We examined if HSRs can be caused by interaction of PLD with antibodies that bind to poly(ethylene glycol) (PEG) in the corona of the PLD. histamine and platelet-activating factor. Anti-PEG IgG also induced hypothermia in mice after administration of other PEGylated liposomes, nanoparticles, or proteins. Humanized anti-PEG IgG promoted binding of PEGylated nanoparticles to human immune cells and induced secretion of histamine from human basophils in the presence of PLD. Anti-PEG IgE could also induce hypersensitivity reactions in mice after administration of PLD. Our results demonstrate an ASTX-660 important role for IgG antibodies in induction of HSRs to PEGylated nanomedicines through interaction with KL-1 Fc receptors on innate immune cells and provide a deeper understanding of HSRs to PEGylated nanoparticles and macromolecular drugs that may facilitate development of safer nanomedicines. Keywords:poly(ethylene glycol), immunogenicity, anti-PEG antibodies, hypersensitivity reactions, PEGylated liposomal doxorubicin, innate immune cells, basophils == Introduction == PEGylated nanoparticles and macromolecular drugs can induce infusion-related hypersensitivity reactions (HSRs) in some patients, particularly during their first exposure to the medicine.13HSRs usually occur immediately after administration of PEGylated nanomedicines and biomolecules with symptoms that include flushing and facial swelling, breathing difficulties, head and back pain, tightness in the chest or throat, hypothermia, ASTX-660 hypotension, and death in the most severe cases.4This is typified by PEGylated liposomal doxorubicin (PLD), a liposomal formulation of doxorubicin hydrochloride (Doxil, Caelyx, and their generic versions) used in almost one million cancer patients but which can cause infusion related hypersensitivity reactions (HSRs) in 5% to 10% of patients.57HSRs can force discontinuation of beneficial treatment, place a substantial burden on the medical ASTX-660 system, and may prevent successful translation of PEGylated nanomedicines to the clinic. HSRs are therefore recognized as a major barrier to the development of nanomedicines.8,9 HSRs to nanomedicines have been linked to complement activation-related pseudoallergy (CARPA), in which liberation of complement products stimulate innate immune cells to secrete vasoactive and inflammatory mediators.6,10Indeed, the levels of plasma C terminal complex (SC5b-9) is dose-dependently increased in patients experiencing HSR.5,11Rapid uptake of nanoparticles into phagocytic cells such as macrophages and activation of innate immune cells independently of complement activation have also been proposed to cause infusion related adverse reactions to nanoparticles.12,13However, the actual mechanism of HSRs caused by nanomedicines remains controversial.3,13,14 Poly(ethylene glycol) (PEG) is physically attached to many macromolecular drugs, nanoparticles, and liposomes to achieve desirable pharmacokinetic properties and enhance their biological activity. Antibodies against PEG, however, are naturally present in many normal individuals, possibly due to exposure to PEG in ASTX-660 a wide range of household products such as lotions, creams, and shampoos, which is why they are also referred to as pre-existing anti-PEG antibodies.15,16Some PEGylated drugs also induce the production of antibodies against PEG.1720Anti-PEG antibodies can accelerate drug clearance from the circulation, alter drug biodistribution, activate complement, destabilize the integrity of PEGylated nanomedicines, and reduce drug therapeutic efficacy.2127Anti-PEG antibodies can also induce hypersensitivity reactions in patients receiving PEGylated medicines.5,6,28,29 Here we examine the hypothesis that anti-PEG antibodies bound to PEGylated nanoparticles and macromolecules can interact with Fc receptors on immune cells to initiate HSRs.30We demonstrate that anti-PEG IgG but not IgM antibodies induce hypersensitivity-like symptoms against PLD and other PEGylated nanoparticles and macromolecules in mice that depend primarily on neutrophils, macrophages, and basophils. HSR symptoms are alleviated by blocking Fc receptors and elaboration of histamine and platelet-activating factor. Human anti-PEG IgG can induce histamine secretion from human basophils in the presence of PLD, consistent with an important role for Fc receptor-mediated responses to antibodies bound to PEGylated nanoparticles in HSRs. == Materials and Methods == == Animals == Nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice (NOD.CB17-Prkdcscid/NcrCrl, 812 weeks old) were obtained from BioLASCO Taiwan Co., Ltd. BALB/c and C57BL/6JNarl mice (812 weeks old) were obtained from the National Laboratory Animal Center, Taipei, Taiwan. Sash (c-Kitw-sh) mice were a kind gift from Dr. Ya-Jen.