Mass spectrometric detection of antigen-specific immunoglobulin peptides in paraneoplastic patient sera

Maat, Peter; VanDuijn, Martijn M.; Brouwer, Eric; Dekker, Lennard J. M.; Zeneyedpour, Lona; Luider, Theo M.; Smitt, Peter Sillevis

doi:10.1016/j.jaut.2012.02.002

Cited by 16 publications

(14 citation statements)

References 27 publications

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“…The old paradigm in immunological sciences states that antigen specificity is determined by a completely random process which will result in unique antigen binding peptide sequence regions in antibodies (of similar target) in different individuals. A number of studies, several of which were MS-based, have in recent years challenged this dogma [42]–[47]. Even though MS methods are not as sensitive as genomic studies, MS-based methodologies may still give a truer picture of the expression levels of the abundant IgG-peptide repertoire.…”

Section: Discussionmentioning

confidence: 99%

IgG Antibodies to Cyclic Citrullinated Peptides Exhibit Profiles Specific in Terms of IgG Subclasses, Fc-Glycans and a Fab-Peptide Sequence

et al. 2014

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The Fc-glycan profile of IgG1 anti-citrullinated peptide antibodies (ACPA) in rheumatoid arthritis (RA) patients has recently been reported to be different from non-ACPA IgG1, a phenomenon which likely plays a role in RA pathogenesis. Herein we investigate the Fc-glycosylation pattern of all ACPA-IgG isotypes and simultaneously investigate in detail the IgG protein-chain sequence repertoire. IgG from serum or plasma (S/P, n = 14) and synovial fluid (SF, n = 4) from 18 ACPA-positive RA-patients was enriched using Protein G columns followed by ACPA-purification on cyclic citrullinated peptide-2 (CCP2)-coupled columns. Paired ACPA (anti-CCP2 eluted IgG) and IgG flow through (FT) fractions were analyzed by LC-MS/MS-proteomics. IgG peptides, isotypes and corresponding Fc-glycopeptides were quantified and interrogated using uni- and multivariate statistics. The Fc-glycans from the IgG4 peptide EEQFNSTYR was validated using protein A column purification. Relative to FT-IgG4, the ACPA-IgG4 Fc-glycan-profile contained lower amounts (p = 0.002) of the agalacto and asialylated core-fucosylated biantennary form (FA2) and higher content (p = 0.001) of sialylated glycans. Novel differences in the Fc-glycan-profile of ACPA-IgG1 compared to FT-IgG1 were observed in the distribution of bisected forms (n = 5, p = 0.0001, decrease) and mono-antennnary forms (n = 3, p = 0.02, increase). Our study also confirmed higher abundance of FA2 (p = 0.002) and lower abundance of afucosylated forms (n = 4, p = 0.001) in ACPA-IgG1 relative to FT-IgG1 as well as lower content of IgG2 (p = 0.0000001) and elevated content of IgG4 (p = 0.004) in ACPA compared to FT. One λ-variable peptide sequence was significantly increased in ACPA (p = 0.0001). In conclusion, the Fc-glycan profile of both ACPA-IgG1 and ACPA-IgG4 are distinct. Given that IgG1 and IgG4 have different Fc-receptor and complement binding affinities, this phenomenon likely affects ACPA effector- and immune-regulatory functions in an IgG isotype-specific manner. These findings further highlight the importance of antibody characterization in relation to functional in vivo and in vitro studies.

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Section: Discussionmentioning

confidence: 99%

IgG Antibodies to Cyclic Citrullinated Peptides Exhibit Profiles Specific in Terms of IgG Subclasses, Fc-Glycans and a Fab-Peptide Sequence

et al. 2014

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“…In another study on Sjö gren syndrome, secreted human Ro52 antibody from unrelated patients was found to share public V region sequences (27). Finally, our group observed 28 common Ig-derived sequences in paraneoplastic anti-Hu-syndrome that were specific for autoantigen and were found exclusively in samples from single autoantibody-defined clinical neurological entities (12).…”

Section: Discussionmentioning

confidence: 71%

“…However, this view has recently been challenged (9 -11). Both after vaccination and in paraneoplastic syndromes such as anti-Hu, strikingly identical shared complementarity determining region (CDR) motifs were observed among patients (12). Of note is also a surprising study in which it was observed that malignant chronic lymphocytic leukemia B cells in different patients all recognized a single fungal antigen and showed shared use of CDR3 sequences among different individuals (13).…”

mentioning

confidence: 98%

Cerebrospinal-fluid-derived Immunoglobulin G of Different Multiple Sclerosis Patients Shares Mutated Sequences in Complementarity Determining Regions

