Transforming growth factor beta orchestrates PD-L1 enrichment in tumor-derived exosomes and mediates CD8 T-cell dysfunction regulating early phosphorylation of TCR signalome in breast cancer.
Tumor cells promote immune evasion through upregulation of programmed death-ligand 1 (PD-L1) that binds with programmed cell death protein 1 (PD1) on cytotoxic T cells and promote dysfunction. Though therapeutic efficacy of anti-PD1 antibody has remarkable effects on different type of cancers it is less effective in breast cancer (BC). Hence, more details understanding of PD-L1-mediated immune evasion is necessary. Here, we report BC cells secrete extracellular vesicles in form of exosomes carry PD-L1 and are highly immunosuppressive. Transforming growth factor beta (TGF-β) present in tumor microenvironment orchestrates BC cell secreted exosomal PD-L1 load. Circulating exosomal PD-L1 content is highly correlated with tumor TGF-β level. The later also found to be significantly associated with CD8+CD39+, CD8+PD1+ T-cell phenotype. Recombinant TGF-β1 dose dependently induces PD-L1 expression in Texos in vitro and blocking of TGF-β dimmed exosomal PD-L1 level. PD-L1 knocked down exosomes failed to suppress effector activity of activated CD8 T cells like tumor exosomes. While understanding its effect on T-cell receptor signaling, we found siPD-L1 exosomes failed to block phosphorylation of src family proteins, linker for activation of T cells and phosphoinositide phospholipase Cγ of CD8 T cells more than PD-L1 exosomes. In vivo inhibition of exosome release and TGF-β synergistically attenuates tumor burden by promoting Granzyme and interferon gamma release in tumor tissue depicting rejuvenation of exhausted T cells. Thus, we establish TGF-β as a promoter of exosomal PD-L1 and unveil a mechanism that tumor cells follow to promote CD8 T-cell dysfunction.
Chatterjee S
，Chatterjee A
，Jana S
，Dey S
，Roy H
，Das MK
，Alam J
，Adhikary A
，Chowdhury A
，Biswas A
，Manna D
，Bhattacharyya A
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Inhibition of T-cell-mediated immune response via the PD-1/ PD-L1 axis in cholangiocarcinoma cells.
Cholangiocarcinoma (CCA) is a malignant biliary tract epithelium tumor. The programmed death-1 (PD-1)/programmed receptor-ligand 1 (PD-L1) signaling pathway has been implicated as an immune escape mechanism in several cancers. The present study aimed to assess the expression of PD-L1 on human CCA cell lines and its potential role in suppressing CD8+ T- cell function. A panel of intrahepatic CCA cell lines was evaluated for immune regulatory checkpoint ligands and inflammation markers. Effects of pro-inflammatory cytokine, interferon gamma (IFN-γ), on the expression of immune regulatory checkpoint ligands and inflammation markers were determined. The PD-L1 function was measured by co-culturing CCA cells with lymphocytes. Most of the selected Thai CCA cell lines, including HuCCA-1, RMCCA-1, KKU-100, and KKU-213, expressed higher PD-L1 than normal cholangiocyte MMNK-1 and ANK-1 cells. Both PD-L1 and cyclooxygenase-2 (COX-2) expressions were highest in HuCCA-1 cells. A 48 h treatment with IFN-γ increased the expression of PD-L1 and COX-2 in CCA cells. The expression of CTLA-4 ligands, including H7-1 and H7-2, did not change after IFN-γ treatment. Rofecoxib, a specific COX-2 inhibitor, mitigated IFN-γ-induced PD-L1 expression. After 48 h co-incubation, CD8+ T-cell apoptosis was increased as compared to the control group. Pretreatment of CCA cells with IFN-γ further increased CD8+ T-cell apoptosis. Pembrolizumab, an anti-PD-1 antibody, mitigated CCA cell escape phenomenon. The inhibition of T-cell-mediated immune response via the PD-L1/PD-1 axis are evidenced in intrahepatic CCA. Immunotherapy with checkpoint inhibitor offers a potentially therapeutic strategy for CCA patients; however, further in vivo and clinical studies are required.
Suriyo T
，Fuangthong M
，Artpradit C
，Ungtrakul T
，Sricharunrat T
，Taha F
，Satayavivad J
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A radiomics approach to assess tumour-infiltrating CD8 cells and response to anti-PD-1 or anti-PD-L1 immunotherapy: an imaging biomarker, retrospective multicohort study.
