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Noninvasive Early Biomarkers in Ovarian Cancer
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Platinum resistance is a major cause of treatment failure in ovarian cancer. We previously identified matrix metalloproteinase 9 (MMP-9) as a potential therapeutic target of chemoresistant disease. A2780cis (cisplatin-resistant) and A2780 (cisplatin-sensitive) ovarian carcinoma cell lines were used. The cytotoxic effect of MMP-9/MMP-2 inhibitor, (2R)-2-[(4-Biphenylsulfonyl) amino]-3 phenylpropionic acid (C21H19NO4S) alone or in combination with cisplatin was determined using high content screening. Protein expression was examined using immunohistochemistry and ELISA. Co-incubation of cisplatin and an MMP-9/MMP-2 inhibitor, (2R)-2-[(4-Biphenylsulfonyl) amino]-3 phenylpropionic acid (C21H19NO4S) resulted in significantly greater cytotoxicity as compared to either treatment alone in a cisplatin resistant MMP-9 overexpressing cell line; A2780cis. In addition, pre-incubating with MMP-9i prior to cisplatin further enhances the cytotoxic effect. No significant difference was observed in MMP-9 protein in tissue but a trend towards increased MMP-9 was observed in recurrent serum. We propose that MMP-9/MMP-2i may be utilized in the treatment of recurrent/chemoresistant ovarian cancers that overexpress MMP-9 mRNA but its role in vivo remains to be evaluated.
Background:
Tumor-derived DNA can be found in the plasma of cancer patients. In this study, we explored the use of shotgun massively parallel sequencing (MPS) of plasma DNA from cancer patients to scan a cancer genome noninvasively.
Methods:
Four hepatocellular carcinoma patients and a patient with synchronous breast and ovarian cancers were recruited. DNA was extracted from the tumor tissues, and the preoperative and postoperative plasma samples of these patients were analyzed with shotgun MPS.
Results:
We achieved the genomewide profiling of copy number aberrations and point mutations in the plasma of the cancer patients. By detecting and quantifying the genomewide aggregated allelic loss and point mutations, we determined the fractional concentrations of tumor-derived DNA in plasma and correlated these values with tumor size and surgical treatment. We also demonstrated the potential utility of this approach for the analysis of complex oncologic scenarios by studying the patient with 2 synchronous cancers. Through the use of multiregional sequencing of tumoral tissues and shotgun sequencing of plasma DNA, we have shown that plasma DNA sequencing is a valuable approach for studying tumoral heterogeneity.
Conclusions:
Shotgun DNA sequencing of plasma is a potentially powerful tool for cancer detection, monitoring, and research.
The aim of this study was to assess the clinical value of preoperative blood levels of HE4 as a predictor of overall survival in patients with ovarian cancer and to validate previous data of HE4 and the ROMA algorithm including HE4 and CA125 in discriminating benign and malignant ovarian tumors.
The preoperative plasma levels of HE4 and CA125 were analyzed with ELISA in 312 patients with adnexal lesions. Tumors were classified as benign (n= 206), borderline (i.e. low malignant potential tumors) (n= 25), and well (n= 14), moderately (n= 15), and poorly (n= 51) differentiated malignant.
In univariate Cox regression analyses high levels (dichotomized at the median) of HE4, CA125, increased age (continuous variable), advanced-stage of disease 2-4, histological grade 3 and non-optimal tumor debulking at primary surgery were all significantly associated with shorter overall survival. A multivariate Cox regression model including pre-operative available covariates HE4 and CA125 both dichotomized at median in addition to age as continuous variable showed that high levels of HE4 was an independent prognostic marker for worse prognosis HR 2.02 (95% CI 1.1-3.8). In postmenopausal women the ROMA algorithm gave the highest AUC of 0.94 (95% CI, 0.90-0.97) which was higher than the separate markers HE4 AUC 0.91 (95% CI 0.86-0.95) and CA125 AUC 0.91(95% CI 0.87-0.96).
High concentration of plasma HE4 is an independent preoperative marker of poor prognosis in patients with ovarian cancer. The algorithm ROMA discriminates in postmenopausal women between malignant and benign tumors with an AUC of 0.94.
Risk for Ovarian Malignancy Algorithm (ROMA) and Human epididymis protein 4 (HE4) appear to be promising predictors for epithelial ovarian cancer (EOC), however, conflicting results exist in the diagnostic performance comparison among ROMA, HE4 and CA125.
Remote databases (MEDLINE/PUBMED, EMBASE, Web of Science, Google Scholar, the Cochrane Library and ClinicalTrials.gov) and full texts bibliography were searched for relevant abstracts. All studies included were closely assessed with the QUADAS-2 (Quality Assessment of Diagnostic Accuracy Studies-2). EOC predictive value of ROMA was systematically evaluated, and comparison among the predictive performances of ROMA, HE4 and CA125 were conducted within the same population. Sensitivity, specificity, DOR (diagnostic odds ratio), LR ± (positive and negative likelihood ratio) and AUC (area under receiver operating characteristic-curve) were summarized with a bivariate model. Subgroup analysis and sensitivity analysis were used to explore the heterogeneity.
Data of 7792 tests were retrieved from 11 studies. The overall estimates of ROMA for EOC predicting were: sensitivity (0.89, 95% CI 0.84-0.93), specificity (0.83, 95% CI 0.77-0.88), and AUC (0.93, 95% CI 0.90-0.95). Comparison of EOC predictive value between HE4 and CA125 found, specificity: HE4 (0.93, 95% CI 0.87-0.96) > CA125 (0.84, 95% CI 0.76-0.90); AUC: CA125 (0.88, 95% CI 0.85-0.91) > HE4 (0.82, 95% CI 0.78-0.85). Comparison of OC predictive value between HE4 and CA125 found, AUC: CA125 (0.89, 95% CI 0.85-0.91) > HE4 (0.79, 95% CI 0.76-0.83). Comparison among the three tests for EOC prediction found, sensitivity: ROMA (0.86, 95%CI 0.81-0.91) > HE4 (0.80, 95% CI 0.73-0.85); specificity: HE4 (0.94, 95% CI 0.90-0.96) > ROMA (0.84, 95% CI 0.79-0.88) > CA125 (0.78, 95%CI 0.73-0.83).
ROMA is helpful for distinguishing epithelial ovarian cancer from benign pelvic mass. HE4 is not better than CA125 either for EOC or OC prediction. ROMA is promising predictors of epithelial ovarian cancer to replace CA125, but its utilization requires further exploration.
Tumor-associated autoantibodies (AAbs) have been described in patients with lung cancer, and the EarlyCDT®-Lung test that measures such AAbs is available as an aid for the early detection of lung cancer in high-risk populations. Improvements in specificity would improve its cost-effectiveness, as well as reduce anxiety associated with false positive tests. Samples from 235 patients with newly diagnosed lung cancer and matched controls were measured for the presence of AAbs to a panel of six (p53, NY-ESO-1, CAGE, GBU4-5, Annexin I, and SOX2) or seven (p53, NY-ESO-1, CAGE, GBU4-5, SOX2, HuD, and MAGE A4) antigens. Data were assessed in relation to cancer type and stage. The sensitivity and specificity of these two panels were also compared in two prospective consecutive series of 776 and 836 individuals at an increased risk of developing lung cancer. The six-AAb panel gave a sensitivity of 39 % with a specificity of 89 %, while the seven-AAb panel gave a sensitivity of 41 % with a specificity of 91 % which, once adjusted for occult cancers in the population, resulted in a specificity of 93 %. Analysis of these AAb assays in the at-risk population confirmed that the seven-AAb panel resulted in a significant increase in the specificity of the test from 82 to 90 %, with no significant change in sensitivity. The change from a six- to a seven-AAb assay can improve the specificity of the test and would result in a PPV of 1 in 8 and an overall accuracy of 92 %.
