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High-Throughput Discovery of T Cell Epitopes Using Cytokine Capture

A high-throughput assay identifies epitopes that activate T cells by capturing cytokines on the surface of barcoded antigen-presenting cells (APCs).

  • Diagnostics

Project Overview

  • Variation in human leukocyte (HLA) and T cell receptor (TCR) genes is associated with risk of infection and autoimmunity and can influence patient survival following cancer immunotherapy.
  • Identification of specific complexes between HLA molecules, epitopes, and TCRs can thus provide fundamental information about disease pathogenesis and lead to the generation of T cell-based treatments such as TCR-based cellular therapies. However, there remains technological barriers to identifying which HLA, epitopes, and TCRs lead to T cell activation. 
  • Researchers at Dana-Farber have developed a high-throughput assay for identifying epitopes that activate T cells by capturing cytokines on the surface of barcoded antigen-presenting‑ cells (APCs).
  • Dana-Farber Cancer Institute is looking for the right partner with an interest in licensing these assets for further development. 

Technology

Traditional epitope identification technologies such as ELISPOT (enzyme-linked immunospot) or ICS (intracellular cytokine staining) are widely used to detect HLA epitope-TCR complexes. These methods rely on the capture of T cell activation dependent cytokines and have had broad applications such as identifying epitopes targeted by CD8 or CD4 T cells in disease such as cancer. However, these assays are particularly limited in assessing large epitope candidate sets due to the high costs of peptide synthesis. Development of a high-throughput epitope identification technology that combines both HLA and peptide diversity may enable identification of HLA-epitopeTCR complexes whose identification is intractable using existing assays. 

Researchers at Dana-Farber have developed a high-throughput assay for identifying epitopes that activate T cells by capturing cytokines on the surface of barcoded antigen-presenting‑ cells (APCs). This is a strategy in which a signal is generated by cytokine capture – similar to traditional functional assays – but in which the anti-cytokine antibodies are directly expressed by the APCs along with an encoded peptide and multiple HLA class I or II genes. After APC/T cell co-culture, APCs coated with cytokine are sorted and the encoded epitopes are read out by next-generation sequencing (NGS). 

This technology allows pooled screening of thousands of encoded peptides to enable epitope discovery for orphan TCRs. 

Further Details: 

Team Members: Matthew L. Meyerson, MD, PhD, Mark Lee, MD, PhD

Applications

  • This method has the sensitivity to detect a single epitope from a pool of thousands of peptide-encoding oligonucleotides and can identify epitopes for orphan CD8+ and CD4+ TCRs. 
  • The development of engineered APCs that link T cell-secreted cytokine signal with DNA that encodes the presented peptide allows oligonucleotide pools with complexities in at least the thousands to be filtered for T cell epitopes. 
  • Unlike other DNA-based screenings, signal generation occurs through an unmodified HLA molecule, and the signal is common to both class I- and II-restricted T cells. 
  • Inexpensive scaling to many thousands of encoded peptides. 
  • Encoded anti-cytokine antibody that can be interchanged, allowing selection of T cell subsets through their differential cytokine secretion. 
  • Robust epitope selection strategy, which could help greatly expand the number of known class II epitopes/TCR pairings. 

Opportunities

Dana-Farber Cancer Institute is looking for the right partner with an interest in licensing these assets for further development. 

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Team Members