Bioresponsive Reversibly Crosslinked Hyaluronic Acid-Polypeptide Nanoparticles For Active Tumor-Targeted Cancer Therapy

Biodegradable polymereric nanoparticles can remarkably enhance the water solubility of anticancer drugs, effectively prolong blood circulation time of drugs in vivo, greatly improve tumor accumulation of anticancer drugs owing to the enhanced permeability and retention (EPR) effect, and enormously reduce systemic toxicity, thus have emerged as one of the most promising nanoplatforms for tumor-targeted therapy. Quite a few nanotherapeutics based on biodegradable polymeric nanopartics have been advanced to clinical practices or clinical trials. However, the results of the clinical trials indicate that the therapeutic efficacy of these nanotherapeutics is far from expectations due to the low stability in vivo leading to premature drug release and low tumor accumulation, inefficient tumor cell uptake; and inadequate or slow intracellular drug release following uptake by tumor cells. In Chapter 1, we first summarized biodegradable polymeric nanotherapeutics currently used in clinic and clinical trials as well as their advantages and pratical use on cancer therapy, and then highlight the recent advanced strategies on enhancing in vivo stability, improving cellular uptake and accelerating intracellular drug release of nanotherapeutics.In Chapter 2, bioresponsive reversibly crosslinked hyaluronic acid-polypeptide nanoparticles were developed from hyaluronic acid-g-poly(?-benzyl-L-glutamate)-lipoic acid(HA-g-PBLG-LA) for active targeted delivery of doxorubicin (DOX) to human breast tumor xenograft in vivo. DOX-loaded nanoparticles following auto-crosslinking (DOX-CLNPs) are highly stable with little drug leakage under physiological conditions while quickly release ca.92% DOX in 30 h under a cytoplasmic-mimicking reductive environment. The in vitro assays reveal that DOX-CLNPs possess a superior selectivity and antitumor activity to clinically used pegylated liposomal doxorubicin hydrochloride (DOX-LPs) in CD44 receptor overexpressing MCF-7 human breast cancer cells. Strikingly, DOX-CLNPs exhibit a superb tolerated dose of over 100 mg DOX equiv./kg, which is more than 5 times higher than DOX-LPs, and an extraordinary breast tumor accumulation of 8.6%ID/g in mice. The in vivo therapeutic studies in MCF-7 human breast tumor-bearing nude mice show that DOX-CLNPs effectively inhibit tumor growth, improve survival rate, and significantly decrease adverse effects as compared to DOX-LPs. DOX-CLNPs based on natural endogenous materials with high drug loading, great stability and CD44-targetability are highly promising for precision cancer chemotherapy,In Chapter 3, pH and reduction dual-responsive reversibly crosslinked nanoparticles were developed from hyaluronic acid-g-poly(?-diisopropylethylenediamine-L-aspartate)-lipoic acid (HA-g-P(Asp-DIP)-LA) for active targeted delivery and triggered intracellular release of anticancer proteins. HA-g-P(Asp-DIP)-LA following interaction with cytochrome C (CC) via electrostatic effect could self-assembled in aqueous solution into nanoparticles with an average diameter of about 166-185 nm, achieving high protein loading content (up to 26.9%) and protein loading efficiency (up to 92.1%). In vitro drug release results indicated that the CC-loaded crosslinked nanoparticles (CC-CLNPs) had high stability at physiological environment (pH 7.4,37?), in which less than 15% of CC was released in 24 h. On the contrary,36.4% of CC was released in 24 h at endosomal pH of 5.0 mainly due to the swell of nanoparticles resulted from complete protonation of DIP segements. Moreover, rapid CC release was observed under a reductive condition (10 mM GSH), in which 65.2% and 100% of CC were released at pH 7.4 and 5.0 in 24 h, respectively, indicating that CC-CLNPs had pH and reduction dual-sensitivity. MTT assays demonstrated that CC-CLNPs caused obvious cell apoptosis with IC50 (inhibitory concentration to produce 50% cell death) of 33.8?g CC equiv/mL in CD44 overexpressed MCF-7 cells. Confocal laser scan microscopy (CLSM) observation showed strong fluorescence of FITC labeled CC (FITC-CC) was observed in the cytoplasm after the incubation of CC-CLNPs with MCF-7 cells for 8 h. The inhibition experiments demonstrated that the fluorescence of FITC-CC in MCF-7 cells was negligible by pretreating the cells with free HA (5 mg/mL) prior to incubating with CC-CLNPs, confirming that CC-CLNPs was taken up by MCF-7 cells via a receptor-mediated mechanism. These dual-responsive reversibly crosslinked nanoparticles based on HA-polypeptide have appeared as a highly potential nanoplatform for active targeted delivery of anticancer proteins.