Point-of-care (POC) implementation of early detection and screening methodologies for ovarian cancer may enable improved survival rates through early intervention. Current laboratory-confined immunoanalyzers have long turnaround times and are often incompatible with multiplexing and POC implementation. Rapid, sensitive, and multiplexable POC diagnostic platforms compatible with promising early detection approaches for ovarian cancer are needed. To this end, we report the adaptation of the programmable bio-nano-chip (p-BNC), an integrated, microfluidic, and modular (programmable) platform for CA125 serum quantitation, a biomarker prominently implicated in multimodal and multimarker screening approaches. In the p-BNCs, CA125 from diseased sera (Bio) is sequestered and assessed with a fluorescence-based sandwich immunoassay, completed in the nano-nets (Nano) of sensitized agarose microbeads localized in individually addressable wells (Chip), housed in a microfluidic module, capable of integrating multiple sample, reagent and biowaste processing, and handling steps. Antibody pairs that bind to distinct epitopes on CA125 were screened. To permit efficient biomarker sequestration in a three-dimensional microfluidic environment, the p-BNC operating variables (incubation times, flow rates, and reagent concentrations) were tuned to deliver optimal analytical performance under 45 minutes. With short analysis times, competitive analytical performance (inter- and intra-assay precision of 1.2% and 1.9% and limit of detection of 1.0 U/mL) was achieved on this minisensor ensemble. Furthermore, validation with sera of patients with ovarian cancer (n = 20) showed excellent correlation (R(2) = 0.97) with gold-standard ELISA. Building on the integration capabilities of novel microfluidic systems programmed for ovarian cancer, the rapid, precise, and sensitive miniaturized p-BNC system shows strong promise for ovarian cancer diagnostics.
Background
Tumor-infiltrating CD8+ T cells are correlated with prolonged progression-free and overall survival in epithelial ovarian cancer (EOC). A significant fraction of EOC patients mount autoantibody responses to various tumor antigens, however the relationship between autoantibodies and tumor-infiltrating T cells has not been investigated in EOC or any other human cancer. We hypothesized that autoantibody and T cell responses may be correlated in EOC and directed toward the same antigens.
Methodology and Principal Findings
We obtained matched serum and tumor tissue from 35 patients with high-grade serous ovarian cancer. Serum samples were assessed by ELISA for autoantibodies to the common tumor antigen NY-ESO-1. Tumor tissue was examined by immunohistochemistry for expression of NY-ESO-1, various T cell markers (CD3, CD4, CD8, CD25, FoxP3, TIA-1 and Granzyme B) and other immunological markers (CD20, MHC class I and MHC class II). Lymphocytic infiltrates varied widely among tumors and included cells positive for CD3, CD8, TIA-1, CD25, FoxP3 and CD4. Twenty-six percent (9/35) of patients demonstrated serum IgG autoantibodies to NY-ESO-1, which were positively correlated with expression of NY-ESO-1 antigen by tumor cells (r = 0.57, p = 0.0004). Autoantibodies to NY-ESO-1 were associated with increased tumor-infiltrating CD8+, CD4+ and FoxP3+ cells. In an individual HLA-A2+ patient with autoantibodies to NY-ESO-1, CD8+ T cells isolated from solid tumor and ascites were reactive to NY-ESO-1 by IFN-γ ELISPOT and MHC class I pentamer staining.
Conclusion and Significance
We demonstrate that tumor-specific autoantibodies and tumor-infiltrating T cells are correlated in human cancer and can be directed against the same target antigen. This implies that autoantibodies may collaborate with tumor-infiltrating T cells to influence clinical outcomes in EOC. Furthermore, serological screening methods may prove useful for identifying clinically relevant T cell antigens for immunotherapy.
Ovarian cancer has one of the highest mortalities in malignancies in women, but little is known of its tumour progression properties and there is still no effective molecule that can monitor its growth or therapeutic responses. MSLN (mesothelin), a secreted protein that is overexpressed in ovarian cancer tissues with a poor clinical outcome, has been previously identified to activate PI3K (phosphoinositide 3-kinase)/Akt signalling and inhibit paclitaxel-induced apoptosis. The present study investigates the correlation between MSLN and MMP (matrix metalloproteinase)-7 in the progression of ovarian cancer, and the mechanism of MSLN in enhancing ovarian cancer invasion. The expression of MSLN correlated well with MMP-7 expression in human ovarian cancer tissues. Overexpressing MSLN or ovarian cancer cells treated with MSLN showed enhanced migration and invasion of cancer cells through the induction of MMP-7. MSLN regulated the expression of MMP-7 through the ERK (extracellular-signal-regulated kinase) 1/2, Akt and JNK (c-Jun N-terminal kinase) pathways. The expression of MMP-7 and the migrating ability of MSLN-treated ovarian cancer cells were suppressed by ERK1/2- or JNK-specific inhibitors, or a decoy AP-1 (activator protein 1) oligonucleotide in in vitro experiments, whereas in vivo animal experiments also demonstrated that mice treated with MAPK (mitogen-activated protein kinase)/ERK- or JNK-specific inhibitors could decrease intratumour MMP-7 expression, delay tumour growth and extend the survival of the mice. In conclusion, MSLN enhances ovarian cancer invasion by MMP-7 expression through the MAPK/ERK and JNK signal transduction pathways. Blocking the MSLN-related pathway could be a potential strategy for inhibiting the growth of ovarian cancer.
According to extensive epidemiologic data, infertility is associated with increased ovarian cancer risk. Previous studies showed that both women with infertility and those with ovarian cancer have autoantibodies to ovarian antigens. The objective was to determine if women with infertility have antibodies to mesothelin, a well-characterized ovarian cancer antigen.
Sera were obtained from women with infertility (n = 109), ovarian cancer (n = 28), benign ovarian tumors or cysts (n = 24), and from healthy women (n = 152). Infertility included those with a risk for ovarian cancer; endometriosis (n = 23), ovulatory dysfunction (n = 17), premature ovarian failure (POF; n = 25) and unexplained infertility (n = 44). Sera were assayed for mesothelin antibodies and for circulating mesothelin antigen by immunoassay and compared with assay control sera (n = 16) to determine a positive result.
Mesothelin antibodies were significantly more frequent in women with prematurely reduced ovarian function including ovulatory dysfunction (59%), ovarian failure (44%) and unexplained infertility (25%) compared with controls. In contrast, women with endometriosis, who also have a high risk for ovarian cancer, did not have mesothelin antibodies. Serum levels of mesothelin were rarely elevated in women with infertility but were high in most patients with ovarian cancer.
We show for the first time that antibodies to mesothelin, a well-characterized ovarian cancer antigen, occur in some women with epidemiologic risk for ovarian cancer. The results suggest it may be possible to identify which women with infertility have ovarian cancer risk.
Screening for ovarian cancer with cancer antigen 125 (CA-125) and transvaginal ultrasound has an unknown effect on mortality.
To evaluate the effect of screening for ovarian cancer on mortality in the Prostate, Lung, Colorectal and Ovarian (PLCO) Cancer Screening Trial.
