ORCID Profile
0000-0003-2165-7813
Current Organisation
University of South Australia
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Publisher: Wiley
Date: 18-04-2023
DOI: 10.1002/YEA.3851
Abstract: Beer refermentation in bottles is an industrial process utilized by breweries where yeast and fermentable extract are added to green beer. The beer is refermented for a minimum of 2 weeks before distribution, with the physiological state of the yeast a critical factor for successful refermentation. Ideally, fresh yeast that is propagated from a dedicated propagation plant should be used for refermentation in bottles. Here, we explored the applicability of the fluorescent and redox‐sensitive dye, resazurin, to assess cellular metabolism in yeast and its ability to differentiate between growth stages. We applied this assay, with other markers of yeast physiology, to evaluate yeast quality during a full‐scale industrial propagation. Resazurin allowed the discrimination between the different growth phases in yeast and afforded a more in‐depth understanding of yeast metabolism during propagation. This assay can be used to optimize the yeast propagation process and cropping time to improve beer quality.
Publisher: Portland Press Ltd.
Date: 15-06-2004
DOI: 10.1042/BJ20031780
Abstract: Two alternatively spliced forms of the human protein tyrosine phosphatase TCPTP (T-cell protein tyrosine phosphatase) exist: a 48 kDa form that is targeted to the endoplasmic reticulum (TC48) and a shorter 45 kDa form that is targeted to the nucleus (TC45). In this study we have identified Ser-304 (Phe301-Asp-His-Ser304-Pro-Asn-Lys307) as a major TCPTP phosphory-lation site and demonstrate that TC45, but not TC48, is phosphorylated on this site in vivo. Phosphorylation of TC45 on Ser-304 was cell cycle-dependent, and increased as cells progressed from G2 into mitosis, but subsided upon mitotic exit. Ser-304 phosphorylation was increased when cells were arrested in mitosis by microtubule poisons such as nocodazole, but remained unaltered when cells were arrested at the G2/M checkpoint by adriamycin. Phosphorylation of Ser-304 did not alter significantly the phosphatase activity or the protein stability of TC45, and had no apparent effect on TC45 localization. Ser-304 phosphorylation was ablated when cells were treated with the CDK (cyclin-dependent protein kinase) inhibitors roscovitine or SU9516, but remained unaltered when ERK1/2 activation was inhibited with the MEK (mitogen-activated protein kinase/extracellular-signal-regulated kinase kinase) inhibitor PD98059. In addition, recombinant CDKs, but not the Polo-like kinase Plk1, phosphorylated Ser-304 in vitro. Our studies identify Ser-304 as a major phosphorylation site in human TCPTP, and the TC45 variant as a novel mitotic CDK substrate.
Publisher: Wiley
Date: 09-05-2016
Abstract: Applying MALDI-MS imaging to tissue microarrays (TMAs) provides access to proteomics data from large cohorts of patients in a cost- and time-efficient way, and opens the potential for applying this technology in clinical diagnosis. The complexity of these TMA data-high-dimensional low s le size-provides challenges for the statistical analysis, as classical methods typically require a nonsingular covariance matrix that cannot be satisfied if the dimension is greater than the s le size. We use TMAs to collect data from endometrial primary carcinomas from 43 patients. Each patient has a lymph node metastasis (LNM) status of positive or negative, which we predict on the basis of the MALDI-MS imaging TMA data. We propose a variable selection approach based on canonical correlation analysis that explicitly uses the LNM information. We apply LDA to the selected variables only. Our method misclassifies 2.3-20.9% of patients by leave-one-out cross-validation and strongly outperforms LDA after reduction of the original data with principle component analysis.
Publisher: Wiley
Date: 07-2010
Abstract: The quality of MALDI-TOF mass spectrometric analysis is highly dependent on the matrix and its deposition strategy. Although different matrix-deposition methods have specific advantages, one major problem in the field of proteomics, particularly with respect to quantitation, is reproducibility between users or laboratories. Compounding this is the varying crystal homogeneity of matrices depending on the deposition strategy used. Here, we describe a novel optimised matrix-deposition strategy for LC-MALDI-TOF/TOF MS using an automated instrument that produces a nebulised matrix "mist" under controlled atmospheric conditions. Comparisons of this with previously reported strategies showed the method to be advantageous for the atypical matrix, 2,5-DHB, and improved phosphopeptide ionisation when compared with deposition strategies for CHCA. This optimised DHB matrix-deposition strategy with LC-MALDI-TOF/TOF MS, termed EZYprep LC, was subsequently optimised for phosphoproteome analysis and compared to LC-ESI-IT-MS and a previously reported approach for phosphotyrosine identification and characterisation. These methods were used to map phosphorylation on epidermal growth factor-stimulated epidermal growth factor receptor to gauge the sensitivity of the proposed method. EZYprep DHB LC-MALDI-TOF/TOF MS was able to identify more phosphopeptides and characterise more phosphorylation sites than the other two proteomic strategies, thus proving to be a sensitive approach for phosphoproteome analysis.
