ORCID Profile
0000-0001-9039-8259
Current Organisations
Queensland University of Technology
,
Griffith University Griffith Sciences
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Analytical Chemistry | Electrochemistry | Sensor Technology (Chemical aspects) | Electroanalytical Chemistry
Expanding Knowledge in the Chemical Sciences | Scientific Instruments |
Publisher: Informa UK Limited
Date: 21-05-2013
DOI: 10.3109/07388551.2012.687361
Abstract: Human saliva harbours proteins of clinical relevance and about 30% of blood proteins are also present in saliva. This highlights that saliva can be used for clinical applications just as urine or blood. However, the translation of salivary biomarker discoveries into clinical settings is h ered by the dynamics and complexity of the salivary proteome. This review focuses on the current status of technological developments and achievements relating to approaches for unravelling the human salivary proteome. We discuss the dynamics of the salivary proteome, as well as the importance of s le preparation and processing techniques and their influence on downstream protein applications post-translational modifications of salivary proteome and protein: protein interactions. In addition, we describe possible enrichment strategies for discerning post-translational modifications of salivary proteins, the potential utility of selected-reaction-monitoring techniques for biomarker discovery and validation, limitations to proteomics and the biomarker challenge and future perspectives. In summary, we provide recommendations for practical saliva s ling, processing and storage conditions to increase the quality of future studies in an emerging field of saliva clinical proteomics. We propose that the advent of technologies allowing sensitive and high throughput proteome-wide analyses, coupled to well-controlled study design, will allow saliva to enter clinical practice as an alternative to blood-based methods due to its simplistic nature of s ling, non-invasiveness, easy of collection and multiple collections by untrained professionals and cost-effective advantages.
Publisher: Springer Science and Business Media LLC
Date: 05-2021
DOI: 10.1007/S11033-021-06398-7
Abstract: Oral cavity cancer (OCC) is the predominant subtype of head and neck cancer (HNC) and has up to 50% mortality. Genome-wide microRNA (miR) sequencing data indicates overexpression of miR-9-5p in HNC tumours, however, the biological role of miR-9-5p in OCC is complex it can either act as a tumour suppressor or an oncomir, regulating many target genes at the post-transcriptional level. We have investigated the overexpression of miR-9-5p in three OCC cell lines. We have evaluated its expression levels and Galectin-3 as potential biomarkers in saliva s les collected from controls and OCC patients. We found that over expression of miR-9-5p in OCC cell lines resulted in a significant reduction in cell proliferation and migration, and an increase in apoptosis, which was paralleled by an increase in Galectin-3 secretion and export of Galectin-3 protein. Our data are consistent with miR-9-5p being a modulator of Galectin-3 via the AKT/γ-catenin pathway. In addition, the positive correlation between the levels of miR-9-5p expression and secreted Galectin-3 in saliva reflects a similar relationship in vivo, and supports the utility of their integrative evaluation in OCC. Our findings indicate that both miR-9-5p and Galectin-3 are critical biomolecules in the progression of OCC.
Publisher: Frontiers Media SA
Date: 31-03-2020
Publisher: American Association for Cancer Research (AACR)
Date: 15-06-2020
DOI: 10.1158/1557-3265.AACRAHNS19-A03
Abstract: Metastasis in head and neck cancer (HNC) patients is reflected by measurable levels of circulating tumor cells (CTCs) in the blood of patients. CTCs represent cancer cells from the primary and metastatic sites, thereby providing a comprehensive representation of the tumor burden of an in idual patient. Recent advancements have shown that PD-1/PD-L1 immune checkpoint therapies have durable responses in HNC. Our study was designed to use multiple CTC enrichment platforms for the capture of CTCs and novel culture formulations for the ex vivo expansion of CTCs. n=300 head and neck cancer patients were recruited to investigate the prognostic role of CTCs. We evaluated multiple CTC isolation technologies (CellSearch, filtration, CD45 depletion, inertial microfluidics) using matched patient s les, which showed that epitope-independent CTC isolation captured a greater proportion of CTCs. Molecular alterations present in the primary tissue were confirmed in the CTCs by 3D-DNA FISH (EGFR- lification). The presence of CTC clusters was associated with the development of distant metastatic disease (P=0.0313). HNC CTC-positive patients had shorter progression-free survival (PFS) (Hazard ratio [HR]: 4.946 95% confidence interval [CI]:1.571-15.57 P=0.0063) and PD-L1-positive CTCs were found to be significantly associated with worse outcome ([HR]:5.159 95% [CI]:1.011-26.33 P=0.0485). In a proof-of-principle study, we were able to demonstrate for the first time short-term patient-derived CTC cultures outside the patient’s body from 7/18 HNC s les (4/7 HPV-positive). Recently, we have preliminary data that suggest that PD-L1 is frequently expressed on CTCs in HNC and an immunoscore may be able to identify patients likely to benefit from immunotherapy—a current unmet clinical need. Expanding CTCs outside the patient’s body allows for the recapitulation of the molecular ersity present within the tumor, understanding of the disease progression, and testing of therapies. Citation Format: Arutha Kulasinghe, Joanna Kapeleris, Liz Kenny, Majid Warkiani, Ian Vela, Jean Paul Thiery, Ken O’Byrne, Chamindie Punyadeera. Isolation, characterization, and expansion of circulating tumor cells in head and neck cancers [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research 2019 Apr 29-30 Austin, TX. Philadelphia (PA): AACR Clin Cancer Res 2020 (12_Suppl_2):Abstract nr A03.
Publisher: Springer Science and Business Media LLC
Date: 23-06-2022
DOI: 10.1007/S13402-022-00681-W
Abstract: Local recurrence and metastasis remain the major causes of death in head and neck cancer (HNC) patients. Circulating tumour cells (CTCs) are shed from primary and metastatic sites into the circulation system and have been reported to play critical roles in the metastasis and recurrence of HNC. Here, we explored the use of CTCs to predict the response to treatment and disease progression in HNC patients. Blood s les were collected at diagnosis from HNC patients (n = 119). CTCs were isolated using a spiral microfluidic device and were identified using immunofluorescence staining. Correlation of baseline CTC numbers to 13-week PET-CT data and multidisciplinary team consensus data were conducted. CTCs were detected in 60/119 (50.4%) of treatment naïve HNC patients at diagnosis. Baseline CTC numbers were higher in stage III vs. stage I-II p16-positive oropharyngeal cancers (OPCs) and other HNCs ( p = 0.0143 and 0.032, respectively). In addition, we found that baseline CTC numbers may serve as independent predictors of treatment response, even after adjusting for other conventional prognostic factors. CTCs were detected in 10 out of 11 patients exhibiting incomplete treatment responses. We found that baseline CTC numbers are correlated with treatment response in patients with HNC. The expression level of cell-surface vimentin (CSV) on CTCs was significantly higher in patients with persistent or progressive disease, thus providing additional prognostic information for stratifying the risk at diagnosis in HNC patients. The ability to detect CTCs at diagnosis allows more accurate risk stratification, which in the future may be translated into better patient selection for treatment intensification and/or de-intensification strategies.
Publisher: Elsevier BV
Date: 07-2022
Publisher: Impact Journals, LLC
Date: 09-08-2016
Publisher: Future Medicine Ltd
Date: 03-2016
DOI: 10.2217/BMM.16.2
Abstract: The early detection of head and neck squamous cell carcinoma (HNSCC) continues to be a challenge to the clinician. Saliva as a diagnostic medium carries significant advantages including its close proximity to the region of interest, ease of collection and noninvasive nature. While the identification of biomarkers continues to carry significant diagnostic and prognostic utility in HNSCC, epigenetic alterations present a novel opportunity to serve this purpose. With the developments of novel and innovative technologies, epigenetic alterations are now emerging as attractive candidates in HNSCC. As such, this review will focus on two commonly aberrant epigenetic alterations: DNA methylation and microRNA expression in HNSCC and their potential clinical utility. Identification and validation of these salivary epigenetic biomarkers would not only enable early diagnosis but will also facilitate in the clinical management.
Publisher: Springer New York
Date: 2019
DOI: 10.1007/978-1-4939-9769-5_6
Abstract: Oral premalignant disorders (OPMD) have relatively high malignant transformation rates to Oral Cancers (OC). Oral carcinogenesis is a multistep process that originates as epithelial hyperplasia followed by epithelial dysplasia, leading to fully malignant phenotypes. Early detection can be lifesaving but is currently not possible due to the lack of early diagnostic tools. The current diagnostic methods such as biopsy s ling, tumor tissue staining, and imaging techniques require skilled personnel and are invasive, painful, time-consuming, and expensive. Saliva has gained momentum as the diagnostic fluid of the future due to its noninvasive nature, ease of s ling, multiple s les can be collected with ease and more importantly does not require skilled personnel. The use of saliva in cancer diagnostics is an emerging and an expanding field. MicroRNA (miRNA) play a role in cancer initiation and progression and the expression changes of miRNA have been investigated as a potential biomarker in cancer studies. In this chapter, we describe a robust and cost-effective protocol to isolate and enrich miRNA from saliva s les. Profiling miRNAs in saliva s les can form part of the clinical management of OPMD and OC patients in the future.
Publisher: Frontiers Media SA
Date: 19-01-2021
Abstract: Immune checkpoint inhibitors (ICI) have shown durable and long-term benefits in a subset of head and neck squamous cell carcinoma (HNSCC) patients. To identify patient-responders from non-responders, biomarkers are needed which are predictive of outcome to ICI therapy. Cues in the tumor microenvironment (TME) have been informative in understanding the tumor-immune contexture. In this preliminary study, the NanoString GeoMx™ Digital Spatial Profiling (DSP) technology was used to determine the immune marker and compartment specific measurements in a cohort of HNSCC tumors from patients receiving ICI therapy. Our data revealed that markers involved with immune cell infiltration (CD8 T-cells) were not predictive of outcome to ICI therapy. Rather, a number of immune cell types and protein markers (CD4, CD68, CD45, CD44, CD66b) were found to correlate with progressive disease. Cross platform comparison with the Opal Vectra (Perkin Elmer) for a number of markers across similar regions of interest demonstrated concordance for pan-cytokeratin, CD8, and PD-L1. This study, to our knowledge, represents the first digital spatial analysis of HNSCC tumors. A larger cohort of HNSCC will be required to orthogonally validate the findings.
Publisher: Wiley
Date: 07-08-2020
DOI: 10.1111/CAS.14585
Publisher: Elsevier BV
Date: 09-2022
Publisher: Springer New York
Date: 2019
DOI: 10.1007/978-1-4939-9769-5_8
Abstract: Metastasis is responsible for 90% of cancer-related deaths. The study of circulating tumor cells (CTCs) enables the study of the units of disease responsible for the process of metastasis. While the biology of the primary tissue is relatively known, little is understood about the cells en route to distant sites. Here we describe the isolation of CTCs using the spiral microfluidic technology for the efficient sorting of CTCs from head and neck cancer (HNC) patient blood s les. Furthermore, the molecular characterization of CTCs can aid in stratifying patients for targeted therapy such as immunotherapy, which is having a profound impact in the treatment of metastatic HNC.
Publisher: Future Science Ltd
Date: 09-2013
DOI: 10.4155/BIO.13.190
Abstract: Background: Plasma D-dimer tests are currently used to exclude deep vein thrombosis and pulmonary embolism. Human saliva has numerous advantages over blood as a diagnostic s le. The aims of our study were to develop a reliable immunoassay to detect D-dimer levels in saliva, and to determine the correlation between salivary and blood D-dimer levels. Results/methodology: Saliva and blood s les were collected from 40 healthy volunteers. We developed a AlphaLISA ® immunoassay with acceptable analytical performances to quantify D-dimer levels in the s les. The median salivary D-dimer levels were 138.1 ng/ml (morning) and 140.7 ng/ml (afternoon), and the plasma levels were 75.0 ng/ml. Salivary D-dimer levels did not correlate with plasma levels (p = 0.61). Conclusion: For the first time, we have quantified D-dimer levels and found twofold increase in saliva (p 0.05) than in plasma. Further studies are required to demonstrate the clinical relevance/utility of salivary D-dimer in patients with confirmed deep vein thrombosis and/or pulmonary embolism.
Publisher: Springer Science and Business Media LLC
Date: 17-08-2017
DOI: 10.1038/S41598-017-07885-3
Abstract: Saliva has attracted attention as a diagnostic fluid due to the association of oral microbiota with systemic diseases. However, the lack of standardised methods for saliva collection has led to the slow uptake of saliva in microbiome research. The aim of this study was to systematically evaluate the potential effects on salivary microbiome profiles using different methods of saliva collection, storage and gDNA extraction. Three types of saliva fractions were collected from healthy in iduals with or without the gDNA stabilising buffer. Subsequently, three types of gDNA extraction methods were evaluated to determine the gDNA extraction efficiencies from saliva s les. The purity of total bacterial gDNA was evaluated using the ratio of human β-globin to bacterial 16S rRNA PCR while 16S rRNA gene licon sequencing was carried out to identify the bacterial profiles present in these s les. The quantity and quality of extracted gDNA were similar among all three gDNA extraction methods and there were no statistically significant differences in the bacterial profiles among different saliva fractions at the genus-level of taxonomic classification. In conclusion, saliva s ling, processing and gDNA preparation do not have major influence on microbiome profiles.
Publisher: Wiley
Date: 15-01-2022
Publisher: Springer Science and Business Media LLC
Date: 08-10-2014
DOI: 10.1007/S10529-013-1341-0
Abstract: The measurements of plasma natriuretic peptides (NT-proBNP, proBNP and BNP) are used to diagnose heart failure but these are expensive to produce. We describe a rapid, cheap and facile production of proteins for immunoassays of heart failure. DNA encoding N-terminally His-tagged NT-proBNP and proBNP were cloned into the pJexpress404 vector. ProBNP and NT-proBNP peptides were expressed in Escherichia coli, purified and refolded in vitro. The analytical performance of these peptides were comparable with commercial analytes (NT-proBNP EC50 for the recombinant is 2.6 ng/ml and for the commercial material is 5.3 ng/ml) and the EC50 for recombinant and commercial proBNP, are 3.6 and 5.7 ng/ml respectively). Total yield of purified refolded NT-proBNP peptide was 1.75 mg/l and proBNP was 0.088 mg/l. This approach may also be useful in expressing other protein analytes for immunoassay applications. To develop a cost effective protein expression method in E. coli to obtain high yields of NT-proBNP (1.75 mg/l) and proBNP (0.088 mg/l) peptides for immunoassay use.
