title: The CEA/CD3-bispecific antibody MEDI-565 (MT111) binds a nonlinear epitope in the full-length but not a short splice variant of CEA.
abstract: MEDI-565 (also known as MT111) is a bispecific T-cell engager (BiTE®) antibody in development for the treatment of patients with cancers expressing carcinoembryonic antigen (CEA). MEDI-565 binds CEA on cancer cells and CD3 on T cells to induce T-cell mediated killing of cancer cells. To understand the molecular basis of human CEA recognition by MEDI-565 and how polymorphisms and spliced forms of CEA may affect MEDI-565 activity, we mapped the epitope of MEDI-565 on CEA using mutagenesis and homology modeling approaches. We found that MEDI-565 recognized a conformational epitope in the A2 domain comprised of amino acids 326-349 and 388-410, with critical residues F(326), T(328), N(333), V(388), G(389), P(390), E(392), I(408), and N(410). Two non-synonymous single-nucleotide polymorphisms (SNPs) (rs10407503, rs7249230) were identified in the epitope region, but they are found at low homozygosity rates. Searching the National Center for Biotechnology Information GenBank® database, we further identified a single, previously uncharacterized mRNA splice variant of CEA that lacks a portion of the N-terminal domain, the A1 and B1 domains, and a large portion of the A2 domain. Real-time quantitative polymerase chain reaction analysis of multiple cancers showed widespread expression of full-length CEA in these tumors, with less frequent but concordant expression of the CEA splice variant. Because the epitope was largely absent from the CEA splice variant, MEDI-565 did not bind or mediate T-cell killing of cells solely expressing this form of CEA. In addition, the splice variant did not interfere with MEDI-565 binding or activity when co-expressed with full-length CEA. Thus MEDI-565 may broadly target CEA-positive tumors without regard for expression of the short splice variant of CEA. Together our data suggest that MEDI-565 activity will neither be impacted by SNPs nor by a splice variant of CEA.
authors.affiliation: Department of Antibody Discovery and Protein Engineering, MedImmune LLC, Gaithersburg, Maryland, United States of America.
authors.forename: Li
authors.initials: L
authors.lastname: Peng
authors.forename: Michael D
authors.initials: MD
authors.lastname: Oberst
authors.forename: Jiaqi
authors.initials: J
authors.lastname: Huang
authors.forename: Philip
authors.initials: P
authors.lastname: Brohawn
authors.forename: Chris
authors.initials: C
authors.lastname: Morehouse
authors.forename: Kristen
authors.initials: K
authors.lastname: Lekstrom
authors.forename: Patrick A
authors.initials: PA
authors.lastname: Baeuerle
authors.forename: Herren
authors.initials: H
authors.lastname: Wu
authors.forename: Yihong
authors.initials: Y
authors.lastname: Yao
authors.forename: Steven R
authors.initials: SR
authors.lastname: Coats
authors.forename: William
authors.initials: W
authors.lastname: Dall'Acqua
authors.forename: Melissa
authors.initials: M
authors.lastname: Damschroder
authors.forename: Scott A
authors.initials: SA
authors.lastname: Hammond
mesh_terms: D000328:Adult; D000368:Aged; D000595:Amino Acid Sequence; D000818:Animals; D018033:Antibodies, Bispecific / Q000276:immunology*; D061067:Antibodies, Monoclonal, Humanized / Q000276:immunology; D000918:Antibody Specificity; D017252:CD3 Complex / Q000276:immunology*; D016466:CHO Cells; D002272:Carcinoembryonic Antigen / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D006224:Cricetinae; D003412:Cricetulus; D018604:Epitope Mapping; D000939:Epitopes / Q000276:immunology*; D015972:Gene Expression Regulation, Neoplastic; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D008875:Middle Aged; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000235:genetics / Q000473:pathology; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology; D055815:Young Adult
chemical_list: D018033:Antibodies, Bispecific; D061067:Antibodies, Monoclonal, Humanized; D017252:CD3 Complex; D002272:Carcinoembryonic Antigen; D000939:Epitopes; D020033:Protein Isoforms; D012333:RNA, Messenger