Breast and prostate cancer patients differ significantly in their serum Thymidine kinase 1 (TK1) specific activities compared with those hematological malignancies and blood donors: implications of using serum TK1 as a biomarker.

BACKGROUND: Thymidine kinase 1 (TK1) is a cellular enzyme involved in DNA precursor synthesis, and its activity has been used as a proliferation marker for monitoring malignant diseases. Here, for the first time, we evaluated both TK1 activity and protein levels in sera from patients with different malignancies. METHODS: Serum samples from patients with myelodysplastic syndrome (MDS, n = 22), breast cancer (n = 42), prostate cancer (n = 47) and blood donors (n = 30) were analyzed for TK1 protein and activity levels, using a serum TK1 (STK1) protein assay based on antibodies and an activity assay that measured [(3)H]-deoxythymidine (dThd) phosphorylation. The molecular forms of TK1 in sera from some of these patients were analyzed using size-exclusion chromatography. RESULTS: Mean STK1 activities in sera from MDS, breast and prostate cancer were 11 ± 17.5, 6.7 ± 19 and 1.8 ± 1.4 pmol/min/mL, differing significantly from blood donors (mean ± standard deviation (SD) = 1.1 ± 0.9 pmol/min/mL). Serum TK1 protein (25 kDa polypeptide) levels were also significantly higher in MDS, breast, prostate cancer compared to blood donors (mean ± SD = 19 ± 9, 22 ± 11, 20 ± 12, and 5 ± 3.5 ng/mL, respectively). The STK1 specific activities of sera from patients with MDS and blood donors were significantly higher when compared with activities in sera from breast and prostate cancer patients. Size-exclusion analysis of sera from breast and prostate cancer showed that the detected active TK1 was primarily a high molecular weight complex, similar to the forms found in sera from MDS patients and blood donors. However, Western blotting demonstrated high TK1 25 kDa protein levels in fractions lacking TK1 activity in sera from cases with breast and prostate cancer. CONCLUSIONS: These results demonstrate that there are differences in the specific activities and the subunit compositions of STK1 in hematological malignancies compared with breast and prostate cancer. This fact has several important implications for the use of STK1 as a tumor biomarker. One is that STK1 protein assays may differentiate early-stage tumor development in breast and prostate cancer more effectively than STK1 activity assays.

Accuracy of GFR estimating equations combining standardized cystatin C and creatinine assays: a cross-sectional study in Sweden.

BACKGROUND: The recently established international cystatin C calibrator makes it possible to develop non-laboratory specific glomerular filtration rate (GFR) estimating (eGFR) equations. This study compares the performance of the arithmetic mean of the revised Lund-Malmö creatinine and CAPA cystatin C equations (MEANLM-REV+CAPA), the arithmetic mean of the Chronic Kidney Disease Epidemiology Collaboration equation (CKD-EPI) creatinine and cystatin C equations (MEANCKD-EPI), and the composite CKD-EPI equation (CKD-EPICREA+CYSC) with the corresponding single marker equations using internationally standardized calibrators for both cystatin C and creatinine. METHODS: The study included 1200 examinations in 1112 adult Swedish patients referred for measurement of GFR (mGFR) 2008-2010 by plasma clearance of iohexol (median 51 mL/min/1.73 m2). Bias, precision (interquartile range, IQR) and accuracy (percentage of estimates ±30% of mGFR; P30) were compared. RESULTS: Combined marker equations were unbiased and had higher precision and accuracy than single marker equations. Overall results of MEANLM-REV+CAPA/MEANCKD-EPI/CKD-EPICREA+CYSC were: median bias -2.2%/-0.5%/-1.6%, IQR 9.2/9.2/8.8 mL/min/1.73 m2, and P30 91.3%/91.0%/91.1%. The P30 figures were about 7-14 percentage points higher than the single marker equations. The combined equations also had a more stable performance across mGFR, age and BMI intervals, generally with P30 ≥90% and never <80%. Combined equations reached P30 of 95% when the difference between eGFRCREA and eGFRCYSC was <10% but decreased to 82% at a difference of ≥40%. CONCLUSIONS: Combining cystatin C and creatinine assays improves GFR estimations with P30 ≥90% in adults. Reporting estimates of both single and combined marker equations in clinical settings makes it possible to assess the validity of the combined equation based on the agreement between the single marker equations.

