Single-cell cytokine profiling of tumor-infiltrating T cells to measure patient responses to anti-PD-1 therapy.

49Background: Functional alteration of tumor-infiltrating T lymphocytes (TILs) may serve as a predictor for clinical outcome in cancer patients receiving immunotherapy.To evaluate TILs function unleashed by anti-PD-1 blocking, we employed a single-cell technology integrated with automated bioinformatics to simultaneously measure 17 cytokines secreted by single TILs, permitting the full spectrum delineation of anti-tumor T cell functions in patients with metastatic melanoma. Methods: TILs were enriched from biopsied melanoma tissues digested with enzymes, stimulated with anti-CD3 at 37°C, 5% CO2 for 24 hrs and loaded into a single-cell barcode chip (SCBC) containing ~12000 microchambers. Each chamber (~1.2 nl) was pre-patterned with a complete copy of a 17-plex antibody array. Cells on the SCBC were imaged and incubated for 16 hrs at 37°C, 5% CO2; single-cell cytokine signals were captured with a microarray scanner. The polyfunctional expression (2+ cytokines per cell) of single TILs was evaluated across 4...