Profilin 1 is a potential biomarker for bladder cancer aggressiveness

Of the most important clinical needs for bladder cancer (BC) management is the identification of biomarkers for disease aggressiveness. Urine is a "gold mine" for biomarker discovery, nevertheless, with multiple proteins being in low amounts, urine proteomics becomes challenging. In the present study we applied a fractionation strategy of urinary proteins based on the use of immobilized metal affinity chromatography for the discovery of biomarkers for aggressive BC. Urine samples from patients with non invasive (two pools) and invasive (two pools) BC were subjected to immobilized metal affinity chromatography fractionation and eluted proteins analyzed by 1D-SDS-PAGE, band excision and liquid chromatography tandem MS. Among the identified proteins, multiple corresponded to proteins with affinity for metals and/or reported to be phosphorylated and included proteins with demonstrated association with BC such as MMP9, fibrinogen forms, and clusterin. In agreement to the immobilized metal affinity chromatography results, aminopeptidase N, profilin 1, and myeloblastin were further found to be differentially expressed in urine from patients with invasive compared with non invasive BC and benign controls, by Western blot or Elisa analysis, nevertheless exhibiting high interindividual variability. By tissue microarray analysis, profilin 1 was found to have a marked decrease of expression in the epithelial cells of the invasive (T2+) versus high risk non invasive (T1G3) tumors with occasional expression in stroma; importantly, this pattern strongly correlated with poor prognosis and increased mortality. The functional relevance of profilin 1 was investigated in the T24 BC cells where blockage of the protein by the use of antibodies resulted in decreased cell motility with concomitant decrease in actin polymerization. Collectively, our study involves the application of a fractionation method of urinary proteins and as one main result of this analysis reveals the association of profilin 1 with BC paving the way for its further investigation in BC stratification.     

lncRNA TUC338 is a potential diagnostic biomarker for bladder cancer

Although long noncoding RNA TUC338 has been characterized as an oncogene, its role in bladder cancer is unknown. The purpose of the present study is to investigate the function of TUC338 in bladder cancer. We found that TUC338 was upregulated in early-stage bladder cancer patients and showed early diagnostic values. After surgical resection, plasma levels of TUC338 were significantly downregulated. Moreover, microRNA 10b (miR-10b) was also upregulated in bladder cancer patients. TUC338 and miR-10b were positive and significantly correlated in bladder cancer patients, but not in healthy controls. Bladder cancer cells with TUC338 overexpression showed upregulated miR-10b, while miR-10b overexpression failed to significantly affect TUC338. TUC338 and miR-10b overexpression significantly promoted bladder cancer cell invasion and migration. Therefore, TUC338 may promote bladder cancer at least partially by upregulating miR-10b.     

Assessment of DNA flow cytometry as a surrogate end point biomarker in a bladder cancer chemoprevention trial

Although conventional cytology represents the most widely performed cytometric analysis of bladder cancer cells, DNA flow cytometry has, over the past decade, been increasingly used to evaluate cell proliferation and DNA ploidy in cells from bladder washings. We have investigated whether DNA flow cytometry and conventional cytology of epithelial cells obtained from bladder washings provide reliable surrogate endpoint biomarkers in clinical chemoprevention trials. We used cytometric and clinical data from a chemoprevention trial of the synthetic retinoid Fenretinide on 99 patients with superficial bladder cancer. A total of 642 bladder washing specimens obtained from the patients at 4 month intervals was analyzed. Intra-individual agreement and correlation of flow cytometric DNA ploidy (diploid vs. aneuploid), DNA Index, Hyper-Diploid-Fraction (proportion of cells with DNA content higher than 2C), and conventional cytologic examination, as assessed by kappa statistics and Spearman's correlation test, were poor from baseline through 24 months. Moreover, no correlation was found between DNA ploidy and cytology at each time point. The same results were obtained when the analyses were stratified by treatment group. In addition, the association between the results of bladder washing (by either DNA flow cytometry or cytology) and concomitant tumor recurrence was significant only for abnormal cytology, while neither biomarker was predictive of tumor recurrence at the subsequent visit. During the time of this study only four patients progressed to muscle-invasive bladder cancer, indicating the "low-risk" features of the patient population. We conclude that DNA flow cytometry and conventional cytology on epithelial cells obtained from bladder washings do not appear to provide suitable surrogate endpoint biomarkers during the early stages of bladder carcinogenesis.     

