Functional studies of TP63-affected ESCC tumor immune microenvironment
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ABSTRACT: Our preliminary data from unbiased analyses of both ESCC patient samples and cell lines identified interferon-γ (IFN-γ) signaling pathway as the most significantly enriched pathways suppressed by TP63. To validate the activation of IFN pathways and the immune responses upon silencing of TP63, we utilized immune-competent mice to conduct allograft experiments. To analyze the effect of TP63 on ESCC tumor microenvironment, resected tumors were then collected to perform scRNA-seq.
Project description:To understand the difference of protein expression between paired esophageal squamous cell carcinoma (ESCC) and adjacent normal tissues, we collected 10 paired ESCC and normal tissues from surgical resected specimems for high-throughput proteomic experiments. From comparative analysis, the dysregulated signaling pathways in ESCC could be uncovered.
Project description:To discover ESCC related proteins, we used SWATH to quantify the protein abundance between ESCC and adjacent tissues. Briefly, we pooled 10 ESCCtissues and their corresponding adjacent tissues for SWATH acquisition with three replicates.Three DDA repeats were also acquired with the pooled 10-paired ESCC tissue.The trypsin digested peptide mixture was analyzed by AB SCIEX 5600 (AB SCIEX).The database searching procedure was achieved using ProteinPilot v4.5 (AB Sciex). The database is IPI_homo_sapiens_V3.87.
Project description:The source of IFN-γ in ovarian cancer microenvironment and its biological effect to the tumor cells is unclear. The immortalized human ovarian surface epithelial cell line, HOSE-E7/hTERT (HOSE) was treated with IFN-γ and expression microarray analysis was performed, and probes showing significantly higher values in IFN-γ-added group were termed “IFN-γ signature genes (295 probes)”. We then applied this signature to our ovarian cancer microarray data, which included 75 ovarian cancer clinical samples, by means of ss-GSEA. IFN-γ signature score was strongly correlated to the number of infiltrating CD4-positive or CD8-positive lymphocytes in the tumors. These data suggest that the IFN-γ in the ovarian cancer microenvironment is derived from lymphocytes, and an IFN-γ-rich microenvironment is strongly correlated to a lymphocyte-rich microenvironment.
Project description:Purpose: To identify TP63 expression regulated pathways in HNSCC Methods: A recombinant lentivirus encoding either NS shRNA or TP63 shRNA was introduced into a HNSCC cell line, FaDu. SCCs were gene generated by implanting either FaDu-NS shRNA (n=3) or FaDu-TP63 shRNA into the tongue of athymic nude mice. Tongue SCCs harvested at the end of study were used for transcriptome analysis
Project description:Patients with lymphoma harboring TP63 rearrangements have aggressive clinical course and dismal prognosis with no target therapy available. Thus, there is an urgent need to elucidate the molecular mechanisms and to develop novel therapeutic options for these patients. We then generated a TBL1XR1::TP63 fusion knockin trangenic mouse model and crossed with CD2/iCre mouse. This project is to study the role and function of TP63 fusion in mouse lymphomagenesis.
Project description:Background & Aims: Lineage-specific expression of long non-coding RNAs (lncRNAs) has been observed recently. However, the underlying mechanism of such specific transcription regulation is unclear. The aim of this study is to identify squamous cell carcinoma (SCC) lineage-specific lncRNAs and to investigate the mechanisms for their expression and function. Methods: Expression characteristics and functions of four candidate SCC-specific lncRNAs were explored. qRT-PCR assays were employed to measure the expression of LINC01503 in 113 tumor/normal matched esophageal SCC (ESCC) cases and its association with survival was determined. The mechanisms underlying LINC01503 function and regulation in ESCC cells were examined using molecular biological methods. Results: Using SCC as a model, we identified a novel super-enhancer (SE)-driven lncRNA, LINC01503, which was uniquely expressed in SCCs such as those from esophagus (ESCC) and head and neck (HNSC). LINC01503 was up-regulated in SCCs and its high expression correlated with poor clinical outcomes. SCC master transcriptional factor TP63 directly bound to a SE at LINC01503 locus and activated its transcription. LINC01503 exhibited strong oncogenic functions in ESCC cell models both in vitro and in vivo. ERK2 and EBP-1 were identified as interacting proteins mediating the effects of LINC01503. Specifically, LINC01503 protected ERK2 from dephosphorylation by DUSP-6, leading to the activation of ERK/MAPK pathway. Similarly, LINC01503 interfered the interaction between EBP-1 and PI3K p85, enhancing Akt signaling pathway. Conclusions: These results demonstrated that LINC01503 is a lineage-specific oncogene in ESCC, which may serve as a potential biomarker and therapeutic target for SCC patients.