Singh

Stoop

Stingl

et al. 2013

Molecular & Cellular Proteomics

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B lymphocytes play a pivotal role in multiple sclerosis pathology, possibly via both antibody-dependent and -independent pathways. Intrathecal immunoglobulin G in multiple sclerosis is produced by clonally expanded B-cell populations. Recent studies indicate that the complementarity determining regions of immunoglobulins specific for certain antigens are frequently shared between different individuals. In this study, our main objective was to identify specific proteomic profiles of mutated complementarity determining regions of immunoglobulin G present in multiple sclerosis patients but absent in healthy controls. To achieve this objective, we purified immunoglobulin G from the cerebrospinal fluid of 29 multiple sclerosis patients and 30 healthy controls and separated the corresponding heavy and light chains via SDS-PAGE. Subsequently, bands were excised, trypsinized, and measured with high-resolution mass spectrometry. We sequenced 841 heavy and 771 light chain variable region peptides. We observed 24 heavy and 26 light chain complementarity determining regions that were solely present in a number of multiple sclerosis patients. Using stringent criteria for the identification of common peptides, we found five complementarity determining regions shared in three or more patients and not in controls. Interestingly, one complementarity determining region with a single mutation was found in six patients. Additionally, one other patient carrying a similar complementarity determining region with another mutation was observed. In addition, we found a skew in the -to-ratio and in the usage of certain variable heavy regions that was previously observed at the transcriptome level. At the protein level, cerebrospinal fluid immunoglobulin G shares common characteristics in the antigen binding region among different multiple sclerosis patients. The indication of a shared fingerprint may indicate common antigens for B-cell activation. Molecular & Cellular Proteomics 12:

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“…To validate the utility and specificity of our secondary antibody (goat anti-human IgG (HþL)), we designed a similar sandwich ELISA using as a positive control a different human protein and sera that were known to contain autoantibodies to it. Patients with paraneoplastic neurologic syndromes (PNS) can generate autoantibodies against the onconeural antigen HuD (23). Recombinant HuD antigen (ELAV4) was applied to serial dilutions of patient serum samples, known to be positive for anti-HuD antibodies.…”

Section: Detection Of Anti-survivin Antibodies By Elisamentioning

confidence: 99%

Survivin Autoantibodies Are Not Elevated in Lung Cancer When Assayed Controlling for Specificity and Smoking Status

Broodman

VanDuijn

Stingl

et al. 2016

Cancer Immunology Research

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The high mortality rate in lung cancer is largely attributable to late diagnosis. Case-control studies suggest that autoantibodies to the survivin protein are potential biomarkers for early diagnosis. We tested the hypothesis that sandwich ELISA can detect autoantibodies to survivin before radiologic diagnosis in patients with early-stage non-small cell lung cancer (NSCLC). Because previous studies assayed survivin autoantibodies with the direct antigencoating ELISA (DAC-ELISA), we first compared that assay with the sandwich ELISA. Based on the more robust results from the sandwich ELISA, we used it to measure survivin autoantibodies in the serum of 100 individuals from a well-controlled population study [the Dutch-Belgian Lung Cancer Screening Trial (NELSON) trial] composed of current and former smokers (50 patients with NSCLC, both before and after diagnosis, and 50 matched, smoking-habit control subjects), and another 50 healthy nonsmoking control subjects. We found no difference in specific autoantibodies to survivin in NSCLC patients, although nonspecific median optical densities were 24% higher (P < 0.001) in both NSCLC patients and smokers, than in healthy nonsmokers. Finally, we confirmed the ELISA results with Western blot analysis of recombinant and endogenous survivin (HEK-293), which showed no anti-survivin reactivity in patient sera. We conclude that specific anti-survivin autoantibody reactivity is most likely not present in sera before or after diagnosis. Autoantibody studies benefit from a comparison to a well-controlled population, stratified for smoking habit.

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Mass spectrometric detection of antigen-specific immunoglobulin peptides in paraneoplastic patient sera

Cited by 16 publications

References 27 publications

IgG Antibodies to Cyclic Citrullinated Peptides Exhibit Profiles Specific in Terms of IgG Subclasses, Fc-Glycans and a Fab-Peptide Sequence

IgG Antibodies to Cyclic Citrullinated Peptides Exhibit Profiles Specific in Terms of IgG Subclasses, Fc-Glycans and a Fab-Peptide Sequence

Cerebrospinal-fluid-derived Immunoglobulin G of Different Multiple Sclerosis Patients Shares Mutated Sequences in Complementarity Determining Regions

Survivin Autoantibodies Are Not Elevated in Lung Cancer When Assayed Controlling for Specificity and Smoking Status

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