Publisher: Elsevier BV
Date: 05-2014
DOI: 10.1016/J.BBAPAP.2014.01.018
Abstract: The timely detection of gastric cancer will contribute significantly towards effective treatment and is aided by the availability and reliability of appropriate biomarkers. A combination of several biomarkers can improve the sensitivity and specificity of cancer detection and this work reports results from a panel of 4 proteins. By combining a validated preclinical mouse model with a proteomic approach we have recently discovered novel biomarkers for the detection of gastric cancer. Here, we investigate the specificity of four of those biomarkers (afamin, clusterin, VDBP and haptoglobin) for the detection of gastric cancer using two independent methods of validation: ELISA, and a non antibody based method: Multiple Reaction Monitoring with High Resolution Mass Spectrometry (MRM-HR). All four biomarkers reliably differentiated GC from benign patient serum, and also in a small cohort of 11 early stage cases. We also present a novel isoform specific biomarker alpha-1-antitrypsin (A1AT) that was identified using a mouse model for gastric cancer. This isoform is distinct in charge and mobility in a pH gradient and was validated using human s les by isoelectric focussing and Western-blot (IEF-WB). This article is part of a Special Issue entitled: Biomarkers: A Proteomic Challenge.
Publisher: Wiley
Date: 10-05-2016
Abstract: Metastasis is a crucial step of malignant progression and is the primary cause of death from endometrial cancer. However, clinicians presently face the challenge that conventional surgical-pathological variables, such as tumour size, depth of myometrial invasion, histological grade, lymphovascular space invasion or radiological imaging are unable to predict with accuracy if the primary tumour has metastasized. In the current retrospective study, we have used primary tumour s les of endometrial cancer patients diagnosed with (n = 16) and without (n = 27) lymph node metastasis to identify potential discriminators. Using peptide matrix assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI), we have identified m/z values which can classify 88% of all tumours correctly. The top discriminative m/z values were identified using a combination of in situ sequencing and LC-MS/MS from digested tumour s les. Two of the proteins identified, plectin and α-Actin-2, were used for validation studies using LC-MS/MS data independent analysis (DIA) and immunohistochemistry. In summary, MALDI-MSI has the potential to identify discriminators of metastasis using primary tumour s les.
Publisher: Elsevier BV
Date: 12-2001
Publisher: MDPI AG
Date: 06-04-2023
Abstract: Eighty percent of ovarian cancer patients initially respond to chemotherapy, but the majority eventually experience a relapse and die from the disease with acquired chemoresistance. In addition, 20% of patients do not respond to treatment at all, as their disease is intrinsically chemotherapy resistant. Data-independent acquisition nano-flow liquid chromatography–mass spectrometry (DIA LC-MS) identified the three protein markers: gelsolin (GSN), calmodulin (CALM1), and thioredoxin (TXN), to be elevated in high-grade serous ovarian cancer (HGSOC) tissues from patients that responded to chemotherapy compared to those who did not the differential expression of the three protein markers was confirmed by immunohistochemistry. Analysis of the online GENT2 database showed that mRNA levels of GSN, CALM1, and TXN were decreased in HGSOC compared to fallopian tube epithelium. Elevated levels of GSN and TXN mRNA expression correlated with increased overall and progression-free survival, respectively, in a Kaplan–Meier analysis of a large online repository of HGSOC patient data. Importantly, differential expression of the three protein markers was further confirmed when comparing parental OVCAR-5 cells to carboplatin-resistant OVCAR-5 cells using DIA LC-MS analysis. Our findings suggest that GSN, CALM1, and TXN may be useful biomarkers for predicting chemotherapy response and understanding the mechanisms of chemotherapy resistance. Proteomic data are available via ProteomeXchange with identifier PXD033785.
