Transforming Growth Factor Beta Induced Expression Research Articles

Integrated Analysis Identified TGFBI as a Biomarker of Disease Severity and Prognosis Correlated with Immune Infiltrates in Patients with Sepsis.

Sepsis is a major contributor to morbidity and mortality among hospitalized patients. This study aims to identify markers associated with the severity and prognosis of sepsis, providing new approaches for its management and treatment. Data were mined from the Gene Expression Omnibus (GEO) databases and were analyzed by multiple statistical methods like the Spearman correlation coefficient, Kaplan-Meier analysis, Cox regression analysis, and functional enrichment analysis. Candidate indicator' associations with immune infiltration and roles in sepsis development were evaluated. Additionally, we employed techniques such as flow cytometry and neutral red staining to evaluate its impact on macrophage functions like polarization and phagocytosis. Twenty-eight genes were identified as being closely linked to the severity of sepsis, among which transforming growth factor beta induced (TGFBI) emerged as a distinct marker for predicting clinical outcomes. Notably, reductions in TGFBI expression during sepsis correlate with poor prognosis and rapid disease progression. Elevated expression of TGFBI has been observed to mitigate abnormalities in sepsis-related immune cell infiltration that are critical to the pathogenesis and prognosis of the disease, including but not limited to type 17 T helper cells and activated CD8 T cells. Moreover, the protein-protein interaction network revealed the top ten genes that interact with TGFBI, showing significant involvement in the regulation of the actin cytoskeleton, extracellular matrix-receptor interactions, and phagosomes. These are pivotal elements in the formation of phagocytic cups by macrophages, squaring the findings of the Human Protein Atlas. Additionally, we discovered that TGFBI expression was significantly higher in M2-like macrophages, and its upregulation was found to inhibit lipopolysaccharide-induced polarization and phagocytosis in M1-like macrophages, thereby playing a role in preventing the onset of inflammation. TGFBI warrants additional exploration as a promising biomarker for assessing illness severity and prognosis in patients with sepsis, considering its significant association with immunological and inflammatory responses in this condition.

LncRNA H19 inhibits proliferation and enhances apoptosis of nephroblastoma cells by regulating the miR-675/TGFBI axis.

To investigate the influences of long non-coding ribonucleic acid (lncRNA) H19 on proliferation and apoptosis of nephroblastoma cells. A total of 5 pairs of nephroblastoma tissues and paraneoplastic tissues were obtained. Gene expression levels of lncRNA H19, microRNA (miR)-675, and transforming growth factor beta induced (TGFBI) were detected via quantitative Reverse Transcription-Polymerase Chain Reaction (qRT-PCR). Their regulatory effects on the viability of nephroblastoma cells were examined by Cell Counting Kit-8 (CCK-8) assay. Finally, the apoptosis level in each group was detected through TUNEL assay, and the protein expressions of TGFBI and Caspase-8 were examined using Western blotting (WB) assay. The gene expression levels of lncRNA H19 and miR-675 were markedly downregulated in nephroblastoma tissues (p<0.05), while that of TGFBI was notably upregulated (p<0.05). LncRNA H19 could reduce the proliferative ability of HFWT cells (p<0.05) and stimulates apoptosis rate (p<0.05). It upregulated the expressions of miR-675 and Caspase-8 (p<0.05), and downregulated TGFBI (p<0.05). Besides, miR-675 was able to upregulate Caspase-8 (p<0.05) and downregulate TGFBI (p<0.05). In addition, the protein expression of Caspase-8 was downregulated (p<0.05), while that of TGFBI was upregulated (p<0.05) after the knockdown of miR-675 in HFWT cells. LncRNA H19 may inhibit TGFBI expression by regulating miR-675 level, so as to weaken the proliferation and enhance the apoptosis of nephroblastoma cells.

Synergistic inhibitory effects of 5-aza-2'-deoxycytidine and cisplatin on urothelial carcinoma growth via suppression of TGFBI-MAPK signaling pathways.

