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Thermo Scientific B·R·A·H·M·S CT-proAVP LIA for use in endocrinology February 2011.

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Presentation on theme: "Thermo Scientific B·R·A·H·M·S CT-proAVP LIA for use in endocrinology February 2011."— Presentation transcript:

1 Thermo Scientific B·R·A·H·M·S CT-proAVP LIA for use in endocrinology February 2011

2 2 Vasopressin & CT-proAVP - FAQs  What is Vasopressin (CT-proAVP) and where is it produced?  What is the physiological role of Vasopressin?  Why not simply measure Vasopressin?  Is CT-proAVP produced together with Vasopressin? Do both analytes show the same kinetics?  Which CT-proAVP levels should be expected in normals?  Thermo Scientific B·R·A·H·M·S CT-proAVP LIA in the Differential Diagnosis of Diabetes insipidus  What about the performance of the Thermo Scientific B·R·A·H·M·S CT-proAVP LIA Assay?

3 3 What is Vasopressin and where is it produced?

4 4 Structure of Vasopressin O NH 2 NH 2 - O NH 2 -C Arginine-Vasopressin (AVP)  synonym: Vasopressin or antidiuretic hormone (ADH)  peptide hormone  9 amino acids  Disulfide bridge between two cysteine amino acids  C-terminal amidation

5 5 Synthesis of Vasopressin Figures adapted from: Golenhofen, Basislehrbuch Physiologie, Urban & Fischer; and Morgenthaler NG et al.: Clin Chem 2006 Information: Russel IC and Glover PJ: Critical Care and Resuscitation 2002; Ranger GS: IJCP 2002; Oghlakian G and Klapholz M: Cardiology in Review 2009  Synthesis as a precursor hormone (pre-pro-vasopressin) in the hypothalamus  Cleavage and transport in granules down the axons  Storage in granules in the posterior pituitary  Release into nearby capillaries upon appropriate stimulation

6 6 What is the physiological role of Vasopressin?

7 7 Vasopressin - physiological role AVP: acts via V 2 -receptors in the kidney -> water retention Main role: Regulation of water balance Figure adapted from: Knoers NV N Engl J Med. 2005 May 5;352(18):1847-50 - Increased plasma osmolality - Decreased arterial circulating volume AVP: Synthesis in the Hypothalamus

8 8 receptorlocationeffect V2kidneywater retention V1 a vascular smooth muscle cells strong vasoconstriction V1 b endocrine cells (e.g. pituitary) regulation of ACTH secretion during stress Vasopressin (AVP) effects Effects of AVP dependent on concentration :  maximal antidiuretic effect: below 15 pg/ml  vasoconstrictor effect at higher concentrations  very little effect on blood pressure at physiological levels! Singh Ranger G, Int J Clin Pract 2002; 56(10):777-782

9 9 Vasopressin in stress situation ACTH AVP STRESS Cortisol Myocardial infarction

10 10 Why not simply measure Vasopressin?

11 11 Quantification of Vasopressin is difficult Vasopressin

12 12 Quantification of Vasopressin is difficult Vasopressin Receptor

13 13 Quantification of Vasopressin is difficult Vasopressin Platelets Receptor

14 14 Quantification of Vasopressin is difficult Vasopressin Platelets Protease Receptor

15 15 Quantification of Vasopressin is difficult Vasopressin Platelets Protease Receptor Further problem: very unstable ex vivo (even frozen)

16 16 Quantification of Vasopressin is difficult Vasopressin Platelets Protease Receptor Only specialized labs measure AVP (time to results several days) Not a single FDA approved AVP assay on the market LIMITED CLINICAL USE Further problem: very unstable ex vivo (even frozen)

17 17 Morgenthaler NG et al., Clin Chem. 2006 Prohormone processing and assay SignalVasopressin Neurophysin II CT-proAVP

