Papillary Thyroid Cancer: Genetic Alterations and Molecular Biomarker Investigations.

Papillary thyroid cancer (PTC) is the most prevalent form of malignancy among all cancers of the thyroid. It is also one of the few cancers with a rapidly increasing incidence. PTC is usually contained within the thyroid gland and generally biologically indolent. Prognosis of the cancer is excellent, with less than 2% mortality at 5 years. However, more than 25% of patients with PTC developed a recurrence during a long term follow-up. The present article provides an updated condensed overview of PTC, which focuses mainly on the molecular alterations involved and recent biomarker investigations.

iTRAQ analysis of urinary proteins: Potential use of gelsolin and osteopontin to distinguish benign thyroid goiter from papillary thyroid carcinoma.

BACKGROUND: Benign thyroid goiter (BTG) and papillary thyroid carcinoma (PTC) are often interchangeably misdiagnosed. METHODS: Pooled urine samples of patients with BTG (n=10), patients with PTC (n=9) and healthy controls (n=10) were subjected to iTRAQ analysis and immunoblotting. RESULTS: The ITRAQ analysis of the urine samples detected 646 proteins, 18 of which showed significant altered levels (p<0.01; fold-change>1.5) between patients and controls. Whilst four urinary proteins were commonly altered in both BTG and PTC patients, 14 were unique to either BTG or PTC. Amongst these, four proteins were further chosen for validation using immunoblotting, and the enhanced levels of osteopontin in BTG patients and increased levels of a truncated gelsolin fragment in PTC patients, relative to controls, appeared to corroborate the findings of the iTRAQ analysis. CONCLUSION: The data of the present study is suggestive of the potential application of urinary osteopontin and gelsolin to discriminate patients with BTG from those with PTC non-invasively. However, this needs to be further validated in studies of individual urine samples.

Lectins: an effective tool for screening of potential cancer biomarkers.

In recent years, the use of lectins for screening of potential biomarkers has gained increased importance in cancer research, given the development in glycobiology that highlights altered structural changes of glycans in cancer associated processes. Lectins, having the properties of recognizing specific carbohydrate moieties of glycoconjugates, have become an effective tool for detection of new cancer biomarkers in complex bodily fluids and tissues. The specificity of lectins provides an added advantage of selecting peptides that are differently glycosylated and aberrantly expressed in cancer patients, many of which are not possibly detected using conventional methods because of their low abundance in bodily fluids. When coupled with mass spectrometry, research utilizing lectins, which are mainly from plants and fungi, has led to identification of numerous potential cancer biomarkers that may be used in the future. This article reviews lectin-based methods that are commonly adopted in cancer biomarker discovery research.

Melicope ptelefolia leaf extracts exhibit antioxidant activity and exert anti-proliferative effect with apoptosis induction on four different cancer cell lines.

