Prognostic factors of renal cell carcinoma.

Renal cell carcinoma represents 23% of all adult malignancies and its incidence in the Czech Republic is one of the highest worldwide. Until late stages this disease often remains asymptomatic, which makes its diagnosis difficult. Despite an increasing proportion of small, incidentally detected tumours on imaging, approximately one third of patients are still diagnosed with advanced disease. Moreover, a relapse occurs in up to 40% of patients after surgery for localized tumour. Increased availability of imaging investigations allowing an early detection of kidney carcinoma and advances in systemic treatment have favourably affected the outcome of patients with this type of tumour. Nevertheless, mortality of renal cell carcinoma remains the highest among urological malignancies. The individual course of the disease and its response to systemic treatment are difficult to predict. A number of prognostic factors of renal cell carcinoma have been identified, of which TNM classification and tumour grade remain the most important. Recently, several multivariate prognostic models have become available, allowing a more accurate prediction of the disease course. In localized disease, they are useful in identifying patients at higher risk of recurrence and allow optimization of follow-up after surgery. In metastatic disease, they are routinely used to stratify patients into risk groups for targeted treatment. There has been a long-term effort to identify a suitable biomarker useful for an early detection and assessment of the prognosis of renal cell carcinoma. At the same time, such a biomarker could improve the accuracy of established prognostic systems. This text presents an overview of prognostic factors of renal cell carcinoma, including a summary of potential biomarkers.

Time and Temperature Stability of TGF-ß1, EGF and IGF-1 in 20% and 100% Human Serum.

Autologous serum eye drops (ASEDs) are used as a treatment for severe dry eye disease. The concentration and stability of various growth factors in ASEDs is determinative for their efficiency. We therefore assessed the concentrations of transforming growth factor beta 1 (TGF-ß1), epidermal growth factor (EGF) and insulin-like growth factor 1 (IGF-1) in ASEDs following storage at 4-8, -20, -80 and -156 °C. Twenty % and 100% sera from eight healthy volunteers were analysed by the sandwich enzyme immunoassay at different time intervals up to seven months. The mean levels of TGF-ß1 and EGF in undiluted and 20% serum did not differ significantly from the baseline levels in fresh serum for any storage conditions after 7 days at 4-8 °C, as well as after 4- and 7-month preservation at sub-zero temperatures. In 20% serum, no IGF-1 concentration decrease was found following 7 days of preservation at 4-8 °C. However, a decrease to 78 % and 81 % (P < 0.01) of baseline values was found in 20% serum after 4-month storage at -20 °C and 7-month storage at -156 °C, respectively. A more pronounced decrease in IGF-1 was observed in undiluted serum. All assessed growth factors present in 20% frozen serum remained stable for up to 7 months. The highest stability was achieved at -80 °C. At -20 and -156 °C, some decrease in IGF-1 occurred. Our results indicate that 20% ASEDs can be stored frozen up to 7 months under proper conditions.

Circulating Cell-Free DNA Extraction from Liquid Biopsy for Cancer Research.

As the number of cancer patients globally increases, a need for reliable biomarkers including circulating tumour DNA from liquid biopsy for diagnosis, prognosis and monitoring of the disease is rising. Currently, mainly tissue samples from biopsy are used, but there are certain limitations: firstly, it is an invasive technique, and secondly, in some cases it is almost impossible to obtain an acceptable tissue sample. This could be changed by using circulating cell-free DNA from liquid biopsy, which also gives the possibility of repeated examination. Here, we focus on the options of isolating circulating cell-free DNA from plasma samples using two isolation techniques: precision manual QIAamp Circulating Nucleic Acid Kit and automatic MagNA Pure Compact (MPC) using Nucleic Acid Isolation Kit I. Manual extraction gave significantly better yields of circulating tumour DNA (P < 0.05). This DNA also had less contaminants (organic compounds or proteins). DNA obtained by both tested methods of isolation is suitable for subsequent molecular genetic methods.

The Effects of Different Storage Conditions and Repeated Freeze/Thaw Cycles on the Concentration, Purity and Integrity of Genomic DNA.

