Airway epithelial CD47 plays a critical role in inducing influenza virus-mediated bacterial super-infection.

Respiratory viral infection increases host susceptibility to secondary bacterial infections, yet the precise dynamics within airway epithelia remain elusive. Here, we elucidate the pivotal role of CD47 in the airway epithelium during bacterial super-infection. We demonstrated that upon influenza virus infection, CD47 expression was upregulated and localized on the apical surface of ciliated cells within primary human nasal or bronchial epithelial cells. This induced CD47 exposure provided attachment sites for Staphylococcus aureus, thereby compromising the epithelial barrier integrity. Through bacterial adhesion assays and in vitro pull-down assays, we identified fibronectin-binding proteins (FnBP) of S. aureus as a key component that binds to CD47. Furthermore, we found that ciliated cell-specific CD47 deficiency or neutralizing antibody-mediated CD47 inactivation enhanced in vivo survival rates. These findings suggest that interfering with the interaction between airway epithelial CD47 and pathogenic bacterial FnBP holds promise for alleviating the adverse effects of super-infection.

NgR1 is an NK cell inhibitory receptor that destabilizes the immunological synapse.

The formation of an immunological synapse (IS) is essential for natural killer (NK) cells to eliminate target cells. Despite an advanced understanding of the characteristics of the IS and its formation processes, the mechanisms that regulate its stability via the cytoskeleton are unclear. Here, we show that Nogo receptor 1 (NgR1) has an important function in modulating NK cell-mediated killing by destabilization of IS formation. NgR1 deficiency or blockade resulted in improved tumor control of NK cells by enhancing NK-to-target cell contact stability and regulating F-actin dynamics during IS formation. Patients with tumors expressing abundant NgR1 ligand had poor prognosis despite high levels of NK cell infiltration. Thus, our study identifies NgR1 as an immune checkpoint in IS formation and indicates a potential approach to improve the cytolytic function of NK cells in cancer immunotherapy.

Stepwise molecular mechanisms responsible for chemoresistance in bladder cancer cells.

Chemotherapy resistance is an obstacle to cancer therapy and is considered a major cause of recurrence. Thus, understanding the mechanisms of chemoresistance is critical to improving the prognosis of patients. Here, we have established a stepwise gemcitabine-resistant T24 bladder cancer cell line to understand the molecular mechanisms of chemoresistance within cancer cells. The characteristics of the stepwise chemoresistance cell line were divided into 4 phases (parental, early, intermediate, and late phases). These four phase cells showed increasingly aggressive phenotypes in vitro and in vivo experiments with increasing phases and revealed the molecular properties of the biological process from parent cells to phased gemcitabine-resistant cell line (GRC). Taken together, through the analysis of gene expression profile data, we have characterized gene set of each phase indicating the response to anticancer drug treatment. Specifically, we identified a multigene signature (23 genes including GATA3, APOBEC3G, NT5E, MYC, STC1, FOXD1, SMAD9) and developed a chemoresistance score consisting of that could predict eventual responsiveness to gemcitabine treatment. Our data will contribute to predicting chemoresistance and improving the prognosis of bladder cancer patients.

The Role of IL-7 and IL-7R in Cancer Pathophysiology and Immunotherapy.

Interleukin-7 (IL-7) is a multipotent cytokine that maintains the homeostasis of the immune system. IL-7 plays a vital role in T-cell development, proliferation, and differentiation, as well as in B cell maturation through the activation of the IL-7 receptor (IL-7R). IL-7 is closely associated with tumor development and has been used in cancer clinical research and therapy. In this review, we first summarize the roles of IL-7 and IL-7Rα and their downstream signaling pathways in immunity and cancer. Furthermore, we summarize and discuss the recent advances in the use of IL-7 and IL-7Rα as cancer immunotherapy tools and highlight their potential for therapeutic applications. This review will help in the development of cancer immunotherapy regimens based on IL-7 and IL-7Rα, and will also advance their exploitation as more effective and safe immunotherapy tools.

YAP1 activation is associated with the progression and response to immunotherapy of non-muscle invasive bladder cancer.

