Measurement of aberrant glycosylation of prostate specific antigen can improve specificity in early detection of prostate cancer

Introduction

We previously identified prostate cancer (PCa)-associated aberrant glycosylation of PSA, where α2,3-linked sialylation is an additional terminal N-glycan on free PSA (S2,3PSA). We then developed a new assay system measuring S2,3PSA using a magnetic microbead-based immunoassay. We compared the diagnostic accuracy of conventional PSA and percent-free PSA (%fPSA) tests.

Methods

We used MagPlex beads to measure serum S2,3PSA levels using anti-human fPSA monoclonal antibody (8A6) for capture and anti-α2,3-linked sialic acid monoclonal antibody (HYB4) for detection. We determined the cutoff values in a training test and measured serum S2,3PSA levels in 314 patients who underwent biopsy, including 138 PCa and 176 non-PCa patients with PSA of <10.0 ng/ml. Serum S2,3PSA levels were presented as mean fluorescence intensity (MFI). Receiver operating characteristic curves were used to evaluate the diagnostic accuracy of total PSA, %fPSA, and S2,3PSA.

Results

We determined an MFI cutoff value of 1130 with a sensitivity of 95.0% and specificity of 72.0% for the diagnosis of PCa in the training test. In the validation study, the area under the curve for the detection of PCa with S2,3PSA was 0.84, which was significantly higher than that with PSA or %fPSA.

Conclusions

Although the present study is small and preliminary, these results suggest that the measurement of serum S2,3PSA using a magnetic microbead-based immunoassay may improve the accuracy of early detection of PCa and reduce unnecessary prostate biopsy.     

An approach to the detection and management of early breast cancer

The mortality rate for breast cancer has not changed for 40 years. The ultimate results using the various surgical techniques differ very little. Local tumour excision produces survival rates comparable with those for more radical operations. Ideal treatment should give optimal quality and quantity of survival with minimal trauma. Survival is mainly determined by the ability to live in symbiosis with the disease and in most cases clinically obvious tumours are already incurable. Clinical examination alone will not detect disease at a stage early enough for more effective treatment. Mammography and thermography can detect occult breast cancer and, in particular, thermography can be used for serial examinations in high-risk patients because the procedure is simple and without hazard.     

A rapid method to improve protein detection by indirect ELISA

The enzyme-linked immunosorbant assay (ELISA) is a rapid, high-throughput, quantitative immunoassay for the selective detection of target antigens. The general principle behind an ELISA is antibody mediated capture and detection of an antigen with a measurable substrate. Numerous incarnations of the ELISA have resulted in its commercialization for sensitive diagnostic applications using a variety of detection platforms. Many of these applications require a pair of antibodies necessary for the capture and detection of a specific antigen (cELISA) in defined substrates. However, the availability of cELISA for target antigens is limited and thus restricts the use of this technique for quantitative measure of antigens during discovery. Alternatively, the indirect ELISA (iELISA) requires only a single antibody directed against a target antigen that has been immobilized to a surface. Unlike the cELISA, which uses an immobilized capture antibody that can bind a native antigen in solution followed by a detector antibody that binds captured antigen, the iELISA uses an antibody the binds directly to an immobilized antigen for detection. Although the iELISA may lack the sensitivity of a cELISA, its requirement of only a single antigen specific antibody makes it a simple technique for evaluating the relative difference in the level of target protein expression between samples. However, many antibodies that work effectively to detect protein antigens in other immunoassays such as Western blotting or immunohistochemistry fail to work in microplate based iELISA. Although these alternate immunoassay methods are useful for qualitative determination of target antigens, they provide limited quantitative information, limiting the assessment of sample specific differences in protein expression. We hypothesized that protein conformation following adsorption on the plastic surface of microplates impedes antibody epitope binding and this restriction could be overcome by a short chemical denaturation step. In this report we define a rapid method to assess the utility of an antibody for iELISA application and demonstrate a significant improvement in both qualitative and quantitative protein detection after chemical denaturation using defined assay conditions.     

