Hyaluronan Reduces Cationic Liposome-Induced Toxicity and Enhances the Antitumor Effect of Targeted Gene Delivery in Mice.

Cationic nanocarriers are reported to induce cell necrosis, especially in the lungs upon systemic administration. The release of damage-associated molecular patterns, such as mitochondrial DNA from the injured cell may result in the inflammatory toxicity of the nanocarrier, which has largely limited its clinical application. Partial blocking of the surface charge of cationic nanocarriers might improve their safety. As hyaluronan (HA) is an anionic polysaccharide that is widely used for specific binding to CD44 to improve the cellular uptake efficiency in tumor-targeting therapy, in this study, we modified cationic liposomes (LP) with the negatively charged HA at a mass ratio of 10% to prepare targeted HA-modified cationic liposomes (HALP). Cationic liposomes modified with hyaluronan showed significantly less cytotoxicity due to the blockage of their surface charge than the unmodified liposomes. In addition, HA modification helped to reduce cell necrosis in lung tissue and reduced the amount of mitochondria subsequently released, which alleviated pulmonary inflammation in mice. HA-modified liposomes also improved the survival of mice injected with a fatal dose of HALP compared with mice injected with cationic LP. In addition, both serological biochemical analysis and histological examination proved that a liposome modified with HA is a safer carrier for systemic administration than an unmodified liposome. Furthermore, HALP/survivin exhibited an enhanced antitumor effect by inhibiting tumor growth and promoting tumor cell apoptosis compared with the unmodified LP group. In conclusion, compared to the properties of cationic liposomes, liposomes modified with 10% HA (HALP) might be gene vectors with lower toxicity and higher tumor targeting efficiency.

Cationic nanocarriers induce cell necrosis through impairment of Na(+)/K(+)-ATPase and cause subsequent inflammatory response.

Nanocarriers with positive surface charges are known for their toxicity which has limited their clinical applications. The mechanism underlying their toxicity, such as the induction of inflammatory response, remains largely unknown. In the present study we found that injection of cationic nanocarriers, including cationic liposomes, PEI, and chitosan, led to the rapid appearance of necrotic cells. Cell necrosis induced by cationic nanocarriers is dependent on their positive surface charges, but does not require RIP1 and Mlkl. Instead, intracellular Na(+) overload was found to accompany the cell death. Depletion of Na(+) in culture medium or pretreatment of cells with the Na(+)/K(+)-ATPase cation-binding site inhibitor ouabain, protected cells from cell necrosis. Moreover, treatment with cationic nanocarriers inhibited Na(+)/K(+)-ATPase activity both in vitro and in vivo. The computational simulation showed that cationic carriers could interact with cation-binding site of Na(+)/K(+)-ATPase. Mice pretreated with a small dose of ouabain showed improved survival after injection of a lethal dose of cationic nanocarriers. Further analyses suggest that cell necrosis induced by cationic nanocarriers and the resulting leakage of mitochondrial DNA could trigger severe inflammation in vivo, which is mediated by a pathway involving TLR9 and MyD88 signaling. Taken together, our results reveal a novel mechanism whereby cationic nanocarriers induce acute cell necrosis through the interaction with Na(+)/K(+)-ATPase, with the subsequent exposure of mitochondrial damage-associated molecular patterns as a key event that mediates the inflammatory responses. Our study has important implications for evaluating the biocompatibility of nanocarriers and designing better and safer ones for drug delivery.