New biomarkers: added clinical value explained Can biomarkers improve clinical decisions?

Transcription

1 New biomarkers: added clinical value explained Can biomarkers improve clinical decisions? Alex Lefevre UZ Leuven 30/11/2012

2 Agenda Introduction Sensitivity Specificity - ROC Biomarkers with added medical value: IL-6 Biomarkers with added medical value: S100 Biomarkers with added medical value: TnT hs Biomarkers with added medical value: NT-proBNP Biomarkers with added medical value: Cystatin C Conclusion

3 Agenda Introduction Sensitivity Specificity - ROC Biomarkers with added medical value: IL-6 Biomarkers with added medical value: S100 Biomarkers with added medical value: TnT hs Biomarkers with added medical value: NT-proBNP Biomarkers with added medical value: Cystatin C Conclusion

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5 Status 2010

6 Biomarkers: an ongoing evolution

7 The value of diagnostics 1 Costs vs. decision-making Hospital costs Healthcare decision-making Diagnostics While diagnostics comprise less than 5% of hospital costs and about 1,6% of all medical care costs, their findings influence as much as 60% - 70% of healthcare decision-making 1 Source: The Value of diagnostics July 2005 Sept 2009, The Lewing Group

8 Definition of Biomarker Biological Marker (Biomarker): A characteristic that is objectively measured and evaluated as an indicator of normal biological processes, pathogenic processes, or pharmacologic responses to a therapeutic intervention. Biomarkers Definitions Working Group, Clin. Pharmacol. Ther2001; 69, Purpose: Biomarkers are used in clinical practice to identify risk for disease, diagnose disease and its severity, guide intervention strategies, and monitor patient responses to therapy. When used in a clinical research setting, biomarkers may predict whether a drug or other intervention is safe and effective in a shorter time and at lower cost than clinical outcomes studies. CA Altar, ClinPharmacolTher2008; 83:

9 Characteristics of an ideal biomarker Safe to measure Easy to measure Low cost to measure Cost of follow-up tests is relatively low Proven treatment to modify biomarker exists Biomarker modification proven to protect against disease/outcome Consistent across genders and ethnic groups

10 Example : Point-of-care testing in Neonatology Saving time, money and blood Results on the spot Up to 50 % less transfusions Simplification blood collection : capillary blood

11 Example : NT-ProBNP Prognostic value in chronic heart failure Reduce length of stay in hospital Less re-hospitalisations

12 Agenda Introduction Sensitivity Specificity - ROC Biomarkers with added medical value: IL-6 Biomarkers with added medical value: S100 Biomarkers with added medical value: TnT hs Biomarkers with added medical value: NT-proBNP Biomarkers with added medical value: Cystatin C Conclusion

13 What about new diagnostic tests?

14 The sensible use of biomarkers in clinical practice Healthy people Cutoff ill patients Marker concentration Ideal situation: does not exist

15 The sensible use of biomarkers in clinical practice Cut-off Healthy people ill patients Marker concentration False negative and false positive findings are a function of the chosen cut-off: the higher the specificity, the lower the sensitivity and vice versa.

16 The sensible use of biomarkers in clinical practice Cut-off Healthy people ill patients High specificity less healthy are false positive, but more patients having a disease are false negative Marker concentration

17 The sensible use of biomarkers in clinical practice Healthy people Cutoff ill patients High sensitivity all people having a disease are being detected, but more false positives Marker concentration

18 Sensitivity Specificity NPV -PPV Biomarker Disease Present Absent Positive A B A + B Negative C D C + D A + C B + D A + B + C +D Disease prevalence: A + C / A + B + C + D Specificity: D / B + D Sensitivity: A / A + C Negative likelihood ratio (LR-): (1 sensitivity)/specificity Negative predictive value: D / C + D Positive likelihood ratio (LR+): sensitivity/(1-specificity) Positive predictive value: A / A +B

19 The Area Under an ROC Curve The accuracy of the test depends on how well the test separates the group being tested into those with and without the disease in question = excellent (A) = good (B) = fair (C) = poor (D) = fail (F)