Publisher: Impact Journals, LLC
Date: 16-09-2016
Publisher: Informa UK Limited
Date: 27-01-2020
Publisher: Wiley
Date: 06-06-2023
DOI: 10.1002/CAM4.6185
Abstract: Despite the rising incidence, particularly of the human papillomavirus (HPV)‐associated fraction of oropharyngeal cancer (OPC), there are no early detection methods for OPC. Considering the close association between saliva and head and neck cancers, this study was designed to investigate salivary micro RNA (miRNAs) associated with OPC, especially focusing on HPV‐positive OPC. Saliva was collected from OPC patients at diagnosis and patients were clinically followed up ≤5 years. Salivary small RNA isolated from HPV‐positive OPC patients ( N = 6), and HPV‐positive ( N = 4) and negative controls ( N = 6) were analysed by next‐generation sequencing to identify dysregulated miRNAs. Discovered miRNAs were validated by quantitative PCR using two different assays in a separate cohort of patients (OPC = 91, controls = 92). The relative expression was calculated considering SNORD‐96A as the normalizer. Candidate miRNAs with diagnostic and prognostic potential were evaluated by generalized logistic regression. A panel consisting of nine miRNAs was identified to have the best diagnostic performance to discriminate HPV‐positive OPC from HPV‐positive controls (AUC‐ validation‐1 = 94.8%, validation‐2 = 98%). Further, a panel consisting of six miRNAs were identified to discriminate OPC from controls regardless of the HPV status (AUC‐ validation‐1 = 77.2%, validation‐2 = 86.7%). In addition, the downregulation of hsa‐miR‐7‐5p was significantly associated with poor overall survival of OPC patients (HR = 0.638). A panel consisting of nine miRNAs were identified for the prediction of the overall survival of the OPC patients (log‐rank test‐ p = 0.0008). This study highlights that salivary miRNAs can play an essential role in the detection and prognostication of OPC.
Publisher: Elsevier BV
Date: 12-2020
Publisher: Elsevier BV
Date: 09-2022
DOI: 10.1016/J.BBCAN.2022.188784
Abstract: Head and neck squamous cell carcinomas (HNSCCs) are aggressive and clinically challenging tumours that require a multidisciplinary management approach. Despite significant therapy improvements, HNSCC patients have a poor prognosis with a 5-year survival rate of about 65%. As recently recognised key players in cancer, exosomes are extracellular vesicles (EVs) with a diameter of nearly 50-120 nm which transport information from one cell to another. Exosomes are actively involved in various aspects of tumour initiation, development, metastasis, immune regulation, therapy resistance, and therapeutic applications. However, current knowledge of the role of exosomes in the pathophysiological processes of HNSCC is still in its infancy, and additional studies are needed. In this review, we summarise and discuss the relevance of exosomes in mediating local immunosuppression and therapy resistance of HNSCC. We also review the most recent studies that have explored the therapeutic potential of exosomes as cancer vaccines, drug carriers or tools to reverse the drug resistance of HNSCC.
Publisher: Sciedu Press
Date: 14-12-2015
DOI: 10.5430/JST.V6N1P28
Publisher: Wiley
Date: 08-2023
DOI: 10.1002/CTM2.1341
Publisher: AME Publishing Company
Date: 10-2020
DOI: 10.21037/TLCR-20-521
Publisher: Elsevier BV
Date: 10-2023
Publisher: Springer Science and Business Media LLC
Date: 15-01-2018
DOI: 10.1038/S41598-017-19117-9
Abstract: Distant metastasis (DM) from head and neck cancers (HNC) portends a poor patient prognosis. Despite its important biological role, little is known about the cells which seed these DM. Circulating tumour cells (CTCs) represent a transient cancer cell population, which circulate in HNC patients’ peripheral blood and seed at distant sites. Capture and analysis of CTCs offers insights into tumour metastasis and can facilitate treatment strategies. Whilst the data on singular CTCs have shown clinical significance, the role of CTC clusters in metastasis remains limited. In this pilot study, we assessed 60 treatment naïve HNC patients for CTCs with disease ranging from early to advanced stages, for CTC clusters utilizing spiral CTC enrichment technology. Single CTCs were isolated in 18/60–30% (Ranging from Stage I-IV), CTC clusters in 15/60–25% (exclusively Stage IV) with 3/15–20% of CTC clusters also containing leukocytes. The presence of CTC clusters associated with the development of distant metastatic disease( P = 0.0313). This study demonstrates that CTC clusters are found in locally advanced patients, and this may be an important prognostic marker. In vivo and in vitro studies are warranted to determine the role of these CTC clusters, in particular, whether leukocyte involvement in CTC clusters has clinical relevance.
Publisher: Elsevier BV
Date: 2022
DOI: 10.2139/SSRN.4057071
Publisher: Public Library of Science (PLoS)
Date: 09-09-2016
Publisher: Wiley
Date: 02-2015
DOI: 10.1002/CAM4.424
Publisher: Elsevier BV
Date: 2020
DOI: 10.1016/J.JPROT.2019.103532
Abstract: Saliva has become one of the more attractive body fluids for protein biomarker discovery studies because of its ease of collection, non-invasiveness and multiple s les can be collected from an in idual at a single time point. With the development of modern data acquisition strategies, such as Sequential Window Acquisition of all Theoretical Mass Spectrometry (SWATH-MS), the quality of saliva s le preparation has become a crucial factor for a successful protein identification and quantification. Several s le preparation methods have been proposed, but there has been no systematic evaluation conducted to date that compared each of these methods. We have therefore, performed an extensive assessment using technical and biological repeats to evaluate the number of protein IDs and repeatability of three most commonly used techniques, in-solution digestion, filter-aided s le preparation (FASP) and in stage-tip (iST) digestion. We discovered that in the case of human saliva s le, FASP provided the highest number of proteins (human and microbial) identifiable from a pool saliva s le, and there were no significant differences in terms of repeatability among the three methods investigated.
Publisher: Elsevier BV
Date: 07-2012
DOI: 10.1016/J.CCA.2012.02.020
Abstract: Human saliva mirrors the body's health and can be collected non-invasively, does not require specialized skills and is suitable for large population based screening programs. The aims were twofold: to evaluate the suitability of commercially available saliva collection devices for quantifying proteins present in saliva and to provide levels for C-reactive protein (CRP), myoglobin, and immunoglobin E (IgE) in saliva of healthy in iduals as a baseline for future studies. Saliva was collected from healthy volunteers (n=17, ages 18-33years). The following collection methods were evaluated: drool Salimetrics® Oral Swab (SOS) Salivette® Cotton and Synthetic (Sarstedt) and Greiner Bio-One Saliva Collection System (GBO SCS®). We used AlphaLISA® assays to measure CRP, IgE and myoglobin levels in human saliva. Significant (p<0.05) differences in the salivary flow rates were observed based on the method of collection, i.e. salivary flow rates were significantly lower (p<0.05) in unstimulated saliva (i.e. drool and SOS), when compared with mechanically stimulated methods (p<0.05) (Salivette® Cotton and Synthetic) and acid stimulated method (p<0.05) (SCS®). Saliva collected using SOS yielded significantly (p<0.05) lower concentrations of myoglobin and CRP, whilst, saliva collected using the Salivette® Cotton and Synthetic swab yielded significantly (p<0.05) lower myoglobin and IgE concentrations respectively. The results demonstrated significantly relevant differences in analyte levels based on the collection method. Significant differences in the salivary flow rates were also observed depending on the saliva collection method. The data provide preliminary baseline values for salivary CRP, myoglobin, and IgE levels in healthy participants and based on the collection method.
Publisher: Springer Science and Business Media LLC
Date: 21-12-2016
DOI: 10.1007/S11126-015-9415-X
Abstract: Sleep dysfunction is a pervasive issue in schizophrenia and psychosis. Current knowledge is drawn almost exclusively from studies using quantitative research methodologies that include measures and tools developed in healthy population groups. Qualitative studies investigating the first-person perspectives of sleep problems are therefore important for designing better assessment and treatment tools to meet consumer needs. Focus groups were conducted to elicit detailed information regarding the personal experience of sleep problems, their antecedents and impact, in 14 in iduals with schizophrenia-spectrum disorder who experienced insomnia during their illness. Thematic analysis was applied to examine the data and draw treatment implications for sleep management. Insomnia was ubiquitous and frequently co-occurred with other sleep difficulties (nightmares, sleep walking, acting out dreams, etc.) in this group. Discussions revealed themes common across insomnia populations (role of negative mood states and cognitive intrusions) and also new themes on factors contributing to sleep problems in schizophrenia: (1) beliefs that sleep problems cannot be changed (2) trauma and adversity (3) lifestyle choices and lack of motivation and (4) medication side effects. Sleep problems also had profound impact on daytime dysfunctions and disability. The findings point to novel issues that may benefit from consideration in the treatment of sleep problems in schizophrenia. Unhelpful cognitions and behaviours about sleep can be addressed with psychological interventions, activity scheduling and motivational interviewing techniques. Seeking a first-person perspective is vital for identifying issues that will impact on treatment success and recovery.
Publisher: Springer Science and Business Media LLC
Date: 22-01-2013
DOI: 10.1007/S13402-012-0122-4
Abstract: Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cause of cancer mortality in the world and the 5th most commonly occurring cancer. Tobacco smoking, alcohol consumption and human papilloma virus (HPV) infections have been associated with the occurrence of HNSCC. Despite advances that have been made in HNSCC treatment, smoking-associated HNSCC patients still exhibit a poor 5 year survival rate (30-50 %) and a concomitant poor quality of life. The major clinical challenge to date lies in the early detection of dysplastic lesions,which can progress to malignancy. In addition, there are currently no tools available to monitor HNSCC patients for early stages of local recurrences or distant metastases. In the recent past, micro-RNAs (miRNA) have been assessed for their role in cancer initiation and progression, including HNSCC. It is now well-established that deregulation of these single stranded, small non-coding, 19-25 nt RNAs can e.g. enhance the expression of oncogenes or subdue the expression of tumor suppressor genes. The aims of this review are three-fold: first to retrieve from the literature miRNAs that have specifically been associated with HNSCC, second to group these miRNAs into those regulating tumor initiation, progression and metastasis, and third to discern miRNAs related to smoking-associated HNSCC versus HPV-associated HNSCC development. This review gives an overview on the miRNAs regulating the development of head and neck cancers. The ultimate establishment of miRNA expression profiles that are HNSCC specific, and miRNAs that orchestrate altered gene and protein expression levels in HNSCC, could pave the way for a better understanding of the mechanism underlying its pathogenesis and the development of novel, targeted therapies.
Publisher: Impact Journals, LLC
Date: 31-07-2017
Publisher: MDPI AG
Date: 29-11-2018
DOI: 10.3390/DIAGNOSTICS8040079
Abstract: Background: PIK3CA pathways are the most frequently mutated oncogenic pathway in head and neck squamous cell carcinoma (HNSCC), including virally driven HNCs. PIK3CA is involved in the PI3K-PTEN-mTOR signalling pathway. PIK3CA has been implicated in HNSCC progression and PIK3CA mutations may serve as predictive biomarkers for therapy selection. Circulating tumour DNA (ctDNA) derived from necrotic and apoptotic tumour cells are thought to harbour tumour-specific genetic alterations. As such, the detection of PIK3CA alterations detected by ctDNA holds promise as a potential biomarker in HNSCC. Methods: Blood s les from treatment naïve HNSCC patients (n = 29) were interrogated for a commonly mutated PIK3CA hotspot mutation using low cost allele-specific Plex-PCRTM technology. Results: In this pilot, cross sectional study, PIK3CA E545K mutation was detected in the plasma s les of 9/29 HNSCC patients using the Plex-PCRTM technology. Conclusion: The results of this pilot study support the notion of using allele-specific technologies for cost-effective testing of ctDNA, and further assert the potential utility of ctDNA in HNSCC.
Publisher: Elsevier BV
Date: 10-2021
Publisher: Frontiers Media SA
Date: 14-08-2018
Publisher: Oxford University Press (OUP)
Date: 10-2013
DOI: 10.1373/CLINCHEM.2012.200204
Abstract: The use of nonstandardized N-terminal pro–B-type natriuretic peptide (NT-proBNP) assays can contribute to the misdiagnosis of heart failure (HF). Moreover, there is yet to be established a common consensus regarding the circulating forms of NT-proBNP being used in current assays. We aimed to characterize and quantify the various forms of NT-proBNP in the circulation of HF patients. Plasma s les were collected from HF patients (n = 20) at rest and stored at −80 °C. NT-proBNP was enriched from HF patient plasma by use of immunoprecipitation followed by mass spectrometric analysis. Customized homogeneous sandwich AlphaLISA® immunoassays were developed and validated to quantify 6 fragments of NT-proBNP. Mass spectrometry identified the presence of several N- and C-terminally processed forms of circulating NT-proBNP, with physiological proteolysis between Pro2-Leu3, Leu3-Gly4, Pro6-Gly7, and Pro75-Arg76. Consistent with this result, AlphaLISA immunoassays demonstrated that antibodies targeting the extreme N or C termini measured a low apparent concentration of circulating NT-proBNP. The apparent circulating NT-proBNP concentration was increased with antibodies targeting nonglycosylated and nonterminal epitopes (P & 0.05). In plasma collected from HF patients, immunoreactive NT-proBNP was present as multiple N- and C-terminally truncated fragments of the full length NT-proBNP molecule. Immunodetection of NT-proBNP was significantly improved with the use of antibodies that did not target these terminal regions. These findings support the development of a next generation NT-proBNP assay targeting nonterminal epitopes as well as avoiding the central glycosylated region of this molecule.