Project description:Background & Aims: Lineage-specific expression of long non-coding RNAs (lncRNAs) has been observed recently. However, the underlying mechanism of such specific transcription regulation is unclear. The aim of this study is to identify squamous cell carcinoma (SCC) lineage-specific lncRNAs and to investigate the mechanisms for their expression and function. Methods: Expression characteristics and functions of four candidate SCC-specific lncRNAs were explored. qRT-PCR assays were employed to measure the expression of LINC01503 in 113 tumor/normal matched esophageal SCC (ESCC) cases and its association with survival was determined. The mechanisms underlying LINC01503 function and regulation in ESCC cells were examined using molecular biological methods. Results: Using SCC as a model, we identified a novel super-enhancer (SE)-driven lncRNA, LINC01503, which was uniquely expressed in SCCs such as those from esophagus (ESCC) and head and neck (HNSC). LINC01503 was up-regulated in SCCs and its high expression correlated with poor clinical outcomes. SCC master transcriptional factor TP63 directly bound to a SE at LINC01503 locus and activated its transcription. LINC01503 exhibited strong oncogenic functions in ESCC cell models both in vitro and in vivo. ERK2 and EBP-1 were identified as interacting proteins mediating the effects of LINC01503. Specifically, LINC01503 protected ERK2 from dephosphorylation by DUSP-6, leading to the activation of ERK/MAPK pathway. Similarly, LINC01503 interfered the interaction between EBP-1 and PI3K p85, enhancing Akt signaling pathway. Conclusions: These results demonstrated that LINC01503 is a lineage-specific oncogene in ESCC, which may serve as a potential biomarker and therapeutic target for SCC patients.
Project description:In response to genotoxic stress the TP53 tumour suppressor activates target gene expression to induce cell cycle arrest or apoptosis depending on the extent of DNA damage. These canonical activities can be repressed by TP63 in normal stratifying epithelia to maintain proliferative capacity or drive proliferation of squamous cell carcinomas, where TP63 is frequently overexpressed/amplified. Here we use ChIP-sequencing, integrated with microarray analysis, to define the genome wide interplay between TP53 and TP63 in response to genotoxic stress in normal cells. We reveal that TP53 and TP63 bind to overlapping, but distinct cistromes of sites through utilization of distinctive consensus motifs and that TP53 is constitutively bound to a number of sites. We demonstrate that cisplatin and adriamycin elicit distinct effects on TP53 and TP63 binding events, through which TP53 can induce or repress transcription of an extensive network of genes by direct binding and/or modulation of TP63 activity. Collectively, this results in a global TP53 dependent repression of cell cycle progression, mitosis and DNA damage repair concomitant with activation of anti-proliferative and pro-apoptotic canonical target genes. Further analyses reveals that in the absence of genotoxic stress TP63 plays an important role in maintaining expression of DNA repair genes, loss of which results in defective repair Examination of p63 and p53 binding sites in neonatal foreskin keratinocytes in response to adriamycin or cisplatin treatment
Project description:We detected DNA methylation of 15 ESCC samples' tumor tissues and their pericarcinomatous tissues to identify the aberrant DNA methylation status in ESCC.