Prediction of an HLA-DR-binding peptide derived from Wilms’ tumour 1 protein and demonstration of in vitro immunogenicity of WT1(124–138)-pulsed dendritic cells generated according to an optimised protocol

The Wilms' tumour 1 (WT1) protein is over-expressed in several types of cancer including leukaemias and might therefore constitute a novel target for immunotherapy. Recently, human leucocyte antigen (HLA) class I-binding WT1 peptides have been identified and shown to stimulate CD8(+) T cells in vitro. For maximal CD8 cell efficacy, CD4(+) helper T cells responding to major histocompatibility complex (MHC) class II-binding epitopes are required. Here, we report that scanning the WT1 protein sequence using an evidence-based predictive computer algorithm (SYFPEITHI) yielded a peptide WT1(124-138) predicted to bind the HLA-DRB1*0401 molecule with high affinity. Moreover, synthetic WT1(124-138)-peptide-pulsed dendritic cells (DC), generated according to a protocol optimised in the present study, sensitised T cells in vitro to proliferate and secrete interferon-gamma (IFN-gamma) when rechallenged with specific peptide-pulsed DC, but not with peripheral blood mononuclear cells (PBMC). These results suggest that the WT1 protein may yield epitopes immunogenic to CD4 as well as CD8 T cells, and therefore constitute a novel potential target for specific immunotherapy.     

Glucose-induced gradual phenotypic modulation of cultured human glomerular epithelial cells may be independent of Wilms’ tumor 1 (WT1)

Background

Melatonin, a hormone-like substance involved in the regulation of the circadian rhythm, has been demonstrated to protect cells against oxidative DNA damage and to inhibit tumorigenesis.

Results

In the current study, we investigated the effect of melatonin on DNA strand breaks using the alkaline DNA comet assay in breast cancer (MCF-7) and colon cancer (HCT-15) cell lines. Our results demonstrated that cells pretreated with melatonin had significantly shorter Olive tail moments compared to non-melatonin treated cells upon mutagen (methyl methanesulfonate, MMS) exposure, indicating an increased DNA repair capacity after melatonin treatment. We further examined the genome-wide gene expression in melatonin pretreated MCF-7 cells upon carcinogen exposure and detected altered expression of many genes involved in multiple DNA damage responsive pathways. Genes exhibiting altered expression were further analyzed for functional interrelatedness using network- and pathway-based bioinformatics analysis. The top functional network was defined as having relevance for “DNA Replication, Recombination, and Repair, Gene Expression, [and] Cancer”.

Conclusions

These findings suggest that melatonin may enhance DNA repair capacity by affecting several key genes involved in DNA damage responsive pathways.     

Wilms’ Tumor Gene 1 (WT1) Silencing Inhibits Proliferation of Malignant Peripheral Nerve Sheath Tumor sNF96.2 Cell Line

Wilms’ tumor gene 1 (WT1) plays complex roles in tumorigenesis, acting as tumor suppressor gene or an oncogene depending on the cellular context. WT1 expression has been variably reported in both benign and malignant peripheral nerve sheath tumors (MPNSTs) by means of immunohistochemistry. The aim of the present study was to characterize its potential pathogenetic role in these relatively uncommon malignant tumors. Firstly, immunohistochemical analyses in MPNST sNF96.2 cell line showed strong WT1 staining in nuclear and perinuclear areas of neoplastic cells. Thus, we investigated the effects of silencing WT1 by RNA interference. Through Western Blot analysis and proliferation assay we found that WT1 knockdown leads to the reduction of cell growth in a time- and dose-dependent manner. siWT1 inhibited proliferation of sNF96.2 cell lines likely by influencing cell cycle progression through a decrease in the protein levels of cyclin D1 and inhibition of Akt phosphorylation compared to the control cells. These results indicate that WT1 knockdown attenuates the biological behavior of MPNST cells by decreasing Akt activity, demonstrating that WT1 is involved in the development and progression of MPNSTs. Thus, WT1 is suggested to serve as a potential therapeutic target for MPNSTs.     

Single Nucleotide Polymorphisms in the Wilms’ Tumour Gene 1 in Clear Cell Renal Cell Carcinoma

The Wilms’ tumour gene 1 (WT1) single nucleotide polymorphism (SNP) rs16754 has recently been described as an independent prognostic factor in acute myeloid leukaemia (AML) patients. It is of great interest to test whether WT1 SNPs can be used as a molecular marker in other cancer types in order to improve risk and treatment stratification. We performed sequencing analysis on all 10 exons of the WT1 gene in a total of 182 patients with clear cell renal cell carcinoma (ccRCC). Six different SNPs were identified, in descending order for minor allele frequency: rs2234582, rs16754, rs1799925, rs5030315, rs2234583, and rs2234581. At least one minor allele for WT1 SNP was identified in 61% of ccRCC patients. In the entire study population, only 6% carried two copies of the minor allele. The genotypes of WT1 SNPs in 78 tumour-free kidney tissue specimens were found to be in 95% concordance with corresponding tumour samples. No correlation was observed between WT1 SNP genotypes and RNA expression level. WT1 SNP genotypes did not associate with clinical and pathological characteristics. We found favourable outcomes associated with the homozygous minor allele for WT1 SNP. However, SNP genotypes did not show to be of prognostic significance when comparing wild-type versus homozygous or heterozygous for the minor allele in the entire cohort. None of the previously reported WT1 mutations in AML was found in the present study. A novel WT1 missense mutation was identified in only one patient. Our data suggest that common WT1 mutations are not involved in ccRCC. Due to too few cases harbouring the homozygous minor allele, the prognostic impact needs to be verified in larger study populations.     