Publisher: Wiley
Date: 12-2002
DOI: 10.1038/OBY.2002.165
Abstract: Lower lipid and insulin levels are found during a glucose-tolerance test in obese black than obese white South African women. Therefore, beta-cell function and lipid metabolism were compared in these populations during a mixed meal. Blood concentrations of glucose, free fatty acids (FFAs), insulin, lipograms, and in vivo FFA oxidation were determined at fasting and for 7 hours after oral administration of a mixed emulsion containing glucose-casein-sucrose-lipid and [1-(13)C] palmitic acid in 8 lean black women (LBW), 10 obese black women (OBW), 9 lean white women (LWW), and 10 obese white women (OWW). Subcutaneous and visceral fat mass was assessed by computerized tomography. Visceral fat area was higher in OWW (152.7 +/- 17.0 cm(2)) than OBW (80.0 +/- 6.7 cm(2) p < 0.01). In OBW, 30-minute insulin levels were higher (604.3 +/- 117.6 pM) than OWW (311.0 +/- 42.9 pM p < 0.05). Total triglyceride was higher in OWW (706.7 +/- 96.0 mM x 7 hours) than OBW (465.7 +/- 48.2 mM x 7 hours p < 0.05) and correlated with visceral fat area (beta = 0.38, p = 0.05). Palmitate oxidation was higher in lean than obese women in both ethnic groups and correlated negatively with fat mass (beta = -0.58, p < 0.005). The higher 30-minute insulin response in OBW may reflect a higher insulinotropic effect of FFAs or glucose. The elevated triglyceride level of OWW may be due to their higher visceral fat mass and possibly reduced clearance by adipose tissue.
Publisher: Oxford University Press (OUP)
Date: 2004
Abstract: Menstrual effluent affects mesothelial cell (MC) morphology. We evaluated whether these changes were consistent with epithelial-mesenchymal transitions (EMT). Monolayer cultures of MC were incubated overnight in conditioned media, prepared from cells isolated form menstrual effluent, with or without kinase and ATP inhibitors. Changes in cell morphology were monitored using time-lapse video microscopy and immunohistochemistry. Effects on the expression of EMT-associated molecules were evaluated using real-time RT-PCR and/or Western blot analysis. Incubation in conditioned media disrupted cell-cell contacts, and increased MC motility. The changes were reversible. During the changes the distribution of cytokeratins, fibrillar actin and alpha-tubulin changed. Sodium azide, an inhibitor of ATP production, and Genistein, a general tyrosine kinase inhibitor, antagonized these effects. Wortmannin, a phosphatidylinositol 3-kinase inhibitor, and SU6656, an Src tyrosine kinase inhibitor, only partially antagonized the effect. The expression of Snail and vimentin was markedly up-regulated, whereas the expression of E-cadherin was decreased and cytokeratins were altered. In MC, menstrual effluent initiates a reversible, energy-dependent transition process from an epithelial to a mesenchymal phenotype. Involvement of the (Src) tyrosine kinase signalling pathway and the changes in the expression of cytokeratins, Snail, vimentin and E-cadherin demonstrate that the morphological changes are EMT.
Publisher: Elsevier
Date: 2019
Publisher: Wiley
Date: 12-08-2023
DOI: 10.1002/WRNA.1754
Abstract: Oral cancer (OC) is the most prevalent subtype of cancer arising in the head and neck region. OC risk is mainly attributed to behavioral risk factors such as exposure to tobacco and excessive alcohol consumption, and a lesser extent to viral infections such as human papillomaviruses and Epstein-Barr viruses. In addition to these acquired risk factors, heritable genetic factors have shown to be associated with OC risk. Despite the high incidence, biomarkers for OC diagnosis are lacking and consequently, patients are often diagnosed in advanced stages. This delay in diagnosis is reflected by poor overall outcomes of OC patients, where 5-year overall survival is around 50%. Among the biomarkers proposed for cancer detection, noncoding RNA (ncRNA) can be considered as one of the most promising categories of biomarkers due to their role in virtually all cellular processes. Similar to other cancer types, changes in expressions of ncRNAs have been reported in OC and a number of ncRNAs have diagnostic, prognostic, and therapeutic potential. Moreover, some ncRNAs are capable of regulating gene expression by various mechanisms. Therefore, elucidating the current literature on the four main types of ncRNAs namely, microRNA, lncRNA, snoRNA, piwi-RNA, and circular RNA in the context of OC pathogenesis is timely and would enable further improvements and innovations in diagnosis, prognosis, and treatment of OC. This article is categorized under: RNA in Disease and Development > RNA in Disease RNA in Disease and Development > RNA in Development.
Publisher: Oxford University Press (OUP)
Date: 05-2011
DOI: 10.1373/CLINCHEM.2010.153767
Abstract: Over the past 10 years, the use of saliva as a diagnostic fluid has gained attention and has become a translational research success story. Some of the current nanotechnologies have been demonstrated to have the analytical sensitivity required for the use of saliva as a diagnostic medium to detect and predict disease progression. However, these technologies have not yet been integrated into current clinical practice and work flow. As a diagnostic fluid, saliva offers advantages over serum because it can be collected noninvasively by in iduals with modest training, and it offers a cost-effective approach for the screening of large populations. Gland-specific saliva can also be used for diagnosis of pathology specific to one of the major salivary glands. There is minimal risk of contracting infections during saliva collection, and saliva can be used in clinically challenging situations, such as obtaining s les from children or handicapped or anxious patients, in whom blood s ling could be a difficult act to perform. In this review we highlight the production of and secretion of saliva, the salivary proteome, transportation of biomolecules from blood capillaries to salivary glands, and the diagnostic potential of saliva for use in detection of cardiovascular disease and oral and breast cancers. We also highlight the barriers to application of saliva testing and its advancement in clinical settings. Saliva has the potential to become a first-line diagnostic s le of choice owing to the advancements in detection technologies coupled with combinations of biomolecules with clinical relevance.
Publisher: Springer Science and Business Media LLC
Date: 03-03-2016
Publisher: MDPI AG
Date: 22-11-2019
DOI: 10.3390/BIOM9120766
Abstract: Screening for systolic heart failure (SHF) has been problematic. Heart failure management guidelines suggest screening for structural heart disease and SHF prevention strategies should be a top priority. We developed a multi-protein biomarker panel using saliva as a diagnostic medium to discriminate SHF patients and healthy controls. We collected saliva s les from healthy controls (n = 88) and from SHF patients (n = 100). We developed enzyme linked immunosorbent assays to quantify three specific proteins eptide (Kallikrein-1, Protein S100-A7, and Cathelicidin antimicrobial peptide) in saliva s les. The analytical and clinical performances and predictive value of the proteins were evaluated. The analytical performances of the immunoassays were all within acceptable analytical ranges. The multi-protein panel was able to significantly (p 0.001) discriminate saliva s les collected from patients with SHF from controls. The multi-protein panel demonstrated good performance with an overall diagnostic accuracy of 81.6% (sensitivity of 79.2% and specificity of 85.7%) when distinguishing SHF patients from healthy in iduals. In conclusion, we have developed immunoassays to measure the salivary concentrations of three proteins combined as a panel to accurately distinguish SHF patients from healthy controls. While this requires confirmation in larger cohorts, our findings suggest that this three-protein panel has the potential to be used as a biomarker for early detection of SHF.
Publisher: Informa UK Limited
Date: 17-02-2016
DOI: 10.1586/14737159.2016.1144474
Abstract: Heart failure (HF) affects approximately 23 million in iduals worldwide and this number is increasing, due to an aging and growing population. Early detection of HF is crucial in the management of this debilitating disease. Current diagnostic methods for HF rely heavily on clinical imaging techniques and blood analysis, which makes them less than ideal for population-based screening purposes. Studies focusing on developing novel biomarkers for HF have utilized various techniques and biological fluids, including urine and saliva. Promising results from these studies imply that these body fluids can be used in evaluating the clinical manifestation of HF and will one day be integrated into a clinical workflow and facilitate HF management.
Publisher: Oxford University Press (OUP)
Date: 07-2013
DOI: 10.1373/CLINCHEM.2012.197863
Abstract: The use of salivary diagnostics is increasing because of its noninvasiveness, ease of s ling, and the relatively low risk of contracting infectious organisms. Saliva has been used as a biological fluid to identify and validate RNA targets in head and neck cancer patients. The goal of this study was to develop a robust, easy, and cost-effective method for isolating high yields of total RNA from saliva for downstream expression studies. Oral whole saliva (200 μL) was collected from healthy controls (n = 6) and from patients with head and neck cancer (n = 8). The method developed in-house used QIAzol lysis reagent (Qiagen) to extract RNA from saliva (both cell-free supernatants and cell pellets), followed by isopropyl alcohol precipitation, cDNA synthesis, and real-time PCR analyses for the genes encoding β-actin (“housekeeping” gene) and histatin (a salivary gland–specific gene). The in-house QIAzol lysis reagent produced a high yield of total RNA (0.89–7.1 μg) from saliva (cell-free saliva and cell pellet) after DNase treatment. The ratio of the absorbance measured at 260 nm to that at 280 nm ranged from 1.6 to 1.9. The commercial kit produced a 10-fold lower RNA yield. Using our method with the QIAzol lysis reagent, we were also able to isolate RNA from archived saliva s les that had been stored without RNase inhibitors at −80 °C for & years. Our in-house QIAzol method is robust, is simple, provides RNA at high yields, and can be implemented to allow saliva transcriptomic studies to be translated into a clinical setting.
Publisher: Wiley
Date: 22-11-2018
DOI: 10.1002/CAM4.1832
Publisher: Elsevier
Date: 2013
Publisher: Informa UK Limited
Date: 15-09-2014
DOI: 10.1586/14737159.2014.960519
Abstract: Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer in the world with 600,000 new cases diagnosed annually. Tobacco and alcohol use have been associated as the principal etiological factors of this pathogenesis. The incidence of smoking-associated HNSCC has declined, while human papilloma virus (HPV)-associated HNSCC is on the rise. There are currently no clinically validated biomarkers to detect this cancer at an early stage (cancers independent of HPV status). It is well-established that the aberrant expression of miRNAs can lead to tumorigenesis. miRNA expression differences have also been demonstrated in HPV-positive and HPV-negative HNSCC tumor tissues as well as in body fluids. Therefore, miRNAs have the potential to provide an unprecedented insight into the pathogenesis of HNSCC and serve as potential biomarkers. This review addresses HNSCC disease burden and the regulation of miRNA by HPV viral oncoproteins, potential miRNA biomarkers and future perspectives. miRNA provides an unique opportunity to fulfill the current clinical challenge in HNSCC patient management by enabling early detection followed by targeted interventions, leading to a significant reduction in mortality and morbidity.
Publisher: Elsevier BV
Date: 03-2003
DOI: 10.1016/S0960-0760(03)00061-X
Abstract: Our understanding of the mechanisms of the actions of oestrogens and progestins have evolved from the simple concept of nuclear receptor-mediated regulation of transcription to a highly sophisticated, finely tuned interplay between various coregulators, other signaling cascades and transcription factors. The net result of these complex regulatory mechanisms is a steroid-, cell-, or tissue-specific action of oestrogens and progestins, their antagonists or selective modulators of their receptors. In this review, we have attempted to shed some light on the regulation of the actions of oestrogens and progestins on the human endometrium.
Publisher: Springer Science and Business Media LLC
Date: 09-08-2023
Publisher: MDPI AG
Date: 03-04-2019
Abstract: The incidence of human papillomavirus (HPV)-positive oropharyngeal cancer (OPC) is rising in high-income countries, including Australia. Increasing evidence suggests that accurate HPV testing is pivotal for clinical decision making and treatment planning in these patients. Recently, the eighth edition of the American Joint Committee on Cancer/Union for International Cancer Control (AJCC/UICC) tumor–node–metastasis (TNM) staging system for OPC (based on the p16INK4a (p16) status) was proposed and has been implemented. However, the applicability of this new staging system is still far from clear. In our study, n = 127 OPC patients from Queensland, Australia were recruited, and the tumor p16 expression in these patients was examined using immunohistochemical (IHC) analysis. HPV-16 genotyping, viral load, and physical status (episomal versus integrated) in the saliva s les of OPC patients were determined using the qPCR method. A good inter-rater agreement (k = 0.612) was found between tumor p16 expression and oral HPV-16 infection in OPC. Importantly, according to the eighth edition staging system, HPV-16 DNA viral load ( copies/50 ng) was significantly associated with the advanced stages of OPC. In concordance with previous studies, a mixed HPV-16 form (partially or fully integrated) was predominately found in OPC patients. Taken together, our data support HPV-16 detection in saliva as a screening biomarker to identify people within the community who are at risk of developing OPC.
Publisher: Croatian Society for Medical Biochemistry and Laboratory Medicine
Date: 2014
DOI: 10.11613/BM.2014.028
Publisher: Wiley
Date: 07-08-2019
DOI: 10.1002/CNCR.32435
Abstract: Fanconi anemia (FA) is a rare inherited genetic condition that may lead to bone marrow failure, leukemia, and/or solid tumors. It is caused by the loss of function of at least 1 gene of the FA/BRCA pathway, which is necessary for DNA repair. Patients with FA have a 200-fold to 1000-fold risk of developing head and neck cancer, mainly oral squamous cell carcinoma (OSCC), and of doing so at a much younger age than in iduals within the general population. Also, patients who have FA with OSCC have poor overall survival rates, reinforcing the necessity to detect OSCC early. The scope of the current review is to provide an update on OSCC in patients with FA.
Publisher: Future Science Ltd
Date: 03-2014
DOI: 10.4155/BIO.14.21
Abstract: Background: Dysregulation of salivary immunoglobulins has been implicated in illnesses ranging from periodontal disease to HIV aids and malignant cancers. Despite these advances there is a lack of agreement among studies with regard to the salivary immunoglobulin levels in healthy controls. Methodology: Resting and mechanically stimulated saliva s les and matching serum s les were collected from healthy in iduals (n = 33 40–55 years of age gender: 23 female, 10 male). A matrix-matched AlphaLISA ® assay was developed to determine the concentrations of IgG1 and IgG4 in serum and saliva s les. Conclusion: Clear relationships were observed in the flow rate and concentration of each immunoglobulin in the two types of saliva. This study affirms the need to establish and standardize collection methods before salivary IgGs are used for diagnostic purposes.
Publisher: Wiley
Date: 23-12-2018
DOI: 10.1002/HED.25578
Abstract: Accumulating evidence has suggested the utility of salivary oral rinse as a diagnostic fluid to detect oral human papillomavirus (HPV) DNA, but there are many methods for collecting saliva. Salivary oral rinse and unstimulated whole mouth saliva s les were collected from 45 oropharyngeal cancer (OPC) patients. We show a positive correlation of HPV-16 E2 (r = 0.95, P < 0.0001) and E6/7 (r = 0.93, P < 0.0001) relative copy number as well as HPV genotypes in both s le methods. There was a significant correlation between the two s le methods in the ratio of HPV16 E2 to E6/7 DNA (r = 0.46, P < 0.01). Consistent with previous studies, a mixed HPV-16 form (episomal and integrated) was commonly found in both saliva and tumor s les. Detection of HPV in saliva s les collected by either method yielded comparable results, and showed good sensitivity for detection of HPV derived from OPC.
