Conventional and hydrolysable PEGs: comparative evaluation of tissue accumulation, biodistribution and toxicological...

PEG has been commonly considered safe and is widely used in pharmaceutical and personal care products. Concerns about its safety arised in relation to studies demonstrating its accumulation in some organs of animals treated with very high PEG dosages (e.g., 200 mg per kg per week for up to 12 weeks). Although these dosages are substantially higher (200-100 000 times) than those of PEG-containing drugs, the concerns might still be valid for chronic therapies if the polymer is not cleared after the accumulation. In this study, in addition to investigate the phenomena of PEG accumulation/elimination and its effects on full blood count, plasma biochemistry and behavioral performance of treated animals, we also studied new hydrolysable PEGs (HyPEGs) for reducing PEG accumulation. HyPEGs contain a cleavable linker that, upon in vivo hydrolysis, lowers by half the size of the polymer. We here evaluated the in vitro effect of 20- and 40 kDa conventional PEGs on macrophages' phenotype in THP1 cells, and the potential in vivo toxicity, biodistribution, and vacuolation in liver, kidneys, spleen, choroid plexus, heart, and lungs of a small library of 20- and 40 kDa-conventional and hydrolysable PEGs, with both linear and branched structures, administered twice weekly for 12 weeks at the high dosage of 200 mg per kg per week followed by an 8-week washout period (no PEG administration). We demonstrated that PEG internalization by macrophages resulted in molecular weight-dependent cellular responses. In vivo, all PEGs accumulated in the liver, spleen, kidneys, and choroid plexus, and in general accumulation of HyPEGs was less evident. The 8-week washout period allowed for the clearance of most of the accumulated PEGs in all organs except for the kidneys, likely because the kidneys are the terminal organs for PEG elimination. Overall, all PEGs did not show any hematotoxicity by analyzing biochemical- and hematological-parameters. Cytoplasmic PEG-negative vacuoles were present in the choroid plexus, suggesting adaptive cellular responses, but no signs of neurotoxicity were observed. Our results indicate that, although the different PEGs elicited distinct macrophage responses in vitro, all hydrolysable PEGs showed limited tissue accumulation in vivo without signs of toxicity, highlighting their safety.