20 Agenda Introduction Sensitivity Specificity - ROC Biomarkers with added medical value: IL-6 Biomarkers with added medical value: S100 Biomarkers with added medical value: TnT hs Biomarkers with added medical value: NT-proBNP Biomarkers with added medical value: Cystatin C Conclusion

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22 IL-6: An early, sensitive marker of inflammation often the first indication of critical disease

23 IL-6 Biochemistry - Summary 184 amino acids glyco-protein and has a molecular weight of about 26 kd. Released by many cells. Multiple biological actions on different types of target cells via receptors and gp130. Substantial impact on the immune system, hematopoesis, and inflammation. Longest half-life of the pro-inflammatory mediators in the systemic circulation. Activation time: very short Half-life time: about 1 hour IL-6 triggers the production and release of CRP in the liver. IL-6 triggers to a certain extent procalcitonin. Early and sensitive alarm marker for (systemic) inflammation and sepsis and neonatal sepsis.

24 IL-6 Procalcitonin -CRP Polytrauma patient gallbladder surgery Day 4-5 : sepsis Patient 205, male, 65 years data Feldkirch IL-6 CRP PCT IL-6 [pg/ml] CRP [mg/l] PCT [ng/m L] IL CRP PCT day

25 IL-6 Procalcitonin -CRP Day 5: severe sepsis Day 6-8: signs of sepsis Patient 210, male, 70 years data Feldkirch IL-6 CRP PCT IL-6 [pg/ml] CRP [mg/l] PCT [ng/ml] IL CRP PCT day

26 IL-6 Procalcitonin -CRP Day 1-3 sepsis Patient 204, male, 77 years data Feldkirch IL-6 CRP PCT IL-6 [pg/ml] CRP [mg/l] PCT [ng/ml] IL CRP PCT day

27 IL-6: Values of Crit. Care Patients IL-6 (pg/ml) 7 pg/ml * Median Mean Min Max N SIRS * Reference Healthy Sepsis Severe Sepsis Septic Shock n= 281 IL-6 values increase steadily with severity of the disease.

28 Procalcitonin: Values of Crit. Care Patients PCT (ng/ml) 0.05 ng/ml * Median Mean Min Max N SIRS * Reference Healthy Sepsis Severe Sepsis Septic Shock n= 281 Procalcitonin distinct increase are seen only from severe sepsis on.

29 Sepsis Markers IL-6 is an early and sensitive alarm marker for (systemic) inflammation and sepsis. Initiated by IL-1 and TNFα appearenance assumed within 1hour half-life ~ 1 hour (hs)crp Initiated by IL-6 appearance ~ h after IL-6 half-life ~ approx. 20 h PCT due to its later increase and longer half-life is a confirmation of (bacterial) sepsis. Initiated by endotoxins, IL-6 and (??) not known appearance ~ 24 hours after IL-6 half-life ~ approx. 24 h The three markers do not replace each other. In contrary -used combined they add significant information regarding the status and the severity of the disease.

30 Agenda Introduction Sensitivity Specificity - ROC Biomarkers with added medical value: IL-6 Biomarkers with added medical value: S100 Biomarkers with added medical value: TnT hs Biomarkers with added medical value: NT-proBNP Biomarkers with added medical value: Cystatin C Conclusion

31 S100 The Parameter: The unfractionated protein mixture isolated from bovine brain was solublein 100% saturated ammonium sulfate solution.

32 S100: The Parameter S100 mrna Distribution S100 belongs to a family of calcium-binding proteins. Up to now 21 members are identified It s molecular weight is approximately 10.5 kd. S100 A1(α) and S100 B (β) occur mostly as homo- or hetero-dimers. S100A0 S100A1 colon, S100A2 muscle, S100A4 spleen, stomach, S100A7 thymus, S100A8 S100A9 S100A10 S100B colon, skin, S100C S100P CaLB3 S100F kidney, heart brain, aorta, bladder, fat, heart, kidney, intestine, pancreas,skeletal muscle, spleen, testes, uterus, melanoma cells kidney, lung, skeletal mammary epithelial lung, skeletal muscle, nerve, thymus, fibroblasts, bone marrow, lymphocytes heart, lung, stomach, fibroblast, neuroblasts (monocytes monocytes kidney, nerve brain, aorta, bladder, fat, intestine, pancreas, skeletal muscle, testes, nerve, melanoma cells heart, kidney, lung, adrenal gland, fibroblasts placenta liver, intestine skin

