title: Selectivity of binding of PEGs and PEG-like oligomers to anti-PEG antibodies induced by methoxyPEG-proteins.
abstract: The use of methoxypoly(ethylene glycol) (mPEG) in PEG conjugates of proteins and non-protein therapeutic agents has led to the recognition that the polymer components of such conjugates can induce anti-PEG antibodies (anti-PEGs) that may accelerate the clearance and reduce the efficacy of the conjugates. Others have classified anti-PEGs as "methoxy-specific" or "backbone-specific". The results of our previous research on anti-PEGs in the sera of rabbits immunized with mPEG or hydroxyPEG (HO-PEG) conjugates of three unrelated proteins were consistent with that classification (Sherman, M.R., et al., 2012. Bioconjug. Chem. 23, 485-499). Enzyme-linked immunosorbent assays (ELISAs) were performed on rabbit antisera and rabbit monoclonal anti-PEGs with competitors including 10 kDa mPEG, 10 kDa PEG diol and six linear or cyclic oligomers of oxyethylene (CH2CH2O), with molecular weights of ca. 150-264 Da. Our results demonstrate that (1) the binding affinities of anti-mPEGs depend more on the backbone lengths of the polymers and the hydrophobicities of their end-groups than on their resemblance to the methoxy terminus of the immunogenic polymer; (2) anti-PEGs raised against HO-PEG-proteins are not directed against the terminal hydroxy group, but against the backbone; (3) rabbit anti-PEGs bind to and distinguish among PEG-like oligomers with as few as three oxyethylene groups; and (4) none of the monoclonal or polyclonal anti-PEGs was absolutely "methoxy-specific" or "backbone-specific", but displayed distinct relative selectivities. If these results are relevant to human immune responses, the clinical use of stable conjugates of HO-PEG with proteins and non-protein therapeutic agents would be expected to produce fewer and less intense immune responses than those induced by conjugates with mPEG or PEGs with larger alkoxy groups.
authors.affiliation: Mountain View Pharmaceuticals, Inc., 3475 Edison Way, Suite S, Menlo Park, CA 94025-1821, United States.
authors.forename: Mark G P
authors.initials: MG
authors.lastname: Saifer
authors.forename: L David
authors.initials: LD
authors.lastname: Williams
authors.forename: Monika A
authors.initials: MA
authors.lastname: Sobczyk
authors.forename: Shawnya J
authors.initials: SJ
authors.lastname: Michaels
authors.forename: Merry R
authors.initials: MR
authors.lastname: Sherman
mesh_terms: D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000097:blood / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000032:analysis / Q000276:immunology; D000915:Antibody Affinity / Q000276:immunology*; D006801:Humans; D011092:Polyethylene Glycols / Q000737:chemistry / Q000378:metabolism*; D011506:Proteins / Q000276:immunology*; D011817:Rabbits
chemical_list: D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D011506:Proteins; D011092:Polyethylene Glycols; C028210:monomethoxypolyethylene glycol
keywords: 20kDa mPEG-KLH; 5kDa mPEG-KLH; AAALAC; Albumin; Anti-PEG; Anti-PEGs; Association for the Assessment and Accreditation of Laboratory Animal Care; Backbone-specific; D(50); ELISA; EtO-TEG; H and L chains; HO-PEG; IACUC; IC(50); IFN-α; Institutional Animal Care and Use Committee; KLH; Methoxy-specific; Monoclonal; NPC; PEG; PEG20K-KLH; Poly(ethylene glycol); Polyclonal; RI; RSD; SOD; TEG diol and TetraEG are defined in Table 1; UV; anti-PEG antibodies; dilution of serum that corresponds to 50% of maximal binding in a direct ELISA; enzyme-linked immunosorbent assay; heavy and light immunoglobulin chains; human serum albumin; hydroxyPEG; inhibitor concentration that reduces binding to 50% of maximal binding in a competitive ELISA; interferon-α; keyhole limpet hemocyanin; mAU/min; mAb; mPEG; mPEG-KLH; mTEG; milli-absorbance units per minute; monoclonal antibody; monomethoxypoly(ethylene glycol); n-BuO-TEG; p-nitrophenylcarbonate; p-nitrophenylchloroformate; pNPCOCl; poly(ethylene glycol); porcine Cu–Zn superoxide dismutase; refractive index; relative standard deviation=s.d./mean; ultraviolet absorbance. Crown ether