scholarly journals Pregnancy-Associated Plasma Protein-A (PAPP-A) in Ewing Sarcoma: Role in Tumor Growth and Immune Evasion

2019 ◽  
Vol 111 (9) ◽  
pp. 970-982 ◽  
Author(s):  
Sabine Heitzeneder ◽  
Elena Sotillo ◽  
Jack F Shern ◽  
Sivasish Sindiri ◽  
Peng Xu ◽  
...  
Keyword(s):  
Cell Surface ◽  
Cell Lines ◽  
Plasma Protein ◽  
Immune Evasion ◽  
Ewing Sarcoma ◽  
Antigen Processing ◽  
Expression Profiles ◽  
Statistical Tests ◽  
Protein A ◽  
Differentially Expressed

AbstractBackgroundEwing sarcoma (EWS) manifests one of the lowest somatic mutation rates of any cancer, leading to a scarcity of druggable mutations and neoantigens. Immunotherapeutics targeting differentially expressed cell surface antigens could provide therapeutic benefit for such tumors. Pregnancy-associated plasma protein A (PAPP-A) is a cell membrane-associated proteinase produced by the placenta that promotes fetal growth by inducing insulinlike growth factor (IGF) signaling.MethodsBy comparing RNA expression of cell surface proteins in EWS (n = 120) versus normal tissues (n = 42), we comprehensively characterized the surfaceome of EWS to identify highly differentially expressed molecules. Using CRISPR/Cas-9 and anti-PAPP-A antibodies, we investigated biological roles for PAPP-A in EWS in vitro and in vivo in NSG xenograft models and performed RNA-sequencing on PAPPA knockout clones (n = 5) and controls (n = 3). All statistical tests were two-sided.ResultsEWS surfaceome analysis identified 11 highly differentially overexpressed genes, with PAPPA ranking second in differential expression. In EWS cell lines, genetic knockout of PAPPA and treatment with anti-PAPP-A antibodies revealed an essential survival role by regulating local IGF-1 bioavailability. MAb-mediated PAPPA inhibition diminished EWS growth in orthotopic xenografts (leg area mm2 at day 49 IgG2a control (CTRL) [n = 14], mean = 397.0, SD = 86.1 vs anti-PAPP-A [n = 14], mean = 311.7, SD = 155.0; P = .03; median OS anti-PAPP-A = 52.5 days, 95% CI = 46.0 to 63.0 days vs IgG2a = 45.0 days, 95% CI = 42.0 to 52.0 days; P = .02) and improved the efficacy of anti-IGF-1R treatment (leg area mm2 at day 49 anti-PAPP-A + anti-IGF-1R [n = 15], mean = 217.9, SD = 148.5 vs IgG2a-CTRL; P < .001; median OS anti-PAPP-A + anti-IGF1R = 63.0 days, 95% CI = 52.0 to 67.0 days vs IgG2a-CTRL; P < .001). Unexpectedly, PAPPA knockout in EWS cell lines induced interferon (IFN)-response genes, including proteins associated with antigen processing/presentation. Consistently, gene expression profiles in PAPPA-low EWS tumors were enriched for immune response pathways.ConclusionThis work provides a comprehensive characterization of the surfaceome of EWS, credentials PAPP-A as a highly differentially expressed therapeutic target, and discovers a novel link between IGF-1 signaling and immune evasion in cancer, thus implicating shared mechanisms of immune evasion between EWS and the placenta.

scholarly journals Proteomic Study of HPV-Positive Head and Neck Cancers: Preliminary Results

2014 ◽  
Vol 2014 ◽  
pp. 1-16 ◽  
Author(s):  
Géraldine Descamps ◽  
Ruddy Wattiez ◽  
Sven Saussez
Keyword(s):  
Head And Neck ◽  
Cell Lines ◽  
Expression Profiles ◽  
Elongation Factor ◽  
Head And Neck Carcinoma ◽  
Hpv Infection ◽  
Differentially Expressed ◽  
Hpv Status ◽  
Clinical Series ◽  
Protein Expression Profiles

