Identification of shared TCR sequences from T cells in human breast cancer using emulsion RT-PCR

Proc Natl Acad Sci U S A. 2016 Jul 19;113(29):8272-7. doi: 10.1073/pnas.1606994113. Epub 2016 Jun 15.


Infiltration of T cells in breast tumors correlates with improved survival of patients with breast cancer, despite relatively few mutations in these tumors. To determine if T-cell specificity can be harnessed to augment immunotherapies of breast cancer, we sought to identify the alpha-beta paired T-cell receptors (TCRs) of tumor-infiltrating lymphocytes shared between multiple patients. Because TCRs function as heterodimeric proteins, we used an emulsion-based RT-PCR assay to link and amplify TCR pairs. Using this assay on engineered T-cell hybridomas, we observed ∼85% accurate pairing fidelity, although TCR recovery frequency varied. When we applied this technique to patient samples, we found that for any given TCR pair, the dominant alpha- or beta-binding partner comprised ∼90% of the total binding partners. Analysis of TCR sequences from primary tumors showed about fourfold more overlap in tumor-involved relative to tumor-free sentinel lymph nodes. Additionally, comparison of sequences from both tumors of a patient with bilateral breast cancer showed 10% overlap. Finally, we identified a panel of unique TCRs shared between patients' tumors and peripheral blood that were not found in the peripheral blood of controls. These TCRs encoded a range of V, J, and complementarity determining region 3 (CDR3) sequences on the alpha-chain, and displayed restricted V-beta use. The nucleotides encoding these shared TCR CDR3s varied, suggesting immune selection of this response. Harnessing these T cells may provide practical strategies to improve the shared antigen-specific response to breast cancer.

Keywords: T-cell receptors; T-cell repertoire profiling; breast cancer; emulsion RT-PCR; high-throughput sequencing.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Base Sequence
  • Breast Neoplasms / genetics*
  • Cell Line
  • Emulsions
  • Female
  • Humans
  • Polymerase Chain Reaction / methods
  • Receptors, Antigen, T-Cell / genetics*
  • T-Lymphocytes / metabolism*


  • Emulsions
  • Receptors, Antigen, T-Cell