NUPR1 induces autophagy and promotes the progression of Esophageal squamous cell carcinoma via the MAPK-mTOR pathway.

PURPOSE: Esophageal squamous cell carcinoma (ESCC) is a dominant pathological type in China. NUPR1 is a complex molecule implicated in various physiological and biological functions whose expression is upregulated in response to stress. Furthermore, autophagy is a vital physiological mechanism in the onset and metastasis of malignancies. This study aims to uncover the influence of NUPR1 on ESCC occurrence and development by regulating autophagy while also exploring its association with the MAPK signaling pathway. METHODS: First, the differences in NUPR1 between ESCC and normal tissues were analyzed through online databases. Subsequently, the pathological tissues of clinical samples were stained and scored using immunohistochemistry. And NUPR1 expression in ESCC cells was investigated, as was the function of NUPR1 in the modulation of ESCC's malignant behavior. Furthermore, a nude mouse ESCC xenograft model was developed. Finally, RNA sequencing was performed on NUPR1-downregulated ESCC cells, which was verified using WB. RESULTS: Our findings initially uncovered differences in the expression of NUPR1 in ESCC and normal tissues. In vitro experiments demonstrated that NUPR1 downregulation significantly inhibited ESCC cell proliferation, invasion, and migration, as well as promoted their apoptosis. Our xenograft model exhibited significant inhibition of ESCC tumors upon NUPR1 downregulation. Subsequently, RNA sequencing uncovered that NUPR1 regulates its malignant biological behavior through MAPK-mTOR signaling pathway. Finally, we found that NUPR1 downregulation can inhibit autophagic flux in ESCC. CONCLUSION: Collectively, our findings show that NUPR1 enhances the progression of ESCC by triggering autophagy and is associated with the MAPK-mTOR signaling pathway.

JAML promotes the antitumor role of tumor-resident CD8+ T cells by facilitating their innate-like function in human lung cancer.

Tissue-resident memory CD8+T cells (CD8+TRMs) are thought to play a crucial role in cancer immunosurveillance. However, the characteristics of CD8+TRMs in the tumor microenvironment (TME) of human non-small cell lung cancer (NSCLC) remain unclear. Here, we report that CD8+TRMs accumulate explicitly and exhibit a unique gene expression profile in the TME of NSCLC. Interestingly, these tumor-associated CD8+TRMs uniquely exhibit an innate-like phenotype. Importantly, we found that junction adhesion molecule-like (JAML) provides an alternative costimulatory signal to activate tumor-associated CD8+TRMs via combination with cancer cell-derived CXADR (CXADR Ig-like cell adhesion molecule). Furthermore, we demonstrated that activating JAML could promote the expression of TLR1/2 on CD8+TRMs, inhibit tumor progression and prolong the survival of tumor-bearing mice. Finally, we found that higher CD8+TRMs and JAML expression in the TME could predict favorable clinical outcomes in NSCLC patients. Our study reveals an intrinsic bias of CD8+TRMs for receiving the tumor-derived costimulatory signal in the TME, which sustains their innate-like function and antitumor role. These findings will shed more light on the biology of CD8+TRMs and aid in the development of potential targeted treatment strategies for NSCLC.

Inflammatory dendritic cells restrain CD11b+CD4+ CTLs via CD200R in human NSCLC.

CD4+ cytotoxic T lymphocytes (CD4+ CTLs) are suggested to play a crucial role in inflammatory diseases, including cancer, but their characteristics in human non-small cell lung cancer (NSCLC) remain unknown. Here, using the cell surface marker CD11b, we identify CD11b+CD4+ CTLs as a cytotoxic subset of CD4+ T cells in multiple tissues of NSCLC patients. In addition, tumor-infiltrating CD11b+CD4+ CTLs show a dysfunctional phenotype with elevated expression of CD200 receptor (CD200R), a negatively immunomodulatory receptor. CD4+ regulatory T (Treg) cells restrain the anti-tumor role of CD11b+CD4+ CTLs via CD200. Mechanistically, inflammatory dendritic cells promote the CD200R expression of CD11b+CD4+ CTLs by secreting interleukin-1ß (IL-1ß). Finally, we demonstrate that CD200 blockade can revive the tumor-killing role of CD11b+CD4+ CTLs and prolong the survival of tumor-bearing mice. Taken together, our study identifies CD11b+CD4+ CTLs in NSCLC with decreased cytotoxicity that can be reinvigorated by CD200 blockade, suggesting that targeting CD200 is a promising immunotherapy strategy in NSCLC.

Tissue-resident CD69+ CXCR6+ Natural Killer cells with exhausted phenotype accumulate in human non-small cell lung cancer.

