Epithelial and Stromal Cell Urokinase Plasminogen Activator Receptor Expression Differentially Correlates with Survival in Rectal Cancer Stages B and C Patients

Urokinase plasminogen activator receptor (uPAR) has been proposed as a potential prognostic factor for colorectal cancer (CRC) patient survival. However, CRC uPAR expression remains controversial, especially regarding cell types where uPAR is overexpressed (e.g., epithelium (uPAR^E) or stroma-associated cells (uPAR^S)) and associated prognostic relevance. In this study, two epitope-specific anti-uPAR monoclonal antibodies (MAbs) could discriminate expression of uPAR^E from uPAR^S and were used to examine this association with survival of stages B and C rectal cancer (RC) patients. Using immunohistochemistry, MAbs #3937 and R4 were used to discriminate uPAR^E from uPAR^S respectively in the central and invasive frontal regions of 170 stage B and 179 stage C RC specimens. Kaplan-Meier and Cox regression analyses were used to determine association with survival. uPAR expression occurred in both epithelial and stromal compartments with differential expression observed in many cases, indicating uPAR^E and uPAR^S have different cellular roles. In the central and invasive frontal regions, uPAR^E was adversely associated with overall stage B survival (HR = 1.9; p = 0.014 and HR = 1.5; p = 0.031, respectively) reproducing results from previous studies. uPAR^S at the invasive front was associated with longer stage C survival (HR = 0.6; p = 0.007), reflecting studies demonstrating that macrophage peritumoural accumulation is associated with longer survival. This study demonstrates that different uPAR epitopes should be considered as being expressed on different cell types during tumour progression and at different stages in RC. Understanding how uPAR^E and uPAR^S expression affects survival is anticipated to be a useful clinical prognostic marker of stages B and C RC.     

Array-comparative genomic hybridization reveals loss of SOCS6 is associated with poor prognosis in primary lung squamous cell carcinoma

Background

Primary tumor recurrence commonly occurs after surgical resection of lung squamous cell carcinoma (SCC). Little is known about the genes driving SCC recurrence.

Methods

We used array comparative genomic hybridization (aCGH) to identify genes affected by copy number alterations that may be involved in SCC recurrence. Training and test sets of resected primary lung SCC were assembled. aCGH was used to determine genomic copy number in a training set of 62 primary lung SCCs (28 with recurrence and 34 with no evidence of recurrence) and the altered copy number of candidate genes was confirmed by quantitative PCR (qPCR). An independent test set of 72 primary lung SCCs (20 with recurrence and 52 with no evidence of recurrence) was used for biological validation. mRNA expression of candidate genes was studied using qRT-PCR. Candidate gene promoter methylation was evaluated using methylation microarrays and Sequenom EpiTYPER analysis.

Results

18q22.3 loss was identified by aCGH as being significantly associated with recurrence (p = 0.038). Seven genes within 18q22.3 had aCGH copy number loss associated with recurrence but only SOCS6 copy number was both technically replicated by qPCR and biologically validated in the test set. SOCS6 copy number loss correlated with reduced mRNA expression in the study samples and in the samples with copy number loss, there was a trend for increased methylation, albeit non-significant. Overall survival was significantly poorer in patients with SOCS6 loss compared to patients without SOCS6 loss in both the training (30 vs. 43 months, p = 0.023) and test set (27 vs. 43 months, p = 0.010).

Conclusion

Reduced copy number and mRNA expression of SOCS6 are associated with disease recurrence in primary lung SCC and may be useful prognostic biomarkers.     

