HK1197933B - Diagnosis and risk stratification by means of the novel marker ct-proadm - Google Patents

Description

Diagnosis and risk stratification using the novel marker CT-proADM

The application is a divisional application of an invention patent application with the application date of 2007, 20/12 and the application number of 200780047509.X and the invention name of 'diagnosis and risk stratification by using a novel marker CT-proADM'.

The invention relates to a novel diagnostic marker CT-proADM (C-terminal fragment of preproADM, SEQ ID No. 1) for diagnosing and/or stratifying the risk of diseases. The invention also relates to methods for diagnosing and/or risk stratification for diseases, in particular cardiovascular diseases, cardiac insufficiency, infections and/or inflammations of the lungs and respiratory tract. In the method, the CT-proADM (SEQ ID No. 1) marker, or a peptide fragment or fragment thereof, or the marker comprised in a marker combination (marker panel, marker cluster) is determined in a patient to be tested. The invention also relates to a diagnostic device and a kit for carrying out said method.

The prior art describes how proadrenomedullin (proADM) and adrenomedullin (EP0622458B1, Lewis LK, Smith MW, Yandle TG, Richards AM, Nichols MG. Adrenomedullin (1-52) measured in human serum by radioimmunoassay: plasma center, adsorption, and range. Clin Chem1998;44:571-7; Ueda S, Nishio K, Minamin N, Kubo A, Akai Y, Kaawa K et al, incorporated plasma levels of human serum in tissue with system in tissue system, and vice versa; 9980. J.E.K.E. K.E. K.E.A. J.R. CrCare 1999; 99160: 132-6; K.E.K.E.K.E.K.E.K.E.K.E.K. K. K.E.E. K. Biochemical company, K.K.K.E.E.E.K. K. 1996, K.E.E.E.E.E.E.E.E.E.E.E.E.E.A. K. A. K. A. may be used in diagnosis, et 4. incorporated by human animal tissue culture system in diagnosis, et 4. A. may be determined in diagnosis, and the present in diagnosis, and the invention may be a diagnosis of human animal tissue culture system may be used in the invention. In particular for the purpose of sepsis diagnosis (EPl121600B 1). N-terminal fragments of (pro) pro-adrenomedullin for use in diagnostics are also described in EP0622458B1, e.g.PAMP (Hashidas, Kitamura K, Nagatomo Y, Shibata Y, Imamura T, Yamada K et al, Development of an ultra-sensitive enzyme assay for human adadenoduct N-terminal20peptide and direct assay of two molecular assays of PAMP in plant from chemical subjects and tissues with cardiac disease Biotin 2004;37: 14-21).

Furthermore, other fragments of proadrenomedullin for diagnostic purposes, so-called intermediate Adrenomedullin segment (MR-proADM) (Structure J, Tao C, Morgethaler NG, bergmann A. identification of an Adrenomedulalin preceding fragment in a plant of diseases. Peptides2004;25:1369-72; Morgethaler NG, Structure J, Alonso C, bergmann A. measurement of central acquired nuclear in a plant with immune nuclear assay in Clin M2005; 51:1823-9; Christ-Craiton M, Morgethalerin NG, Muller C, Biisserver R, Harmoni R et al, Harmoni R2005, stress transporter binding J. promoter, Morgetalr binding protein J. promoter III, Morgetalr C, Morgetalr D, Muller C, Morgethalerin R, Harmoni et al, Morgetrien. promoter J. Pro, Morgetable promoter, protein, hormone, protein.

However, there is a great need for a reliable diagnosis or (risk) stratification of diseases, in particular cardiovascular diseases, cardiac insufficiency, infections and/or inflammations of the lungs and respiratory tract, especially for further clinical decisions, and especially for the severity of the disease, in particular cardiovascular diseases, cardiac insufficiency, infections and/or inflammations of the lungs and respiratory tract.

It is an object of the present invention to provide novel markers.

It is a further object of the present invention to provide improved methods for diagnosis and/or risk stratification for diseases, in particular cardiovascular diseases, cardiac insufficiency, infections and/or inflammations of the lung and respiratory tract.