Publisher: Springer Science and Business Media LLC
Date: 21-11-2020
Publisher: Wiley
Date: 14-04-2003
DOI: 10.1002/IJC.11085
Abstract: In de novo glioblastoma multiforme, loss of the tumour suppressor protein PTEN can coincide with the expression of a naturally occurring mutant epidermal growth factor receptor known as deltaEGFR. DeltaEGFR signals constitutively via the phosphatidylinositol 3-kinase (PI3K) rotein kinase Akt and mitogen-activated protein kinase pathways. In human U87MG glioblastoma cells that lack PTEN, deltaEGFR expression enhances tumourigenicity by increasing cellular proliferation. Inhibition of PI3K signaling with the pharmacologic inhibitor wortmannin, or by the reconstitution of physiological levels of PTEN to dephosphorylate the lipid products of PI3K, negated the growth advantage imparted by deltaEGFR on U87MG cells. PTEN reconstitution suppressed the elevated PI3K signaling, without affecting mitogen-activated protein kinase signaling and caused a delay in G1 cell cycle progression that was concomitant with increased cyclin-dependent protein kinase inhibitor p21CIP1/WAF1 protein levels. Our study provides insight into the mechanism by which deltaEGFR may contribute to glioblastoma development.
Publisher: SPIE
Date: 07-2015
DOI: 10.1117/12.2186140
Publisher: MDPI AG
Date: 10-07-2022
DOI: 10.3390/MPS5040057
Abstract: The molecular analysis of small or rare patient tissue s les is challenging and often limited by available technologies and resources, such as reliable antibodies against a protein of interest. Although targeted approaches provide some insight, here, we describe the workflow of two complementary mass spectrometry approaches, which provide a more comprehensive and non-biased analysis of the molecular features of the tissue of interest. Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) generates spatial intensity maps of molecular features, which can be easily correlated with histology. Additionally, liquid chromatography tandem mass spectrometry (LC-MS/MS) can identify and quantify proteins of interest from a consecutive section of the same tissue. Here, we present data from concurrent precancerous lesions from the endometrium and fallopian tube of a single patient. Using this complementary approach, we monitored the abundance of hundreds of proteins within the precancerous and neighboring healthy regions. The method described here represents a useful tool to maximize the number of molecular data acquired from small s le sizes or even from a single case. Our initial data are indicative of a migratory phenotype in these lesions and warrant further research into their malignant capabilities.
Publisher: Elsevier BV
Date: 05-2011
DOI: 10.1016/J.JAUT.2011.02.006
Abstract: The Class IA phosphoinositide 3-kinase delta (PI3Kδ) has been implicated in multiple signaling pathways involved in leukocyte activation and hence is an attractive target in many human autoimmune diseases, including multiple sclerosis (MS). Here, using mice expressing a catalytically inactive form of the PI3Kδ subunit p110δ, we show that signaling through PI3Kδ is required for full and sustained pathology of experimental autoimmune encephalomyelitis (EAE), a Th17-driven model of MS. In p110δ-inactivated mice, T cell activation and function during EAE was markedly reduced and fewer T cells were observed in the central nervous system (CNS). The decrease in T cell activation is unlikely to be due to defects in dendritic cell (DC) function, as p110δ-inactivated DCs migrate and present antigen normally. However, significant increases in the proportion of T cells undergoing apoptosis at early stages of EAE were evident in the absence of PI3Kδ activity. Furthermore, a profound defect in Th17 cellular responses during EAE was apparent in the absence of PI3Kδ activity while Th1 responses were less affected. A highly selective PI3Kδ inhibitor, IC87114, also had greater inhibitory effects on Th17 cell generation in vitro than it did on Th1 cell generation. Thus, PI3Kδ plays an important role in Th17 responses in EAE, suggesting that small molecule inhibitors of PI3Kδ may be useful therapeutics for treatment of MS and other autoimmune diseases.
Publisher: Wiley
Date: 15-10-2019
Abstract: Malignant ascites is a fluid, which builds up in the abdomen and contains cancer cells in the form of single cells or multicellular clusters called spheroids. Malignant ascites has been observed in patients suffering from ovarian, cervical, gastric, colorectal, pancreatic, endometrial, or primary liver cancer. The spheroids are believed to play a major role in chemo resistance and metastasis of the cancer. To ease the discomfort of patients, malignant ascites (MA) is often drained from the abdomen using a procedure called paracentesis. MA retrieved via this minimal invasive procedure is a great source for cancer spheroids, which can be used for testing chemotherapeutic drugs and drug combinations. Herein, the existing workflow is adapted to make concurrent monitoring of drug accumulation, drug response, and drug metabolites feasible using primary spheroids or spheroids grown without a scaffolding matrix. To achieve this, those spheroids are embedded in matrigel, before fixing them with formalin. This makes it possible to process, store, and ship s les at room temperature. This new approach might be used to choose the best targeted therapy for each patient and thereby facilitate personalized medicine.
