Edie Dullaghan

The role of multiple SOS boxes upstream of the M. tuberculosis lexA gene – identification of a novel DNA-damage inducible gene.

2002-02-01

Four potential binding sites for LexA were identified upstream of the Mycobacterium tuberculosis lexA gene. A mutational analysis of these sites in a lexA-lacZ reporter construct revealed that only one of these SOS boxes was required for DNA-damage-mediated regulation of lexA expression. A novel DNA-damage-inducible gene, Rv2719c, was identified that was divergently transcribed relative to lexA; the other three SOS boxes were found to be involved in regulating expression of this novel mycobacterial-specific gene. The SOS boxes lay in the respective promoter regions of the genes that they regulated.

Two-dimensional DNA displays for comparisons of bacterial genomes.

2003-02-01

We developed two whole genome-scanning techniques to aid in the discovery of polymorphisms as well as horizontally acquired
genes in prokaryotic organisms. First, two-dimensional bacterial genomic display (2DBGD) was developed using restriction enzyme
fragmentation to separate genomic DNA based on size, and then employing denaturing gradient gel electrophoresis (DGGE) in the
second dimension to exploit differences in sequence composition.

An anti-infective peptide that selectively modulates the innate immune response.

We show that an innate defense–regulator peptide (IDR-1) was protective in mouse models of infection with important
Gram-positive and Gram-negative pathogens, including methicillin-resistant Staphylococcus aureus, vancomycin-resistant
Enterococcus and Salmonella enterica serovar Typhimurium.

The Commensal Streptococcus salivarius K12 Downregulates the Innate Immune Responses of Human Epithelial Cells and Promotes Host-Microbe Homeostasis.

2008-08-01

Streptococcus salivarius is an early colonizer of human oral and nasopharyngeal epithelia, and strain K12 has
reported probiotic effects. An emerging paradigm indicates that commensal bacteria downregulate immune
responses through the action on NF-
B signaling pathways, but additional mechanisms underlying probiotic
actions are not well understood. Our objective here was to identify host genes specifically targeted by K12 by
comparing their responses with responses elicited by pathogens and to determine if S. salivarius modulates
epithelial cell immune responses.

A Quantitative Method for the Specific Assessment of Caspase-6 Activity in Cell Culture.

2011-11-01

Aberrant activation of caspase-6 has recently emerged as a major contributor to the pathogeneses of neurodegenerative disorders such as Alzheimer's and Huntington disease. Commercially available assays to measure caspase-6 activity commonly use the VEID peptide as a substrate. However these methods are not well suited to specifically assess caspase-6 activity in the presence of other, confounding protease activities, as often encountered in cell and tissue samples. Here we report the development of a method that overcomes this limitation by using a protein substrate, lamin A, which is highly specific for caspase-6 cleavage at amino acid 230. Using a neo-epitope antibody against cleaved lamin A, we developed an electrochemiluminescence-based ELISA assay that is suitable to specifically detect and quantify caspase-6 activity in highly apoptotic cell extracts. The method is more sensitive than VEID-based assays and can be adapted to a high-content imaging platform for high-throughput screening. This method should be useful to screen for and characterize caspase-6 inhibitor compounds and other interventions to decrease intracellular caspase-6 activity for applications in neurodegenerative disorders.

Hormonal modulators of glial ABCA1 and apoE levels.

2013-09-01

Apolipoprotein E (apoE) is the major lipid carrier in the central nervous system. As apoE plays a major role in the pathogenesis of Alzheimer disease (AD) and also mediates repair pathways after several forms of acute brain injury, modulating the expression, secretion, or function of apoE may provide potential therapeutic approaches for several neurological disorders. Here we show that progesterone and a synthetic progestin, lynestrenol, significantly induce apoE secretion from human CCF-STTG1 astrocytoma cells, whereas estrogens and the progesterone metabolite allopregnanolone have negligible effects. Intriguingly, lynestrenol also increases expression of the cholesterol transporter ABCA1 in CCF-STTG1 astrocytoma cells, primary murine glia, and immortalized murine astrocytes that express human apoE3. The progesterone receptor inhibitor RU486 attenuates the effect of progestins on apoE expression in CCF-STTG1 astrocytoma cells but has no effect on ABCA1 expression in all glial cell models tested, suggesting that the progesterone receptor (PR) may participate in apoE but does not affect ABCA1 regulation.These results suggest that selective reproductive steroid hormones have the potential to influence glial lipid homeostasis through liver X receptor-dependent and progesterone receptor-dependent pathways.