BACKGROUND: Melicope ptelefolia is a well-known herb in a number of Asian countries. It is often used as vegetable salad and traditional medicine to address various ailments. However, not many studies have been currently done to evaluate the medicinal benefits of M. ptelefolia (MP). The present study reports antioxidant, anti-proliferative, and apoptosis induction activities of MP leaf extracts. METHOD: Young MP leaves were dried, powdered and extracted sequentially using hexane (HX), ethyl acetate (EA), methanol (MeOH) and water (W). Antioxidant activity was evaluated using ferric reducing antioxidant power (FRAP), 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 1,1-Diphenyl-2-picryl-hydrazyl (DPPH) radicals scavenging and cellular antioxidant activity (CAA) assays. Anti-proliferative activity was evaluated through cell viability assay, using the following four human cancer cell lines: breast (HCC1937, MDA-MB-231), colorectal (HCT116) and liver (HepG2). The anti-proliferative activity was further confirmed through cell cycle and apoptosis assays, including annexin-V/7-aminoactinomycin D staining and measurements of caspase enzymes activation and inhibition. RESULT: Overall, MP-HX extract exhibited the highest antioxidant potential, with IC50 values of 267.73 ± 5.58 and 327.40 ± 3.80 µg/mL for ABTS and DPPH radical-scavenging assays, respectively. MP-HX demonstrated the highest CAA activity in Hs27 cells, with EC50 of 11.30 ± 0.68 µg/mL, while MP-EA showed EC50 value of 37.32 ± 0.68 µg/mL. MP-HX and MP-EA showed promising anti-proliferative activity towards the four cancer cell lines, with IC50 values that were mostly below 100 µg/mL. MP-HX showed the most notable anti-proliferative activity against MDA-MB-231 (IC50 = 57.81 ± 3.49 µg/mL) and HCT116 (IC50 = 58.04 ± 0.96 µg/mL) while MP-EA showed strongest anti-proliferative activity in HCT116 (IC50 = 64.69 ± 0.72 µg/mL). The anticancer potential of MP-HX and MP-EA were also demonstrated by their ability to induce caspase-dependent apoptotic cell death in all of the cancer cell lines tested. Cell cycle analysis suggested that both the MP-HX and MP-EA extracts were able to disrupt the cell cycle in most of the cancer cell lines. CONCLUSIONS: MP-HX and MP-EA extracts demonstrated notable antioxidant, anti-proliferative, apoptosis induction and cancer cell cycle inhibition activities. These findings reflect the promising potentials of MP to be a source of novel phytochemical(s) with health promoting benefits that are also valuable for nutraceutical industry and cancer therapy.

Tissue and serum samples of patients with papillary thyroid cancer with and without benign background demonstrate different altered expression of proteins.

BACKGROUND: Papillary thyroid cancer (PTC) is mainly diagnosed using fine-needle aspiration biopsy. This most common form of well-differentiated thyroid cancer occurs with or without a background of benign thyroid goiter (BTG). METHODS: In the present study, a gel-based proteomics analysis was performed to analyse the expression of proteins in tissue and serum samples of PTC patients with (PTCb; n = 6) and without a history of BTG (PTCa; n = 8) relative to patients with BTG (n = 20). This was followed by confirmation of the levels of proteins which showed significant altered abundances of more than two-fold difference (p < 0.01) in the tissue and serum samples of the same subjects using ELISA. RESULTS: The data of our study showed that PTCa and PTCb distinguish themselves from BTG in the types of tissue and serum proteins of altered abundance. While higher levels of alpha-1 antitrypsin (A1AT) and heat shock 70 kDa protein were associated with PTCa, lower levels of A1AT, protein disulfide isomerase and ubiquitin-conjugating enzyme E2 N seemed apparent in the PTCb. In case of the serum proteins, higher abundances of A1AT and alpha 1-beta glycoprotein were detected in PTCa, while PTCb was associated with enhanced apolipoprotein A-IV and alpha 2-HS glycoprotein (AHSG). The different altered expression of tissue and serum A1AT as well as serum AHSG between PTCa and PTCb patients were also validated by ELISA. DISCUSSION: The distinctive altered abundances of the tissue and serum proteins form preliminary indications that PTCa and PTCb are two distinct cancers of the thyroid that are etiologically and mechanistically different although it is currently not possible to rule out that they may also be due other reasons such as the different stages of the malignant disease. These proteins stand to have a potential use as tissue or serum biomarkers to discriminate the three different thyroid neoplasms although this requires further validation in clinically representative populations.

Contrasting increased levels of serum amyloid A in patients with three different bone sarcomas: An indicator of tumor malignancy?