33 S100B: The Parameter S100 A1 und B is predominantly found in astrocytes, glia cells, and Schwann cells It can be detected in the systemic circulation when the BBB opens temporarily. S100 S100 Urban & Fischer 2003 Roche Lexikon Medizin

34 S100: Three distinct clinical indications Monitoring melanoma patients stage III and IV Exclusion of minor traumatic brain injury (MTBI) Monitoring primary/secondary brain injury in severe traumatic brain injury intracranial hemorrhage e.g. after severe stroke or subarachnoidal hemorrhage patients after cardiac or bypass surgery sepsis patients

35 S100 Values in Mild Traumatic Brain Injury Patients and Healthy patients Results

36 S100 Values in Mild Traumatic Brain Injury Patients and Healthy patients Results

37 Prospective Study Results: Contingency Table Patients GCS 13 15: N = 1309 [µg/l] S100-Value * S µg/l S100- < 0.10 µg/l CCT + CCT * CCT- *= p<0.001 U-Test CCT+ Sensitivity: 98.9 % ( % CI 95) Specificity: 31.6 % ( % CI 95) NPV: 99.7 % ( % CI 95) PPV: 10 % ( % CI 95) Biberthaler P. et al. SHOCK, Vol. 25, No. 5, pp , 2006 *= µg/l

38 Prospective Study Results S100 Values by GCS Score CCT+ CCTcut-off 0.1µg/L N =1309 S-100B in [µg/l] * * * # *=p<0.001 U-test CCT+ vs. CCT- #=p<0.05 ANOVA/Dunn s GCS 15 vs. 13/14 GCS Score Biberthaler P. et al. SHOCK, Vol. 25, No. 5, pp , 2006

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40 Methodology Studies containing adult patients with nonpenetrating head injury GCS score 13 S100B serum & cranial CT scan within 24 hours of injury 272 abstracts & 49 full manuscripts were chosen 12 studies were found eligible and selected containing 2466 patientsfrom Sweden, Norway, Germany, France, Slovenia, Brazil, and the United States. Time from injury to S100B sampling ranged from less than 3 hours to less than 24 hours. All were emergency departments Cutoff S100B = 0,10 µg/l 40

41 Table showing the 12 eligible studies with true positives (S100+, CT+), false positives (S100+, CT ), false negatives (S100, CT+), true negatives (S100, CT ). NPV = 99,2% The studies were graded and assigned a quality rating with respect to the key question according to the Centre of Evidence Based Medicine (CEBM) criteria. The final recommendation (A to D) is based on CEBM criteria; grade A referring to consistent class 1 studies, grade B for consistent level 2 or 3 studies (or extrapolations from level 1 studies), grade C for level 4 studies (or extrapolations from level 2 or 3 studies), and grade D for level 5 studies. 41

42 The 12 eligible studies show similar results with high individual sensitivities. The pooled sensitivity for all studies was 97%(95% CI 91% 99%) and the pooled specificity 40%(95% CI 30% 51%). 42

43 Conclusions: Very high individual NPVs (90% 100%) for the ability of a negative (under cutoff level = 0,10µg/L) S100B level to predict a normal CT scan. Sensitivity NPV D Dimer Pulmonary embolism 95% 99% D Dimer Deep vein thrombosis 88/90/92% 99/96/84% Myoglobine Acute Myocard infarction 61% 87% Troponin T Acute Myocard infarction 73% 91% S100B Minor Head Injury 97% 99% S100B (half-life ): should therefore be taken within 3 hours of injury. S100B does is not affected by alcohol intoxication in head-injured patients. S100B has to be used as a rule out marker for minor head injuriesand by this helps to reduce the number of routine CT examinations. S100B gives additional information to CCT scan in axonal injury or early after accident 43