Natural killer (NK) cells with tissue-residency features (trNK cells) are a new subpopulation of NK cells, which plays an important role in tissue homeostasis. However, the characteristics of trNK cells in the tumor microenvironment (TME) of human cancers remain unclear. Using multicolor flow cytometry, we investigated the quantity, phenotype, and function of trNK cells in biospecimens freshly resected from 60 non-small cell lung cancer (NSCLC) patients. We successfully identified a new CD69+ CXCR6+ trNK subset with an immunomodulatory-like and exhausted phenotype, specifically accumulated in the TME of NSCLC. In vitro experiments showed that CD69+ CXCR6+ trNK cells more readily secreted IFN-γ and TNF-α spontaneously. Furthermore, the production of IFN-γ and TNF-α by tumor-infiltrating CD69+ CXCR6+ trNK cells was not induced by their reactivation in vitro, which is analogous to T-cell exhaustion. Finally, we demonstrated that the dysfunction of CD69+ CXCR6+ trNK cells could be partly ameliorated by PD-1 and CTLA-4 blockade. In summary, we identified a new dysfunctional CD69+ CXCR6+ trNK cell subset that specifically accumulates in the TME of NSCLC. Our findings suggest that CD69+ CXCR6+ trNK cells are a promising target for immune checkpoint inhibitors in the treatment of NSCLC.

The immune landscape of human thymic epithelial tumors.

Human thymic epithelial tumors (TET) are common malignancies in the anterior mediastinum with limited biological understanding. Here we show, by single cell analysis of the immune landscape, that the developmental pattern of intra-tumoral T-cells identify three types within TETs. We characterize the developmental alterations and TCR repertoires of tumor-infiltrating T cells in the context of the distinguishing epithelial tumor cell types. We demonstrate that a subset of tumor cells, featuring medullary thymic epithelial cell (TEC) phenotype and marked by KRT14/GNB3 expression, accumulate in type 1 TETs, while T-cell positive selection is inhibited. Type 2 TETs are dominated by CCL25+ cortical TEC-like cells that appear to promote T-cell positive selection. Interestingly, the CHI3L1+ medullary TEC-like cells that are the characteristic feature of type 3 TETs don't seem to support T-cell development, however, they may induce a tissue-resident CD8+ T cell response. In summary, our work suggests that the molecular subtype of epithelial tumour cells in TETs determine their tumour immune microenvironment, thus GNB3 and CHI3L1 might predict the immunological behavior and hence prognosis of these tumours.

Oncolytic adenovirus promotes vascular normalization and nonclassical tertiary lymphoid structure formation through STING-mediated DC activation.

Inducing a full antitumor immune response in the tumor microenvironment (TME) is essential for successful cancer immunotherapy. Here, we report that an oncolytic adenovirus carrying mIL-15 (Ad-IL15) can effectively induce antitumor immune response and inhibit tumor growth in a mouse model of cancer. We found that Ad-IL15 facilitated the activation and infiltration of immune cells, including dendritic cells (DCs), T cells and natural killer (NK) cells, in the TME. Unexpectedly, we observed that Ad-IL15 also induced vascular normalization and tertiary lymphoid structure formation in the TME. Moreover, we demonstrated these Ad-IL15-induced changes in the TME were depended on the Ad-IL15-induced activation of the STING-TBK1-IRF3 pathway in DCs. Taken together, our findings suggest that Ad-IL15 is a candidate for cancer immunotherapy that promotes immune cell activation and infiltration, tumor vascular normalization and tertiary lymphoid structure formation in the TME.

CD38 identifies pre-activated CD8+ T cells which can be reinvigorated by anti-PD-1 blockade in human lung cancer.

BACKGROUND: CD38 has been observed expressing in activated T cells, while the features and functions of CD38+ T cells in human NSCLC are still unclear. METHODS: Here we uncovered the correlation between CD38 expression and survival and immune infiltration levels in tumor of NSCLC. Then, we collected samples from 51 NSCLC patients to study the biological feature and response to anti-PD-1 of tumor-infiltrating CD38+ CD8+ T cells in vitro. RESULTS: We found CD38 expression correlated with the survival and immune infiltration levels of NSCLC. It is interesting that CD38+ CD8+ T cells enriched in the tumors expressed higher level of cytotoxic molecule, cytokines and PD-1 than CD38- CD8+ T cells. Moreover, PD-1+ subset in tumor-infiltrating CD38+ CD8+ T cells expressed higher level of activated markers than PD-1+ CD38- CD8+ T cells. Next, we found tumor-infiltrating CD38+ CD8+ T cells expressed higher level of CD103, IFN-γ, TNF-α and perforin than CD38- CD8+ T cells when were reactivated in vitro. Finally, we observed that CD38+ CD8+ T cells isolated from tumors could be reinvigorated by anti-PD-1 in vitro. CONCLUSIONS: Our findings demonstrate that CD38 expression defines a subset of CD8+ T cells enriched in tumors of NSCLC which have paradoxical phenotypes and response to anti-PD-1. Our results suggest a pre-priming of these cells is may exist in tumor and consequentially facilitate it acquiring both anti-tumor potency and exhausted phenotype which can be reinvigorated by PD-1 blockade.

