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David Elashoff,
Hui Zhou,
Jean Reiss,
Jianghua Wang,
Hua Xiao,
Bradley Henson, Shen Hu,
Martha Arellano,
Uttam Sinha,
Anh Le, [......],
Vishad Nabili,
Mark Lingen,
Darly Morris,
Timothy Randolph,
Ziding Feng,
David Akin,
Dragana A Kastratovic,
David Chia,
Elliot Abemayor,
David T W Wong
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ABSTRACT: Oral cancer is the sixth most common cancer with a 5-year survival rate of approximately 60%. Presently, there are no scientifically credible early detection techniques beyond conventional clinical oral examination. The goal of this study is to validate whether the seven mRNAs and three proteins previously reported as biomarkers are capable of discriminating patients with oral squamous cell carcinomas (OSCC) from healthy subjects in independent cohorts and by a National Cancer Institute (NCI)-Early Detection Research Network (EDRN)-Biomarker Reference Laboratory (BRL).
Three hundred and ninety-five subjects from five independent cohorts based on case controlled design were investigated by two independent laboratories, University of California, Los Angeles (Los Angeles, CA) discovery laboratory and NCI-EDRN-BRL.
Expression of all seven mRNA and three protein markers was increased in OSCC versus controls in all five cohorts. With respect to individual marker performance across the five cohorts, the increase in interleukin (IL)-8 and subcutaneous adipose tissue (SAT) was statistically significant and they remained top performers across different cohorts in terms of sensitivity and specificity. A previously identified multiple marker model showed an area under the receiver operating characteristic (ROC) curve for prediction of OSCC status ranging from 0.74 to 0.86 across the cohorts.
The validation of these biomarkers showed their feasibility in the discrimination of OSCCs from healthy controls. Established assay technologies are robust enough to perform independently. Individual cutoff values for each of these markers and for the combined predictive model need to be further defined in large clinical studies.
Salivary proteomic and transcriptomic biomarkers can discriminate oral cancer from control subjects.
Cancer Epidemiology Biomarkers & Prevention 03/2012; 21(4):664-72. · 4.12 Impact Factor
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ABSTRACT: Sjögren's syndrome (SS) is an autoimmune inflammatory disorder of exocrine glands. SS particularly affects the lacrimal and salivary glands. Dry mouth and dry eyes are frequently proffered as presenting symptoms, but nonspecific symptoms such as malaise and fatigue, and extraglandular manifestations like purpura, polyneuropathy and arthritis are also often present. Moreover, lymphomas develop in about 7.5% of SS patients, mostly marginal zone B-cell lymhomas. Futhermore, SS has a very substantial impact on the patients' quality of life and their daily activities. Recently, many breakthroughs have been seen in salivary diagnostics, which not only can be used for diagnosis but also for monitoring of disease activity and disease progression as well as for objectively scoring the effect of intervention treatment with biologicals. In addition, salivary proteomics, genomics and system biology have been shown to be very promising tools in unravelling the pathophysiology of SS, thus providing in depth insight in its underlying mechanisms which could give clues for intervention therapies with biologicals. The latter is of particular interest as B cell depletion therapy has been shown a very promising therapy for a subgroup of SS patients. When applying salivary diagnostics in combination with instruments to rate disease activity and progression in SS, one might be able to select those SS patients who respond to a particular type of biological. These topics are addressed in this review and promises for the near future are described.
Current pharmaceutical biotechnology 01/2012; 13(10):2026-45. · 3.40 Impact Factor
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PROTEOMICS - CLINICAL APPLICATIONS 10/2011; 5(9-10):553. · 1.81 Impact Factor
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ABSTRACT: Sjögren’s syndrome (SS) is a chronic, progressive autoimmune disease primarily affecting women. Diagnosis of SS requires an invasive salivary gland tissue biopsy and a long delay from the start of the symptoms to final diagnosis has been frequently observed. In this study,we aim to identify salivary autoantibody biomarkers for primary SS (pSS) using a protein microarray approach. Immune-response protoarrays were used to profile saliva autoantibodies from patients with pSS (n = 514), patients with systemic lupus erythematosus(SLE, n = 513), and healthy control subjects (n = 513). We identified 24 potential autoantibody biomarkers that can discriminate patients with pSS from both patients with SLE and healthy individuals. Four saliva autoantibody biomarkers, anti-transglutaminase, anti-histone, anti-SSA, and anti-SSB, were further tested in independent pSS (n = 534), SLE (n = 534), and healthy control (n = 534) subjects and all were successfully validated with ELISA. This study has demonstrated the potential of a high-throughput protein microarray approach for the discovery of autoantibody biomarkers. The identified saliva autoantibody biomarkers may lead to a clinical tool for simple, noninvasive detection of pSS at low cost.