44 Agenda Introduction Sensitivity Specificity - ROC Biomarkers with added medical value: IL-6 Biomarkers with added medical value: S100 Biomarkers with added medical value: TnT hs Biomarkers with added medical value: NT-proBNP Biomarkers with added medical value: Cystatin C Conclusion

45 History of markers for acute myocardial infarction AST enzyme in AMI CK enzyme in AMI CK-MB enzyme inhition activity CK / LD isoenzymes electrophor. Myoglobin RIA CK-MB mass immunoassay ctnt ctni hs-tns precommercial TnT hs launched st WHO criteria for AMI th WHO criteria for AMI 1979 WHO MONICA criteria for AMI Redifinition of AMI 2007 Universal definition of MI ESC guidelines - NSTEMI 2010

46 Temporal pattern of release after the onset of AMI Kinetic Magnitude of elevation varies with infarct size French, J. and White, H. (2004). Heart 90(1): ULRR = upper limit of reference range 30 November 2012 page Roche

47 Biomarkers in Acute Coronary Syndrome Detection, sensitivity and specificity for AMI Markers First Detection Duration of Detection Sensitivity Specificity Myoglobin h 8-12 h CK-MB 2 3 h 1-2 d Troponin I 3 4 h 7-10 d Troponin T 3 4 h 7-14 d CK Total 4 6 h 2-3 d Individuals can deviate from this pattern, it has been reported that 15% of patients have discordant TnT and CK-MB results, perhaps due to timing but also inter and intraindividual variations. Successful reperfusion in AMI patients accelerates the cardiac marker release. *TIMI: Thrombolysis in Myocardial Infarction French JK, White HD. Clinical implications of the new definition of myocardial infarction. Heart (2004) 90: November 2012 page Roche

48 Cardiac Troponin T and Troponin I Major circulating forms Ternary complex Binary complex Free form Both ctnt and ctni are found circulating in different forms: as ternary complex, binary complex (mainly TnI-TnC) and free forms 1 Free TnT is the major TnT circulating form 2 Cardiac TnI is highly susceptible to proteolysis and enzymatic modification. Multiple circulating TnI forms are described 3 : Proteolysed forms Complexed to heparin or not Phosphorylate forms Oxidized and reduced forms Degradation can occur both in vitro and in vivo challenging consistency of TnI measurements 1 Korff S et al. Heart, 2006; 92: ; 2 Wu A et al. Clin Chem 1998; 44: ; 3 Katrukha A et al. Clin Chem 1997; 43:

49 Universal definition of myocardial Expert consensus document by the joined ESC/ACCF/AHA/WHF task force infarction, 2007 The term myocardial infarction should be used when there is evidence of is evidence of myocardial ischemia. Under these conditions any one of the following criteria meets the diagnosis for myocardial infarction: Detection of rise and/or fallof cardiac biomarkers (preferably troponin) with at least one value above the 99 th percentile in combination with at least one of the following: Symptoms of ischemia ECG changes indicative for ischemia : ST-T changes or new LBBB or new Q-waves Imaging evidence of new loss of viable myocardium or new regional wall motion abnormality Optimal precision (coefficient of variation [CV]) at the 99 th percentile upper reference limit [URL]) should be defined as 10%. The use of assays that do not have independent validation of optimal precision (CV 10%) is not recommended. Thygesen K et al. JACC 2007; 50 (22):

50 Universal definition of MI By the joined ESC/ACCF/AHA/WHF task force MI definition Increase of Troponin Thygesen K et al. Eur Heart J 2012

51 TnT-hs distribution of values: Reference population 90 The overall imprecision of the ctnt-hs assay fulfils current guideline recommendations th percentile URL value from 533 healthy volunteers : 14 pg/ml and 10% CV at 13 ng/l 60 Frequency th percentile 14,2 ng/l (reference limit) Elecsys TnT-hs [ng/l] Saenger AK et al. Clin Chim Acta 2011; 412(9-10): The limit of blank and limit of detection were 3 and 5 ng/l, respectively