The above object is solved on the one hand by providing the diagnostic marker CT-proADM (C-terminal fragment of Pro-adrenomedullin, SEQ ID No. 1), or a peptide or fragment thereof, and on the other hand by a method for in vitro diagnosis and/or risk stratification of a disease. In the method, the CT-proADM (SEQ ID No. 1) marker, or a peptide fragment or fragment thereof, or the marker comprised in a marker combination (marker panel, marker cluster) is determined in a patient to be tested (hereinafter referred to as the method according to the invention).

According to the present invention, the term "risk stratification" comprises the discovery of diseased patients with a poorer prognosis in order to carry out a more intensive diagnosis and (subsequent) treatment/management of the disease with the aim of obtaining a disease course as favourable as possible.

It is therefore particularly advantageous that reliable diagnosis and/or risk stratification can be carried out by means of the method according to the invention. The method according to the invention enables clinical decisions to be made which lead to a faster diagnosis. Such clinical decisions also include further treatment by application of drugs for treatment or therapy of diseases.

The invention therefore also relates to a method for risk stratification of a patient, in particular for risk stratification of a patient for clinical decision-making, in particular in time-critical medicine or emergency medicine, and for admission of a patient.

In a further preferred embodiment of the method according to the invention, diagnosis and/or risk stratification is performed for prognosis, for differential diagnostic early diagnosis and detection, for severity assessment, and for disease progression assessment of concomitant therapy.

In another embodiment of the method according to the invention, a body fluid or body tissue, preferably blood, optionally whole blood, serum or available plasma, is extracted from the patient to be tested and the diagnosis is performed ex vivo, i.e. outside the human or animal body. As a result of the determination of the marker CT-proADM (SEQ ID No. 1), or a peptide or fragment thereof, and the amount thereof present in at least one patient sample, a diagnosis and/or risk stratification can be carried out.

Within the scope of the present invention, "diseases" are understood as diseases of humans or animals, in particular mammals. These diseases, especially human diseases, are described in Pschyrembel, De Gruyter, Berlin 2004.

However, it is particularly advantageous within the scope of the present invention to diagnose and/or stratify cardiovascular diseases, cardiac insufficiency, infections and/or inflammations of the lungs and respiratory tract.

The term "cardiovascular disease" according to the present invention includes all diseases of the heart and blood circulation, especially such indications as hypertension, coronary heart disease, in particular acute coronary syndrome, (acute) myocardial infarction, angina pectoris.

The term "acute coronary syndrome" includes the different stages of coronary heart disease that are directly life threatening. In particular, it relates to emergency medicine, i.e. acute myocardial infarction and/or angina pectoris and sudden cardiac death. In addition to acute myocardial infarction, which according to the WHO standard (WHO (1979) is defined as an acute chest pain event lasting more than 20 minutes with ST elevation and/or elevation of the myocardial enzymes, according to the general and criterion of diagnosis, report of the Joint International Society and Federation, Circulation59(3):607-609), which is defined as an acute chest pain event, which is accompanied by ST elevation and/or elevation of the myocardial enzymes, the term includes unstable Angina (AP), which according to the invention can be included in the term "acute coronary syndrome" (Hamm: Leitinen: Aktuses Koronronronronronronronronronronronrondrom (ACS) -Te 1: ACS patent medicine property ST-Health. Z (K93: 72).

Within the scope of the present invention, "cardiac insufficiency" is understood to mean the acute or chronic inability of the heart to supply sufficient blood and thus to provide sufficient oxygen to the tissues in order to ensure their metabolism at rest or under load (stress). Clinically, cardiac insufficiency occurs in the presence of typical symptoms (dyspnea, fatigue, fluid retention) due to cardiac malfunction in the sense of systolic or diastolic malfunction. Chronic cardiac insufficiency (CHF) is also included in the present invention (kardiologitecompact, published by christian means, Reimer Riessen und Ioakim spinodonos, 2. unrefined, Thieme 2006). The reasons for cardiac insufficiency may be: heart valve defects (e.g., long-term consequences of rheumatic fever), myocarditis (myocardial inflammation), cardiac arrhythmias, heart attacks in addition to hypertension (hypertension) and/or additional coronary arteriosclerosis (calcification) (coronary heart disease). The invention also includes the following: hypertensive heart disease with (congestive) cardiac insufficiency, hypertensive heart disease and kidney disease with (congestive) cardiac insufficiency, primary right ventricular insufficiency, secondary right ventricular insufficiency, unadapted left ventricular insufficiency (NYHA (new york heart association) class I), left ventricular insufficiency with discomfort under increased load (NYHA class II), left ventricular insufficiency with discomfort under lighter load (NYHA class III), left ventricular insufficiency with discomfort under resting state (NYHA class IV) and cardiogenic shock.