This study aimed to reveal the gene transcriptional alterations, possible molecular mechanisms, and pathways involved in the synergy of 5-aza-2'-deoxycytidine (DAC) and Cisplatin (CDDP) in urothelial carcinoma (UC). Two UC cell lines, 5637 and T24, were used in the present study. A cDNA microarray was performed to identify critical genes involved in the synergistic mechanism of both agents against UC cells. The results showed that several key regulatory genes, such as interleukin 24 (IL24), fibroblast growth factor 1 (FGF1), and transforming growth factor beta induced (TGFBI), may play critical roles in the synergy of DAC and CDDP in UC. Pathway enrichment suggested that many carcinogenesis-related pathways, such as the ECM-receptor interaction and MAPK signaling pathways, may participate in the synergy of both agents. Our results suggest that TGF-β1 stimulates the phosphorylation of ERK1/2 and p38 by increasing TGFBI expression, and that the TGFBI-MAPK signaling pathway plays an important role in the synergy of DAC and CDDP against UC. Therefore, we revealed the synergistic mechanism of DAC and CDDP in UC. Several key regulatory genes play critical roles in the synergy of combined treatment, and the TGFBI-MAPK signaling pathway may be an important potential target of these two agents.

Pan-Cancer Analysis of the Associations of TGFBI Expression With Prognosis and Immune Characteristics.

Transforming growth factor-beta-induced (TGFBI) protein has important roles in tumor growth, metastasis, and immunity. However, there is currently no pan-cancer evidence regarding TGFBI. In this study, we conducted a pan-cancer analysis of TGFBI mRNA and protein expression and prognoses of various cancer types using public databases. We also investigated the associations of TGFBI expression with tumor microenvironment (TME) components, immune cell infiltration, tumor mutational burden (TMB), and microsatellite instability (MSI), along with the TGFBI genetic alteration types. The results showed that TGFBI expression varied among different cancer types, and it was positively or negatively related to prognosis in various cancers. TGFBI expression was also significantly correlated with TME components, TMB, MSI, immune cell infiltration, and immunoinhibitory and immunostimulatory gene subsets. These findings indicate that TGFBI participates in various immune responses and it may function as a prognostic marker in various cancers. The findings may be useful for developing immunotherapies that target TGFBI.

Pan-Cancer Analysis of the Associations of TGFBI Expression with Prognosis and Immune Characteristics

Transforming growth factor-beta-induced (TGFBI) protein has important roles in tumor growth, metastasis, and immunity. However, there is currently no pan-cancer evidence regarding TGFBI. Here, we conducted a pan-cancer analysis of TGFBI mRNA and protein expression and prognoses of various cancer types using public databases. We also explored the associations of TGFBI expression with tumor microenvironment (TME) components, tumor mutational burden (TMB), microsatellite instability (MSI), and immune cell infiltration, along with the TGFBI genetic alteration types. The results showed that TGFBI expression varied among different cancer types, and it was positively or negatively related to prognosis in various cancers. TGFBI expression was also significantly correlated with TME components, TMB, MSI, immune cell infiltration, and immunoinhibitory and immunostimulatory gene subsets. These findings indicate that TGFBI participates in various immune responses and it may function as a prognostic marker in various cancers. The findings may be useful for developing immunotherapies that target TGFBI.

TGFBI modulates tumour hypoxia and promotes breast cancer metastasis.

Breast cancer metastasis is a complex process that depends not only on intrinsic characteristics of metastatic stem cells, but also on the particular microenvironment that supports their growth and modulates the plasticity of the system. In search for microenvironmental factors supporting cancer stem cell (CSC) growth and tumour progression to metastasis, we here investigated the role of the matricellular protein transforming growth factor beta induced (TGFBI) in breast cancer. We crossed the MMTV-PyMT model of mammary gland tumorigenesis with a TgfbiΔ/Δ mouse and studied the CSC content of the tumours. We performed RNAseq on wt and ko tumours, and analysed the tumour vasculature and the immune compartment by IHC and FACS. The source of TGFBI expression was determined by qPCR and by bone marrow transplantation experiments. Finally, we performed insilico analyses using the METABRIC cohort to assess the potential prognostic value of TGFBI. We observed that deletion of Tgfbi led to a dramatic decrease in CSC content and lung metastasis. Our results show that lack of TGFBI resulted in tumour vessel normalisation, with improved vessel perfusion and decreased hypoxia, a major factor controlling CSCs and metastasis. Furthermore, human data mining in a cohort of breast cancer patients showed that higher expression of TGFBI correlates with poor prognosis and is associated with the more aggressive subtypes of breast cancer. Overall, these data reveal a novel biological mechanism controlling metastasis that could potentially be exploited to improve the efficacy and delivery of chemotherapeutic agents in breast cancer.

