ADM-gly as marker for early predicting sepsis

Abstract

The present invention relates to diagnosis and monitoring the extent of sepsis and / or septic shock; early assessing the risk of getting sepsis and / or septic shock; predicting or determining or monitoring need of therapy or intervention of a subject. The invention relates to a method that comprises determining a level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT -proADM or fragments thereof of at least five amino acids or the total-ADM or a ratio of a level of said biomarker to a level of ADM-Nffzin the sample of the subject, and wherein the level of said biomarker or said ratio is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or is predictive for an enhanced risk of getting sepsis and / or septic shock, or is indicative for the need of therapy or intervention of sepsis and / or septic shock of said subject.

Description

[0001] ADM-Gly as marker for early predicting sepsis

[0002] The present invention relates to diagnosis and monitoring the extent of sepsis and / or septic shock; early assessing the risk of getting sepsis and / or septic shock; predicting or determining or monitoring need of therapy or intervention of a subject. The invention relates to a method that comprises determining a level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT -proADM or fragments thereof of at least five amino acids or the total-ADM or a ratio of a level of said biomarker to a level of ADM-NH2 in the sample of the subject, and wherein the level of said biomarker or said ratio is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or is predictive for an enhanced risk of getting sepsis and / or septic shock, or is indicative for the need of therapy or intervention of sepsis and / or septic shock of said subject.

[0003] Background of the invention

[0004] The incidence of sepsis has continued to increase in hospitalized patients1, comprising mortality rates in the range of 10% and 54%, depending on the level of disease severity, definition of organ dysfunction used2. An early and accurate assessment and prediction of the patient’s risk to develop sepsis or septic shock is therefore of crucial importance. Especially the prediction of sepsis in order to make prompt and reliable decisions concerning early treatment strategies and treatment monitoring is of high unmet medical need. Predicting patient prognosis may affect various aspects of patient management including treatment, diagnostic tests (e.g., microbiology, blood chemistry, radiology etc), and admission. Timely identification of patients with higher chance for poor prognosis may result in more efficient patient management procedures including for example, intensive care unit (ICU) admission, advanced therapeutics, invasive diagnostics or surgical intervention, which could reduce complications and mortality.

[0005] Adrenomedullin is one of the best studied peptide hormones. The Adrenomedullin (ADM) mRNA encodes a preprohormone of 185 amino acids (SEQ ID No. 1), the pre-pro-Adrenomedullin that is enzymatically converted into Proadrenomedullin by cleavage of the N-terminal signal peptide. Proadrenomedullin is then further process by several prohormone convertases to result in four peptides, namely

[0006] PAMP (SEQ ID No. 2): Proadrenomedullin amino-terminal peptide or Proadrenomedullin N- terminal 20 peptide3,4,5with a C-terminal glycine residue

[0007] MR-proADM (SEQ ID No. 6): Mid-regional Proadrenomedullin, a stable and inert peptide.

[0008] ADM-Gly (SEQ ID No. 4): C-terminally glycine extended, inactive precursor of biologically active ADM (bio-ADM (SEQ ID No. 5))

[0009] CT-proADM (SEQ ID No. 3): C-terminal Proadrenomedullun or Adrenotenisn CT-proADM: Little is known about the physiological role and behaviour in clinical settings of CT- proADM, sometimes referred to as Adrenotensin. It was shown to increase pulmonary arterial tension, is considered as a vasoconstriction and vasopressor peptide, that can induce proliferation of vascular smooth muscle cells6.

[0010] PAMP: In order to gain its full biological activity, the C-terminally glycine extended PAMP needs to be amidated by the enzyme peptidylglycine alpha-amidating monooxygenase (PAM). PAM recognizes the C-terminal glycine and catalyzes a sequential two-step reaction also referred to as amidation or C- terminal amidation. Thereby glyoxylate, originating from the c-terminal glycine, is released and an amidated carboxyl terminal arginine structure in PAMP is formed, which is essential for PAMP’s biological activity. Amidated or mature PAMP (m-PAMP; SEQ ID No 20) is a potent vasoactive and angiogenetic factor with actions complementing or antagonizing bio-ADM signalling. In rats, PAMP has a potent hypotensive effect causing vasodilation. Levels of PAMP-Gly and m-PAMP in healthy human subjects are 1.15 pmol / L and 0.51 pmol / L, respectively4,7 l(). In a rat-model of septic shock induced by LPS, plasma concentration of m-PAMP increased 30 minutes after LPS injection and was doubled after a 4 hours observation period when compared to baseline11. Even though originating from the same propeptide, levels of bio-ADM started to change after 2 hours in the same experiment. Further, plasma levels of PAMP may be increased in several clinical conditions, such as essential hypertension9chronic renal failure12, congestive heart failure13, and chronic glomerulonephritis14. When compared to bio-ADM, plasma levels of PAMP are relatively lower, indicating a differential role between bio-ADM and PAMP on pathophysiology in these disease conditions.