Publisher: Springer Science and Business Media LLC
Date: 09-10-2020
DOI: 10.1007/S00392-019-01557-0
Abstract: Patients with HF are at a higher risk of rehospitalisation and, as such, significant costs to our healthcare system. A non-invasive method to collect body fluids and measure Gal-3 could improve the current management of HF. In this study, we investigated the potential prognostic utility of salivary Galectin-3 (Gal-3) in patients with heart failure (HF). We collected saliva s les from patients with HF (n = 105) either at hospital discharge or during routine clinical visits. Gal-3 concentrations in saliva s les were measured by ELISA. The Kaplan-Meier survival curve analysis and Cox proportional regression model were used to determine the potential prognostic utility of salivary Gal-3 concentrations. The primary end point was either cardiovascular death or hospitalisation. Salivary Gal-3 concentrations were significantly higher (p 172.58 ng/mL had a significantly (p 172.58 ng/mL demonstrated a higher cumulative risk of the primary outcome compared to those with lower Gal-3 levels, even after adjusting for other variables. Confirming our findings in a larger multi-centre clinical trial in the future would enable salivary Gal-3 measurements to form part of routine management for patients with HF.
Publisher: Springer Science and Business Media LLC
Date: 23-09-2016
Publisher: Elsevier BV
Date: 07-2023
Publisher: Oxford University Press (OUP)
Date: 07-2007
Abstract: To date, research into the biological processes and molecular mechanisms associated with endometrial receptivity and embryo implantation has been a focus of attention, whereas the complex events that occur in the human endometrium during the menstrual and proliferative phase under the influence of estrogen have received little attention. The objective of this review is to provide an update of our current understanding of the actions of estrogen on both human and rodent endometrium, with special emphasis on the regulation of uterine growth and cell proliferation, and the value of global gene expression analysis, in increasing understanding of these processes.
Publisher: MDPI AG
Date: 18-03-2019
Abstract: Objectives: In non-small cell lung cancers (NSCLC), tumour biopsy can often be an invasive procedure. The development of a non-invasive methodology to study genetic changes via circulating tumour cells (CTCs) is an appealing concept. Whilst CTCs typically remain as rare cells, improvements in epitope-independent CTC isolation techniques has given rise to a greater capture of CTCs. In this cross sectional study, we demonstrate the capture and characterization of NSCLC CTCs for the clinically actionable markers epidermal growth factor receptor (EGFR) alterations, anaplastic lymphoma kinase (ALK) rearrangements and programmed death ligand-1 (PD-L1) expression. The study identified CTCs/CTC clusters in 26/35 Stage IV NSCLC patients, and subsequently characterized the CTCs for EGFR mutation, ALK status and PD-L1 status. This pilot study demonstrates the potential of a non-invasive fluid biopsy to determine clinically relevant biomarkers in NSCLC.
Publisher: Mary Ann Liebert Inc
Date: 02-2017
Abstract: Saliva is an easily accessible s le and offers practical and noninvasive biomarker solutions as an alternative to blood and urine-based diagnostics. Saliva contains a plethora of biomolecules such as nucleic acids, hormones, proteins, and electrolytes. On the other hand, little is known on the extent to which the biomolecules in saliva vary over time within a given person. This baseline information is crucial for future development of robust saliva-based diagnostics. We have collected unstimulated whole mouth saliva from 20 healthy young men at four times during the day, including before and after a meal. We measured the salivary cortisol, testosterone, C-reactive protein (CRP), stability of genomic DNA (gDNA) and DNA methylation levels of APC, P16
Publisher: Oxford University Press (OUP)
Date: 26-04-2006
Abstract: Regeneration and growth of the human endometrium after shedding of the functional layer during menstruation depends on an adequate angiogenic response. We analysed the mRNA expression levels of all known vascular endothelial growth factor (VEGF) ligands and receptors in human endometrium collected in the menstrual and proliferative phases of the menstrual cycle. In addition, we evaluated the expression of VEGF-A, VEGF-R2 and NRP-1 at the protein level. Two periods of elevated mRNA expression of ligands and receptors were observed, separated by a distinct drop at cycle days (CDs) 9 and 10. Immunohistochemical staining showed that VEGF and VEGF-R2 were expressed in epithelial, stromal and endothelial cells. NRP-1 was mainly confined to stroma and blood vessels only in late-proliferative endometrium, epithelial staining was also observed. Except for endothelial VEGF-R2 expression in CDs 6-8, there were no significant differences in the expression of VEGF, VEGF-R2 or NRP-1 in any of the cell compartments. In contrast, VEGF release by cultured human endometrium explants decreased during the proliferative phase. This output was significantly reduced in menstrual and early-proliferative endometrium by estradiol (E2) treatment. Western blot analysis indicated that part of the VEGF-A was trapped in the extracellular matrix (ECM). Changes in VEGF ligands and receptors were associated with elevated expression of the hypoxia markers HIF1alpha and CA-IX in the menstrual and early proliferative phases. HIF1alpha was also detected in late-proliferative phase endometrium. Our findings indicate that VEGF-A exerts its actions mostly during the first half of the proliferative phase. Furthermore, VEGF-A production appears to be triggered by hypoxia in the menstrual phase and subsequently suppressed by estrogen during the late proliferative phase.
Publisher: MDPI AG
Date: 03-02-2020
DOI: 10.3390/BIOM10020223
Abstract: The role of human papillomavirus type 16 (HPV16) in oral potentially malignant disorders (OPMD) and oral cavity carcinoma (OC) is still under debate. We investigated HPV16 prevalence in unstimulated saliva, oral rinse s les, oral swabs and tumour biopsies collected from OPMD (n = 83) and OC (n = 106) patients. HPV16 genotype, viral load, physical status (episomal vs. integrated) and tumour p16INK4a expression were determined. Oral HPV16 prevalence was higher in OC than in OPMD, but this difference was not statistically significant (7.5% (8/106) versus 3.6% (3/83), odds ratio (OR): 2.18, 95% confidence interval (CI): 0.56, 8.48, p = 0.26). There was a significant association (p 0.05) between oral HPV16 infection and heavy tobacco consumption. Real-time PCR results indicated that no integration events occurred in either OPMD or OC cases based on the HPV16 E2/E6 ratio. HPV16 positive OPMD and OC patients had similar HPV16 E2 and E6 viral loads. The inter-rater agreement between tumour p16INK4a expression and oral HPV16 infection was considered as fair (k = 0.361) for OC. Our data suggest that the involvement of HPV16 in oral carcinogenesis is limited.
Publisher: Elsevier BV
Date: 02-2015
DOI: 10.1016/J.IJCARD.2014.12.052
Abstract: We have previously demonstrated that circulating NT-proBNP is truncated at the N and C termini. Aims of this study are three-fold: firstly to determine whether the NT-proBNP levels correlate with NYHA functional classes when measuring with different antibody pairs secondly to evaluate the diagnostic potential of ProBNP and thirdly to investigate whether combining NT-proBNP assays with or without ProBNP would lead to better diagnostic accuracies. Plasma s les were collected from healthy controls (n=52) and HF patients (n=46). Customized AlphaLISA® immunoassays were developed and validated to measure the concentrations of proBNP and NT-proBNP (with antibodies targeting 13-45, 13-76, 28-76). The diagnostic performance and predictive value of proBNP and NT-proBNP assays and their combinations were evaluated. Plasma proBNP assay showed acceptable diagnostic performance. NT-proBNP13-76 assay is useful in diagnosing and stratifying HF patients. The diagnostic performance of NT-proBNP13-76 demonstrated improvement over commercial NT-proBNP tests. The combination of NT-proBNP13-76 with NT-proBNP28-76 assays gave the best diagnostic assay performance. Our results demonstrate that while there is major heterogeneity in circulating NT-proBNP, specific epitopes of the peptides are extraordinarily stable, providing ideal targets for clinically useful diagnostic assays. Future new clinical diagnostic clinical trials should include a multimarker approach rather than using a single marker to diagnose HF.
Publisher: Wiley
Date: 05-09-2012
Publisher: Springer Science and Business Media LLC
Date: 26-08-2014
DOI: 10.1007/S13402-014-0188-2
Abstract: MicroRNAs (miRNAs) are known to play an important role in cancer development by post-transcriptionally affecting the expression of critical genes. The aims of this study were two-fold: (i) to develop a robust method to isolate miRNAs from small volumes of saliva and (ii) to develop a panel of saliva-based diagnostic biomarkers for the detection of head and neck squamous cell carcinoma (HNSCC). Five differentially expressed miRNAs were selected from miScript™ miRNA microarray data generated using saliva from five HNSCC patients and five healthy controls. Their differential expression was subsequently confirmed by RT-qPCR using saliva s les from healthy controls (n = 56) and HNSCC patients (n = 56). These s les were ided into two different cohorts, i.e., a first confirmatory cohort (n = 21) and a second independent validation cohort (n = 35), to narrow down the miRNA diagnostic panel to three miRNAs: miR-9, miR-134 and miR-191. This diagnostic panel was independently validated using HNSCC miRNA expression data from The Cancer Genome Atlas (TCGA), encompassing 334 tumours and 39 adjacent normal tissues. Receiver operating characteristic (ROC) curve analysis was performed to assess the diagnostic capacity of the panel. On average 60 ng/μL miRNA was isolated from 200 μL of saliva. Overall a good correlation was observed between the microarray data and the RT-qPCR data. We found that miR-9 (P <0.0001), miR-134 (P <0.0001) and miR-191 (P <0.001) were differentially expressed between saliva from HNSCC patients and healthy controls, and that these miRNAs provided a good discriminative capacity with area under the curve (AUC) values of 0.85 (P <0.0001), 0.74 (P < 0.001) and 0.98 (P < 0.0001), respectively. In addition, we found that the salivary miRNA data showed a good correlation with the TCGA miRNA data, thereby providing an independent validation. We show that we have developed a reliable method to isolate miRNAs from small volumes of saliva, and that the saliva-derived miRNAs miR-9, miR-134 and miR-191 may serve as novel biomarkers to reliably detect HNSCC.
Publisher: Wiley
Date: 27-01-2014
DOI: 10.1002/RCM.6806
Abstract: Diseases including cancer and congenital disorders of glycosylation have been associated with changes in the site-specific extent of protein glycosylation. Saliva can be non-invasively s led and is rich in glycoproteins, giving it the potential to be a useful biofluid for the discovery and detection of disease biomarkers associated with changes in glycosylation. Saliva was collected from healthy in iduals and glycoproteins were enriched using phenylboronic acid based glycoprotein enrichment resin. Proteins were deglycosylated with peptide-N-glycosidase F and digested with AspN or trypsin. Desalted peptides and deglycosylated peptides were separated by reversed-phase liquid chromatography and detected with on-line electrospray ionization quadrupole-time-of-flight mass spectrometry using a 5600 TripleTof instrument. Site-specific glycosylation occupancy was semi-quantitatively determined from the abundance of deglycosylated and nonglycosylated versions of each given peptide. Glycoprotein enrichment identified 67 independent glycosylation sites from 24 unique proteins, a 3.9-fold increase in the number of glycosylation sites identified. Enrichment of glycoproteins rather than glycopeptides allowed detection of both deglycosylated and nonglycosylated versions of each peptide, and thereby robust measurement of site-specific occupancy at 21 asparagines. Healthy in iduals showed limited biological variability in occupancy, with partially modified sites having characteristics consistent with inefficient glycosylation by oligosaccharyltransferase. Inclusion of negative controls without enzymatic deglycosylation controlled for spontaneous chemical deamidation, and identified asparagines previously incorrectly annotated as glycosylated. We developed a s le preparation and mass spectrometry detection strategy for rapid and efficient measurement of site-specific glycosylation occupancy on erse salivary glycoproteins suitable for biomarker discovery and detection of changes in glycosylation occupancy in human disease.
Publisher: Springer Science and Business Media LLC
Date: 03-08-2023
Publisher: Springer Science and Business Media LLC
Date: 06-01-2021
Publisher: Oxford University Press (OUP)
Date: 03-2005
DOI: 10.1530/EJE.1.01872
Abstract: Objective : It has been suggested that adiponectin regulates plasma free fatty acid (FFA) clearance by stimulating FFA uptake and/or oxidation in muscle. We aimed to determine changes in plasma adiponectin concentration and adiponectin receptor 1 and 2 mRNA expression in skeletal muscle during and after prolonged exercise under normal, fasting conditions (high FFA trial HFA) and following pharmacological inhibition of adipose tissue lipolysis (low FFA trial LFA). Furthermore, we aimed to detect and locate adiponectin in skeletal muscle tissue. Methods : Ten subjects performed two exercise trials (120 min at 50% VO 2max ). Indirect calorimetry was used to determine total fat oxidation rate. Plasma s les were collected at rest, during exercise and during post-exercise recovery to determine adiponectin, FFA and glycerol concentrations. Muscle biopsies were taken to determine adiponectin protein and adiponectin receptor 1 and 2 mRNA expression and to localise intramyocellular adiponectin. Results : Basal plasma adiponectin concentrations averaged 6.57±0.7 and 6.63±0.8 mg/l in the HFA and LFA trials respectively, and did not change significantly during or after exercise. In the LFA trial, plasma FFA concentrations and total fat oxidation rates were substantially reduced. However, plasma adiponectin and muscle adiponectin receptor 1 and 2 mRNA expression did not differ between trials. Immunohistochemical staining of muscle cross-sections showed the presence of adiponectin in the sarcolemma of in idual muscle fibres and within the interfibrillar arterioles. Conclusion : Plasma adiponectin concentrations and adiponectin receptor 1 and 2 mRNA expression in muscle are not acutely regulated by changes in adipose tissue lipolysis and/or plasma FFA concentrations. Adiponectin is abundantly expressed in muscle, and, for the first time, it has been shown to be present in/on the sarcolemma of in idual muscle fibres.
