ABSTRACT
Neoantigens are peptides derived from non-synonymous mutations presented by human leukocyte antigens (HLAs), which are recognized by antitumour T cells1-14. The large HLA allele diversity and limiting clinical samples have restricted the study of the landscape of neoantigen-targeted T cell responses in patients over their treatment course. Here we applied recently developed technologies15-17 to capture neoantigen-specific T cells from blood and tumours from patients with metastatic melanoma with or without response to anti-programmed death receptor 1 (PD-1) immunotherapy. We generated personalized libraries of neoantigen-HLA capture reagents to single-cell isolate the T cells and clone their T cell receptors (neoTCRs). Multiple T cells with different neoTCR sequences (T cell clonotypes) recognized a limited number of mutations in samples from seven patients with long-lasting clinical responses. These neoTCR clonotypes were recurrently detected over time in the blood and tumour. Samples from four patients with no response to anti-PD-1 also demonstrated neoantigen-specific T cell responses in the blood and tumour to a restricted number of mutations with lower TCR polyclonality and were not recurrently detected in sequential samples. Reconstitution of the neoTCRs in donor T cells using non-viral CRISPR-Cas9 gene editing demonstrated specific recognition and cytotoxicity to patient-matched melanoma cell lines. Thus, effective anti-PD-1 immunotherapy is associated with the presence of polyclonal CD8+ T cells in the tumour and blood specific for a limited number of immunodominant mutations, which are recurrently recognized over time.
Subject(s)
Antigens, Neoplasm , CD8-Positive T-Lymphocytes , Immune Checkpoint Inhibitors , Immunotherapy , Melanoma , Humans , Antigens, Neoplasm/immunology , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , Melanoma/drug therapy , Melanoma/genetics , Melanoma/immunology , Melanoma/pathology , Receptors, Antigen, T-Cell/immunology , Receptors, Antigen, T-Cell/metabolism , Immune Checkpoint Inhibitors/pharmacology , Immune Checkpoint Inhibitors/therapeutic use , HLA Antigens/immunology , Neoplasm Metastasis , Precision Medicine , Gene Editing , CRISPR-Cas Systems , MutationABSTRACT
We used optical tweezers to tie individual DNA molecules in knots. Although these knots become highly localized under tension, they remain surprisingly mobile and undergo thermal diffusion with classical random walk statistics. The diffusion constants of knots with different complexities correlate with theoretical calculations of knot sizes. We show that this correlation can be explained by a simple hydrodynamical model of "self-reptation" of the knot along a polymer.