Randomized controlled trial of 78,216 women aged 55 to 74 years assigned to undergo either annual screening (n = 39,105) or usual care (n = 39,111) at 10 screening centers across the United States between November 1993 and July 2001. Intervention The intervention group was offered annual screening with CA-125 for 6 years and transvaginal ultrasound for 4 years. Participants and their health care practitioners received the screening test results and managed evaluation of abnormal results. The usual care group was not offered annual screening with CA-125 for 6 years or transvaginal ultrasound but received their usual medical care. Participants were followed up for a maximum of 13 years (median [range], 12.4 years [10.9-13.0 years]) for cancer diagnoses and death until February 28, 2010.
Mortality from ovarian cancer, including primary peritoneal and fallopian tube cancers. Secondary outcomes included ovarian cancer incidence and complications associated with screening examinations and diagnostic procedures.
Ovarian cancer was diagnosed in 212 women (5.7 per 10,000 person-years) in the intervention group and 176 (4.7 per 10,000 person-years) in the usual care group (rate ratio [RR], 1.21; 95% confidence interval [CI], 0.99-1.48). There were 118 deaths caused by ovarian cancer (3.1 per 10,000 person-years) in the intervention group and 100 deaths (2.6 per 10,000 person-years) in the usual care group (mortality RR, 1.18; 95% CI, 0.82-1.71). Of 3285 women with false-positive results, 1080 underwent surgical follow-up; of whom, 163 women experienced at least 1 serious complication (15%). There were 2924 deaths due to other causes (excluding ovarian, colorectal, and lung cancer) (76.6 per 10,000 person-years) in the intervention group and 2914 deaths (76.2 per 10,000 person-years) in the usual care group (RR, 1.01; 95% CI, 0.96-1.06).
Among women in the general US population, simultaneous screening with CA-125 and transvaginal ultrasound compared with usual care did not reduce ovarian cancer mortality. Diagnostic evaluation following a false-positive screening test result was associated with complications. Trial Registration clinicaltrials.gov Identifier: NCT00002540.
This paper reviews current screening techniques as well as novel biomarkers and their potential role in early detection of ovarian cancer. Ovarian cancer is one of the most common reproductive cancers and has the highest mortality rate amongst gynecologic cancers. Because most ovarian cancer diagnoses occur in the late stages of the disease, five-year survival rates fall below 20%. To improve survival rates and to lower mortality rates for ovarian cancer, improved detection at early stages of the disease is needed. Current screening approaches include tumor markers, ultrasound, or a combination. Efforts are underway to discover new biomarkers of ovarian cancer in order to surmount the obstacles in early-stage diagnosis. Among serum protein markers, HE4 and mesothelin can augment CA125 detection providing higher sensitivity and specificity due to the presence of these proteins in early-stage ovarian cancer. Detection testing that includes methylation of the MCJ gene and increased expression of vascular endothelial growth factor is correlated to poor prognosis and may predict patient survival outcome. Detection testing of biomarkers with long-term stability and combination panels of markers, will likely lead to effective screening strategies with high specificity and sensitivity for early detection of ovarian cancer.
Recently, a Risk of Ovarian Malignancy Algorithm (ROMA) utilising human epididymis secretory protein 4 (HE4) and CA125 successfully classified patients as presenting a high or low risk for epithelial ovarian cancer (EOC). We validated this algorithm in an independent prospective study.
Women with a pelvic mass, who were scheduled to have surgery, were enrolled in a prospective study. Preoperative serum levels of HE4 and CA125 were measured in 389 patients. The performance of each of the markers, as well as that of ROMA, was analysed.
When all malignant tumours were included, ROMA (receiver operator characteristic (ROC)-area under curve (AUC)=0.898) and HE4 (ROC-AUC)=0.857) did not perform significantly better than CA125 alone (ROC-AUC=0.877). Using a cutoff for ROMA of 12.5% for pre-menopausal patients, the test had a sensitivity of 67.5% and a specificity of 87.9%. With a cutoff of 14.4% for post-menopausal patients, the test had a sensitivity of 90.8% and a specificity of 66.3%. For EOC vs benign disease, the ROC-AUC of ROMA increased to 0.913 and for invasive EOC vs benign disease to 0.957.
This independent validation study demonstrated similar performance indices to those recently published. However, in this study, HE4 and ROMA did not increase the detection of malignant disease compared with CA125 alone. Although the initial reports were promising, measurement of HE4 serum levels does not contribute to the diagnosis of ovarian cancer.
Circulating tumor cells (CTCs) shed from primary and metastatic cancers are admixed with blood components and are thus rare, making their isolation and characterization a major technological challenge. CTCs hold the key to understanding the biology of metastasis and provide a biomarker to noninvasively measure the evolution of tumor genotypes during treatment and disease progression. Improvements in technologies to yield purer CTC populations amenable to better cellular and molecular characterization will enable a broad range of clinical applications, including early detection of disease and the discovery of biomarkers to predict treatment responses and disease progression.
Autoantibodies (AAbs) to tumor-associated antigens (TAAs) have been identified in the circulation of patients with cancer. This paper will focus on recent knowledge related to circulating AAbs to TAAs in breast carcinoma. So far, the following TAAs have been identified to elicit circulating AAbs in breast carcinoma: p53, MUC-1, heat shock proteins (HSP-27, HSP-60, and HSP-90), HER2/neu/c-erb B2, GIPC-1, c-myc, c-myb, cancer-testis antigens (NY-ESO-1), BRCA1, BRCA2, endostatin, lipophilin B, cyclin B1, cyclin D1, fibulin, insulin-like growth factor binding protein 2 (IGFBP-2), topoisomerase II alpha (TOPO2α), and cathepsin D. Measurement of serum AAbs to one specific TAA only is of little value for screening and early diagnosis of breast carcinoma; however, assessment of AAbs to a panel of TAAs may have promising diagnostic potential.
Metabolomics is a new, rapidly expanding field dedicated to the global study of metabolites in biological systems. In this article metabolomics is applied to find urinary biomarkers for breast and ovarian cancer.
Urine samples were collected from early- and late-stage breast and ovarian cancer patients during presurgical examinations and randomly from females with no known cancer. After quantitatively measuring a set of metabolites using nuclear magnetic resonance spectroscopy, both univariate and multivariate statistical analyses were employed to determine significant differences.
Metabolic phenotypes of breast and ovarian cancers in comparison with normal urine and with each other revealed significance at Bonferroni-corrected significance levels resulting in unique metabolite patterns for breast and ovarian cancer. Intermediates of the tricarboxylic acid cycle and metabolites relating to energy metabolism, amino acids, and gut microbial metabolism were perturbed.
The results presented here illustrate that urinary metabolomics may be useful for detecting early-stage breast and ovarian cancer.
Ovarian cancer is a heterogeneous, rapidly progressive, highly lethal disease of low prevalence. The etiology remains poorly understood. Numerous risk factors have been identified, the most prominent involving an inherited predisposition in 10% of cases. Women with germline mutations associated with Hereditary Breast/Ovarian Cancer and Lynch syndromes have dramatically elevated risks (up to 46% and 12%, respectively). Risk-reducing salpingo-oophorectomy is the best method to prevent ovarian cancer in these high-risk women. Significant risk reduction is also seen in the general population who use oral contraceptives. Since up to 89% patients with early-stage disease have symptoms prior to diagnosis, increased awareness of the medical community may facilitate further workup in patients who otherwise would have had a delay. Despite enormous effort, there is no proof that routine screening for ovarian cancer in either the high-risk or general populations with serum markers, sonograms, or pelvic examinations decreases mortality. Further evaluation is needed to determine whether any novel biomarkers, or panels of markers, have clinical utility in early detection. Prospective clinical trials have to be designed and completed prior to offering of any of these new diagnostic tests. CA125 is currently the only biomarker recommended for monitoring of therapy as well as detection of recurrence. This commentary provides an overview on the background, screening and surveillance of ovarian cancer.