Proteomics 02/2011; 11(8):1499-507. · 4.43 Impact Factor
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ABSTRACT: Sjögren’s syndrome (SS) is a systemic autoimmune disease with a variety of presenting symptoms that may delay its diagnosis. We previously discovered a number of candidate salivary biomarkers for primary SS using both mass spectrometry and expression microarray analysis. In the current study, we aimed to verify these candidate biomarkers in independent patient populations and to evaluate their predictive values for primary SS detection.
In total, 34 patients with primary SS, 34 patients with systemic lupus erythematosus (SLE), and 34 healthy individuals were enrolled for the validation studies. Salivary protein biomarkers were measured using either Western blotting or enzyme-linked immunosorbent assay, and the messenger RNA (mRNA) biomarkers were measured using quantitative polymerase chain reaction. Statistical analysis was performed using R software, version 2.9.
Three protein biomarkers (cathepsin D [CPD], α-enolase, and ß₂-microglobulin [ß₂m]) and 3 mRNA biomarkers (myeloid cell nuclear differentiation antigen [MNDA], guanylate binding protein 2 [GBP-2], and low-affinity IIIb receptor for the Fc fragment of IgG) were significantly elevated in patients with primary SS compared with both SLE patients and healthy controls. The combination of 3 protein biomarkers, CPD, α-enolase, and ß₂m, yielded a receiver operating characteristic (ROC) value of 0.99 in distinguishing primary SS from healthy controls. The combination of protein biomarkers ß₂m and 2 mRNA biomarkers, MNDA and GBP-2, reached an ROC of 0.95 in discriminating primary SS from SLE.
We have successfully verified a panel of protein and mRNA biomarkers that can discriminate primary SS from both SLE and healthy controls. If further validated in patients with primary SS and those with sicca symptoms but no autoimmune disease, these biomarkers may lead to a simple yet highly discriminatory clinical tool for diagnosis of primary SS.
Arthritis care & research. 11/2010; 62(11):1633-8.
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ABSTRACT: Saliva harbors a wide spectrum of proteins that may reflect the health/disease status in the human body. Profiling of the proteins in saliva from a disease population can potentially yield valuable clinical parameters to be used for diagnosis and prognosis of the disease. Advances in proteomic technologies have enabled comprehensive profiling of protein expression in cells, tissue, and body fluids. When applied to readily accessible saliva samples from disease patients for biomarker study, such a global approach allows attaining the most discriminatory protein biomarkers that can best predict the disease status. In this chapter, we describe the protocols for proteomic analysis of saliva using 2D gel electrophoresis, Western blotting, and LC-MS/MS.
Methods in molecular biology (Clifton, N.J.) 01/2010; 666:31-41.
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ABSTRACT: Pancreatic cancer is one of the most aggressive human tumors due to its high potential of local invasion and metastasis. The aim of this study was to characterize the membrane proteomes of pancreatic ductal adenocarcinoma (PDAC) cells of primary and metastatic origins, and to identify potential target proteins related to metastasis of pancreatic cancer.
Membrane/membrane-associated proteins were isolated from AsPC-1 and BxPC-3 cells and identified with a proteomic approach based on SDS-PAGE, in-gel tryptic digestion and liquid chromatography with tandem mass spectrometry (LC-MS/MS). X! Tandem was used for database searching against the SwissProt human protein database.
We identified 221 & 208 proteins from AsPC-1 and BxPC-3 cells, respectively, most of which are membrane or membrane-associated proteins. A hundred and nine proteins were found in both cell lines while the others were present in either AsPC-1 or BxPC-3 cells. Differentially expressed proteins between two cell lines include modulators of cell adhesion, cell motility or tumor invasion as well as metabolic enzymes involved in glycolysis, tricarboxylic acid cycle, or nucleotide/lipid metabolism.
Membrane proteomes of AsPC-1 (metastatic) and BxPC-3 (primary) cells are remarkably different. The differentially expressed membrane proteins may serve as potential targets for diagnostic and therapeutic interventions.
Journal of Biomedical Science 01/2010; 17:74. · 2.01 Impact Factor
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ABSTRACT: Oral cancer, predominantly oral squamous cell carcinoma, is considered as the 6th most common human cancer in the world. The
American Cancer Society estimated that 35,310 new cases of oral cancer were diagnosed in 2008 and 7,590 patients died from
the disease in the US. Worldwide, oral cancer is a major cancer problem, with an estimated 300,000 new cases diagnosed annually.
