title: Russell body phenotype is preferentially induced by IgG mAb clones with high intrinsic condensation propensity: relations between the biosynthetic events in the ER and solution behaviors in vitro.
abstract: The underlying reasons for why some mAb (monoclonal antibody) clones are much more inclined to induce a Russell body (RB) phenotype during immunoglobulin biosynthesis remain elusive. Although RBs are morphologically understood as enlarged globular aggregates of immunoglobulins deposited in the endoplasmic reticulum (ER), little is known about the properties of the RB-inducing mAb clones as secretory cargo and their physical behaviors in the extracellular space. To elucidate how RB-inducing propensities, secretion outputs, and the intrinsic physicochemical properties of individual mAb clones are interrelated, we used HEK293 cells to study the biosynthesis of 5 human IgG mAbs for which prominent solution behavior problems were known a priori. All 5 model mAbs with inherently high condensation propensities induced RB phenotypes both at steady state and under ER-to-Golgi transport block, and resulted in low secretion titer. By contrast, one reference mAb that readily crystallized at neutral pH in vitro produced rod-shaped crystalline bodies in the ER without inducing RBs. Another reference mAb without notable solution behavior issues did not induce RBs and was secreted abundantly. Intrinsic physicochemical properties of individual IgG clones thus directly affected the biosynthetic steps in the ER, and thereby produced distinctive cellular phenotypes and influenced IgG secretion output. The findings implicated that RB formation represents a phase separation event or a loss of colloidal stability in the secretory pathway organelles. The process of RB induction allows the cell to preemptively reduce the extracellular concentration of potentially pathogenic, highly aggregation-prone IgG clones by selectively storing them in the ER.
authors.affiliation: a Department of Therapeutic Discovery; Amgen ; Seattle , WA USA.
authors.forename: Haruki
authors.initials: H
authors.lastname: Hasegawa
authors.forename: Christopher E
authors.initials: CE
authors.lastname: Woods
authors.forename: Francis
authors.initials: F
authors.lastname: Kinderman
authors.forename: Feng
authors.initials: F
authors.lastname: He
authors.forename: Ai Ching
authors.initials: AC
authors.lastname: Lim
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D015153:Blotting, Western; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D003460:Crystallization; D004721:Endoplasmic Reticulum / Q000276:immunology / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000378:metabolism; D007147:Immunoglobulin Light Chains / Q000235:genetics / Q000378:metabolism; D002479:Inclusion Bodies / Q000276:immunology / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D013314:Stress, Mechanical; D013696:Temperature; D014162:Transfection
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains
keywords: BFA, Brefeldin A; CB, crystalline body; DIC, differential interference contrast; ER, endoplasmic reticulum;; Fab, fragment antigen binding; HC, heavy chain; HEK, human embryonic kidney; IgG, immunoglobulin G; LC, light chain;; RB, Russell body; Russell body; VH, heavy chain variable domain; VL, light chain variable domain; crystalline body; endoplasmic reticulum; gelation; immunoglobulin; mAb, monoclonal antibody; phase separation; protein aggregation; protein condensation; protein crystallization