52 Troponin assay precision requirements The first criteria of Fred Apple s scorecard 1 Analytical criterion recommended by the international guidelines: a total imprecision equal to or less than 10% at the 99 th percentile value 2,3 Total imprecision at the 99 th percentile, CV % 10 Acceptance designation Guideline acceptable >10 to 20 Clinically usable > 20 Not acceptable 1. Apple FS. A new season for cardiac troponin assays: It s time to keep a scorecard. Clin Chem 2009;55: Thygesen K, Alpert JS, White HD; Joint ESC/ACCF/AHA/WHF Task Force for the Redefinition of Myocardial Infarction: Universal definition of myocardial infarction. JJAm Coll Cardiol 2007; 50, Morrow DA, Cannon CP, Jesse RL et al. National Academy of Clinical Chemistry laboratory medicine practice guidelines; clinical characteristics and utilization of biochemical markers in acute coronary syndromes. Clin Chem 53,

53 Progress in cardiology Influence of sensitivity and precision on early detection Normal ctn turnover range of Tn rise 2-6 h post event 6-8 h post event Adapted from: Hochholzer W et al. Am Heart J 2010; 160(4):

54 Acute chest pain AMI 2 problems: Rule in & Rule out O h 1 h 2 h 3 h 4 h 5 h 6 h 7 h ECG 1 CTn 2 CTn Delay in adequate therapy Morbidity Cost Patient: Anxiety,

55 Clinical validation Non STEMI patients ctnt-hs (% positive) depending on time after onset of symptom in patients developing NSTEMI 100 % ctnt-hs all ctnt-hs early (< 4h) ctnt- hs late (>4h) The ctnt-hs assay identified 20% more patients with a final diagnosis of NSTEMI than ctnt Gen 4 Time to diagnosis of NSTEMI was made a mean of 175 min (2h55 min) earlier with ctnths than with ctnt Gen reference ctnt Gen 4 Admission1h 2h 3h 4h 5h 6h 24h A 2 cd measurement within 3-5 h after admission considerably improved ctnths performance Adapted from: Giannitsis et al. Clin Chem 2010; 56 (2), November 2012 page Roche hours after admission

56 Clinical added value Earlier diagnostic of AMI, the 1 st supporting facts The diagnostic performanceof the novel Tn assays was very high at presentationand significantly higher than the standard TnT gen 4 assay TnT-hs improved by 21%the earlier diagnostic of AMI TnT-hs demonstrated the highest sensitivity (95%) The high NPV (99%)confirmed that the novel sensitive Tn assays may help for the reliable rule out of AMI 30 November 2012 page Roche

57 Cardiac TnT-hs to diagnose MI Using the 99 th percentile and criteria for fall and rise of ctn Patient with symptoms of MI* Baseline ctnt-hs <14 ng/l 14 to < 53 ng/l 53 ng/l Retest ctnt-hs 6 hours later to rule out MI Retest ctnt-hs 3 hours later Retest ctnt -hs 3 hours later Change <50% Change 50% Change <20% Change 20% Adverse prognosis Retest ctnt-hs at 6, 12 hours Myocardial infarction * To make the diagnosis of MI, there must also be evidence of ischemia manifesting as symptoms or evident on electrocardiogram, imaging evidence of new loss of viable myocardium or new regional wall motion abnormality Adapted from: White H Am Heart J 2010;159(6): November 2012 page Roche

58 20 Keller T et al. New England J Med 2009; 361: Riechlin T et al. New England J Med 2009; 361: Giannitsis et al. Clin Chem 2010; 56: Weber M et al. Am Heart J 2011; 162: Source: Eur Heart J 2011; 32(23): November 2012 page Roche

59 Absolute and relative changes in cardiac troponin Reichlin T et al. Circ 2011; 124(2): ROC curves of 1-hour (A) and 2-hour (B) ctn changes ctnt-hs absolute change Sensitivity Sensitivity ctni Ultra absolute change ctnt-hs relative change ctni Ultra relative change specificity A specificity B ROC-optimised delta change for ctnt-hs: 7 ng/l The accuracy of absolute ctn changes for the diagnosis of AMI was high for both assays after both 1 and 2 hours, and was significantly higher than that of relative changes at both time points 30 November 2012 page Roche

