title: A novel assay to measure B cell responses to keyhole limpet haemocyanin vaccination in healthy volunteers and subjects with systemic lupus erythematosus.
abstract: The aim of the study was to characterize performance of a complementary set of assays to measure antigen-specific immune responses in subjects immunized with a neoantigen. Healthy volunteers (HV) (n = 8) and patients with systemic lupus erythematosus (SLE) (n = 6) were immunized with keyhole limpet haemocyanin (KLH) on days 1 and 29. Serum antibodies were detected using a flow cytometric bead array (CBA) that multiplexed the KLH response alongside pre-existing anti-tetanus antibodies. Peripheral blood mononuclear cells were studied by B cell ELISPOT. These assays were built upon precedent assay development in cynomolgus monkeys, which pointed towards their utility in humans. Primary anti-KLH IgG responses rose to a mean of 65-93-fold above baseline for HV and SLE patients, respectively, and secondary responses rose to a mean of 260-170-fold above baseline. High levels of anti-tetanus IgG were detected in pre-immunization samples and their levels did not change over the course of study. Anti-KLH IgG1-4 subclasses were characterized by a predominant IgG1 response, with no significant differences in subclass magnitude or distribution between HV and SLE subjects. Anti-KLH IgM levels were detectable, although the overall response was lower. IgM was not detected in two SLE subjects whodid generate an IgG response. All subjects responded to KLH by B cell ELISPOT, with no significant differences observed between HV and SLE subjects. The CBA and B cell ELISPOT assays reliably measured anti-KLH B cell responses, supporting use of this approach and these assays to assess the pharmacodynamic and potential safety impact of marketed/investigational immune-therapeutics.
authors.affiliation: Department of Medical Sciences, Amgen, Inc, Thousand Oaks, CA 91320, USA. jferbas@amgen.com
authors.forename: John
authors.initials: J
authors.lastname: Ferbas
authors.forename: Shelley S
authors.initials: SS
authors.lastname: Belouski
authors.forename: Michelle
authors.initials: M
authors.lastname: Horner
authors.forename: Arunan
authors.initials: A
authors.lastname: Kaliyaperumal
authors.forename: Li
authors.initials: L
authors.lastname: Chen
authors.forename: Malcolm
authors.initials: M
authors.lastname: Boyce
authors.forename: C Bernie
authors.initials: CB
authors.lastname: Colaço
authors.forename: Neil
authors.initials: N
authors.lastname: McHugh
authors.forename: Vanessa
authors.initials: V
authors.lastname: Quick
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Nicholl
authors.forename: Gerald
authors.initials: G
authors.lastname: Siu
authors.forename: James
authors.initials: J
authors.lastname: Chung
mesh_terms: D000276:Adjuvants, Immunologic / Q000008:administration & dosage*; D001402:B-Lymphocytes / Q000187:drug effects / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006433:Hemocyanins / Q000008:administration & dosage* / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D008180:Lupus Erythematosus, Systemic / Q000276:immunology / Q000517:prevention & control*; D014611:Vaccination
chemical_list: D000276:Adjuvants, Immunologic; D007074:Immunoglobulin G; D006433:Hemocyanins; C032808:keyhole-limpet hemocyanin
keywords: ELISPOT immunoassay; flow cytometric bead immunoassay; keyhole limpet haemocyanin; systemic lupus erythematosus; vaccine response