Human papillomavirus (HPV) was recently recognized as a new risk factor for head and neck squamous cell carcinoma. For oropharyngeal cancers, an HPV+ status is associated with better prognosis in a subgroup of nonsmokers and nondrinkers. However, HPV infection is also involved in the biology of head and neck carcinoma (HNC) in patients with a history of tobacco use and/or alcohol consumption. Thus, the involvement of HPV infection in HN carcinogenesis remains unclear, and further studies are needed to identify and analyze HPV-specific pathways that are involved in this process. Using a quantitative proteomics-based approach, we compared the protein expression profiles of two HPV+ HNC cell lines and one HPV− HNC cell line. We identified 155 proteins that are differentially expressed (P<0.01) in these three lines. Among the identified proteins, prostate stem cell antigen (PSCA) was upregulated and eukaryotic elongation factor 1 alpha (EEF1α) was downregulated in the HPV+ cell lines. Immunofluorescence and western blotting analyses confirmed these results. Moreover, PSCA and EEF1αwere differentially expressed in two clinical series of 50 HPV+ and 50 HPV− oral cavity carcinomas. Thus, our study reveals for the first time that PSCA and EEF1αare associated with the HPV-status, suggesting that these proteins could be involved in HPV-associated carcinogenesis.

scholarly journals Gene expression profiles of progressive pancreatic endocrine tumours and their liver metastases reveal potential novel markers and therapeutic targets

2006 ◽  
Vol 13 (2) ◽  
pp. 541-558 ◽  
Author(s):  
G Capurso ◽  
S Lattimore ◽  
T Crnogorac-Jurcevic ◽  
F Panzuto ◽  
M Milione ◽  
...  
Keyword(s):  
Gene Expression ◽  
Liver Metastases ◽  
Cell Lines ◽  
Differentially Expressed Genes ◽  
Expression Profiles ◽  
Therapeutic Targets ◽  
Gene Expression Profiles ◽  
Differentially Expressed ◽  
Striking Similarity ◽  
Metastatic Lesions

The intrinsic nature of tumour behaviour (stable vs progressive) and the presence of liver metastases are key factors in determining the outcome of patients with a pancreatic endocrine tumour (PET). Previous expression profile analyses of PETs were limited to non-homogeneous groups or to primary lesions only. The aim of this study was to investigate the gene expression profiles of a more uniform series of sporadic, non-functioning (NF) PETs with progressive disease and, for the first time, their liver metastases, on the Affymetrix human genome U133A and B GeneChip set. Thirteen NF PET samples (eight primaries and five liver metastases) from ten patients with progressive, metastatic disease, three cell lines (BON, QGP and CM) and four purified islet samples were analysed. The same samples were employed for confirmation of candidate gene expression by means of quantitative RT-PCR, while a further 37 PET and 15 carcinoid samples were analysed by immunohistochemistry. Analysis of genes differentially expressed between islets and primaries and metastases revealed 667 up- and 223 down-regulated genes, most of which have not previously been observed in PETs, and whose gene ontology molecular function has been detailed. Overexpression of bridging integrator 1 (BIN1) and protein Z dependent protease inhibitor (SERPINA10) which may represent useful biomarkers, and of lymphocyte specific protein tyrosine kinase (LCK) and bone marrow stromal cell antigen (BST2) which could be used as therapeutic targets, has been validated. When primary tumours were compared with metastatic lesions, no significantly differentially expressed genes were found, in accord with cluster analysis which revealed a striking similarity between primary and metastatic lesions, with the cell lines clustering separately. We have provided a comprehensive list of differentially expressed genes in a uniform set of aggressive NF PETs. A number of dysregulated genes deserve further in-depth study as potentially promising candidates for new diagnostic and treatment strategies. The analysis of liver metastases revealed a previously unknown high level of similarity with the primary lesions.