Sarcoma is a malignant tumor that originates from the bone or soft tissue. In this study, abundances of serum amyloid A (SAA) in patients with pleomorphic sarcoma (PS), chondrosarcoma (CS), and osteosarcoma (OS) were analyzed and compared with those from their respective age-matched healthy control subjects. Results obtained from our analysis by 2DE showed that the levels of SAA were markedly elevated in patients with PS and OS, which are highly metastatic, while in patients with CS, which is a less aggressive sarcoma, the increase appeared less pronounced. A similar trend of altered abundances was also observed when the levels of SAA in the subjects were estimated using Western blot, ELISA, and multiple-reaction monitoring analyses. Absolute quantification using multiple-reaction monitoring further demonstrated that the increased abundance of SAA in patients with PS, OS, and CS was mainly attributed to isoform SAA1. In view of the different degrees of tumor malignancy in PS, OS, and CS, our data suggest their apparent correlation with the levels of SAA in the patients.

Unmasking Heavily O-Glycosylated Serum Proteins Using Perchloric Acid: Identification of Serum Proteoglycan 4 and Protease C1 Inhibitor as Molecular Indicators for Screening of Breast Cancer.

Heavily glycosylated mucin glycopeptides such as CA 27.29 and CA 15-3 are currently being used as biomarkers for detection and monitoring of breast cancer. However, they are not well detected at the early stages of the cancer. In the present study, perchloric acid (PCA) was used to enhance detection of mucin-type O-glycosylated proteins in the serum in an attempt to identify new biomarkers for early stage breast cancer. Sensitivity and specificity of an earlier developed sandwich enzyme-linked lectin assay were significantly improved with the use of serum PCA isolates. When a pilot case-control study was performed using the serum PCA isolates of normal participants (n = 105) and patients with stage 0 (n = 31) and stage I (n = 48) breast cancer, higher levels of total O-glycosylated proteins in sera of both groups of early stage breast cancer patients compared to the normal control women were demonstrated. Further analysis by gel-based proteomics detected significant inverse altered abundance of proteoglycan 4 and plasma protease C1 inhibitor in both the early stages of breast cancer patients compared to the controls. Our data suggests that the ratio of serum proteoglycan 4 to protease C1 inhibitor may be used for screening of early breast cancer although this requires further validation in clinically representative populations.

Application of SELDI-TOF in N-glycopeptides profiling of the urine from patients with endometrial, ovarian and cervical cancer.

PURPOSE: Endometrial (ECa), ovarian (OCa) and cervical (CCa) cancers are among 10 of the most common cancers affecting women worldwide. Cancers are known to cause some proteins to be differentially glycosylated or aberrantly excreted in the urine, which can be used as biomarkers. Since ECa, OCa and CCa are difficult to diagnose at the early stage, the aim of the present study was to identify a panel of new biomarkers for early detection of the cancers using surface-enhanced laser desorption/ionization-time-of-flight (SELDI-TOF) technology. Identification of early biomarkers that are specific and efficient can increase the survival rate of the patients. EXPERIMENTAL DESIGN: Digested urinary proteins from patients with ECa, OCa and CCa were incubated on the champedak mannose-binding (CMB) lectin-immobilized PS10 chip. The lectin-captured glycopeptides were detected with SELDI-TOF mass spectrometry and followed by biomarker wizard analysis. RESULTS: Peaks m/z 1201 and 1449 were detected as potential group discriminators. The peak m/z 1201 could distinguish OCa from CCa and ECa and its sensitivity and specificity were 100%. For m/z 1449, it was able to differentiate ECa from the other two types of cancer. CONCLUSIONS: The findings of this study suggest urinary glycopeptides m/z 1201 and 1449 may serve as potential biomarkers for the early detection of ECa, OCa and CCa, although this requires further extensive validation on clinically representative populations.

Chronotherapeutic effect of fisetin on expression of urea cycle enzymes and inflammatory markers in hyperammonaemic rats.