Generation of a new cystatin C-based estimating equation for glomerular filtration rate by use of 7 assays standardized to the international calibrator.

BACKGROUND: Many different cystatin C-based equations exist for estimating glomerular filtration rate. Major reasons for this are the previous lack of an international cystatin C calibrator and the nonequivalence of results from different cystatin C assays. METHODS: Use of the recently introduced certified reference material, ERM-DA471/IFCC, and further work to achieve high agreement and equivalence of 7 commercially available cystatin C assays allowed a substantial decrease of the CV of the assays, as defined by their performance in an external quality assessment for clinical laboratory investigations. By use of 2 of these assays and a population of 4690 subjects, with large subpopulations of children and Asian and Caucasian adults, with their GFR determined by either renal or plasma inulin clearance or plasma iohexol clearance, we attempted to produce a virtually assay-independent simple cystatin C-based equation for estimation of GFR. RESULTS: We developed a simple cystatin C-based equation for estimation of GFR comprising only 2 variables, cystatin C concentration and age. No terms for race and sex are required for optimal diagnostic performance. The equation, [Formula: see text] is also biologically oriented, with 1 term for the theoretical renal clearance of small molecules and 1 constant for extrarenal clearance of cystatin C. CONCLUSIONS: A virtually assay-independent simple cystatin C-based and biologically oriented equation for estimation of GFR, without terms for sex and race, was produced.

The age related association is more pronounced for cystatin C estimated GFR than for creatinine estimated GFR in primary care patients.

OBJECTIVES: There is an age associated change in GFR but this association may be influenced by the method used. The aims of the present study were to assess the association between age and cystatin C and creatinine based glomerular filtration rate estimates in primary care patients, and to determine the proportion of patients with clinically important renal impairment. MATERIALS AND METHODS: 1552 samples with simultaneous requests for creatinine and cystatin C from 1552 primary care patients in the county of Uppsala, Sweden were analysed. MDRD, CKD-EPI and cystatin C equations were used to calculate glomerular filtration rate (GFR) and the associations between GFR and age were explored. RESULTS: The yearly change in cystatin C estimated GFR was 1.24 mL/min/1.73 m(2) while the corresponding decline for creatinine estimated GFR was 0.76 mL/min/1.73 m(2) for MDRD and 0.99 mL/min/1.73 m(2) for CKD-EPI. CONCLUSIONS: The age related association with GFR estimates is smaller for creatinine estimates than for cystatin C estimates. This leads to differences in the number of patients with reduced eGFR detected with the three estimates and the patient treatment will depend on the estimate used. This is not coherent with a good patient care and we thus need to develop new eGFR equations with better agreement between the estimates.

Risk factors for cardiovascular mortality in patients with systemic lupus erythematosus, a prospective cohort study.

INTRODUCTION: Systemic lupus erythematosus (SLE) is a chronic autoimmune disease. Cardiovascular disease (CVD) is common and a major cause of mortality. Studies on cardiovascular morbidity are abundant, whereas mortality studies focusing on cardiovascular outcomes are scarce. The aim of this study was to investigate causes of death and baseline predictors of overall (OM), non-vascular (N-VM), and specifically cardiovascular (CVM) mortality in SLE, and to evaluate systematic coronary risk evaluation (SCORE). METHODS: 208 SLE patients were included 1995-1999 and followed up after 12 years. Clinical evaluation, CVD risk factors, and biomarkers were recorded at inclusion. Death certificates and autopsy protocols were collected. Causes of death were divided into CVM (ischemic vascular and general atherosclerotic diseases), N-VM and death due to pulmonary hypertension. Predictors of mortality were investigated using multivariable Cox regression. SCORE and standardized mortality ratio (SMR) were calculated. RESULTS: During follow-up 42 patients died at mean age of 62 years. SMR 2.4 (CI 1.7-3.0). 48% of deaths were caused by CVM. SCORE underestimated CVM but not to a significant level. Age, high cystatin C levels and established arterial disease were the strongest predictors for all- cause mortality. After adjusting for these in multivariable analyses, only smoking among traditional risk factors, and high soluble vascular cell adhesion molecule-1 (sVCAM-1), high sensitivity C-reactive protein (hsCRP), anti-beta2 glycoprotein-1 (abeta2GP1) and any antiphospholipid antibody (aPL) among biomarkers, remained predictive of CVM. CONCLUSION: With the exception of smoking, traditional risk factors do not capture the main underlying risk factors for CVM in SLE. Rather, cystatin C levels, inflammatory and endothelial markers, and antiphospholipid antibodies (aPL) differentiate patients with favorable versus severe cardiovascular prognosis. Our results suggest that these new biomarkers are useful in evaluating the future risk of cardiovascular mortality in SLE patients.

