Detection In Peripheral Blood Research Articles

Detection and Clinical Significance of Circulating Tumor Cells in Colorectal Cancer--20 Years of Progress.

Circulating tumor cells (CTC) may be defined as tumor- or metastasis-derived cells that are present in the bloodstream. The CTC pool in colorectal cancer (CRC) patients may include not only epithelial tumor cells, but also tumor cells undergoing epithelial-mesenchymal transition (EMT) and tumor stem cells. A significant number of patients diagnosed with early stage CRC subsequently relapse with recurrent or metastatic disease despite undergoing "curative" resection of their primary tumor. This suggests that an occult metastatic disease process was already underway, with viable tumor cells being shed from the primary tumor site, at least some of which have proliferative and metastatic potential and the ability to survive in the bloodstream. Such tumor cells are considered to be responsible for disease relapse in these patients. Their detection in peripheral blood at the time of diagnosis or after resection of the primary tumor may identify those early-stage patients who are at risk of developing recurrent or metastatic disease and who would benefit from adjuvant therapy. CTC may also be a useful adjunct to radiological assessment of tumor response to therapy. Over the last 20 years many approaches have been developed for the isolation and characterization of CTC. However, none of these methods can be considered the gold standard for detection of the entire pool of CTC. Recently our group has developed novel unbiased inertial microfluidics to enrich for CTC, followed by identification of CTC by imaging flow cytometry. Here, we provide a review of progress on CTC detection and clinical significance over the last 20 years.

Research of Xinfeng Capsule on Pulmonary Function in Adjuvant Arthritis Rats Based on the Effects of B, T Cell Immunity

Objective: to observe the effects of Xinfeng Capsule on adjuvant arthritis rat model of pulmonary function, B, T cell immunity. Methods 40 rats were randomly divided into normal, model, Tripterygium Glycosides tablet and XFC group. Except the normal group, the other rat’s right rear paw intradermal injection of Freund’s complete adjuvant induced arthritis in 0.1 mL. Inflammation induced 19 d after administration, the normal group, the model groups were given physiological saline, rats in the treatment group were given of Xinfeng Capsule (2.4 g/kg), Tripterygium Glycosides tablet (10 mg/kg), once a day, continuous administration of 30D. Joint inflammation in rats was observed performance, flow cytometry detection of peripheral blood B, T lymphocyte attenuator (BTLA), regulatory T cells (Treg) and surface markers of forkhead transcription factor (FoxP3) expression was detected by Western blotting, synovial membrane, lung tissue BTLA, FoxP3. Results compared with the normal group, model group of rat paw swelling degree, arthritis index increased; pulmonary function, reduce the expression levels of peripheral blood BTLA, Tregs and BTLA in pulmonary tissue, synovium, FoxP3 (P <0.05 or P <0.01). While the XFC group rat paw swelling and arthritis index lower than the model group, pulmonary function, BTLA, Tregs and FoxP3 increased (P <0.05 or P <0.01); Xinfeng capsule group is better than control group (P <0.05) medicine Tripterygium wilfordii. Conclusion of Xinfeng capsule may be through upregulation of BTLA, Tregs, and FoxP3 expression, reduce the stimulation of inflammatory mediators in the lung tissue and improve the lung function of rats with adjuvant arthritis.

Concordance and Prognostic Significance of Minimal Residual Disease Detection in Peripheral Blood and Bone Marrow Samples of Infants with MLL-rearranged Acute Lymphoblastic Leukemia Treated By MLL-Baby Protocol

