| 1. |
- Winerdal, Malin E., et al.
(författare)
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Urinary Bladder Cancer Tregs Suppress MMP2 and Potentially Regulate Invasiveness
- 2018
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Ingår i: CANCER IMMUNOLOGY RESEARCH. - : American Association for Cancer Research (AACR). - 2326-6066 .- 2326-6074. ; 6:5, s. 528-538
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Tidskriftsartikel (refereegranskat)abstract
- Regulatory T cells (Treg) have long been considered one-sided suppressors of antitumor immune responses and hence associated with poor patient outcome in cancer. However, evidence is mounting of a paradoxical positive prognostic effect of Tregs on certain malignancies, including urinary bladder cancer (UBC). This discrepancy has partly been attributed to the shear misidentification of Tregs, but also to the inflammatory profile of the tumor. Our aim was to determine whether tumor-infiltrating Forkhead box P3+ (FOXP3+) cells confer a stable Treg phenotype and to investigate putative beneficial Treg functions, focusing on tumor-promoting inflammatory pathways in UBC. Patients (n = 52) with suspected UBC were prospectively included. We show, by using a broad range of analytical approaches, that tumor-infiltrating CD4+FOXP3+ T cells in UBC phenotypically, functionally, and epigenetically represent a true Treg population. At the invasive front of UBC tumors, we found an inverse relationship between Treg frequency and expression of matrix metalloproteinase 2 (MMP2), a key proinvasive factor induced by tumor-promoting inflammation. Correspondingly, a significant, dose-dependent Treg-mediated downregulation of MMP2 protein and mRNA expression was observed in both macrophages and UBC cells. Also, we found that Treg frequency specifically at the invasive front positively correlated with survival. Thus, we identify Treg-mediated suppression of MMP2 in the tumor microenvironment as a mechanism explaining the paradoxical positive prognostic impact of tumor-infiltrating Tregs in UBC.
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| 2. |
- Zhang, Lu, et al.
(författare)
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Detection of micro-metastases by flow cytometry in lymph nodes from patients with penile cancer
- 2018
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Ingår i: BMC Urology. - : BioMed Central (BMC). - 1471-2490. ; 18:1
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: The tumor draining lymph node concept was first described in penile cancer for staging. Immunohistochemistry and histopathology evaluations are routinely used in clinical practice to examine lymph nodes for metastasis. However, these methods are time-consuming with low diagnostic accuracy and micro-metastases might be missed. In this study, we aim to evaluate detection of metastatic cells in draining lymph nodes by flow cytometry.METHODS: To assess the sensitivity of micro-metastasis detection by FACS (Fluorescence-activated cell sorting), HeLa cells were titrated into Peripheral blood mononuclear cells (PBMCs) and expression of pan-cytokeratin AE1/AE3 was analyzed. Single cell suspensions were separately prepared from 10 regional lymph nodes obtained from 5 patients with invasive penile cancer undergoing radical surgery and lymph node dissection. Lymph node dereived cells were examined for cell surface expression of EpCAM, E-cadherin and intracellular expression of pan-cytokeratin AE1/AE3 by FACS.RESULTS: Ten lymph nodes from 5 penile cancer patients were investigated in a head-to-head comparison between FACS and pathology examination of sections. All metastatic lymph nodes verified by pathology examination were also identified by FACS. Two additional lymph nodes with micro-metastases were diagnosed by FACS only.CONCLUSIONS: FACS analyses of pan-cytokeratin AE1/AE3 stained single cells from tumor draining lymph nodes can be used to detect micro-metastases in patients with penile cancer patients.
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| 3. |
- Ahlén Bergman, Emma, et al.
(författare)
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Increased CD4+ T cell lineage commitment determined by CpG methylation correlates with better prognosis in urinary bladder cancer patients
- 2018
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Ingår i: Clinical Epigenetics. - : BMC. - 1868-7083 .- 1868-7075. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Urinary bladder cancer is a common malignancy worldwide. Environmental factors and chronic inflammation are correlated with the disease risk. Diagnosis is performed by transurethral resection of the bladder, and patients with muscle invasive disease preferably proceed to radical cystectomy, with or without neoadjuvant chemotherapy. The anti-tumour immune responses, known to be initiated in the tumour and draining lymph nodes, may play a major role in future treatment strategies. Thus, increasing the knowledge of tumour-associated immunological processes is important. Activated CD4+ T cells differentiate into four main separate lineages: Th1, Th2, Th17 and Treg, and they are recognized by their effector molecules IFN-γ, IL-13, IL-17A, and the transcription factor Foxp3, respectively. We have previously demonstrated signature CpG sites predictive for lineage commitment of these four major CD4+ T cell lineages. Here, we investigate the lineage commitment specifically in tumour, lymph nodes and blood and relate them to the disease stage and response to neoadjuvant chemotherapy.RESULTS: Blood, tumour and regional lymph nodes were obtained from patients at time of transurethral resection of the bladder and at radical cystectomy. Tumour-infiltrating CD4+ lymphocytes were significantly hypomethylated in all four investigated lineage loci compared to CD4+ lymphocytes in lymph nodes and blood (lymph nodes vs tumour-infiltrating lymphocytes: IFNG -4229 bp p < 0.0001, IL13 -11 bp p < 0.05, IL17A -122 bp p < 0.01 and FOXP3 -77 bp p > 0.05). Examination of individual lymph nodes displayed different methylation signatures, suggesting possible correlation with future survival. More advanced post-cystectomy tumour stages correlated significantly with increased methylation at the IFNG -4229 bp locus. Patients with complete response to neoadjuvant chemotherapy displayed significant hypomethylation in CD4+ T cells for all four investigated loci, most prominently in IFNG p < 0.0001. Neoadjuvant chemotherapy seemed to result in a relocation of Th1-committed CD4+ T cells from blood, presumably to the tumour, indicated by shifts in the methylation patterns, whereas no such shifts were seen for lineages corresponding to IL13, IL17A and FOXP3.CONCLUSION: Increased lineage commitment in CD4+ T cells, as determined by demethylation in predictive CpG sites, is associated with lower post-cystectomy tumour stage, complete response to neoadjuvant chemotherapy and overall better outcome, suggesting epigenetic profiling of CD4+ T cell lineages as a useful readout for clinical staging.