Screening is an unsolved problem for ovarian cancer (OvCA). As late detection is equivalent to poor prognosis, we analysed whether OvCA patients show diagnostically meaningful microRNA (miRNA) patterns in blood cells.
Blood-borne whole miRNome profiles from 24 patients with OvCA and 15 age- and sex-matched healthy controls were biostatistically evaluated.
Student's t-test revealed 147 significantly deregulated miRNAs before and 4 after Benjamini-Hochberg adjustment. Although these included miRNAs already linked to OvCA (e.g., miR-16, miR-155), others had never before been connected to specific diseases. A bioinformatically calculated miRNA profile allowed for discrimination between blood samples of OvCA patients and healthy controls with an accuracy of >76%. When only cancers of the serous subtype were considered and compared with an extended control group (n=39), accuracy, specificity and sensitivity all increased to >85%.
Our proof-of-principle study strengthens the hypothesis that neoplastic diseases generate characteristic miRNA fingerprints in blood cells. Still, the obtained OvCA-associated miRNA pattern is not yet sensitive and specific enough to permit the monitoring of disease progression or even preventive screening. Microarray-based miRNA profiling from peripheral blood could thus be combined with other markers to improve the notoriously difficult but important screening for OvCA.
Heat‐shock proteins promote cell survival under adverse environmental conditions. Synthesis of the 27‐kDa (HSP27), 70‐kDa (HSP70), and 90‐kDa (HSP90) heat‐shock proteins is increased in malignantly transformed cells and has been associated with tumor proliferation, metastasis, and resistance to chemotherapeutic agents. The increased expression of heat‐shock proteins and their association with tumor‐specific antigens may result in local immunity to the heat‐shock proteins. We examined the occurrence of IgA antibodies to HSP27, HSP70, and HSP90 in the lower genital tracts of women with possible gynecologic cancers. Cervical samples were obtained from 119 consecutive women being evaluated for a gynecologic malignancy or returning for a follow‐up examination following cancer treatment. Aliquots were tested for IgA anti‐heat‐shock protein antibodies by ELISA. Aliquots were also tested for IgG antibodies to HSP27 as well as for human papillomavirus. Anti‐HSP27 IgA was detected in 85.7% of 21 women with endometrial cancer tested prior to diagnosis and in 41.1% of 17 women tested after treatment. In women with ovarian cancer, 77.8% of 9 women tested prior to diagnosis and 75.0% of 24 women evaluated after treatment were anti‐HSP27 IgA‐positive. Of 6 women with cervical cancer tested prior to diagnosis, 5 were positive for this antibody. None of 25 women with benign diagnoses or 46 healthy women were cervical IgA anti‐HSP27‐positive (P < 0.0001). In contrast, anti‐HSP27 IgG was not associated with a gynecologic malignancy. HSP27 cervical antibodies were not associated with the presence of human papillomavirus. Cervical IgA antibodies to HSP90 were associated with ovarian cancer; antibodies to HSP70 were not cancer‐associated. We conclude that cervical IgA antibodies to HSP27 may be indicators of a gynecologic malignancy. Int. J. Cancer 87:824–828, 2000. © 2000 Wiley‐Liss, Inc.
Of all of the epithelial ovarian cancers, clear cell carcinoma (CCC) of the ovary has the worst prognosis. We applied the oligonucleotide array technique to identify genes generally involved in CCC. Of the ∼12,600 genes that were analyzed, 28 were expressed significantly differently between four CCC and seven non-CCC cell lines. Among 16 up-regulated genes in CCC, we further investigated a transcription factor, hepatocyte nuclear factor-1β (HNF-1β). We validated up-regulation of HNF-1β in CCC in terms of both mRNA and protein level using real-time quantitative reverse transcriptase-polymerase chain reaction and immunoblotting. Immunohistochemical analysis of 83 surgically resected ovarian cancers showed that almost all CCC specimens (21 of 22 cases) had nuclear staining for HNF-1β, whereas most non-CCC specimens (60 of 61 cases) showed no immunostaining or only focal and faint staining in the nucleus. Furthermore, we investigated the significance of HNF-1β expression in CCC using RNA interference. The reduction of HNF-1β expression by RNA interference induced apoptotic cell death in ovarian CCC cells, which was confirmed by terminal dUTP nick-end labeling and fluorescence-activated cell-sorting analyses. Our results suggest that HNF-1β is not only an excellent CCC-specific molecular marker but also a molecular target for therapy of ovarian CCC.
Estimates of the contribution of BRCA1 and BRCA2 to breast cancer incidence in outbred populations have been based on studies that are either small or have selected for cases diagnosed at an early age. Only one of these has reported an estimate of the breast cancer risk associated with a mutation in these genes, and there is no published ovarian cancer risk estimate derived from a population-based case series. We screened a population-based series of breast cancer cases diagnosed before the age of 55 for mutations in BRCA1 and BRCA2. Pedigree information from the mutation carriers was used to estimate penetrance and the proportion of familial risk of breast cancer due to BRCA1 and BRCA2. We identified eight (0.7%) BRCA1 and 16 (1.3%) BRCA2 mutation carriers in 1220 breast cancer cases (actual sample size 1435 adjusted for 15% polymerase chain reaction failure rate). Mutation prevalence was substantially higher in cases diagnosed before 35 years-of-age and with increasing number of relatives affected with breast or ovarian cancer. However, most mutation carriers were diagnosed in the older age groups, and a minority reported a first-degree relative with breast cancer. Breast cancer penetrance by age 80 was estimated to be 48% (95% CI 7–82%) for BRCA1 mutation carriers and 74% (7–94%) for BRCA2 mutation carriers. Ovarian cancer penetrance for BRCA1 and BRCA2 combined was 22% (6–65%) by age 80. 17% of the familial risk of breast cancer was attributable to BRCA1 and BRCA2. At birth, the estimated prevalence of BRCA1 mutation carriers was 0.07% or 0.09% depending on the penetrance function used for the calculation. For BRCA2 the birth prevalence estimates were 0.14% and 0.22%. Mutations in the genes BRCA1 and BRCA2 are rare in the population and account for a small fraction of all breast cancer in the UK. They account for less than one fifth of the familial risk of breast cancer. Eligibility criteria for BRCA1 and BRCA2 mutation testing based on family history and age of onset will identify only a small proportion of mutation carriers. © 2000 Cancer Research Campaign
As a promising biomarker, human epididymis protein 4 (HE4) has been widely used for the early detection and differential diagnosis of ovarian cancer. This study evaluated the function of HE4 in the carcinogenesis and progression of ovarian cancer. An enzyme immunometric assay, used to detect HE4 in the serum of ovarian cancer patients, showed that the protein could discriminate between malignant and benign ovarian tumours with high specificity. An exogenous HE4 gene was transfected into ovarian cancer cell lines and an immortalized ovarian epithelial cell line. Compared with the controls, HE4 overexpression significantly promoted cell apoptosis and adhesion. Overexpression of HE4 also led to significant inhibition of cell proliferation, migration and invasiveness in vitro, as well as xenograft tumour formation in vivo. This is the first report to demonstrate the functional importance of HE4 in multiple cellular processes and indicates that HE4 may play a protective role in the progression of ovarian cancer.