Publisher: MDPI AG
Date: 19-02-2021
Abstract: Synthetic polypeptides and polymer-peptide hybrid materials have been successfully implemented in an array of biomedical applications owing to their biocompatibility, biodegradability and ability to mimic natural proteins. In addition, these materials have the capacity to form complex supramolecular structures, facilitate specific biological interactions, and incorporate a erse selection of functional groups that can be used as the basis for further synthetic modification. Like conventional synthetic polymers, polypeptide-based materials can be designed to respond to external stimuli (e.g., light and temperature) or changes in the environmental conditions (e.g., redox reactions and pH). In particular, pH-responsive polypeptide-based systems represent an interesting avenue for the preparation of novel drug delivery systems that can exploit physiological or pathological pH variations within the body, such as those that arise in the extracellular tumour microenvironment, intracellularly within endosomes/lysosomes, or during tissue inflammation. Here, we review the significant progress made in advancing pH-responsive polypeptides and polymer-peptide hybrid materials during the last five years, with a particular emphasis on the manipulation of ionisable functional groups, pH-labile linkages, pH-sensitive changes to secondary structure, and supramolecular interactions.
Publisher: Elsevier BV
Date: 09-2005
DOI: 10.1016/J.EJCB.2005.04.002
Abstract: In the cell cycle the transition from G2 phase to cell ision (M) is strictly controlled by protein phosphorylation-dephosphorylation reactions effected by several protein kinases and phosphatases. Although much indirect and direct evidence point to a key role of protein phosphatase 2A (PP2A) at the G2/M transition, the control of the enzyme activity prior to and after the transition are not fully clarified. Using synchronized HeLa cells we determined the PP2A activity (i.e. the increment sensitive to inhibition by 2nM okadaic acid) in immunoprecipitates obtained with antibodies raised against a conserved peptide sequence (residues 169-182, Ab(169/182)) of the PP2A catalytic subunit (PP2A C). Two different substrates were offered: the phospho-peptide KR(p)TIRR and histone H1 phosphorylated by means of the cyclin-dependent protein kinase p34(cdc2). The results indicate that in HeLa cells the specific activity of PP2A towards both substrates goes through a minimum in late G2 phase and stays low until metaphase. Treatment of G2 cells with TPA (10(-7) M) caused a reactivation of the downregulated PP2A activity within 20 min, i.e. the same time frame within which TPA was shown earlier to block HeLa cells at the transition from G2 to mitosis [Kinzel et al., 1988. Cancer Res. 48, 1759-1762]. Activation of PP2A was also induced by TPA in mitotic cells. The low activity of PP2A in mitotic cells was accompanied by a strong reaction of mitotic PP2A C with anti-P-Tyr antibodies in Western blots, which was reversed by treatment of mitotic cells with TPA. The results suggest that the activity of cellular PP2A requires downregulation for the transition from G2 phase to mitosis. Unscheduled reactivation of PP2A induced by TPA in late G2 phase appears to inhibit the progress into mitosis.
Publisher: Frontiers Media SA
Date: 25-09-2020
Publisher: Wiley
Date: 09-02-2018
Abstract: Multicellular tumor spheroids (MCTS) are a powerful biological in vitro model, which closely mimics the 3D structure of primary avascularized tumors. Mass spectrometry (MS) has established itself as a powerful analytical tool, not only to better understand and describe the complex structure of MCTS, but also to monitor their response to cancer therapeutics. The first part of this review focuses on traditional mass spectrometry approaches with an emphasis on elucidating the molecular characteristics of these structures. Then the mass spectrometry imaging (MSI) approaches used to obtain spatially defined information from MCTS is described. Finally the analysis of primary spheroids, such as those present in ovarian cancer, and the great potential that mass spectrometry analysis of these structures has for improved understanding of cancer progression and for personalized in vitro therapeutic testing is discussed.
Publisher: MDPI AG
Date: 27-10-2021
Abstract: Matrix assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) can determine the spatial distribution of analytes such as protein distributions in a tissue section according to their mass-to-charge ratio. Here, we explored the clinical potential of machine learning (ML) applied to MALDI MSI data for cancer diagnostic classification using tissue microarrays (TMAs) on 302 colorectal (CRC) and 257 endometrial cancer (EC)) patients. ML based on deep neural networks discriminated colorectal tumour from normal tissue with an overall accuracy of 98% in balanced cross-validation (98.2% sensitivity and 98.6% specificity). Moreover, our machine learning approach predicted the presence of lymph node metastasis (LNM) for primary tumours of EC with an accuracy of 80% (90% sensitivity and 69% specificity). Our results demonstrate the capability of MALDI MSI for complementing classic histopathological examination for cancer diagnostic applications.
