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Whole-Proteome Peptide Microarrays for Profiling Autoantibody Repertoires within Multiple Sclerosis and Narcolepsy
KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0003-1242-0873
KTH, School of Biotechnology (BIO), Centres, Albanova VinnExcellence Center for Protein Technology, ProNova. KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0002-5248-8568
KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, School of Biotechnology (BIO), Nano Biotechnology (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.ORCID iD: 0000-0001-8141-8449
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2017 (English)In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 16, no 3, p. 1300-1314Article in journal (Refereed) Published
Abstract [en]

The underlying molecular mechanisms of autoimmune diseases are poorly understood. To unravel the autoimmune processes across diseases, comprehensive and unbiased analyses of proteins targets recognized by the adaptive immune system are needed. Here we present an approach starting from high-density peptide arrays to characterize autoantibody repertoires and to identify new autoantigens. A set of ten plasma and serum samples from subjects with multiple sclerosis, narcolepsy, and without any disease diagnosis were profiled on a peptide array representing the whole proteome, hosting 2.2 million 12-mer peptides with a six amino acid lateral shift. On the basis of the IgG reactivities found on these whole-proteome peptide micro arrays, a set of 23 samples was then studied on a targeted array with 174 000 12-mer peptides of single amino acid lateral shift. Finally, verification of IgG reactivities was conducted with a larger sample set (n = 448) using the bead-based peptide microarrays. The presented workflow employed three different peptide microarray formats to discover and resolve the epitopes of human autoantibodies and revealed two potentially new autoantigens: MAP3K7 in multiple sclerosis and NRXN1 in narcolepsy. The presented strategy provides insights into antibody repertoire reactivity at a peptide level and may accelerate the discovery and validation of autoantigens in human diseases.

Place, publisher, year, edition, pages
American Chemical Society (ACS), 2017. Vol. 16, no 3, p. 1300-1314
Keywords [en]
peptide microarrays, autoantibody profiling, epitope mapping narcolepsy, multiple sclerosis
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
URN: urn:nbn:se:kth:diva-205512DOI: 10.1021/acs.jproteome.6b00916ISI: 000395726200017PubMedID: 28121444Scopus ID: 2-s2.0-85014691057OAI: oai:DiVA.org:kth-205512DiVA, id: diva2:1098496
Funder
VINNOVAKnut and Alice Wallenberg Foundation
Note

QC 20170524

Available from: 2017-05-24 Created: 2017-05-24 Last updated: 2017-09-05Bibliographically approved
In thesis
1. Array-based Autoantibody Profiling and Epitope Mapping
Open this publication in new window or tab >>Array-based Autoantibody Profiling and Epitope Mapping
2017 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Antibodies are a class of proteins that are made by the immune system to recognize harmful organisms and molecules. Their exceptional capability of specifically recognizing molecules has been investigated for over a century and information thereof has been utilized for a variety of applications including vaccine and generation of therapeutic antibodies. Occasionally, instead of protecting the host against pathogens, antibodies can recognize constituents of the host and thereby cause an autoimmune reaction that eventually can lead to a disease. Therefore, it is of great interest to understand what the antibodies bind to and their specificities.

 

The last decades of technical development and availability of protein and peptide microarrays have enabled large-scale profiling of antibodies and precise determination of their specificities through epitope mapping. In this thesis the aim was to use affinity proteomics tools to profile antibodies, determine their specificities, and discover potential associations of autoantigens to disease by analyzing blood-derived samples with microarray-based methods.

 

In Paper I, 57 serum samples from patients with the suggested autoimmune disease narcolepsy, were analyzed on planar antigen microarrays with 10,846 human protein fragments. Verification on an independent sample collection consisting of serum samples from 176 individuals, revealed METTL22 and NT5C1A as two potential autoantigens. In Paper II, antibodies from 53 plasma samples from patients with first-episode psychosis, a condition suggested to have a partial autoimmune component, were analyzed on planar antigen microarrays with 2,304 human protein fragments. After a follow-up study of the patients, antibodies toward an antigen representing the three proteins, PAGE2, PAGE2B, PAGE5, was found associated to an increased risk of developing schizophrenia. In Paper III, serum and plasma samples from patients with the autoimmune diseases multiple sclerosis and narcolepsy, were epitope mapped on high-density peptide microarrays with approximately 2.2 million peptides. Technical and biological verification, by using other microarray technology and analyzing  samples from 448 patients, revealed one peptide for multiple sclerosis and narcolepsy, representing the proteins MAP3K7 and NRXN1, with higher antibody reactivity towards in each group, respectively. In Paper IV, purified polyclonal antibodies raised against a surface antigen found on malaria-infected erythrocytes, were profiled on the peptide microarrays representing all proteins found on malaria-infected erythrocytes derived from Plasmodium falciparum. Then, different Plasmodium falciparum strains were analyzed by immunofluorescence microscopy and western blots, using the epitope mapped antibodies. The performance of the immunoassays were compared to the identified epitopes, and validated by RNA sequencing.

 

In conclusion, these investigations describe multiplex methods to identify and characterize antibodies, their disease association and epitopes. Follow-up studies are needed to determine their potential use and clinical value.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2017. p. 89
Series
TRITA-BIO-Report, ISSN 1654-2312 ; 2017:19
Keywords
antibody, antigens, peptide, epitope mapping, autoimmunity, autoantibodies, microarrays
National Category
Medical Biotechnology
Research subject
Biotechnology
Identifiers
urn:nbn:se:kth:diva-213689 (URN)978-91-7729-499-3 (ISBN)
Public defence
2017-10-06, Air & Fire, Tomtebodavägen 23A, Solna, 10:00 (English)
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Supervisors
Note

QC 20170905

Available from: 2017-09-05 Created: 2017-09-04 Last updated: 2017-09-05Bibliographically approved

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Zandian, ArashForsström, BjörnSchwenk, Jochen M.Uhlén, MathiasNilsson, PeterAyoglu, Burcu

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Zandian, ArashForsström, BjörnHäggmark-Månberg, AnnaSchwenk, Jochen M.Uhlén, MathiasNilsson, PeterAyoglu, Burcu
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Proteomics and NanobiotechnologyScience for Life Laboratory, SciLifeLabAlbanova VinnExcellence Center for Protein Technology, ProNovaProteomics (closed 20130101)Nano Biotechnology (closed 20130101)
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Journal of Proteome Research
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