The crucial requirement of molecular genetic methods is high-quality input material. The key question is "how to preserve DNA during long-term storage." Biobanks are recommended to aliquot isolated DNA into provided volumes. The aim of this study was to analyse the effect of repeated freezing and thawing on the genomic DNA integrity, quality and concentration. The aliquoted DNA isolated from blood cells using the automatic MagNA system and manual salting out method underwent freeze/thaw cycles at different storage conditions (-20 °C, -80 °C and liquid nitrogen). The average initial concentrations were 270.6 ng/µl (salting out method) and 125.0 ng/µl (MagNA). All concentration deviations relative to the concentration after the first freeze/ thaw cycle were less than 5 % for -20 °C and -80 °C cycling with both isolation methods. The average percentage differences of liquid nitrogen samples were higher, and the MagNA isolation method showed significant differences. There were no significant changes in the DNA purity or quality. The repeating freeze/ thaw up to 100 cycles (through -20 °C and -80 °C, respectively) did not significantly influence the integrity, concentration, or purity of genomic DNA, suggesting that storage of samples in high-volume pools without multiple aliquoting is possible. Storage in a freezer seems to be the most suitable way of long-term DNA preservation, because liquid nitrogen storage leads to formation of DNA clumps.

The effect of surgery on the levels of matrix metalloproteinases in patients with inguinal hernia.

Matrix metalloproteinases (MMPs) are associated with the alteration of extracellular matrix. The purpose of this study was to investigate how the levels of matrix metalloproteinases and their inhibitors - TIMPs are influenced by the presence of inguinal hernia as well as by its surgical treatment. The studied group consisted of 25 patients with inguinal hernia and 21 healthy controls for comparison. Two blood samples - before and after the treatment were collected from patients. Serum concentrations of MMPs and TIMPs were analysed by multiplex immunoassays. There was a difference in circulating levels of MMPs in patients before the surgery compared to healthy controls - the concentrations of MMP-2 and MMP-9 were significantly lower (p=0.026, p=0.018, respectively). After the surgery, the levels of MMPs, especially MMP-2 (p<0.0001), were significantly decreased in patients compared to the preoperative values, apart from MMP-9. On the contrary, MMP-9 showed significant increase after the surgery (p<0.0001). Circulation levels of TIMP-2 in patients were significantly decreased in comparison with controls (p=0.004), whereas levels of TIMP-1 were similar to controls. Both tested metalloproteinase inhibitors showed a significant decrease in detected levels (TIMP-1 p=0.0004; TIMP-2 p<0.0001) after the procedure compared to the preoperative values. The levels of MMPs, especially MMP-2 and MMP-9, and their inhibitors TIMP-1 and TIMP-2 are involved by the presence of inguinal hernia as well as are influenced by the surgery.

Pancreatic stellate cells - rising stars in pancreatic pathologies.

Pluripotent pancreatic stellate cells (PSCs) receive growing interest in past decades. Two types of PSCs are recognized -vitamin A accumulating quiescent PSCs and activated PSCs- the main producents of extracellular matrix in pancreatic tissue. PSCs plays important role in pathogenesis of pancreatic fibrosis in pancreatic cancer and chronic pancreatitis. PSCs are intensively studied as potential therapeutical target because of their important role in developing desmoplastic stroma in pancreatic cancer. There also exists evidence that PSC are involved in other pathologies like type-2 diabetes mellitus. This article brings brief characteristics of PSCs and recent advances in research of these cells.

Can gram-negative-like biomarker values in Streptococcus pyogenes sepsis negatively influence right choice of initial antibiotic therapy?