Molecular biomarker-based screening for early detection of cervical cancer

OBJECTIVE: To evaluate the effectiveness of a molecular biomarker-based screening method for early detection of cervical cancer. STUDY DESIGN: Fluorescent immunochemical labeling was used to classify cervical cytology specimens as probably normal or probably abnormal. These specimens were then Papanicolaou stained and evaluated twice by a cytotechnologist and by a pathologist when appropriate. The first evaluation was performed as conventional Pap screening to assign a reference per-specimen diagnosis. The second evaluation assigned a cytologic diagnosis to each cell in the specimen. The fluorescence results were correlated with those from each of the two morphologic evaluations to determine the sensitivity and specificity of the method on a per-specimen and a per-cell basis. Where available, in cases of positive morphologic analysis, biopsy was compared to fluorescence results. RESULTS: The per-specimen sensitivity and specificity were 87.5% and 81.8%, respectively, when using atypical squamous cells of undetermined significance and "above" as the decision threshold. For the same specimens evaluated on a cell-by-cell basis, the corresponding sensitivity and specificity were 71.4% and 66.3% when the same threshold was used. The per-specimen sensitivity for high grade squamous intraepithelial lesions was 100% under these conditions. CONCLUSION: This method appears to be a robust and reliable means of detecting cervical dysplasia and is now being evaluated in additional clinical studies.     

Genome-specific higher-order background models to improve motif detection

Motif detection based on Gibbs sampling is a common procedure used to retrieve regulatory motifs in silico. Using a species-specific background model was previously shown to increase the robustness of the algorithm. Here, we demonstrate that selecting a non-species-adapted background model can have an adverse effect on the results of motif detection. The large differences in the average nucleotide composition of prokaryotic sequences exacerbate the problem of exchanging background models. Therefore, we have developed complex background models for all prokaryotic species with available genome sequences.     

Determination of plasma microRNA for early detection of gastric cancer

Gastric cancer is the fourth most prevalent malignancy worldwide and remains the second most common cause of cancer-related death globally. Understanding the molecular structure of gastric carcinogenesis might identify new diagnostic and therapeutic strategies for this disease. Thus, early detection of gastric cancer is a key measure to reduce the mortality and improve the prognosis of gastric cancer. There have recently been several reports that microRNAs (miRNAs) circulate in highly stable, cell-free forms in blood. Because serum and plasma miRNAs are relatively easy to access, circulating miRNAs also have great potential to serve as non-invasive biomarkers. Although a number of miRNAs associated with gastric cancer have been identified, the underlying mechanism of these miRNAs in tumorigenesis and tumor progression remains to be investigated. The purpose of this study is to identify the potential of serum miRNAs as biomarkers for early detection of gastric cancer patients. RNA was isolated using the High Pure miRNA Isolation Kit (Roche) following the manufacturer’s protocol. cDNA and preamplification protocols were obtained from the isolated plasma miRNAs. The BioMark? 96.96 Dynamic Array (Fluidigm Corporation) for real-time qPCR was used to simultaneously quantite the expression of 740 miRNAs. All statistical analyses were performed using the Biogazelle’s qbase PLUS 2.0 software. In this study, among 740 miRNAs that we analyzed only miR-195-5p was significantly (p < 0.05, fold changes = 13, 3) down-regulated in gastric cancer patients compared with control. We demonstrated that miR-195-5p is a novel tumor suppressor miRNA and may contribute to gastric carcinogenesis. The miRNA expression profile described in this study should contribute to future studies on the role of miRNAs in gastric cancer.     