Carcinogen-DNA adducts as a biomarker for cancer risk

Carcinogen-DNA adducts are thought to be a useful biomarker in epidemiologic studies seeking to show that environmental exposures to xenobiotics cause cancer. This paper reviews the literature in this field from an epidemiologic perspective and identifies several common problems in the epidemiologic design and analysis of these studies. Carcinogen-DNA adducts have been used in studies attempting to link xenobiotic exposures to hepatocellular carcinoma, smoking related cancers and breast cancer. Adduct measurements have been useful in further implicating aflatoxin exposure in the etiology of hepatocellular carcinoma. For smoking related cancers, associations with carcinogen-DNA adducts are commonly seen in current smokers but less so in ex- or non-smokers. In breast cancer the associations have been inconsistent and weak and there is little evidence that carcinogen-DNA adducts implicate xenobiotic exposures in the etiology of breast cancer. Methodological issues common to these studies are the use of target versus surrogate tissues and how this choice impacts control selection, disease effects on adduct levels, the time period reflected by adduct levels, the use of inappropriate statistical analyses and small sample sizes. It is unclear whether the lack of association between carcinogen-DNA adducts and cancer reflects a lack of association between the xenobiotic exposure of interest and cancer or the effects of these methodological issues. A greater focus needs to be placed on designs that allow measurements of adduct levels in tissues collected years prior to cancer diagnosis, there is little need for further hospital based case-control studies in which adducts are measured at the time of or after diagnosis. New designs that address these issues are suggested in the paper.     

p16INK4a and p14ARF methylation as a potential biomarker for human bladder cancer

Promoter hypermethylation is one of the putative mechanisms underlying the inactivation of negative cell-cycle regulators. We examined whether the methylation status of p16(INK4a) and p14(ARF), genes located upstream of the RB and p53 pathway, is a useful biomarker for the staging, clinical outcome, and prognosis of human bladder cancer. Using methylation-specific PCR (MSP), we examined the methylation status of p16(INK4a) and p14(ARF) in 64 samples from 45 bladder cancer patients (34 males, 11 females). In 19 patients with recurrent bladder cancer, we examined paired tissue samples from their primary and recurrent tumors. The methylation status of representative samples was confirmed by bisulfite DNA sequencing analysis. The median follow-up duration was 34.3 months (range 27.0-100.1 months). The methylation rate for p16(INK4a) and p14(ARF) was 17.8% and 31.1%, respectively, in the 45 patients. The incidence of p16(INKa) and p14(ARF) methylation was significantly higher in patients with invasive (>or=pT2) than superficial bladder cancer (pT1) (p=0.006 and p=0.001, respectively). No MSP bands for p16(INK4a) and p14(ARF) were detected in the 8 patients with superficial, non-recurrent tumors. In 19 patients with tumor recurrence, the p16(INK4a) and p14(ARF) methylation status of the primary and recurrent tumors was similar. Of the 22 patients who had undergone cystectomy, 8 (36.4%) manifested p16(INKa) methylation; p16(INK4a) was not methylated in 23 patients without cystectomy (p=0.002). Kaplan-Meier analysis revealed that patients with p14(ARF) methylation had a significantly poorer prognosis than those without (p=0.029). This is the first study indicating that MSP analysis of p16(INK4a) and p14(ARF) genes is a useful biomarker for the pathological stage, clinical outcome, and prognosis of patients with bladder cancer.     

Discovery of Apo-A1 as a potential bladder cancer biomarker by urine proteomics and analysis

Bladder cancer is clinically characterized by high recurrent rate and poor prognosis and thereby patients need regular re-examinations which are invasive, unpleasant, and expensive. A noninvasive and less expensive method for detecting and monitoring bladder cancer would thus be advantageous. In this study, by using the two-dimensional electrophoresis (2-DE) approach with subsequent mass spectrometry (MS), we demonstrated the increased expression of apolipoprotein-A1 (Apo-A1) in individual urine from patients with bladder cancer, which was confirmed by Western blot results. A further analysis of the urinary Apo-A1 levels by an enzyme-linked immunosorbent assay yielded results that were consistent with the Western blot, and suggested Apo-A1 could provide diagnostic utility to distinguish patients with bladder cancer from healthy controls at 19.21 ng/ml. Further validation assay in a larger number of urine samples (n = 379) showed that Apo-A1 could be used as a biomarker to diagnosis bladder cancer with a sensitivity and specificity of 89.2% and 84.6% respectively. Moreover, the application of exfoliative urinary cytology in combination with the urine Apo-A1 detection could significantly increased the sensitivity in detecting bladder cancer. Our data showed a significant relationship of expressed Apo-A1 was established between bladder cancer and normal controls. Apo-A1 could be a potential biomarker for the diagnosis of bladder cancer.     