18 18 Morgenthaler NG et al., Clin Chem. 2006 Prohormone processing and assay SignalVasopressin Neurophysin II CT-proAVP Signal Peptidase Vasopressin Neurophysin II CT-proAVP

19 19 Morgenthaler NG et al., Clin Chem. 2006 Prohormone processing and assay SignalVasopressin Neurophysin II CT-proAVP Signal Peptidase Vasopressin Neurophysin II CT-proAVP Vasopressin Prohormone Convertase CT-proAVP Neurophysin II

20 20 Morgenthaler NG et al., Clin Chem. 2006 Prohormone processing and assay SignalVasopressin Neurophysin II CT-proAVP Signal Peptidase Vasopressin Neurophysin II CT-proAVP Vasopressin Prohormone Convertase CT-proAVP Neurophysin II

21 21 Morgenthaler NG et al., Clin Chem. 2006 Prohormone processing and assay SignalVasopressin Neurophysin II CT-proAVP Signal Peptidase Vasopressin Neurophysin II CT-proAVP Vasopressin Prohormone Convertase CT-proAVP Neurophysin II CT-proAVP very stable ex vivo

22 22 Morgenthaler NG et al., Clin Chem. 2006 Prohormone processing and assay SignalVasopressin Neurophysin II CT-proAVP Signal Peptidase Vasopressin Neurophysin II CT-proAVP Vasopressin Prohormone Convertase CT-proAVP Neurophysin II CT-proAVP very stable ex vivo

23 23 Is CT-proAVP produced together with Vasopressin? Do both analytes show the same kinetics in vivo?

24 24 r = 0.78 LIA Assay Morgenthaler NG et al., Clin Chem. 2006 Jan;52(1):112-9. Jochberger S et al., Schock 2009 31: 132-138 Validation in: Jochberger S et al., Intensive Care Med 2009 35:489-497 Correlation of Vasopressin and CT-proAVP

25 25 t 1/2 : few minutes CT-proAVP – like Vasopressin – is rapidly degraded in vivo Morgenthaler et al. Clin Chem 2006

26 26 t 1/2 : few minutes CT-proAVP – like Vasopressin – is rapidly degraded in vivo Morgenthaler et al. Clin Chem 2006

27 27 97.5 % percentile normals: t 1/2 : few minutes CT-proAVP – like Vasopressin – is rapidly degraded in vivo Morgenthaler et al. Clin Chem 2006

28 28 Control Hypotonic saline infusion Hypertonic saline infusion / thirsting n=8 CT-proAVP – Stimulation via osmoreceptors CT-proAVP behaves like AVP Szinnai et al. JCEM (2007)

29 29 Balanescu S. et.al. JCEM 2011 in press CT-proAVP correlates better with osmolality than Vasopressin

30 30 CT-proAVP- stimulation via baroreceptors/ hemorrhagic shock, model CT-proAVP behaves like AVP Morgenthaler et al. Shock 2007

31 31 Which CT-proAVP levels should be expected in normals?

32 32 Morgenthaler NG et al., Clin Chem. 2006 Jan;52(1):112-9 Normal distribution CT-proAVP is not age-related

33 33 Bhandari SS et al, Clinical Science (2009) 116, 257–263 706 healthy volunteers Significantly higher levels in males CT-proAVP levels dependent on gender

34 34 Morgenthaler NG et al., Clin Chem. 2006 Jan;52(1):112-9 CT-proAVP: Influence of exercise

35 35 Morgenthaler NG et al., Clin Chem. 2006 Jan;52(1):112-9 CT-proAVP: Influence of exercise 97.5 % percentile normals:

36 36 CT-proAVP LIA in the differential diagnosis of Diabetes insipidus

37 37 What is Diabetes insipidus ? Diabetes Insipidus (DI) is a disorder in which there is an abnormal increase in urine output, fluid intake and often thirst (polyuria-polydipsia-syndrome). Urine output is increased because it is not concentrated normally -> the urine is not yellow but pale, colorless or watery. Diabetes Insipidus is divided into three types, each of which has a different cause and must be treated differently.