HMGB1: an important regulator of myeloid differentiation and acute myeloid leukemia as well as a promising therapeutic target

High mobility group box 1 (HMGB1) is a non-histone nuclear protein which has been intensively studied in various physiological and pathological processes including leukemia. Here in this study, we further demonstrated that HMGB1 presents higher expression in the bone marrow mononuclear cells of acute myeloid leukemia (AML) patients compared with the normal controls and contributes to the AML pathogenesis and progression by inhibiting apoptosis, facilitating proliferation, and inducing myeloid differentiation blockade of AML cells. Mechanistic investigation revealed that transforming growth factor beta-induced (TGFBI) acts as a potential downstream target of HMGB1 and lentivirus-mediated knockdown of TGFBI expression impaired phorbol-12-myristate-13-acetate (PMA) and all-trans retinoic acid (ATRA)–induced myeloid differentiation of AML cell lines. On the other hand, chidamide, an orally histone deacetylase inhibitor, decreases HMGB1 expression significantly in AML cells with concomitant upregulation of TGFBI expression, and confers therapeutic effect on AML by inducing cell differentiation, apoptosis and inhibiting cell proliferation. In conclusion, our findings provide additional insights that HMGB1 is a promising therapeutic target of AML, and also present experimental evidence for the clinical application of chidamide as a novel agent in AML therapy by downregulating HMGB1 expression.Key messagesHMGB1 induces cell proliferation and myeloid differentiation blockade and inhibits apoptosis of AML cells.TGFBI acts as a potential target of HMGB1.Chidamide, a selective HDAC inhibitor, confers promising therapeutic effect for AML via downregulating HMGB1 expression.

Comprehensively investigating the expression levels and the prognostic role of transforming growth factor beta-induced (TGFBI) in glioblastoma multiforme.

BackgroundTransforming growth factor beta-induced (TGFBI) protein has been found expressed in several cancer types, and expression levels of TGFBI can affect the cancer patients’ outcomes, but the role of TGFBI in glioblastoma multiforme (GBM) remains obscure.MethodsThe TGFBI expression levels in GBM were performed via Gene Expression Profiling Interactive Analysis (GEPIA) and UALCAN databases. Further, the mutations types of TGFBI were analyzed by using the cBioportal dataset. LinkedOmics selected correlated genes, kinases, and microRNA (miRNA) targets of TGFBI. GEPIA conducted the prognostic value of TGFBI and correlated genes. Then, the relationship between TGFBI and immune infiltrates was performed by Tumor Immune Estimation Resource (Timer). We compared the TGFBI protein expression levels in GBM and control samples through the Human Protein Atlas (HPA). Finally, the GSCAlite was used to achieve the drugs, and molecules target the TGFBI and significantly correlated genes.ResultsTGFBI is significantly overexpressed in GBM, but the clinical features do not have considerable influence on TGFBI expression levels. Overexpression of TGFBI acts as an adverse biomarker of GBM. The enrichment function of TGFBI showed that the main biological functions, including extracellular matrix (ECM) organization, angiogenesis, leukocyte migration, T cell activation, cell cycle G2/M phase transition, and growth factor binding. About the significant correlated genes, overexpression of mitogen-activated protein kinase 13 (MAPK13) [Log-rank P=0.08 HR (high) =1.4], myosin IG (MYO1G) [Log-rank P=0.06 HR (high) =1.4], plasminogen activator urokinase receptor (PLAUR) [Log-rank P=0.03 HR (high) =1.5], thrombomodulin (THBD) [Log-rank P=0.028 HR (high) =1.5] indicated the poor prognosis of GBM. Further, TGFBI had a significant association with dendritic cell (DC) infiltrates (cor =0.516, P=9.00e−30). The higher the DC infiltration, the shorter survival of GBM. TGFBI protein expression levels were not significantly different in GBM and normal tissue. Finally, TGFBI is associated with resistance to belinostat, LAQ824, CAY10603, CUDC-101, methotrexate, 5-fluorouracil, and navitoclax.ConclusionsIn the present study, we showed TGFBI was overexpressed in GBM, and TGFBI is associated with DC cell infiltrates. Overexpression of TGFBI and high DC infiltration might be an adverse biomarker of GBM. Finally, TGFBI is associated with tumor multi-drug resistance.

Identification of transforming growth factor beta induced (TGFBI) as an immune-related prognostic factor in clear cell renal cell carcinoma (ccRCC).