Publisher: Springer Science and Business Media LLC
Date: 10-02-2023
DOI: 10.1038/S41416-023-02167-4
Abstract: Head and neck cancers (HNC) are the seventh most prevalent cancer type globally. Despite their common categorisation, HNCs are a heterogeneous group of malignancies arising in various anatomical sites within the head and neck region. These cancers exhibit different clinical and biological manifestations, and this heterogeneity also contributes to the high rates of treatment failure and mortality. To evaluate patients who will respond to a particular treatment, there is a need to develop in vitro model systems that replicate in vivo tumour status. Among the methods developed, patient-derived cancer organoids, also known as tumouroids, recapitulate in vivo tumour characteristics including tumour architecture. Tumouroids have been used for general disease modelling and genetic instability studies in pan-cancer research. However, a limited number of studies have thus far been conducted using tumouroid-based drug screening. Studies have concluded that tumouroids can play an essential role in bringing precision medicine for highly heterogenous cancer types such as HNC.
Publisher: Springer Science and Business Media LLC
Date: 25-02-2019
DOI: 10.1038/S41378-019-0045-6
Abstract: Circulating tumor cells (CTCs) carry a wealth of information on primary and metastatic tumors critical for precise cancer detection, monitoring, and treatment. Numerous microfluidic platforms have been developed in the past few years to capture these rare cells in patient bloodstream for deciphering the critical information needed. However, the practical need for a high-quality method of CTC isolation remains to be met. Herein, we demonstrate a novel multi-flow microfluidic device that is able to sensitively provide high purity ( %) of separation outcome without labeling. Our device is constructed and configured based on the phenomenal effect of size-dependent inertial migration. The recovery rate of % has been achieved using spiked cancer cells at clinically relevant concentrations (10 cells per 5 mL and above). We have also successfully detected CTCs from 6 out of 8 non-small-cell-lung-cancer (NSCLC) patients, while none for 5 healthy control subjects. With these results, we envision our approach is a promising alternative for reliable CTC capture, and thus for facilitating the progress of extracting information from CTCs to personalize treatment strategies for solid tumor patients.
Publisher: Elsevier BV
Date: 05-2012
Publisher: Oxford University Press (OUP)
Date: 2021
Abstract: Biological markers that evaluate physical healing as well as psychological impact of a burn are essential for effective treatment of paediatric burns. The objective of this review is to summarize the evidence supporting the use of biomarkers in children with burns. An extensive review of the literature was performed using PubMed. A total of 59 biomarkers were identified relating to burn presence, specifically relating to processes involved in inflammation, wound healing, growth and metabolism. In addition, biomarkers involved in the stress response cascade following a burn trauma were also identified. Although many biomarkers have been identified that are potentially associated with burn-related physical and psychological trauma, an understanding of burn biology is still lacking in children. We propose that future research in the field of children’s burns should be conducted using broad screening methods for identifying potential biomarkers, examine the biological interactions of different biomarkers, utilize child-appropriate biological fluids such as urine or saliva, and include a range of different severity burns. Through further research, the biological response to burn injury may be fully realized and clinically relevant diagnostic tests and treatment therapies utilizing these biomarkers could be developed, for the improvement of healing outcomes in paediatric burn patients.
Publisher: Springer Berlin Heidelberg
Date: 2015
Publisher: Ivyspring International Publisher
Date: 2017
DOI: 10.7150/THNO.21804
Publisher: Springer Science and Business Media LLC
Date: 06-08-2022
DOI: 10.1007/S00432-022-04242-4
Abstract: Circulating tumour cells (CTCs) are a rare cell subpopulation regulated by the tumour microenvironment. In hypoxic conditions, CTCs are able to invade the lymphatic and circulatory systems leading to metastasis at distant sites. To mimic in vivo oxygen variations and effects on CTCs, we have cultured five non-small cell lung cancer (NSCLC) cell lines under normoxic and hypoxic conditions, followed by a pulse of reoxygenation for 4 h. Proliferation, spheroid-formation and colony formation ability under varying O 2 levels were investigated. Proliferation rate was not altered when cells were cultured in 2D models under hypoxic conditions. However, we observed that hypoxia enhanced in vitro formation of tumour-spheres and accelerated clonogenicity of NSCLC cell lines. In addition, cells exposed to hypoxia and reoxygenation conditions showed altered expression of epithelial-mesenchymal transition (EMT) related genes in NSCLC cell lines both at mRNA (AKT1, CAMK2NH1, DESI1, VIM, MAP1B, EGFR, ZEB1, HIF1α) and protein levels (Vimentin, Pan-cytokeratin). Our data suggest that when investigating CTCs as a prognostic biomarker in NSCLC, it is also essential to take into consideration EMT status to obtain a comprehensive overview of CTCs in circulation.
Publisher: MDPI AG
Date: 10-08-2018
DOI: 10.3390/MI9080397
Abstract: There is growing awareness for the need of early diagnostic tools to aid in point-of-care testing in cancer. Tumor biopsy remains the conventional means in which to s le a tumor and often presents with challenges and associated risks. Therefore, alternative sources of tumor biomarkers is needed. Liquid biopsy has gained attention due to its non-invasive s ling of tumor tissue and ability to serially assess disease via a simple blood draw over the course of treatment. Among the leading technologies developing liquid biopsy solutions, microfluidics has recently come to the fore. Microfluidic platforms offer cellular separation and analysis platforms that allow for high throughout, high sensitivity and specificity, low s le volumes and reagent costs and precise liquid controlling capabilities. These characteristics make microfluidic technology a promising tool in separating and analyzing circulating tumor biomarkers for diagnosis, prognosis and monitoring. In this review, the characteristics of three kinds of circulating tumor markers will be described in the context of cancer, circulating tumor cells (CTCs), exosomes, and circulating tumor DNA (ctDNA). The review will focus on how the introduction of microfluidic technologies has improved the separation and analysis of these circulating tumor markers.
Publisher: BMJ
Date: 24-05-2016
Publisher: SAGE Publications
Date: 2014
DOI: 10.4137/BMI.S16199
Abstract: Head and neck cancers (HNCs) represent a significant and ever-growing burden to the modern society, mainly due to the lack of early diagnostic methods. A significant number of HNCs is often associated with drinking, smoking, chewing beetle nut, and human papilloma virus (HPV) infections. We have analyzed DNA methylation patterns in tumor and normal tissue s les collected from head and neck squamous cell carcinoma (HNSCC) patients who were smokers. We have identified novel methylation sites in the promoter of the mediator complex subunit 15 ( MED15/PCQAP) gene (encoing a co-factor important for regulation of transcription initiation for promoters of many genes), hyper methylated specifically in tumor cells. Two clusters of CpG dinucleotides methylated in tumors, but not in normal tissue from the same patients, were identified. These CpG methylation events in saliva s les were further validated in a separate cohort of HNSCC patients (who developed cancer due to smoking or HPV infections) and healthy controls using methylation-specific PCR (MSP). We used saliva as a biological medium because of its non-invasive nature, close proximity to the tumors, easiness and it is an economically viable option for large-scale screening studies. The methylation levels for the two identified CpG clusters were significantly different between the saliva s les collected from healthy controls and HNSCC in iduals (Welch's t-test returning P 0.05 and Mann–Whitney test P 0.01 for both). The developed MSP assays also provided a good discriminative ability with AUC values of 0.70 ( P 0.01) and 0.63 ( P 0.05). The identified novel CpG methylation sites may serve as potential non-invasive biomarkers for detecting HNSCC.
Publisher: Springer Science and Business Media LLC
Date: 15-02-2021
Publisher: American Chemical Society (ACS)
Date: 27-05-2009
DOI: 10.1021/AC801244D
Abstract: We have developed a new protein microarray (ImmunoFlow Protein Platform, IFPP) that utilizes a porous nitrocellulose (NC) membrane with printed spots of capture probes. The s le is pumped actively through the NC membrane, to enhance binding efficiency and introduce stringency. Compared to protein microarrays assayed with the conventional incubation-shaking method the rate of binding is enhanced on the IFPP by at least a factor of 10, so that the total assay time can be reduced drastically without compromising sensitivity. Similarly, the sensitivity can be improved. We demonstrate the detection of 1 pM of C-reactive protein (CRP) in 70 microL of plasma within a total assay time of 7 min. The small s le and reagent volumes, combined with the speed of the assay, make our IFPP also well-suited for a point-of-care/near-patient setting. The potential clinical application of the IFPP is demonstrated by validating CRP detection both in human plasma and serum s les against standard clinical laboratory methods.
Publisher: Research Square Platform LLC
Date: 19-08-2020
DOI: 10.21203/RS.3.RS-55052/V1
Abstract: Despite advances in cancer treatment, the five-year mortality rate for oral cancers (OC) is 40%, mainly due to the lack of early diagnostics. To advance early diagnostics for high-risk and average-risk populations, we developed and evaluated machine-learning (ML) classifiers using metatranscriptomic data from saliva s les (n=433) collected from oral premalignant disorders (OPMD), OC patients (n=71) and normal controls (n=171). Our diagnostic classifiers yielded a receiver operating characteristics (ROC) area under the curve (AUC) up to 0.9, sensitivity up to 83% (92.3% for stage 1 cancer) and specificity up to 97.9%. Our metatranscriptomic signature incorporates both taxonomic and functional microbiome features, and reveals a number of previously known and novel taxa and functional pathways associated with OC. For the first time, we demonstrate the potential clinical utility of an AI/ML model for diagnosing OC early, opening a new era of non-invasive diagnostics, enabling early intervention and improved patient outcomes.
Publisher: Informa UK Limited
Date: 31-05-2001
Publisher: Elsevier BV
Date: 10-2012
DOI: 10.1593/TLO.12232
Publisher: Wiley
Date: 09-1997
DOI: 10.1002/(SICI)1097-4539(199709)26:5<249::AID-XRS188>3.0.CO;2-5
Publisher: Informa UK Limited
Date: 31-05-2001
Publisher: BMJ
Date: 23-06-2016
Publisher: Elsevier BV
Date: 09-2008
DOI: 10.1016/J.CONTRACEPTION.2008.05.003
Abstract: Fundamental and genetic differences between women in the endometrium may cause some to develop endometriosis, whereas others do not. Oral contraceptives (OC) may have an effect on the endometrium, rendering the development of endometriosis less likely. Endometrium from women using OC (OCE) and menstrual endometrium (ME) from normal cycling women were transplanted onto the chicken chorioallantoic membrane (CAM), and endometriosis-like lesion formation was evaluated. Microarray gene expression profiling was performed to identify differentially expressed genes in the endometrium from these groups. Microarray data were validated by real-time PCR. Less endometriosis-like lesions were formed after transplantation of OCE than after transplantation of ME (p<.05). Most of the differentially expressed genes belong to the TGFbeta superfamily. Real-time PCR validation revealed that inhibin betaA (INHBA) expression was significantly decreased in OCE as compared to ME. OC use affects the characteristics of endometrium, rendering it less potent to develop into endometriosis.
Publisher: Impact Journals, LLC
Date: 10-10-2017
Publisher: Elsevier BV
Date: 09-2018
DOI: 10.1016/J.ORALONCOLOGY.2018.07.001
Abstract: Over the past decade, there has been emerging research in the field of extracellular vesicles, especially those originating from endosomes, referred to as 'exosomes. Exosomes are membrane-bound nanovesicles secreted by most cell types upon fusion of multivesicular bodies (MVBs) to the cell plasma membrane. These vesicles are present in almost all body fluids such as blood, urine, saliva, breast milk, cerebrospinal and peritoneal fluids. Exosomes participate in intercellular communication by transferring the biologically active molecules like proteins, nucleic acids, and lipids to neighboring cells. Exosomes are enriched in the tumour microenvironment and growing evidence demonstrates that exosomes mediate cancer progression and metastasis. Given the important biological role played by these nanovesicles in cancer pathogenesis, these can be used as ideal non-invasive biomarkers in detecting and monitoring tumours as well as therapeutic targets. The scope of the current review is to provide an overview of exosomes with a special focus on salivary exosomes as potential biomarkers in head and neck cancers.
Publisher: Wiley
Date: 09-04-2023
DOI: 10.1002/CAM4.5857
Abstract: Despite aggressive treatment, more than 90% of glioblastoma (GBM) patients experience recurrences. GBM response to therapy is currently assessed by imaging techniques and tissue biopsy. However, difficulties with these methods may cause misinterpretation of treatment outcomes. Currently, no validated therapy response biomarkers are available for monitoring GBM progression. Metabolomics holds potential as a complementary tool to improve the interpretation of therapy responses to help in clinical interventions for GBM patients. Saliva and blood from GBM patients were collected pre and postoperatively. Patients were stratified conforming their progression‐free survival (PFS) into favourable or unfavourable clinical outcomes ( months or PFS ≤ 9 months, respectively). Analysis of saliva (whole‐mouth and oral rinse) and plasma s les was conducted utilising LC‐QqQ‐MS and LC‐QTOF‐MS to determine the metabolomic and lipidomic profiles. The data were investigated using univariate and multivariate statistical analyses and graphical LASSO‐based graphic network analyses. Altogether, 151 metabolites and 197 lipids were detected within all saliva and plasma s les. Among the patients with unfavourable outcomes, metabolites such as cyclic‐AMP, 3‐hydroxy‐kynurenine, dihydroorotate, UDP and cis‐aconitate were elevated, compared to patients with favourable outcomes during pre‐and post‐surgery. These metabolites showed to impact the pentose phosphate and Warburg effect pathways. The lipid profile of patients who experienced unfavourable outcomes revealed a higher heterogeneity in the abundance of lipids and fewer associations between markers in contrast to the favourable outcome group. Our findings indicate that changes in salivary and plasma metabolites in GBM patients can potentially be employed as less invasive prognostic biomarkers/biomarker panel but validation with larger cohorts is required.
Publisher: Medknow
Date: 2010
Publisher: Elsevier BV
Date: 10-2022
Publisher: Informa UK Limited
Date: 07-11-2019
Publisher: Elsevier BV
Date: 12-2017
DOI: 10.1016/J.ORALONCOLOGY.2017.10.011
Abstract: Immune checkpoint inhibitors have gained traction over the last few years in the treatment of metastatic/recurrent head and neck squamous cell carcinoma (HNSCC) patients. Monoclonal antibodies that block the programmed death 1 (PD-1) receptor and its major ligand, PD-L1, have shown durable responses and low toxicity profiles. There are currently no validated predictive biomarkers to select patients likely to respond to anti-PD-1/PD-L1 therapy to avoid unwanted side effects and to reduce healthcare expenditure. A circulating tumour cell (CTC) PD-L1 assay could be developed as a companion diagnostic tool to potentially predict the efficacy of immune checkpoint blockade treatments.