38 38 Types of Diabetes insipidus Central Diabetes Insipidus (also known as neurogenic DI): The most common type of DI is caused by a lack of vasopressin. Treatment: various drugs including a modified vasopressin known as desmopressin or DDAVP Nephrogenic Diabetes insipidus (also known as renal DI): is caused by an inability of the kidneys to respond to the "antidiuretic effect" of normal amounts of vasopressin. Treatment: It cannot be treated with DDAVP and, depending on the cause, may or may not be curable by eliminating the offending drug or disease.

39 39 Types of Diabetes insipidus Central Diabetes Insipidus (also known as neurogenic DI): The most common type of DI is caused by a lack of vasopressin. Treatment: various drugs including a modified vasopressin known as desmopressin or DDAVP Nephrogenic Diabetes insipidus (also known as renal DI): is caused by an inability of the kidneys to respond to the "antidiuretic effect" of normal amounts of vasopressin. Treatment: It cannot be treated with DDAVP and, depending on the cause, may or may not be curable by eliminating the offending drug or disease. Diagnostic Challenge: All types of Diabetes insipidus also as partial forms existing!

40 40 Types of Diabetes insipidus (II) primary polydipsia : occurs when vasopressin is suppressed by excessive intake of fluids. most common type of polyuria-polydipsia-syndrome most often caused by an abnormality in the part of the brain that regulates thirst or by psychogenic illnesses (psychogenic polydipsia) difficult to differentiate from central DI because it mimics DI. Interested in more?: http://www.diabetesinsipidus.org http://www.diabetesinsipidus.org Also in French and Spanish language

41 41  Clinical Challenges: Differential diagnosis of patients with polyuria-polydipsia syndrome  State-of-the art diagnosis: 1. Stimulation of AVP release via a Water deprivation test 2. Indirect measurement of AVP release by monitoring of urine osmolality and - volume during water deprivation (ability to concentrate urine). 3. Additional Desmopressin administration to differentiate nephrogenic DI from central DI.  Direct AVP measurement becomes not the diagnostic reference standard because of its methological limitations (instability of analyte and uncomfortable assay handling) Differential Diagnosis of Diabetes insipidus

42 42 CT-proAVP for Differential diagnosis of Diabetes insipidus central DI primary Polidipsia Nephrogenic DI Urine Volume/ fluid intake Excessive Urine- Osmolalitylow CT-proAVP basallow (< 2.6 pmol/l) low (~3 pmol/l) high (>20 pmol/l) CT-proAVP increase after thirsting noyessmall State-of-the-art diagnosis ability to concentrate urine during water deprivation, indirect measurement via urine- volume and – osmolality ability to respond to desmopressin intake

43 43 Differential diagnosis of Diabetes insipidus central DI primary Polidipsia Nephrogenic DI Urine Volume/ fluid intake Excessive Urine- Osmolalitylow CT-proAVP basallow (< 2.6 pmol/l) low (~3 pmol/l) high (>20 pmol/l) CT-proAVP increase after thirsting noyessmall State-of-the-art diagnosis Diagnosis without water deprivation and Desmopressin stimulation possible! ability to concentrate urine during water deprivation, indirect measurement via urine- volume and - osmolality

44 44 Differential diagnosis of Diabetes insipidus central DI primary Polidipsia Nephrogenic DI Urine Volume/ fluid intake Excessive Urine- Osmolalitylow CT-proAVP basallow (< 2.6 pmol/l) low (~3 pmol/l) high (>20 pmol/l) CT-proAVP increase after thirsting noyessmall State-of-the-art diagnosis Diagnosis without water deprivation possible! Differential diagnosis of partial DI possible ability to concentrate urine during water deprivation, indirect measurement via urine- volume and - osmolality

45 45 CT-proAVP course during water deprivation Mean value of CT-proAVP in primary polydipsia Mean value of CT-proAVP in central DI

