Single-cell analysis of a progressive Rosai-Dorfman disease affecting the cerebral parenchyma: a case report.

Neurologic Rosai-Dorfman disease (RDD) is a rare type of non-Langerhans cell histiocytosis that affects the central nervous system. Most neurologic RDDs grow like meningiomas, have clear boundaries, and can be completely resected. However, a few RDDs are invasive and aggressive, and no effective treatment options are available because the molecular mechanisms involved remain unknown. Here, we report a case of deadly and glucocorticoid-resistant neurologic RDD and explore its possible pathogenic mechanisms via single-cell RNA sequencing. First, we identified two distinct but evolutionarily related histiocyte subpopulations (the C1Q+ and SPP1+ histiocytes) that accumulated in the biopsy sample. The expression of genes in the KRAS signaling pathway was upregulated, indicating gain-of-function of KRAS mutations. The C1Q+ and SPP1+ histiocytes were highly differentiated and arrested in the G1 phase, excluding the idea that RDD is a lympho-histio-proliferative disorder. Second, although C1Q+ histiocytes were the primary RDD cell type, SPP1+ histiocytes highly expressed several severe inflammation-related and invasive factors, such as WNT5A, IL-6, and MMP12, suggesting that SPP1+ histiocytes plays a central role in driving the progression of this disease. Third, oligodendrocytes were found to be the prominent cell type that initiates RDD via MIF and may resist glucocorticoid treatment via the MDK and PTN signaling pathways. In summary, in this case, we report a rare presentation of neurologic RDD and provided new insight into the pathogenic mechanisms of progressive neurologic RDD. This study will also offer evidence for developing precision therapies targeting this complex disease.

Endoscopic features of esophageal high-grade intraepithelial neoplasia dominated by cytological atypia.

Little is known about esophageal high-grade intraepithelial neoplasia dominated by cytological atypia (HGINc). We aimed to elucidate the endoscopic features of HGINc compared with esophageal high-grade intraepithelial neoplasia dominated by architectural atypia (HGINa). All patients pathologically diagnosed as esophageal high-grade intraepithelial neoplasia after endoscopic submucosal dissection at our center between January 2018 and December 2019 were included in this study. According to the pathological diagnosis, the patients were divided into two groups: HGINa group and HGINc group. Basic characteristics and endoscopic information were collected in detail. Data were analyzed statistically. Binary logistic regression was performed and a predictive model for HGINc was established. Then we evaluated its predictive value and built a nomogram for clinical application. A total of 175 patients were included in this study (126 with HGINa and 49 with HGINc). Among 228 lesions found in all patients, there were 148 HGINa and 80 HGINc. The independent relevant factors for HGINc were tobacco and alcohol usage, color, and gross type. To predict risk of HGINc, a three-factor model (TFM) was established with a highest area under curve (AUC) as 0.869 (95% CI, 0.852, 0.939). When the cut-off value was set as 0.3569184, the diagnostic accuracy, sensitivity, specificity, positive predictive value, and negative predictive value for HGINc was 81.14%, 88.75%, 77.03%, 67.62%, and 92.68%, respectively. HGINc differs greatly in endoscopic features from HGINa in our study. It's important to reduce misdiagnosis that our model was established with good predictive value for clinical application.

Single-cell transcriptomics provides insights into the origin and microenvironment of human oesophageal high-grade intraepithelial neoplasia.

BACKGROUND: High-grade intraepithelial neoplasia (HIN) is the precursor of oesophageal squamous cell carcinoma. The molecular and functional properties of HIN are determined by intrinsic origin cells and the extrinsic microenvironment. Yet, these factors are poorly understood. METHODS: We performed single-cell RNA sequencing of cells from HINs and adjacent tissues from the human oesophagus. We analysed the heterogeneity of basal layer cells and confirmed it using immunostaining. Aneuploid cells in HIN were studied using primary cell culture combined with karyotype analysis. We reconstructed the lineage relationship between tumour and normal populations based on transcriptome similarity. Integration analysis was applied to our epithelial data and published invasive cancer data, and results were confirmed by immunostaining and 3D organoid functional experiments. We also analysed the tumour microenvironment of HIN. RESULTS: The basal layer contained two cell populations: KRT15high STMN1low and KRT15high STMN1high cells, which were located mainly in the interpapillary and papillary zones, respectively. The KRT15high STMN1low population more closely resembled stem cells and transcriptome similarity revealed that HIN probably originated from these slow-cycling KRT15high STMN1low cells. 3D Organoid experiments and RNA-sequencing showed that basal-cell features and the differentiation ability of the normal epithelium were largely retained in HIN, but may change dramatically in tumour invasion stage. Moreover, the tumour microenvironment of HIN was characterised by both inflammation and immunosuppression. CONCLUSIONS: Our study provides a comprehensive single-cell transcriptome landscape of human oesophageal HIN. Our findings on the origin cells and unique microenvironment of HIN will allow for the development of strategies to block tumour progression and even prevent cancer initiation.