Cytokines and memory across the mature life span of women.

Increasing evidence suggests a role of the immune system in modulation of cognition, but details on affected memory systems are largely lacking. We therefore aimed to study the relation between selected cytokines and subsets of memory, and the impact of age in these relations. From a random population-based sample (the Betula Prospective Cohort Study), 298 women (age 45-90) were studied in terms of episodic recall and recognition, semantic fluency and knowledge, and prospective memory. Circulating cytokines of relevance for cognition and aging were measured with ELISA. Levels of interleukin (IL)-6 and sIL-2R were significantly and negatively associated with most cognitive variables, while the opposite was true for IL-1ß. Age shared substantial variance with both cytokines and memory, and turned most correlations non-significant when controlled for together with education, BMI and presence of disease. Interactions between age and cytokines were further analyzed in multiple regressions. For IL-6, significant negative interactions with age were found for semantic fluency (p<0.05) and prospective memory (p<0.01), and for sIL-2R in predicting semantic knowledge (p<0.05), indicating an increased negative impact of these cytokines on memory with increasing age. In conclusion, the study indicates a relation between cytokines and memory that appears to be largely mediated by age, and supports the suggestion that cytokine dysregulation with higher age may interact with cognitive aging.

Comparison of cardiovascular prognosis by 3 serum cystatin C methods in the Heart and Soul Study.

BACKGROUND: Cystatin C is a promising new biomarker to determine the estimated glomerular filtration. However, the Siemens' cystatin C assay (Siemens), used in many longitudinal studies, has had limited clinical applicability because it requires a specific, dedicated instrument. Other companies, including Gentian and Roche, have developed cystatin C assays that can be used with most routine clinical chemistry analyzers. METHODS: We compared the agreement of Gentian and Roche with Siemens assays in 948 participants at the baseline visit of the Heart and Soul Study, a cohort of participants with established coronary artery disease who were followed for an average of 8 years. We then compared associations of all 3 cystatin C measures and eGFR-Modification of Diet in Renal Disease (MDRD) with clinical outcomes. RESULTS: The Gentian assay had higher correlation with Siemens (r = 0.96) than did Roche (r = 0.93, P < 0.001). After cross-tabulating quartiles of each cystatin C measure, agreements (κ statistic) were higher for Siemens and Gentian (0.73, 95% CI 0.72-0.75) than for Roche and Siemens (0.64, 0.63-0.66) or for Roche and Gentian (0.69, 0.65-0.71). These differences in agreement had minimal impact on associations with clinical outcomes; the hazard ratios (HRs) for mortality comparing the high vs low quartiles were 3.2 (95% CI 2.1-4.8) for Siemens, 3.1 (CI 2.1-4.7) for Gentian, 3.1 (CI 2.1-4.7) for Roche, and 1.6 (CI 1.1-2.3) for eGFR-MDRD, after multivariate adjustment. CONCLUSIONS: In summary, agreement with the Siemens' assay was modestly higher for the Gentian compared with the Roche assay, although all 3 methods for cystatin C measurement had similar utility as predictors of clinical outcomes.