Within the scope of the present invention, the term "lung and respiratory tract infections" refers in particular to such infections caused by bacteria, viruses, fungi or parasites, for example indications such as deep respiratory tract infections (LRTI: lower respiratory tract infections), bronchitis, pneumonia, sarcoidosis, bronchiectasis, non-cardiogenic pulmonary edema.

Furthermore, according to the present invention, deep respiratory tract infection (LRTI: lower respiratory tract infection), bronchitis, septic bronchitis, and pneumonia are preferable. Pneumonia, particularly community-acquired pneumonia (CAP: community-associated pneumonia), and deep respiratory tract infection (LRTI: lower respiratory tract infection) are particularly preferred.

Pneumonia is understood within the scope of the present invention as acute or chronic inflammation of the lung tissue. The infection is caused by bacteria, viruses or fungi, in few cases by inhalation of toxic substances or by immunization. For clinicians, pneumonia constitutes a diverse group of symptoms (fever or hypothermia, shivering, cough, pleurisy chest pain, increased sputum production, increased respiratory rate, voiced soundsBronchial respiratory sounds, high frequency aristo sounds (ohrnahe)) Pleural frication), in combination with at least one chest X-ray visible infiltration (harrisons lnnere Medizin, herausgeben von Manfred diesel, Norbert sutttorp und MartinZeitz, ABW wissenschafts verlag 2005).

Within the scope of the present invention, the term "infectious diseases of the lung and respiratory tract" or "inflammatory diseases of the lung and respiratory tract" refers to the following indications: for example, interstitial lung diseases and pulmonary fibrosis, Chronic Obstructive Pulmonary Disease (COPD), in particular exacerbation of COPD infection, bronchial asthma, in particular exacerbation of bronchial asthma infection, bronchial cancer.

According to the present invention, COPD refers to a group of chronic diseases characterized by cough, profuse sputum and dyspnea under load. First, chronic obstructive bronchitis and emphysema are mentioned. The clinical manifestations of both diseases are characterized inter alia by respiratory obstruction. Moreover, the main symptom of COPD is colloquially referred to as "cough in smokers". The invention is particularly advantageous in acute exacerbation situations.

Within the scope of the present invention, "CT-proADM" is understood as an free human protein or polypeptide consisting of 33 amino acids having the amino acid sequence SEQ ID No. 1: SLPEAGPGRTLVSSKPQAHGAPAPPSGSAPHFL, respectively; or a fragment having the amino acid sequence 153-185 (Ser at position 153 and Leu at position 185) of proadrenomedullin SEQ ID No.2 (FIG. 1) (Kitamura K, Sakata J, Kangawa K, Kojima M, Matsuo H, Eto T. cloning and characterization of cDNAencoding adapter for human adorenomedulin Biochem Biophys Res Commun1993;194: 720-725); or a peptide fragment thereof and/or a fragment thereof. Such fragments may be, for example, the amino acid sequences 1 to 15 of SEQ ID No.1 and/or 153 to 167 of proadrenomedullin (FIG. 1) or 19 to 33 of SEQ ID No.1 and/or 171 to 185 of proadrenomedullin (FIG. 1) (see examples). CT-proADM is also known as adrenotensin (adrenetenin) due to its vasoconstrictive effect (Gumusel B, Chang JK, Hyman A, Lippton H.adrenetenin: an ADM genetic product with the disposal effects of ADM.Life Sci1995;57: PL 87-90).

Furthermore, "CT-proADM" according to the invention may have modifications, such as glycolation or glycosylation, lipidation or derivatization.

In another embodiment, "CT-proADM" (SEQ ID No. 1) can additionally also be determined together with other markers, wherein "CT-proADM" is comprised in a marker combination (marker panel, marker cluster), i.e.preferably such a marker combination which already indicates a disease. However, such markers are preferred, which in turn indicate preferred indications/diseases within the scope of the present invention, and may bring about synergistic effects.