BACKGROUND: Elevated blood ammonia leads to hyperammonaemia that affects vital central nervous system (CNS) functions. Fisetin, a naturally occurring flavonoid, exhibits therapeutic benefits, such as anti-cancer, anti-diabetic, anti-oxidant, anti-angiogenic, neuroprotective and neurotrophic effects. METHODS: In this study, the chronotherapeutic effect of fisetin on ammonium chloride (AC)-induced hyperammonaemic rats was investigated, to ascertain the time point at which the maximum drug effect is achieved. The anti-hyperammonaemic potential of fisetin (50mg/kg b.w. oral) was analysed when administered to AC treated (100mg/kg b.w. i.p.) rats at 06:00, 12:00, 18:00 and 00:00h. Amelioration of pathophysiological conditions by fisetin at different time points was measured by analysing the levels of expression of liver urea cycle enzymes (carbamoyl phosphate synthetase-I (CPS-I), ornithine transcarbamoylase (OTC) and argininosuccinate synthetase (ASS)), nuclear transcription factor kappaB (NF-κB p65), brain glutamine synthetase (GS) and inducible nitric oxide synthase (iNOS) by Western blot analysis. RESULTS: Fisetin increased the expression of CPS-I, OTC, ASS and GS and decreased iNOS and NF-κB p65 in hyperammonaemic rats. Fisetin administration at 00:00h showed more significant effects on the expression of liver and brain markers, compared with other time points. CONCLUSIONS: Fisetin could exhibit anti-hyperammonaemic effect owing to its anti-oxidant and cytoprotective influences. The temporal variation in the effect of fisetin could be due to the (i) chronopharmacological, chronopharmacokinetic properties of fisetin and (ii) modulations in the endogenous circadian rhythms of urea cycle enzymes, brain markers, redox enzymes and renal clearance during hyperammonaemia by fisetin. However, future studies in these lines are necessitated.

Aberrant proteins featured in the saliva of habitual betel quid chewers: an indication of early oral premalignancy?

Prolonged chewing of betel quid is known to cause oral diseases, including cancer. The present study was performed to screen for aberrant proteins in the saliva of habitual betel quid chewers compared to nonchewers. Saliva of female subjects (n = 10) who had been chewing betel quid for more than 20 years and nonbetel quid chewers (n = 10) of the same gender and range of age was analyzed by gel-based proteomics. Increased structural microheterogeneity of saliva haptoglobin beta chains indicated by shifts of focused spots similar to that earlier reported in patients with oral squamous cell carcinoma, and their relatively higher abundance compared to nonbetel quid chewers, were detected in saliva protein profiles of all chewers. In addition, the majority of the betel quid chewers also showed significant higher abundance of hemopexin, alpha-1B glycoprotein, alpha1-antitrypsin, complement C3, and transthyretin. These proteins had previously been associated with several different cancers. Our data demonstrated different forms of protein aberration in the saliva of betel quid chewers, which may be indicative of early oral precancerous conditions.

Role of Bacopa monnieri in the temporal regulation of oxidative stress in clock mutant (cryb) of Drosophila melanogaster.

Accruing evidences imply that circadian organization of biochemical, endocrinological, cellular and physiological processes contribute to wellness of organisms and in the development of pathologies such as malignancy, sleep and endocrine disorders. Oxidative stress is known to mediate a number of diseases and it is notable to comprehend the orchestration of circadian clock of a model organism of circadian biology, Drosophila melanogaster, under oxidative stress. We investigated the nexus between circadian clock and oxidative stress susceptibility by exposing D. melanogaster to hydrogen peroxide (H2O2) or rotenone; the reversibility of rhythms following exposure to Bacopa monnieri extract (ayurvedic medicine rich in antioxidants) was also investigated. Abolishment of 24h rhythms in physiological response (negative geotaxis), oxidative stress markers (protein carbonyl and thiobarbituric acid reactive substances) and antioxidants (superoxide dismutase, catalase, glutathione-S-transferase and reduced glutathione) were observed under oxidative stress. Furthermore, abolishment of per mRNA rhythm in H2O2 treated wild type flies and augmented susceptibility to oxidative stress in clock mutant (cry(b)) flies connotes the role of circadian clock in reactive oxygen species (ROS) homeostasis. Significant reversibility of rhythms was noted following B. monnieri treatment in wild type flies than cry(b) flies. Our experimental approach revealed a relationship involving oxidative stress and circadian clock in fruit fly and the utility of Drosophila model in screening putative antioxidative phytomedicines prior to their use in mammalian systems.