pHLIP and acidity as a universal biomarker for cancer

Of great importance to clinical cancer diagnosis is the use of organic biomarkers. The detection of RNA, DNA, and protein antigen are all established methods for identifying specific cancer types and instrumental in promoting greater survivorship of the patient. Despite many decades of intense cancer research, we have yet to identify a "universal" protein or nucleic acid that allows us to diagnose more than a small subset of cancers at a time. In this review, we examine the use of localized cellular acidity as a universal marker for solid tumors, outlining some successes with a small peptide we call pHLIP, a pH-sensitive biosensor that allows us to label tumor tissue in live mice.     

Calreticulin as a potential diagnostic biomarker for lung cancer

Calreticulin (CRT) is an endoplasmic reticulum luminal Ca(2+)-binding chaperone protein. By immunizing mice with recombinant fragment (rCRT/39-272), six clones of monoclonal antibodies (mAbs) were generated and characterized. Based on these mAbs, a microplate chemiluminescent enzyme immunoassay (CLEIA) system with a measured limit of detection of 0.09?ng/ml was developed. Using this CLEIA system, it was found that soluble CRT (sCRT) level in serum samples from 58 lung cancer patients was significantly higher than that from 40 healthy individuals (only 9 were detectable, P?相似文献   

The survivin molecule as a double-edged sword in cellular physiologic and pathologic conditions and its role as a potential biomarker and therapeutic target in cancer

Survivin is a member of the family of apoptosis inhibitory proteins with increased expression level in most cancerous tissues. Evidence shows that survivin plays regulatory roles in proliferation or survival of normal adult cells, principally vascular endothelial cells, T lymphocytes, primitive hematopoietic cells, and polymorphonuclear neutrophils. Survivin antiapoptotic role is, directly and indirectly, related to caspase proteins and shows its role in cell division through the chromosomal passenger complex. Survivin contains many genetic polymorphisms that the role of some variations has been proven in several cancers. The −31G/C polymorphism is one of the most important survivin mutations which is located in the promoter region on a CDE/CHR motif. This polymorphism can upregulate the survivin messenger RNA. In addition, its allele C can increase the risk of cancers in 1.27-fold than allele G. Considering the fundamental role of survivin in different cancers, this protein could be considered as a new therapeutic target in cancer treatment. For this purpose, various strategies have been designed including the prevention of survivin expression through inhibition of mRNA translation using antagonistic molecules, inhibition of survivin gene function through small inhibitory molecules, gene therapy, and immunotherapy. In this study, we describe the structure, played roles in physiological and pathological states and genetic polymorphisms of survivin. Finally, the role of survivin as a potential target in cancer therapy given challenges ahead has been discussed.     

Metallothionein as a prognostic biomarker in breast cancer

Breast cancer is the most common cancer in women, with a general upward trend in incidence. Basic and clinical breast cancer research has continued at a rapid pace, in the endeavor to understand the biology of the disease so as to improve management of patients. Besides traditional pathological indicators, expression of molecular markers in breast cancer has also been comprehensively investigated. This paper will focus on the prognostic utility of metallothioneins (MTs), a family of low molecular weight metal binding proteins encoded by at least 10 functional MT genes that are associated with cell proliferation in breast cancer. Evidence that MT is a potential prognostic biomarker for breast cancer is supported by many reports in the literature. Expression of the MT protein has been detected by immunohistochemistry in a significant portion of invasive ductal breast cancers. MT expression has also been well studied in association with traditional clinico-pathological parameters of breast cancers. Generally, higher MT expression in breast cancers is predictive of worse patient outcomes. The relationship of MT isoforms to histological grade, estrogen receptor (ER) status, and prognosis will also be discussed.     

