SOX2 Is an Oncogene Activated by Recurrent 3q26.3 Amplifications in Human Lung Squamous Cell Carcinomas

Squamous cell carcinoma (SCC) of the lung is a frequent and aggressive cancer type. Gene amplifications, a known activating mechanism of oncogenes, target the 3q26-qter region as one of the most frequently gained/amplified genomic sites in SCC of various types. Here, we used array comparative genomic hybridization to delineate the consensus region of 3q26.3 amplifications in lung SCC. Recurrent amplifications occur in 20% of lung SCC (136 tumors in total) and map to a core region of 2 Mb (Megabases) that encompasses SOX2, a transcription factor gene. Intense SOX2 immunostaining is frequent in nuclei of lung SCC, indicating potential active transcriptional regulation by SOX2. Analyses of the transcriptome of lung SCC, SOX2-overexpressing lung epithelial cells and embryonic stem cells (ESCs) reveal that SOX2 contributes to activate ESC-like phenotypes and provide clues pertaining to the deregulated genes involved in the malignant phenotype. In cell culture experiments, overexpression of SOX2 stimulates cellular migration and anchorage-independent growth while SOX2 knockdown impairs cell growth. Finally, SOX2 over-expression in non-tumorigenic human lung bronchial epithelial cells is tumorigenic in immunocompromised mice. These results indicate that the SOX2 transcription factor, a major regulator of stem cell function, is also an oncogene and a driver gene for the recurrent 3q26.33 amplifications in lung SCC

Advances in tenascin-C biology

Tenascin-C is an extracellular matrix glycoprotein that is specifically and transiently expressed upon tissue injury. Upon tissue damage, tenascin-C plays a multitude of different roles that mediate both inflammatory and fibrotic processes to enable effective tissue repair. In the last decade, emerging evidence has demonstrated a vital role for tenascin-C in cardiac and arterial injury, tumor angiogenesis and metastasis, as well as in modulating stem cell behavior. Here we highlight the molecular mechanisms by which tenascin-C mediates these effects and discuss the implications of mis-regulated tenascin-C expression in driving disease pathology

Cartographie haute-résolution des amplifications de la région 3q25-qter pour l'identification d'oncogènes dans différents types tumoraux,Application à des fibro-histiocytomes malins et des carcinomes épidermoïdes (poumon et voies aéro-digestives supérieur