Patients with OSCC are often diagnosed at a late stage, and there is a high recurrence rate after treatment, especially in
those with neck lymph node metastasis. The overall 5-year survival rates for oral cancer have remained low and stagnant during
the past few decades. The high mortality rate can be attributed to factors including nonresponsiveness to chemotherapy and
radiation therapy, late presentation of the lesions, and the lack of biological markers for the early detection of these lesions.
In this chapter, we have provided a brief introduction on current genomic and proteomic technologies and review their applications
in molecular analysis of oral cancer. A precise molecular portrait of oral carcinogenesis will improve diagnosis, treatment,
and monitoring of the disease.
11/2009: pages 105-120;
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Fang Wei,
Prabhudas Patel,
Wei Liao,
Kishore Chaudhry,
Lei Zhang,
Martha Arellano-Garcia, Shen Hu,
David Elashoff,
Hui Zhou,
Shilin Shukla,
Franky Shah,
Chih-Ming Ho,
David T Wong
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ABSTRACT: Multiplexing assay of biomarkers at the point-of-care is an elusive goal for molecular diagnostics.
Here, we report an electrochemical (EC) sensor for oral cancer detection based on the simultaneous detection of two salivary biomarkers: interleukin (IL)-8 mRNA and IL-8 protein.
Under the multiplexing mode, the limit of detection of salivary IL-8 mRNA reaches to 3.9 fM and 7.4 pg/mL for IL-8 protein in saliva. Multiplex assay of these 2 biomarkers directly from 28 cancer and 28 matched control saliva samples shows significant difference between the two groups. From the receiver operating characteristic analysis, the EC sensor yields around 90% sensitivity and specificity for both IL-8 mRNA and IL-8 protein, which are very close to the data measured by traditional assays (ELISA and PCR) with the same group of saliva. Combined IL-8 mRNA and protein show better AUC compared with single biomarker.
We show, for the first time, concurrently multiplexing detection of salivary mRNA and protein biomarkers using point-of-care EC sensor.
Clinical Cancer Research 07/2009; 15(13):4446-52. · 7.74 Impact Factor
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ABSTRACT: Saliva (oral fluids) is an emerging biofluid poised for detection of clinical diseases. Although the rationale for oral diseases applications (e.g. oral cancer) is intuitive, the rationale and relationship between systemic diseases and saliva biomarkers are unclear.
In this study, we used mouse models of melanoma and non-small cell lung cancer and compared the transcriptome biomarker profiles of tumor-bearing mice to those of control mice. Microarray analysis showed that salivary transcriptomes were significantly altered in tumor-bearing mice vs. controls. Significant overlapping among transcriptomes of mouse tumors, serum, salivary glands and saliva suggests that salivary biomarkers have multiple origins. Furthermore, we identified that the expression of two groups of significantly altered transcription factors (TFs) Runx1, Mlxipl, Trim30 and Egr1, Tbx1, Nr1d1 in salivary gland tissue of melanoma-bearing mice can potentially be responsible for 82.6% of the up-regulated gene expression and 62.5% of the down-regulated gene expression, respectively, in the saliva of melanoma-bearing mice. We also showed that the ectopic production of nerve growth factor (NGF) in the melanoma tumor tissue as a tumor-released mediator can induce expression of the TF Egr-1 in the salivary gland.
Taken together, our data support the conclusion that upon systemic disease development, significant changes can occur in the salivary biomarker profile. Although the origins of the disease-induced salivary biomarkers may be both systemic and local, stimulation of salivary gland by mediators released from remote tumors plays an important role in regulating the salivary surrogate biomarker profiles.
PLoS ONE 02/2009; 4(6):e5875. · 4.09 Impact Factor
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ABSTRACT: Lectin microarray is a new technology that utilizes a panel of lectins immobilized on well-defined substrate for high-throughout analysis of glycans and glycoproteins. In this article, we have reviewed the fabrication and detection schemes in lectin microarray and discussed its novel applications in glycomics. We have also demonstrated a lectin array on PDMS with MALDI-TOF-MS for glycoprotein analysis. This method has been demonstrated for differential analysis of serum glycoproteins in oral cancer and healthy control subjects.
PROTEOMICS - CLINICAL APPLICATIONS 02/2009; 3(2):148-154. · 1.81 Impact Factor
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ABSTRACT: Saliva samples from certain disease patients harbor a wide spectrum of proteins, mRNAs, DNAs and other molecules that may be associated with the disease phenotype. If successfully discovered and validated, these informative molecules may serve as biomarkers, leading to the use of non-invasive biofluid for detecting and monitoring the diseases. This article summarizes the current advances in searching for potential biomarkers in saliva for human cancers, especially head and neck/oral cancers. With the new molecular profiling technologies such as microarray and proteomics, we are expecting to reveal highly discriminatory genomic and proteomic targets that can best detect the disease status.