Background. Minimal residual disease (MRD) is powerful tool for prediction of treatment outcome in leukemia patients of various age groups, including infants with acute lymphoblastic leukemia (ALL). In the vast majority of cases only bone marrow (BM) samples are used for MRD detection.Objective. To estimate prognostic significance of MRD in peripheral blood (PB) and BM by qualitative detection of different MLL fusion gene transcripts in infant ALL enrolled into MLL-Baby protocol.Methods. Fifty three infants (20 boys and 33 girls) with median age of 5.3 months (range 0.03-11.80) and defined MLL rearrangements were included in the current study. Among them there were 25 patients (47.2%) carrying MLL-AFF1 fusion gene transcripts, 10 (18.9%) MLL-MLLT3-positive cases, 9 (17.0%) MLL-MLLT1-positive cases, 5 (9.4%) MLL-MLLT10-positive cases and 4 (7.5%) MLL-EPS15-positive ones. MRD evaluation was performed by detection of MLL fusion gene transcripts in BM and PB samples using real-time PCR and nested RT-PCR with sensitivity non-less than 1E-04. MRD-negativity was defined as absence of fusion gene transcripts in both assays. Median of follow-up period in the observed group was 5.2 years. Time points (TP) for MRD assessment were as follows: day 15 of remission induction (TP1), at the end of remission induction (TP2), after each course of ATRA administration (TP3-TP7). Informed consent was obtained in all cases.Results. We estimated 142 paired BM/PB samples. 77 samples were double positive, 43 were double negative Thus concordance between MRD results in BM and PB samples achieved 84.5%. Concordance varied between different TPs of MLL-Baby protocol from 79.0% to 100%. The highest concordance rate was at TP4 and TP7 (92.3% and 100%, respectively). Interestingly, all discrepant results (22 samples 15.5%) were BM-positive/PB-negative. Median level of ABL gene, used for normalization, was similar in BM and PB samples (4.85E+04 vs 4.95E+04, respectively, p=0.760). Evaluation of prognostic significance of MRD in BM in TP1-TP7 revealed that TP4 was the earliest TP when discriminative data between MRD-positive and MRD-negative patients were obtained. MRD-positivity at TP4 in BM led to unfavorable outcome. Event-free survival was significantly lower in MRD-positive group (n=22) in comparison to MRD-negative one (n=31) (0.06±0.06 vs 0.70±0.09 p=0.0001), while cumulative incidence of relapse in MRD-positive patients was remarkably higher (0.92±0.01 vs 0.29±0.08, p<0.0001). MRD-positivity at this TP in BM was the only significant factor in the diagnostic model where initial risk factors (age at diagnosis, initial WBC count, immunophenotype, CNS disease, presence of MLL-AF4) were combined to response criteria (number of blast cells at day 8 of dexamethasone prophase and MRD in BM at TP4) (Table). The only TP when MRD data obtained from PB samples had prognostic value was TP6. In this TP cumulative incidence of relapse in MRD-positive patients was significantly higher in comparison to MRD-negative ones (0.88±0.11 vs 0.25±0.13, respectively, p=0.003). However these data did not bring any extra advantages as compared to TP4 in BM.Conclusions. Despite high qualitative concordance rate between MRD detection in BM and PB samples we could not show prognostic value of MRD monitoring in PB by fusion gene transcripts. Univariate and multivariate analysis revealed that MRD-positivity at TP4 in BM was the only significant and independent prognostic factor of unfavorable outcome in the observed group of patients. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Peripheral Blood for Epidermal Growth Factor Receptor Mutation Detection in Non-Small Cell Lung Cancer Patients

OBJECTIVE: It is important to analyze and track Epidermal Growth Factor Receptor (EGFR) mutation status for predicting efficacy and monitoring resistance throughout EGFR-tyrosine kinase inhibitors (TKIs) treatment in non-small cell lung cancer (NSCLC) patients. The objective of this study was to determine the feasibility and predictive utility of EGFR mutation detection in peripheral blood. METHODS: Plasma, serum and tumor tissue samples from 164 NSCLC patients were assessed for EGFR mutations using Amplification Refractory Mutation System (ARMS). RESULTS: Compared with matched tumor tissue, the concordance rate of EGFR mutation status in plasma and serum was 73.6% and 66.3%, respectively. ARMS for EGFR mutation detection in blood showed low sensitivity (plasma, 48.2%; serum, 39.6%) but high specificity (plasma, 95.4%; serum, 95.5%). Treated with EGFR-TKIs, patients with EGFR mutations in blood had significantly higher objective response rate (ORR) and insignificantly longer progression-free survival (PFS) than those without mutations (ORR: plasma, 68.4% versus 38.9%, P = 0.037; serum, 75.0% versus 39.5%, P = 0.017; PFS: plasma, 7.9 months versus 6.1 months, P = 0.953; serum, 7.9 months versus 5.7 months, P = 0.889). In patients with mutant tumors, those without EGFR mutations in blood tended to have prolonged PFS than patients with mutations (19.7 months versus 11.0 months, P = 0.102). CONCLUSIONS: EGFR mutations detected in blood may be highly predictive of identical mutations in corresponding tumor, as well as showing correlations with tumor response and survival benefit from EGFR-TKIs. Therefore, blood for EGFR mutation detection may allow NSCLC patients with unavailable or insufficient tumor tissue the opportunity to benefit from personalized treatment. However, due to the high false negative rate in blood samples, analysis for EGFR mutations in tumor tissue remains the gold standard.

