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. 2022 Nov 2;12:18509. doi: 10.1038/s41598-022-23016-z

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© The Author(s) 2022

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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/.

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SC79 therapy decreases Treg infiltration with an increase in IFN-γ expression on CD4 Treg TILs. (a) Heat map derived from FlowSOM/UMAP analysis of CD4+ TILs (n = 3). (b) Histogram showing the per cent of cells in each cluster as seen in (c) involving tSNE analysis of TILs. c, tSNE analysis of TILs. (d) Analytical FACS profiles showing the frequency FoxP3 x CD25 expression (left panel). Histogram showing the numbers of CD25^hi Tregs in the TIL population (right panel) (n = 3). (e) Analytical FACS profiles showing the frequency of FoxP3 x ICOS expression, Histogram showing the numbers of ICOS^hi Tregs (right panel) (n = 3). (f) FACS profiles showing the frequency of CD25 x ICOS cells in the TIL population. Histogram showing the numbers of CD25^hi/ICOS^hi Tregs (right panel). (g) Profiles showing the pSTAT5 x CD25 expression in the TIL population. Histogram showing the numbers of pSTAT5^hi x CD25^hi Tregs (right panel) (n = 3). (h) Profiles showing the frequency of FoxP3+ cells expressing IFN-γ in TIL populations. responder and non-responder tumors (n = 3). Analysis of Treg expressing IFN-γ frequency from CTL (n = 4), SC79 R (n = 3) and SC79 NR (n = 3) TILs, i, Percent of IFN-γ Tregs in the TIL populations. The percentage increased in responder but not non-responder tumors. j, Fold increase of Treg expressing IFN-γ from TILs after 48 h of in vitro activation with SC79 (n = 3). (k) SC79 reverses the ability of T-cells to suppress anti-CD3 proliferation as measured by an increase in the presence of SC79 after 72 h of culture (n = 3). Results from the figure (a) to (g) were obtained after CyTOF acquisition from a pool of 3 tumors in each group. For statistical analyses, parametric unpaired t-tests and one-way ANOVA were used. *p < 0.05, **p < 0.01. The R package CATALYST v1.12.1 was used to perform unsupervised clustering, and generate the heatmaps and tSNE plots. https://bioconductor.org/packages/release/bioc/html/CATALYST.html. https://f1000research.com/articles/6-748.