BACKGROUND: Despite the availability of several treatments for non-muscle-invasive bladder cancer (NMIBC), many patients are still not responsive to treatments, and the disease progresses. A new prognostic classifier can differentiate between treatment response and progression, and it could be used as a very important tool in patient decision-making regarding treatment options. In this study, we focused on the activation of Yes-associated protein 1 (YAP1), which is known to play a pivotal role in tumour progression and serves as a factor contributing to the mechanism of resistance to various relevant therapeutic agents. We further evaluated its potential as a novel prognostic agent. METHODS: We identified YAP1-associated gene signatures based on UC3-siYAP1 cells (n=8) and NMIBC cohort (n=460). Cross-validation was performed using 5 independent bladder cancer patient cohorts (n=1006). We also experimentally validated the changes of gene expression levels representing each subgroup. FINDINGS: The 976-gene signature based on YAP1-activation redefined three subgroups and had the benefits of Bacillus Calmette-Guérin (BCG) treatment in patients with NMIBC (hazard ratio 3.32, 95% CI 1.29-8.56, p = 0.01). The integrated analysis revealed that YAP1 activation was associated with the characterization of patients with high-risk NMIBC and the response to immunotherapy. INTERPRETATION: This study suggests that YAP1 activation has an important prognostic effect on bladder cancer progression and might be useful in the selection of immunotherapy. FUNDING: A funding list that contributed to this research can be found in the Acknowledgements section.

Transcriptional Profiling of Advanced Urothelial Cancer Predicts Prognosis and Response to Immunotherapy.

Recent investigations reported that some subtypes from the Lund or The Cancer Genome Atlas (TCGA) classifications were most responsive to PD-L1 inhibitor treatment. However, the association between previously reported subtypes and immune checkpoint inhibitor (ICI) therapy responsiveness has been insufficiently explored. Despite these contributions, the ability to predict the clinical applicability of immune checkpoint inhibitor therapy in patients remains a major challenge. Here, we aimed to re-classify distinct subtypes focusing on ICI responsiveness using gene expression profiling in the IMvigor 210 cohort (n = 298). Based on the hierarchical clustering analysis, we divided advanced urothelial cancer patients into three subgroups. To confirm a prognostic impact, we performed survival analysis and estimated the prognostic value in the IMvigor 210 and TCGA cohort. The activation of CD8+ T effector cells was common for patients of classes 2 and 3 in the TCGA and IMvigor 210 cohort. Survival analysis showed that patients of class 3 in the TCGA cohort had a poor prognosis, while patients of class 3 showed considerably prolonged survival in the IMvigor 210 cohort. One of the distinct characteristics of patients in class 3 is the inactivation of the TGFß and YAP/TAZ pathways and activation of the cell cycle and DNA replication and DNA damage (DDR). Based on our identified transcriptional patterns and the clinical outcomes of advanced urothelial cancer patients, we constructed a schematic summary. When comparing clinical and transcriptome data, patients with downregulation of the TGFß and YAP/TAZ pathways and upregulation of the cell cycle and DDR may be more responsive to ICI therapy.

Prevention of abdominal aortic aneurysm by anti-microRNA-712 or anti-microRNA-205 in angiotensin II-infused mice.

OBJECTIVE: Abdominal aortic aneurysm (AAA) is characterized as a progressive dilation and degradation of the aortic wall, associated with activation of matrix metalloproteinases (MMPs) and inflammation. Emerging evidence indicates a role for microRNAs (miRNAs) in AAA pathogenesis, but it is unclear whether abdominal aortic endothelial miRNAs play a role in the disease process. We aimed to identify miRNAs in the abdominal aortic endothelium that play a critical role in AAA development. APPROACH AND RESULTS: The mouse model of AAA induced by angiotensin II infusion was used in this study. Through a miRNA array and validation study, we initially identified the murine-specific miR-712 and subsequently its human/murine homolog miR-205 as angiotensin II-induced miRNAs in the abdominal aortic endothelium in vivo and in vitro. Mechanistically, miR-712 stimulated MMP activity in the aortic wall by directly targeting 2 MMP inhibitors: tissue inhibitor of metalloproteinase 3 (TIMP3) and reversion-inducing cysteine-rich protein with kazal motifs (RECK). Silencing of miR-712 and miR-205 by using anti-miR-712 and anti-miR-205, respectively, significantly decreased the aortic MMP activity and inflammation, preventing AAA development in angiotensin II-infused ApoE(-/-) mice. Further, upregulation of 4 angiotensin II-sensitive miRNAs, miR-205, -21, -133b, and -378, identified in this murine study were confirmed in human AAA samples compared with nondiseased control. CONCLUSIONS: Our results demonstrate that angiotensin II-sensitive miR-712 and its human homolog miR-205 downregulate TIMP3 and RECK, which in turn stimulate aortic MMP activity and inflammation, leading to AAA development. Targeting these miRNAs may be a novel therapeutic strategy to prevent AAA.