Expression of MAGE genes peripheral blood of gastric cancer patients and its relation to lymph node metastasis and prognosis

Objective In the detection of peripheral blood in patients with gastric cancer the melanoma antigen A1 (MAGEA1) and the melanoma antigen A3 (MAGEA3) gene expression,combined with the conditions of patients with postoperative follow-up analysis MAGE gene expression and the relationship between gastric cancer metastasis and prognosis.Methods Peripheral blood was obtained from 40 patients with gastric carcinoma and 20 patients with benign diseases.The mRNA of the MAGEA1 and MAGEA3 genes in the peripheral blood mononuclear cells(PBMC) was detected by RT-PCR,meanwhile the MAGEA1 and MAGEA3 transcripts in gastric carcinoma tissues and corresponding nomal tissue were detected.In contrast with CEA gene,we detected its expression by nested RT-PCR.Results Of the 40 gastric carcinoma patients,MAGEA1 and MAGEA3 mRNA were positive in 47.5％ (17/40) and 25.0％ (10/40) of PBMC,respectively,and in 65.0％ (26/40) and 30.0％ (12/40) of gastric carcinoma tissues,the corresponding paracancerous gastric tissues were not expressed,of which 4 cases of peripheral blood both expression.In the PBMC of 40 gastric carcinoma patients,62.5％ (25/40) samples were detected to express at least one type of MAGE mRNA,the detection of CEA gene mRNA in peripheral blood of the patients in PBMCwas 65.0％ (26/40),20 patients with non neoplastic diseases of PBMC were not detected in the MAGE gene mRNA.There were 22.5％ (9/40) of 40 gastric carcinoma patients with CEA gene expression in PBMC.The detection rate of the MAGE gene of mRNA PBMC in patients with gastric cancer was not related with the lymph node metastasis,TNM stage,and the degree of tumor differentiation,depth of invasion,tumor size,distant metastasis.Conclusion Gastric cancer patients with peripheral blood MAGE gene expression is closely related to the tumor lymph node metastasis,TNM staging,express positive patients with poor prognosis. Key words: Gastric neoplasms ; Melanoma antigen gene ; Peripheral blood mononuclear cells; Prognosis

Role of estradiol, prolactin, and inhibin B in normal male ejaculation

With a lack of published hormonal parameters associated with ejaculation in healthy men, we studied hormones with negative feedback on the hypothalamic-pituitary-gonadal (HPG) axis. Prolactin, inhibin B, and estradiol, which may exert rapid effects via G-coupled receptors (GPR30) during ejaculation (i.e. epididymal motility regulation), were investigated since peripheral blood detection may affect central spinal cord motor generator or bulbocavernosus muscle (BCM).

Serum tryptase correlates with the KIT D816V mutation burden in adults with indolent systemic mastocytosis

