapto-benzoic acid (TNB), 2- mercaptonicotinic acid (2-MNA), and mercaptopyridine (MPD), were tested for their efficiency in forming adducts with hyaluronic acid (HA) through disulfide bonds. Our results showed that MPD exhibited the highest disulfide exchange efficiency when the reaction was conducted at the optimal pH value. The synthesized hyaluronic acid-SS-mercaptopurine (HA-SS-MP) was characterized by 1H-nuclear magnetic resonance spectroscopy. The HA-SS-MP micelle showed a particle size of 264.4 nm in aqueous solution. An in vitro release experiment demonstrated that the releasing rate of 6-MP from HA-SS-MP in the release medium containing glutathione (GSH) was much higher than that in the release medium without GSH. Cell uptake experiments demonstrated the CD44 targeting of hyaluronic acid. Cytotoxicity tests in mouse melanoma cells (B16F10) showed that HA-SS-MP can kill melanoma cells effectively and with lower cytotoxicity, compared with the active ingredient, 6-MP.