Clear cell renal cell carcinoma (ccRCC) is the most common subtype among kidney cancer, which has poor prognosis. The aim of this study was to screen out novel prognostic biomarkers and therapeutic targets for immunotherapy, and some novel molecule drugs for ccRCC treatment. Immune scores ranged from -1109.36 to 2920.81 and stromal scores ranged from -1530.11 to 1955.39 were firstly calculated by applying ESTIMATE algorithm. Then 17 DEGs associated with immune score and stromal score were further identified. 6 candidate hub genes were screened out by performing overall survival (OS) and disease-free survival analyses based on TCGA-KIRC data, one of which including TGFBI was further regarded as hub gene associated with prognosis by calculating the R2 (R2 = 0.011, P = 0.018) and AUC (AUC = 0.874). The prognostic value of TGFBI was validated by performing OS, CSS, and PFS analyses based on GSE29609 and E-MTAB-3267. CMap analysis suggested that 3 molecule drugs might be novel choice for ccRCC treatment. Further analysis demonstrated that CNVs of TGFBI was associated with OS of patients with ccRCC. TGFBI expression was also correlated with histologic grade, pathologic stage, and immune infiltration level, significantly. TGFBI was the most relevant gene with OS among the candidate hub genes, which might be novel DNA methylation biomarkers for ccRCC. In conclusion, our findings indicated that TGFBI was correlated with prognosis of patients with ccRCC, which might be novel prognostic biomarkers, and targets for immunotherapy in ccRCC. Three small molecule drugs were also identified, which showed strong potential for ccRCC treatment.

Tumor-associated macrophages promote ovarian cancer cell migration by secreting transforming growth factor beta induced (TGFBI) and tenascin C

A central and unique aspect of high-grade serous ovarian carcinoma (HGSC) is the extensive transcoelomic spreading of tumor cell via the peritoneal fluid or malignant ascites. We and others identified tumor-associated macrophages (TAM) in the ascites as promoters of metastasis-associated processes like extracellular matrix (ECM) remodeling, tumor cell migration, adhesion, and invasion. The precise mechanisms and mediators involved in these functions of TAM are, however, largely unknown. We observed that HGSC migration is promoted by soluble mediators from ascites-derived TAM, which can be emulated by conditioned medium from monocyte-derived macrophages (MDM) differentiated in ascites to TAM-like asc-MDM. A similar effect was observed with IL-10-induced alternatively activated m2c-MDM but not with LPS/IFNγ-induced inflammatory m1-MDM. These observations provided the basis for deconvolution of the complex TAM secretome by performing comparative secretome analysis of matched triplets of different MDM phenotypes with different pro-migratory properties (asc-MDM, m2c-MDM, m1-MDM). Mass spectrometric analysis identified an overlapping set of nine proteins secreted by both asc-MDM and m2c-MDM, but not by m1-MDM. Of these, three proteins, i.e., transforming growth factor beta-induced (TGFBI) protein, tenascin C (TNC), and fibronectin (FN1), have been associated with migration-related functions. Intriguingly, increased ascites concentrations of TGFBI, TNC, and fibronectin were associated with short progression-free survival. Furthermore, transcriptome and secretome analyses point to TAM as major producers of these proteins, further supporting an essential role for TAM in promoting HGSC progression. Consistent with this hypothesis, we were able to demonstrate that the migration-inducing potential of asc-MDM and m2c-MDM secretomes is inhibited, at least partially, by neutralizing antibodies against TGFBI and TNC or siRNA-mediated silencing of TGFBI expression. In conclusion, the present study provides the first experimental evidence that TAM-derived TGFBI and TNC in ascites promote HGSC progression.

PPAR Gamma-Regulated MicroRNA 199a-5p Underlies Bone Marrow Adiposity in Aplastic Anemia

Increased propensity of bone marrow-derived mesenchymal stem cells (BM-MSCs) toward adipogenic differentiation has been implicated in the fatty bone marrow and defective hematopoiesis of aplastic anemia (AA). However, the underlying molecular mechanism remains to be investigated. In this study, we found that microRNA 199a-5p (miR-199a-5p) exhibits significantly higher expression in AA BM-MSCs compared with the normal control and is demonstrated to facilitate adipogenic differentiation of BM-MSCs through lentivirus-mediated miR-199a overexpression. Mechanistic investigation reveals that miR-199a-5p could be regulated by PPAR gamma (PPARγ) in a transcription-independent manner and regulates adipogenic differentiation by targeting the expression of transforming growth factor beta induced (TGFBI), which is subsequently validated as a negative regulator of adipogenesis. Besides, the positive correlation between PPARγ and miR-199a-5p expression as well as the inverse relationship between miR-199a-5p and TGFBI expression in normal and AA BM-MSCs was observed. Altogether, our work demonstrates that PPARγ-regulated miR-199a-5p promotes adipogenesis of BM-MSCs by inhibiting TGFBI expression, which might be a novel mechanism underlying the bone marrow adiposity in AA, and provides promising therapeutic targets for AA treatment.