[0011] ADM-Gly, bio-ADM and MR-proADM: In general, ADM is upregulated by LPS and proinflammatory cytokines and plasma or serum levels of adrenomedullin are increased in sepsis and are associated with the severity of sepsis, mortality, and organ failure15. However, the term “Adrenomedullin” (ADM), as used in many publications is imprecise and in terms of this invention it is important to distinguish between the most prominent forms of ADM, which are MR-proADM and bio-ADM, but also ADM- Gly:

[0012] ADM-Gly is the direct, inactive16biosynthetic precursor of bio-ADM, often referred to as the intermediate form of ADM, and represents the dominating circulating form of Adrenomedullin in humans17. To gain its biological activity, ADM-Gly is activated by PAM17in the same manner as it was described for the activation of m-PAMP, resulting in the biologically active product bio-ADM, harbouring an amidated carboxyl terminal tyrosine structure.

[0013] Little is known about behaviour of ADM-Gly or it’s levels in clinical settings such as sepsis or septic shock. ADM-Gly was shown to be elevated in sepsis and septic shock before and had predictive value with respect to outcome and mortality in sepsis or shock (WO2021170752A1). The ratio of ADM-Gly / bio-ADM was reported to be predictive for mortality in septic patients (WO2021170763 Al). Using an ICTEIA (Immune Complex Transfer Enzyme Immunoassay) ADM-Gly was determined in plasma and tissue from a LPS model in rats18showing an ADM-Gly increase to 225.1 fmol / mL (1370 pg / mL) in comparison to 5.22 fmol / mL (32 pg / mL) in control animals, while bio-ADM increased to 107.8 fmol / mL (650 pg / mL)18, however, these measurements were performed 3 hours after LPS infusion and no time resolved ADM determinations were done directly after start of LPS infusion. The ADM-Gly / bio-ADM ratio (ADM ratio) was lower in the LPS animals (2.35) than in the control animals (10.81)18. Further, indirectly determining ADM-Gly it was shown that total plasma ADM (tADM), which is the sum of bio-ADM and ADM-Gly, correlates with the surgical stress score (SSS) as well as is elevated in several surgical groups (Cardiopulmonary bypass, abdominal, cervical laminoplasty) directly after surgery19. A comparison of tADM levels at day of ICU-admission and post ICU-admission at day 3 in patients with sepsis revealed elevated levels of tADM at admission as well as a prognostic value for tADM with respect to severity and outcome of sepsis, comparable to bio- ADM20. Further, ADM-Gly levels are elevated in pericardial fluid during ischemic heart disease21, in plasma of patients with essential hypertension and chronic renal failure, but ADM-Gly was reported to increase in parallel with bio-ADM in these indications22. In obese, an increase of plasma ADM-Gly was reported to be elevated in parallel with bio-ADM23.

[0014] The physiological function of ADM-Gly during sepsis remains unclear. It was reported that proinflammatory cytokines (i.a. AL- 1 , TNF and LPS) induce enhanced transcription of the ADM gene24, which would imply an early bio-ADM elevation during LPS infusion, as reported for m-PAMP11, that itself arises from PAMP-Gly. Surprisingly this is not the case for bio-ADM. As shown in example 2 of this invention, bio-ADM as the product of PAM mediated amidation of ADM-Gly, appears to have a delayed response to systemic inflammation for an unknown reason. Instead, an unexpectedly early ADM-Gly elevation takes place leading to an early elevation of the ADM-Gly / bio-ADM ratio.

[0015] Bio-ADM functions as a neurotransmitter and as a paracrine and autocrine signalling factor. As ADM- Gly, it belongs to the calcitonin gene-related peptide (CGRP) / calcitonin family of peptides hormone family25. Bio-ADM level in patients with congestive heart failure, myocardial infarction, kidney diseases, hypertensive disorders, diabetes mellitus, in the acute phase of shock and in sepsis and septic shock are significantly elevated, although to different extents4,26 31(WO2022234111A1). In sepsis and septic shock, bio-ADM levels are associated with severity of sepsis, organ failure and mortality. Due to impaired circulatory function and thereby impaired blood-flow in sepsis, oxygen supply to various organs is reduced causing hypoxia and multi organ failure. Elevation of bio-ADM is additionally induced by hypoxia, however, it is not known whether the same mechanisms can be applied to ADM- Gly. Bio-ADM reduces endothelial permeability leading to the reduction of hypoxia and extravasation of plasma components, such as leukocytes, from circulation. Furthermore, administration of bioactive ADM leads to reduced mortality in a rodent model of sepsis. It is widely accepted that the action of mortality reduction happens via reduction of endothelial permeability.