Publisher: Elsevier
Date: 2017
DOI: 10.1016/BS.ACR.2016.11.002
Abstract: Pathologists play an essential role in the diagnosis and prognosis of benign and cancerous tumors. Clinicians provide tissue s les, for ex le, from a biopsy, which are then processed and thin sections are placed onto glass slides, followed by staining of the tissue with visible dyes. Upon processing and microscopic examination, a pathology report is provided, which relies on the pathologist's interpretation of the phenotypical presentation of the tissue. Targeted analysis of single proteins provide further insight and together with clinical data these results influence clinical decision making. Recent developments in mass spectrometry facilitate the collection of molecular information about such tissue specimens. These relatively new techniques generate label-free mass spectra across tissue sections providing nonbiased, nontargeted molecular information. At each pixel with spatial coordinates (x/y) a mass spectrum is acquired. The acquired mass spectrums can be visualized as intensity maps displaying the distribution of single m/z values of interest. Based on the s le preparation, proteins, peptides, lipids, small molecules, or glycans can be analyzed. The generated intensity maps/images allow new insights into tumor tissues. The technique has the ability to detect and characterize tumor cells and their environment in a spatial context and combined with histological staining, can be used to aid pathologists and clinicians in the diagnosis and management of cancer. Moreover, such data may help classify patients to aid therapy decisions and predict outcomes. The novel complementary mass spectrometry-based methods described in this chapter will contribute to the transformation of pathology services around the world.
Publisher: Informa UK Limited
Date: 03-2003
Publisher: Elsevier BV
Date: 03-2024
Publisher: Wiley
Date: 11-2010
DOI: 10.1002/0471140864.PS1311S62
Abstract: This unit describes methods to detect, identify, and characterize tyrosine phosphorylation in proteins by mass spectrometry, including s le preparation methods, enrichment strategies using phosphotyrosine‐specific antibodies, and chromatographic separation methods. Curr. Protoc. Protein Sci . 62:13.11.1‐13.11.26. © 2010 by John Wiley & Sons, Inc.
Publisher: Hindawi Limited
Date: 11-07-2016
DOI: 10.1002/HUMU.23032
Abstract: Ectrodactyly/split hand-foot malformation is genetically heterogeneous with more than 100 syndromic associations. Acinar dysplasia is a rare congenital lung lesion of unknown etiology, which is frequently lethal postnatally. To date, there have been no reports of combinations of these two phenotypes. Here, we present an infant from a consanguineous union with both ectrodactyly and autopsy confirmed acinar dysplasia. SNP array and whole-exome sequencing analyses of the affected infant identified a novel homozygous Fibroblast Growth Factor Receptor 2 (FGFR2) missense mutation (p.R255Q) in the IgIII domain (D3). Expression studies of Fgfr2 in development show localization to the affected limbs and organs. Molecular modeling and genetic and functional assays support that this mutation is at least a partial loss-of-function mutation, and contributes to ectrodactyly and acinar dysplasia only in homozygosity, unlike previously reported heterozygous activating FGFR2 mutations that cause Crouzon, Apert, and Pfeiffer syndromes. This is the first report of mutations in a human disease with ectrodactyly with pulmonary acinar dysplasia and, as such, homozygous loss-of-function FGFR2 mutations represent a unique syndrome.
Publisher: American Chemical Society (ACS)
Date: 09-2020
Publisher: Wiley
Date: 03-0001
Abstract: Cytoskeletal proteins are essential building blocks of cells. More than 100 cytoskeletal and cytoskeleton-associated proteins are known and for some, their function and regulation are understood in great detail. Apart from cell shape and support, they facilitate many processes such as intracellular signaling and transport, and cancer related processes such as proliferation, migration, and invasion. During the last decade, comparative proteomic studies have identified cytoskeletal proteins as in vitro markers for tumor progression and metastasis. Here, these results are summarized and a number of unrelated studies are highlighted, identifying the same cytoskeletal proteins as potential biomarkers. These findings might indicate that the abundance of these potential markers of tumor progression is associated with the biological outcome and are independent of the cancer origin. This correlates well with recently published results from the Cancer Genome Atlas, indicating that cancers show remarkable similarities in their analyzed molecular information, independent of their organ of origin. It is postulated that the quantification of cytoskeletal proteins in healthy tissues, tumors, in adjacent tissues, and in stroma, is a great source of molecular information, which might not only be used to classify tumors, but more importantly to predict patients' outcome or even best treatment choices.