Thus, the present invention relates to such embodiments of the method according to the invention, wherein the test patient is determined additionally together with at least one further marker selected from the group consisting of: an inflammation marker, a cardiovascular marker, a neurohormonal marker, or an ischemia marker.

according to the invention, the inflammatory marker can be selected from at least one of the markers C-reactive protein (CRP), cytokines such as, for example, TNF- α, interleukins such as, for example, IL-6, procalcitonin (1-116,3-116), and adhesion molecules such as VCAM (vascular cell adhesion molecule) or ICAM (intercellular adhesion molecule), and the cardiovascular marker is selected from at least one of the markers creatine kinase, myeloperoxidase, and peptin, myoglobin, natriuretic proteins, in particular ANP (or ANF), proANP, NT-proANP, BNP, proBNP, NT-proBNP or in each case partial sequences thereof, cardiac troponin, CRP.

The ischemic marker may be selected from at least one of the following: troponin I and T, CK-MB. Furthermore, the neurohormonal marker can be at least one natriuretic protein, in particular ANP (or ANF), proANP, NT-proANP, BNP, proBNP, NT-proBNP or in each case a partial sequence thereof.

Particularly preferred is the combination of CT-proADM with a prohormone, in particular proBNP, NT-proBNP marker.

In another embodiment of the invention, the method according to the invention can be carried out by means of parallel or simultaneous determination of markers (e.g. a 96-well or more multi-well titer plate), wherein the determination is performed on at least one patient sample.

Furthermore, the method according to the invention and its determination can also be carried out by means of an automated analytical device, in particular Kryptor (http:// www.kryptor.net /).

In another embodiment, the method according to the invention and its determination can be performed by means of a rapid Test, such as a lateral-flow Test, whether a single-parameter or a multi-parameter determination.

The invention furthermore relates to the use of CT-proADM (SEQ ID No. 1) or a peptide fragment or fragment thereof for the in vitro diagnosis and/or risk stratification of diseases, in particular cardiovascular circulatory diseases, cardiac insufficiency, infections and/or inflammations of the lung and respiratory tract.

In another embodiment, the invention relates to the use of CT-proADM (SEQ ID No. 1) or a peptide fragment or fragment thereof for cardiac diagnostics.

The invention furthermore relates to the use of CT-proADM (SEQ ID No. 1) or peptide fragments or fragments thereof, or comprised in a marker combination (marker panel, marker cluster), for the in vitro diagnosis and/or risk stratification of diseases, in particular cardiovascular circulatory diseases, cardiac insufficiency, infections and/or inflammations of the lung and respiratory tract, and in particular in view of the above embodiments. The marker combination may optionally comprise additional suitable markers.

Another object is to provide a suitable diagnostic device or to apply said device for carrying out the method according to the invention.

Within the scope of the present invention, such a diagnostic device is understood to mean in particular an array or an analytical instrument (e.g. immunoassay, ELISA, etc.), which in a broad sense is a device for carrying out the method according to the invention.

The invention also relates to a kit or the use of such a kit for the in vitro diagnosis and/or risk stratification for diseases, in particular cardiovascular diseases, cardiac insufficiency, infections and/or inflammations of the lungs and respiratory tract, in which method, in particular taking into account the above-described embodiments, the CT-proADM (SEQ ID No. 1) marker, or a peptide fragment or fragment thereof, or the marker comprised in a marker combination (marker panel, marker cluster) is determined for a patient to be tested. Such detection reagents include, for example, antibodies and the like.

The invention therefore relates to antibodies, in particular monoclonal antibodies, for the detection of CT-proADM, which preferably bind to CT-proADM individually, independently of one another, in each case at the binding sites 1-15 or 19-33 of SEQ ID No. 1; or to bind the CT-proADM at one of these two binding sites or at both binding sites.

The following examples and drawings are intended to explain the invention in more detail, without limiting the invention to these examples and drawings.

Drawings

FIG. 1 shows the amino acid sequence of prepro-adrenomedullin (preproADM).

FIG. 2 shows that CT-proADM concentration is related to the severity of cardiac insufficiency.

Figure 3 shows that CT-proADM concentration increases with PSI.

Example (b):

immunoassay

Peptide synthesis

Two regions (positions 153-167, 171-185) were selected from the known amino acid sequence of preproADM. The regions, each supplemented with an N-terminal cysteine residue, were chemically synthesized, purified, mass detected by means of mass spectrometry and reverse phase HPLC, and lyophilized as soluble peptide in aliquots according to standard methods (Firma JPT, Berlin, Deutschland). The amino acid sequence of the peptide is as follows:

PSK16CSLPEAGPGRTLVSSK position 153-167

PHL16 CHGAPAPPPSGSAPHFL positions 171-185

Furthermore, peptides were synthesized comprising regiopositions 153-185 of preproADM (PSL 33 SLPEAGPGRTLVSSKPQAAGPPSGSAPHFL).