60 Absolute and relative changes in cardiac troponin Mueller M et al. ClinChem2012; 58(1): Prediction of non-stemi in the entire study population A 1.0 B 1.0 Prediction of non-stemi in ACS patients Sensitivity Absolute change (AUC=0.898)* Relative change (AUC=0.752) Baseline ctnt-hs (AUC=0.731) Peak ctnt-hs (AUC=0.830) Sensitivity Absolute change (AUC=0.941)* Relative change (AUC=0.741) Baseline ctnt-hs (AUC=0.836) Peak ctnt-hs (AUC=0.894) specificity 1 specificity ROC-optimised absolute delta change: 9.2 ng/l ROC-optimised absolute delta change : 6.9 ng/l Absolute ctnt-hs changes had the highest AUC for the prediction of non-stemi 30 November 2012 page Roche

61 Elevated ctnt-hs in patients with ACS Celik S. et al. Clin Res Cardiol 2011;100(12): Results: Kaplan-Meier survival A: survival B : event free survival (composite endpoint, death/mi) among patients with ctnt Gen4 below the 99 th percentile (<0.01 µg/l) according to ctnt-hs level (A) 100 Survival probability (%) ctnt-hs <14 ng/l (n=285; events=1) ctnt-hs 14 ng/l (n=169; events=9) Logrank p= (B) Survival probability (%) ctnt-hs 14 ng/l (n=169; events=12) Logrank 6.42 p=0.011 ctnt-hs <14 ng/l (n=285; events=6) Time (days) Time (days) Rates of both all-cause of death and the composite of death/non-fatal MI were significantly higher when ctnt-hsvalues exceeded the 99 th percentile, particularly in patients with ctntgen4 <99 th percentile

62 Cardiac TnT-hs predicts risk in general population In coronary heart disease, death & heart failure -ARIC study ctnt-hs levels are associated with incident CHD, mortality and HF in individuals from a general population without known cardiovascular disease (n=9698 individuals) Risk of Coronary Heart Disease over time 30 by ctnt-hs level 30 Risk of death over time by ctnt-hs level 30 Risk of Heart Failure over time by ctnt-hs level ctnt-hs quintile Quintile Quintile 4 Risk (%) Quintile 3 Quintile 2 Quintile Follow-up years Follow-up years Follow-up years The ctnt-hs assay identifies patients at risk of future cardiovascular events Source: Saunders, JT. et al. (2011) Circ 23(13): November 2012 page Roche

63 Conclusions The 99th percentile hs-ctn URL value should be used as the decision limit for the diagnosis of AMI. The ctnt-hs assay meets the 10% CV criterion at the 99 th percentile URL. The diagnosis of acute myocardial necrosis requires a significant change with serial testing. At low baseline ctn (around the 99th percentile): marked change is required (> 50% ) In case of markedly elevated baseline ctn: change of > 20% is required The improved sensitivity leads to an earlier diagnosis of AMI: Sensitive troponin assays detect patients with AMI already at 2 hours after onset of symptoms The new ctnt-hs assay helps for risk stratification of ACS The ctnt-hs assay identifies patients at risk of future cardiovascular events As recommended by the guidelines, ctn results have to interpreted with the clinical presentation of the patient; and acute changes in serial samples are required to differentiate acute from chronic myocardial damage

64 Agenda Introduction Sensitivity Specificity - ROC Biomarkers with added medical value: IL-6 Biomarkers with added medical value: S100 Biomarkers with added medical value: TnT hs Biomarkers with added medical value: NT-proBNP Biomarkers with added medical value: Cystatin C Conclusion

65 Heart failure in Belgium: HF has a significant impact on the health care costs in Belgium. The mean in-hospital stayfor HF = 13 days 1 Total in-hospital stay for HF of 254,826 days. HF accounts for 5% of all emergencymedical admissionsto a hospital 1 30%of the patients are re-admissed within 6 months and50% within a year, depending on the level of severity % hospitalisation costs are due to rehospitalisations. The mortality rate is ~50% over 5 year. 30/11/ Data RIZIV Data Unamec 2011

66 Diagnosis of heart failure Overview HF is a diagnostic challenge 50 % of the patients referred to cardiologists have originally been misdiagnosed Patients usually are older and have more than one disease Diagnosis of underlying disease is crucial for treatment Diagnostic procedure Is it heart failure? What is the underlying disease? Diagnostic measures Symptoms Physical findings, clinical signs Blood marker (NT-proBNP) Chest X-ray Echocardiography Braunwald E, 2001.