L'approche de clonage positionnel a été appliquée à la cartographie haute résolution des aberrations du chromosome 3 dans différents types tumoraux afin d'identifier de nouveaux candidats oncogènes du bras long du chromosome 3 (3q), amplifiés et sur-exprimés en conséquence.Une première partie a été dédiée aux Fibro-Histiocytomes Malins (MFH). Des amplicons chevauchants du locus 3q28 ont été caractérisés dans une lignée cellulaire et deux tumeurs de MFH. A ce locus, nous avons identifié un oncogène candidat, hsa-miR-28, codant pour un microARN ainsi que trois de ses cibles, impliquées dans la régulation du cycle cellulaire.Une seconde partie concerne les carcinomes épidermoïdes (SCC).Nous avons analysé les aberrations du chromosome 3 dans 25 SCC du poumon par hybridation génomique comparative sur puces à ADN (CGH array) et défini une région consensus de 2 Mb au locus 3q26.3 amplifiée à haut niveau dans 20% des cas. L'analyse des conséquences transcriptionnelles pour 9 gènes a montré que la plupart sont sur-exprimés de manière récurrente mais deux, SOX2 et SOX2OT, représentent vraisemblablement les cibles des amplifications du locus.Nous avons également isolé le gène cyclin L1 au locus 3q25.3 comme oncogène candidat dans les SCC des Voies Aéro-Digestives Supérieures (VADS). Le nombre de copies du gène est très fréquemment augmenté dans ces tumeurs et le gène surexprimé. Finalement, nous avons caractérisé par CGH array différentes amplifications du 3q dans une série de 25 SCC des VADS, définissant une région commune avec les SCC du poumon suggérant ainsi que des gènes similaires du locus 3q26.3 sont impliqués dans les SCC indépendamment de leur localisation.En conclusion, nous avons isolé sur le bras long du chromosome 3 différents candidats oncogènes dans plusieurs types tumoraux, ce qui illustre la puissance de l'approche de clonage positionnel appliquée aux génomes tumoraux. Des travaux sont encore nécessaires pour vérifier leur statut oncogénique.This thesis work specifically applied the positional cloning strategy to the high resolution mapping of chromosome 3 aberrations in different tumour types to identify new candidate oncogenes located at 3q, amplified and consequently over-expressed. A first part was dedicated to Malignant Fibrous Histiocytomas (MFH). We first characterized overlapping amplicons at 3q28 in a MFH cell line and two other MFH primary tumours. From this region, we identified a microRNA encoding gene, hsa-miR-28 as an oncogene candidate and further identified three targets of miR-28 which all play roles in cell cycle regulation. A second part was dedicated to Squamous Cell Carcinoma (SCC). We analyzed chromosome 3 aberrations in a series of 25 lung SCC using array CGH and delineated a common region of high-level amplifications at 3q26.3 for 20% of the tumours. Further high-resolution mapping pinpoint a 2 Mb consensus region. Analyses of the transcriptional consequences of these high-level amplifications were carried out for 9 genes. Most of them are recurrently over-expressed but two, SOX2 and SOX2OT, likely represent the 3q26.3 amplification driver genes. Our strategy also enabled us to isolate cyclin L1 (CCNL1) gene at 3q25.3 as a candidate oncogene in head and neck SCC (HNSCC). Investigations of CCNL1 gene alterations in a large series of HNSCC revealed consistent copy number gains and over-expression. Using array CGH we finally delineated several high-level amplifications at 3q in a series of 25 analyzed HNSCC, defining a common region overlapping with the consensus region defined for lung SCC thus suggesting that similar genes at 3q26.3 may be involved in SCC pathogenesis independently of the localization.In conclusion we were able to isolate several candidate oncogenes located at 3q in different tumour types, illustrating the power of the positional cloning strategy applied to tumour genomes. Further investigations are needed to assess their oncogenic status

Towards the identification of oncogenes by high resolution mapping of gene amplifications of the 3q25-qter region in Malignant Fibrous Histiocytoma and Squamous Cell Carcinoma

Cancer cells are characterized by a certain number of hallmarks. Among these, genetic instability is admitted to be a near universal one. Consequently, most of tumour cells presents genetic alterations and particularly at the chromosomal level. Actually, interstitial deletions, low-level copy number gains and high-level copy number amplifications of genomic regions are recurrently observed in tumour genomes, and produce deregulations in the expression of tumour suppressor genes and oncogenes. We used a positional cloning strategy by mapping of chromosomal aberrations to define common genomic intervals that are altered in different tumours and identify candidate oncogenes. This thesis work specifically applied this strategy to chromosome 3 aberrations in different tumour types and particularly to identify new candidate oncogenes located at 3q, amplified and consequently over-expressed. A first part was dedicated to the study of some soft-tissue sarcomas, Malignant Fibrous Histiocytomas (MFH). In a first approach, we characterized an amplicon at 3q28 in a MFH cell line. This region was also found to be amplified in two other MFH primary tumours but no obvious candidate oncogene was isolated (Hussenet et al., submitted). From this region, we finally identified a microRNA encoding gene, hsa-miR-28 representing an oncogene candidate: its inhibition indeed slows down the proliferation of 2 MFH cell lines. We further identified three targets of miR-28 (DP2, E2F6 and CDC14A): all play roles in the regulation of the cell cycle and interestingly the hsa-miR-28 gene expression itself seems to be cell cycle regulated (Hussenet et al., in preparation). A second part was dedicated to Squamous Cell Carcinoma (SCC) of two localizations (lung & head and neck). We analyzed chromosome 3 aberrations in a series of 25 lung SCC using array CGH. This work allowed us to delineate three regions of deletions at 3p, low-level copy number gain of the 3q26-qter region in a majority of tumours, as well as a common region of high-level amplifications at 3q26.3 for 20% of the tumours. Further high-resolution mapping of these amplifications using array CGH with an array providing tiling coverage of the 3q26.3 locus pinpoint a consensus region that is approximately 2 Mb in size. Analyses of the transcriptional consequences of these high-level amplifications were carried out for 9 genes of the region. Most of them are recurrently over-expressed but two, SOX2 and SOX2OT, likely represent the 3q26.3 amplification driver genes in these tumours. Ongoing in vitro functional assays will assess the effect of over-expression of candidate oncogenes located in the consensus region of high-level amplifications at 3q26.3 (Hussenet et al., in preparation). Our positional cloning strategy also enabled us to isolate the cyclin L1 (CCNL1) gene at 3q25.3 as a candidate oncogene, amplified and over-expressed in a head and neck SCC (HNSCC) cell line, as well as over-expressed in HNSCC tumours (Redon et al., 2002). Further investigations of CCNL1 gene alterations in a larger series of HNSCC revealed consistent low-level copy number gains and over-expression in these tumours (Muller et al., submitted). Finally we also screened a series of 25 HNSCC for chromosome 3 aberrations using array CGH. We were able to delineate several recurrent sites of deletions at 3p, low-level gains and high-level amplifications at 3q. A common region of high-level amplifications in HNSCC is overlapping with the consensus region we have previously defined for lung SCC and suggests that similar genes at 3q26.3 may be involved in SCC pathogenesis independently of the localization. In conclusion, this work illustrates the power of the positional cloning strategy applied to tumour genomes through the use of high resolution methods and tools to map the recurrent sites of genomic alterations. We indeed were able to isolate several oncogene candidate located at 3q that are amplified and consequently over-expressed in different tumour types. Further investigations are still needed to assess their oncogenic status as well as putative relationships with clinical parameters