Publisher: Springer Science and Business Media LLC
Date: 24-01-2017
DOI: 10.1038/TP.2016.261
Abstract: Meta-analyses of genome-wide association studies (meta-GWASs) and candidate gene studies have identified genetic variants associated with cardiovascular diseases, metabolic diseases and mood disorders. Although previous efforts were successful for in idual disease conditions (single disease), limited information exists on shared genetic risk between these disorders. This article presents a detailed review and analysis of cardiometabolic diseases risk (CMD-R) genes that are also associated with mood disorders. First, we reviewed meta-GWASs published until January 2016, for the diseases ‘type 2 diabetes, coronary artery disease, hypertension’ and/or for the risk factors ‘blood pressure, obesity, plasma lipid levels, insulin and glucose related traits’. We then searched the literature for published associations of these CMD-R genes with mood disorders. We considered studies that reported a significant association of at least one of the CMD-R genes and ‘depression’ or ‘depressive disorder’ or ‘depressive symptoms’ or ‘bipolar disorder’ or ‘lithium treatment response in bipolar disorder’, or ‘serotonin reuptake inhibitors treatment response in major depression’. Our review revealed 24 potential pleiotropic genes that are likely to be shared between mood disorders and CMD-Rs. These genes include MTHFR , CACNA1D , CACNB2 , GNAS , ADRB1 , NCAN , REST , FTO , POMC , BDNF , CREB , ITIH4 , LEP , GSK3B , SLC18A1 , TLR4 , PPP1R1B , APOE , CRY2 , HTR1A , ADRA2A , TCF7L2 , MTNR1B and IGF1 . A pathway analysis of these genes revealed significant pathways: corticotrophin-releasing hormone signaling , AMPK signaling , cAMP-mediated or G-protein coupled receptor signaling , axonal guidance signaling , serotonin or dopamine receptors signaling , d opamine-DARPP32 feedback in cAMP signaling , circadian rhythm signaling and leptin signaling . Our review provides insights into the shared biological mechanisms of mood disorders and cardiometabolic diseases.
Publisher: SPIE
Date: 09-02-2012
DOI: 10.1117/12.909681
Publisher: Springer Science and Business Media LLC
Date: 23-06-2018
DOI: 10.1038/LEU.2017.196
Publisher: MDPI AG
Date: 15-08-2023
Abstract: Nanoparticle-based therapeutics are being clinically translated for treating cancer. Even when thought to be biocompatible, nanoparticles are being increasingly identified as altering cell regulation and homeostasis. Antioxidant pathways are important for maintaining cell redox homeostasis and play important roles by maintaining ROS levels within tolerable ranges. Here, we sought to understand how a model of a relatively inert nanoparticle without any therapeutic agent itself could antagonize a cancer cell lines’ antioxidant mechanism. A label-free protein expression approach was used to assess the glutathione-thioredoxin antioxidative pathway in a prostate cancer cell line (PC-3) after exposure to gold nanoparticles conjugated with a targeting moiety (transferrin). The impact of the nanoparticles was also corroborated through morphological analysis with TEM and classification of pro-apoptotic cells by way of the sub-G0/G1 population via the cell cycle and annexin V apoptosis assay. After a two-hour exposure to nanoparticles, major proteins associated with the glutathione-thioredoxin antioxidant pathway were downregulated. However, this response was acute, and in terms of protein expression, cells quickly recovered within 24 h once nanoparticle exposure ceased. The impact on PRDX-family proteins appears as the most influential factor in how these nanoparticles induced an oxidative stress response in the PC-3 cells. An apparent adaptive response was observed if exposure to nanoparticles continued. Acute exposure was observed to have a detrimental effect on cell viability compared to continuously exposed cells. Nanoparticle effects on cell regulation likely provide a compounding therapeutic advantage under some circumstances, in addition to the action of any cytotoxic agents however, any therapeutic advantage offered by nanoparticles themselves with regard to vulnerabilities specific to the glutathione-thioredoxin antioxidative pathway is highly temporal.
Publisher: Hindawi Limited
Date: 11-08-2016
DOI: 10.1002/HUMU.23058
Publisher: Wiley
Date: 09-11-2019
Abstract: Protein glycosylation, particularly N-linked glycosylation, is a complex posttranslational modification (PTM), which plays an important role in protein folding and conformation, regulating protein stability and activity, cell-cell interaction, and cell signaling pathways. This review focuses on analytical techniques, primarily MS-based techniques, to qualitatively and quantitatively assess N-glycosylation while successfully characterizing compositional, structural, and linkage features with high specificity and sensitivity. The analytical techniques explored in this review include LC-ESI-MS/MS and MALDI time-of-flight MS (MALDI-TOF-MS), which have been used to analyze clinical s les, such as serum, plasma, ascites, and tissue. Targeting the aberrant N-glycosylation patterns observed in MALDI-MS imaging (MSI) offers a platform to visualize N-glycans in tissue-specific regions. The studies on the intra-patient (i.e., a comparison of tissue-specific regions from the same patient) and inter-patient (i.e., a comparison of tissue-specific regions between different patients) variation of early- and late-stage ovarian cancer (OC) patients identify specific N-glycan differences that improve understanding of the tumor microenvironment and potentially improve therapeutic strategies for the clinic.