The ‘curse’ of Westermarck 1

This article consists of a critical application of recent theory of nationalism to the unusual case of Mauritius. It is a poly‐ethnic and poly‐religious society whose inhabitants neither share a mythical distant past nor consider themselves culturally identical, but where nation‐building has nevertheless been moderately successful since the late 1960s. Through an examination of the processes of institutional and cultural nation‐building in the island, it is argued that the current Eurocentric view of the nation (notably as a culturally homogeneous imagined community) must be modified to fit cases like this one. Thus, the value of nationalism and nationhood as comparative concepts is questioned.     

Morphoproteomic Profiling of the Mammalian Target of Rapamycin (mTOR) Signaling Pathway in Desmoplastic Small Round Cell Tumor (EWS/WT1), Ewing’s Sarcoma (EWS/FLI1) and Wilms’ Tumor(WT1)

Background

Desmoplastic small round cell tumor (DSRCT) is a rare sarcoma in adolescents and young adults. The hallmark of this disease is a EWS-WT1 translocation resulting from apposition of the Ewing’s sarcoma (EWS) gene with the Wilms’ tumor (WT1) gene. We performed morphoproteomic profiling of DSRCT (EWS-WT1), Ewing’s sarcoma (EWS-FLI1) and Wilms’ tumor (WT1) to better understand the signaling pathways for selecting future targeted therapies.

Methodology

This pilot study assessed patients with DSRCT, Wilms’ tumor and Ewing’s sarcoma. Morphoproteomics and immunohistochemical probes were applied to detect: p-mTOR (Ser2448); p-Akt (Ser473); p-ERK1/2 (Thr202/Tyr204); p-STAT3 (Tyr 705); and cell cycle-related analytes along with their negative controls.

Principal Findings

In DSRCT the PI3K/Akt/mTOR pathway is constitutively activated by p-Akt (Ser 473) expression in the nuclear compartment of the tumor cells and p-mTOR phosphorylated on Ser 2448, suggesting mTORC2 (rictor+mTOR) as the dominant form. Ewing’s sarcoma had upregulated p-Akt and p-mTOR, predominantly mTORC2. In Wilm’s tumor, the mTOR pathway is also activated with most tumor cells moderately expressing p-mTOR (Ser 2448) in plasmalemmal and cytoplasmic compartments. This coincides with the constitutive activation of one of the downstream effectors of the mTORC1 signaling pathway, namely p-p70S6K (Thr 389). There was constitutive activation of the Ras/Raf/ERK pathway p-ERK 1/2 (Thr202/Tyr204) expression in the Wilms tumor and metastatic Ewing’s sarcoma, but not in the DSRCT.

Conclusion

Morphoproteomic tumor analyses revealed constitutive activation of the mTOR pathway as evidenced by: (a) expression of phosphorylated (p)-mTOR, p-p70S6K; (b) mTORC 2 in EWS and DSRCT; (c) ERK signaling was seen in the advanced setting indicating these as resistance pathways to IGF1R related therapies. This is the first morphoproteomic study of such pathways in these rare malignancies and may have potential therapeutic implications. Further study using morphoproteomic assessments of these tumors are warranted.     

Role of Cl− channels in primary brain tumour

There is tight interplay between Ca²⁺ and Cl⁻ flux that can influence brain tumour proliferation, migration and invasion. Glioma is the predominant malignant primary brain tumour, accounting for ˜80% of all cases. Voltage-gated Cl⁻ channel family (ClC) proteins and Cl⁻ intracellular channel (CLIC) proteins are drastically overexpressed in glioma, and are associated with enhanced cell proliferation, migration and invasion. Ca²⁺ also plays fundamental roles in the phenomenon. Ca²⁺-activated Cl⁻ channels (CaCC) such as TMEM16A and bestrophin-1 are involved in glioma formation and assist Ca²⁺ movement from intracellular stores to the plasma membrane. Additionally, the transient receptor protein (TRP) channel TRPC1 can induce activation of ClC-3 by increasing intracellular Ca²⁺concentrations and activating Ca²⁺/calmodulin-dependent protein kinase II (CaMKII). Therefore, Ca²⁺ and Cl⁻currents can concurrently mediate brain tumour cellular functions. Glioma also expresses volume regulated anion channels (VRACs), which are responsible for the swelling-induced Cl⁻ current, I_Cl,swell. This current enables glioma cells to perform regulatory volume decrease (RVD) as a survivability mechanism in response to hypoxic conditions within the tumour microenvironment. RVD can also be exploited by glioma for invasion and migration. Effective treatment for glioma is challenging, which can be in part due to prolonged chemotherapy leading to mutations in genes associated with multi-drug resistances (MRP1, Bcl-2, and ABC family). Thus, a potential therapeutic strategy for treatment of glioma can be through the inhibition of selected Cl⁻ channels.     