Cartographie haute-résolution des amplifications de la région 3q25-qter pour l'identification d'oncogènes dans différents types tumoraux (Application à des fibro-histiocytomes malins et des carcinomes épidermoïdes (poumon et voies aéro-digestives supérieures))

L approche de clonage positionnel a été appliquée à la cartographie haute résolution des aberrations du chromosome 3 dans différents types tumoraux afin d identifier de nouveaux candidats oncogènes du bras long du chromosome 3 (3q), amplifiés et sur-exprimés en conséquence.Une première partie a été dédiée aux Fibro-Histiocytomes Malins (MFH). Des amplicons chevauchants du locus 3q28 ont été caractérise s dans une lignée cellulaire et deux tumeurs de MFH. A ce locus, nous avons identifié un oncogène candidat, hsa-miR-28, codant pour un microARN ainsi que trois de ses cibles, impliquées dans la régulation du cycle cellulaire.Une seconde partie concerne les carcinomes épidermoïdes (SCC).Nous avons analysé les aberrations du chromosome 3 dans 25 SCC du poumon par hybridation génomique comparative sur puces à ADN (CGH array) et défini une région consensus de 2 Mb au locus 3q26.3 amplifiée à haut niveau dans 20% des cas. L analyse des conséquences transcriptionnelles pour 9 gènes a montré que la plupart sont sur-exprimés de manière récurrente mais deux, SOX2 et SOX2OT, représentent vraisemblablement les cibles des amplifications du locus.Nous avons également isolé le gène cyclin L1 au locus 3q25.3 comme oncogène candidat dans les SCC des Voies Aéro-Digestives Supérieures (VADS). Le nombre de copies du gène est très fréquemment augmenté dans ces tumeurs et le gène surexprimé. Finalement, nous avons caractérisé par CGH array différentes amplifications du 3q dans une série de 25 SCC des VADS, définissant une région commune avec les SCC du poumon suggérant ainsi que des gènes similaires du locus 3q26.3 sont impliqués dans les SCC indépendamment de leur localisation.En conclusion, nous avons isolé sur le bras long du chromosome 3 différents candidats oncogènes dans plusieurs types tumoraux, ce qui illustre la puissance de l approche de clonage positionnel appliquée aux génomes tumoraux. Des travaux sont encore nécessaires pour vérifier leur statut oncogénique.This thesis work specifically applied the positional cloning strategy to the high resolution mapping of chromosome 3 aberrations in different tumour types to identify new candidate oncogenes located at 3q, amplified and consequently over-expressed. A first part was dedicated to Malignant Fibrous Histiocytomas (MFH). We first characterized overlapping amplicons at 3q28 in a MFH cell line and two other MFH primary tumours. From this region, we identified a microRNA encoding gene, hsa-miR-28 as an oncogene candidate and further identified three targets of miR-28 which all play roles in cell cycle regulation. A second part was dedicated to Squamous Cell Carcinoma (SCC). We analyzed chromosome 3 aberrations in a series of 25 lung SCC using array CGH and delineated a common region of high-level amplifications at 3q26.3 for 20% of the tumours. Further high-resolution mapping pinpoint a 2 Mb consensus region. Analyses of the transcriptional consequences of these high-level amplifications were carried out for 9 genes. Most of them are recurrently over-expressed but two, SOX2 and SOX2OT, likely represent the 3q26.3 amplification driver genes. Our strategy also enabled us to isolate cyclin L1 (CCNL1) gene at 3q25.3 as a candidate oncogene in head and neck SCC (HNSCC). Investigations of CCNL1 gene alterations in a large series of HNSCC revealed consistent copy number gains and over-expression. Using array CGH we finally delineated several high-level amplifications at 3q in a series of 25 analyzed HNSCC, defining a common region overlapping with the consensus region defined for lung SCC thus suggesting that similar genes at 3q26.3 may be involved in SCC pathogenesis independently of the localization.In conclusion we were able to isolate several candidate oncogenes located at 3q in different tumour types, illustrating the power of the positional cloning strategy applied to tumour genomes. Further investigations are needed to assess their oncogenic status.STRASBOURG-Sc. et Techniques (674822102) / SudocSudocFranceF