Publisher: Wiley
Date: 15-12-2016
Abstract: This review discusses the current status of proteomics technology in endometrial cancer diagnosis, treatment and prognosis. The first part of this review focuses on recently identified biomarkers for endometrial cancer, their importance in clinical use as well as the proteomic methods used in their discovery. The second part highlights some of the emerging mass spectrometry based proteomic technologies that promise to contribute to a better understanding of endometrial cancer by comparing the abundance of hundreds or thousands of proteins simultaneously.
Publisher: American Chemical Society (ACS)
Date: 19-09-2016
DOI: 10.1021/ACS.JPROTEOME.6B00053
Abstract: Although acetylation is regarded as a common protein modification, a detailed proteome-wide profile of this post-translational modification may reveal important biological insight regarding differential acetylation of in idual proteins. Here we optimized a novel peptide IEF fractionation method for use prior to LC-MS/MS analysis to obtain a more in depth coverage of N-terminally acetylated proteins from complex s les. Application of the method to the analysis of the serous ovarian cancer cell line OVCAR-5 identified 344 N-terminally acetylated proteins, 12 of which are previously unreported. The protein peptidyl-prolyl cis-trans isomerase A (PPIA) was detected in both the N-terminally acetylated and unmodified forms and was further analyzed by data-independent acquisition in carboplatin-responsive parental OVCAR-5 cells and carboplatin-resistant OVCAR-5 cells. This revealed a higher ratio of unacetylated to acetylated N-terminal PPIA in the parental compared with the carboplatin-resistant OVCAR-5 cells and a 4.1-fold increase in PPIA abundance overall in the parental cells relative to carboplatin-resistant OVCAR-5 cells (P = 0.015). In summary, the novel IEF peptide fractionation method presented here is robust, reproducible, and can be applied to the profiling of N-terminally acetylated proteins. All mass spectrometry data is available as a ProteomeXchange repository (PXD003547).
Publisher: Elsevier BV
Date: 07-2017
DOI: 10.1016/J.BBAPAP.2016.10.010
Abstract: The prediction of lymph node metastasis using clinic-pathological data and molecular information from endometrial cancers lacks accuracy and is therefore currently not routinely used in patient management. Consequently, although only a small percentage of patients with endometrial cancers suffer from metastasis, the majority undergo radical surgery including removal of pelvic lymph nodes. Upon analysis of publically available data and published research, we compiled a list of 60 proteins having the potential to display differential abundance between primary endometrial cancers with versus those without lymph node metastasis. Using data dependent acquisition LC-ESI-MS/MS we were able to detect 23 of these proteins in endometrial cancers, and using data independent LC-ESI-MS/MS the differential abundance of five of those proteins was observed. The localization of the differentially expressed proteins, was visualized using peptide MALDI MSI in whole tissue sections as well as tissue microarrays of 43 patients. The proteins identified were further validated by immunohistochemistry. Our data indicate that annexin A2 protein level is upregulated, whereas annexin A1 and α actinin 4 expression are downregulated in tumours with lymph node metastasis compared to those without lymphatic spread. Moreover, our analysis confirmed the potential of these markers, to be included in a statistical model for prediction of lymph node metastasis. The predictive model using highly ranked m/z values identified by MALDI MSI showed significantly higher predictive accuracy than the model using immunohistochemistry data. In summary, using publicly available data and complementary proteomics approaches, we were able to improve the prediction model for lymph node metastasis in EC.
Publisher: Wiley
Date: 28-04-2015
DOI: 10.1038/ICB.2015.35
Abstract: Phosphoinositide 3-kinase γ (PI3Kγ) consists of the catalytic subunit p110γ that forms a mutually exclusive heterodimer with one of the two adaptor subunits, p101 or p84. Although activation of PI3Kγ is necessary for cell migration downstream of G-protein-coupled receptor engagement, particularly within the immune system, aberrant PI3Kγ signalling has been associated with transformation, increased migration and the progression of multiple cancer types. Regulation of PI3Kγ signal activation and duration is critical to controlling and maintaining coordinated cellular migration however, the mechanistic basis for this is not well understood. We have recently demonstrated that, in contrast to the tumour-promoting potential of p110γ and p101, p84 possesses tumour-suppressor activity, suggesting a negative regulatory role within PI3Kγ signalling. The present study investigated the role of p84 phosphorylation in the context of PI3Kγ signalling, cell migration and p84-mediated tumour suppression. Two putative phosphorylation sites were characterised within p84, Ser358 and Thr607. Expression of wild-type p84 reduced the oncogenic potential of MDA.MB.231 cells and inhibited metastatic lung colonisation in vivo, effects that were dependent on Thr607. Furthermore, loss of Thr607 enhanced migration of MDA.MB.231 cells in vitro and prevented the induction of p84 110γ dimers. The dimerisation of wild-type p84 with p110γ was not detected at the plasma membrane, indicating an inhibitory interaction preventing PI3Kγ lipid-kinase activity. In contrast, Ser358 phosphorylation was not determined to be critical for p84 activity in the context of migration. Our findings suggest that p84 binding to p110γ may represent a novel negative feedback signal that terminates PI3Kγ activity.