Coupling and immunization

Polypeptides PSK16 and PHL16 were coupled to the carrier protein KLH (keyhole limpet hemocyanin) by means of MBS (m-maleimidobenzoyl-N-hydroxysuccinimide ester) (see the instruction manual "NHS Esters-MaleimideProsslinkers (NHS ester-MaleimideProssoinker" PIERCE, Rockford, IL, USA.) sheep were immunized using these conjugates by initially receiving 100 μ g of conjugate per sheep (mass data based on the peptide part of the conjugate) and subsequently receiving 50 μ g of conjugate every four weeks (mass data based on the peptide part of the conjugate). starting four months after the start of immunization, 700ml of blood was collected from each sheep every 4 weeks and antiserum was obtained by centrifugation.

Purification of antibodies

In the one-step method, peptide-specific antibodies are prepared from antisera that have been extracted from four months after immunization.

For this, the peptides PSK16 and PHL16 were first coupled to SulfoLink gel (see the instructions "SulfoLink kit", PIERCE company, Rockford, IL, USA). In this method, coupling is performed by providing 5ml of gel per 5mg of peptide.

Affinity purification of peptide-specific antibodies from sheep antiserum against peptides was performed as follows:

the peptide column was first washed three times in alternation, using 10ml each of elution buffer (50 mM citric acid, pH 2.2) and binding buffer (100mM sodium phosphate, 0.1% Tween, pH 6.8). 100ml of antiserum was filtered through 0.2 μm and mixed with the existing column material. For this purpose, the gel was washed quantitatively from the column using 10ml of binding buffer. Incubations were performed at room temperature with shaking overnight. The sediment was quantitatively transferred to an empty column (NAP25, Pharmacia, empty). The passage was discarded. Then washed with 250ml binding buffer to remove proteins (eluent protein content <0.02a280 nm). Elution buffer was added to the washed column and fractions of one ml each were collected. The protein content of each fraction was determined by the BCA-method (see the instructions, PIERCE, Rockford, IL, USA). Fractions with protein concentration >0.8mg/ml were pooled. The pools were subjected to protein assay using the BCA method, resulting in a yield of 34mg of anti-PSK 16 antibody and 48mg of anti-PHL 16 antibody.

Marking

Mu.l of the purified anti-PSK 16 antibody (see above) was buffered thoroughly in 1ml of 100mM potassium phosphate buffer (pH 8.0) via NAP-5 gel filtration column (Pharmacia) according to the protocol. The protein concentration of the antibody solution was adjusted to 1.5mg/ml with 100mM potassium phosphate buffer (pH 8.0).

For chemiluminescent labeling, the antibodies were further processed as follows: with 10. mu.l of MA 70-acridine67 μ l of the antibody solution was treated with-NHS-ester (1 mg/ml; Behring, HOECHST) and incubated for 15 minutes at room temperature. Then 423. mu.l of 1M glycine were added and incubated for a further 10 minutes. Next, the labeled sediment was sufficiently buffered in 1ml of eluent A (50 mM potassium phosphate, 100mM sodium chloride, pH7.4) by means of a NAP-5 gel filtration column (Pharmacia) according to the instructions of the procedure, and in the process, the low molecular weight components were removed. To isolate the final labeled residue not attached to the antibody, gel filtration HPLC (column: Waters Protein Pak SW 300) was performed. The test was performed and chromatography was performed with eluent A at a flow rate of 1 ml/min. The wavelengths at 368nm and 280nm were measured using a flow rate photometer. The absorbance ratio at 368nm/280nm, as a measure of the degree of antibody labeling, peaked at 0.10 +/-0.01. Fractions containing monomeric antibody (retention time 8-10 minutes) were collected and collected in 3ml of 100mM sodium phosphate, 150mM sodium chloride, 5% bovine serum albumin, 0.1% sodium azide in a liquid at pH 7.4.