Publisher: American Association for Cancer Research (AACR)
Date: 07-2018
DOI: 10.1158/1538-7445.AM2018-5572
Abstract: Background: Metastasis in HNC patients is reflected by measurable levels of circulating tumor cells (CTCs) in the peripheral blood of cancer patients. CTCs represent cancer cells from the primary and metastatic sites, thereby providing a comprehensive representation of the tumor burden of an in idual patient. For patients without CTCs at presentation, the detection of CTCs in the blood and analysis of biomarkers within them provide an opportunity to identify patients "at risk" of developing overt metastasis, accelerating targeted treatment in addition to routing care with the clear aim of improving cure. Methods: Our study aimed to assess whether CTCs from the blood of HNC patients attending the Princess Alexandra Hospital and Royal Brisbane and Women's Hospital provided early cues of distant metastases (n=250). Results: With significant advances in CTC isolation technologies, we could demonstrate a higher CTC capture efficiency using epitope-independent platforms. By assessment of single and clustered CTCs, our data showed that HNC patients can be identified 4-6 months prior to developing clinical/radiographically evident metastasis. In these patients, a window for treatment escalation could become a possibility. In a proof-of-principle study, using novel culture formulations and hypoxic conditions (1-2% O2), we were able to demonstrate, for the first time, short-term patient-derived CTC cultures ex vivo from 7/18 HNC s les (4/7 HPV-positive, oropharyngeal) in a clinically relevant time period. Recent advancements have shown that PD-1 immune checkpoint therapies have durable responses in metastatic HNC patients that fail 1st- and 2nd-line therapy. Our preliminary data suggest PD-L1 is frequently expressed on HNC CTCs, and an immunoscore may be able to stratify patients likely to respond to immunotherapy. Conclusion: Expanding CTCs outside the patient's body allows for the recapitulation of the molecular ersity present within the tumor, understanding the disease progression and testing of therapies. Patients with a high percentage of PD-L1+ CTCs could be potential candidates for anti-PD-L1 therapy, a promising new immunotherapy. Citation Format: Arutha Kulasinghe, Chris Perry, Liz Kenny, Tony Blick, Majid Warkiani, Ian Vela, Ken O'Byrne, Jean-Paul Thiery, Erik Thompson, Colleen Nelson, Chamindie Punyadeera. Circulating tumor cells: The tumor trail left in the blood [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018 2018 Apr 14-18 Chicago, IL. Philadelphia (PA): AACR Cancer Res 2018 (13 Suppl):Abstract nr 5572.
Publisher: Springer Science and Business Media LLC
Date: 10-12-2020
DOI: 10.1007/S00392-019-01583-Y
Abstract: The original version of this article unfortunately contained a mistake.
Publisher: Public Library of Science (PLoS)
Date: 31-10-2012
Publisher: Elsevier BV
Date: 07-2013
DOI: 10.1016/J.JCHROMB.2013.05.001
Abstract: Saliva contains a number of biochemical components which may be useful for diagnosis/monitoring of metabolic disorders, and as markers of cancer or heart disease. Saliva collection is attractive as a non-invasive s ling method for infants and elderly patients. We present a method suitable for saliva collection from neonates. We have applied this technique for the determination of salivary nucleotide metabolites. Saliva was collected from 10 healthy neonates using washed cotton swabs, and directly from 10 adults. Two methods for saliva extraction from oral swabs were evaluated. The analytes were then separated using high performance liquid chromatography (HPLC) with tandem mass spectrometry (MS/MS). The limits of detection for 14 purine yrimidine metabolites were variable, ranging from 0.01 to 1.0μM. Recovery of hydrophobic purine yrimidine metabolites from cotton tips was consistently high using water/acetonitrile extraction (92.7-111%) compared with water extraction alone. The concentrations of these metabolites were significantly higher in neonatal saliva than in adults. Preliminary ranges for nucleotide metabolites in neonatal and adult saliva are reported. Hypoxanthine and xanthine were grossly raised in neonates (49.3±25.4 30.9±19.5μM respectively) compared to adults (4.3±3.3 4.6±4.5μM) nucleosides were also markedly raised in neonates. This study focuses on three essential details: contamination of oral swabs during manufacturing and how to overcome this weighing swabs to accurately measure small saliva volumes and methods for extracting saliva metabolites of interest from cotton swabs. A method is described for determining nucleotide metabolites using HPLC with photodiode array or MS/MS. The advantages of utilising saliva are highlighted. Nucleotide metabolites were not simply in equilibrium with plasma, but may be actively secreted into saliva, and this process is more active in neonates than adults.
Publisher: MDPI AG
Date: 12-04-2019
DOI: 10.3390/BIOM9040148
Abstract: Silencing of tumor-suppressor genes (TSGs) by DNA promoter hypermethylation is an early event in carcinogenesis hence, TSGs may serve as early tumor biomarkers. We determined the promoter methylation levels of p16INK4a, RASSF1A, TIMP3, and PCQAP/MED15 TSGs in salivary DNA from oral cancer (OC) and oropharyngeal cancer (OPC) patients, using methylation-specific PCR coupled with densitometry analysis. We assessed the association between DNA methylation of in idual TSGs with OC and OPC risk factors. The performance and the clinical validity of this quadruple-methylation marker panel were evaluated in discriminating OC and OPC patients from healthy controls using the CombiROC web tool. Our study reports that RASSF1A, TIMP3, and PCQAP/MED15 TSGs were significantly hypermethylated in OC and OPC cases compared to healthy controls. DNA methylation levels of TSGs were significantly augmented by smoking, alcohol use, and betel quid chewing, indicating the fact that frequent exposure to risk factors may drive oral and oropharyngeal carcinogenesis through TSG promoter hypermethylation. Also, this quadruple-methylation marker panel of p16INK4a, RASSF1A, TIMP3, and PCQAP/MED15 TSGs demonstrated excellent diagnostic accuracy in the early detection of OC at 91.7% sensitivity and 92.3% specificity and of OPC at 99.8% sensitivity and 92.1% specificity from healthy controls.
Publisher: Oxford University Press (OUP)
Date: 11-09-2007
Abstract: To identify specific markers of rectovaginal endometriotic nodule vasculature, highly enriched preparations of vascular endothelial cells and pericytes were obtained from endometriotic nodules and control endometrial and myometrial tissue by laser capture microdissection (LCM), and gene expression profiles were screened by microarray analysis. Of the 18 400 transcripts on the arrays, 734 were significantly overexpressed in vessels from fibromuscular tissue and 923 in vessels from stromal tissue of endometriotic nodules, compared with vessels dissected from control tissues. The most frequently expressed transcripts included known endothelial cell-associated genes, as well as transcripts with little or no previous association with vascular cells. The higher expression in blood vessels was further corroborated by immunohistochemical staining of six potential markers, five of which showed strong expression in pericytes. The most promising marker was matrix Gla protein, which was found to be present in both glandular epithelial cells and vascular endothelial cells of endometriotic lesions, although it was barely expressed at all in normal endometrium. LCM, combined with microarray analysis, constitutes a powerful tool for mapping the transcriptome of vascular cells. After immunohistochemical validation, markers of vascular endothelial and perivascular cells from endometriotic nodules could be identified, which may provide targets to improve early diagnosis or to selectively deliver therapeutic agents.
Publisher: Springer Science and Business Media LLC
Date: 27-10-2022
DOI: 10.1007/S00216-022-04376-X
Abstract: Glycosylation is the most common post-translational modification of proteins, and glycosylation changes at cell surfaces are frequently associated with malignant epithelia including head and neck squamous cell carcinoma (HNSCC). In HNSCC, 5-year survival remains poor, averaging around 50% globally: this is partly related to late diagnosis. Specific protein glycosylation signatures on malignant keratinocytes have promise as diagnostic and prognostic biomarkers and as therapeutic targets. Nevertheless, HNSCC-specific glycome is to date largely unknown. Herein, we tested six established HNSCC cell lines to capture the qualitative and semi-quantitative N-glycome using porous graphitized carbon liquid chromatography coupled to electrospray ionisation tandem mass spectrometry. Oligomannose-type N-glycans were the predominant features in all HNSCC cell lines analysed (57.5–70%). The levels of sialylated N-glycans showed considerable cell line-dependent differences ranging from 24 to 35%. Importantly, α2-6 linked sialylated N-glycans were dominant across most HNSCC cell lines except in SCC-9 cells where similar levels of α2-6 and α2-3 sialylated N-glycans were observed. Furthermore, we found that HPV-positive cell lines contained higher levels of phosphorylated oligomannose N-glycans, which hint towards an upregulation of lysosomal pathways. Almost all fucose-type N-glycans carried core-fucose residues with just minor levels ( 4%) of Lewis-type fucosylation identified. We also observed paucimannose-type N-glycans (2–5.5%), though in low levels. Finally, we identified oligomannose N-glycans carrying core-fucose residues and confirmed their structure by tandem mass spectrometry. This first systematic mapping of the N-glycome revealed erse and specific glycosylation features in HNSCC, paving the way for further studies aimed at assessing their possible diagnostic relevance.
Publisher: Elsevier BV
Date: 12-2012
DOI: 10.1016/J.JCHROMB.2012.10.023
Abstract: Saliva is a crucial biofluid for oral health and is also of increasing importance as a non-invasive source of disease biomarkers. Salivary alpha-amylase is an abundant protein in saliva, and changes in amylase expression have been previously associated with a variety of diseases and conditions. Salivary alpha-amylase is subject to a high ersity of post-translational modifications, including physiological proteolysis in the oral cavity. Here we developed methodology for rapid s le preparation and non-targeted LC-ESI-MS/MS analysis of saliva from healthy subjects and observed an extreme ersity of alpha-amylase proteolytic isoforms. Our results emphasize the importance of consideration of post-translational events such as proteolysis in proteomic studies, biomarker discovery and validation, particularly in saliva.
Publisher: MDPI AG
Date: 03-2017
Publisher: Elsevier BV
Date: 10-2016
DOI: 10.1016/J.ORALONCOLOGY.2016.07.014
Abstract: Developing non-invasive diagnostic tools in the field of head and neck oncology has been a challenge. Analysis of circulating tumour derivatives in a patient's blood has been explored in other solid cancers. This includes analysis of circulating tumour DNA, intact circulating tumour cells (CTCs) and exosomes. These circulating tumour derivatives provide avenues of investigation which can be representative of a patient's primary tumour signature and can be assessed from a patient's blood s le. In advanced stage cancer patients, these tumour derivatives are found in higher amounts, attributed to higher cellular turnover (apoptosis, autophagy), lysed CTCs and sloughing from necrotic tumours. Head and neck squamous cell carcinoma (HNSCC) patients often present with advanced disease associated with a poor 5-year survival of <50%. Outside of sophisticated imaging and clinical examination, there is a lack of available biomarkers to measure disease burden, and/or response to therapy. Implementation of a liquid biopsy in HNSCC through serial blood s les has the potential to detect metastatic events earlier, thereby allowing better selection of appropriate treatment choices, predict prognosis in patients with potentially curable disease, monitor systemic therapies and residual disease post-treatment.
Publisher: Springer Science and Business Media LLC
Date: 08-12-2021
DOI: 10.1038/S41525-021-00257-X
Abstract: Despite advances in cancer treatment, the 5-year mortality rate for oral cancers (OC) is 40%, mainly due to the lack of early diagnostics. To advance early diagnostics for high-risk and average-risk populations, we developed and evaluated machine-learning (ML) classifiers using metatranscriptomic data from saliva s les ( n = 433) collected from oral premalignant disorders (OPMD), OC patients ( n = 71) and normal controls ( n = 171). Our diagnostic classifiers yielded a receiver operating characteristics (ROC) area under the curve (AUC) up to 0.9, sensitivity up to 83% (92.3% for stage 1 cancer) and specificity up to 97.9%. Our metatranscriptomic signature incorporates both taxonomic and functional microbiome features, and reveals a number of taxa and functional pathways associated with OC. We demonstrate the potential clinical utility of an AI/ML model for diagnosing OC early, opening a new era of non-invasive diagnostics, enabling early intervention and improved patient outcomes.
Publisher: Frontiers Media SA
Date: 15-03-2022
Abstract: Despite efforts to improve earlier diagnosis of non-small cell lung cancer (NSCLC), most patients present with advanced stage disease, which is often associated with poor survival outcomes with only 15% surviving for 5 years from their diagnosis. Tumour tissue biopsy is the current mainstream for cancer diagnosis and prognosis in many parts of the world. However, due to tumour heterogeneity and accessibility issues, liquid biopsy is emerging as a game changer for both cancer diagnosis and prognosis. Liquid biopsy is the analysis of tumour-derived biomarkers in body fluids, which has remarkable advantages over the use of traditional tumour biopsy. Circulating tumour cells (CTCs) and circulating tumour DNA (ctDNA) are two main derivatives of liquid biopsy. CTC enumeration and molecular analysis enable monitoring of cancer progression, recurrence, and treatment response earlier than traditional biopsy through a minimally invasive liquid biopsy approach. CTC-derived ex-vivo cultures are essential to understanding CTC biology and their role in metastasis, provide a means for personalized drug testing, and guide treatment selection. Just like CTCs, ctDNA provides opportunity for screening, monitoring, treatment evaluation, and disease surveillance. We present an updated review highlighting the prognostic and therapeutic significance of CTCs and ctDNA in NSCLC.
Publisher: Informa UK Limited
Date: 18-01-2016
DOI: 10.1586/14737159.2016.1127758
Abstract: Head and neck cancer patients often present with advanced metastatic disease resulting in a poor 5-year survival. Therefore, there is a need for non-invasive diagnostic tools that could complement conventional imaging to inform clinicians of patient outcomes and treatment responses. A liquid biopsy addresses this unmet clinical need a simple peripheral blood draw could provide information about the disseminated disease in terms of circulating tumor cells and circulating tumor DNA. Moreover, detectable tumor DNA in the saliva of head and neck cancer patients could signify the early signs of the disease and present an opportunity for clinical intervention. This review provides an overview of the current literature with regard to the feasibility of such a test in the head and neck cancer field and highlights the need for such a test.