Performance evaluation of a turbidimetric cystatin C assay on different high-throughput platforms.

OBJECTIVE: The goal with this study was to evaluate the analytical performance of a new cystatin C immunoassay (Tina-quant a Cystatin C, Roche Diagnostics GmbH). The evaluation was carried out at four centers according to a standardized protocol. MATERIAL AND METHODS: The Tina-quant a Cystatin C is a latex particle-enhanced immunoturbidimetric assay. Roche cobas 6000, MODULAR ANALYTICS SWA and COBAS INTEGRA instruments were included in the study. Method comparison studies were carried out against two turbidimetric methods (Dako Cystatin C, Gentian Cystatin C), and one nephelometric method (Siemens N-Latex Cystatin C). RESULTS: Linearity was proven throughout the measuring range from 0.4 to 8 mg/L. Within-run CVs ranged from 0.7-2.8%, and total CVs from 1.4-4.7 % (concentration range 0.6-3.9 mg/L). Comparable results were obtained with paired serum and Li-heparinate plasma samples. Good agreement was achieved in the comparisons between the Tina-quant a Cystatin C assay and the other commercially available cystatin C assays, two different turbidimetric methods (slope range 0.88-1.04, intercept < 0.17 mg/L, r > or = 0.993) and one nephelometric assay (slope range 0.90-1.05, intercept < 0.21 mg/L, r > or = 0.986). CONCLUSIONS: The Tina-quant a Cystatin C assay was shown to be precise and accurate with proven linearity over the measuring range. Good comparability was obtained with other commercially available assays for the determination of cystatin C. The Tina-quant a Cystatin C assay is very well suited for clinical use on routine clinical chemistry analysers to detect renal dysfunction with a 24 h availability.

Cystatin C and modification of diet in renal disease (MDRD) estimated glomerular filtration rate differ during normal pregnancy.

OBJECTIVE: To calculate normal values for estimation of the glomerular filtration rate (eGFR) for pregnant females. eGFR is used to monitor patients with suspected kidney disease and to optimize the dosage of drugs that are eliminated by the kidneys. Plasma creatinine and cystatin C are the two most widely used GFR markers. Both markers are recommended to be automatically reported as estimated GFR. DESIGN: Retrospective study. SETTING: Tertiary university hospital. POPULATION: We have studied creatinine (eGFR(MDRD)) (MDRD, modified diet in renal disease) and cystatin C (eGFR(cystc)) estimated GFR during 52 normal pregnancies from pregnancy week 10 to delivery and postpartum. METHODS: Each woman was sampled repeatedly and the samples were grouped according to gestational age into the following periods: week 7-16; week 18-24; week 24-28; week 28-31; week 31-34; week 34-38; -2-0 weeks prior to delivery and postpartum (> 6 weeks after delivery). MAIN OUTCOME MEASURES: The 2.5 and 97.5 percentiles for these markers were calculated according to the recommendations of the International Federation of Clinical Chemistry on the statistical treatment of reference values. RESULTS: In healthy pregnant females eGFR(cystc) was higher in the first two trimesters and lower prior to delivery in comparison with eGFR(MDRD). eGFR(cystc) and eGFR(MDRD) give different results. No significant correlations between the two estimates were found in any of the time groups. CONCLUSIONS: It is important to distinguish between the two GFR estimates and use separate reference intervals for pregnant females.

Predictors of the first cardiovascular event in patients with systemic lupus erythematosus - a prospective cohort study.