Systemic mastocytosis (SM) is characterized by the growth of neoplastic mast cells (MCs). Most adults with indolent SM carry the KIT D816V mutation. We recently introduced the D816V+ allele fraction as a disease marker in SM using a sensitive and quantitative KIT D816V mutation analysis that consistently allows mutation detection in peripheral blood (PB) and bone marrow (BM). The D816V+ allele fraction represents a quantitative measure which allows KIT D816V-positivity to be analyzed as a continuous variable instead of a categorical variable (negative/positive) as previously described. Serum tryptase represents an established disease marker in SM, and it remains to be tested whether tryptase and the D816V+ allele fraction are associated or represent independent disease markers. In this study, correlation analysis between serum tryptase, the D816V+ allele fraction in PB and BM, and the MC fraction was performed in 57 indolent systemic mastocytosis (ISM) patients. We detected significant correlations between the D816V+ allele fraction, the mature neoplastic MC fraction, and serum tryptase which represent three different biological measures of disease burden. Mutation analysis was performed in two or more PB samples in 39 patients, and the results demonstrated high stability with no overall tendency to increasing D816V+ allele fractions over time. Considerable variation was nevertheless observed in the correlation analyses. Serum tryptase reflects the mature MC load, whereas the D816V+ allele fraction includes cells other than mature MCs to a variable extent. We conclude that tryptase and the D816V+ allele fraction represent different, although correlated, measures of disease status in SM.

Multiple Marker Detection in Peripheral Blood for NSCLC Diagnosis

BackgroundNon-invasive early detection of lung cancer could reduce the number of patients diagnosed with advanced disease, which is associated with a poor prognosis. We analyzed the diagnostic accuracy of a panel of peripheral blood markers in detecting non small cell lung cancer (NSCLC).Methods100 healthy donors and 100 patients with NSCLC were enrolled onto this study. Free circulating DNA, circulating mRNA expression of peptidylarginine deiminase type 4 (PAD4/PADI4), pro-platelet basic protein (PPBP) and haptoglobin were evaluated using a Real-Time PCR-based method.ResultsFree circulating DNA, PADI4, PPBP and haptoglobin levels were significantly higher in NSCLC patients than in healthy donors (p<0.0001, p<0.0001, p = 0.0002 and p = 0.0001, respectively). The fitted logistic regression model demonstrated a significant direct association between marker expression and lung cancer risk. The odds ratios of individual markers were 6.93 (95% CI 4.15–11.58; p<0.0001) for free DNA, 6.99 (95% CI 3.75–13.03; p<0.0001) for PADI4, 2.85 (95% CI 1.71–4.75; p<0.0001) for PPBP and 1.16 (95% CI 1.01–1.33; p = 0.031) for haptoglobin. Free DNA in combination with PPBP and PADI4 gave an area under the ROC curve of 0.93, 95% CI = 0.90–0.97, with sensitivity and specificity over 90%.ConclusionsFree circulating DNA analysis combined with PPBP and PADI4 expression determination appears to accurately discriminate between healthy donors and NSCLC patients. This non-invasive multimarker approach warrants further research to assess its potential role in the diagnostic or screening workup of subjects with suspected lung cancer.

Evaluating different methods of Toxoplasma gondii detection in peripheral blood

Introduction: The mechanisms involved in the virulence and recurrences of ocular toxoplasmosis are poorly understood. New detection methods are necessary to diagnose atypical cases and to better understand recurrences. A sensitive method to identify Toxoplasma gondii in peripheral blood of patients is necessary. Purpose: To detect the presence of T. gondii in peripheral human blood using immunohistochemistry and PCR. Methods: Yellow fluorescent protein RH strain of T. gondii was cultured on fibroblast monolayer for 5 weeks. Cytospins and whole blood smears from spiked blood were stained using monoclonal antibodies on an automated Ventana. For real time PCR, different concentrations of T. gondii were added to human blood and DNA was extracted. PCR was performed targeting a 62bp fragment of the B1 gene or the 529bp fragment. The primers used were: B1 Forward 5’- CTA GTA TCG GTG CGG CAA TGT -3’and Reverse 5’- GGC AGC GTC TCT TCC TCT TTT -3’. For the 529 bp fragment primers used were 5’- CGC TGC AGG GAG GAA GAC GAA AGT TG- 3’ and reverse 5´- CGC TGC AGA CAC AGT GCA TCT GAA TT- 3’. All assays were run in triplicate. Results: The qPCR was able to detect fewer parasites than immunohistochemistry. Moreover, qPCR targeting the 529 bp fragment was better than the B1 gene, being able to detect parasites in concentrations as low as 1 toxo/mL. Conclusions: The qPCR method using the 529bp target proved to be more sensitive than qPCR using the B1 gene target and cytospins or blood smears using immunohistochemistry.