From tumour to tuber; tumour cell characteristics and chromosome numbers of crown gall-derived tetraploid potato plants (Solanum tuberosum cv. ‘Maris Bard’)

Theoretical and Applied Genetics - Agrobacterium tumefaciens strains, known to induce tobacco crown galls that spontaneously develop shoots, were used to induce galls on cultured shoots of a...     

Exogenous tumour necrosis factor α induces suppression of autoimmune arthritis

Introduction  

Our previous studies showed that arthritic Lewis (LEW) rats produced the highest levels of tumour necrosis factor (TNF)α in the recovery phase of adjuvant arthritis (AA), suggesting a correlation between high TNFα levels and reduced severity of arthritis. To further explore this correlation, we compared the TNFα secretion profile of the AA-resistant Wistar Kyoto (WKY) rats with that of LEW rats, determined the effect of exogenous TNFα on the course of AA in LEW rats, and examined various mechanisms involved in TNFα-induced disease modulation.     

Wilms‘瘤的分子生物学

Ｗｉｌｍｓ’瘤，即肾母细胞瘤，是儿童泌尿系统最常见的肿瘤，常合并其他泌尿生殖系统遣传病，本综述了Ｗｉｌｍｓ‘瘤相关的基因－－ＷＴＩ基因的结构、功能、失活机制，以及Ｗｉｌｍｓ瘤和合并Ｗｉｌｍｓ’瘤的一些泌尿生殖系统遗传病的分子生物学发展机理。     

Molecular interactions of ‘high risk’ human papillomaviruses E6 and E7 oncoproteins: implications for tumour progression

The aetiology of cervical cancer has been primarily attributed to human papillomaviruses (HPVs). These are characterized by the persistent expression of the two oncogenes, E6 and E7. Experimental studies show that E6 and E7 genes of the high risk HPVs deregulate key cell cycle controls. Recent work has uncovered new cellular partners for these proteins that throw light on many of the pathways and processes in which these viral proteins intervene. This review focuses on the regulation of host proteins by the viral oncoproteins and consequence of such interactions on cell survival, proliferation, differentiation and apoptosis.     

Implication of TGF-β as a survival factor during tumour development

Transforming growth factor (TGF)-β is a pleiotropic secretory protein which inhibits and potentiates tumour progression during early and late stage of tumourigenicity, respectively. However, it still remains veiled how TGF-β signalling reveals its two faces. Hoshino et al. (Autocrine TGF-β protects breast cancer cells from apoptosis through reduction of BH3-only protein, Bim, J. Biochem. 2011;149:55-65) demonstrated a new aspect of TGF-β as a survival factor in highly metastatic breast cancer cells from which TGF-β1 and TGF-β3 are abundantly expressed. They found that TGF-β suppressed the expression of BH3-only protein Bim which promotes programmed death signalling via release of cytochrome c from mitochondria. Further interestingly, forkhead box C1 (Foxc1) whose expression is suppressed upon TGF-β stimulation is involved in the expression of Bim. Based on their results, autocrine TGF-β signalling in certain breast cancers promotes cell survival via inhibition of apoptotic signalling. Thus, the inhibitors for activin receptor-like kinase (ALK)5 kinase might exert a curative influence on certain types of metastatic breast cancers.     

Effect of 1-methylcyclopropene on postharvest physiological changes of ‘Zaohong’ plum

Plum is a highly perishable fruit and postharvest fruit softening limits its shelf life. The aim of this work was to study the specific effects of 1-methylcyclopropene (1-MCP) treatment on physiological changes in ‘Zaohong’ plums. Plums were treated with 500 nL L⁻¹ 1-MCP at 20°C for 18 h followed by 20°C storage. The results showed that 1-MCP treatment significantly reduced endogenous ethylene production and the activities of ethylene biosynthetic enzymes’ (1-aminocyclopropane-1-carboxylic acid synthase, ACS and 1-aminocyclopropane-1-carboxylic acid oxidase, ACO) in plum fruit during storage when compared with untreated fruit. Although 1-MCP treatment inhibited ethylene production and 1-aminocyclopropane-1-carboxylic acid (ACC) accumulation, it did not inhibit the accumulation of N-malonyl-ACC (MACC). Higher firmness was also found in 1-MCP-treated plums than in controls. During storage, superoxide anion (O₂^−·) and hydrogen peroxide (H₂O₂) levels decreased in 1-MCP-treated fruit. 1-MCP treatment also regulated superoxide dismutase (SOD) and catalase (CAT) activities during storage. Xylanase activity was upregulated while activities of polygalacturonase (PG), pectin methyl esterase (PME) and cellulase enzymes in the fruit were downregulated by 1-MCP treatment. In conclusion, 1-MCP might be a potent compound for extending both storage period and shelf life of ‘Zaohong’ plums by suppressing ethylene biosynthesis, regulating cell wall degradation enzymes and reducing fruit softening.