Úvod: Biomarkery jsou u septických pacientů využívány jak k diagnostice sepse, tak k antibiotickému stewardshipu. Sepse vyvolaná gramnegativními bakteriemi mívá odlišné charakteristiky, především vysoký prokalcitonin vs C-reaktivní protein v porovnání se sepsí vyvolanou grampozitivními bakteriemi. Avšak jednotlivá infekční agens, především Streptococcus pyogenes, nemusí do tohoto schématu zapadat, což může vest k nesprávné iniciální volbě antibiotika. Metody: Retrospektivní analýza biomarkerů, iniciální volby antibiotické léčby a výsledků léčby u pacientů se sepsí vyvolanou S. pyogenes, Escherichia coli a Staphylococcus aureus. Hodnoty biomarkerů byly porovnány pomocí Kruskal-Wallis testu s následným Dunn post-Hoc testem s prahem p < 0,05. Výsledky: Hodnoty prokalcitoninu byly nejvyšší u sepse vyvolané S. pyogenes (12,51 ng/ml, IQR: 6,26-48,38 ng/ml) oproti sepsi vyvolaná E. coli (4,30 ng/ml, IQR: 1,50-10,00 ng/ml, p < 0,001) a S. aureus (0,75 ng/ml, QR: 0,34-1,62 ng/ml, p < 0,001). Poměr neutrofilů a lymfocytů vykazoval stejné charakteristiky jako prokalcitonin. Správná iniciální antibiotická léčba byla v souboru S. pyogenes 11,29 % v porovnání s 99,3 % a 100 % u S. aureus a E. coli skupin. Závěr: Oproti předchozím studiím byly v našem souboru pozorovány nejvyšší hodnoty prokalcitoninu u pacientů se sepsí vyvolanou S. pyogenes spíše než gramnegativními bakteriemi. Vysoké hodnoty prokalcitoninu imitující gramnegativní zánětlivou odpověď přispěli k ovlivnění výběru iniciální antibiotické léčby (bez znalosti původce), což mohlo vést k vyšší mortalitě u této skupiny pacientů. Proto doporučujeme přehodnocení významu prokalcitoninu v diagnostice sepse pro zlepšení přežití i kvality života pacientů.

Plasma osteopontin levels, but not its myocardial expression, reflect heart failure severity in recently diagnosed dilated cardiomyopathy.

BACKGROUND: Elevated levels of the extracellular matrix glycoprotein osteopontin (OPN) may be detected in both myocardium and plasma under various pathological conditions affecting the heart. Several studies demonstrated increased plasma OPN levels in patients with heart failure due to dilated cardiomyopathy (DCM), while other studies showed high OPN expression levels in the myocardium of such patients. However, very little is known about OPN levels in both plasma and myocardium of the same individual with DCM. Therefore, we aimed to compare plasma OPN levels and levels of myocardial OPN expression in patients with recent-onset DCM (Ro-DCM). METHODS: We examined plasma OPN as well as creatinine, C­reactive protein (CRP), brain natriuretic peptide (BNP), and troponin I levels in 25 patients with Ro-DCM. Furthermore, all subjects underwent transthoracic echocardiography, selective coronary angiography, and endomyocardial biopsy (EMB) for the assessment of myocardial OPN expression. RESULTS: No significant correlation between myocardial OPN expression and clinical, biochemical, or echocardiographic parameters was found. In log transformation analysis, plasma OPN levels correlated significantly with BNP levels (r = 0.46, p = 0.031), with CRP levels (r = 0.52, p = 0.015), and with early diastolic mitral annular velocity (r = -0.57, p = 0.009). There was a borderline association between the plasma OPN log value and New York Heart Association class (p = 0.053). CONCLUSION: Plasma OPN levels reflect heart failure severity in patients with Ro-DCM. Myocardial OPN expression is not associated with either plasma OPN levels or markers of heart failure in these individuals.

The significance of calprotectin, CD147, APOA4 and DJ-1 in non-invasive detection of urinary bladder carcinoma.

Aim of the study is to define the diagnostic accuracy of selected urinary protein biomarkers in the non-invasive detection of primary and recurrent urothelial carcinoma of the urinary bladder. The urinary levels of calprotectin, CD147, APOA4 and protein deglycase DJ-1 were examined in 255 individuals, including 60 controls with non-malignant urological disease, 61 patients with a history of urinary bladder cancer with negative cytology and negative cystoscopy and 134 patients with urinary bladder cancer. Urinary concentrations of biomarkers were determined by Enzyme-Linked Immunosorbent Assay (ELISA). During the follow-up of patients with non-muscle invasive bladder cancer (NMIBC), a group of 44 patients with cancer recurrence was compared to the group of 61 patients with a history of NMIBC but with no evidence of disease. Urinary concentrations of the evaluated markers did not reveal any significant difference between these groups. During the primary diagnosis, a group of 90 patients with primary bladder cancer and 60 subjects with benign disease were compared. Urinary levels of CD147 were not significantly higher in patients with tumors. The greatest diagnostic accuracy was observed in APOA4 (sensitivity 55.6, specificity 83.3, AUC 0.75), and lesser in calprotectin (sensitivity 39.4, specificity 87.7, AUC 0.66) and in DJ-1 (sensitivity 61.1, specificity 66.7, AUC 0.64), respectively. Apolipoprotein A4 may be used potentially as a supplemental urinary marker in the diagnosis of primary bladder cancer.