MUC2 mRNA detection in peripheral blood and bone marrow of breast cancer patients reveals micrometastasis

Tumor dissemination to distant organ is the main cause of death. Therefore there is urgent need to set up sensitive methods for early detection of circulating tumor cells (CTCs) and disseminated tumor cells (DTCs) in peripheral blood (PB) and bone marrow (BM) specimens of breast cancer patients. We aim to detect MUC2 mRNA positive cells in PB and BM of breast cancer patients; to relate this to patient relapse. In this study to detect MUC2 mRNA positive cells (tumor marker), PB and BM samples were collected from 50 breast cancer patients after operation and before adjuvant therapy with 20 PB from healthy individuals as negative controls. Chi-square test was used to analyze data. MUC2 mRNA by using Real-time PCR was detected in 9 (18%) of PB and in 10 (20%) of BM samples and none of the healthy individuals. The relapse rate among MUC2-positive patients was significance in BM (P

Circulating Tumor Cells as Predictors of Response and Failure in Breast Cancer Patients Treated with Preoperative Chemotherapy

To explore the significance of circulating tumor cells (CTCs) detection in the course of preoperative chemotherapy (PC) and their effect on the outcomes. Fifty-five patients with stage II/III invasive breast cancer were enrolled into a preoperative clinical trial. Patients were given PC with sequential single-agent doxorubicin and paclitaxel vs paclitaxel followed by doxorubicin. Blood samples (8 mL) were collected from patients before PC, after each phase, and at 6 months intervals during follow-up. Peripheral blood mononuclear cells were isolated and enriched for epithelial cells. Quantitative RT-PCR was used to determine the presence of cytokeratin 19 (CK19) mRNA. Samples were considered positive when the PCR curve crossed the standard threshold curve. After the first phase of chemotherapy, there was a 59% overall reduction in the median tumor volume. The percentage of volume reduction did not differ between patients who presented with detectable CTCs at baseline and those who did not (p=0.89). After the second phase of chemotherapy, there was a further decrease in the median tumor volume to 93% from baseline. There was no correlation between the lack of response and the presence of CTCs either after the first (p=0.36) or second (p=0.5391) phases of PC. The presence of CTCs was a predictor of local or distant relapse (p=0.0411). The detection of CTCs did not affect overall survival (p=0.2569). CTCs can be used as predictors of relapse after definitive treatment of locally advanced breast cancer; however, CTCs detection in peripheral blood during the course of PC does not implicate a particular pattern of response to treatment.

Early Adaptive Immune Responses in the Respiratory Tract of Foot-and-Mouth Disease Virus-Infected Cattle

Foot-and-mouth disease (FMD) is a highly contagious viral disease which affects both domestic and wild biungulate species. This acute disease, caused by the FMD virus (FMDV), usually includes an active replication phase in the respiratory tract for up to 72 h postinfection, followed by hematogenous dissemination and vesicular lesions at oral and foot epithelia. The role of the early local adaptive immunity of the host in the outcome of the infection is not well understood. Here we report the kinetics of appearance of FMDV-specific antibody-secreting cells (ASC) in lymphoid organs along the respiratory tract and the spleen in cattle infected by aerosol exposure. While no responses were observed for up to 3 days postinfection (dpi), all animals developed FMDV-ASC in all the lymphoid organs studied at 4 dpi. Tracheobronchial lymph nodes were the most reactive organs at this time, and IgM was the predominant isotype, followed by IgG1. Numbers of FMDV-ASC were further augmented at 5 and 6 dpi, with an increasing prevalence in upper respiratory organs. Systemic antibody responses were slightly delayed compared with the local reaction. Also, IgM was the dominant isotype in serum at 5 dpi, coinciding with a sharp decrease of viral RNA detection in peripheral blood. These results indicate that following aerogenous administration, cattle develop a rapid and vigorous genuine local antibody response throughout the respiratory tract. Time course and isotype profiles indicate the presence of an efficient T cell-independent antibody response which drives the IgM-mediated virus clearance in cattle infected by FMDV aerosol exposure.