Identification of proteins of altered abundance in oil palm infected with Ganoderma boninense.

Basal stem rot is a common disease that affects oil palm, causing loss of yield and finally killing the trees. The disease, caused by fungus Ganoderma boninense, devastates thousands of hectares of oil palm plantings in Southeast Asia every year. In the present study, root proteins of healthy oil palm seedlings, and those infected with G. boninense, were analyzed by 2-dimensional gel electrophoresis (2-DE). When the 2-DE profiles were analyzed for proteins, which exhibit consistent significant change of abundance upon infection with G. boninense, 21 passed our screening criteria. Subsequent analyses by mass spectrometry and database search identified caffeoyl-CoA O-methyltransferase, caffeic acid O-methyltransferase, enolase, fructokinase, cysteine synthase, malate dehydrogenase, and ATP synthase as among proteins of which abundances were markedly altered.

Effect of melatonin on antioxidant status and circadian activity rhythm during hepatocarcinogenesis in mice.

AIM: Alteration of circadian systems can cause cancer and affects its development and response to therapeutics. The present study investigates whether cancer can disrupt circadian locomotor rhythms and evaluated the influence of melatonin (MLT) and oxaliplatin on the levels of antioxidants and circadian locomotor activity rhythms in N-nitrosodiethylamine (NDEA)-induced liver tumor in Indian field mouse (Mus booduga). MATERIALS AND METHODS: Effects of NDEA, NDEA, and MLT, as well as NDEA and oxaliplatin, on levels of mice liver marker enzymes and antioxidants and their circadian locomotor activity rhythm were assessed. RESULTS: Treatment of mice with NDEA caused significant alteration of their liver marker enzymes [aspartate transaminase and alanine transaminase; P<0.05 Duncan's multiple range test (DMRT) test] antioxidant levels (superoxide dismutase, catalase, glutathione peroxidase, glutathione-S-transferase; P<0.05, DMRT test] and circadian locomotor activity rhythm, which were abrogated when the animals were also given MLT or the anticancer drug, oxaliplatin. CONCLUSION: Our study demonstrated that the circadian clock was disturbed by hepatocarcinogenesis and the effects could be reversed by the chronobiotic, MLT.

Enhanced expression of a 35 kDa fragment of inter-alpha-trypsin inhibitor H4 in sera of healthy pregnant women and patients with hydatidiform mole.

BACKGROUND: Accumulated data from previous studies appear to suggest a link between the overexpression of a 35 kDa fragment of serum inter-alpha-trypsin inhibitor H4 (ITIH4) with cancers that are associated with up-regulated levels of oestrogens. The truncated fragment was postulated to be a product of oestrogen-induced action of kallikrein on native ITIH4. The present lectin-based proteomic analyses were performed to assess the specificity of the 35 kDa fragment of ITIH4 as a potential cancer biomarker and determine whether it was also overexpressed in the sera of cancer-negative pregnant women who are known to have high levels of plasma oestrogens. RESULTS: Our results demonstrated that the 35 kDa fragment of ITIH4 was overexpressed in healthy pregnant women and patients with hydatidiform mole, relative to the controls. The serum oestradiol levels of both groups of pregnant subjects were also confirmed to be higher than those of the control women who were not pregnant. CONCLUSIONS: Overexpression of the 35 kDa fragment of ITIH4 was not restrictive to patients with cancers but also occurred in women who were pregnant and those diagnosed with hydatidiform mole. Our data implicate the limitation of the 35 kDa ITIH4 fragment as a cancer biomarker and its correlation with serum oestrogen levels.

Infection of Burkholderia cepacia induces homeostatic responses in the host for their prolonged survival: the microarray perspective.