Secreted TGF-beta-induced protein promotes aggressive progression in bladder cancer cells.

Background: Transforming growth factor-beta-induced (TGFBI) is an exocrine protein, which has been found to be able to promote the development of nasopharyngeal carcinoma, glioma, pancreatic cancer, and other tumors. However, there is currently no report concerning the relationship between TGFBI and invasive progression of bladder cancer (BCa).Methods: IHC staining, qRT-PCR and Western blot were used to analyze TGFBI and EMT markers levels. In vivo tumorigenesis was performed by xenograft tumor model.Results: In this study, we found that both mRNA and protein levels of TGFBI were significantly up-regulated in muscle invasive bladder cancer (MIBC) tissues compared with non-muscle-invasive bladder cancer (NMIBC) tissues. The high expression level of TGFBI was positively correlated with high histological grade and advanced clinical stage, and BCa patients with high TGFBI levels exhibited poor prognoses. We further confirmed that high expression level of TGFBI can promote proliferation, invasive progression, and epithelial-to-mesenchymal transition (EMT) of BCa cells in vitro, as well as promote tumor growth and EMT in vivo, while silencing of TGFBI inhibited these malignant phenotypes. TGFBI was involved in the up-regulation of EMT by inducing the expression level of Slug, Vimentin, Snail, MMP2, and MMP9 genes, while it down-regulated the expression level of E-cadherin. Moreover, Western blot analysis was carried out to demonstrate that BCa cell lines stably transfected with expression of TGFBI, a secreted protein. Furthermore, conditional medium containing TGFBI protein also resulted in enhanced EMT and malignant phenotype of BCa cells.Conclusion: Our results indicate that high expression level of TGFBI promotes EMT, proliferation, and invasive progression of BCa cells, and TGFBI is a potential therapeutic target and prognostic marker for BCa.

Abstract 163: Identification and characterization of TGFBI in circulating tumor cell subline from pancreatic cancer cell line

Abstract Distant metastasis to liver, lung, brain, or bone occurs via circulating tumor cells (CTCs). We hypothesized that a subset of CTCs had more malignant features than tumor cells at the primary site. We established a highly malignant cell line, Panc-1-CTC, derived from the human pancreatic cancer cell line Panc-1 (Panc-1-P) using an in vivo selection method. Panc-1-CTC cells exhibited more profound migratory and invasive abilities than Panc-1-P cells in vitro. In addition, Panc-1-CTC cells had a higher tumor-forming ability than Panc-1-P cells in vivo. To investigate whether a difference in malignant phenotypes exists between Panc-1-P and -CTC cells, we performed comprehensive gene expression array analysis. As a result, Panc-1-CTC significantly expressed transforming growth factor beta-induced (TGFBI), an extracellular matrix protein, more abundantly than did Panc-1-P cells. TGFBI is considered to regulate cell adhesion, but its functions remain unclear. In the present study, knockdown of TGFBI reduced cell migration and invasion abilities, whereas overexpression of TGFBI increased both abilities. Moreover, elevated expression of TGFBI was associated with poor prognosis in patients with pancreatic cancer. Taken together, TGFBI might be targeted for cancer therapy and its expression could be a biomarker for the prediction of prognosis in pancreatic cancer. Citation Format: Tomoki Muramatsu, Taku Sato, Minoru Tanabe, Johji Inazawa. Identification and characterization of TGFBI in circulating tumor cell subline from pancreatic cancer cell line [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 163.

TGFBI Promotes Tumor Growth and is Associated with Poor Prognosis in Oral Squamous Cell Carcinoma.