Serum tumor markers have a major role in the screening, diagnosis, and monitoring of most of the gynecologic cancers. Ovarian cancer is one of the deadliest of the group because it is so frequently asymptomatic until it has advanced to an untreatable stage. Even serum cancer antigen-125 (CA-125), clinically one of the most reliable serum markers for ovarian cancer, is elevated in only half of early-stage still-treatable tumors. Because of the very low prevalence of ovarian cancer in the general population, at present, there is no cost-effective imaging or simple microscopic screening test for ovarian cancer as there is for breast and cervical cancers. However, recent proteomics and nucleic acid-based analyses have shown great promise for the discovery of new and more useful serum biomarkers, which cumulatively might provide such a screening tool. In this review, we will discuss both the currently used serum tumor markers for screening, diagnosis, monitoring of ovarian cancer, and the novel biomarkers that are now under investigation and validation. Cancer Epidemiol Biomarkers Prev; 1-11. ©2012 AACR.
There has been concern about current ovarian cancer screening strategies after the publication showing a lack of benefit from screening in the ovarian part of the Prostate, Lung, Colorectal and Ovarian Cancer Screening Trial. The ongoing United Kingdom Collaborative Trial of Ovarian Cancer Screening involves 202,638 low-risk postmenopausal women. The performance characteristics on prevalence screen in sensitivity, specificity, and stage distribution have been encouraging. Screening differs from the Prostate, Lung, Colorectal and Ovarian Cancer Screening Trial in the use of algorithms to interpret CA125 and ultrasound imaging and well-defined, centrally coordinated management of screen-detected abnormalities with protocols for intervention based on screening findings. There is a possibility that these essential differences may be sufficient to alter the natural history of ovarian cancer and ultimately lead to demonstration of a mortality benefit from screening when the trial reports in 2014/2015.
Human epididymis protein 4 (HE4), a precursor of human epididymis protein, has been recently identified as a new promising serum biomarker for ovarian carcinoma. We performed a systematic review of studies that investigated the use of HE4 in the diagnosis of ovarian cancer in patients with pelvic or gynecological masses. We also evaluated the diagnostic performance of HE4 for differentiating between patients with benign gynecological disease and those with ovarian cancer.
We searched PubMed database (1990-2011) to collect articles in English that evaluated the diagnostic value of HE4 in patients with gynecological or pelvic masses. Two reviewers independently assessed the methodological quality of each study using the quality assessment of diagnostic accuracy studies tool. The data were analyzed using Meta-Disc1.4 software. Meta-analysis of the reported sensitivity and specificity of each study and summary receiver operating characteristic (SROC) curve were performed.
A total of 9 studies involving 1807 women were included. When the control group was composed of healthy women, the pooled sensitivity and specificity for HE4 in diagnosing ovarian cancer were 83% (95% confidence interval [CI], 77%-88%) and 90% (95% CI, 87%-92%), respectively. The area under the SROC curve was 0.9271. When the control group was composed of women with benign disease, the pooled sensitivity and specificity for HE4 were 74% (95% CI, 69%-78%) and 90% (95% CI, 87%-92%). The area under the SROC curve was 0.8853.
The current analysis indicated that HE4 may be a valuable marker in the diagnosis of ovarian carcinoma. Serum HE4 detection is not only a useful preoperative test for predicting the benign or malignant nature of pelvic masses but has a potential to be used as an initial step in ovarian cancer screening strategy.
BACKGROUND
Ovarian carcinoma is apparently restricted for a long time to the peritoneal cavity. However, about 50% of patients with a surgically documented complete intraabdominal response experience later recurrence. Occult hematogenous micrometastases are common to most epithelial malignancies and have recently been found in 30% of bone marrow samples of ovarian carcinoma patients, as examined by immunocytochemistry. Moreover, these findings were associated with poor progression-free and overall survival. The aim of the current study was to evaluate the possible prognostic significance of tumor cells detected in the peripheral blood and bone marrow of ovarian carcinoma patients by an immunomagnetic method.METHODS
In a total of 90 patients with histologically proven epithelial ovarian carcinoma, blood and (in 73 cases) bone marrow samples were taken. Tumor cells were identified by a microbead coated with the antibody MOC-31, which recognizes an epitope regularly expressed on ovarian carcinoma cells.RESULTSThe authors detected carcinoma cells in the bone marrow in 21% of ovarian carcinoma patients, and in the peripheral blood in 12% of patients. Mean overall survival was 25 and 28 months for patients with or without circulating tumor cells, respectively.CONCLUSIONS
Ovarian carcinoma cells seem to reach peripheral circulation more frequently than expected. However, in contrast to an earlier report, detection of tumor cells in the bone marrow and/or blood was not associated with poor prognosis in ovarian carcinoma patients. This discrepancy remains unexplained, but characterization of circulating ovarian carcinoma cells for their malignant and metastatic capacity is clearly warranted. Cancer 2002;94:707–12. © 2002 American Cancer Society.DOI 10.1002/cncr.10250
The search for clinically relevant tissue and body fluid–borne biomarkers has never been more important to the war on cancer. New -omic technologies can measure hundreds to tens of thousands of analytes at once. The field is intoxicated with the prospect of coupling these platforms, and the
Recent microarray studies have shown that the expression of hepatocyte nuclear factor-1β (HNF-1β) was significantly up-regulated in clear cell carcinoma (CCC) of the ovary. HNF-1β may be a useful marker for CCC in peritoneal fluid cytology. We designed an experimental study using three CCC cell lines to evaluate the influence of alcohol fixation or air drying on immunocytochemistry for HNF-1β. Each cell line was cultured on chamber slides or transplanted into the abdominal cavity of nude mice, then the slides or ascites smears of nude mice were immunostained with or without microwave-mediated epitope retrieval. Immunoreactivity with HNF-1β, which was either alcohol-fixed or air-dried, was noticeably improved after microwave heating. In contrast, two serous adenocarcinoma cell lines never showed immunoreactivity. Based on these results, 21 archival Papanicolaou-stained slides of peritoneal fluid (5 CCCs, 12 serous, 2 mucinous, and 2 endometrioid adenocarcinomas) were decolorized and immunostained under heating pretreatment. Five of 5 CCCs were distinctively positive for HNF-1β, whereas all non-CCC ovarian cancers were negative for this protein. The present study clearly demonstrated that heating epitope retrieval improved the immunoreactivity for a nuclear protein in alcohol-fixed or air-dried cytology specimens. HNF-1β is likely to be helpful for the diagnosis of CCC in the peritoneal fluid. Diagn. Cytopathol. 2007;35:193–197. © 2007 Wiley-Liss, Inc.
Plasma of cancer patients contains cell-free tumor DNA that carries information on tumor mutations and tumor burden. Individual mutations have been probed using allele-specific assays, but sequencing of entire genes to detect cancer mutations in circulating DNA has not been demonstrated. We developed a method for tagged-amplicon deep sequencing (TAm-Seq) and screened 5995 genomic bases for low-frequency mutations. Using this method, we identified cancer mutations present in circulating DNA at allele frequencies as low as 2%, with sensitivity and specificity of >97%. We identified mutations throughout the tumor suppressor gene TP53 in circulating DNA from 46 plasma samples of advanced ovarian cancer patients. We demonstrated use of TAm-Seq to noninvasively identify the origin of metastatic relapse in a patient with multiple primary tumors. In another case, we identified in plasma an EGFR mutation not found in an initial ovarian biopsy. We further used TAm-Seq to monitor tumor dynamics, and tracked 10 concomitant mutations in plasma of a metastatic breast cancer patient over 16 months. This low-cost, high-throughput method could facilitate analysis of circulating DNA as a noninvasive "liquid biopsy" for personalized cancer genomics.