An adult tissue-specific stem cell molecular phenotype is activated in epithelial cancer stem cells and correlated to patient outcome

Recent studies have shown that embryonic stem cell-like molecular phenotypes are commonly activated in human epithelial primary tumors and are linked to adverse patient prognosis.(1,2) However it remains unclear whether these correlations to outcome are linked to the differentiation status of the human primary tumors(1) or represent molecular reminiscences of epithelial cancer stem cells.(2) In addition, while it has been demonstrated that leukemic cancer stem cells re-acquire an embryonic stem cell-like phenotype,(3,4) the molecular basis of stem cell function in epithelial cancer stem cells has not been investigated. Here we show that a normal adult tissue-specific stem cell molecular phenotype is commonly activated in epithelial cancer stem cells and for the first time provide evidence that enrichment in cancer stem cells-specific molecular signatures are correlated to highly aggressive tumor phenotypes in human epithelial cancers

The laminin response in inflammatory bowel disease: Protection or malignancy?

Laminins (LM), basement membrane molecules and mediators of epithelial-stromal communication, are crucial in tissue homeostasis. Inflammatory Bowel Diseases (IBD) are multifactorial pathologies where the microenvironment and in particular LM play an important yet poorly understood role in tissue maintenance, and in cancer progression which represents an inherent risk of IBD. Here we showed first that in human IBD colonic samples and in murine colitis the LMα1 and LMα5 chains are specifically and ectopically overexpressed with a concomitant nuclear p53 accumulation. Linked to this observation, we provided a mechanism showing that p53 induces LMα1 expression at the promoter level by ChIP analysis and this was confirmed by knockdown in cell transfection experiments. To mimic the human disease, we induced colitis and colitis-associated cancer by chemical treatment (DSS) combined or not with a carcinogen (AOM) in transgenic mice overexpressing LMα1 or LMα5 specifically in the intestine. We demonstrated that high LMα1 or LMα5 expression decreased susceptibility towards experimentally DSS-induced colon inflammation as assessed by histological scoring and decrease of pro-inflammatory cytokines. Yet in a pro-oncogenic context, we showed that LM would favor tumorigenesis as revealed by enhanced tumor lesion formation in both LM transgenic mice. Altogether, our results showed that nuclear p53 and associated overexpression of LMα1 and LMα5 protect tissue from inflammation. But in a mutation setting, the same LM molecules favor progression of IBD into colitis-associated cancer. Our transgenic mice represent attractive new models to acquire knowledge about the paradoxical effect of LM that mediate either tissue reparation or cancer according to the microenvironment. In the early phases of IBD, reinforcing basement membrane stability/organization could be a promising therapeutic approach

Predictors of negative first SARS-CoV-2 RT-PCR despite final diagnosis of COVID-19 and association with outcome.