Hsp90 Inhibitors Are Efficacious against Kaposi Sarcoma by Enhancing the Degradation of the Essential Viral Gene LANA,of the Viral Co-Receptor EphA2 as well as Other Client Proteins

Heat-shock protein 90 (Hsp90) inhibitors exhibit activity against human cancers. We evaluated a series of new, oral bioavailable, chemically diverse Hsp90 inhibitors (PU-H71, AUY922, BIIB021, NVP-BEP800) against Kaposi sarcoma (KS). All Hsp90 inhibitors exhibited nanomolar EC₅₀ in culture and AUY922 reduced tumor burden in a xenograft model of KS. KS is associated with KS-associated herpesvirus (KSHV). We identified the viral latency associated nuclear antigen (LANA) as a novel client protein of Hsp90 and demonstrate that the Hsp90 inhibitors diminish the level of LANA through proteasomal degradation. These Hsp90 inhibitors also downregulated EphA2 and ephrin-B2 protein levels. LANA is essential for viral maintenance and EphA2 has recently been shown to facilitate KSHV infection; which in turn feeds latent persistence. Further, both molecules are required for KS tumor formation and both were downregulated in response to Hsp90 inhibitors. This provides a rationale for clinical testing of Hsp90 inhibitors in KSHV-associated cancers and in the eradication of latent KSHV reservoirs.     

Effect of fur on pyrC gene expression

The promoter region of pyrC (dihydroorotase) gene of Escherichia coli was shown to have Fur protein binding properties by gel retardation assay. In vivo regulation of the pyrC expression was studied by measuring dihydroorotase activity and beta-galactosidase level in the fur+ and fur- genetic background. The expression of chromosomal dihydroorotase activity and beta-galactosidase activity of pyrC-lacZ fusion plasmid was repressed to about 30% and 17%, respectively in the fur+ strain compared to those in the fur- strain. Divalent ions such as Fe2+ and Zn2+ were not required for the repression. PyrC expression was also reduced to one half by 1 mM uracil. The effect of uracil was independent on the fur gene.     

Characterization of Cadmium-Binding Ligands from Roots of <Emphasis Type="Italic">Echinochloa crusgalli</Emphasis> var. <Emphasis Type="Italic">frumentacea</Emphasis>

Herbaceous Echinochloa crusgalli var. frumentacea is highly resistant to a wide range of heavy metal concentrations. However, its detoxification mechanism has not been reported yet. We exposed plants to 100 μM CdCl₂ for 7 days then examined cadmium accumulation and its subcellular distribution in binding to ligands. Cd concentration increased over the exposure period to a saturation point at day 5 when its level reached 1,732.41 μg g⁻¹ dry weight, an amount about 820 times greater than that found in the control. In the roots, most Cd was distributed in the insoluble fraction, perhaps because of an absorption mechanism at the root surface. Our profile for distribution revealed two Cd-binding ligand peaks: a high molecular weight of 60 kDa and a 2.5-kDa Cd-binding ligand. The latter increased with time and had a typical phytochelatin (PC) amino acid composition of predominantly cysteine, glutamate, and glycine (16.5%, 16.6%, and 11.9%, respectively). The ratio of glutamate/cysteine/glycine was 1.4:1.4:1.0, which is similar to that for other typical PCs.     

Glucose oxidase: natural occurrence, function, properties and industrial applications

Glucose oxidase (GOX) from Aspergillus niger is a well-characterised glycoprotein consisting of two identical 80-kDa subunits with two FAD co-enzymes bound. Both the DNA sequence and protein structure at 1.9 Ǻ have been determined and reported previously. GOX catalyses the oxidation of d-glucose (C₆H₁₂O₆) to d-gluconolactone (C₆H₁₀O₆) and hydrogen peroxide. GOX is produced naturally in some fungi and insects where its catalytic product, hydrogen peroxide, acts as an anti-bacterial and anti-fungal agent. GOX is Generally Regarded As Safe, and GOX from A. niger is the basis of many industrial applications. GOX-catalysed reaction removes oxygen and generates hydrogen peroxide, a trait utilised in food preservation. GOX has also been used in baking, dry egg powder production, wine production, gluconic acid production, etc. Its electrochemical activity makes it an important component in glucose sensors and potentially in fuel cell applications. This paper will give a brief background on the natural occurrence, functions as well as the properties of glucose oxidase. A good coverage on the diverse uses of glucose oxidase in the industry is presented with a brief outline on the working principles in the various settings. Furthermore, food grade GOX preparations are relatively affordable and widely available; the readers may be encouraged to explore other potential uses of GOX. One example is that GOX-catalysed reaction generates significant amount of heat (∼200 kJ/mol), and this property has been mostly neglected in the various applications described so far.