I , J ) 10 - 12 days following MC38 tumour inoculation and treatment with anti - PD - 1 antibody or isotype control , CD4cre Ptpn6 / 11wt / wt and CD4cre Ptpn6fl / fl / Ptpn11fl / flWbm mice were sacrificed , and TILs analysed . Graphs depict frequencies of CD8 + T cells ( gated as CD45 + TCRβ + CD8 + ) expressing IFN - γ and TNFα upon re - stimulation ( G ) and percentages of CD8 + T cells ( H ) .

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I, J) 10-12 days following MC38 tumour inoculation and treatment with anti-PD-1 antibody or isotype control, CD4cre Ptpn6/11wt/wt and CD4cre Ptpn6fl/fl/Ptpn11fl/flWbm mice were sacrificed, and TILs analysed. Graphs depict frequencies of CD8+ T cells (gated as CD45+ TCRβ+ CD8+) expressing IFN-γ and TNFα upon re-stimulation (G) and percentages of CD8+ T cells (H).


B ) Ptpn6 and Ptpn11 copy number , as estimated through high resolution mass spectrometry in GzmBcre Ptpn6 / 11fl / fl and control CTLs .

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B) Ptpn6 and Ptpn11 copy number, as estimated through high resolution mass spectrometry in GzmBcre Ptpn6/11fl/fl and control CTLs.


C and D ) GzmBcre R26R - YFP reporter mice were injected subcutaneously with MC38 cells and analysed after 11 days . Graphs show the percentage of YFP + cells among CD45 + CD8 + TILs ( C ) and the percentage of PD - 1 + cells among CD45 + CD8 + YFP + TILs ( D ) . Results depict n = 4 mice ( C and D ) .

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C and D) GzmBcre R26R-YFP reporter mice were injected subcutaneously with MC38 cells and analysed after 11 days. Graphs show the percentage of YFP+ cells among CD45+ CD8+ TILs (C) and the percentage of PD-1+ cells among CD45+ CD8+ YFP+ TILs (D). Results depict n = 4 mice (C and D).


GzmBcre Ptpn6 / 11fl / fl and control Ptpn6 / 11fl / fl mice were subcutaneously injected with MC38 cells . Survival curves and statistical comparisons between different groups are shown ( E ) .

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GzmBcre Ptpn6/11fl/fl and control Ptpn6/11fl/fl mice were subcutaneously injected with MC38 cells. Survival curves and statistical comparisons between different groups are shown (E).


The tumour start depicts the time ( days ) following tumour engraftment after which the tumour is palpable , while tumour endpoint depicts the time ( days ) from tumour start until the reaching the maximal allowed tumour size ( F ) .

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The tumour start depicts the time (days) following tumour engraftment after which the tumour is palpable, while tumour endpoint depicts the time (days) from tumour start until the reaching the maximal allowed tumour size (F).


C ) Numbers of CTLs ( gated on CD8 + T cells ) from Ptpn6 / 11fl / fl or GzmBcre Ptpn6 / 11fl / fl mice were counted daily by flow cytometry with the addition of DAPI to monitor dead cells .

C) Numbers of CTLs (gated on CD8+ T cells) from Ptpn6/11fl/fl or GzmBcre Ptpn6/11fl/fl mice were counted daily by flow cytometry with the addition of DAPI to monitor dead cells.


D ) Quantitative high - resolution mass spectrometry was used to resolve the proteome of Ptpn6 / 11fl / fl and GzmBcre Ptpn6 / 11fl / fl CTLs ( day 7 ) . The plot depicts protein copy number in control and GzmBcre Ptpn6 / 11fl / fl cells . Significantly altered proteins are indicated in red ; selected examples are annotated . Results are based on n = 3 biological replicates and two - tailed , unequal - variance t - test on log10 transformed copy number per cell values was used to compare differences between experimental groups .

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D) Quantitative high-resolution mass spectrometry was used to resolve the proteome of Ptpn6/11fl/fl and GzmBcre Ptpn6/11fl/fl CTLs (day 7). The plot depicts protein copy number in control and GzmBcre Ptpn6/11fl/fl cells. Significantly altered proteins are indicated in red; selected examples are annotated. Results are based on n = 3 biological replicates and two-tailed, unequal-variance t-test on log10 transformed copy number per cell values was used to compare differences between experimental groups.


G ) Graph illustrates the percentage of DAPI + dead cells of Ptpn6 / 11fl / fl and GzmBcre Ptpn6 / 11fl / fl CTLs ( gated on CD8 + T cells ) at the indicated days as measured by flow cytometry .

G) Graph illustrates the percentage of DAPI+ dead cells of Ptpn6/11fl/fl and GzmBcre Ptpn6/11fl/fl CTLs (gated on CD8+ T cells) at the indicated days as measured by flow cytometry.


H ) Protein copy number of Tnfrsf1b and Mcl - 1 in GzmB cre Ptpn6 / 11fl / fl and control CTLs , as estimated through high resolution mass spectrometry .

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H) Protein copy number of Tnfrsf1b and Mcl-1 in GzmB cre Ptpn6/11fl/fl and control CTLs, as estimated through high resolution mass spectrometry.