Simultaneous analysis of the expression of 14 genes with individual prognostic value in myelodysplastic syndrome patients at diagnosis: WT1 detection in peripheral blood adversely affects survival

Several studies have evaluated the prognostic value of the individual expression of certain genes in patients with myelodysplastic syndromes (MDS). However, none of them includes their simultaneous analysis by quantitative polymerase chain reaction (PCR). We evaluated relative expression levels of 14 molecular markers in 193 peripheral blood samples from untreated MDS patients using real-time PCR. Detectable WT1 expression levels, low TET2, and low IER3 gene expression were the only markers showing in univariate analysis a poor prognostic value for all treatment-free (TFS), progression-free (PFS), and overall survival (OS). In multivariate analysis, molecular parameters associated with a shorter TFS were: WT1 detection (p = 0.014), low TET2 (p = 0.002), and low IER3 expression (p = 0.025). WT1 detection (p = 0.006) and low TET2 (p = 0.006) expression were associated with a shorter PFS when multivariate analysis was carried out by including only molecular markers. Molecular values with an independent value in OS were: WT1 detection (p = 0.003), high EVI1 expression (p = 0.001), and undetectatable p15-CDKN2B (p = 0.037). WT1 expressers were associated with adverse clinical-biological features, high IPSS and WPSS scoring, and unfavorable molecular expression profile. In summary, detectable WT1 expression levels, and low TET2 and low IER3 expression in peripheral blood showed a strong association with adverse prognosis in MDS patients at diagnosis. However, WT1 was the only molecular marker displaying an independent prognostic value in both OS and TFS.

Diagnosis of viral myocarditis by cardiac magnetic resonance and viral genome detection in peripheral blood

In patients with acute myocarditis, viral genome can be detected in plasma and peripheral leukocytes. Its relationship with active myocardial inflammation, however, is not well understood. Myocardial edema as a feature of inflammation and myocardial necrosis or fibrosis can be frequently observed in patients with acute myocarditis by cardiovascular magnetic resonance (CMR). We assessed the association of viral genome presence in peripheral blood samples with myocardial edema and irreversible injury. We examined consecutive patients with clinically suspected myocarditis after an episode of viral illness. State-of-the-art methods were used for detecting myocardial edema and irreversible injury using CMR and viral genome applying reverse transcribed, nested polymerase chain reaction in peripheral blood samples. The specificity of viral amplification products was confirmed by automatic DNA sequencing. Of a total of 55 patients (53.5 ± 15.6 years), 21 were positive for viral genome in peripheral leukocytes. Interestingly, 18 (86%) of these patients also showed global myocardial edema, as compared to only 7/34 (21%) without PCR evidence for viral genome. The overall agreement between CMR criteria for edema and viral PCR was 84%. In contrast, there was no significant relationship of viral genome presence with myocardial necrosis or scars. In patients with clinically suspected myocarditis, myocardial edema but not irreversible myocardial injury is associated with the presence of viral genome in peripheral blood.

BCR-ABL fusion protein detection in peripheral blood and bone marrow samples of adult precursor B-cell acute lymphoblastic leukemia patients using the flow cytometric immunobead assay

The ability to detect the BCR-ABL fusion gene in precursor B-cell acute lymphoblastic leukemia (pB-ALL) is essential for making accurate treatment decisions. We used a new flow cytometric immunobead assay for BCR-ABL fusion protein detection in peripheral blood and/or bone marrow samples from 38 adult pB-ALL patients and the results were compared with polymerase chain reaction (PCR) detection of BCR-ABL transcript. The fusion protein was detected in peripheral blood and bone marrow samples from seven of the 38 (18%) patients, and results for both the p190 and p210 were confirmed by PCR. One case, which was positive by cytogenetics and fluorescence in situ hybridization (FISH), was negative by PCR but positive by flow cytometry. Another case, which was positive by PCR and negative by flow cytometry, was from a patient on steroid treatment. The cytometric immunobead assay for BCR-ABL fusion protein detection was found to be suitable for the investigation of pB-ALL patients. This assay is reliable, rapid and simple to use for peripheral blood and bone marrow samples.