Publisher: MDPI AG
Date: 08-07-2016
DOI: 10.3390/IJMS17071088
Publisher: MDPI AG
Date: 10-05-2022
Abstract: The five-year survival rate for women with ovarian cancer is very poor despite radical cytoreductive surgery and chemotherapy. Although most patients initially respond to platinum-based chemotherapy, the majority experience recurrence and ultimately develop chemoresistance, resulting in fatal outcomes. The current administration of cytotoxic compounds is h ered by dose-limiting severe adverse effects. There is an unmet clinical need for targeted drug delivery systems that transport chemotherapeutics selectively to tumor cells while minimizing off-target toxicity. G protein-coupled receptors (GPCRs) are the largest family of membrane receptors, and many are overexpressed in solid tumors, including ovarian cancer. This review summarizes the progress in engineered nanoparticle research for drug delivery for ovarian cancer and discusses the potential use of GPCRs as molecular entry points to deliver anti-cancer compounds into ovarian cancer cells. A newly emerging treatment paradigm could be the personalized design of nanomedicines on a case-by-case basis.
Publisher: American Chemical Society (ACS)
Date: 29-10-2019
DOI: 10.1021/ACS.ANALCHEM.9B03091
Abstract: The strength of MALDI-MSI is to analyze and visualize spatial intensities of molecular features from an intact tissue. The distribution of the intensities can then be visualized within a single tissue section or compared in between sections, acquired consecutively. This method can be reliably used to reveal physiological structures and has the potential to identify molecular details, which correlate with biological outcomes. MALDI-MSI implementation in clinical laboratories requires the ability to ensure method quality and validation to meet diagnostic expectations. To be able to get consistent qualitative and quantitative results, standardized s le preparation and data acquisition are of highest priority. We have previously shown that the deposition of internal standards onto the tissue section during s le preparation can be used to improve the mass accuracy of monitored
Publisher: Wiley
Date: 16-03-2021
Abstract: The applicability of mass spectrometry imaging (MSI) has exponentially increased with the improvement of s le preparation, instrumentation (spatial resolution) and data analysis. The number of MSI publications listed in PubMed continues to grow with 378 published articles in 2020‐2021. Initially, MSI was just sensitive enough to identify molecular features correlating with distinct tissue regions, similar to the resolution achieved by visual inspection after standard immunohistochemical staining. Although the spatial resolution was limited compared with other imaging modalities, the molecular intensity mapping added a new exciting capability. Over the past decade, significant improvements in every step of the workflow and most importantly in instrumentation were made, which now enables the molecular analysis at a cellular and even subcellular level. Here, we summarize the latest developments in MSI, with a focus on the latest approaches for tissue‐based imaging described in 2020.
Publisher: Frontiers Media SA
Date: 15-12-2020
Abstract: Serous endometrial cancer (SEC) and high grade serous ovarian cancer (HGSOC) are aggressive gynecological malignancies with high rates of metastasis and poor prognosis. Endometrial intraepithelial carcinoma (EIC), the precursor for SEC, and serous tubal intraepithelial carcinoma (STIC), believed to be the precursor lesion for HGSOC, can also be associated with intraabdominal spread. To provide insight into the etiology of these precancerous lesions and to explore the potential molecular mechanisms underlying their metastatic behavior, we performed a proteomic mass spectrometry analysis in a patient with synchronous EIC and STIC. Through histological and molecular identification of precancerous lesions followed by laser capture microdissection, we were able to identify over 450 proteins within the precancerous lesions and adjacent healthy tissue. The proteomic analysis of STIC and EIC showed remarkable overlap in the proteomic patterns, reflecting early neoplastic changes in proliferation, loss of polarity and attachment. Our proteomic analysis showed that both EIC and STIC, despite being regarded as premalignant lesions, have metastatic potential, which correlates with the common presentation of invasive serous gynecological malignancies at advanced stage.
Start Date: 2000
End Date: 2002
Funder: German Research Foundation
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