[0016] MR-proADM’s physiological function is unidentified, yet, but MR-proADM was identified as a prognostic marker for risk assessment and stratification for patients with sepsis or septic shock, pneumonia, myocardial infarction and heart failure25. Changes of MR-proADM levels as a result of LPS treatment in human specimen were reported to take place 90 to 120 minutes after begin of LPS administration32,33. In patients with adverse medical emergencies, MR-proADM was suitable for prediction of death and improved clinical models for intensive care unit admission34. When determined within of 24 hours after ICU admission, MR-proADM increases in line with the APACHE-II score and adds prognostic and diagnostic management value for severe sepsis and septic shock, as well as is a predictor of 28 day mortality35,36. In addition, levels of MR-proADM are associated with organ failure37. Further, MR-proADM was prognostic for 90 days mortality in heart failure38and levels of MR-proADM correlated with disease severity in chronic heart failure and were associated with increased risk of death at 12 month follow-up39,40. W02019077082 describes a method for monitoring a therapy in a subject, wherein the subject is under treatment with a binder against adrenomedullin by determining the level of a fragment of pre-pro-Adrenomedullin selected from the group comprising MR-proADM, C-terminal Proadrenomedullin (CT-pro DM) and / or Proadrenomedullin N-terminal 20 peptide (PAMP) or fragments thereof (but not bio-ADM or ADM-Gly) in a bodily fluid obtained from said subject and correlating said level of the fragment of pre-pro-Adrenomedullin with the requirement for adapting therapeutic measures of said patient, where said therapeutic measures is selected from the group comprising hydrocortisone.

[0017] The surprising finding of this invention is the early elevation of ADM-Gly levels several hours before sepsis onset and several hours before the onset of bio-ADM elevation. This allows for prediction of the transition of a patient into the state of sepsis on the one hand and allows for early decision making on patient’s therapy options with respect to standard sepsis therapy on the other hand. Further, the surprising early elevation of ADM-Gly levels allows for early decisions with respect to patient’s treatment using Adrenomedullin targeting therapies, such as usage of anti-ADM antibodies to achieve an early enrichment of bio-ADM and counteract sepsis related endothelial damage, to enhance conversion of ADM-Gly to bio-ADM (WO2021170763A1) or the use of therapeutic PAM (peptidylglycine alpha-amidating monooxygenase) enzyme to convert the early arising ADM-Gly excess into bio-ADM (WO2021170816A1).

[0018] It was the surprising finding of the present invention that ADM-Gly or fragments thereof are early biomarker(s) for the prediction that a patient will later develop sepsis, severe sepsis and septic shock. Therefore, the technical problem underlying the invention is the provision of means and methods to provide a fast and reliable way to predict the risk of getting sepsis and / or septic shock or to diagnose sepsis and / or septic shock in a subject.

[0019] The technical problem is solved by provision of the embodiments provided herein below and as characterized in the appended claims.

[0020] Description of the invention

[0021] Subject matter of the present invention is a method for: a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject or, the method comprising:

[0022] Determining the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the sum of the levels of ADM-Gly according to SEQ ID No. 4 and ADM-NH2according SEQ ID No. 5 ("total-ADM") in a bodily fluid obtained from said subject; and

[0023] A) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with the extent of sepsis and / or septic shock in said subject or diagnosing sepsis and / or septic shock in said subject, wherein an elevated level above a certain threshold is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or,

[0024] B) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with risk for getting sepsis and / or septic shock, wherein an elevated level above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock, or C) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with need of therapy or intervention of sepsis and / or septic shock, wherein an elevated level above a certain threshold is indicative for the need of therapy or intervention of sepsis and / or septic shock.

[0025] The person skilled in the art understands that the method according to the present invention comprises the following steps:

[0026] Subject matter of the present invention is a method for: a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject, the method comprising:

[0027] Determining the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the sum of the levels of ADM-Gly according to SEQ ID No. 4 and ADM-NH2 according SEQ ID No. 5 ("total-ADM") in a bodily fluid obtained from said subject; and

[0028] A) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total- ADM with the extent of sepsis and / or septic shock in said subject or diagnosing sepsis and / or septic shock in said subject, wherein an elevated level above a certain threshold is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or,

[0029] B) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total- ADM with risk for getting sepsis and / or septic shock, wherein an elevated level above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock, or C) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total - ADM with need of therapy or intervention of sepsis and / or septic shock, wherein an elevated level above a certain threshold is indicative for the need of therapy or intervention of sepsis and / or septic shock.

[0030] The same applies, mutatis mutandis, to the following corresponding embodiments of the invention.