46 46 Superiority of CT-proAVP in diagnosing Diabetes insipidus Conclusion: Current state-of -the art - method WDT gives no reliable results in the differential diagnosis of polyuria-polydipsia syndrome! CT-proAVP is superior to the current method of choice and revives the concept of the direct test in the polyuria- polydipsia syndrome. Fenske W. et.al. Copeptin in the differential diagnosis of the polyuria- polydipsia syndrome – revisiting the direct and indirect water deprivation tests. JCEM accepted January 2011

47 47 CT-proAVP: Diagnosis of central DI totalis and nephrogenic DI in the 1 st blood draw Fenske W. et.al. Copeptin in the differential diagnosis of the polyuria- polydipsia syndrome – revisiting the direct and indirect water deprivation tests. JCEM accepted January 2011 basal CT-proAVP [pmol/l] (fasting, in the morning after 8h dehydration) 20 Sensitivity (%)95100 Specificity (%)100100 Central Diabetes nephrogenic insipidus totalis Diabetes insipidus

48 48 Best separation of primary polydipsia and partial central DI (in contrast to current methods including AVP measurements) specificity 100% sensitivity 86% Poster: Fenske W: 14 th Annual meeting of the neuroendocrinology section of the DGE October 15, 2010 (Munich) Paper accepted at JCEM Jan. 2011 Differential diagnosis of unclear cases after water deprivation erum -Na +

49 49 Index Δ CT-proAVP [8h-16h] x 1000 [pmol/L/mmol/L] S-Na+ [16h] 20 Sensitivity (%)100 86 Specificity (%) 86100 central Diabetes primary insipidus partialis polydipsia 2 nd blood draw: Stimulated CT-proAVP differentiates safe between central DI partialis and Primary Polydipsia Fenske W. et.al. Copeptin in the differential diagnosis of the polyuria- polydipsia syndrome – revisiting the direct and indirect water deprivation tests. JCEM accepted January 2011

50 50 Reliable differential diagnosis of DI with the help of CT-proAVP Suspicion of Diabetes insipidus with Polyurie-Polydipsie-Syndrome CT-proAVP basal (in the morning, fasting, after 8h dehydration) CT-proAVP <2,6 pmol/L CT-proAVP >20 pmol/L Central Diabetes insipidus totalis Renal Diabetes insipidus CT-proAVP >=2,6 - 20 pmol/L CT-proAVP-Index <20 CT-proAVP-Index >=20 Central Diabetes insipidus partalis Primary Polydipsia Ratio of CT-proAVP-Delta (8 to16h) and Serum-Na + (16h) = CT-proAVP-Index CT-proAVP stimulated and Serum-Na + (after 16 hours dehydration)

51 51 Advantages for the diagnostic routine Significantly higher diagnostic accuracy for all variations of Diabetes insipidus and primary Polydipsia Considerably eased differential diagnosis of Polyuria- Polydipsia-Syndrome Reduced physical and psychical exposure of the patient due to simplified WDT and redundancy of desmopressin stimulation Support of safe therapeutic decisions with highly sensitive measurement values Overall cost reduction due to reduced complexity, less lab consulting and no prescription of desmopressin

52 52 What about the performance of the LIA assay?

53 53 Reminder: Why not measure AVP directly? AVP is very unstable in plasma even at -20 °C storage (sample transport frozen or blood collection directly in the lab) AVP is largely attached to platelets AVP assays performed with the required accuracy are available in only a few selected laboratories (non of them FDA cleared) Sample extraction necessary Time to result up to 72 hours Sample volume 1-4 ml plasma No reliable clinical results

54 54 Advantages Thermo Scientific B·R·A·H·M·S CT-proAVP LIA sample volume only 50µl for plasma and serum one-step procedure (time to result 3 hours) stable analyte (at room temperature) highest sensitivity sandwich-immunoassay clinical superiority shown

55 55 CT-proAVP LIA assay parameters Data taken from IFU (instructions for use)


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