Preclinical safety and hepatotoxicity evaluation of biomineralized copper sulfide nanoagents.

Albumin-biomineralized copper sulfide nanoparticles (Cu2-xS NPs) have attracted much attention as an emerging phototheranostic agent due to their advantages of facile preparation method and high biocompatibility. However, comprehensive preclinical safety evaluation is the only way to meet its further clinical translation. We herein evaluate detailedly the safety and hepatotoxicity of bovine serum albumin-biomineralized Cu2-xS (BSA@Cu2-xS) NPs with two different sizes in rats. Large-sized (LNPs, 17.8 nm) and small-sized (SNPs, 2.8 nm) BSA@Cu2-xS NPs with great near-infrared absorption and photothermal conversion efficiency are firstly obtained. Seven days after a single-dose intravenous administration, SNPs distributed throughout the body are cleared primarily through the feces, while a large amount of LNPs remained in the liver. A 14-day subacute toxicity study with a 28-day recovery period are conducted, showing long-term hepatotoxicity without recovery for LNPs but reversible toxicity for SNPs. Cellular uptake studies indicate that LNPs prefer to reside in Kupffer cells, leading to prolonged and delayed hepatotoxicity even after the cessation of NPs administration, while SNPs have much less Kupffer cell uptake. RNA-sequencing analysis for gene expression indicates that the inflammatory pathway, lipid metabolism pathway, drug metabolism-cytochrome P450 pathway, cholesterol/bile acid metabolism pathway, and copper ion transport/metabolism pathway are compromised in the liver by two sizes of BSA@Cu2-xS NPs, while only SNPs show a complete recovery of altered gene expression after NPs discontinuation. This study demonstrates that the translational feasibility of small-sized BSA@Cu2-xS NPs as excellent nanoagents with manageable hepatotoxicity.