MBD3 promotes hepatocellular carcinoma progression and metastasis through negative regulation of tumour suppressor TFPI2.

BACKGROUND: The mechanism of recurrence and metastasis of hepatocellular carcinoma (HCC) is complex and challenging. Methyl-CpG binding domain protein 3 (MBD3) is a key epigenetic regulator involved in the progression and metastasis of several cancers, but its role in HCC remains unknown. METHODS: MBD3 expression in HCC was detected by immunohistochemistry and its association with clinicopathological features and patient's survival was analysed. The effects of MBD3 on hepatoma cells growth and metastasis were investigated, and the mechanism was explored. RESULTS: MBD3 is significantly highly expressed in HCC, associated with the advanced tumour stage and poor prognosis in HCC patients. MBD3 promotes the growth, angiogenesis and metastasis of HCC cells by inhibiting the tumour suppressor tissue factor pathway inhibitor 2 (TFPI2). Mechanistically, MBD3 can inhibit the TFPI2 transcription via the Nucleosome Remodeling and Deacetylase (NuRD) complex-mediated deacetylation, thus reactivating the activity of matrix metalloproteinases (MMPs) and PI3K/AKT signaling pathway, leading to the progression and metastasis of HCC CONCLUSIONS: Our results unravel the novel regulatory function of MBD3 in the progression and metastasis of HCC and identify MBD3 as an independent unfavourable prognostic factor for HCC patients, suggesting its potential as a promising therapeutic target as well.

Impaired Cytolytic Activity and Loss of Clonal Neoantigens in Elderly Patients With Lung Adenocarcinoma.

INTRODUCTION: Whether the efficacy of programmed death 1 (PD-1)/programmed death ligand 1 (PD-L1) inhibitors declines with senescence remains controversial for lung adenocarcinoma (LUAD). Responsiveness to anti-PD-1/PD-L1 therapy is thought to rely on neoantigen exposure and immune elements in the tumor microenvironment. In this study, we explored the features of the tumor immune microenvironment in elderly patients with treatment-naïve LUAD. METHODS: Transcriptome profiles and clinical characteristics of patients with LUAD were retrieved from The Cancer Genome Atlas as a discovery cohort. Immune cell infiltration (quantified by a single-sample gene set enrichment analysis), immunoregulatory molecule expression, and mutational patterns (from The Cancer Immunome Atlas) were compared between young and elderly patients. Immune cell infiltration was verified by immunohistochemistry using a validation cohort including 105 treatment-naïve patients with LUAD. A tissue microarray consisting of 120 LUAD patients was used in the immunohistochemistry validation. RESULTS: Activated CD8+ T cell numbers increased slightly with age, but cytolytic molecules in T cells (granzyme B [GZMB], perforin 1 [PRF1], granzyme A [GZMA], granzyme M [GZMM], and granulysin [GNLY]) gradually declined. PD-L1 expression was not associated with age; however, a number of immunosuppressive elements beyond PD-L1 were upregulated in aging patients, including regulatory T cells and co-inhibitory molecules, for example, TIM-3, TIGIT, and HHLA2. Finally, senescence was accompanied by a loss of clonal neoantigens, which is believed to be correlated with responsiveness to immune checkpoint inhibitors. CONCLUSIONS: Elderly patients were characterized by increased numbers of CD8+ T cells and impaired cytolytic molecule expression. The observed immune signature was also associated with a loss of clonal neoantigens and the accumulation of immunosuppressive elements. These findings show a unique immune microenvironment in senescence and support biomarker-guided candidate identification for anti-PD-1/PD-L1 therapeutic strategies for elderly patients with LUAD.