Coupling of

Irradiated 5ml small polystyrene tubes (Greiner) were coated with purified anti-PHL 16 antibody as follows: the antibody was diluted to a concentration of 6.6. mu.g/ml in 50mM Tris, 100mM sodium chloride, pH 7.8. 300 μ l of the solution was inhaled into each vial. The vials were incubated at 22 degrees celsius for 20 hours. The solution was aspirated. Each vial was then filled with 4.2ml of 10mM sodium phosphate, 2% Karion FP, 0.3% bovine serum albumin, pH 6.5. After 20 hours the solution was aspirated. Finally the vials were dried in a vacuum desiccator.

Implementation and analysis of immunoassays

Peptide PSL33 was used as a standard material in serial dilutions in normal horse Serum (SIGMA). Concentrations according to the weight of the starting peptide were assigned to the standards prepared in this way.

The sandwich immunoassay was performed as follows: mu.l of standard or test substance and 200. mu.l of detection buffer (100mM sodium phosphate, 150mM sodium chloride, 5% bovine serum albumin, 0.1% nonspecific sheep IgG, 0.1% sodium azide, pH7.4) containing 100 million RLU (relative light units) of MA 70-labeled antibody were pipetted into small test tubes each coated with antibody. Incubation was performed at 22 ℃ for 2 hours with shaking. The individual vials were then washed 4 times each with 1ml of washing solution (0.1% Tween 20), dripped off and the chemiluminescence attached to the vial was measured in a luminometer (BERTHOLD, LB952T; Basisreagen BRAHMS AG). The CT-proADM concentration of the test substances was read off on a standard curve with the aid of MultiCalc software (Spline Fit).

The analyte that can be determined using this assay is called C-terminal proadrenomedullin (CT-proADM).

Clinical value

Normal range

The concentration of CT-proADM was determined in a test substance (n = 200) of healthy control persons. The median was 77.6pmol/L, the lowest measurement was 46.6, the highest was 136.2pmol/L, and the 95% percentile was 58.6 or 113.8pmol/L, respectively.

Cardiac insufficiency/severity

The concentration of CT-proADM is determined in patients suffering from chronic or acute decompensated cardiac insufficiency. CT-proADM concentrations are associated with the severity of cardiac insufficiency: the mean value of the CT-proADM concentrations in the 4 severity NYHA classes I-IV is: 85. 107, 140.4 or 242.7pmol/L (see FIG. 2).

Chronic cardiac insufficiency/diagnosis

CT-proADM values were determined for a group of 316 patients with chronic cardiac insufficiency and for a group of 200 healthy controls. receptor-Operator-Characteristics analysis (receptor-operators-Characteristics analysis) showed an AUC of 0.79. A threshold value (cut-off Wert) of 122pmol/ml gives a sensitivity of 39.7% and a specificity of 98%. At a threshold of 113pmol/L, a sensitivity of 46.3% and a specificity of 95% were obtained.

Chronic cardiac insufficiency/prediction (prognosis)

CT-proADM values were determined for a group of 316 patients with chronic cardiac insufficiency. Patients were observed over an average period of 360 days. During this period 42 patients died and 274 survived. The optimal threshold for predicting mortality (defined as the maximum product of sensitivity and specificity) was determined by recipient effect profile analysis: 119.7 pmol/L. At this threshold, the predicted sensitivity was 73.2% and the specificity was 62.2%. The death likelihood ratio at threshold 119.7pg/ml is: 1.9.

	<119.7pmol/L	>119.7pmol/L
			alive	172	103
Is dead	11	30

Acute cardiac insufficiency/diagnosis

CT-proADM values were determined for a group of 125 patients with acute dyspnea. Of these 125 patients 69 patients suffered from cardiac insufficiency. Receiver effect profile analysis for differential diagnosis of cardiac insufficiency showed AUC to be 0.75. At a threshold of 410pmol/L, a sensitivity of 12.4% and a specificity of 98% were obtained. At a threshold of 315pmol/L, a sensitivity of 18.3% and a specificity of 95% were obtained.

Acute cardiac insufficiency/prediction (prognosis)

CT-proADM values were determined for a group of 69 patients with acute decompensated cardiac insufficiency. The patient was observed over a period of 360 days. During this period, 21 patients died and 48 survived. The optimal threshold for predicting mortality (defined as the maximum product of sensitivity and specificity) was determined by recipient effect profile analysis: 192 pmol/L. At this threshold, the predicted sensitivity was 66.6% and specificity was 75%. The mortality likelihood ratio at the threshold 192pmol/L is: 2.5.