Frontiers in bioscience (Scholar edition) 02/2009; 1:296-303.
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ABSTRACT: Lectin microarray is a new technology that utilizes a panel of lectins immobilized on well-defined substrate for high-throughout analysis of glycans and glycoproteins. In this article, we have reviewed the fabrication and detection schemes in lectin microarray and discussed its novel applications in glycomics. We have also demonstrated a lectin array on PDMS with MALDI-TOF-MS for glycoprotein analysis. This method has been demonstrated for differential analysis of serum glycoproteins in oral cancer and healthy control subjects.
PROTEOMICS - CLINICAL APPLICATIONS 01/2009; 3(2):148 - 154. · 1.81 Impact Factor
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Shen Hu,
Michael Zhou,
Jiang Jiang,
Jianghua Wang,
David Elashoff,
Sven Gorr,
Sara A Michie,
Fred K L Spijkervet,
Hendrika Bootsma,
Cees G M Kallenberg,
Arjan Vissink,
Steve Horvath,
David T Wong
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ABSTRACT: To identify key target genes and activated signaling pathways associated with the pathogenesis of Sjögren's syndrome (SS) by conducting a systems analysis of parotid glands manifesting primary SS or primary SS/mucosa-associated lymphoid tissue (MALT) lymphoma phenotypes.
A systems biology approach was used to analyze parotid gland tissue samples obtained from patients with primary SS, patients with primary SS/MALT lymphoma, and subjects without primary SS (non-primary SS controls). The tissue samples were assessed concurrently by gene-expression microarray profiling and proteomics analysis, followed by weighted gene-coexpression network analysis.
Gene-coexpression modules related to primary SS and primary SS/MALT lymphoma were significantly enriched with genes known to be involved in the immune/defense response, apoptosis, cell signaling, gene regulation, and oxidative stress. Detailed functional pathway analyses indicated that primary SS-associated modules were enriched with genes involved in proteasome degradation, apoptosis, signal peptides of the class I major histocompatibility complex (MHC), complement activation, cell growth and death, and integrin-mediated cell adhesion, while primary SS/MALT lymphoma-associated modules were enriched with genes involved in translation, ribosome biogenesis and assembly, proteasome degradation, class I MHC signal peptides, the G13 signaling pathway, complement activation, and integrin-mediated cell adhesion. Combined analyses of gene expression and proteomics data implicated 6 highly connected "hub" genes for distinguishing primary SS from non-primary SS, and 8 hub genes for distinguishing primary SS/MALT lymphoma from primary SS.
Systems biology analyses of the parotid glands from patients with primary SS and those with primary SS/MALT lymphoma revealed pathways and molecular targets associated with disease pathogenesis. The identified gene modules/pathways provide further insights into the molecular mechanisms of primary SS and primary SS/MALT lymphoma. The identified disease-hub genes represent promising targets for therapeutic intervention, diagnosis, and prognosis.
Arthritis & Rheumatism 01/2009; 60(1):81-92. · 7.87 Impact Factor
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Weihong Yan,
Rolf Apweiler,
Brian M Balgley,
Pinmanee Boontheung,
Jonathan L Bundy,
Benjamin J Cargile,
Steve Cole,
Xueping Fang,
Mireya Gonzalez-Begne,
Timothy J Griffin, [......],
David States,
James L Stephenson,
Shawn Than,
John R Yates,
Weixia Yu,
Hongwei Xie,
Yongming Xie,
Gilbert S Omenn,
Joseph A Loo,
David T Wong
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ABSTRACT: The proteome of human salivary fluid has the potential to open new doors for disease biomarker discovery. A recent study to comprehensively identify and catalog the human ductal salivary proteome led to the compilation of 1166 proteins. The protein complexity of both saliva and plasma is large, suggesting that a comparison of these two proteomes will provide valuable insight into their physiological significance and an understanding of the unique and overlapping disease diagnostic potential that each fluid provides. To create a more comprehensive catalog of human salivary proteins, we have first compiled an extensive list of proteins from whole saliva (WS) identified through MS experiments. The WS list is thereafter combined with the proteins identified from the ductal parotid, and submandibular and sublingual (parotid/SMSL) salivas. In parallel, a core dataset of the human plasma proteome with 3020 protein identifications was recently released. A total of 1939 nonredundant salivary proteins were compiled from a total of 19 474 unique peptide sequences identified from whole and ductal salivas; 740 out of the total 1939 salivary proteins were identified in both whole and ductal saliva. A total of 597 of the salivary proteins have been observed in plasma. Gene ontology (GO) analysis showed similarities in the distributions of the saliva and plasma proteomes with regard to cellular localization, biological processes, and molecular function, but revealed differences which may be related to the different physiological functions of saliva and plasma. The comprehensive catalog of the salivary proteome and its comparison to the plasma proteome provides insights useful for future study, such as exploration of potential biomarkers for disease diagnostics.