Acetylation Blocks cGAS Activity and Inhibits Self-DNA-Induced Autoimmunity.

The presence of DNA in the cytoplasm is normally a sign of microbial infections and is quickly detected by cyclic GMP-AMP synthase (cGAS) to elicit anti-infection immune responses. However, chronic activation of cGAS by self-DNA leads to severe autoimmune diseases for which no effective treatment is available yet. Here we report that acetylation inhibits cGAS activation and that the enforced acetylation of cGAS by aspirin robustly suppresses self-DNA-induced autoimmunity. We find that cGAS acetylation on either Lys384, Lys394, or Lys414 contributes to keeping cGAS inactive. cGAS is deacetylated in response to DNA challenges. Importantly, we show that aspirin can directly acetylate cGAS and efficiently inhibit cGAS-mediated immune responses. Finally, we demonstrate that aspirin can effectively suppress self-DNA-induced autoimmunity in Aicardi-Goutières syndrome (AGS) patient cells and in an AGS mouse model. Thus, our study reveals that acetylation contributes to cGAS activity regulation and provides a potential therapy for treating DNA-mediated autoimmune diseases.

Tumor Microenvironment Properties are Associated With Low CD68-positive Cell Infiltration and Favorable Disease-free Survival in EGFR-mutant Lung Adenocarcinoma.

BACKGROUND: The benefits of immune checkpoint inhibitors for first-line treatment in patients with lung adenocarcinoma harboring EGFR mutations are unclear. The effects of ICIs depend on the tumor microenvironment (TME). Differences in TME properties between mutant and wild-type EGFR have not been fully characterized. PATIENTS AND METHODS: We collected 105 surgically resected (50 EGFR mutated and 55 EGFR wild-type), treatment-naïve lung adenocarcinoma tissues with clinical data to investigate the landscape and compartmentalization of tumor-infiltrating immune cells with respect to EGFR status by immunohistochemistry. The normalized FPKM values of data for 531 patients were obtained from The Cancer Genome Atlas (TCGA) Data Portal (https://portal.gdc.cancer.gov/). RESULTS: CD68-positive cells within the tumor niche exhibited more intensive infiltration in wild-type EGFR than in mutations, and was related to lymph node invasion. In the RNA-Seq analysis, MMP9 and VEGFA showed higher levels in wild-type EGFR than in mutant cases. The EGFR mutation independently predicted a favorable disease-free survival. CONCLUSION: The CD68-positive cells play a crucial role in discriminating the TME between different EGFR statuses.

CUE domain-containing protein 2 promotes the Warburg effect and tumorigenesis.

Cancer progression depends on cellular metabolic reprogramming as both direct and indirect consequence of oncogenic lesions; however, the underlying mechanisms are still poorly understood. Here, we report that CUEDC2 (CUE domain-containing protein 2) plays a vital role in facilitating aerobic glycolysis, or Warburg effect, in cancer cells. Mechanistically, we show that CUEDC2 upregulates the two key glycolytic proteins GLUT3 and LDHA via interacting with the glucocorticoid receptor (GR) or 14-3-3ζ, respectively. We further demonstrate that enhanced aerobic glycolysis is essential for the role of CUEDC2 to drive cancer progression. Moreover, using tissue microarray analysis, we show a correlation between the aberrant expression of CUEDC2, and GLUT3 and LDHA in clinical HCC samples, further demonstrating a link between CUEDC2 and the Warburg effect during cancer development. Taken together, our findings reveal a previously unappreciated function of CUEDC2 in cancer cell metabolism and tumorigenesis, illustrating how close oncogenic lesions are intertwined with metabolic alterations promoting cancer progression.

A case report: delayed high fever and maculopapules during Sorafenib treatment of ectopic hepatocellular carcinoma.