Systemic Immune Dysregulation Correlates With Clinical Features of Early Non-Small Cell Lung Cancer.

Background: Systemic immune dysregulation correlates with cancer progression. However, the clinical implications of systemic immune dysregulation in early non-small cell lung cancer (NSCLC) remain unclear. Methods: Using a panel of 9 markers to identify 12 parameters in the peripheral blood of 326 patients (34 in the discovery group and 292 in the validation group), we investigated systemic immune dysregulation in early NSCLC. Then, we analyzed the impact of surgery on the systemic immune state of these patients. Finally, we analyzed correlations between systemic immune dysregulation and the clinical features of early NSCLC. Results: We found striking systemic immune dysregulation in the peripheral blood of early NSCLC patients. This dysregulation was characterized by a significant decrease in total lymphocytes, T cells, quiescent T cells, CD4+ T cells, and NKT cells. We also observed increased proportions of activated lymphocytes and activated T cells. Systemic immune dysregulation was increased after surgery. Furthermore, systemic immune dysregulation was correlated with multiple clinical features, such as sex, age, smoking history, pathological type, tumor stage, surgical approach, tumor differentiation, and epidermal growth factor receptor (EGFR) mutation. Finally, we observed that systemic immune dysregulation was correlated with complications and systemic inflammatory response syndrome (SIRS) in early NSCLC patients. Conclusions: Our results reveal systemic immune dysregulation occurring in early NSCLC and demonstrate the correlation between these dysregulations and clinical features. Our findings suggest that systemic immune dysregulation is involved in cancer development and may be a promising candidate for high-risk screening and treatment strategies for early NSCLC.

CD8+ T Cells Form the Predominant Subset of NKG2A+ Cells in Human Lung Cancer.

Background: NKG2A is an inhibitory receptor of both T cells and natural killer (NK) cells. Persistent activation promotes T cells and NK cells to express NKG2A and results in the progression of chronic infection and cancer. However, the characteristics and subsets of NKG2A+ lymphocytes in human lung cancer are still unclear. Methods: Here, we used the Tumor Immune Estimation Resource database and immune profiling of paired biospecimens to uncover the correlation between NKG2A expression and immune infiltration levels in human cancer as well as the characteristics of NKG2A+ lymphocytes in human lung cancer. Results: We found that KLRC1 expression was especially correlated with CD8+ T-cell infiltration levels in 34 types of human cancer through the Tumor Immune Estimation Resource database. Moreover, NKG2A+ CD8+ T cells were the predominant subset of NKG2A+ lymphocytes in human lung cancer. In contrast, the NKG2A+ NK cells were decreased in tumors compared with the paired normal lung tissue. Tumor-infiltrating NKG2A+ CD8+ T cells expressed tissue-resident memory T cell (TRM cell) and exhausted T-cell markers. Cytokines and cytotoxic molecules secreted by tumor-infiltrating NKG2A+ CD8+ T cells were significantly lower than those secreted by NKG2A- CD8+ T cells in vitro. When stimulated with T-cell receptor activator, tumor-infiltrating NKG2A+ CD8+ T cells could secrete large amounts of granzyme B. Conclusions: Our findings demonstrate that tumor-infiltrating NKG2A+ CD8+ T cells form the predominant subset of NKG2A+ cells in human lung cancer and suggest that targeting NKG2A+ CD8+ T cells is a promising approach for future anti-lung cancer immunotherapy.

Inhibition of MUC1-C regulates metabolism by AKT pathway in esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is one of the most common digestive tumors worldwide. The Mucin 1 (MUC1) heterodimeric protein has been confirmed that is overexpressed in ESCC and induced adverse outcomes. However, the detailed mechanism(s) remained challenging. So, we investigated the relationship between MUC1-C and metabolism in ESCC cells. In the results, TP53-induced glycolysis and apoptosis regulator (TIGAR) was overexpressed and correlative with MUC1-C positively in ESCC tissue. Targeting MUC1-C inhibits AKT-mTORC-S6K1 signaling and blocks TIGAR translation. We found that the inhibitory effect of GO-203 on TIGAR was mediated by inhibition of AKT-mTOR-S6K1 pathway. The findings also demonstrated that the suppressive effect of GO-203 on TIGAR is related to the decrease of glutathione level, the increase of reactive oxygen species and the loss of mitochondrial transmembrane membrane potential. In xenograft tissues, GO-203 inhibited the growth of ESCC cells and lead to the low expression of transmembrane C-terminal subunit (MUC1-C) and TIGAR. This evidence supports the contention that MUC1-C is significant for metabolism in ESCC and indicated that MUC1-C is a potential target for the treatment of ESCC.