Publisher: Ivyspring International Publisher
Date: 2023
DOI: 10.7150/THNO.78872
Publisher: Elsevier BV
Date: 11-2018
Publisher: Wiley
Date: 2000
DOI: 10.1002/(SICI)1097-4539(200001/02)29:1<53::AID-XRS405>3.0.CO;2-S
Publisher: Frontiers Media SA
Date: 03-06-2021
Abstract: Glioblastoma (GBM) is the most common and aggressive type of tumour arising from the central nervous system. GBM remains an incurable disease despite advancement in therapies, with overall survival of approximately 15 months. Recent literature has highlighted that GBM releases tumoural content which crosses the blood-brain barrier (BBB) and is detected in patients’ blood, such as circulating tumour cells (CTCs). CTCs carry tumour information and have shown promise as prognostic and predictive biomarkers in different cancer types. Currently, there is limited data for the clinical utility of CTCs in GBM. Here, we report the use of spiral microfluidic technology to isolate CTCs from whole blood of newly diagnosed GBM patients before and after surgery, followed by characterization for GFAP, cell-surface vimentin protein expression and EGFR lification. CTCs were found in 13 out of 20 patients (9/20 before surgery and 11/19 after surgery). Patients with CTC counts equal to 0 after surgery had a significantly longer recurrence-free survival (p=0.0370). This is the first investigation using the spiral microfluidics technology for the enrichment of CTCs from GBM patients and these results support the use of this technology to better understand the clinical value of CTCs in the management of GBM in future studies.
Publisher: Elsevier BV
Date: 08-2021
Publisher: Springer Science and Business Media LLC
Date: 02-06-2021
DOI: 10.1007/S40291-021-00538-2
Abstract: Increasing evidence supports the notion that human papillomavirus (HPV) DNA integration onto the human genome can influence and alter the molecular cargo in the exosomes derived from head and neck cancer cells. However, the molecular cargo of salivary exosomes derived from HPV-driven oropharyngeal cancer (HPV-driven OPC) remains unelucidated. Salivary exosomes morphology and molecular characterizations were examined using the nanoparticle tracking (NTA), western blot analysis, transmission electron microscopy (TEM) and mass spectrometry analysis. We report that HPV16 DNA was detected (80%) in isolated salivary exosomes of HPV-driven OPC patients. Importantly, we demonstrate elevated protein levels of six main glycolytic enzymes [i.e., aldolase (ALDOA), glyceraldehye-3-phosphate dehydrogenase (GAPDH), lactate dehydrogenase A/B (LDHA and LDHB), phosphoglycerate kinase 1 (PGK1) and pyruvate kinase M1/2 (PKM)] in isolated salivary exosomes of HPV-driven OPC patients, suggesting a novel mechanism underlying the potential role of salivary exosomes in mediating the reciprocal interplay between glucose metabolism and HPV-driven OPC. Our data demonstrate the potential diagnostic value of HPV16 DNA and glycolytic enzymes in salivary exosomes in discriminating healthy controls from HPV-driven OPC patients, thereby opening new avenues in the future for clinical translation studies.
Publisher: Wiley
Date: 17-02-2022
DOI: 10.1002/CNCR.34148
Abstract: Although the majority of human papillomavirus (HPV) infections are cleared by the immune system, a small percentage of them progress to develop HPV-driven cancers. Cervical cancer studies highlight that HPV persistence and cancer risk are associated with genetic factors, especially at the human leukocyte antigen (HLA) genes. This study was conducted to investigate such associations in head and neck cancer (HNC). In all, 192 patients with HNC and 384 controls were genotyped with the Infinium Global Screening Array (Illumina, Inc). HLA variants were imputed with SNP2HLA, and an association analysis was performed by logistic regression. HPV-positive HNCs were significantly associated with single-nucleotide polymorphisms (SNPs) at DRB1_32660090 (P = 1.728 × 10 HPV-positive HNC risk is associated with distinct HLA variants, and some of them are shared by both cervical cancer and HPV-positive HNC. Human papillomavirus (HPV)-positive head and neck cancer (HNC) risk is associated with distinct human leukocyte antigen variants, and some of them are shared by both cervical cancer and HPV-positive HNC. Cervical cancer studies highlight that human papillomavirus (HPV)-driven cancer risk is linked with human leukocyte antigen (HLA) polymorphism. Hence, the current study was designed to investigate the HLA associations in HPV-positive and HPV-negative head and neck cancer (HNC) and compare these associations with cervical cancer. Several lead signals reported by previous HNC and cervical genome-wide association studies were replicated in the current study. However, these associations were limited to the HPV-positive HNC group, and this suggests that HPV-positive HNC risk is associated with distinct HLA variants, and some of them are shared by both cervical cancer and HPV-positive HNC.
Publisher: AMPCo
Date: 29-09-2020
DOI: 10.5694/MJA2.50797
Publisher: Springer Science and Business Media LLC
Date: 13-07-2018
Publisher: Cold Spring Harbor Laboratory
Date: 31-07-2022
DOI: 10.1101/2022.07.30.22278239
Abstract: Oral squamous cell carcinoma (OSCC) and oropharyngeal squamous cell carcinoma (OPSCC) are the two major subtypes of head and neck cancer (HNC) that can go undetected resulting in late detection and poor outcomes. We describe the development and validation of a convenient and easy-to-use test, called CancerDetect for Oral & Throat cancer™ (CDOT), to detect markers of OSCC and/or OPSCC within a high-risk population using salivary metatranscriptomics. We collected saliva s les from 1,175 unique in iduals who were 50 years or older, or adults who had a history of tobacco use. All saliva s les were processed through a metatranscriptomic method to isolate microbial organisms and functions, as well as human transcripts. Of the 1175 s les, 945 were used to train a classifier using machine learning methods, resulting in a salivary RNA metatranscriptomic signature. The classifier was then independently validated on the 230 remaining s les unseen by the classifier, consisting of 20 OSCC (all stages), 76 OPSCC (all stages), and 134 negatives (including 14 pre-malignant). On the validation cohort, the specificity of the CDOT test was 94%, sensitivity was 90% for participants with a histopathological diagnosis of OSCC, and 84.2% for participants with a diagnosis of OPSCC. Similar classification results were observed among people in early stage (stages I & II) vs late stage (stages III & IV) of OSCC and OPSCC. CDOT is a non-invasive test that can be easily administered in dentist offices, primary care centers and specialized cancer clinics for early detection of OPSCC and OSCC. This test, having received breakthrough designation by the US Food and Drug Administration (FDA), will broadly enable early diagnosis of OSCC and OPSCC, saving lives and significantly reducing healthcare expenditure.
Publisher: Elsevier BV
Date: 2018
DOI: 10.2139/SSRN.3237005
Publisher: Frontiers Media SA
Date: 03-08-2018
Publisher: Impact Journals, LLC
Date: 09-08-2021
Publisher: Springer Science and Business Media LLC
Date: 31-10-2019
DOI: 10.1038/S41416-019-0603-6
Abstract: Gliomas are the most common tumours of the central nervous system and the most aggressive form is glioblastoma (GBM). Despite advances in treatment, patient survival remains low. GBM diagnosis typically relies on imaging techniques and postoperative pathological diagnosis however, both procedures have their inherent limitations. Imaging modalities cannot differentiate tumour progression from treatment-related changes that mimic progression, known as pseudoprogression, which might lead to misinterpretation of therapy response and delay clinical interventions. In addition to imaging limitations, tissue biopsies are invasive and most of the time cannot be performed over the course of treatment to evaluate ‘real-time’ tumour dynamics. In an attempt to address these limitations, liquid biopsies have been proposed in the field. Blood s ling is a minimally invasive procedure for a patient to endure and could provide tumoural information to guide therapy. Tumours shed tumoural content, such as circulating tumour cells, cell-free nucleic acids, proteins and extracellular vesicles, into the circulation, and these biomarkers are reported to cross the blood–brain barrier. The use of liquid biopsies is emerging in the field of GBM. In this review, we aim to summarise the current literature on circulating biomarkers, namely circulating tumour cells, circulating tumour DNA and extracellular vesicles as potential non-invasively s led biomarkers to manage the treatment of patients with GBM.
Publisher: Elsevier BV
Date: 12-2022
Publisher: Wiley
Date: 11-08-2015
DOI: 10.1002/IJC.29108
Abstract: Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer with 650,000 new cases p/a worldwide. HNSCC causes high morbidity with a 5-year survival rate of less than 60%, which has not improved due to the lack of early detection (Bozec et al. Eur Arch Otorhinolaryngol. 2013 : 2745-9). Metastatic disease remains one of the leading causes of death in HNSCC patients. This review article provides a comprehensive overview of literature over the past 5 years on the detection of circulating tumour cells (CTCs) in HNSCC CTC biology and future perspectives. CTCs are a hallmark of invasive cancer cells and key to metastasis. CTCs can be used as surrogate markers of overall survival and progression-free survival. CTCs are currently used as prognostic factors for breast, prostate and colorectal cancers using the CellSearch® system. CTCs have been detected in HNSCC, however, these numbers depend on the technique applied, time of blood collection and the clinical stage of the patient. The impact of CTCs in HNSCC is not well understood, and thus, not in routine clinical practice. Validated detection technologies that are able to capture CTCs undergoing epithelial-mesenchymal transition are needed. This will aid in the capture of heterogeneous CTCs, which can be compiled as new targets for the current food and drug administration-cleared CellSearch® system. Recent studies on CTCs in HNSCC with the CellSearch® have shown variable data. Therefore, there is an immediate need for large clinical trials encompassing a suite of biomarkers capturing CTCs in HNSCC, before CTCs can be used as prognostic markers in HNSCC patient management.
Publisher: MDPI AG
Date: 15-06-2020
DOI: 10.3390/CELLS9061465
Abstract: Tumor tissue biopsy is often limited for non-small cell lung cancer (NSCLC) patients and alternative sources of tumoral information are desirable to determine molecular alterations such as anaplastic lymphoma kinase (ALK) rearrangements. Circulating tumor cells (CTCs) are an appealing component of liquid biopsies, which can be s led serially over the course of treatment. In this study, we enrolled a cohort of ALK-positive (n = 8) and ALK-negative (n = 12) NSCLC patients, enriched for CTCs using spiral microfluidic technology and performed DNA fluorescent in situ hybridization (FISH) for ALK. CTCs were identified in 12/20 NSCLC patients ranging from 1 to 26 CTCs/7.5 mL blood. Our study revealed that 3D imaging of CTCs for ALK translocations captured a well-defined separation of 3′ and 5′ signals indicative of ALK translocations and overlapping 3′/5′ signal was easily resolved by imaging through the nuclear volume. This study provides proof-of-principle for the use of 3D DNA FISH in the determination of CTC ALK translocations in NSCLC.
Publisher: Informa UK Limited
Date: 2018
Publisher: Elsevier
Date: 2022
Publisher: Wiley
Date: 02-02-2021
DOI: 10.1002/HED.26619
Publisher: American Society of Clinical Oncology (ASCO)
Date: 20-05-2020
DOI: 10.1200/JCO.2020.38.15_SUPPL.E21692
Abstract: e21692 Background: Tumour tissue-based information is limited. Liquid biopsy can provide valuable real-time information through circulating tumour cells (CTCs). Profiling and expanding CTCs may provide avenues to study transient metastatic disease. Methods: 70 NSCLC patients were recruited. CTCs were enriched using the spiral microfluidic chip and a RosetteSep™ using bloods from NSCLC patients. CTC cultures were carried out using the Clevers media under hypoxic conditions. CTCs were characterized using immunofluorescence and mutation-specific antibodies for s les with known mutation profiles. Exome sequencing was used to characterized CTC cultures. Results: CTCs ( 2 cells) were detected in 38/70 (54.3%) of patients ranging from 0-385 CTCs per 7.5ml blood. In 4/5 patients where primary tumours harboured an EGFR exon 19 deletion, this EGFR mutation was also captured in CTCs. ALK translocation was confirmed on CTCs from a patient harbouring an ALK-rearrangement in the primary tumour. Short term CTC cultures were successfully generated in 9/70 NSCLC patients. Whole exome sequencing confirmed the presence of somatic mutations in the CTC cultures with mutational signatures consistent with NSCLC. Conclusions: This study demonstrates a workflow for ex vivo culture of CTCs from NSCLC blood s les.
Publisher: Springer Science and Business Media LLC
Date: 08-2001
Abstract: The effects of free fatty acids (FFA), leptin, tumour necrosis factor (TNF) alpha and body fat distribution on in vivo oxidation of a glucose load were studied in two South African ethnic groups. Anthropometric and various metabolic indices were measured at fasting and during a 7 h oral glucose tolerance test (OGTT). Body composition was measured using bioelectrical impedance analysis and subcutaneous and visceral fat mass was assessed using a five- and two-level CT-scan respectively. Glucose oxidation was evaluated by measuring the ratio of (13)CO(2) to (12)CO(2) in breath following ingestion of 1-(13)C-labelled glucose. Ten lean black women (LBW), ten obese black women (OBW), nine lean white women (LWW) and nine obese white women (OWW) were investigated after an overnight fast. Visceral fat levels were significantly higher (P<0.01) in obese white than black women, despite similar body mass indexes (BMIs). There were no ethnic differences in glucose oxidation however in the lean subjects of both ethnic groups the area under the curve (AUC) was higher than in obese subjects (P<0.05 for both) and was found to correlate negatively with weight (r=-0.69, P<0.01) after correcting for age. Basal TNF alpha concentrations were similar in all groups. Percentage suppression of FFAs at 30 min of the OGTT was 24+/-12% in OWW and -38+/-23% (P<0.05) in OBW, ie the 30 min FFA level was higher than the fasting level in the latter group. AUC for FFAs during the late postprandial period (120--420 min) was significantly higher in OWW than OBW (P<0.01) and LWW (P<0.01) and correlated positively with visceral fat mass independent of age (r=0.78, P<0.05) in the OWW only. Leptin levels were higher (P<0.01) both at fasting and during the course of the OGTT in obese women from both ethnic groups compared to the lean women. Glucose oxidation is reduced in obese subjects of both ethnic groups inter- and intra-ethnic differences were observed in visceral fat mass and FFA production and it is possible that such differences may play a role in the differing prevalences of obesity-related disorders that have been reported in these two populations.