Objectives:
The invasive growth of circulating tumor cells (CTCs) propagates cancer metastasis. The aims of this study were to evaluate the association of invasive CTCs, detected by a novel cell invasion assay, with disease stage, CA-125 level and patient survival.
Methods:
Peripheral blood samples from 71 patients undergoing evaluation for ovarian malignancy were assessed for the presence of invasive CTCs using a cell invasion assay that enriches and identifies tumor cells with a cell adhesion matrix (CAM). Invasive CTCs were identified as cells exhibiting CAM invasion (CAM+) and expressing standard epithelial markers (Epi+).
Results:
43 (60.6%) patients had detectable CTCs: 0/5 benign patients, 1/10 (10%) early stage, 39/52 (73.1%) late stage and 3/4 (75%) unstaged patients (p-value <0.001). CTC counts ranged from 0-149 CTCs/ml with stage III/IV patients exhibiting significantly higher mean counts (41.3 CTCs/ml) than stage I/II patients (6.0 CTCs/ml) and benign patients (0 CTCs/ml, p-value=0.001). A positive correlation between CTC count and CA-125 level was observed (Spearman correlation coefficient r=0.309, p-value=0.035). Kaplan-Meier curves revealed a significant decrease in disease-free survival in patients with detectable CTCs (median survival 15.0 months vs. 35.0 months, log-rank p-value=0.042). Tumor grade and tumor histology did not influence CTC detection.
Conclusions:
Invasive CTCs can be detected in a majority of epithelial ovarian cancer patients and may predict shorter disease-free survival. Furthermore, higher CTC counts may reflect later stage disease and higher CA-125 levels.
Autoantibodies represent an attractive biomarker for diagnostic assays principally due to the stability of immunoglobulin in patient serum facilitating measurement with conventional assays. Immune responses to tumorigenesis may facilitate detection of ovarian cancer in the early stages of the disease with identification of a panel of tumour specific autoantibodies. Despite the reporting of many tumour associated autoantibodies using arrays of tumour antigens, this has not led to the advance in diagnostic capability as rapidly as was initially expected. Here we examine the potential diagnostic utility of candidate autoantibody biomarkers identified via screening of serum samples on a high content human protein array from a unique cohort of early stage and late stage ovarian cancer patients. We analyse the performance of autoantibodies to the tumour suppressor protein p53 and the novel autoantigens alpha adducin and endosulfine alpha identified in our array screen. Each antigen has different performance characteristics using conventional ELISA format and Western blot immunoassay. The high attrition rate of promising autoantigens identified by array screening can in part be explained by the presentation of the epitope of the antigen in the subsequent method of validation and this study provides directions on maximising the potential of candidate biomarkers. This article is part of a Special Issue entitled: Translational Proteomics.
Ovarian cancer is the most lethal gynecological cancer. Due to few early symptoms and a lack of early detection strategies, most patients are diagnosed with advanced-stage disease. Most of these patients, although initially responsive, eventually develop drug resistance. In this chapter, epigenetic changes in ovarian cancer are described. Various epigenetic changes including CpG island methylation and histone modification have been identified in ovarian cancer. These aberrations are associated with distinct disease subtypes and present in circulating serum of ovarian cancer patients. Several epigenetic changes have shown promise for their diagnostic, prognostic, and predictive capacity but still need further validation.
In contrast to DNA mutations and deletions, epigenetic modifications are potentially reversible by epigenetic therapies. Promising preclinical studies show epigenetic drugs to enhance gene re-expression and drug sensitivity in ovarian cancer cell lines and animal models.
Discrimination between epithelial ovarian cancer (EOC) and benign ovarian tumor (BOT) has always been difficult in clinical practice. We investigated the application of metabolomics in distinguishing EOC and BOT and tried to discover valuable biomarkers.
Plasma metabolomic profiling was performed using ultra-performance liquid chromatography mass spectrometry (UPLC/MS). Partial least-squares discriminant analysis was employed to classify EOC and BOT, and reveal their metabolic differences. The area under the receiver-operating characteristic curve (AUC) was utilized to evaluate the predictive performance of the metabolic profiles for external validation set.
The metabolomic profiles consisting of 535 metabolites revealed a clear separation between EOC and BOT, with AUC of 0.86 for the external validation set. 6 metabolic biomarkers were identified, and the plasma concentrations of the 4 ascertained biomarkers (L-tryptophan, LysoPC(18:3), LysoPC(14:0), and 2-Piperidinone) were lower in EOC patients than those in BOT patients. Among them, tryptophan and LysoPC have been suspected to participate in cancer progression, and 2-Piperidinone might be a novel biomarker for EOC.
Metabolomics could be used to discriminate EOC from BOT in clinical practice, and the identified metabolic biomarkers might be important on investigating the biological mechanisms of EOC.
One of the deadly hallmarks of cancer is its ability to prosper within the constraints of the host immune system. Recent advances in immunoproteomics and high-throughput technologies have lead to profiling of the antibody repertoire in cancer patients. This in turn has lead to the identification of tumour associated antigens/autoantibodies. Autoantibodies are extremely attractive and promising biomarker entities, however there has been relatively little discussion on how to interpret the humoral immune response. It may be that autoantibody profiles hold the key to ultimately uncovering neoplastic associated pathways and through the process of immunosculpting the tumour may have yielded an immune response in the early stages of malignant tumour development. The aim of this review is to discuss the utility of the autoantibody response that is elicited as a result of malignancy and discuss the advantages and limitations of autoantibody profiling. This article is part of a Special Issue entitled: Translational Proteomics.
In this study, electrochemical immunoassay was introduced into the recently proposed microfluidic paper-based analytical device (μPADs). To improve the performance of electrochemical immunoassay on μPAD for point-of-care testing (POCT), a novel wax-patterned microfluidic paper-based three-dimensional electrochemical device (3D-μPED) was demonstrated based on the multi-walled carbon nanotubes (MWCNTs) modified μPAD. Using typical HRP-O-Phenylenediamine-H(2)O(2) electrochemical system, a sandwich immunoassay on this 3D-μPED for sensitive diagnosis of two tumor markers simultaneously in real clinical serum samples was developed with a linear range of 0.001-75.0 UmL(-1) for cancer antigen 125 and 0.05-50.0 ngmL(-1) for carcinoembryonic antigen. In addition, this 3D-μPED can be easily integrated and combined with the recently emerging paper electronics to further develop simple, sensitive, low-cost, disposable and portable μPAD for POCT, public health and environmental monitoring in remote regions, developing or developed countries.
Evaluate and compare the effectiveness of CA125, HE4, Mesothelin and MMP7 marker levels to monitor ovarian cancer patients after surgery and chemotherapy. Evaluate the lead time of a rise of marker levels before recurrence.
The study consists of 23 patients with advanced stage ovarian/fallopian tube cancer. Blood was drawn after front line surgery and chemotherapy treatment and at 3 month intervals thereafter. One patient had chemoresistant disease, two patients remained in remission and 20 patients had recurring disease and were used for marker evaluation.