Multiple Myeloma Cells Modulate ICAM-3 To Evade Natural Killer Cell-Mediated Lysis

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3105-3105
Author(s):  
Tarun K. Garg ◽  
Ricky D Edmondson ◽  
Shweta S. Chavan ◽  
Junaid Khan ◽  
Susann Szmania ◽  
...  
Keyword(s):  
Amino Acids ◽  
Flow Cytometry ◽  
High Risk ◽  
Cell Surface ◽  
Natural Killer ◽  
Cell Lines ◽  
Myeloma Cell ◽  
Differentially Expressed ◽  
Myeloma Cells ◽  
Chromium Release

Abstract Introduction Ex vivo activated/expanded natural killer (ENK) cells can induce myeloma cell lysis both in vitro and in murine models and are currently being studied clinically in the setting of high-risk relapsing disease and asymptomatic disease at high risk of progression. This prompted us to study, in myeloma cell lines, whether intrinsic resistance to ENK cell lysis exists, whether repeated challenge with ENK leads to increased resistance, and what the underlying mechanisms of resistance are. Of 11 myeloma cell lines tested in standard 4h chromium release assays, 8 were avidly killed (78-89% lysis, E:T Ratio 10:1) whereas 3 lines were less sensitive (41-65% lysis). Repeated exposure to ENK challenge decreased sensitivity in 4 of 11 lines, that was at least in part due to down-regulation of Tumor Necrosis Factor-Related Apoptosis Inducing Ligand-Receptors on the myeloma cell surface (Garg et al, Blood 2012, 120:4020). In this study we investigated the resistance issue further via metabolomics, gene expression profiling (GEP) and flow cytometry analysis of OPM2, which was intrinsically resistant and developed further resistance after challenge with ENK cells. Methods Metabolomics was studied using a quantitative proteomic strategy entailing stable isotope labeling with amino acids in cell culture – mass spectrometry (SILAC-MS). Resistant and parental OPM2 cells were grown either in medium with heavy amino acids (13C6 L-Lysine and 13C6 L-Arginine) or with light amino acids (12C6 L-Lysine and 12C6L-Arginine). Reverse labeling with heavy or light amino acids was also done to confirm the results. Cell lysates from heavy and light amino acid labeled cells were pooled, simultaneously resolved on SDS-PAGE, protein bands were excised and analyzed on a mass spectrometer after trypsin digestion. GEP was performed using the Affymetrix U133 Plus 2.0 microarray platform (Santa Clara, CA). The fold change of signal intensity for genes and proteins in resistant vs. parental OPM2 was calculated. The most differentially expressed genes (top 150-fold up or down) and proteins (up or down by 1.3-fold) were compared for commonality. Cell surface protein expression was determined via flow cytometry. The ability of ENK to lyse myeloma cell targets in the presence of isotype control or ICAM-3 blocking antibody was tested in 4h chromium release assays. Results Metabolomics identified >3800 proteins and revealed that the abundance of 352 proteins was significantly altered in resistant myeloma cells. These altered proteins were mainly associated with cell cycle, morphology, organization, cellular compromise, immune response, and survival. Further, a comparison of these differentially expressed proteins with GEP data revealed 3 commonly up-regulated molecules: TBC1D8B, HSPA1A and IFI16; and 2 down-regulated molecules: intercellular adhesion molecule (ICAM-3) and BAI3. Of these, ICAM-3, a ligand for leukocyte function-associated antigen-1 (LFA-1) and a potent signaling molecule, was selected for further studies. Flow cytometry confirmed that ICAM-3 cell surface expression was > 8-fold lower on resistant versus parental OPM2 cells. Further, blocking of ICAM-3 in cytotoxicity assays resulted in decreased lysis (43% blocked, E:T ratio 5:1), suggesting that this molecule is functionally important and takes part in ENK cell-mediated killing. Conclusion In conclusion, quantitative proteomic analysis demonstrated dynamic changes in the ENK-resistant OPM2 myeloma cells that correlated with GEP and differences in ICAM-3 expression may have functional implications. Studies evaluating the expression of ICAM-3 in myeloma patients at diagnosis and relapse are in progress. Myeloma cells may down-regulate ICAM-3 as a mechanism of escape from immune surveillance and therefore, ICAM-3 may be a useful biomarker to predict sensitivity to ENK cell-mediated killing and aid in the selection of patients most likely to benefit from ENK cell therapy. Disclosures: Barlogie: Celgene: Consultancy, Honoraria, Research Funding; Myeloma Health, LLC: Patents & Royalties.