BACKGROUND: Sorafenib is the standard first-line therapy for hepatocellular carcinoma (HCC) and probably ectopic hepatocellular carcinoma (EHCC) as well. No report involves a side effect of delayed high fever of sorafenib. This manuscript describes a case of EHCC in the thoracic and abdominal cavities, who showed a delayed high fever and maculopapules during sorafenib treatment. CASE PRESENTATION: The patient is a 63-year-old Chinese male with advanced EHCC, taking sorafenib 400 mg twice daily. On the tenth day, red maculopapules appeared all over the body. On the same day, the patient began to suffer from continuous high fever. Due to these effects, the patient was asked to cease sorafenib treatment, and the high fever and maculopapules were alleviated quickly. However, the symptoms were present again upon re-challenge of sorafenib. Prednisone was then administered to control the symptoms, with the dosage gradually reduced from 30 to 5 mg/day in 1.5 months. No recurrence of fever or maculopapules has been found. Tumor response reached partial response (PR) and progression free survival (PFS) reached 392 days + by the date of Apr. 14th, 2016. CONCLUSION: EHCC could be treated like orthotopic HCC by oral administration of sorafenib, which shows good tumor response and survival benefit. Delayed high fever and maculopapules are potential, rare and severe side effects of sorafenib, and could be effectively controlled by glucocorticoid.

microRNA-214 promotes epithelial-mesenchymal transition and metastasis in lung adenocarcinoma by targeting the suppressor-of-fused protein (Sufu).

Distant metastasis is the major cause of cancer-related deaths in patients with lung adenocarcinoma (LAD). Emerging evidence reveals that miRNA is critical for tumor metastasis. miR-214 expression has been associated with LAD progression. However, whether and how miR-214 is involved in the development and metastasis of LAD remain unaddressed. Here, we found that the expression of miR-214 was elevated in LAD and correlated positively with LAD metastasis and epithelial-mesenchymal transition (EMT). In addition, we found that miR-214 enhanced the molecular program controlling the EMT of LAD cells and promoted LAD cell metastasis both in vitro and in vivo. This study thus provides the first evidence to show that the miR-214 expression by LAD cells contributes to the EMT and metastasis of LAD. Mechanistically, Sufu was identified as an important miR-214 functional target for the EMT and metastasis of LAD, ectopic expression of Sufu alleviated miR-214 promoted EMT and metastasis. Importantly, the expression of Sufu inversely correlated with the expression of miR-214 and vimentin and positively associated with the expression of E-cadherin in the tumor cells from human LAD patients. Collectively, this study uncovers a previously unappreciated miR-214-Sufu pathway in controlling EMT and metastasis of LAD and suggests that interfering with miR-214 and Sufu could be a viable approach to treat late stage metastatic LAD patients.

Clear cell tumor of the lung: a case report and literature review.

Clear cell tumor of the lung is a rare and benign pulmonary tumor; only sporadic cases have been reported. Here, we report the case of a 38-year-old man with recurrent cough, blood-streaked sputum and left chest pain. A chest computed tomography scan showed a round, homogeneous pulmonary mass in the left lower lobe, which exhibited intense heterogeneous enhancement in the arterial phase and homogeneous in the delay phase after injecting a contrast agent. The patient underwent a fine-needle aspiration biopsy and was diagnosed as having a benign clear cell tumor of the lung. The clinical presentation and radiographic investigation of this tumor are summarized in this paper to recognize this rare disease. Interestingly, we found some differences with previously reported cases.

Screening biomarkers of prostate cancer by integrating microRNA and mRNA microarrays.

OBJECTIVE: In this study, we screened microRNA (miRNA) target genes of prostate cancer by integrating miRNA and mRNA expression profiles after target prediction and performed function enrichment analysis for selected candidate genes. METHODS: The miRNA expression profile (GSE36802) and mRNA expression profile (GSE36801) were downloaded from the Gene Expression Omnibus database. We processed data and identified the differentially expressed miRNAs and mRNAs with R packages. Verified targets of miRNAs were identified through miRecods and miRTarBase. Then, software of Search Tool for the Retrieval of Interacting Genes was used to construct the interaction network of target genes. Finally, we performed function enrichment analysis for genes in the interaction network with the Functional Classification Tool. RESULTS: A total of 22 upregulated and 8 downregulated miRNAs were detected in this study, of which, hsa-mir-31 was the most overexpressed miRNA in prostate cancer. Both ITGA5 and RDX, two target genes of hsa-mir-31, were found to be differentially expressed from mRNA profiles by overexpressing hsa-mir-31. The cell adhesion molecule was found to be the most significant pathway enriched by ITGA5 and RDX. CONCLUSION: Overexpression of hsa-mir-31 can be a significant marker to distinguish cancer tissues from benign tissues. The targets such as ITGA5 and RDX regulated by hsa-mir-31 are candidate genes of prostate cancer, which provide new treatment strategies for its gene therapy.

