Unexpected Toxicity of Monolayer Protected Gold Clusters Eliminated by PEG-Thiol Place Exchange Reactions

Abstract

Monolayer protected clusters (MPCs) are small, metal nanoparticles capped with thiolate ligands that have been widely studied for their size-dependent properties and for their ability to be functionalized for biological applications. Common water-soluble MPCs, functionalized by 2-mercaptopropanoyl) amino acetic acid (tiopronin) or glutathione, have been used previously to interface with biological systems. These MPCs are ideal for biological applications not only due to their water-solubility but also their small size (< 5 nm). These characteristics are expected to enable easy biodistribution and clearance. In this report we show an unexpected toxicity is associated with the tiopronin monolayer protected cluster (TMPC), making it incompatible for potential in vivo applications. This toxicity is linked to significant histological damage to the renal tubules, causing mortality at concentrations above 20 μM. We further show how the incorporation of poly-ethylene glycol (PEG) by simple place-exchange reaction eliminates this toxicity. We analyzed gold content within blood and urine and found an increased lifetime of the particle within the bloodstream due to the creation of the mixed monolayer. Also shown was the elimination of kidney damage with the use of the mixed-monolayer particle via Multistix™ analysis, MALDI-TOF MS analysis, and histological examination. Final immunological analysis showed no effect on white blood cell (WBC) count for the unmodified particle and a surprising increase in WBC count with injection of mixed monolayer particles at concentrations higher than 30 μM, suggesting that there may be an immune response to these mixed monolayer nanoparticles at high concentrations; therefore, special attention should be focused on selecting the best capping ligands for use in vivo. These findings make the mixed monolayer an excellent candidate for further biological applications using water-soluble nanoparticles.

Acute hematologic, hepatologic, and nephrologic changes after intraperitoneal injections of 18 nm gold nanoparticles in hamsters

“…The leukocytosis observed in GNP-injected hamsters is in agreement with Simpson et al, 22 Zhang et al, 47 and Sengupta et al 48 The elevation in total leukocytic count might be attributed to the immunogenic effect of GNPs 15 or to their ability to trigger an inflammatory response by stimulating the release of cytokines. 49 Previously reported leukocytosis after injection of 10 nm and 60 nm particles in mice was attributed to an inflammatory response.…”

“…[17][18][19] The particle surface chemistry and charge also have important roles in the induced changes due to their immunogenicity and effects on bioclearance. 13,15,[20][21][22][23][24] The route of administration has a major impact on GNP-induced effects. Intravenous route was shown to be the safest 12,25 followed by subcutaneous application.…”

Inductively Coupled Plasma‐Mass Spectrometry in Biodistribution Studies of (Engineered) Nanoparticles

“…Distribution studies have shown that coating Au NPs with polyethylene glycol (PEG) considerably increases blood circulation times compared to bare particles (10,22) and decreases their toxic effects on the kidney. (7) Prolongation of blood circulation was also observed when Au nanorods were coated with PEG. (23) An increase in blood circulation time and conjugation of Au NPs to ligands, such as TNFα, improves their accumulation in tumor tissue.…”

“…(6) In addition, Au NPs modified with certain thiol monolayers, such as tiopronin, are not as inert as previously assumed, causing renal complications and morbidity in mice. (7) Conducting biodistribution studies is therefore of paramount importance to ensure that the NPs reach their target tissue in the body, and leave the body in due time after their job is done, without causing damage.…”

“…(22) Unfortunately, PEG-coated Au NPs have been shown to increase inflammation markers in blood and liver and lead to lower cellular uptake and binding interactions with protein disease targets, reducing their potential as drug carriers. (7,10,24) As an alternative to PEG-coating, two recent studies by Simpson et al (25) and Zhang et al (6) have investigated biodistribution and clearance of 1.2 and 2.1 nm Au NPs and nanoclusters, respectively, coated with glutathione (GSH) using ICPMS. Blood circulation times of GSHcoated Au NPs were considerably longer compared to PEG-coated or bovine serum albumin (BSA) coated particles.…”