INTRODUCTION: Cardiovascular disease (CVD) is a major cause of premature mortality among Systemic lupus erythematosus (SLE) patients. Many studies have measured and evaluated risk factors for premature subclinical atherosclerosis, but few studies are prospective and few have evaluated risk factors for hard endpoints, i.e. clinically important cardiovascular events (CVE). We investigated the impact of traditional and lupus associated risk factors for the first ever CVE in a longitudinal cohort of SLE patients. METHODS: A total of 182 SLE patients (mean age 43.9 years) selected to be free of CVE were included. Cardiovascular and autoimmune biomarkers were measured on samples collected after overnight fasting at baseline. Clinical information was collected at baseline and at follow up. End point was the first ever CVE (ischemic heart, cerebrovascular or peripheral vascular disease or death due to CVD). Impact of baseline characteristics/biomarkers on the risk of having a first CVE was evaluated with Cox regression. RESULTS: Follow up was 99.5% after a mean time of 8.3 years. Twenty-four patients (13%) had a first CVE. In age-adjusted Cox regression, any positive antiphospholipid antibody (aPL), elevated markers of endothelial activation (von Willebrand factor (vWf), soluble vascular cellular adhesion molecule-1 (sVCAM-1)) and fibrinogen predicted CVEs. Of SLE manifestations, arthritis, pleuritis and previous venous occlusion were positively associated with future CVEs while thrombocytopenia was negatively associated. Among traditional risk factors only age and smoking were significant predictors. In a multivariable Cox regression model age, any positive aPL, vWf and absence of thrombocytopenia were all predictors of the first CVE. CONCLUSIONS: In addition to age, positive aPL, biomarkers indicating increased endothelial cell activity/damage, and absence of thrombocytopenia were independent predictors of CVEs in this prospective study. Our results indicate that activation of the endothelium and the coagulation system are important features in SLE related CVD. Furthermore, we observed that the risk of CVEs seems to differ between subgroups of SLE patients.

Monitoring therapy in canine malignant lymphoma and leukemia with serum thymidine kinase 1 activity--evaluation of a new, fully automated non-radiometric assay.

Thymidine kinase 1 (TK), which is involved in the synthesis of DNA precursors, is only expressed in S-G2 cells. Serum TK levels correlate to the proliferative activity of tumor disease. Determinations of TK levels have so far relied on radio enzyme assay (REA) and experimental ELISA methods, which have limited the clinical use of this biomarker, although recent studies in dogs with malignant lymphoma (ML) demonstrate its wide potential. A non-radiometric method based on a competitive immunoassay with specific anti-3'-azido-deoxythymidine monophosphate (AZTMP) antibodies has been further developed into the fully automated Liaison TK assay (DiaSorin). Sera from healthy dogs (n=30), and dogs with leukemia (LEUK) (n=35), ML (n=84), non-hematological tumors (n=50), and inflammatory disease (n=14) were tested using both methods. Lymphoma and LEUK samples were available before and during chemotherapy. The coefficients of variation for the Liaison TK assay in this study were 6.3 and 3.4% (low/high TK, respectively), and the correlation between TK REA (X) and the Liaison TK assay (Y) was y=0.9203x+1.3854 (R2=0.9501). The TK1 levels measured during chemotherapy gave very clear differences between dogs in complete remission and dogs out of remission. A Tukey-Kramer analysis showed that all LEUKs and MLs out of remission differed significantly from the other groups. The Liaison TK assay showed high precision, high sensitivity and a good correlation to the TK REA. The Liaison TK assay provides valuable clinical information in the treatment and management of canine LEUK and ML, with a potential to be further validated in human trials.

Circadian variability of cystatin C, creatinine, and glomerular filtration rate (GFR) in healthy men during normal sleep and after an acute shift of sleep.

The estimation of the glomerular filtration rate (GFR) is essential for the evaluation of patients with kidney disease and for the treatment of patients with medications that are eliminated by the kidneys. Plasma cystatin C has been shown in several studies to be superior to plasma creatinine for the estimation of GFR. However, there is limited information on the circadian variation of cystatin C and estimated GFR using cystatin C (eGFR(CystC)) or "The Modification of Diet in Renal Disease Study" (MDRD) (eGFR(MDRD)) equations. We studied the circadian variation of cystatin C and creatinine during night- and day-sleep conditions in seven healthy volunteers. Serum samples were collected every hour (48 samples per individual) to evaluate the effect of different sampling times on the test results. The median intra-individual coefficients of variations for the studied markers were 4.2% for creatinine, 4.7% for eGFR(MDRD), 5.5% for cystatin C, and 7.7% for eGFR(CystC). Neither cystatin C nor creatinine differed significantly between the night- and day-sleep conditions. Cystatin C differed significantly with time of day (p=.0003), but this was not the case for creatinine (p=.11). The circadian variation of cystatin C was minor. Small but significant increases in creatinine values and a decrease of eGFR(MDRD) were observed after food intake. Thus, cystatin C and creatinine sampling does not have to be restricted to specific times of the day.