Purpose: In a previous study, we found that transforming growth factor beta-induced (TGFBI) is a hub gene strongly associated with oral squamous cell carcinoma (OSCC), using gene chip meta-analysis and PPI network analysis. Thus, the present study was established to explore the role of TGFBI in the pathogenesis of OSCC and to define the underlying mechanisms.Methods: The correlations between TGFBI expression and the clinicopathological features and prognosis of OSCC were analyzed. Then, TGFBI-knockout HSC-3 cell lines were constructed using the CRISPR/Cas9 system. Cell proliferation, migration, and invasion in vitro were determined by cell counting, CCK-8, colony formation, and Transwell assays. Moreover, a xenograft animal study was implemented to determine the tumorigenicity and metastatic ability associated with TGFBI in vivo. The genes and pathways differentially expressed after TGFBI knockout were determined using transcriptional sequencing and bioinformatics.Results: TGFBI expression was significantly higher in OSCC than in normal tissue. Its high expression was also correlated with high stage and was predictive of poor prognosis, as we expected. Knockout of TGFBI inhibited cell proliferation and clone formation, and enhanced cell migration and invasion in vitro. Besides, the xenograft animal study showed that TGFBI knockout suppressed tumor growth and metastasis in vivo. Furthermore, transcriptome sequencing revealed that genes associated with cell proliferation, metastasis, and inflammatory responses exhibited a change of expression upon TGFBI knockout. GO and KEGG analyses indicated that the function of TGFBI is related to responses to bacteria and inflammatory responses.Conclusions: TGFBI overexpression can promote OSCC and is associated with poor prognosis in OSCC patients. TGFBI knockout can inhibit cell proliferation and metastasis in vivo. TGFBI may alter cell responses to bacteria, which causes an imbalance in the immune inflammatory response and promotes the development of OSCC.

Identification and characterization of transforming growth factor beta-induced in circulating tumor cell subline from pancreatic cancer cell line.

Distant metastasis to liver, lung, brain, or bone occurs by circulating tumor cells (CTC). We hypothesized that a subset of CTC had features that are more malignant than tumor cells at the primary site. We established a highly malignant cell line, Panc‐1‐CTC, derived from the human pancreatic cancer cell line Panc‐1 using an in vivo selection method. Panc‐1‐CTC cells showed greater migratory and invasive abilities than its parent cell line in vitro. In addition, Panc‐1‐CTC cells had a higher tumor‐forming ability than parent cells in vivo. To examine whether a difference in malignant phenotypes exists between Panc‐1‐CTC cells and parent cells, we carried out comprehensive gene expression array analysis. As a result, Panc‐1‐CTC significantly expressed transforming growth factor beta‐induced (TGFBI), an extracellular matrix protein, more abundantly than did parent cells. TGFBI is considered to regulate cell adhesion, but its functions remain unclear. In the present study, knockdown of TGFBI reduced cell migration and invasion abilities, whereas overexpression of TGFBI increased both abilities. Moreover, elevated expression of TGFBI was associated with poor prognosis in patients with pancreatic cancer.

Transforming growth factor beta induced (TGFBI) is a potential signature gene for mesenchymal subtype high-grade glioma.

Previous study revealed that higher expression of transforming growth factor beta induced (TGFBI) is correlated to poorer cancer-specific survival and higher proportion of tumor necrosis and Fuhrman grades III and IV in clear cell renal cell carcinomas. However, the relationships between TGFBI expression and malignant phenotypes of gliomas remain unclear. We downloaded and analyzed data from seven GEO datasets (GSE68848, GSE4290, GSE13041, GSE4271, GSE83300, GSE34824 and GSE84010), the TCGA database and the REMBRANDT database to investigate whether TGFBI could be a biomarker of glioma. From microarray data (GSE68848, GSE4290) and RNA-seq data (TCGA), TGFBI expression levels were observed to correlate positively with pathological grade, and TGFBI expression levels were significantly higher in gliomas than in normal brain tissues. Furthermore, in GSE13041, GSE4271 and the TCGA cohort, TGFBI expression in the mesenchymal (Mes) subtype high-grade glioma (HGG) was significantly higher than that in the proneural subtype. Kaplan-Meier survival analysis of GBM patients in the GSE83300 dataset, REMBRANDT and TCGA cohort revealed that patients in the top 50% TGFBI expression group survived for markedly shorter periods than those in the bottom 50%. Analysis of grade III gliomas showed that the median survival time was significantly shorter in the TGFBI high expression group than in the TGFBI low expression group. In addition, we found that TGFBI expression levels might relate to several classical molecular characterizations of glioma, such as, IDH mutation, TP53 mutation, EGFR amplification, etc. These results suggest that TGFBI expression positively correlates with glioma pathological grades and that TGFBI is a potential signature gene for Mes subtype HGG and a potential prognostic molecule.