Flow-dependent epigenetic DNA methylation regulates endothelial gene expression and atherosclerosis.

In atherosclerosis, plaques preferentially develop in arterial regions of disturbed blood flow (d-flow), which alters endothelial gene expression and function. Here, we determined that d-flow regulates genome-wide DNA methylation patterns in a DNA methyltransferase-dependent (DNMT-dependent) manner. Induction of d-flow by partial carotid ligation surgery in a murine model induced DNMT1 in arterial endothelium. In cultured endothelial cells, DNMT1 was enhanced by oscillatory shear stress (OS), and reduction of DNMT with either the inhibitor 5-aza-2'-deoxycytidine (5Aza) or siRNA markedly reduced OS-induced endothelial inflammation. Moreover, administration of 5Aza reduced lesion formation in 2 mouse models of atherosclerosis. Using both reduced representation bisulfite sequencing (RRBS) and microarray, we determined that d-flow in the carotid artery resulted in hypermethylation within the promoters of 11 mechanosensitive genes and that 5Aza treatment restored normal methylation patterns. Of the identified genes, HoxA5 and Klf3 encode transcription factors that contain cAMP response elements, suggesting that the methylation status of these loci could serve as a mechanosensitive master switch in gene expression. Together, our results demonstrate that d-flow controls epigenomic DNA methylation patterns in a DNMT-dependent manner, which in turn alters endothelial gene expression and induces atherosclerosis.

Involvement of pro-phenoloxidase 3 in lamellocyte-mediated spontaneous melanization in Drosophila.

Phenoloxidase (PO), a melanin-forming enzyme around the foreign bodies, is an important component of the host defense system in invertebrates. Pro-PO is the enzymatically inactive zymogen form of PO. In the Drosophila genome, three Pro-PO isoforms have been identified to date. These include Pro-PO1 and 2, which are primarily expressed in crystal cells, and Pro-PO3, which is predominantly found in the lamellocytes. In this study, we demonstrated that Drosophila Pro-PO3, but not Pro-PO1 or 2, is enzymatically active in its zymogen form. These findings were evidenced by spectacular melanin forming capacities of various cells and tissues that overexpressed these proenzymes. Furthermore, the melanization phenotype observed in the lamellocyte-enriched hop(Tum-1) mutant was drastically reduced in the absence of PPO3, indicating that PPO3 plays a major role in the lamellocyte-mediated spontaneous melanization process. Taken together, these findings indicate that the biochemical properties, activation mode and in vivo role of Pro-PO3 are likely distinct from those of the other two Pro-PO enzymes involved in Drosophila physiology.

Homeodomain protein CDX2 regulates COX-2 expression in colorectal cancer.

CDX2 is an intestine-specific tumor suppressor gene encoding homeodomain-containing transcription factor, which is involved in a variety of developmental, proliferating, and differentiating processes. Moreover, the expression of CDX2 is reduced in a subset of primary colorectal cancers. In contrast, cyclooxygenase-2 (COX-2) is often up-regulated in human colorectal cancers. However, the molecular relationship between CDX2 down-regulation and COX-2 up-regulation is unknown. Here we show that CDX2 down-regulates COX-2 promoter activity by interacting with NF-kappaB. The ectopic expression of CDX2 was found to suppress PMA-induced COX-2 promoter activity in a dose-dependent manner. In addition, the treatment of colorectal cancer cells with PMA resulted in significant reduction in the level of endogenous CDX2 and a significant increase in the level of endogenous COX-2, in a dose-dependent manner. Furthermore, CDX2 was found to co-immunoprecipitate with the p65 subunit of NF-kappaB and to inhibit p65-induced NF-kappaB minimal promoter activity in colon cancer cells. These results suggest that reduced CDX2 expression may be involved in colorectal carcinogenesis by enhancing NF-kappaB-mediated inflammatory genes such as COX-2.