The Mutation-Associated Neoantigen Functional Expansion of Specific T Cells (MANAFEST) Assay: A Sensitive Platform for Monitoring Antitumor Immunity

Cancer Immunol Res. 2018 Aug;6(8):888-899. doi: 10.1158/2326-6066.CIR-18-0129. Epub 2018 Jun 12.


Mutation-associated neoantigens (MANA) are a target of antitumor T-cell immunity. Sensitive, simple, and standardized assays are needed to assess the repertoire of functional MANA-specific T cells in oncology. Assays analyzing in vitro cytokine production such as ELISpot and intracellular cytokine staining have been useful but have limited sensitivity in assessing tumor-specific T-cell responses and do not analyze antigen-specific T-cell repertoires. The FEST (Functional Expansion of Specific T cells) assay described herein integrates T-cell receptor sequencing of short-term, peptide-stimulated cultures with a bioinformatic platform to identify antigen-specific clonotypic amplifications. This assay can be adapted for all types of antigens, including MANAs via tumor exome-guided prediction of MANAs. Following in vitro identification by the MANAFEST assay, the MANA-specific CDR3 sequence can be used as a molecular barcode to detect and monitor the dynamics of these clonotypes in blood, tumor, and normal tissue of patients receiving immunotherapy. MANAFEST is compatible with high-throughput routine clinical and lab practices. Cancer Immunol Res; 6(8); 888-99. ©2018 AACR.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antigens, Neoplasm / immunology*
  • CD8-Positive T-Lymphocytes / immunology*
  • Carcinoma, Non-Small-Cell Lung / immunology
  • Carcinoma, Non-Small-Cell Lung / therapy
  • Cells, Cultured
  • Computational Biology / methods
  • Epitopes / immunology
  • Humans
  • Immunity, Cellular
  • Immunotherapy
  • Lung Neoplasms / immunology
  • Lung Neoplasms / therapy
  • Lymphocyte Activation / immunology
  • Monitoring, Immunologic / methods*
  • Mutation
  • Neoplasms / immunology*
  • Neoplasms / therapy*
  • Receptors, Antigen, T-Cell, alpha-beta / genetics


  • Antigens, Neoplasm
  • Epitopes
  • Receptors, Antigen, T-Cell, alpha-beta