טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentFridman Anna
SubjectStudy of MHC Class II Epitope Presentation with
Antibodies that Mimic TCR Specificity
DepartmentDepartment of Biology
Supervisor Professor Yoram Reiter
Full Thesis textFull thesis text - English Version


Abstract

MHC class II (MHC-II) molecules are expressed on antigen-presenting cells (APCs) and present peptides derived from extracellular proteins to CD4 T cells. Binding of T cells to MHC via T cell receptor (TCR) initiates signaling cascade resulting in antigen-specific activation of T cell clones. In order to study MHC-II T cell epitopes one can use antibodies that mimic CD4 TCR specificity, while binding MHC-II/peptide complexes. These antibodies, termed T-Cell Receptor-Like (TCR-like) antibodies, combine two arms of the immune system: high-affinity soluble antibody molecules endowed with a fine specificity of T cells receptor.

Recently, novel TCR-like antibodies against autoreactive T-cell epitopes associated with Type 1 diabetes and Multiple Sclerosis autoimmune diseases were isolated in our laboratory by phage display approach. These TCR-like antibodies were characterized and found to be of high significance for two major research lines: (i) for basic molecular studies as a tool to investigate auto-antigen presentation during autoimmune inflammatory processes with the ability to detect and image APCs that present the auto-antigen. (ii) For translational-clinical applications, where these antibodies were shown to inhibit/block the proliferation of autoreactive T cells in-vitro and in-vivo by binding to the disease associated autoantigen with stronger affinity compared to the autoreactive TCRs.

The goal of this study was to broaden our TCR-like antibody approach in order to study MHC-II epitope presentation in two experimental models associated with autoimmunity and immunity to viral infection. These models enabled us to explore and develop novel antigen-specific research tools to study molecular mechanisms related to antigen presentation as well as novel antigen-specific therapeutic strategies.

For the autoimmune model we focused on Rheumatoid Arthritis, a T cell mediated autoimmune disease of the joints. This disease is triggered by autoantigens presentation (e.g. Type II collagen (CII)) on MHC-II molecules by APCs. While presented by HLA-DR4 molecule, residues 261-273 of CII represent an immunodominant T cell epitope.

For the viral model we focused on Influenza infection. After vaccination with inactivated Influenza virus from Hemagglutinin (HA) 3 subtype, one can find elevated frequency of CD4 T cells directed towards immunodominant HLA-DR4/HA-306-318 epitope in immunized HLA-DR4 subjects. Therefore, TCR-like antibodies against HLA-DR4/HA-306-318 viral epitope can be of high significance for the study of antigen presentation on APCs during a natural course of Influenza infection.

By screening of phage display library we isolated TCR-like antibodies against HLA-DR4/CII-261-273 and HLA-DR4/HA-306-318 recombinant complexes. These antibodies, termed P4A11 and H1, respectively, were able to bind native DR4/CII-261-273 or HLA-DR4/HA-306-318 epitopes as presented by peptide-pulsed DR4 APCs. Most significant, P4A11 and H1 inhibited the recognition between CII-261-273, or HA-306-318, pulsed DR4 APCs and their cognate T cell hybridoma and prevent their activation.

In addition, by using these TCR-like antibodies we were able to detect APCs presenting the HLA-DR4/CII-261-273 or HLA-DR4/HA-306-318 epitopes in splenocytes derived from CII or HA immunized DR4 transgenic mice.

In this study we demonstrated a proof of concept for the utility of our TCR-like antibodies for translational-clinical applications and for basic studies related to antigen presentation, as a tool for direct visualization of APCs.