N-glycolylneuraminic acid as a carbohydrate cancer biomarker

One of the forms of aberrant glycosylation in human tumors is the expression of N-glycolylneuraminic acid (Neu5Gc). The only known enzyme to biosynthesize Neu5Gc in mammals, cytidine-5′-monophosphate-N-acetylneuraminic acid (CMAH), appears to be genetically inactivated in humans. Regardless, low levels of Neu5Gc have been detected in healthy humans. Therefore, it is proposed that the presence of Neu5Gc in humans is from dietary acquisition, such as red meat. Notably, detection of elevated Neu5Gc levels has been repeatedly found in cancer tissues, cells and serum samples, thereby Neu5Gc-containing antigens may be exploited as a class of cancer biomarkers. Here we review the findings to date on using Neu5Gc-containing tumor glycoconjugates as a class of cancer biomarkers for cancer detection, surveillance, prognosis and therapeutic targets. We review the evidence that supports an emerging hypothesis of de novo Neu5Gc biosynthesis in human cancer cells as a source of Neu5Gc in human tumors, generated under certain metabolic conditions.     

Proteomic analysis of urinary biomarker candidates for nonmuscle invasive bladder cancer

Nonmuscle invasive tumors of the bladder often recur and thereby bladder cancer patients need regular re-examinations which are invasive, unpleasant, and expensive. A noninvasive and less expensive method, e.g. a urine dipstick test, for monitoring recurrence would thus be advantageous. In this study, the complementary techniques mass spectrometry (MS) and Western blotting (WB)/dot blot (DB) were used to screen the urine samples from bladder cancer patients. High resolving MS was used to analyze and quantify the urinary proteome and 29 proteins had a significantly higher abundance (p<0.05) in bladder cancer samples compared with control urine samples. The increased abundance found in urine from bladder cancer patients compared with controls was confirmed with Western blot for four selected proteins; fibrinogen β chain precursor, apolipoprotein E, α-1-antitrypsin, and leucine-rich α-2-glycoprotein 1. Dot blot analysis of an independent urine sample set pointed out fibrinogen β chain and α-1-antitrypsin as most interesting biomarkers having sensitivity and specificity values in the range of 66-85%. Exploring the Human Protein Atlas (HPA) also revealed that bladder cancer tumors are the likely source of these proteins. They have the potential of being useful in diagnosis, monitoring of recurrence and thus may improve the treatment of bladder tumors, especially nonmuscle invasive tumors.     

Humoral response to cancer as a tool for biomarker discovery

There is an important need to find relevant biomarkers that show high sensitivity and specificity for early diagnosis and prognosis of cancer. An immune response to cancer is elicited in humans, as demonstrated in part by the identification of autoantibodies against a number of tumor-associated antigens in sera from patients with different types of cancer. Identification of tumor-associated antigens and their cognate autoantibodies is a promising strategy for the discovery of relevant biomarkers. During the past few years, proteomic approaches, including SEREX, SERPA and, more recently, protein microarrays, have been the dominant strategies used to identify tumor-associated antigens and their cognate autoantibodies. In this review, we aim to describe advantages, drawbacks, and recent improvements of these approaches for the study of humoral responses.     

Identification of NCAPH as a biomarker for prognosis of breast cancer

Molecular Biology Reports - Non-SMC condensin I complex subunit H (NCAPH) is a structural component of chromosomes during mitosis, which up-regulates in various cancers. However, the role of NCAPH...     

Multiplex targeted proteomic assay for biomarker detection in plasma: a pancreatic cancer biomarker case study

Biomarkers are most frequently proteins that are measured in the blood. Their development largely relies on antibody creation to test the protein candidate performance in blood samples of diseased versus nondiseased patients. The creation of such antibody assays has been a bottleneck in biomarker progress due to the cost, extensive time, and effort required to complete the task. Targeted proteomics is an emerging technology that is playing an increasingly important role to facilitate disease biomarker development. In this study, we applied a SRM-based targeted proteomics platform to directly detect candidate biomarker proteins in plasma to evaluate their clinical utility for pancreatic cancer detection. The characterization of these protein candidates used a clinically well-characterized cohort that included plasma samples from patients with pancreatic cancer, chronic pancreatitis, and healthy age-matched controls. Three of the five candidate proteins, including gelsolin, lumican, and tissue inhibitor of metalloproteinase 1, demonstrated an AUC value greater than 0.75 in distinguishing pancreatic cancer from the controls. In addition, we provide an analysis of the reproducibility, accuracy, and robustness of the SRM-based proteomics platform. This information addresses important technical issues that could aid in the adoption of the targeted proteomics platform for practical clinical utility.     