Abstract 5038: The repression of the extracellular matrix protein TGFBI induces chemoresistance in nasopharyngeal carcinoma

Abstract Introduction: Nasopharyngeal carcinoma (NPC) is a malignancy that is strongly associated with Epstein-Barr virus (EBV). The five-year overall survival (OS) rate of locally-advanced NPC patients is ∼65%. Therapeutic options for such patients are limited; ionizing radiation is the primary mode of therapy for early-stage NPC, while chemoradiotherapy is used for more advanced stages. The use of chemotherapy (combined with radiation) provides a modest benefit in OS, but causes significant toxicities. The major cause of death in NPC is distant metastasis (DM), which remains a clinical challenge. In order to identify novel mechanisms underlying this process, our laboratory has recently completed global RNA sequencing of NPC samples which were fully clinically annotated. The extracellular matrix protein TGFBI (Transforming Growth Factor, Beta-Induced) was identified as a potential marker for chemoresistance. The objective of this study was to elucidate the role of TGFBI, and acquire greater insight into the biological pathways leading to chemoresistance and contributing to DM in NPC. Methods: Nasopharyngeal cell lines including NP69 (normal nasopharyngeal, SV-40 immortalized), and C666-1 (NPC, EBV positive) were used to assess the differential expression of TGFBI between normal and malignant cells. TGFBI under-expressing NP69 cells were engineered using shRNA vectors, and the impact of the loss of TGFBI expression was assessed by cell death, clonogenic, and migration assays. Downstream target gene expression was evaluated by quantitative real-time PCR (qRT-PCR) and Western blot. C666-1 cells, which had no expression of TGFBI (RNA or protein), were used as controls. Results: RNA-seq data identified poorer distant relapse-free survival in chemoradiotherapy-treated patients with low TGFBI expression vs. those with high TGFBI expression. In concordance with this observation, in vitro down-regulation of TGFBI induced cisplatin resistance via the phosphorylation of Akt and PTEN. TGFBI downregulation also increased cellular migration in spheroid cultures. Furthermore, TGFBI was identified to be a target of miR-449b, one of the 4 microRNAs in a DM prognostic signature that was previously identified by our group (Oncotarget 6:4537, 2015). In turn, over-expression of miR-449b in nasopharyngeal cells increased phospho-Akt expression, phenocopying the results observed in the TGFBI experiments. Conclusion: Together, these data suggest that the miR-449∼TGFBI-Akt axis plays a significant role in both treatment resistance and metastasis in NPC. Further studies will allow us to elucidate additional components of this pathway, and identify novel therapeutic targets which can benefit future patients with NPC. Citation Format: Pierre-Antoine Bissey, Jeff W. Bruce, Jacqueline Law, Kenneth W. Yip, Fei-Fei Liu. The repression of the extracellular matrix protein TGFBI induces chemoresistance in nasopharyngeal carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5038.

Identification and validation of TGFBI as a promising prognosis marker of clear cell renal cell carcinoma

ObjectiveTo identify prognostic biomarkers in clear cell renal cell carcinoma (ccRCC) using a proteomic approach. Material and methodsWe performed a comparative proteomic profiling of ccRCC and normal renal tissues from 9 different human specimens. We assessed differential protein expression by iTRAQ (isobaric tagging reagent for absolute quantify) labeling with regard to tumor aggressiveness according to the stage, size, grade, and necrosis (SSIGN) score and confirmed our results using Western blot (9 patients) and immunohistochemistry (135 patients) analysis. ResultsAfter proteomic analysis, 928 constitutive proteins were identified. Among these proteins, 346 had a modified expression in tumor compared with that of normal tissue. Pathway and integrated analyses indicated the presence of an up-regulation of the pentose phosphate pathway in aggressive tumors. In total, 14 proteins were excreted and could potentially become biomarkers. Overexpression of transforming growth factor, beta-induced (TGFBI) in ccRCC was confirmed using Western blot and immunohistochemistry analysis. A significant association was found between the presence of TGFBI expression with tumor category T3–4 (P<0.0001), Fuhrman grades III and IV (P<0.0001), tumor size>4cm (P<0.0001), presence of tumor necrosis (P<0.0001), nodal involvement (n = 0.009), metastasis (P = 0.012), SSIGN score≥5 (P<0.0001), cancer progression (P<0.0001), and cancer-specific death (P<0.0001). Cancer-specific survival was significantly better for patients with no cytoplasmic TGFBI expression (1-, 3-, 5-y cancer-specific survival of 94.7%, 87.8%, and 73.4% vs. 92.9%, 71.2%, and 49.8%, respectively; P<0.0001). ConclusionWe identified 346 proteins involved in renal carcinogenesis and confirmed the presence of a metabolic shift in aggressive tumors. TGFBI was overexpressed in tumors with high SSIGN scores and was significantly associated with oncologic outcomes.