PROTEOMICS - CLINICAL APPLICATIONS 01/2009; 3(1):116-134. · 1.81 Impact Factor
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Weihong Yan,
Rolf Apweiler,
Brian M. Balgley,
Pinmanee Boontheung,
Jonathan L. Bundy,
Benjamin J. Cargile,
Steve Cole,
Xueping Fang,
Mireya Gonzalez-Begne,
Timothy J. Griffin, [......],
David States,
James L. Stephenson Jr,
Shawn Than,
John R. Yates III,
Weixia Yu,
Hongwei Xie,
Yongming Xie,
Gilbert S. Omenn,
Joseph A. Loo Dr,
David T. Wong Dr
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ABSTRACT: The proteome of human salivary fluid has the potential to open new doors for disease biomarker discovery. A recent study to comprehensively identify and catalog the human ductal salivary proteome led to the compilation of 1166 proteins. The protein complexity of both saliva and plasma is large, suggesting that a comparison of these two proteomes will provide valuable insight into their physiological significance and an understanding of the unique and overlapping disease diagnostic potential that each fluid provides. To create a more comprehensive catalog of human salivary proteins, we have first compiled an extensive list of proteins from whole saliva (WS) identified through MS experiments. The WS list is thereafter combined with the proteins identified from the ductal parotid, and submandibular and sublingual (parotid/SMSL) salivas. In parallel, a core dataset of the human plasma proteome with 3020 protein identifications was recently released. A total of 1939 nonredundant salivary proteins were compiled from a total of 19 474 unique peptide sequences identified from whole and ductal salivas; 740 out of the total 1939 salivary proteins were identified in both whole and ductal saliva. A total of 597 of the salivary proteins have been observed in plasma. Gene ontology (GO) analysis showed similarities in the distributions of the saliva and plasma proteomes with regard to cellular localization, biological processes, and molecular function, but revealed differences which may be related to the different physiological functions of saliva and plasma. The comprehensive catalog of the salivary proteome and its comparison to the plasma proteome provides insights useful for future study, such as exploration of potential biomarkers for disease diagnostics.
PROTEOMICS - CLINICAL APPLICATIONS 12/2008; 3(1):116 - 134. · 1.81 Impact Factor
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ABSTRACT: Saliva is a biofluid that can be obtained from individuals without supervision by health care providers. To maximize this clinical advantage, it is highly desirable to have a global salivary analyte stabilizer for proteins, RNA and DNA at ambient temperature.
Whole saliva, saliva supernatant and saliva filtrate (5.0 microm) were treated with RPS at room temperature (RT) for up to 6 days and then subjected to SDS-PAGE. Immunoblotting of beta-actin and cystatin C were used to evaluate protein stability. For salivary DNA/RNA, whole saliva was incubated with RPS at RT for up to 10 weeks. After extracting total DNA/RNA in samples at week 0, 2, 6 and 10, DNA stability was assayed by chromosome 18 DNA qPCR and RNA stability by beta-actin mRNA RT-qPCR.
beta-actin completely degraded in all types of saliva samples after 6-day incubation at RT. However, 24.0%, 91.4% and 89.3% of beta-actin remained intact with RPS for whole saliva, saliva supernatant and filtrate, respectively. Similarly, 70.3% of cystatin C in supernatant remained intact in the presence of RPS. For salivary DNA/RNA, the cycle threshold (Ct) values showed no significant change for chromosome 18 DNA and beta-actin mRNA in RPS-incubated saliva during the 10-week time course while significant increase in Ct values were observed in controls without RPS for both beta-actin mRNA and DNA.
RPS provided effective concurrent stabilization to salivary DNA/RNA in whole saliva for up to 10 weeks and proteins in saliva filtrate for 6 days at RT. We also achieved separation of saliva supernatant from cellular elements by a simple filtration step (bypassing the need for centrifugation).
Archives of oral biology 12/2008; 54(3):268-73. · 1.65 Impact Factor