67 The diagnosis of heart failure was confirmed in 40 (29%) patients. An abnormal ECG did not add any further predictive value to that of NTproBNP. Zaphiriou A. et al. The diagnostic accuracy of plasma BNP and NT-proBNP in patients referred from primary care with suspected heart failure: Results of the UK natriuretic peptide study. Eur J Heart Fail 2005;7: Nielsen et al. N-terminal pro-brain natriuretic peptide for discriminating between cardiac and non-cardiac dyspnoea. Eur J Heart Fail, 2004,6:63-70.

68 NT-proBNP Biology and biochemistry I 1988 Brain natriuretic peptide (BNP) is discovered Natriuretic peptides are released when the heart muscle is stretched: Atrial Natriuretic Peptide (ANP): from myocardial cells of the atrium. Brain Natriuretic Peptide (BNP): from myocardial cells of the ventricles, BNP is also formed in the brain. Other natriuretic peptides C-type natriuretic peptide (CNP): mainly present in vascular tissue Urodilatin: produced by the kidney, function is unclear Spanuth, 2004; Haas, hypertrophy Normal heart ANP NT-proANP BNP NT-proBNP Heart failure ANP NT-proANP BNP NT-proBNP

69 NT-proBNP Biology and biochemistry II Action of BNP: reduces the volume load in the heart by increasing urination (= diuretic) increasing elimination of sodium (= natriuretic) reducing blood pressure i.e., BNP counteracts activated reninangiotensin-aldosterone system Structure 32 amino acids 17-amino acid ring structure essential for activity NT-proBNP is formed by cleavage of probnp pre-probnp probnp N-terminal probnp BNP Half life 120 min. 20 min. Biological effects No Yes Heart muscle Spanuth, 2004; Haas, 2005.

70 NT-proBNP Differences vs. BNP NT-proBNP vs. BNP NT-proBNP Hormonally inactive BNP Hormonally active 76 amino acids 32 amino acids Half-time: min Higher plasma levels, cumulative concentrations reflect whole cardiac situation Up to 3 days stable at room temperature No interference by therapeutics 20 min Lowerplasma levels, depend on the patient's situation at the moment Less stable Interference by therapy with BNP or degradation inhibitors of natriuretic peptides Arguments for NT-proBNP Larger peptide, longer half-life More diagnostic power, less susceptible to clearance mechanisms Higher plasma concentration Easier to measure, more diagnostic resolution Lower intra-individual variation better early detection Analytical differences More stable, more precise, better detection Yeo et al., 2003; Mueller et al., 2004;

71 The natriuretic peptides Biochemistry of NT-proBNP cardiomyocyte -26 amino acid 108 pre-probnp probnp signal NT-proBNP BNP BNP (physiologically active)

72 The Natriuretic Peptides The peptide family 1988 Brain natriuretic peptide (BNP) is discovered Four peptides related with each other regarding biochemisty and physioglogical function: - atrial natriuretic peptide (ANP) - brain (or B-type) natriuretic peptide (BNP) - C-type natriuretic peptide (CNP) - urodilatin, a slightly extended form of ANP Biochemistry - common 17-amino acid ring structure - ring structure highly conserved - 11/17 amino acids are homologous -ring structure essential for physiological activity Physioglogical function diverse effects on tissues involved in sodium regulation and blood pressure homeostasis

73 NT-proBNP correlates with NYHA class Acute phase Chronic phase Januzzi J.L. et al. Use of Amino-Terminal Pro-B-Type Natriuretic Peptide to Guide Outpatient Therapy with Chronic Left Ventricular Systolic Dysfunction. J Am Coll Cardiol 2011;58: Januzzi J. L. et al.: Utility of Amino-Terminal Pro Brain Natriuretic Peptide Testing for Prediction of 1-Year Mortality in Patients With Dyspnea Treated in the Emergency Department. Arch Intern Med. 2006;166:

74 Have natriuretic peptides an added clinical value? Januzzi J.L. et al. The N-Terminal Pro-BNP Investigation of Dyspnea in the Emergency Department (PRIDE) Study. Am J Cardiol 2005;95:

75 Cut-off values for NT-proBNP: chronic heart failure <400 pg/ml 75

76 Cut-off values for NT-proBNP: acute heart failure PPV = 92% PPV = 76% PPV = 82% NPV = 98% 76

77 Negative predictive value

78 RULE-OUT of heart failure = faster

79 Relationmortality/BNP-concentration acute HF

80 Change of NT-proBNP during hospital stay predicts death or readmission P. Bettencourt et al. Circulation 2004; 110: Cumulative survival and no-readmission according to the change of NT-proBNP levels during hospital stay n Events Decrease 82 27% > 30% Change < 30% 49 49% Increase 25 84% > 30%

81 Creatinine values Relationship NT-proBNP values and renal function Hemodynamics Neurogenic reflexes Hormones (RAAS, NP s) N = NT-proBNP values

82 Agenda Introduction Sensitivity Specificity - ROC Biomarkers with added medical value: IL-6 Biomarkers with added medical value: S100 Biomarkers with added medical value: TnT hs Biomarkers with added medical value: NT-proBNP Biomarkers with added medical value: Cystatin C Conclusion

83 Acute Kidney Injury Biomarkers

84 Cystatin C: properties and advantages as a GFR marker Non-glycosylated, single chain protein of 13.3 kda molecular weight. Synthesized by all nucleated cells at a constant rate. Freely filtered by the glomerulus. Reabsorbed by tubular epithelial cells and subsequently degraded. Serum concentration not affected by muscle mass, gender, inflammation, and age. Elimination only via filtration (no secretion, no reentering in circulation). These characteristics make Cystatin C a valuable marker for the estimation of GFR.

85 Kidney Biomarkers The quest for the practical Kidney function marker CREATININE: Well established but, important blind window CYSTATIN-C : Novel marker with improved sensitivity PROTEIN: late marker for protein leakage N-GAL: Marker for acute kidney failure CREATININE blind window Cystatin-C 30 November, 2012 page Roche

86 How to determine kidney function

87 The problem of the Creatinine-blind area in serum- Creatinine based GFR assessments GFR :Kidney Function Cystatin C Creatinine GFR GFR Schematic correlation of GFR with serum Creatinine and Cystatin C concentrations (redrawn from Künzi T (2005)) Creatinine-blind Area The same change in GFR causes a greater change in Cystatin C- than in Creatinine concentration. Thus, early changes in GFR can be detected easily employing Cystatin C-Assays, while may go undetected with Creatininebased GFR assessments. Crea Concentration Cys C Bio-marker concentration

88 Cystatine C vs. Serum Creatinine 30 November, 2012 page Roche

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90 Agenda Introduction Sensitivity Specificity - ROC Biomarkers with added medical value: IL-6 Biomarkers with added medical value: S100 Biomarkers with added medical value: TnT hs Biomarkers with added medical value: NT-proBNP Biomarkers with added medical value: Cystatin C Conclusion

91 Summary IL-6 is an unspecific inflammation marker...but is alsosensitive, an early marker, on-time (due to short half-life) and mirrors the actual situation regarding inflammation. S100 below 0,10 µg/l excludes brain injury with high likelyhood and helps to reduce the number of routine CT scans. ctnt-hs has an improved sensitivity that leads to an earlier diagnosis of AMI and helps for risk stratification of ACS (NT-pro)BNP is a very powerful marker for rule-in/rule-out & prognosis of heart failure and NT-proBNP guided therapy reduces mortality and hospitalization in heart failure patients Cystatin C is a more sensitive marker of GFR than creatinine and is not affected by age, diet or body mass.

92 Biomarkers are only a mark Questions? 92

93 We Innovate Healthcare 93