Counterpoint: Detection of Myocardial Infarction—Is It All Troponin? Role of New Markers

It has become good scientific practice to mention potential conflicts at the beginning of the presentation. My main conflict related to this topic is that I love cardiac troponin, for the very reasons listed by Giannitsis and Katus (1). Cardiac troponin is beyond a doubt the most important biomarker in cardiovascular medicine, even more so after the clinical introduction of sensitive assays (1–6). For the detection of acute myocardial infarction (AMI),2 the room for additional biomarkers, if any, seems to be small and restricted to add-on applications to be used in conjunction with cardiac troponin, clinical assessment, and the 12-lead electrocardiogram. Clinical settings in which there are still unmet needs include: ( a ) the early diagnosis of AMI, ( b ) differentiation of type I and type II AMI, and ( c ) risk stratification for death or recurrent AMI. Cardiac troponins are structural proteins. Their detection in peripheral blood indicates cardiomyocyte damage and likely cell death (1–7). Invariably, there is some time delay between the onset of AMI (coronary plaque rupture and coronary occlusion) and the appearance of cardiac troponin in the peripheral circulation. Because cardiac troponin signals cardiomyocyte damage regardless of the underlying cause, multiple nonischemic conditions can challenge the interpretation of increases in cardiac troponin, particularly mild ones (1, 7). Thus, we could benefit from information provided by additional biomarkers that ( a ) reflect myocardial ischemia (not necrosis), ( b ) indicate plaque rupture or other signals present at the very onset of AMI (e.g., endogenous stress), or ( c ) are associated with a specific pathobiology present in only a subset of AMI patients, thereby allowing more personalized and targeted patient management. Recent evidence suggests that markers fulfilling at least some of these properties might soon become clinically available. Copeptin, the C-terminal part of the vasopressin prohormone, is secreted stoichiometrically …

Aberrant methylation of the PTPRO gene in peripheral blood as a potential biomarker in esophageal squamous cell carcinoma patients

Epigenetic inactivation of protein tyrosine phosphatase receptor-type O (PTPRO), a new member of the PTP family, has been described in several forms of cancer. We evaluated PTPRO promoter hypermethylation as a potential biomarker in esophageal squamous cell carcinoma (ESCC). This alteration was observed in 27 (75%) of 36 primary tumors and correlated significantly with depth of invasion (T-stage, P = 0.013). Among matched peripheral blood samples from ESCC patients, 13 (36.1%) of 36 exhibited detectable methylated PTPRO in plasma, while 15 (41.7%) of 36 had this abnormality in buffy coat. No methylated PTPRO was observed in normal peripheral blood samples from 10 healthy individuals. In addition, demethylation by 5-aza-dC treatment led to gene reactivation in PTPRO-methylated and -silenced ESCC cell lines. To our knowledge, this is the first report of methylated PTPRO as a noninvasive tumor biomarker in peripheral blood. These findings suggest that hypermethylated PTPRO occurs frequently in ESCC. Further, detection in peripheral blood of ESCC patients suggests potential clinical application for noninvasive diagnosis and disease monitoring.

Minimal disease detection in peripheral blood and bone marrow from patients with non-metastatic neuroblastoma

In non-metastatic neuroblastoma (NB), the identification of the cases that require more intensive treatment is still difficult. Minimal disease (MD) and minimal residual disease (MRD) detection in outcome prediction seems to be important in advanced neuroblastoma, but there are not many studies focused on patients with non-metastatic disease. The aim of this study was to determine whether the presence of MD detected at diagnosis could be associated with bad prognosis. Quantitative reverse transcriptase-polymerase chain reaction QRT-PCR was performed on peripheral blood (PB) and bone marrow (BM) samples from patients with non-metastatic NB at diagnosis for tyrosine hydroxylase (TH) and doublecortin (DCX) mRNAs detection. The frequencies of detecting MD in our series of 102 patients with non-metastatic NB were as follows: 6.2% (5/81) PB samples and 10.6% (10/94) BM samples. Overall survival was similar for patients who expressed or not the MD biomarkers at diagnosis. However, patients with MD detected in PB showed lower EFS than patients with negative PB (P=0.038). Minimal disease detection in PB seems to be useful for predicting relapse probabilities in patients with non-metastatic NB. The stages 1 and 2 patients with neuroblastoma showed high survival rates, and MD was detected in a small number of patients probably being non-contributory for predicting patient outcome. For stage 3 patients with NB, MD detection by QRT-PCR in PB at diagnosis could be useful for predicting outcome and for early and sensitive detection of relapsing disease.