Abstract B065: The RSK/YB-1 pathway represents an opportunity for targeting TNBC and holds promise of treating metastases.

2013-10-01

Triple-negative breast cancers (TNBC) account for 15-25% of all breast cancers, have poor outcomes and high rates of relapse. Along with metastatic disease treatment options for TNBC are limited to that of conventional chemotherapy. The lack of targeted therapies for these cancers is a distinct unmet clinical need. YB-1, a transcription factor, is associated with 70% of TNBC and poor prognosis. While YB-1 is not easily druggable, p90 ribosomal S6 kinase (RSK), which lies upstream of YB-1 and activates it by phosphorylation of serine 102, is an ideal candidate. We recently reported that RSK inhibition decreases TNBC cell growth. Also RSK and YB-1 are implicated in invasion and therefore may play a role in metastatic spread. Herein, we asked whether RSK inhibitors would be beneficial for patients with metastatic disease. Our concern for treating metastatic disease was inspired by a study we conducted in 2222 patients with breast cancer. Women who had local, regional or distant metastases were at a much higher risk of dying. Remarkably those with distant metastases were 100 times for likely to die from breast cancer as compared to those without disseminated disease. Further, women with TNBC specifically had the worst outcomes and their time to death was the shortest of any breast cancer subtype. We therefore conducted a screen of 128 compounds, which are in clinical trials, in SUM149 TNBC cells and compared them to the RSK inhibitor BI-D1870. Most of these drugs failed to inhibit TNBC growth; however, BI-D1870 was highly active. These promising results point towards RSK as a potential molecular target for TNBC yet there are no inhibitors available for use in patients at this time.

Discovery of a 1,2-bis(3-indolyl)ethane that selectively inhibits the pyruvate kinase of methicillin-resistant Staphylococcus aureus over human isoforms.

2014-09-04

Methicillin-resistant Staphylococcus aureus pyruvate kinase (MRSA PK) has recently been identified as a target for development of novel antibacterial agents. Testing a series of 1,2-bis(3-indolyl)ethanes against MRSA PK has led to the discovery of a potent inhibitor that is selective over human isoforms.

Discovery of a 1,2-bis(3-indolyl)ethane that selectively inhibits the pyruvate kinase of methicillin-resistant Staphylococcus aureus over human isoforms. Available from: https://www.researchgate.net/publication/265688711_Discovery_of_a_12-bis3-indolylethane_that_selectively_inhibits_the_pyruvate_kinase_of_methicillin-resistant_Staphylococcus_aureus_over_human_isoforms [accessed Mar 24, 2017].

Innate defense regulator peptide 1018 protects against perinatal brain injury.

2014-03-24

There is currently no pharmacological treatment that provides protection against brain injury in neonates. It is known that activation of an innate immune response is a key, contributing factor in perinatal brain injury; therefore, the neuroprotective therapeutic potential of innate defense regulator peptides (IDRs) was investigated.

Results: IDR-1018 suppresses proinflammatory mediators and cell injurious mechanisms in the developing brain, and postinsult treatment is efficacious in reducing LPS-induced hypoxic-ischemic brain damage. IDR-1018 is effective in the brain when given systemically, confers neuroprotection of both gray and white matter, and lacks significant effects on the brain under normal conditions. Thus, this peptide provides the features of a promising neuroprotective agent in newborns with brain injury.

: A Pan-Cancer Analysis of Alternative Splicing Events Reveals Novel Tumor-Associated Splice Variants of Matriptase.

