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. 2022 Nov 25;3(11):1300–1317. doi: 10.1038/s43018-022-00450-6

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© The Author(s) 2022, corrected publication 2022, 2024

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — a, Density plots of the antigen-presenting cluster from Fig. 2a (nonresponders n = 6 patients, responders n = 10 patients). b, Histogram representing the amount of CD86 expressed by cells within the antigen-presenting cluster. c, t-SNEs of CD45⁺ cells in a responder and two nonresponders. Blue represents TME samples taken before treatment, and pink represents TME samples taken after treatment. d, Left, representative VECTRA images of HLA-DR expression (left images with CK expression, right images without CK expression). Right, quantification of VECTRA images of HLA-DR expression on CK⁺ cells within the TME pre-treatment (mean ± s.e.m.) (nonresponders n = 4 patients, responders n = 12 patients). e, Representative VECTRA images of DLNs collected at time of surgery. DC–T cell interactions are highlighted with white arrows. f, Density of CD8⁺ T cells (CD3⁺CD8⁺) in the DLNs (nonresponders n = 3 patients, responders n = 3 patients). g, Density of activated CD8⁺ T cells (CD3⁺CD8⁺IFN-γ⁺) and replicating CD8⁺ T cells (CD3⁺CD8⁺Ki67⁺) in the DLNs (nonresponders n = 3 patients, responders n = 3 patients). h, Density of activated CD4⁺ T cells (CD3⁺CD8⁻Foxp3⁻IFN-γ⁺) in the DLNs (nonresponders n = 3 patients, responders n = 3 patients). i, Percentages of CD3⁺ cells that are CD8⁺ T cells, activated CD8⁺ T cells and activated CD4⁺ T cells (nonresponders n = 3 patients, responders n = 3 patients). j, Quantification of how many T cells were within 15 μm of APCs (CD3⁻HLA⁻DR⁺) (mean ± s.e.m.) (nonresponders n = 3 patients, responders n = 3 patients). k, Average of the top 5 TCR sequences pre- and post-treatment in the blood (nonresponders n = 3 patients, responders n = 5 patients). l, Scatterplot with annotations depicting clones with more than 8 transcripts after treatment in the TME and blood for patients 01-010 and 01-014. Red dots are clones significantly increased in the TME and blue dots are clones that were significantly increased in the blood. Dots along the y and x axes are clones not present in the TME or blood post-treatment, respectively. Statistical significance was determined using a two-tailed paired Student’s t-test. *P < 0.05, **P < 0.01, ***P < 0.001.

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