	<192pmol/L	>192pmol/L
			alive	36	12
Is dead	7	14

Myocardial infarction/prediction (prognosis)

Samples were obtained three days after the onset of the heart attack in 287 patients with an acute heart attack and CT-proADM was determined. The patient was observed over a period of 360 days. During this period, 220 patients did not experience adverse events, 67 were dead or were readmitted due to cardiac insufficiency. The optimal threshold (defined as the maximum product of sensitivity and specificity) for predicting mortality or readmission rate due to cardiac insufficiency was determined by recipient role profiling: 161.9 pmol/L. At this threshold, the predicted sensitivity was 67.2% and specificity was 79.1%. The adverse event likelihood ratio at threshold 161.9pmol/L is: 3.2.

	<161.9pmol/L	>161.9pmol/L
			no adverse events	174	46
Death/cardiac insufficiency	22	45

Pneumonia/severity and prediction (prognosis)

Samples were obtained at admission to 142 patients with community-acquired pneumonia and CT-proADM was determined. The patients were observed over a period of 70 days. 10 patients died during this period. CT-proADM concentrations increased with PSI (pneumonia severity index), a score for disease severity (fig. 3), and with a median of 135pmol/L relative to healthy controls (77 pmol/L). The AUC from the analysis of the receiver's effect profile used to predict mortality was 0.89. The optimal threshold for predicting mortality (defined as the maximum product of sensitivity and specificity) was determined by recipient effect profile analysis: 194.5 pmol/L. At this threshold, the predicted sensitivity was 100% and the specificity was 81.2%. The death likelihood ratio at threshold 194.5pmol/L is: 5.5.

	<194.5pmol/L	>194.5pmol/L
			no adverse events	108	24
Death was caused by death	0	10

Exacerbating COPD

CT-proADM was determined in 53 patients with chronic obstructive pulmonary disease with lower respiratory tract infection. CT-proADM of these patients was 106pmol/L median, higher than healthy controls (77 pmol/L), but lower than pneumonia patients (see above, 135 pmol/L).

Claims (13)

1. Use of an agent for the determination of a diagnostic marker consisting of CT-proADM for the preparation of a kit or a diagnostic device for a method for the diagnosis and/or risk stratification of a disease comprising the determination of CT-proADM in a patient to be tested, wherein the amino acid sequence of CT-proADM is shown in SEQ ID No.1, wherein the disease is selected from the group consisting of cardiac insufficiency, myocardial infarction, pneumonia and advanced chronic obstructive pulmonary disease.

2. Use according to claim 1, wherein the disease is an exacerbated chronic obstructive pulmonary disease.

3. The use according to claim 1, wherein the cardiac insufficiency comprises chronic cardiac insufficiency, acute cardiac insufficiency, hypertensive heart disease with congestive cardiac insufficiency and kidney disease, primary right ventricular insufficiency, secondary right ventricular insufficiency, unadapted left ventricular insufficiency, left ventricular insufficiency with discomfort under increased stress, left ventricular insufficiency with discomfort under lighter stress, left ventricular insufficiency with discomfort under resting state, cardiogenic shock.

4. Use according to any one of claims 1 to 3, characterized in that a parallel or simultaneous determination of the markers is performed.

5. Use according to any one of claims 1 to 3, wherein the assay is performed on at least one patient sample.

6. Use according to any one of claims 1 to 3, characterized in that the assay is carried out using an automated analytical device.

7. Use according to claim 6, characterized in that the assay is performed with the aid of Kryptor.

8. Use according to any one of claims 1 to 3, characterized in that the assay is carried out by means of a rapid test.

9. Use according to claim 8, wherein the assay is performed in a single or multi-parameter assay.

10. Use according to any of claims 1-3, characterized in that stratified diagnosis is performed on patients for clinical decision-making.

11. Use according to claim 10, characterized in that the stratified diagnosis of a patient is used for clinical decision-making for further treatment with the aid of drugs for the treatment or therapy of a disease.

12. Use according to claim 10, characterized in that stratified diagnoses are made on patients for clinical decisions in critical or emergency medicine.

13. Use according to claim 10, wherein the stratified diagnosis of a patient is used for clinical decision making of patient admission.