2014-12-04

High-throughput transcriptome sequencing allows identification of cancer-related changes that occur at the stages of transcription, pre-messenger RNA (mRNA), and splicing. In the current study, we devised a pipeline to predict novel alternative splicing (AS) variants from high-throughput transcriptome sequencing data and applied it to large sets of tumor transcriptomes from The Cancer Genome Atlas (TCGA). We identified two novel tumor-associated splice variants of matriptase, a known cancer-associated gene, in the transcriptome data from epithelial-derived tumors but not normal tissue. Most notably, these variants were found in 69% of lung squamous cell carcinoma (LUSC) samples studied. We confirmed the expression of matriptase AS transcripts using quantitative reverse transcription PCR (qRT-PCR) in an orthogonal panel of tumor tissues and cell lines. Furthermore, flow cytometric analysis confirmed surface expression of matriptase splice variants in chinese hamster ovary (CHO) cells transiently transfected with cDNA encoding the novel transcripts. Our findings further implicate matriptase in contributing to oncogenic processes and suggest potential novel therapeutic uses for matriptase splice variants

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Abstract 2187: Pan-cancer identification and prioritization of cancer-associated differentially expressed genes: A biomarker discovery application.

2017-03-23

Cancer is fundamentally a disease of disordered gene expression. In fact, reversal or neutralization of the changes in gene expression has been shown to be attractive targets for the development of new anti-cancer drugs and therapeutic strategies. New approaches such as antibody drug conjugates (ADCs) also target differentially expressed genes as a mean to recognize tumor cells to selectively deliver toxins to a tumor. In the past decade, the global analysis of gene expression in human cancers have led to the development of a number of potential new biomarkers. For instance, mesothelin (MSLN) was identified as an over-expressed gene in pancreatic cancer and later was proved to be a useful diagnostic marker and so a therapeutic target. Large-scale gene expression analysis, using techniques such as RNA sequencing, provides a powerful tool to identify genes involved in human cancers. In this study, with the ultimate goal being to identify potential novel targets for cancer immunotherapy, we conducted a pan-cancer differential expression analysis in RNA sequencing data from more than 5,000 patients with 25 different cancer types generated by The Cancer Genome Atlas (TCGA). We identified differentially expressed genes (present in at least 5% of samples in a tumor type) in comparison to a large compendium of normal transcriptomes (more than 650 samples, including 30 tissue types) gathered from Genotype-Tissue Expression (GTEx), illumina BodyMap project 2.0, TCGA, and an in-house database. In total, we identified 892 putative tumor-associated differentially expressed genes. In order to further identify novel candidate genes and rank them based on their antigenic potential, we performed an extensive literature search and systematic review to collect the characteristics of an ideal tumor antigen (TA). We developed an Analytic Hierarchy Process (AHP) model - a multiple-criteria decision-making solution - to depict antigen properties for ranking and prioritizing the tumor-associated differentially expressed genes. Our model recognizes the known tumor antigens (such as CA9, Nectin-4, FN1, MSLN and MUC16, which are currently in clinic or pre-clinical studies) in the top 25 of the ranked list. We are currently validating the top-ranked novel antigens in an orthogonal panel of tumor and normal tissues and cell lines using PCR.

Liver-X-Receptor and Progesterone Receptor activation promote apolipoprotein E secretion from CCF-STG1 astrocytoma cells.

2013-11-01

Abstract
Apolipoprotein E is the major lipid carrier in the central nervous system. ApoE plays a major role in the pathogenesis of Alzheimer’s Disease (AD), and also modulates repair pathways after several forms of acute brain injury. Modulating apoE expression, secretion and function may therefore provide potential therapeutic approaches for several neurological disorders. Here we report that natural progesterone and a synthetic progestin, lynestrenol, significantly induce apoE secretion from human astrocytoma cells, whereas estrogens and allopregnanolone have negligible effects. Lynestrenol also increases the expression of the cholesterol transporter ABCA1 at both transcriptional and protein levels via a liver-X-receptor (LXR) dependent pathway. By using a progesterone receptor inhibitor RU486, progesterone receptor (PR) is shown to participate in the regulation of apoE expression in response to lynestrenol and progesterone but has no effect on ABCA1 expression. These results demonstrate the selectivity of certain reproductive hormones to regulate apoE secretion by facilitating both LXR-dependent and PR-dependent pathways in glial cells.

"Discovery and optimization of a new class of pyruvate kinase inhibitors as potential therapeutics for the treatment of methicillin-resistant Staphylococcus aureus infections"

2014-01-24

This paper covers the optimization of a new class of antibiotics for serious Gram+ve infections including MRSA.