title: The impact of glycosylation on the pharmacokinetics of a TNFR2:Fc fusion protein expressed in Glycoengineered Pichia Pastoris.
abstract: OBJECTIVE
  P. pastoris has previously been genetically engineered to generate strains that are capable of producing mammalian-like glycoforms. Our objective was to investigate the correlation between sialic acid content and pharmacokinetic properties of recombinant TNFR2:Fc fusion proteins generated in glycoengineered P. pastoris strains.
  
  
  METHODS
  TNFR2:Fc fusion proteins were generated with varying degrees of sialic acid content. The pharmacokinetic properties of these proteins were assessed by intravenous and subcutaneous routes of administration in rats. The binding of these variants to FcRn were also evaluated for possible correlations between in vitro binding and in vivo PK.
  
  
  RESULTS
  The pharmacokinetic profiles of recombinant TNFR2:Fc produced in P. pastoris demonstrated a direct positive correlation between the extent of glycoprotein sialylation and in vivo pharmacokinetic properties. Furthermore, recombinant TNFR2:Fc produced in glycoengineered Pichia, with a similar sialic acid content to CHO-produced etanercept, demonstrated similar in vivo pharmacokinetic properties to the commercial material. In vitro surface plasmon resonance FcRn binding at pH6.0 showed an inverse relationship between sialic acid content and receptor binding affinity, with the higher affinity binders having poorer in vivo PK profiles.
  
  
  CONCLUSIONS
  Sialic acid content is a critical attribute for modulating the pharmacokinetics of recombinant TNFR2:Fc produced in glycoengineered P. pastoris.
authors.affiliation: Department of Pharmacokinetics, Pharmacodynamics & Drug Metabolism Merck Research Laboratories, West Point, Pennsylvania, 19486, USA.
authors.forename: Liming
authors.initials: L
authors.lastname: Liu
authors.forename: Sujatha
authors.initials: S
authors.lastname: Gomathinayagam
authors.forename: Lora
authors.initials: L
authors.lastname: Hamuro
authors.forename: Thomayant
authors.initials: T
authors.lastname: Prueksaritanont
authors.forename: Weirong
authors.initials: W
authors.lastname: Wang
authors.forename: Terrance A
authors.initials: TA
authors.lastname: Stadheim
authors.forename: Stephen R
authors.initials: SR
authors.lastname: Hamilton
mesh_terms: D000818:Animals; D003001:Cloning, Molecular; D000068800:Etanercept; D005818:Genetic Engineering; D006031:Glycosylation; D015395:Histocompatibility Antigens Class I / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood* / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007166:Immunosuppressive Agents / Q000097:blood* / Q000737:chemistry / Q000378:metabolism; D008297:Male; D019158:N-Acetylneuraminic Acid / Q000032:analysis; D010843:Pichia / Q000235:genetics*; D011485:Protein Binding; D051381:Rats; D017207:Rats, Sprague-Dawley; D011961:Receptors, Fc / Q000378:metabolism; D018124:Receptors, Tumor Necrosis Factor / Q000097:blood* / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D015395:Histocompatibility Antigens Class I; D007074:Immunoglobulin G; D007166:Immunosuppressive Agents; D011961:Receptors, Fc; D018124:Receptors, Tumor Necrosis Factor; D019158:N-Acetylneuraminic Acid; D000068800:Etanercept; C100688:Fc receptor, neonatal