scholarly journals Genes Regulated in Metastatic Osteosarcoma: Evaluation by Microarray Analysis in Four Human and Two Mouse Cell Line Systems

Sarcoma ◽  
2012 ◽  
Vol 2012 ◽  
pp. 1-13 ◽  
Author(s):  
Roman Muff ◽  
Ram Mohan Ram Kumar ◽  
Sander M. Botter ◽  
Walter Born ◽  
Bruno Fuchs
Keyword(s):  
Gene Expression ◽  
Cell Line ◽  
Cell Lines ◽  
Expression Profiles ◽  
Treatment Strategies ◽  
Metastatic Potential ◽  
New Drugs ◽  
Differentially Expressed ◽  
Osteoblastic Cell ◽  
Osteoblastic Cell Line

Osteosarcoma (OS) is a rare bone neoplasm that affects mainly adolescents. It is associated with poor prognosis in case of metastases formation. The search for metastasis predicting markers is therefore imperative to optimize treatment strategies for patients at risk and important for the search of new drugs for the treatment of this devastating disease. Here, we have analyzed by microarray the differential gene expression in four human and two mouse OS cell line systems consisting of parental cell lines with low metastatic potential and derivatives thereof with increased metastatic potential. Using two osteoblastic cell line systems, the most common OS phenotype, we have identified forty-eight common genes that are differentially expressed in metastatic cell lines compared to parental cells. The identified subset of metastasis relevant genes in osteoblastic OS overlapped only minimally with differentially expressed genes in the other four preosteoblast or nonosteoblastic cell line systems. The results imply an OS phenotype specific expression pattern of metastasis regulating proteins and form a basis for further investigation of gene expression profiles in patients’ samples combined with survival analysis with the aim to optimize treatment strategies to develop new drugs and to consequently improve the survival of patients with the most common form of osteoblastic OS.

scholarly journals Clinically Relevant Biomarker Discovery in High-Risk Recurrent Neuroblastoma

2019 ◽  
Vol 18 ◽  
pp. 117693511983291 ◽  
Author(s):  
Peter Utnes ◽  
Cecilie Løkke ◽  
Trond Flægstad ◽  
Christer Einvik
Keyword(s):  
High Risk ◽  
Cell Lines ◽  
Target Genes ◽  
Biomarker Discovery ◽  
Expression Profiles ◽  
Response To Therapy ◽  
Gene Set Enrichment Analysis ◽  
Favorable Outcome ◽  
Differentially Expressed ◽  
Response To Treatment

Neuroblastoma is a pediatric cancer of the developing sympathetic nervous system. High-risk neuroblastoma patients typically undergo an initial remission in response to treatment, followed by recurrence of aggressive tumors that have become refractory to further treatment. The need for biomarkers that can select patients not responding well to therapy in an early phase is therefore needed. In this study, we used next generation sequencing technology to determine the expression profiles in high-risk neuroblastoma cell lines established before and after therapy. Using partial least squares-discriminant analysis (PLS-DA) with least absolute shrinkage and selection operator (LASSO) and leave-one-out cross-validation, we identified a panel of 55 messenger RNAs and 17 long non-coding RNAs (lncRNAs) which were significantly altered in the expression between cell lines isolated from primary and recurrent tumors. From a neuroblastoma patient cohort, we found 20 of the 55 protein-coding genes to be differentially expressed in patients with unfavorable compared with favorable outcome. We further found a twofold increase or decrease in hazard ratios in these genes when comparing patients with unfavorable and favorable outcome. Gene set enrichment analysis (GSEA) revealed that these genes were involved in proliferation, differentiation and regulated by Polycomb group (PcG) proteins. Of the 17 lncRNAs, 3 upregulated ( NEAT1, SH3BP5-AS1, NORAD) and 3 downregulated lncRNAs ( DUBR, MEG3, DHRS4-AS1) were also found to be differentially expressed in favorable compared with unfavorable outcome. Moreover, using expression profiles on both miRNAs and mRNAs in the same cohort of cell lines, we found 13 downregulated and 18 upregulated experimentally observed miRNA target genes targeted by miR-21, -424 and -30e, -29b, -138, -494, - 181a, -34a, -29b, respectively. The advantage of analyzing biomarkers in a clinically relevant neuroblastoma model system enables further studies on the effect of individual genes upon gene perturbation. In summary, this study identified several genes, which may aid in the prediction of response to therapy and tumor recurrence.

