{
  "abstract": "Background Understanding the mechanisms of effector T cell function is critical to the development of safe, effective and durable cell-based therapies. Owing to the highly complex and heterogeneous nature of both tumor and immune cells, researchers are increasingly using single-cell tools to characterize the genotypes, phenotypes and function (G2P2F) of their cell models. Current single-cell OMICs tools do not easily allow the measurement of G2P2F within the same cell. Those that do are very low scale and do not assess the number of cells needed to properly characterize the depth of heterogeneity and identify novel cell types.Methods We evaluated a novel live cell workflow that characterized activated engineered TCR-T cells via flow cytometry, imaging and sequencing at the single-cell level. By using optical cell barcodes to track live cell phenotype and function via flow cytometry and a specially designed optically-registered picowell chip to process these cells for single-cell genomic readout, we were able to obtain matched single-cell data across G2P2F. Expression of activation markers (CD69, CD25, CD137), exhaustion markers (PD1, TIM-3, LAG-3) in TCR T cells were profiled over time in the presence of cognate and non-specific antigen targets expressed by target tumor cells.Results We were able to differentiate specific and non-specific epitope reactivity in TCR-T cells, obtaining novel single-cell multi-omic data combining phenotype, function and transcriptomics for about ~10 4 cells per condition. TCR-T cells activated with their cognate antigen target exhibited higher induction of activation markers; higher cell killing activities and de novo upregulation of activation markers as well as exhaustion markers over time. We also identified differential expression of protein markers that were not concordant with transcriptional profiles, suggestive of post-translational regulation. Deeper analysis is underway to identify key biomarkers pivotal to effector function and T-cell exhaustion.Conclusions This study demonstrated the specific but highly heterogeneous nature of TCR-T cell function post-antigen activation. For the first time, we have demonstrated the ability to connect kinetic changes in phenotype and function within live cells with transcriptomic profiles, offering much more comprehensive analysis of underlying single-cell heterogeneity.",
  "authors": [
    {
      "affiliations": [
        "LASE Innovation, Waltham, MA, USA"
      ],
      "name": "Sheldon Kwok"
    },
    {
      "affiliations": [
        "Flexomics LLC, Waltham, MA, USA"
      ],
      "name": "Magali Soumillon"
    },
    {
      "affiliations": [
        "LASE Innovation, Waltham, MA, USA"
      ],
      "name": "Yulia Shulga"
    },
    {
      "affiliations": [
        "LASE Innovation, Waltham, MA, USA"
      ],
      "name": "Emane Rose Assita"
    },
    {
      "affiliations": [
        "Flexomics LLC, Waltham, MA, USA"
      ],
      "name": "Caroline Sartain"
    },
    {
      "affiliations": [
        "LASE Innovation, Waltham, MA, USA"
      ],
      "name": "Sarah Forward"
    },
    {
      "affiliations": [
        "Flexomics LLC, Waltham, MA, USA"
      ],
      "name": "Amara Alexander"
    },
    {
      "affiliations": [
        "LASE Innovation, Waltham, MA, USA"
      ],
      "name": "Zoe Lin"
    },
    {
      "affiliations": [
        "LASE Innovation, Waltham, MA, USA"
      ],
      "name": "Trevor Brown"
    },
    {
      "affiliations": [
        "Talon Biomarkers, Whippany, NJ, USA"
      ],
      "name": "Pratip Chattopadhyay"
    }
  ],
  "title": "138 Single-cell multi-omic profiling of engineered TCR cells: phenotypes, genotypes and function from the same cells",
  "uid": "a7a0171d-b548-5708-9aaf-8a949d32872c"
}