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| 4. |
- Hartana, Ciputra Adijaya, et al.
(författare)
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Urothelial bladder cancer may suppress perforin expression in CD8+ T cells by an ICAM-1/TGFβ2 mediated pathway
- 2018
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Ingår i: PLOS ONE. - : Public Library Science. - 1932-6203. ; 13:7
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Tidskriftsartikel (refereegranskat)abstract
- The immune system plays a significant role in urothelial bladder cancer (UBC) progression, with CD8+ T cells being capable to directly kill tumor cells using perforin and granzymes. However, tumors avoid immune recognition by escape mechanisms. In this study, we aim to demonstrate tumor immune escape mechanisms that suppress CD8+ T cells cytotoxicity. 42 patients diagnosed with UBC were recruited. CD8+ T cells from peripheral blood (PB), sentinel nodes (SN), and tumor were analyzed in steady state and in vitro-stimulated conditions by flow cytometry, RT-qPCR, and ELISA. Mass spectrometry (MS) was used for identification of proteins from UBC cell line culture supernatants. Perforin was surprisingly found to be low in CD8+ T cells from SN, marked by 1.8-fold decrease of PRF1 expression, with maintained expression of granzyme B. The majority of perforin-deficient CD8+ T cells are effector memory T (TEM) cells with exhausted Tc2 cell phenotype, judged by the presence of PD-1 and GATA-3. Consequently, perforin-deficient CD8+ T cells from SN are low in T-bet expression. Supernatant from muscle invasive UBC induces perforin deficiency, a mechanism identified by MS where ICAM-1 and TGFβ2 signaling were causatively validated to decrease perforin expression in vitro. Thus, we demonstrate a novel tumor escape suppressing perforin expression in CD8+ T cells mediated by ICAM-1 and TGFβ2, which can be targeted in combination for cancer immunotherapy.
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| 5. |
- Krantz, David, et al.
(författare)
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Neoadjuvant Chemotherapy Reinforces Antitumour T cell Response in Urothelial Urinary Bladder Cancer
- 2018
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Ingår i: European Urology. - : Elsevier. - 0302-2838 .- 1873-7560. ; 74:6, s. 688-692
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Tidskriftsartikel (refereegranskat)abstract
- Evidence indicates that neoadjuvant chemotherapy (NAC) may promote antitumour immune responses by activating T cells. The tumour-draining sentinel node (SN) is a key site to study tumour-specific T cell activation, being the primary immunological barrier against the tumour. In this prospective study, we set out to elucidate the effects of NAC on T cell subsets in the SNs of patients with muscle-invasive urothelial bladder cancer. We found that CD8+ effector T (Teff) cell exhaustion was reduced after NAC treatment, while cytotoxicity was increased. Additionally, in complete responders (CR patients), these cells were functionally committed effectors, as displayed by epigenetic analysis. In CD4+ Teffs, NAC treatment was associated with increased clonal expansion of tumour-specific SN-derived cells, as demonstrated by a specific cell reactivity assay. In contrast, we observed an attenuating effect of NAC on regulatory T cells (Tregs) with a dose-dependent decrease in Treg frequency and reduced effector molecule expression in the remaining Tregs. In addition, multicolour flow cytometry analysis revealed that CR patients had higher Teff to activated Treg ratio, promoting antitumoural T cell activation. These results suggest that NAC reinforces the antitumour immune response by activating the effector arm of the T cell compartment and diminishing the influence of suppressive Tregs.PATIENT SUMMARY: In this report, we analysed the effect of chemotherapy on immune cell subsets of 40 patients with advanced bladder cancer. We found that chemotherapy has a positive effect on immune effector T cells, whereas an opposite, diminishing effect was observed for immune-suppressive regulatory T cells. We conclude that chemotherapy reinforces the antitumour immune response in bladder cancer patients.
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