Abstract PD04-07: Monitoring Circulating Tumor Cells (CTC) and Circulating Endothelial Cells (CEC) during Neoadjuvant Combination of Trastuzumab and Bevacizumab with Chemotherapy in HER2 Overexpressing Inflammatory Breast Cancer (IBC): An Ancillary Study of BEVERLY 2 Multicenter Phase II Trial

Abstract Introduction: CTC detection in peripheral blood is an independent prognostic factor in early breast cancer but with a low detection rate of 10% to 23% of the patients (B. Rack, ASCO 2010, FC Bidard, Ann Oncol 2010). Changes in the HER2 status of CTC compared to the primary tumor have been reported (Riethdorf S CCR 2010). Predictive value of CEC for response to anti-angiogenic agents is debated. Material and methods: CTC and CEC were detected in 7.5 ml and 4 ml of blood respectively in IBC (T4d) patients enrolled in the phase II multicenter trial, BEVERLY 2. This study is evaluating the efficacy of bevacizumab (15mg/kg q3w given concurrently) in combination with sequential neoadjuvant chemotherapy of 4 cycles of FEC followed by 4 cycles of Docetaxel-Trastuzumab. Bevacizumab was stopped 4 weeks before and reintroduced 4 weeks (w) after mastectomy. All patients had non metastatic IBC and over expressed HER2 (3+ in IHC or FISH +). The CellSearch™ System, combining EpCAM immunomagnetic selection (IMS) followed by anti-cytokeratin (A45B/B3) and anti-HER2 fluorescently staining for CTC and CD146 IMS and CD105 staining for CEC, was used at baseline, before cycle 5, before and after surgery. Results: From Oct 2008 to Oct 2009, 52 patients were included in this study and 51 were evaluable for CTC. At baseline, 18 patients out of 51 had ≥ one detectable CTC (35.3%, 95%CI 22-48%, range 1 to 92). Pathological complete response rate according to local review was 33/47 (70%) or 15/22 (68%) for centrally reviewed cases. At baseline, CTC level was not correlated with CEC level, neither with other patients and tumor characteristics’ (age, nodal status, PeV) nor pCR (centrally reviewed in 24). All positive cases for CTC detection had HER2 positive CTC. Five pts out of 18 (28%) had both HER2+ and HER2 negative CTCs in their blood. A lower level of CEC (&amp;lt; 20/4ml) before C5 could be associated with a higher probability of pCR (Khi2 test, p=0.053). Conclusion: We observed a high CTC detection rate of 35% in this population of patients with HER2+ IBC and a dramatic drop in CTC level during treatment in concordance with the high efficiency of this combination of chemotherapy and targeted therapy. We observed heterogeneity in the HER2 status of CTC in some patients. CEC levels increased progressively during neoadjuvant treatment and decreased after its interruption. Longer follow-up will show if CTC and CEC variations are early predictive factors for this highly efficient combination in HER2+ IBC. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr PD04-07.

Dual-subtype feline immunodeficiency virus vaccine provides 12 months of protective immunity against heterologous challenge.

The duration of immunity of the dual-subtype feline immunodeficiency virus (FIV) vaccine, Fel-O-Vax FIV, for protection against subtype-B FIV was assessed in this study. Vaccinated cats along with controls were challenged with FIV(FC1), a subtype-B FIV strain, 54 weeks after the final vaccination, and monitored for 46-48 weeks for provirus and viral RNA in peripheral blood, provirus in lymphoid organs, and CD4:CD8 ratios. Results of provirus detection in peripheral blood and lymphoid organs and plasma viral RNA loads showed that 10/14 vaccinated cats were fully protected for 48 weeks against infection with FIV(FC1) whereas 5/5 controls were persistently infected with FIV(FC1). CD4:CD8 inversions were noted in association with FIV infection and viral loads were not significantly different between FIV infected controls and the unprotected vaccinated animals.