Burkholderia cepacia is an opportunistic human pathogen associated with life-threatening pulmonary infections in immunocompromised individuals. Pathogenesis of B. cepacia infection involves adherence, colonisation, invasion, survival and persistence in the host. In addition, B. cepacia are also known to secrete factors, which are associated with virulence in the pathogenesis of the infection. In this study, the host factor that may be the cause of the infection was elucidated in human epithelial cell line, A549, that was exposed to live B. cepacia (mid-log phase) and its secretory proteins (mid-log and early-stationary phases) using the Illumina Human Ref-8 microarray platform. The non-infection A549 cells were used as a control. Expression of the host genes that are related to apoptosis, inflammation and cell cycle as well as metabolic pathways were differentially regulated during the infection. Apoptosis of the host cells and secretion of pro-inflammatory cytokines were found to be inhibited by both live B. cepacia and its secretory proteins. In contrast, the host cell cycle and metabolic processes, particularly glycolysis/glycogenesis and fatty acid metabolism were transcriptionally up-regulated during the infection. Our microarray analysis provided preliminary insights into mechanisms of B. cepacia pathogenesis. The understanding of host response to an infection would provide novel therapeutic targets both for enhancing the host's defences and repressing detrimental responses induced by the invading pathogen.

Aberrant proteins in the saliva of patients with oral squamous cell carcinoma.

Confirmation of oral squamous cell cancer (OSCC) currently relies on histological analysis, which does not provide clear indication of cancer development from precancerous lesions. In the present study, whole saliva proteins of patients with OSCC (n = 12) and healthy subjects (n = 12) were separated by 2DE to identify potential candidate biomarkers that are much needed to improve detection of the cancer. The OSCC patients' 2DE saliva protein profiles appeared unique and different from those obtained from the healthy subjects. The patients' saliva α1-antitrypsin (AAT) and haptoglobin (HAP) ß chains were resolved into polypeptide spots with increased microheterogeneity, although these were not apparent in their sera. Their 2DE protein profiles also showed presence of hemopexin and α-1B glycoprotein, which were not detected in the profiles of the control saliva. When subjected to densitometry analysis, significant altered levels of AAT, complement C3, transferrin, transthyretin, and ß chains of fibrinogen and HAP were detected. The increased levels of saliva AAT, HAP, complement C3, hemopexin, and transthyretin in the OSCC patients were validated by ELISA. The strong association of AAT and HAP with OSCC was further supported by immunohistochemical staining of cancer tissues. The differently expressed saliva proteins may be useful complementary biomarkers for the early detection and/or monitoring of OSCC, although this requires validation in clinically representative populations.

An improved lectin-based method for the detection of mucin-type O-glycans in biological samples.

Mucins and mucin-type glycoproteins, collectively referred to as mucin-type O-glycans, are implicated in many important biological functions and pathological conditions, including malignancy. Presently, there is no reliable method to measure the total mucin-type O-glycans of a sample, which may contain one or more of these macromolecules of unknown structures. We report the development of an improved microassay that is based on the binding of lectins to the unique and constant GalNAc-Ser/Thr structural feature of mucin-type O-glycans. Since the sugar-amino acid linkage in the mucin-type O-glycans is invariably cryptic, we first chemically removed the heterogeneous peripheral and core saccharides of model glycoconjugates before examining for their interactions using an enzyme-linked lectin assay (ELLA). Desialylation of the model glycoconjugates led to maximal binding of the lectins but additional treatments such as Smith degradation did not result in increased binding. Of the lectins tested for their ability to probe the desialylated O-glycans, jacalin showed the highest sensitivity followed by champedak galactose binding (CGB) lectin and Vicia villosa agglutinin. Further improvement in the sensitivity of ELLA was achieved by using microtiter plates that were pre-coated with the CGB lectin, which increased the specificity of the assay to mucin-type O-glycans. Finally, the applicability of the developed sandwich ELLA to crude samples was demonstrated by estimating trace quantities of the mucin-type O-glycans in the human serum.

Gastroprotective mechanism of Bauhinia thonningii Schum.