Sclerostin levels predict cardiovascular mortality in long-term hemodialysis patients: A prospective observational cohort study.

Sclerostin is a protein which is involved in bone metabolism and probably also in vessel wall function. This prospective observational cohort study evaluated the prognostic significance of sclerostin in hemodialysis (HD) patients. In total, 106 HD patients and 25 healthy controls participated in the study. HD patients were prospectively followed up for five years. Sclerostin was measured in serum using standard ELISA kits by Biomedica. Sclerostin concentrations in serum were higher in HD patients compared to the controls (89.2±40.3 pmol/l vs. 32.8±13.0 pmol/l, p<0.001). Sclerostin levels were significant for cardiovascular mortality but not for overall mortality and mortality due to infection. A higher cardiovascular risk was connected to sclerostin concentrations above the median (>84 pmol/l), HR (95 % CI): 2.577 (1.0002-10.207), p=0.04. When sclerostin was evaluated together with residual diuresis in Kaplan-Meier analysis the worst prognosis due to cardiovascular events was observed in the group with high sclerostin and zero residual diuresis compared to all other patients (p=0.007). In summary, serum sclerostin levels in HD patients were increased when compared to healthy subjects. High sclerostin levels were demonstrated as a risk factor for cardiovascular mortality. Further studies are required to clarify the pathophysiological mechanisms of sclerostin action in patients with renal failure before therapeutic measures can be established.

Plasma osteopontin levels in patients with dilated and hypertrophic cardiomyopathy.

BACKGROUND: Osteopontin (OPN) is an extracellular matrix glycoprotein that plays a role in a variety of cellular activities associated with inflammatory and fibrotic responses. Increased OPN levels in myocardium and plasma have been demonstrated in patients with dilated cardiomyopathy (DCM). However, nothing is known about OPN levels in patients with hypertrophic cardiomyopathy (HCM). Therefore, the aim of our study was to compare plasma OPN levels in patients with these two most common cardiomyopathies. PATIENTS AND METHODS: We examined plasma OPN as well as creatinine, C­reactive protein (CRP), brain-type natriuretic peptide (BNP), and troponin I levels in 64 patients with DCM, 43 patients with HCM, and 75 control subjects. Transthoracic echocardiography was also performed on all cardiomyopathy patients. RESULTS: Plasma OPN levels were significantly elevated in patients with DCM compared with HCM patients (95 ± 43 vs. 57 ± 21 ng/ml; p < 0.001) and control subjects (54 ± 19 ng/ml; p < 0.001); however, there was no difference between HCM patients and control subjects. New York Heart Association (NYHA) class III or IV disease was more frequently present in DCM patients than in HCM subjects (44 % vs. 2 %, p < 0.0001). In multivariate analysis, BNP and CRP levels together with NYHA class were found to be significant predictors of plasma OPN levels in DCM patients (p = 0.002, p = 0.029, and p < 0.001 for BNP, CRP, and NYHA, respectively). CONCLUSION: Plasma OPN levels were associated with overall heart failure severity rather than with specific cardiomyopathy subtype in patients suffering from DCM or HCM, respectively.

Matrix Metalloproteinases and Their Tissue Inhibitors: an Evaluation of Novel Biomarkers in ANCA-Associated Vasculitis.

Matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) may play an important role in both inflammation with subsequent fibrosis and in repair and healing in anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). We evaluated the circulating levels of MMPs, including pregnancy-associated plasma protein A (PAPP-A), and TIMPs in patients with AAV. PAPP-A, MMP-2, MMP-3, MMP-7, MMP-9, TIMP-1, TIMP-2 and selected parameters were measured in 100 AAV patients (36 patients with active disease and 64 patients in remission) and 34 healthy subjects. The levels of MMP-2, MMP-3, MMP-7, MMP-9, TIMP-1, TIMP-2, and PAPP-A in AAV were all found to be different to those of the controls. The MMP-7 and PAPP-A concentrations were increased in active disease in comparison to the controls (MMP-7: 13 ±.7 vs. 2 ± 0.6 ng/ml, PAPP-A: 14 ± 18 vs. 6.8 ± 2.6 ng/ml, both P < 0.005). The MMP-2 and TIMP-2 levels were increased in remission when compared to the controls (MMP-2: 242 ± 50 ng/ml vs. 212 ± 26 ng /ml, TIMP-2: 82 ± 14 ng/ml vs. 68 ± 93 ng/ml) and to the active AAV (MMP-2: 242 ± 50 vs. 219 ± 54 ng/ml, TIMP-2: 82 ± 14 ng/ml vs. 73 ± 15 ng/ml, all P < 0.005). MMP-3, MMP-7, TIMP-1, and PAPP-A correlated with serum creatinine. The serum levels of MMPs, TIMPs and PAPP-A are all altered in AAV. MMP-2, MMP-7 and TIMP-2 appear to be promising markers in distinguishing active AAV from remission. MMP-3, MMP-7, TIMP-1, and PAPP-A are associated with kidney function in AAV. Further studies are needed to delineate the exact roles of circulating MMPs, TIMPs and PAPP-A in patients with AAV.

The importance of serum osteopontin and stanniocalcin-1 in renal cell carcinoma.

A total of 56 RCC patients with staging ≥ pT1b were enrolled in a prospective study to assess the prognostic importance of serum levels of osteopontin (OP), stanniocalcin-1 (SC), FGF-23, alpha Klotho and 25-OH-D at the time of diagnosis in renal cell carcinoma (RCC) patients. The relationship between the serum level of the analyzed parameters and recurrence-free survival (RFS), overall survival (OS) and cancer-specific survival (CSS) was examined, and our control group consisted of 20 patients without cancer. The levels of osteopontin, stanniocalcin-1, FGF-23 and alpha Klotho were determined by Enzyme-Linked Immunosorbent Assay (ELISA) and 25-OH-D by chemiluminiscence immunoanalysis (CLIA). The follow-up period median was 46 months. Renal cell carcinoma recurred in 9 patients and 20 patients died during follow-up; 12 of them from RCC. The level of osteopontin and stanniocalcin-1 varied between the control group and RCC patients (at p=0.02 and p=0.0003). Higher levels of stanniocalcin-1 were detected in the metastatic RCC group than in the localized RCC group (p=0.003). Only the stanniocalcin-1 level at the time of surgery was associated with RFS (p=0.0004). Both OS and CCS were associated with the osteopontin, stanniocalcin-1 and FGF preoperative level. Patients with stanniocalcin-1 level over 1,277 pg/ml and osteopontin level over 100 ng/ml had 17.8 times higher and 7.9 times higher risk of dying from RCC progression, respectively (p<0.001 and p=0.002). High levels of osteopontin, stanniocalcin-1 and FGF 23 at the time of surgery are important prognostic factors related to CSS and OS. Patients with high stanniocalcin-1 level were at risk of tumor recurrence.

Low maternal serum concentrations of mannose-binding lectin are associated with the risk of shorter duration of pregnancy and lower birthweight.

Chronic inflammation has been implicated as the underlying mechanism responsible for the pathophysiology of preterm labour. Mannose-binding lectin (MBL) plays a central role in the innate immune response and is thus an important component of the first line of defense. The aim of this study was to investigate whether serum concentrations of MBL correlated with the incidence of preterm birth and low birthweight in a cohort of women with signs of threatened preterm birth. A cohort of 60 patients who presented with regular contractions and/or short cervix (group A) between 24 and 32 weeks of gestation and 20 healthy controls (group B) who had no pregnancy complications and delivered at term were recruited into a prospective study. The following outcomes were recorded: presence of preterm labour and birthweight in all patients. MBL and high sensitivity C-reactive protein levels were measured in all serum samples. The serum concentrations of MBL were significantly reduced in patients with threatened preterm labour (Group A), compared to the control Group B. Furthermore, infants born to Group A mothers with MBL deficiency (n = 13, MBL ≤100 ng/mL) had significantly lower birthweights, compared to those born to Group A women with normal MBL serum concentrations (P < .0001). Our small cohort study demonstrated a strong association between MBL deficiency and preterm delivery, and associated low birthweight. MBL deficiency could thus be considered an important risk factor for preterm birth.

Proteomic approach for identification of IgA nephropathy-related biomarkers in urine.