Proteomic analysis at the bedside: early detection of cancer

Proteomic technologies promise to accelerate rapidly a new era in molecular medicine, especially in the detection and discovery of disease-related biomarkers. These technologies have no bigger impact than in the field of human cancer research. Beyond lifestyle-associated prevention strategies, early detection of cancer has the most profound impact on the ultimate course of the disease: the earlier the cancer is detected, the better the prognosis. Today, new proteomic technologies are being used to discover new diagnostic and prognostic biomarkers for the early detection and treatment of cancer that will have important implications at the bedside.     

SELDI-TOF-based serum proteomic pattern diagnostics for early detection of cancer

Proteomics is more than just generating lists of proteins that increase or decrease in expression as a cause or consequence of pathology. The goal should be to characterize the information flow through the intercellular protein circuitry that communicates with the extracellular microenvironment and then ultimately to the serum/plasma macroenvironment. The nature of this information can be a cause, or a consequence, of disease and toxicity-based processes. Serum proteomic pattern diagnostics is a new type of proteomic platform in which patterns of proteomic signatures from high dimensional mass spectrometry data are used as a diagnostic classifier. This approach has recently shown tremendous promise in the detection of early-stage cancers. The biomarkers found by SELDI-TOF-based pattern recognition analysis are mostly low molecular weight fragments produced at the specific tumor microenvironment.     

Proteomics for early detection of colorectal cancer: recent updates

Introduction: Colorectal cancer (CRC) is a common type of cancer with a relatively poor survival rate. The survival rate of patients could be improved if CRC is detected early. Biomarkers associated with early stages of tumor development might provide useful tools for the early diagnosis of colorectal cancer.

Areas covered: Online searches using PubMed and Google Scholar were performed using keywords and with a focus on recent proteomic studies. The aim of this review is to highlight the need for biomarkers to improve the detection rate of early CRC and provide an overview of proteomic technologies used for biomarker discovery and validation. This review will also discuss recent proteomic studies which focus on identifying biomarkers associated with the early stages of CRC development.

Expert commentary: A large number of CRC biomarkers are increasingly being identified by proteomics using diverse approaches. However, the clinical relevance and introduction of these markers into clinical practice cannot be determined without a robust validation process. The size of validation cohorts remains a major limitation in many biomarker studies.     

A simple vector system to improve performance and utilisation of recombinant antibodies

Background  

Isolation of recombinant antibody fragments from antibody libraries is well established using technologies such as phage display. Phage display vectors are ideal for efficient display of antibody fragments on the surface of bacteriophage particles. However, they are often inefficient for expression of soluble antibody fragments, and sub-cloning of selected antibody populations into dedicated soluble antibody fragment expression vectors can enhance expression.     

Characterization of serum biomarkers for detection of early stage ovarian cancer

We have previously reported the identification of three ovarian cancer biomarker panels comprised of SELDI-TOF-MS peaks representing 14 differentially expressed serum proteins for the diagnosis of ovarian cancer. Using micro-LC-MS/MS, we identified five m/z peaks as transthyretin (TTR 13.9 kDa, TTR fragment 12.9 kDa), beta-hemoglobin (Hb, 15.9 kDa), apolipoprotein AI (ApoAI, 28 kDa) and transferrin (TF, 79 kDa). Western and/or ELISA methods confirmed the differential expression of TTR, Hb, and TF, and multivariate analyses resulted in improving the detection of early stage ovarian tumors (low malignant potential and malignant; receiver operating characteristic, ROC 0.933) as compared to cancer antigen CA125 alone (ROC 0.833). Interestingly, when CA125 was included with our markers in the multivariate analysis, the ROC increased to 0.959. Furthermore, multivariate analysis with only the mucinous subtype of early stage ovarian tumors, showed our markers to greatly improve the detection of disease (ROC 0.959) as compared to CA125 alone (ROC 0.613). Interestingly, the combination of CA125 with our markers did not seem to further improve the detection of mucinous tumors (ROC 0.955). We conclude that TTR, Hb, ApoAI and TF, when combined with CA125 should significantly improve the detection of early stage ovarian cancer.