Because responses of patients with cancer to immunotherapy can vary in success, innovative predictors of response to treatment are urgently needed to improve treatment outcomes. We aimed to develop and independently validate a radiomics-based biomarker of tumour-infiltrating CD8 cells in patients included in phase 1 trials of anti-programmed cell death protein (PD)-1 or anti-programmed cell death ligand 1 (PD-L1) monotherapy. We also aimed to evaluate the association between the biomarker, and tumour immune phenotype and clinical outcomes of these patients.
In this retrospective multicohort study, we used four independent cohorts of patients with advanced solid tumours to develop and validate a radiomic signature predictive of immunotherapy response by combining contrast-enhanced CT images and RNA-seq genomic data from tumour biopsies to assess CD8 cell tumour infiltration. To develop the radiomic signature of CD8 cells, we used the CT images and RNA sequencing data of 135 patients with advanced solid malignant tumours who had been enrolled into the MOSCATO trial between May 1, 2012, and March 31, 2016, in France (training set). The genomic data, which are based on the CD8B gene, were used to estimate the abundance of CD8 cells in the samples and data were then aligned with the images to generate the radiomic signatures. The concordance of the radiomic signature (primary endpoint) was validated in a Cancer Genome Atlas [TGCA] database dataset including 119 patients who had available baseline preoperative imaging data and corresponding transcriptomic data on June 30, 2017. From 84 input variables used for the machine-learning method (78 radiomic features, five location variables, and one technical variable), a radiomics-based predictor of the CD8 cell expression signature was built by use of machine learning (elastic-net regularised regression method). Two other independent cohorts of patients with advanced solid tumours were used to evaluate this predictor. The immune phenotype internal cohort (n=100), were randomly selected from the Gustave Roussy Cancer Campus database of patient medical records based on previously described, extreme tumour-immune phenotypes: immune-inflamed (with dense CD8 cell infiltration) or immune-desert (with low CD8 cell infiltration), irrespective of treatment delivered; these data were used to analyse the correlation of the immune phenotype with this biomarker. Finally, the immunotherapy-treated dataset (n=137) of patients recruited from Dec 1, 2011, to Jan 31, 2014, at the Gustave Roussy Cancer Campus, who had been treated with anti-PD-1 and anti-PD-L1 monotherapy in phase 1 trials, was used to assess the predictive value of this biomarker in terms of clinical outcome.
We developed a radiomic signature for CD8 cells that included eight variables, which was validated with the gene expression signature of CD8 cells in the TCGA dataset (area under the curve [AUC]=0·67; 95% CI 0·57-0·77; p=0·0019). In the cohort with assumed immune phenotypes, the signature was also able to discriminate inflamed tumours from immune-desert tumours (0·76; 0·66-0·86; p<0·0001). In patients treated with anti-PD-1 and PD-L1, a high baseline radiomic score (relative to the median) was associated with a higher proportion of patients who achieved an objective response at 3 months (vs those with progressive disease or stable disease; p=0·049) and a higher proportion of patients who had an objective response (vs those with progressive disease or stable disease; p=0·025) or stable disease (vs those with progressive disease; p=0·013) at 6 months. A high baseline radiomic score was also associated with improved overall survival in univariate (median overall survival 24·3 months in the high radiomic score group, 95% CI 18·63-42·1; vs 11·5 months in the low radiomic score group, 7·98-15·6; hazard ratio 0·58, 95% CI 0·39-0·87; p=0·0081) and multivariate analyses (0·52, 0·35-0·79; p=0·0022).
The radiomic signature of CD8 cells was validated in three independent cohorts. This imaging predictor provided a promising way to predict the immune phenotype of tumours and to infer clinical outcomes for patients with cancer who had been treated with anti-PD-1 and PD-L1. Our imaging biomarker could be useful in estimating CD8 cell count and predicting clinical outcomes of patients treated with immunotherapy, when validated by further prospective randomised trials.
Fondation pour la Recherche Médicale, and SIRIC-SOCRATE 2.0, French Society of Radiation Oncology.
Sun R
，Limkin EJ
，Vakalopoulou M
，Dercle L
，Champiat S
，Han SR
，Verlingue L
，Brandao D
，Lancia A
，Ammari S
，Hollebecque A
，Scoazec JY
，Marabelle A
，Massard C
，Soria JC
，Robert C
，Paragios N
，Deutsch E
，Ferté C
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ResearchGate
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