Membrane progesterone receptor alpha as a potential prognostic biomarker for breast cancer survival: a retrospective study

Classically, the actions of progesterone (P4) are attributed to the binding of nuclear progesterone receptor (PR) and subsequent activation of its downstream target genes. These mechanisms, however, are not applicable to PR- or basal phenotype breast cancer (BPBC) due to lack of PR in these cancers. Recently, the function of membrane progesterone receptor alpha (mPRα) in human BPBC cell lines was studied in our lab. We proposed that the signaling cascades of P4→mPRα pathway may play an essential role in controlling cell proliferation and epithelial mesenchymal transition (EMT) of breast cancer. Using human breast cancer tissue microarrays, we found in this study that the average intensity of mPRα expression, but not percentage of breast cancer with high level of mPRα expression (mPRα-HiEx), was significantly lower in the TNM stage 4 patients compared to those with TNM 1-3 patients; and both average intensities of mPRα expression and mPRα-HiEx rates were significantly higher in cancers negative for ER, as compared with those cancers with ER+. However, after adjusting for age at diagnosis and/or TNM stage, only average intensities of mPRα expression were associated with ER status. In addition, we found that the rates of mPRα-HiEx were significantly higher in cancers with epithelial growth factor receptor-1 (EGFR+) and high level of Ki67 expression, indicating positive correlation between mPRα over expression and EGFR or Ki67. Further analysis indicated that both mPRα-HiEx rate and average intensity of mPRα expression were significantly higher in HER2+ subtype cancers (i.e. HER2+ER-PR-) as compared to ER+ subtype cancers. These data support our hypothesis that P4 modulates the activities of the PI3K and cell proliferation pathways through the caveolar membrane bound growth factor receptors such as mPRα and growth factor receptors. Future large longitudinal studies with larger sample size and survival outcomes are necessary to confirm our findings.     

Identification of Apo-A1 as a biomarker for early diagnosis of bladder transitional cell carcinoma

Background  

Bladder transitional cell carcinoma (BTCC) is the fourth most frequent neoplasia in men, clinically characterized by high recurrent rates and poor prognosis. Availability of urinary tumor biomarkers represents a convenient alternative for early detection and disease surveillance because of its direct contact with the tumor and sample accessibility.     

ELISA microarray technology as a high-throughput system for cancer biomarker validation

A large gap currently exists between the ability to discover potential biomarkers and the ability to assess the real value of these proteins for cancer screening. One major challenge in biomarker validation is the inherent variability in biomarker levels. This variability stems from the diversity across the human population and the considerable molecular heterogeneity between individual tumors, even those that originate from a single tissue. An additional challenge with cancer screening is that most cancers are rare in the general population, meaning that assay specificity must be very high. Otherwise, the number of false positives will be much greater than the number of true positives. Due to these challenges associated with biomarker validation, it is necessary to analyze thousands of samples in order to obtain a clear idea of the utility of a screening assay. Enzyme-linked immunosorbent assay (ELISA) microarray technology can simultaneously quantify levels of multiple proteins and, thus, has the potential to accelerate validation of protein biomarkers for clinical use. This review will discuss current ELISA microarray technology and potential advances that could help to achieve the reproducibility and throughput that are required to evaluate cancer biomarkers.     

Identification of annexin II as a novel secretory biomarker for breast cancer

Early prediction of metastatic breast cancer is important for improvement of prognosis and survival rate. The present study aimed to identify secreted protein biomarkers for detection of invasive breast cancer. To this end, we performed a comparative proteomic analysis by a combination of 2DE and MALDI‐TOF MS analysis of conditioned media from invasive H‐Ras MCF10A human breast epithelial cells and noninvasive MCF10A and N‐Ras MCF10A cells. We identified a list of 25 proteins that were strongly detected in media of H‐Ras MCF10A and focused on annexin II, which was shown to be involved in cell motility. Invasive triple‐negative human breast carcinoma cells, Hs578T, and MDA‐MB‐231, showed increased levels of annexin II in media, demonstrating that secretion of annexin II correlated well with the invasive phenotype of cells. We demonstrated a crucial role of annexin II in breast cell invasion/migration and actin cytoskeleton reorganization required for filopodia formation. Annexin II levels in the plasma samples and breast cancer tissues of breast cancer patients were significantly higher than those of normal groups, providing a clinical relevance to our in vitro findings. Taken together, we identified annexin II as a novel secretory biomarker candidate for invasive breast cancer, especially estrogen receptor‐negative breast cancer.