ETHNOPHARMACOLOGICAL RELEVANCE: Bauhinia thonningii Schum. (Cesalpiniaceae) is locally known as Tambarib and used to treat various diseases including gastric ulcer. AIM OF THE STUDY: The current study aims to evaluate the gastroprotecive mechanism(s) of methanolic (MEBT) and chloroform (CEBT) extracts of Bauhinia thonningii leaves on ethanol-induced gastric ulceration. MATERIALS AND METHODS: Gastric acidity, quantification and histochemistry of mucus, gross and microscopic examination, nitric oxide, lipid peroxidation, 2D gel electrophoresis, mass spectroscopy and biochemical tests were utilized to assess the mechanism(s) underlying the gastroprotective effects of MEBT and CEBT. Effect of these extracts into lipopolysaccharide/interferon-γ stimulated rodent cells were done in vitro. In vitro and in vivo toxicity studies were also conducted. Antioxidant activities of MEBT and CEBT were examined using DPPH, FRAP and ORAC assays. Phytochemical analyses of MEBT and CEBT were conducted using chemical and spectroscopic methods. RESULTS: Gross and histological features confirmed the anti-ulcerogenic properties of Bauhinia thonningii. Gastroprotective mechanism of MEBT was observed to be mediated through the modulation of PAS-reactive substances, MDA and proteomics biomarkers (creatine kinase, malate dehydrogenase, ATP synthase, actin and thioredoxin). MEBT and CEBT showed no significant in vitro and in vivo effects on nitric oxide. Methanolic extract (MEBT) showed superior gastroprotective effects, polyphenolic content and antioxidant activities compared to CEBT. The plant extracts showed no in vitro or in vivo toxicity. CONCLUSION: It could be concluded that MEBT possesses anti-ulcer activity, which could be attributed to the inhibition of ethanol-induced oxidative damage and the intervention in proteomic pathways but not the nitric oxide pathway.

Identification of O-glycosylated proteins that are aberrantly excreted in the urine of patients with early stage ovarian cancer.

Cancer is known to induce or alter the O-glycosylation of selective proteins that may eventually be excreted in the patients' urine. The present study was performed to identify O-glycosylated proteins that are aberrantly excreted in the urine of patients with early stage ovarian cancer (OCa). These urinary glycoproteins are potential biomarkers for early detection of OCa. In this study, urinary proteins of patients with early stage OCa and age-matched OCa negative women were subjected to two-dimensional gel electrophoresis and detection using a lectin that binds to the O-glycosylated proteins. Our analysis demonstrated significant enhanced expression of clusterin and leucine-rich alpha-2-glycoprotein, but lower levels of kininogen in the urine of the OCa patients compared to the controls. The different altered levels of these urinary glycoproteins were further confirmed using competitive ELISA. Our data are suggestive of the potential use of the aberrantly excreted urinary O-glycosylated proteins as biomarkers for the early detection of OCa, although this requires further validation in a large clinically representative population.

Detection of differential levels of proteins in the urine of patients with endometrial cancer: analysis using two-dimensional gel electrophoresis and o-glycan binding lectin.

Cancers can cause some proteins to be aberrantly excreted or released in the urine, which can be used as biomarkers. To screen for potential biomarkers for endometrial cancer (ECa), the urinary proteins from patients who were newly diagnosed with early stage ECa and untreated controls were separated using two-dimensional gel electrophoresis (2-DE) and followed by image analysis. The altered levels of zinc alpha-2 glycoprotein, alpha 1-acid glycoprotein, and CD59 were detected in the patients compared to the controls. In addition, the urine of the ECa patients was also found to contain relatively lower levels of a fragment of nebulin when the 2-DE separated urinary proteins were probed using champedak galactose binding (CGB) lectin. The different levels of the nebulin fragment were further validated by subjecting the urinary protein samples to CGB lectin affinity chromatography and analysis of the bound fractions by LC-MS/MS. Our data is suggestive of the potential use of the differentially expressed urinary proteins as biomarkers for ECa although this requires further extensive validation on clinically representative populations.