β3 integrin modulates transforming growth factor beta induced (TGFBI) function and paclitaxel response in ovarian cancer cells

BackgroundThe extracellular matrix (ECM) has a key role in facilitating the progression of ovarian cancer and we have shown recently that the secreted ECM protein TGFBI modulates the response of ovarian cancer to paclitaxel-induced cell death.ResultsWe have determined TGFBI signaling from the extracellular environment is preferential for the cell surface αvß3 integrin heterodimer, in contrast to periostin, a TGFBI paralogue, which signals primarily via a ß1 integrin-mediated pathway. We demonstrate that suppression of ß1 integrin expression, in ß3 integrin-expressing ovarian cancer cells, increases adhesion to rTGFBI. In addition, Syndecan-1 and −4 expression is dispensable for adhesion to rTGFBI and loss of Syndecan-1 cooperates with the loss of ß1 integrin to further enhance adhesion to rTGFBI. The RGD motif present in the carboxy-terminus of TGFBI is necessary, but not sufficient, for SKOV3 cell adhesion and is dispensable for adhesion of ovarian cancer cells lacking ß3 integrin expression. In contrast to TGFBI, the carboxy-terminus of periostin, lacking a RGD motif, is unable to support adhesion of ovarian cancer cells. Suppression of ß3 integrin in SKOV3 cells increases resistance to paclitaxel-induced cell death while suppression of ß1 integrin has no effect. Furthermore, suppression of TGFBI expression stimulates a paclitaxel resistant phenotype while suppression of fibronectin expression, which primarily signals through a ß1 integrin-mediated pathway, increases paclitaxel sensitivity.ConclusionsTherefore, different ECM components use distinct signaling mechanisms in ovarian cancer cells and in particular, TGFBI preferentially interacts through a ß3 integrin receptor mediated mechanism to regulate the response of cells to paclitaxel-induced cell death.

Massively Parallel Signature Sequencing and Bioinformatics Analysis Identifies Up-Regulation of TGFBI and SOX4 in Human Glioblastoma

BackgroundA comprehensive network-based understanding of molecular pathways abnormally altered in glioblastoma multiforme (GBM) is essential for developing effective therapeutic approaches for this deadly disease.Methodology/Principal FindingsApplying a next generation sequencing technology, massively parallel signature sequencing (MPSS), we identified a total of 4535 genes that are differentially expressed between normal brain and GBM tissue. The expression changes of three up-regulated genes, CHI3L1, CHI3L2, and FOXM1, and two down-regulated genes, neurogranin and L1CAM, were confirmed by quantitative PCR. Pathway analysis revealed that TGF- β pathway related genes were significantly up-regulated in GBM tumor samples. An integrative pathway analysis of the TGF β signaling network identified two alternative TGF−β signaling pathways mediated by SOX4 (sex determining region Y-box 4) and TGFBI (Transforming growth factor beta induced). Quantitative RT-PCR and immunohistochemistry staining demonstrated that SOX4 and TGFBI expression is elevated in GBM tissues compared with normal brain tissues at both the RNA and protein levels. In vitro functional studies confirmed that TGFBI and SOX4 expression is increased by TGF- β stimulation and decreased by a specific inhibitor of TGF- β receptor 1 kinase.Conclusions/SignificanceOur MPSS database for GBM and normal brain tissues provides a useful resource for the scientific community. The identification of non-SMAD mediated TGF−β signaling pathways acting through SOX4 and TGFBI (GENE ID:7045) in GBM indicates that these alternative pathways should be considered, in addition to the canonical SMAD mediated pathway, in the development of new therapeutic strategies targeting TGF−β signaling in GBM. Finally, the construction of an extended TGF- β signaling network with overlaid gene expression changes between GBM and normal brain extends our understanding of the biology of GBM.