Development of an HLA Epitope Database

Rene Duquesnoy

While it is generally accepted that HLA antibodies represent significant risk factors for transplant rejection and graft failure, it has become apparent that such antibodies are specific for epitopes rather than HLA antigens. A distinction between HLA epitopes and antigens is important not only for the determination of mismatch acceptability for sensitized patients but also for a better understanding of the humoral immune response to an HLA mismatch.

This project addresses the determination of HLA class I and class II epitopes recognized by antibodies and what notation system should be used to describe such epitopes.  Because there is so much detailed information about HLA molecular structure and amino acid sequences, it seems likely that HLA epitopes can be defined structurally.  However, their true existence can only be proven with specific antibodies. There is already a considerable literature about antibody-reactive HLA epitopes but there continues to be a need to verify them experimentally and to define new epitopes.

Participants in this project will be identify in their laboratories antibody reactivity patterns of sera which may verify less well defined epitopes or recognize newly defined epitopes. A worldwide effort will likely generate new insights into HLA epitope structure. Altogether the data from participants together with already existing information will be incorporated in an epitope database that will be continually upgraded. 

What methods should be used to determine antibody-reactive epitopes? It seems that antibody binding assays such as Luminex with single allele panels must be used as a minimum but other methods such as Complement-dependent lymphocytotoxicity will be considered.  The “monospecificity” of an antibody seems most relevant to the identification of an epitope.   Human monoclonals are excellent sources especially because circulating antibody-producing B-cells have been cultured from sensitized patients and their supernatants have been tested. Another useful approach is to do absorption-elution studies with informative single alleles.

The determination of a specific epitope depends on the analysis of the antibody reactivity pattern with the HLA panel.  Information about the immunizing event, such as the HLA differences between immunizer and antibody producer enhances the antibody analysis.
It seems likely that epitopes will be described by amino acid polymorphisms in molecular structure models.  HLAMatchmaker and possibly other algorithms that consider such polymorphisms will be made available to analyze antibody reactivity patterns

This project will have an international steering committee:

Rene Duquesnoy (Pittsburgh, PA) Chair, Domenico Adorno (L’Aquila, Italy), Ivan Balasz (Stamford, CT, USA), Frans Claas (Leiden, The Netherlands), Nadim El-Awar (Los Angeles, CA, USA), Marcelo Fernandez-Vina (Houston, TX, USA), Nils Lachmann (Berlin, Germany), Derek Middleton (Liverpool, UK), Arend Mulder (Leiden, The Netherlands), Craig Taylor (Cambridge, UK), Robert Vaughan (London, UK)and Cristina von Glehn (Curitiba, Brazil)

This committee has the following tasks:

1. Determine criteria for the experimental verification of epitopes reacting with informative antibodies.  Address the question of clinically relevant versus irrelevant epitopes.
2. Develop a notation system for Class I and Class II epitope repertoires
3. Verify experimentally defined epitopes submitted by workshop participants
4. Develop a website based HLA epitope database

Please note that this project addresses only antibody-reactive epitopes, not cellular epitopes defined by T-cells.

How to participate in this project?

Participants in this project will look in their laboratories for cases with antibody reactivity patterns that might lead to a determination of new or less well defined HLA Class I, Class II and MICA epitopes. These antibodies would be produced by people sensitized by a transplant, pregnancy or transfusion although mouse monoclonals might also be considered. Antibody sources would be sera and supernatants of antibody-producing cells including EBV-transformed cells.

Participation consists of three steps.

1. Submit information about an antibody reactivity pattern having the potential of identifying a new epitope. Participant provides MFI results with Luminex single allele panels (csv files) together with information about the sensitizing event including HLA types of antibody producer and preferably also immunizer.  The Committee will do a preliminary epitope analysis of the antibody reactivity patterns and depending on the outcome, then requests a specimen for further testing.
2. Submitted specimens will be tested in designated laboratories (which may include Participant) by a standard protocol including parallel testing with Luminex single allele kits from different vendors and ruling out artifacts of the assays. Other antibody detection methods using cell membrane bound HLA panels would be considered. Antibody reactivity has to be monospecific and specimens may be absorbed with informative alleles (based on HLA types of antibody producer and immunizer) and the eluates with be tested for antibody. The reactivity patterns will be analyzed for specificity against new epitopes with distinct amino acid configurations.
3. The committee will review the data of all tested specimens and provide a notation and a structural description of new epitopes to be added to the database posted on the website. Participants will receive detailed reports about the epitopes identified by their antibody specimens for consideration of publication.

This project will be start early 2011 when more details will be posted on the website.

Contacts:
For more information, please contact.

Rene J. Duquesnoy, Ph.D.
Professor Emeritus of Pathology
University of Pittsburgh Medical Center
Room 5712 - PUH/South Tower
200 Lothrop Street
Pittsburgh, PA  15213
Phone: 1-412-647-6148
Mobile phone: 1-412-860-8083
Fax: 1-412-647-1755

Email: Duquesnoyr [at] upmc [dot] edu

Download a PDF of the EFI 2011 Presentation - download PDF file.

Download a PDF of the ASHI 2011 Presentation - download PDF file.

Nov 10, 2011 Posted by Admin