Reverse transcriptase-polymerase chain reaction (RT-PCR) testing is an important tool for diagnosing coronavirus disease 2019 (COVID-19). However, performance concerns have emerged recently, notably regarding sensitivity. We hypothesized that the clinical, biological, and radiological characteristics of patients with a false-negative first RT-PCR test and a final diagnosis of COVID-19 might differ from those of patients with a positive first RT-PCR test. We conducted a multicenter matched case-control study in COVID-19 patients. Patients with a negative first RT-PCR test were matched to patients with a positive first RT-PCR test on age, sex, and initial admission unit (ward or intensive care). We included 80 cases and 80 controls between March 30, and June 22, 2020. Neither mortality at hospital discharge nor hospital stay length differed between the two groups (P = 0.80 and P = 0.54, respectively). By multivariate analysis, two factors were independently associated with a lower risk of a first false-negative test, namely, headache (adjusted OR [aOR], 0.07; 95% confidence interval [95% CI], 0.01-0.49]; P = 0.007) and fatigue/malaise (aOR, 0.16; 95% CI, 0.03-0.81; P = 0.027); two other factors were independently associated with a higher risk of a first false-negative test, namely, platelets > 207·10 mm (aOR, 3.81; 95% CI, 1.10-13.16]; P = 0.034) and C-reactive protein > 79.8 mg·L (aOR, 4.00; 95% CI, 1.21-13.19; P = 0.023). Patients with suspected COVID-19 whose laboratory tests indicating marked inflammation were at higher risk of a first false-negative RT-PCR test. Strategies involving serial RT-PCR testing must be rigorously evaluated

Tenascin-C downregulates wnt inhibitor dickkopf-1, promoting tumorigenesis in a neuroendocrine tumor model

The extracellular matrix molecule tenascin-C (TNC) is a major component of the cancer-specific matrix, and high TNC expression is linked to poor prognosis in several cancers. To provide a comprehensive understanding of TNC's functions in cancer, we established an immune-competent transgenic mouse model of pancreatic β-cell carcinogenesis with varying levels of TNC expression and compared stochastic neuroendocrine tumor formation in abundance or absence of TNC. We show that TNC promotes tumor cell survival, the angiogenic switch, more and leaky vessels, carcinoma progression, and lung micrometastasis. TNC downregulates Dickkopf-1 (DKK1) promoter activity through the blocking of actin stress fiber formation, activates Wnt signaling, and induces Wnt target genes in tumor and endothelial cells. Our results implicate DKK1 downregulation as an important mechanism underlying TNC-enhanced tumor progression through the provision of a proangiogenic tumor microenvironment

Transmembrane Domain Targeting Peptide Antagonizing ErbB2/Neu Inhibits Breast Tumor Growth and Metastasis

Breast cancer is still a deadly disease despite major achievements in targeted therapies designed to block ligands or ligand-binding subunits of major tyrosine kinase receptors. Relapse is significant and metastases deleterious, which demands novel strategies for fighting this disease. Here, we report a proof-of-concept experiment demonstrating that small peptides interfering with the transmembrane domain of the tyrosine kinase epidermal growth factor receptor ErbB2 exhibit anticancer properties when used at micromolar dosages in a genetically engineered mouse model of breast cancer. Different assays demonstrate the specificity of the ErbB2-targeting peptide, which induces long-term reduction of ErbB2 phosphorylation and Akt signaling consistent with reduced tumor cell proliferation and increased survival. Microcomputed tomography analysis established the antimetastatic activity of the peptide and its impact on primary tumor growth. This reveals the interior of the cell membrane as an unexplored dimension for drug design