Transcriptomal analysis of failing and nonfailing human hearts

2003 ◽  
Vol 12 (2) ◽  
pp. 97-112 ◽  
Author(s):  
M. Steenman ◽  
Y.-W. Chen ◽  
M. Le Cunff ◽  
G. Lamirault ◽  
A. Varró ◽  
...  
Keyword(s):  
Gene Expression ◽  
Heart Failure ◽  
Cardiac Function ◽  
Expression Profile ◽  
Large Scale ◽  
Expression Profiles ◽  
Statistical Tests ◽  
Differentially Expressed ◽  
Developmental Gene Expression ◽  
Similar Expression Profile

Heart failure is a multifactorial disease that may result from different initiating events. To contribute to an improved comprehension of normal cardiac function and the molecular events leading to heart failure, we performed large-scale gene expression analysis of failing and nonfailing human ventricle. Our aim was to define and compare expression profiles of 4 specific pathophysiological cardiac situations: 1) left ventricle (LV) from nonfailing heart; 2) LV from failing hearts affected by dilated cardiomyopathy (DCM); 3) LV from failing hearts affected by ischemic CM (ICM); 4) right ventricle (RV) from failing hearts affected by DCM or ICM. We used oligonucleotide arrays representing ∼12,000 human genes. After stringent numerical analyses using several statistical tests, we identified 1,306 genes with a similar expression profile in all 4 cardiac situations, therefore representative of part of the human cardiac expression profile. A total of 95 genes displayed differential expression between failing and nonfailing heart samples, reflecting a reversal to developmental gene expression, dedifferentiation of failing cardiomyocytes, and involvement of apoptosis. Twenty genes were differentially expressed between failing LV and failing RV, identifying possible candidates for different functioning of both ventricles. Finally, no genes were found to be significantly differentially expressed between failing DCM and failing ICM LV, emphasizing that transcriptomal analysis of explanted hearts results mainly in identification of expression profiles of end-stage heart failure and less in determination of expression profiles of the underlying etiology. Taken together, our data resulted in identification of putative transcriptomal landmarks for normal and disturbed cardiac function.

scholarly journals Differentially Expressed Long Noncoding RNAs Involved in FUBP1 Promoting Hepatocellular Carcinoma Cells Proliferation

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Xianpeng Li ◽  
Huaixi Yu ◽  
Feng Xu ◽  
Yifeng Wu ◽  
Jifang Sheng
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Lines ◽  
Noncoding Rna ◽  
Target Genes ◽  
Expression Profiles ◽  
Differentially Expressed ◽  
Cell Counting ◽  
Rt Pcr ◽  
Upstream Element ◽  
Hcc Cells