In five patients HE4 was the only marker to elevate before recurrence with a lead time of up to 4½ months including one patient who did not have a CA125 response at all. In a further two patients, HE4 increased before CA125 did. In four of these seven patients, HE4 levels were consistently at or above threshold during remission when both CA125 and imaging results were negative. MMP7 elevated before recurrence in one patient who was negative for the other markers. Mesothelin elevated in two patients who were also positive for CA125 and HE4.
HE4 can predict ovarian cancer recurrence earlier than CA125 and it can be elevated in patients that do not express CA125 at sufficient levels to make a clinical decision. MMP7 and Mesothelin have lower potential as markers for ovarian cancer recurrence to complement CA125. A failure of HE4 levels to normalize at completion of standard therapy may indicate a poor prognosis.
Ovarian cancer is the most lethal gynecological cancer due to lack of clear symptom and reliable screening biomarker in the early stage. The capability to detect the initiation of malignancy with a sensitive and effective approach is one of the most desirable goals for ovarian cancer therapy. In this study, we spearheaded noninvasive detection of ovarian cancer by salivary transcriptomic biomarkers, and evaluated the clinical utilities of discovered biomarkers using a clinical case-control study. To find salivary mRNA biomarkers, salivary transcriptomes in 11 ovarian cancer patients and 11 matched controls were profiled by Affymetrix HG-U133-Plus-2.0 array. The biomarker candidates selected from the microarray results were then subjected to clinical validation by RT-qPCR using an independent sample cohort including 21 ovarian cancer patients and 35 healthy controls. Seven downregulated mRNA biomarkers were validated. The logistic regression model revealed the combination of five validated biomarkers (AGPAT1, B2M, BASP2, IER3, and IL1B) can significantly discriminate ovarian cancer patients (n = 21) from the healthy controls (n = 35), yielding a receiver operating characteristic plot, area under the curve value of 0.909 with 85.7% sensitivity and 91.4% specificity. In summary, we have demonstrated that the RNA signatures in saliva could serve as biomarkers for detection of ovarian cancer with high sensitivity and specificity. This emerging approach with high-throughput, noninvasive, and effective advantages provides a feasible means for detection of systemic cancer, and opens a new avenue for early disease detection.
This study was designed to determine whether serum HE4 is an independent prognostic factor in ovarian cancer patients.
We measured HE4 in pretreatment serum samples from 80 women with epithelial ovarian cancer, using an enzyme-linked immunosorbent assay. The results were correlated with clinical data.
The median serum HE4 level in ovarian cancer patients was 98.7 (range, 80.3-222.8) pg/ml. Elevated serum HE4 levels before therapy significantly correlated with a poorer progression free survival (log-rank test, P = 0.017). Multivariate analysis revealed serum HE4 to be an independent prognostic factor for progression-free survival (P = 0.036). In multivariate regression analysis, high serum HE4 levels significantly correlated with high tumor grade and serous histology (P = 0.004 and 0.017). In addition, high serum HE4 levels were significantly associated with residual tumor size and operative time (P = 0.003 and 0.033).
Pretreatment serum HE4 seems to be an additional factor for predicting the outcome of patients with epithelial ovarian cancer. Due to its independence from established prognostic factors, serum HE4 may provide additional prognostic information.
It is often difficult to distinguish a benign pelvic mass from a malignancy and tools to help referring physician are needed. The purpose of this study was to validate the Risk of Ovarian Malignancy Algorithm in women presenting with a pelvic mass.
This was a prospective, multicenter, blinded clinical trial that included women who presented to a gynecologist, a family practitioner, an internist, or a general surgeon with an adnexal mass. Serum HE4 and CA 125 were determined preoperatively. A Risk of Ovarian Malignancy Algorithm score was calculated and classified patients into high-risk and low-risk groups for having a malignancy. The sensitivity, specificity, negative predictive value, and positive predictive value of the Risk of Ovarian Malignancy Algorithm were estimated.
A total of 472 patients were evaluated with 383 women diagnosed with benign disease and 89 women with a malignancy. The incidence of all cancers was 15% and 10% for ovarian cancer. In the postmenopausal group, a sensitivity of 92.3% and a specificity of 76.0% and for the premenopausal group the Risk of Ovarian Malignancy Algorithm had a sensitivity of 100% and specificity of 74.2% for detecting ovarian cancer. When considering all women together, the Risk of Ovarian Malignancy Algorithm had a sensitivity of 93.8%, a specificity of 74.9%, and a negative predictive value of 99.0%.
The use of the serum biomarkers HE4 and CA 125 with the Risk of Ovarian Malignancy Algorithm has a high sensitivity for the prediction of ovarian cancer in women with a pelvic mass. These findings support the use of the Risk of Ovarian Malignancy Algorithm as a tool for the triage of women with an adnexal mass to gynecologic oncologists.
II.
Human epididymis protein 4 (HE4) has attracted a lot of interest as a relatively novel biomarker for ovarian carcinoma. Research focus has been directed at HE4 as a diagnostic tool with potential for better triage of women with adnexal masses but the prognostic aspect of HE4 in ovarian cancer patients remains to be elucidated. The aim of the present study was to investigate the prognostic value of prechemotherapy serum HER2, cancer antigen 125 (CA125), and HE4 levels in ovarian cancer patients receiving standard combination chemotherapy.
Serum from 139 patients with newly diagnosed ovarian cancer was analyzed for HER2, CA125, and HE4 using enzyme-linked immunosorbent assay assays. Samples were collected just before first-line chemotherapy, and all patients were treated with carboplatin-paclitaxel combination chemotherapy.
Increasing levels of serum HE4 (grouped into quartiles) was significantly associated with worse progression-free survival (PFS) (P < 10) and overall survival (P < 10). After adjustment in the Cox model, HE4 serum levels remained an independent prognostic parameter for PFS, with a hazard ratio of 1.77 (95% confidence interval, 1.03-3.04; P = 0.040) for patients with HE4 levels above the median compared with patients with HE4 levels below the median. The shorter PFS for patients with high levels of HE4 also translated into an independent significant difference in overall survival (hazard ratio, 3.17 [95% confidence interval, 1.41-7.10]; P = 0.005).Serum HER2 and CA125 levels did not demonstrate an independent prognostic value.
High levels of serum HE4 is a strong and independent indicator of worse prognosis in epithelial ovarian cancer patients.
We sought to assess the ability of human epididymis protein 4 (HE4) and CA-125 to distinguish among benign, borderline, and malignant pelvic masses in premenopausal women.
We conducted a subset analysis of data from a prospective clinical trial that enrolled women undergoing surgery for an adnexal mass. Diagnostic performance of CA-125 and HE4 for epithelial ovarian cancer (EOC) detection in premenopausal women was determined.
Of 229 premenopausal patients, 195 (85%) had benign masses, 18 (8%) had EOC, and 16 (7%) had borderline ovarian tumor. The sensitivity of CA-125 and HE4 for EOC detection was 83.3% and 88.9%, respectively. The specificity of CA-125 and HE4 was 59.5% and 91.8%, respectively. A normal HE4 level ruled out invasive cancer in 98% of women with an elevated CA-125.
HE4 offers superior specificity compared to CA-125 for the differentiation of benign and malignant adnexal masses in premenopausal women.
To examine the value of individual and combinations of ovarian cancer associated blood biomarkers for the discrimination between plasma of patients with type I or II ovarian cancer and disease-free volunteers.