Blocking FGFR4 exerts distinct anti-tumorigenic effects in esophageal squamous cell carcinoma.

BACKGROUND: The FGFR family can be activated by FGFs and plays important roles in regulating cell growth, differentiation, migration, and angiogenesis. Recent studies have suggested that FGFR4 could regulate several processes, including tumor progression. Esophageal squamous cell carcinoma (ESCC) is a malignancy with high global occurrence. However, the molecule mechanism and the potential roles of FGFR4 in ESCC remain unknown. METHODS: Immunohistochemistry and Western blotting were used to detect FGFR4 expression in ESCC samples and cell lines. Cell counting kit-8, and clonogenic, transwell, flow cytometric, and tumor xenograft in nude mice assays were utilized to determine the effect of blocking FGFR4 in proliferation, invasion, migration, and apoptosis of ESCC cells. RESULTS: FGFR4 is frequently overexpressed in ESCC tissue and cell lines. in vitro assays have shown that blocking FGFR4 by a specific blocker, H3B-6527, significantly decreases proliferation, invasion, and migration, and alters epithelial-mesenchymal transition markers in ESCC cells. In addition, FGFR4 blockade is associated with the induction of apoptosis and affects PI3K/Akt and MAPK/ERK pathways. Moreover, FGFR4 blockade could significantly inhibit the growth of xenograft tumors in vivo. CONCLUSION: Our findings suggest that blocking FGFR4 significantly suppresses the malignant behaviors of ESCC and indicate that FGFR4 is a potential target for the treatment of ESCC.

Inhibition of MUC1-C entering nuclear suppresses MYC expression and attenuates malignant growth in esophageal squamous cell carcinoma.

BACKGROUND: The mucin 1 (MUC1) heterodimeric protein (N-terminal subunit and C-terminal subunit) is aberrantly overexpressed in esophageal squamous cell carcinoma (ESCC) and has been linked to poor outcomes in this disease. The detailed mechanism(s), however, remains unclear. In this article, we investigate the effects of the MUC1 C-terminal transmembrane subunit (MUC1-C) through the inhibitor GO-201, which inhibits MUC1-C targeting to nuclear. PATIENTS AND METHODS: The expression of MUC1-C and MYC in the ESCC samples and cell lines was detected by immunohistochemistry, immunofluorescence and western blotting. MYC mRNA level was determined by using quantitative real-time polymerase chain reaction. In addition, Cell Counting Kit-8, clonogenic assay, transwell assay and tumor xenograft in nude mice assay were utilized to determine the role of MUC1-C in proliferation, invasion and migration of ESCC cells. RESULTS: The level of MUC1-C in nuclear and MYC in whole cells in the ESCC tissue is significantly higher than that in the noncancerous tissue. Treatment of MUC1-C-overexpressing ESCC cells with GO-201 was associated with downregulation of MYC expression and induction of apoptosis. Besides, in vitro and in vivo assays have both shown that inhibiting MUC1-C targeting to the nucleus by the GO-201 significantly decreased the abilities of proliferation, invasion and migration in ESCC cells. CONCLUSION: Our findings suggest that MUC1-C targeting to the nucleus plays an important role in suppressing the malignant growth of ESCC and indicate that MUC1-C is a potential target for the treatment of ESCC.

CEP55 promotes the proliferation, migration and invasion of esophageal squamous cell carcinoma via the PI3K/Akt pathway.

BACKGROUND: Centrosomal protein 55 (CEP55) is an important prognostic biomarker that plays an essential role in the proliferation, migration and invasion of multiple tumors. We aimed to investigate the prognostic value of CEP55 in pN0 esophageal squamous cell carcinoma (ESCC) and explore its biological function in ESCC cells. METHODS: We used immunohistochemistry and Western blot analysis to detect the expression of CEP55 in ESCC. Furthermore, both in vitro and in vivo assays were used to determine the effect of CEP55 on malignant behavior in ESCC cells. RESULTS: As expected, we found that CEP55 was overexpressed in ESCC. Univariate and multivariate analyses demonstrated that patients with CEP55 overexpression had a poor prognosis. Additionally, the abilities of proliferation, migration and invasion of cells, as well as the epithelial-mesenchymal transition markers, were all altered with the changed CEP55 expression levels in ESCC cells. Further study elucidated that CEP55 facilitated ESCC via the PI3K/Akt pathway. Blockade of this pathway markedly attenuated CEP55-mediated proliferation, migration, invasion and epithelial-mesenchymal transition of ESCC cells. CONCLUSION: Oncogenic CEP55 correlates with a poor prognosis by regulating tumor cell proliferation, migration and invasion via the PI3K/Akt pathway. It can serve as a promising prognostic biomarker and therapeutic target of pN0 ESCC after Ivor-Lewis esophagectomy.