Background. Far upstream element-binding protein 1 (FUBP1) is reported to be involved in cancer development by regulating the transcription of c-myc gene through binding to far upstream element. Highly expressed FUBP1 was negatively correlated with survival rate of patients with hepatocellular carcinoma (HCC) and could promote the proliferation of HCC cells. However, the downstream mechanism of FUBP1 has not yet been clearly explained. This study is aimed at identifying the expression profiles of long noncoding RNA (lncRNA) in HCC cells in response to FUBP1 overexpression and at investigating the possible lncRNAs that participated in cell proliferation process regulated by FUBP1. Methods. The overexpression of FUBP1 was mediated by lentiviral infection on 3 different types of HCC cell lines (MHCC97-H, MHCC97-L, and Huh-7). The expression of target genes was detected by quantitative reverse transcription-PCR (RT-PCR) and western blotting assays. Microarray and quantitative RT-PCR were applied to screen the differentially expressed lncRNAs in HCC cells after FUBP1 overexpression. The Cell Counting Kit-8 assay was used to confirm the growth vitality of HCC cells. Results. The growth vitality of HCC cells was significantly increased after lentivirus infection. A total of 12 lncRNAs had the same expression trend in the 3 HCC cell lines in response to FUBP1 overexpression, including 3 upregulated lncRNAs and 9 downregulated lncRNAs. Coexpression analysis of dysregulated lncRNAs-mRNAs network showed that lnc-LYZ-2 was the lncRNA most relevant to FUBP1. Inhibition of lnc-LYZ-2 could significantly relieve the proproliferation effect of FUBP1 on HCC cells, suggesting that lnc-LYZ-2 was partially involved in proproliferation regulation of FUBP1. Conclusions. Our results indicated that FUBP1 induced the abnormal expression of lncRNAs and the FUBP1-lncRNAs coexpression network in HCC cells, which could provide theoretical and experimental basis for FUBP1-lncRNAs network involved in HCC development.

scholarly journals Hippo pathway effectors YAP1/TAZ induce a EWS-FLI1-opposing gene signature and associate with disease progression in Ewing Sarcoma

2019 ◽  
Author(s):  
Pablo Rodríguez-Núñez ◽  
Laura Romero-Pérez ◽  
Ana T. Amaral ◽  
Pilar Puerto-Camacho ◽  
Carmen Jordán ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Fusion Protein ◽  
Cell Lines ◽  
Ewing Sarcoma ◽  
Transcriptional Activation ◽  
Transcriptional Repression ◽  
Hippo Pathway ◽  
Protein A ◽  
Gene Signature ◽  
Cell Of Origin

AbstractYAP1 and TAZ (WWTR1) oncoproteins are the final transducers of Hippo tumor suppressor pathway. Deregulation of the pathway leads to YAP1/TAZ activation fostering tumorigenesis in multiple malignant tumor types, including sarcoma. However, oncogenic mutations within the core components of the Hippo pathway are uncommon. Ewing Sarcoma (EwS), a pediatric cancer with low mutation rate, is characterized by a canonical fusion involvingEWSR1gene, andFLI1as the most common partner. The fusion protein is a potent driver of oncogenesis but secondary alterations are scarce, and little is known about other biological factors that determine the risk of relapse or progression. We have observed YAP1/TAZ expression and transcriptional activity in EwS cell lines. Analyses of 55 primary human EwS samples revealed that high YAP1/TAZ expression was associated with progression of the disease and predicted poorer outcome.We did not observe recurrent SNV or copy number gains/losses in Hippo pathway-related loci. However, differential CpG methylation ofRASSF1locus -a regulator of Hippo pathway- was observed in EwS cell lines compared with mesenchymal stem cells, the putative cell of origin of EwS. Hypermethylation ofRASSF1correlated with the transcriptional silencing of the tumor suppressor isoformRASFF1A, and transcriptional activation of the protumorigenic isoformRASSF1Cpromoting YAP1/TAZ activation. Knockdown of YAP1/TAZ decreased proliferation and invasion abilities of EwS cells, and revealed that YAP1/TAZ transcription activity is inversely correlated with the EWS-FLI1 transcriptional signature. This transcriptional antagonism could be partly explained by EWS-FLI1-mediated transcriptional repression of TAZ. Thus, YAP1/TAZ may override the transcriptional program induced by the fusion protein, contributing to the phenotypic plasticity determined by dynamic fluctuation of the fusion protein, a recently proposed model for disease dissemination in EwS.

Sign in / Sign up

Export Citation Format

Share Document