Proteinuria is often used as a surrogate marker in monitoring and predicting outcome in patients with chronic kidney diseases, but it is non-specific. IgAN belongs to the most common primary glomerulonephritis worldwide with serious prognosis. The main aim of this work was to assess differences in urine proteins in patients with IgA nephropathy and to identify abnormal proteins as potential biomarkers of IgA nephropathy or the renal disease. In our pilot project, we selected 20 patients and compared them with 20 healthy volunteers. Protein quantification was performed using iTRAQ (isobaric tag for relative and absolute quantitation) labeling method. The peptides were separated by the isoelectric focusing method (IEF) and nano-LC with C18 column and identified by mass spectrometry using MALDI-TOF/TOF MS. Proteins´ lists obtained from IEF-LC-MS-MS/MS analysis were combined and contained 201 proteins. It was found out that 113 proteins were common in both experiments. 30 urinary proteins were significantly up- or down-regulated in patients with IgA nephropathy. We characterized potential biomarkers such as alpha-1-antitrypsin, apolipoprotein A-I, CD44 antigen or kininogen. Potential biomarkers of IgAN should be validated in further studies.

Contribution of ABCB1 and CYP2D6 genotypes to the outcome of tamoxifen adjuvant treatment in premenopausal women with breast cancer.

Recent pre-clinical evidence suggests that the active metabolite of tamoxifen, endoxifen, is a substrate for efflux pump P-glycoprotein. The aim of our study was to evaluate, if the polymoprhisms within ABCB1 gene alter tamoxifen adjuvant treatment efficacy in premenopausal women. Totally 71 premenopausal women with estrogen receptor positive breast cancer indicated for tamoxifen adjuvant treatment were followed retrospectively for median period of 56 months. The gentic polymorphisms of CYP2D6 and ABCB1 were analyzed and potential covariates as tumor grading, staging, age at the diagnosis, comedication, quantitative positivity of ER or PR were also evaluated. Cox proportional-hazards regression model indicated that patients carrying at least one variant allele in ABCB1 rs1045642 had significantly longer time to event survival compared to wild type subjects. Non-significant trend was noted for better treatment outcome of patients carrying at least one variant allele in the SNP rs2032582, while for the CYP2D6 polymorphism poor metabolizer phenotype resulted in worse outcome in comparison to extensive metabolizers subjects with HR of 4.04 (95 % CI 0.31-52.19). Similarly, patients using CYP2D6 inhibitors had non-significantly shorter time-to-event as compared to never users resulting in hazard ratio of 2.06 (95 % CI 0.40-10.63). ABCB1 polymorphisms may affect outcome of tamoxifen adjuvant treatment in premenopausal breast cancer patiens. This factor should be taken into account in addition to the CYP2D6 polymorphism or phenotypic inhibition of CYP2D6 activity.

RAGE and its ligands in cancer - culprits, biomarkers, or therapeutic targets?

Receptor for advanced glycation end products (RAGE) plays a central role in the regulation of tissue homeostasis, regeneration and resolution of inflammation, but under pathological conditions RAGE-mediated pathways may induce diminished apoptosis, but enhanced autophagy and cell necrosis. These mechanisms may contribute to malignant transformation, cancer progression and metastases. Soluble RAGE may bind natural RAGE ligands and counteract some of the RAGE-mediated effects. Activation of RAGE was demonstrated in different types of cancer (including colon, pancreatic and breast cancer). Expression of RAGE and serum levels of soluble RAGE may serve as cancer biomarkers and strategies aimed at interfering with RAGE signaling might be promising anticancer drugs.

Trefoil factor family (TFF) proteins as potential serum biomarkers in patients with metastatic colorectal cancer.

Trefoil factor family (TFF) is composed of three secretory proteins (TFF1, TFF2 and TFF3) that play an important role in mucosal protection of gastrointestinal tract. Their overexpression in colorectal tumors seems to be associated with more aggressive disease. We collected serum samples from 79 healthy controls and 97 patients with metastatic colorectal cancer at the time of diagnosis or at progression. Serum levels of TTF1-3, CEA and CA19-9 were measured by ELISA. Serum TFF1 and TFF3 levels were significantly higher in patients with colorectal cancer compared to healthy controls (p < 0.0001). Moreover, serum levels of TFF3 correlated with extent of liver involvement in patient without pulmonary metastases and patients with higher TFF3 levels had significantly worse outcome (p < 0.0001). Compared to CEA and CA19-9, TFF3 had higher sensitivity and the same specificity. Our results indicate that TFF3 is an effective biomarker in patients with metastatic colorectal cancer with higher sensitivity than CEA a CA19-9. TFF3 levels strongly correlate with extension of liver disease and seem to have prognostic value.