Stromal cell-derived factor-1 and vascular endothelial growth factor as biomarkers for lymph node metastasis and poor cancer-specific survival in prostate cancer patients after radical prostatectomy.

OBJECTIVES: This study aims to analyze the clinicopathologic significance of stromal cell-derived factor-1 (SDF-1), vascular endothelial growth factor (VEGF), and matrix metalloproteinase-9 (MMP-9) expression in human prostate cancer (CaP), and their involvement in the prognosis of CaP. MATERIALS AND METHODS: The expression of SDF-1, VEGF, and MMP-9 were measured using immunohistochemistry in 148 CaP patients who underwent radical prostatectomy for clinically localized disease and in 10 samples of benign prostatic hyperplasia (BPH). RESULTS: In the CaP group, VEGF and MMP-9 were more strongly expressed in the tumor cells compared with the BPH group. High intensity SDF-1, VEGF, and MMP-9 stains in tumor areas strongly correlated with lymph node metastasis, pathologic stage, and differentiation. Univariate and multivariate analysis showed that SDF-1, VEGF, and lymph node metastasis were independent prognostic factors for prostate cancer-specific survival. High levels of MMP-9, pathologic stage, and differentiation were associated with prostate cancer-specific survival in univariate analysis but the risk estimate was not significant in multivariate analysis. CONCLUSIONS: High expression levels of SDF-1, VEGF, and MMP-9 are more correlated with lymph node metastatic prostate carcinoma compared with non-lymph-node metastatic cancer. High expression levels of SDF-1 and VEGF strongly predict the biochemical progression in CaP patients after radical prostatectomy.

Regulation of VEGF, MMP-9 and metastasis by CXCR4 in a prostate cancer cell line.

To investigate the mechanisms involved in PCa (prostate cancer) metastasis and CXCR4 (CXC chemokine receptor-4)-mediated VEGF (vascular endothelial growth factor) and MMP-9 (matrix metalloproteinase-9) expression, we used lentivirus-mediated RNAi (RNA interference) to reduce the expression of CXCR4 in a PCa cell line. We found that the silencing of CXCR4 led to a significant down-regulation of VEGF and MMP-9 at both the mRNA and protein levels compared with the control in vitro. Using an animal model, we confirmed that CXCR4 silencing via subcutaneous injection could reduce tumour growth as well as inhibit metastasis, particularly bone metastasis, of PCa. Using in vivo immunohistochemistry, we also found that the expression of VEGF and MMP-9 were reduced by the knockdown of CXCR4 in the primary tumours of mice. Collectively, our results indicate that CXCR4 plays an important role in PCa metastasis through the up-regulation of VEGF and MMP-9. These findings may aid future intervention strategies.

Induction of autophagy-dependent apoptosis by the survivin suppressant YM155 in prostate cancer cells.

In this study, we demonstrated that YM155, a novel survivin suppressant, induced both apoptosis, and autophagy that was shown by conversion of cytosolic-associated protein light chain 3 (LC3I) into autophagosome-associated form (LC3II) and a punctate fluorescence pattern of an ectopic GFP-LC3 protein. The lysosomal inhibitor chloroquine further accumulated YM155-induced LC3II, indicating an increase of autophagic flux. Ectopic expression of survivin significantly attenuated YM155-induced apoptosis and autophagy, whereas survivin siRNA induced autophagy. Furthermore, inhibition of either early or late events of autophagy attenuated YM155-induced apoptosis, demonstrating that induction of autophagy proceeds apoptosis. In conclusion, suppression of survivin by YM155 induces autophagy-dependent apoptosis, and YM155-induced autophagy plays a pro-apoptotic role thereby unveiling a novel mechanism of YM155 in prostate cancer cells.