Biochemical effects of consumption of eggs containing omega-3 polyunsaturated fatty acids.

Today, eggs with an increased content of -3 fatty acids are available but there are few publications on the effects of consumption of such eggs on the lipoproteins and acute phase markers in humans. The aim of the present study was to evaluate the effects of consumption of standard eggs and -3 enriched eggs on lipoproteins, glucose and inflammation markers. Nineteen healthy volunteers consumed one extra egg per day of either standard eggs or omega-3 enriched eggs in a double-blind, cross-over study. The duration of each period was 1 month. The effects of the different egg diets on apolipoprotein A1 and B (Apo A1 and B), lipoprotein (a), creatinine, cystatin C, C-reactive protein, serum amyloid protein A, interleukin 6, triglycerides, glucose, total-, high-density lipoprotein and low-density lipo-protein cholesterol concentrations were analyzed. Addition of one regular egg per day to the normal diet had no negative impact on blood lipids or inflammation markers. Consumption of omega-3 enriched eggs resulted in higher levels of ApoA1, lower ApoB/ApoA1 ratio and lower plasma glucose. These effects have been associated in previous studies with a reduced risk for cardiovascular mortality and diabetes.

Patient selection has a strong impact on cystatin C and Modification of Diet in Renal Disease (MDRD) estimated glomerular filtration rate.

OBJECTIVE: Estimation of the glomerular filtration rate (GFR) is essential for the evaluation of patients with kidney disease, and for correct dosage of drugs that are eliminated from the circulation by the kidneys. In most cases GFR is estimated based on serum creatinine and the Modification of Diet in Renal Disease (MDRD) formula. As both cystatin C and creatinine are used for the determination of GFR it is important to investigate if estimated GFR by the two methods differ in various patient groups. DESIGN AND METHODS: We have compared cystatin C and MDRD estimated GFR calculated from the same request from primary care units (n=488), a cardiology ward (n=826), the cardiointensive care unit (n=1026), two oncology wards (n=919 and 1021), and the neurosurgical intensive care unit (n=1515) in an observational cross-sectional study. RESULTS: We found better agreement between the two GFR estimates in samples from primary care patients and patients in the cardiology wards, than in samples from oncology wards or the neurosurgical intensive care unit. In the latter settings there was a pronounced difference between the two GFR estimates. CONCLUSION: The comparisons show that differences in patient selections have a strong impact on the agreement between cystatin C and MDRD estimated glomerular filtration rate.

Comparison between chicken and rabbit antibody based particle enhanced cystatin C reagents for immunoturbidimetry.

We have compared three commercial particle enhanced cystatin C reagents. One of the reagents utilizes chicken antibodies and the other two reagents are rabbit antibody based. We show that the chicken antibody based reagent yields a higher delta absorbance when reacting with the antigen. IgY coupled to latex particles show a strong scatter response even at high antigen concentrations in contrast to the steep decline in scatter previously reported for IgY antibodies in solution. The reagent also showed a low CV for duplicate samples. Laying hens thus seems as an interesting source of antibodies for particle-enhanced immunoassays.

Serum cystatin C is associated with other cardiovascular risk markers and cardiovascular disease in elderly men.

Renal dysfunction is associated with increased cardiovascular morbidity and mortality. The aim of this cross-sectional study was to investigate the relationship between the glomerular filtration marker cystatin C and other cardiovascular risk markers and morbidity in elderly males. Cystatin C was measured in a group of 77-year-old males (n=792) and compared cystatin C with other known risk markers for cardiovascular disease. Cystatin C values were significantly increased in individuals with diabetes (p=0.05) and cardiovascular diseases (p<0.0001). There were significant correlations between cystatin C values and body mass index, HbA1c, insulin, triglycerides and hsCRP.