Publisher: Elsevier BV
Date: 2011
Publisher: Informa UK Limited
Date: 15-06-2017
DOI: 10.1080/14737159.2017.1339603
Abstract: MicroRNAs (miRs) are short (~20 nucleotides) non-coding ribonuecleic acids (ncRNAs) known to be involved in cellular processes such as proliferation, differentiation, immune response, pathogenicity and tumourigenesis, among many others. The regulatory mechanisms exerted by miRs have been implicated in many cancers, including Human Papillomavirus (HPV)-associated cancers. Areas covered: In this review, the authors discuss the involvement of miRs (-143, -375, -21, -200, -296 etc.) that have been shown to be dysregulated in HPV-associated cancers. This review also encompasses both intracellular and exosomal miRs, and their potential as diagnostic biomarkers in saliva and blood. The authors have also attempted to dissect the functional impact of miRs on cellular processes such as changes in cellular polarity, loss of apoptosis and tumour suppression, and unchecked and uncontrolled cell cycle regulation, all of which ultimately lead to aberrant cellular proliferation. Expert commentary: Identification of dysregulated miRs in HPV-associated cancers opens up new opportunities to develop diagnostic, therapeutic and prognostic biomarkers. Studies on global expression patterns of miRs dysregulated in HPV-associated cancers can be instrumental in developing broader therapeutic strategies. Therapies like anti-miR, miR-replacement and those based on alternative natural products targeting miRs, need to be improved and better synchronized to be cost-effective and have better treatment outcomes.
Publisher: Oxford University Press (OUP)
Date: 18-10-2007
Abstract: Estradiol (E(2)) is an important promoter of the growth of both eutopic and ectopic endometrium. The findings with regard to the expression and activity of steroidogenic enzymes in endometrium of controls, in endometrium of endometriosis patients and in endometriotic lesions are not consistent. In this study, we have looked at the mRNA expression and protein levels of a range of steroidogenic enzymes [aromatase, 17beta-hydroxysteroid dehydrogenases (17beta-HSD) type 1, 2 and 4, estrogen sulfotransferase (EST) and steroid sulfatase (STS)] in eutopic and ectopic endometrium of patients (n = 14) with deep-infiltrative endometriosis as well as in disease-free endometrium (n = 48) using real-time PCR and immunocytochemistry. In addition, we evaluated their menstrual cycle-related expression patterns, and investigated their steroid responsiveness in explant cultures. Aromatase and 17beta-HSD type 1 mRNA levels were extremely low in normal human endometrium, while mRNAs for types 2 and 4 17beta-HSD, EST and STS were readily detectable. Only 17beta-HSD type 2 and EST genes showed sensitivity to progesterone in normal endometrium. Types 1 and 2 17beta-HSD and STS protein was detected in normal endometrium using new polyclonal antibodies. In endometriosis lesions, the balance is tilted in favor of enzymes producing E(2). This is due to a suppression of types 2 and 4 17beta-HSD, and an increased expression of aromatase and type 1 17beta-HSD in ectopic endometrium.
Publisher: Elsevier BV
Date: 12-2020
Publisher: Wiley
Date: 31-05-2023
DOI: 10.1002/CNCR.34888
Abstract: Extracellular vesicles (EVs) play a critical role in intercellular communication under physiological and pathological conditions, including cancer. EVs cargo reflects their cell of origin, suggesting their utility as biomarkers. EVs are detected in several biofluids, and their ability to cross the blood–brain barrier has highlighted their potential as prognostic and diagnostic biomarkers in gliomas, including glioblastoma (GBM). Studies have demonstrated the potential clinical utility of plasma‐derived EVs in glioma. However, little is known about the clinical utility of saliva‐derived EVs in GBM. Small EVs were isolated from whole mouth saliva of GBM patients pre‐ and postoperatively. Isolation was performed using differential centrifugation and/or ultracentrifugation. EVs were characterized by concentration, size, morphology, and EVs cell‐surface protein markers. Protein cargo in EVs was profiled using mass spectrometry. There were no statistically significant differences in size and concentration of EVs derived from pre‐ and post GBM patients' saliva s les. A higher number of proteins were detected in preoperative s les compared to postoperative s les. The authors found four highly abundant proteins (aldolase A, 14‐3‐3 protein ε, enoyl CoA hydratase 1, and transmembrane protease serine 11B) in preoperative saliva s les from GBM patients with poor outcomes. Functional enrichment analysis of pre‐ and postoperative saliva s les showed significant enrichment of several pathways, including those related to the immune system, cell cycle and programmed cell death. This study, for the first time, demonstrates the feasibility of isolating and characterizing small EVs from pre‐ and postoperative saliva s les from GBM patients. Preliminary findings encourage further large cohort validation studies on salivary small EVs to evaluate prognosis in GBM.
Publisher: American Association for Cancer Research (AACR)
Date: 15-08-2020
DOI: 10.1158/1538-7445.AM2020-3384
Abstract: Metastasis in cancer patients is reflected by measurable levels of circulating tumor cells (CTCs) in the blood of cancer patients. CTCs represent cancer cells from the primary and metastatic sites, thereby providing a comprehensive representation of the tumor burden of an in idual patient. Recent advancements have shown that PD-1/PD-L1 immune checkpoint therapies have durable responses in a number of solid tumor types. Our study was designed to use multiple CTC enrichment platforms for the capture of CTCs and novel culture formulations for the ex vivo expansion of CTCs. Head and Neck cancer (n=350) and lung cancer (n=150) patients were recruited to investigate the prognostic role of CTCs. In parallel, a subset of HNC tumors were profiled using the NanoString GeoMx Digital Spatial Profiling (DSP) technology using a 44-plex antibody cocktail. We evaluated multiple CTC isolation technologies (CellSearch, filtration, CD45 depletion, Spiral, Straight and novel microfluidic chip technology) using matched patient s les which showed that epitope-independent CTC isolation captured a greater proportion of CTCs. Molecular alterations present in the primary tissue were confirmed in the CTCs by 3D-DNA FISH (EGFR- lification, ALK-translocations). In HNC, the presence of CTC clusters associated with the development of distant metastatic disease (P=0.0313). HNC CTC-positive patients had shorter progression free survival (Hazard ratio [HR]: 4.946 95% [CI]:1.571-15.57 P=0.0063) and PD-L1-positive CTCs were found to be significantly associated with worse outcome ([HR]:5.159 95% [CI]:1.011-26.33 P=0.0485). In a proof of principle study, we were able to demonstrate for the first time, short-term patient derived CTC cultures outside the patient's body and exome sequencing of CTCs cultures confirmed the presence of mutational signatures consistent with The Cancer Genome Atlas (TCGA). Spatial characterization of the tumor microenvironment revealed immune subsets predictive of outcome to immunotherapy. Comprehensive characterization of the tumor microenvironment and liquid biopsy allows for the recapitulation of the molecular ersity present within the tumor, understanding of the disease progression and testing of therapies. Citation Format: Arutha Kulasinghe, Joanna Kapeleris, Liz Kenny, Brett Hughes, Majid Warkiani, Ian Vela, Jean-Paul Thiery, Ken O'Byrne, Chamindie Punyadeera. Characterization of the tumor microenvironment and liquid biopsy in head and neck and non-small cell lung cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR Cancer Res 2020 (16 Suppl):Abstract nr 3384.
Publisher: Springer US
Date: 2022
DOI: 10.1007/978-1-0716-2341-1_8
Abstract: Exosomes are small extracellular vesicles secreted by cells and are known to play a key role in intercellular communication. Several studies have associated exosomes with various roles in tumorigenesis and explored their potential as a source of biomarkers for diagnosis and prognosis in cancer research. Exosomes can be isolated from several body fluids, including those that are noninvasively accessible, such as human saliva. This book chapter provides a step-by-step detailed description of techniques that are used for the isolation, quantification, and characterization of exosomes from saliva. These include ultracentrifugation for the isolation, nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM), and western blot (WB) for characterization of exosomes. The NTA approach explores the Brownian motion and light scattering of particles to predict size and concentration. TEM enables visualization of the exosomes which often present a cup-shaped morphology. Western blot is used to detect commonly expressed exosome-associated proteins. Finally, salivary exosomes isolated using these protocols can further be characterized for downstream analysis according to their cargo (proteins, DNA, RNA, miRNA) and utilized for cancer biomarker discovery.
Publisher: Elsevier BV
Date: 10-2011
DOI: 10.1016/J.JIM.2011.07.013
Abstract: Cardiovascular disease is the leading cause of death in the world. Human C-reactive protein (CRP) has been used in the risk assessment of coronary events. Human saliva mirrors the body's health and well-being and is non-invasive, easy to collect and ideal for third world countries as well as for large patient screening. The aim was to establish a saliva CRP reference range and to demonstrate the clinical utility of salivary CRP levels in assessing the coronary events in a primary health care setting. We have used a homogeneous bead based assay to detect CRP levels in human saliva. We have developed a rapid 15 min (vs 90 min), sequential, one-step assay to detect CRP in saliva. Saliva was collected from healthy volunteers (n=55, ages 20-70 years) as well as from cardiac patients (n=28, ages 43-86 years). The assay incubation time was optimised from 90 min to 15 min and generated a positive correlation (n=29, range 10-2189 pg/mL, r2=0.94 Passing Bablok slope 0.885, Intercept 0, p>0.10), meaning we could decrease the incubation time and produce equivalent results with confidence. The mean CRP level in the saliva of healthy human volunteers was 285 pg/mL and in cardiac patients was 1680 pg/mL (p<0.01). Analysis of CRP concentrations in paired serum and saliva s les from cardiac patients gave a positive correlation (r2=0.84, p<0.001) and the salivary CRP concentration capable of distinguishing healthy from diseased patients. The results suggest that this minimally invasive, rapid and sensitive assay will be useful in large patient screening studies for risk assessment of coronary events.
Publisher: Ivyspring International Publisher
Date: 2017
DOI: 10.7150/THNO.21727
Publisher: MDPI AG
Date: 14-01-2019
Abstract: Circulating tumour cells (CTCs) are the metastatic precursors to distant disease in head and neck cancers (HNCs). Whilst the prognostic and predictive value of single CTCs have been well documented, the role of CTC clusters, which potentially have a higher metastatic capacity are limited. In this study, the authors used a novel straight microfluidic chip to focus and capture CTCs. The chip offers high cell recoveries with clinically relevant numbers (10–500 cells/mL) without the need for further purification. Single CTCs were identified in 10/21 patient s les (range 2–24 CTCs/mL), CTC clusters in 9/21 patient s les (range 1–6 CTC clusters/mL) and circulating tumour microemboli (CTM) in 2/21 s les. This study demonstrated that CTC clusters contain EGFR lified single CTCs within the cluster volume. This novel microfluidic chip demonstrates the efficient sorting and preservation of single CTCs, CTC clusters and CTMs. The authors intend to expand this study to a larger cohort to determine the clinical implication of the CTC subsets in HNC.
Publisher: MDPI AG
Date: 31-05-2020
DOI: 10.3390/CELLS9061359
Abstract: Disruption of DNA methylation patterns is one of the hallmarks of cancer. Similar to other cancer types, human papillomavirus (HPV)-driven head and neck cancer (HNC) also reveals alterations in its methylation profile. The intrinsic ability of HPV oncoproteins E6 and E7 to interfere with DNA methyltransferase activity contributes to these methylation changes. There are many genes that have been reported to be differentially methylated in HPV-driven HNC. Some of these genes are involved in major cellular pathways, indicating that DNA methylation, at least in certain instances, may contribute to the development and progression of HPV-driven HNC. Furthermore, the HPV genome itself becomes a target of the cellular DNA methylation machinery. Some of these methylation changes appearing in the viral long control region (LCR) may contribute to uncontrolled oncoprotein expression, leading to carcinogenesis. Consistent with these observations, demethylation therapy appears to have significant effects on HPV-driven HNC. This review article comprehensively summarizes DNA methylation changes and their diagnostic and therapeutic indications in HPV-driven HNC.
Publisher: Elsevier BV
Date: 03-2016
Abstract: Aberrant glycosylation of proteins is a hallmark of tumorigenesis and could provide diagnostic value in cancer detection. Human saliva is an ideal source of glycoproteins due to the relatively high proportion of glycosylated proteins in the salivary proteome. Moreover, saliva collection is noninvasive and technically straightforward, and the s le collection and storage is relatively easy. Although differential glycosylation of proteins can be indicative of disease states, identification of differential glycosylation from clinical s les is not trivial. To facilitate salivary glycoprotein biomarker discovery, we optimized a method for differential glycoprotein enrichment from human saliva based on lectin magnetic bead arrays (saLeMBA). Selected lectins from distinct reactivity groups were used in the saLeMBA platform to enrich salivary glycoproteins from healthy volunteer saliva. The technical reproducibility of saLeMBA was analyzed with liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify the glycosylated proteins enriched by each lectin. Our saLeMBA platform enabled robust glycoprotein enrichment in a glycoprotein- and lectin-specific manner consistent with known protein-specific glycan profiles. We demonstrated that saLeMBA is a reliable method to enrich and detect glycoproteins present in human saliva.
Publisher: Springer Science and Business Media LLC
Date: 15-02-2017
DOI: 10.1038/SREP42517
Abstract: Whilst locoregional control of head and neck cancers (HNCs) has improved over the last four decades, long-term survival has remained largely unchanged. A possible reason for this is that the rate of distant metastasis has not changed. Such disseminated disease is reflected in measurable levels of cancer cells in the blood of HNC patients, referred to as circulating tumour cells (CTCs). Numerous marker-independent techniques have been developed for CTC isolation and detection. Recently, microfluidics-based platforms have come to the fore to avoid molecular bias. In this pilot, proof of concept study, we evaluated the use of the spiral microfluidic chip for CTC enrichment and subsequent detection in HNC patients. CTCs were detected in 13/24 (54%) HNC patients, representing both early to late stages of disease. Importantly, in 7/13 CTC-positive patients, CTC clusters were observed. This is the first study to use spiral microfluidics technology for CTC enrichment in HNC.
Publisher: Elsevier BV
Date: 10-2023
Location: Australia
Location: Australia
Start Date: 2020
End Date: 12-2023
Amount: $390,000.00
Funder: Australian Research Council
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