Role of interleukin-17 in lymphangiogenesis in non-small-cell lung cancer: Enhanced production of vascular endothelial growth factor C in non-small-cell lung carcinoma cells.

Interleukin-17 (IL-17), a potent pro-inflammatory cytokine, plays an active role in inflammation and cancer. Recently, we found that increased IL-17-producing cells correlate with poor survival and increased lymphangiogenesis in non-small-cell lung cancer (NSCLC), but the mechanism is unknown. Here, we show that IL-17 promotes lymphangiogenesis via inducing vascular endothelial growth factor-C (VEGF-C) production by lung cancer cells. We found that IL-17 receptor (IL-17R) is expressed on the surface of Lewis lung carcinoma (LLC) cells but not on lymphatic endothelial cells (LEC). Moreover, LEC chemotaxis and tube formation (measures of net lymphangiogenic potential) were increased by conditioned medium from recombinant mouse IL-17 (rmIL-17)-stimulated LLC but not by rmIL-17. Interleukin-17 increased production of VEGF-C in lung cancer cell lines. The enhanced chemotaxis and endothelial cord formation in the presence of LLC/rmIL-17 was inhibited by addition of recombinant mouse VEGF R3/Fc chimera. Treatment of the A549 cells with rIL-17 significantly increased VEGF-C expression, which was extracellular signal-regulated protein kinase 1/2 (ERK 1/2) dependent. Importantly, we found significant correlations between IL-17 expression, VEGF-C expression and lymphatic vascular density (LVD) in NSCLC. We conclude that IL-17 is involved in lymphangiogenesis in NSCLC by enhancing production of VEGF-C, and IL-17 may be an important target for the treatment of NSCLC.

Increased IL-17-producing cells correlate with poor survival and lymphangiogenesis in NSCLC patients.

The presence of IL-17-positive cells is observed in a variety of inflammatory associated cancers and IL-17 has been found to be involved in angiogenesis. The aim of this study is to determine the prognostic significance of IL-17 in NSCLC patients and to examine the correlation between IL-17 expression and lymphatic vessel density in NSCLC tissues. The expression of IL-17 was measured by immunohistochemistry in 52 paraffin-embedded tissues with non-small cell lung cancer. The chi(2) test was used to analyze the correlation between IL-17 expression and clinical parameters and lymphatic vessel density (LVD). The Kaplan-Meier method, univariate and multivariate regression analysis was used to analyze the correlation between IL-17 expression and overall survival and disease-free survival. High expression of IL-17 was observed in 25 of 52 lung cancer patients and was associated with smoking status, TNM stage, LVD, overall survival and disease-free survival. Univariate and multivariate analysis showed that IL-17 was an independent prognostic factor for overall survival and disease-free survival. Our results indicate that IL-17 may play a role in the metastasis of lung cancer by promoting lymphangiogenesis. IL-17 expression is an independent prognostic factor in both overall and disease-free survival in NSCLC.

The co-transfection of p16(INK4a) and p14(ARF) genes into human lung cancer cell line A549 and the effects on cell growth and chemosensitivity.

Two functionally and structurally different proteins, p16(INK4a) and p14(ARF), encoded by the gene INK4a/ARF located at 9p21 are cyclin-dependent kinase (cdk) inhibitors and important cell cycle regulators. More and more evidences have been accumulated to show that the exogenous p16(INK4a) or p14(ARF) can inhibit the cell growth and/or induce the apoptosis. But it is still unclear if they can play positive role when combine with the conventional chemotherapy in cancer treatment. Here we show that cationic liposome-mediated gene transfection of INK4a/ARF into lung cancer cell line A549, in which the INK4a/ARF locus was lost, suppressed the growth and induced apoptosis. When treated with five different chemotherapy drugs with different mechanism after the transfection, A549 got an increased chemosensitivity for adriamycin and cisplatin and an unchanged result for topotecan, taxol or vinorelbine. The results indicated that cell cycle redistribution and increased apoptosis index after transfection might be the main explanation for the enhanced chemosensitivity. The combination of gene therapy with conventional chemotherapy is not always better than single chemotherapy. This trial will be of benefit to the treatment of lung cancer when combine the conventional chemotherapy and gene therapy in the future.