Levels of 14 currently promising ovarian cancer-related biomarkers, including CA125, macrophage inhibitory factor-1 (MIF-1), leptin, prolactin, osteopontin (OPN), insulin-like growth factor-II (IGF-II), autoantibodies (AAbs) to eight proteins: p53, NY-ESO-1, p16, ALPP, CTSD, B23, GRP78, and SSX, were measured in the plasma of 151 ovarian cancer patients, 23 with borderline ovarian tumors, 55 with benign tumors and 75 healthy controls.
When examined individually, seven candidate biomarkers (MIF, Prolactin, CA-125, OPN, Leptin, IGF-II and p53 AAbs) had significantly different plasma levels between type II ovarian cancer patients and healthy controls. Based on the receiver operating characteristic (ROC) curves constructed and area under the curve (AUC) calculated, CA125 exhibited the greatest power to discriminate the plasma samples of type II cancer patients from normal volunteers (AUC 0.9310), followed by IGF-II (AUC 0.8514), OPN (AUC 0.7888), leptin (AUC 0.7571), prolactin (AUC 0.7247), p53 AAbs (AUC 0.7033), and MIF (AUC 0.6992). p53 AAbs levels exhibited the lowest correlation with CA125 levels among the six markers, suggesting the potential of p53 AAbs as a biomarker independent of CA125. Indeed, p53 AAbs increased the AUC of ROC curve to the greatest extent when combining CA125 with one of the other markers. At a fixed specificity of 100%, the addition of p53 AAbs to CA125 increased sensitivity from 73.8% to 85.7% to discriminate type II cancer patients from normal controls. Notably, seropositivity of p53 AAbs is comparable in type II ovarian cancer patients with negative and positive CA125, but has no value for type I ovarian cancer patients.
p53 AAbs might be a useful blood-based biomarker for the detection of type II ovarian cancer, especially when combined with CA125 levels.
Serial circulating tumor cell (CTC) counts have demonstrated predictive and prognostic value in patients with metastatic breast, colorectal, and prostate cancer. In a phase III study of pegylated liposomal doxorubicin (PLD) with trabectedin vs. PLD for relapsed ovarian cancer, we evaluated the correlation, if any, between numbers of CTCs and progression free survival, (PFS) and overall survival (OS).
CTCs were isolated from peripheral blood (10 mL) using the CellSearch system and reagents (Veridex). A CTC is defined as EpCAM+, cytokeratin+, CD45-, and is positive for the nuclear stain DAPI. The normal reference range for CellSearch is <2 CTC/7.5 mL of blood. Hazard ratios adjusted for known prognostic factors were estimated by Cox regression.
Two-hundred sixteen patients had baseline CTC measurements of which 111 (51.4%) were randomized to the trabectedin+PLD arm; 143/216 patients (66.2%) were platinum-sensitive. Thirty-one of 216 patients (14.4%) had 2 or more CTCs detected prior to the start of therapy (range 2-566). Univariate Cox regression analyses indicated that patients with ≥2 CTCs prior to therapy had 1.89- (p=0.003) and 2.06-fold (p=0.003) higher risk for progression and death respectively. Multivariate analyses that include baseline CA-125, platinum sensitivity status, largest diameter lesion, number of tumor lesions, ECOG PS, and tumor grade show that patients with elevated baseline CTC had 1.58- (p=0.058) and 1.54-fold (p=0.096) higher risk for progression and death respectively.
Results from this study indicate that elevated numbers of CTCs impart an unfavorable prognosis for ovarian cancer patients.
Disseminated tumor cells (DTCs) in the bone marrow (BM) were shown to be of prognostic significance in gynecological cancers. Bone marrow aspiration is less accepted by patients compared with blood drawing. In this pilot study, we applied the AdnaTest BreastCancer based on immunomagnetic enrichment, targeting common antigens on epithelial gynecological cancers, followed by multiplex reverse transcriptase-polymerase chain reaction for selection and detection of circulating tumor cells (CTCs) in the blood of 122 ovarian cancer patients at primary diagnosis and/or after platinum-based chemotherapy. Results were compared with detection of DTC in BM.
Ten-milliliter blood was obtained before surgery (n=86) and/or after chemotherapy (n=70) and analyzed for CTC with the AdnaTest BreastCancer for the detection of EpCAM-, MUC-1-, and HER-2-transcripts. CA 125 was assessed in an additional single-plex reverse transcriptase-polymerase chain reaction. Bone marrow aspirates were analyzed in duplicate by immunocytochemistry using the pan-cytokeratin antibody A45-B/B3.
Before surgery, CTCs were detected in 19% of patients, expressing EpCAM (31%), MUC-1 (50%), HER-2 (31%), and CA 125 (50%), respectively. After chemotherapy, the overall detection rate for CTC was 27%, thereof EpCAM (68%), MUC-1 (47%), HER2 (21%), and CA 125 (37%). The overall detection rate for DTC in the BM was 35% before surgery and 31% after therapy. A comparison between DTC and CTC resulted in a concordance rate of 59% before surgery and 56% after chemotherapy. CTC positivity significantly correlated with shorter overall survival before surgery (P=0.0054) and after chemotherapy (P=0.047).
This methodological approach might help to identify molecular targets for specific biological therapies. Blood analysis could give additional information complimentary to that obtained by DTC.
The aim of this study was to use a two steps strategy metabolomics to screen/identify and validate novel metabolic biomarker(s) for epithelial ovarian cancer (EOC). In the screening step, serum samples from 27 healthy women, 28 benign ovarian tumors, and 29 EOCs were analyzed by using LC-MS based nontargeted metabolomics. The three groups were separated with OSC filtered PLS-DA model, and six metabolites (27-nor-5β-cholestane-3,7,12,24,25 pentol glucuronide (CPG), phenylalanine, glycocholic acid, propionylcarnitine, Phe-Phe and Lyso PC (18:2)) were considered as potential biomarker candidates. In the validation step, the six metabolites were analyzed in targeted metabolomics by LC-selective ion monitoring mass spectrometry in another 685 serum samples with various clinical backgrounds. As a result, CPG was evaluated to be a potential biomarker and its content was elevated in EOC tissues compared with benign ovarian tumor tissues (p = 0.0005). Besides, CPG levels were found to be up-regulated in early stage EOC and in the three types of EOC histological types. Other variables such as nonovarian diseases, medicine consumption, gynecological inflammations, and menopausal state did not interfere in using CPG as diagnosis marker. CPG was found to be complementary to CA125. Our findings suggest that CPG can be considered a statistical relevant biomarker of EOC, ready for early stage detection.
Multiple omics researches in the past two decades have identified over 200 potential biomarkers for ovarian cancer. Discoveries during the 1990s were more focused on clinicopathology-based biomarkers that were targeted to support diagnosis, but the emphasis has shifted to the identification of prognostic biomarkers in the past 10 years. The post-genomic era has opened the door for personalized cancer treatments and the trend of discovery is moving forward to identify more stratified biomarkers to accurately predict the progression of disease, as well as efficacy biomarkers to precisely determine drug response. To better meet future challenges, biomedical research needs the reformed and standardized infrastructure of tissue banks/biorepositories, with national and international initiatives. Of the hundreds of biomarker candidates for ovarian cancer, only a small number are actively being validated with clinical samples, owing to the lack of biomaterials that are linked with accurate clinical data. The purpose of this article is to present selected biomarkers from the past 20 years of ovarian cancer research, placing special emphasis on biomarkers that are strongly associated with positive or negative clinical outcomes. The article also presents a global view of all known potential biomarkers and mutations for ovarian cancer from NCI's Cancer Gene Index developed by Sophic, and Sanger's Catalogue of Somatic Mutations in Cancer database.