Impact of risk factors on prevalence of anal HPV infection in women with simultaneous cervical lesion.

UNLABELLED: The aim of our study was to determine the risk factors associated with anal HPV infection in HIV-negative women with high-grade cervical lesion. The study group included 172 "high-risk" women who underwent conization for high-grade cervical intraepithelial lesion or microinvasive cervical cancer (CIN 2+). The control group consisted of 100 "low-risk" women with non-neoplastic gynecologic diseases. All participants completed a questionnaire detailing medical history and sexual risk factors and were subjected to anal and cervical HPV genotyping. Concurrent cervical and anal HPV infections were detected in 42.4% (73/172) women of the study group, and in 8.0% (8/100) of women in the control group, respectively. The subgroup with concurrent HPV infections (n=73) dominated women with CIN 3 and microinvasive cancer and anal HPV 16 infections (n=53). Women with concurrent infections more frequently reported any type of sexual contact with the anus including non-penetrative anal sex (OR 2.62, p=0.008). Reporting >5 lifetime sexual partners (OR 2.43, p=0.041), smoking > 60 cigarettes per week (OR 2.33, p=0.048), and a history of penetrative anal intercourse (OR 3.87, p=0.002) were observed as the significant risk factors in women with multiple concurrent HPV infections. Our data support anal HPV testing and anal Pap smear screening in all women with severe cervical lesions caused by HPV 16 and a history of any sexual contact with the anus, heavy smoking and/or more than 5 lifetime sexual partners. KEYWORDS: anal cancer, cervical intraepithelial neoplasia, HPV, risk factor.

Predictive factors for the presence of tumor cells in bone marrow and peripheral blood in breast cancer patients.

UNLABELLED: Simultaneous detection of disseminated tumor cells (DTCs) and circulating tumor cells (CTCs) was shown to be associated with an especially poor prognosis and increased incidence of disease-related deaths in non-metastatic breast cancer patients. We analyzed the occurance of DTCs and CTCs in patients with primary breast cancer and evaluated the correlation of their presence with other prognostic markers and investigated the changes in DTCs/CTCs number at different time points during treatment.Blood of 50 patients with primary breast cancer were used for immunomagnetic separation and detection of circulating tumor cells using the commercial available system the AdnaTest Breast Cancer™ (AdnaGen GmbH, Langenhagen, Germany). Bone marrow aspirates from 50 patients were analyzed for DTCs by immunocytochemistry using the pan-cytokeratin antibody conjugated with FITC (Monoclonal Anti-Cytokeratin antibody F3418, Sigma Aldrich).DTCs were identified in 30% (15/50) and CTCs in 22% (11/50) of patients. We found that DTC positivity could point to a significantly high risk of larger primary tumor size (p-value 0.011) and significantly higher risk of lymph node involvement (p-value 0.002). For CTC positivity, no such relationship was proven. DTCs have shown significantly higher prevalence in ER/PR-negative females and in HER2-positive cases. CTCs were equally prevalent in patients with the presence and absence of standard prognostic and predictive markers such as ER, PR and HER2. We found no correlation between CTCs and DTCs findings (r = -0.097, p = 0.504). We used DTCs/CTCs analysis for therapy monitoring in a small group of 29 patients, who underwent neoadjuvant chemotherapy (NACT). We find out no significant correlation between DTCs/CTCs detection and the primary tumor response to NACT. A pathologic complete response (pCR) was achieved by 31% (9/29) of the patients in our study, however, no association was observed between pCR and the detection of DTCs after NACT.These results support the use of DTCs/CTCs analysis in early breast cancer to generate clinically useful prognostic information. The study of these cells apart from the impact on refining prognosis, has the exciting potential of individualising treatment for women with breast cancer. KEYWORDS: breast cancer, disseminated tumor cells, circulating tumor cells, bone marrow aspiration, prognostic/predictive markers, therapy monitoring.