title: Bridging disulfides for stable and defined antibody drug conjugates.
abstract: To improve both the homogeneity and the stability of ADCs, we have developed site-specific drug-conjugating reagents that covalently rebridge reduced disulfide bonds. The new reagents comprise a drug, a linker, and a bis-reactive conjugating moiety that is capable of undergoing reaction with both sulfur atoms derived from a reduced disulfide bond in antibodies and antibody fragments. A disulfide rebridging reagent comprising monomethyl auristatin E (MMAE) was prepared and conjugated to trastuzumab (TRA). A 78% conversion of antibody to ADC with a drug to antibody ratio (DAR) of 4 was achieved with no unconjugated antibody remaining. The MMAE rebridging reagent was also conjugated to the interchain disulfide of a Fab derived from proteolytic digestion of TRA, to give a homogeneous single drug conjugated product. The resulting conjugates retained antigen-binding, were stable in serum, and demonstrated potent and antigen-selective cell killing in in vitro and in vivo cancer models. Disulfide rebridging conjugation is a general approach to prepare stable ADCs, which does not require the antibody to be recombinantly re-engineered for site-specific conjugation.
authors.affiliation: PolyTherics Ltd, The London Bioscience Innovation Centre , 2 Royal College Street, London NW1 0NH, United Kingdom.
authors.forename: George
authors.initials: G
authors.lastname: Badescu
authors.forename: Penny
authors.initials: P
authors.lastname: Bryant
authors.forename: Matthew
authors.initials: M
authors.lastname: Bird
authors.forename: Korinna
authors.initials: K
authors.lastname: Henseleit
authors.forename: Julia
authors.initials: J
authors.lastname: Swierkosz
authors.forename: Vimal
authors.initials: V
authors.lastname: Parekh
authors.forename: Rita
authors.initials: R
authors.lastname: Tommasi
authors.forename: Estera
authors.initials: E
authors.lastname: Pawlisz
authors.forename: Kosma
authors.initials: K
authors.lastname: Jurlewicz
authors.forename: Monika
authors.initials: M
authors.lastname: Farys
authors.forename: Nicolas
authors.initials: N
authors.lastname: Camper
authors.forename: XiaoBo
authors.initials: X
authors.lastname: Sheng
authors.forename: Martin
authors.initials: M
authors.lastname: Fisher
authors.forename: Ruslan
authors.initials: R
authors.lastname: Grygorash
authors.forename: Andrew
authors.initials: A
authors.lastname: Kyle
authors.forename: Amrita
authors.initials: A
authors.lastname: Abhilash
authors.forename: Mark
authors.initials: M
authors.lastname: Frigerio
authors.forename: Jeff
authors.initials: J
authors.lastname: Edwards
authors.forename: Antony
authors.initials: A
authors.lastname: Godwin
mesh_terms: D061067:Antibodies, Monoclonal, Humanized / Q000737:chemistry* / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004220:Disulfides / Q000737:chemistry*; D004305:Dose-Response Relationship, Drug; D004354:Drug Screening Assays, Antitumor; D006801:Humans; D061986:MCF-7 Cells; D015394:Molecular Structure; D009842:Oligopeptides / Q000737:chemistry* / Q000494:pharmacology*; D013329:Structure-Activity Relationship; D000068878:Trastuzumab
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D004220:Disulfides; D009842:Oligopeptides; D000068878:Trastuzumab; C495575:monomethyl auristatin E