TGFβ suppresses CD8+ T cell expression of CXCR3 and tumor trafficking

Transforming growth factor beta (TGFβ) is a multipotent immunosuppressive cytokine. TGFβ excludes immune cells from tumors, and TGFβ inhibition improves the efficacy of cytotoxic and immune therapies. Using preclinical colorectal cancer models in cell type-conditional TGFβ receptor I (ALK5) knockout mice, we interrogate this mechanism. Tumor growth delay and radiation response are unchanged in animals with Treg or macrophage-specific ALK5 deletion. However, CD8αCre-ALK5flox/flox (ALK5ΔCD8) mice reject tumors in high proportions, dependent on CD8+ T cells. ALK5ΔCD8 mice have more tumor-infiltrating effector CD8+ T cells, with more cytotoxic capacity. ALK5-deficient CD8+ T cells exhibit increased CXCR3 expression and enhanced migration towards CXCL10. TGFβ reduces CXCR3 expression, and increases binding of Smad2 to the CXCR3 promoter. In vivo CXCR3 blockade partially abrogates the survival advantage of an ALK5ΔCD8 host. These data demonstrate a mechanism of TGFβ immunosuppression through inhibition of CXCR3 in CD8+ T cells, thereby limiting their trafficking into tumors.


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Policy information about availability of data All manuscripts must include a data availability statement. This statement should provide the following information, where applicable: -Accession codes, unique identifiers, or web links for publicly available datasets -A list of figures that have associated raw data -A description of any restrictions on data availability Kristina Young 02/17/2020 Flow cytometry data was analyzed using commercially available FlowJo software. Data analysis was performed using Graphpad Prism 7.0 software. Graphpad Prism 7.0 software was used to construct all graphs and calculate statistical significance. FlowJo software was used for FACS analysis and to generate tSNE plots.
All data published is representative of experiments and replicates. RNAseq data was mined from TCGA -colorectal PanCancer Atlas data set on the cBioPortal for cancer genomics (cbioportal.org). SMAD2 and CXCR3 normalized mRNA expression levels were compared by linear regression analysis. Animal models, reagents, and software used are publicly available. There are no restrictions on our data.

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October 2018 Field-specific reporting Please select the one below that is the best fit for your research. If you are not sure, read the appropriate sections before making your selection. The efficacy of therapy on time-to-failure will be determined by Kaplan-Meier survival curves compared using a log-rank test with significance of p<0.05. 8 mice/group provides 80% power to detect a difference in survival at four weeks post treatment of 50% vs. 84%, and difference in median survival of 1 week.
Data was excluded from qPCR if >30 cycles were performed without amplification signal, or if >3 standard deviations from the mean with 3 or more replicates.
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