[0031] Severity of sepsis and / or septic shock, extent of sepsis and / or septic shock, degree of sepsis and / or septic shock, grade of sepsis and / or septic shock and the like are used synonymously throughout this application.

[0032] Mature ADM, bio-ADM and ADM-NH2 is used synonymously throughout this application and is a molecule according to SEQ ID No. 5.

[0033] The present invention solves the above identified technical problem. As documented herein below and in the appended examples, it was unexpectedly found that the level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or total-ADM and / or the ratio of a level of said biomarker and fragments thereof of at least five amino acids or the total-ADM to a level of ADM-NH2 and fragments thereof of at least five amino acids demonstrate(s) a strong statistical relationship with a risk of getting sepsis and / or septic shock and in particular allows early predicting sepsis and / or septic shock with 48 hour advance, preferably 12 hour advance, more preferably 8 hour advance, more preferably 4 hour advance, and at least 2 hour advance the occurrence of sepsis and / or septic shock.

[0034] Occurrence of sepsis and / or septic shock shall mean that a patient develops clinical signs of seps as defined by the SEPSIS-2 definition42, wherein at least two of the following SEPSIS-2 definition42criteria shall be met and the patient or the patient group may be considered patients when with mild symptoms of an infectious disease, or patients with no symptoms of serious infectious disease, e.g., without symptoms of sepsis:

[0035] Development of a body temperature above >38°C or <36°C

[0036] Increase in heart rate >90 beats per minute

[0037] Developing persistent hypotension (drop in mean arterial pressure (MAP) below 60 mmHg, and drop in systolic blood pressure below 90 mmHg or a decrease in systolic blood pressure of at least 40 mmHg)

[0038] Development of any above-mentioned criteria in parallel

[0039] Accordingly, the level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or total-ADM and / or the ratio of a level of said biomarker and fragments thereof of at least five amino acids or the total-ADM to a level of ADM-NH2 and fragments thereof of at least five amino acids predict the risk of geting sepsis and / or septic shock of the subject and thus helps the clinician in the decision which therapy is most suitable. In one embodiment, said biomarker is ADM-Gly and the level of ADM-Gly and fragments thereof of at least five amino acids is higher than 20 pg / mL in a bodily fluid, preferably is higher than 25 pg / ml, more preferably is higher than 30 pg / mL, more preferably is higher than 40 pg / mL, more preferably is higher than 50 pg / mL while the level of bio-ADM is in the range of from 5 to 30 pg / mL and / or said ratio in a bodily fluid is elevated over 1.5, preferably elevated over 1.6, more preferably elevated over 1.7, more preferably is elevated over 1.8, more preferable is elevated over 1.9, more preferably is higher than 2.

[0040] In one embodiment, elevated levels of ADM-Gly means elevation of ADM-Gly by 20% over the given threshold.

[0041] In another embodiment, elevated levels of said ratio means elevation of the ratio by 20% over the given threshold.

[0042] Throughout this application the ratio of a level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or total-ADM to a level of ADM-NH2 and fragments thereof of at least five amino acids is equivalent to a reversed ratio of a level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR- proADM and CT-proADM or fragments thereof of at least five amino acids or total-ADM to a level of ADM-Gly and fragments thereof of at least five amino acids. For example, the ratio of ADM-Gly to ADM-NH2 which is 2 is equivalent to the ratio of ADM-NH2 to ADM-Gly which is 0.5. It shall be understood throughout the specification the determining of the ratio of a level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or total-ADM to a level of ADM-NH2 and fragments thereof of at least five amino acids can be conducted in the way that the ratio of a level of ADM-NH2 and fragments thereof of at least five amino acids to a level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or total-ADM is determined and the corresponding thresholds are adapted to the reversed values. In this case a reduced ratio below a certain threshold is indicative.

[0043] The present invention has, inter alia, the following advantages over the conventional methods: the inventive methods and the kits are fast, objective, easy to use and precise for the prediction of an adverse event: The methods and kits of the invention relate to markers that are easily measurable in routine in hospitals because the levels of ADM-Gly and ADM-NH2 can be determined in routinely obtained blood samples or further biological fluids obtained from a subject. In addition, the determination of the levels of ADM-Gly and ADM-NH2 is very fast. Therefore, the methods and the kits of the invention are suitable for a quick assessment, and diagnosis and prognosis of sepsis and / or septic shock. Furthermore, due to the simple outcome of a biomarker measurement as one specific value, there is no subjective bias of medical staff when using this method or the kits of the invention.

[0044] Accordingly, the herein provided methods and kits are advantageous in the diagnosis, prognosis, risk assessment, and / or risk stratification of sepsis and / or septic shock, of a subject.

[0045] In some embodiments the invention relates to a method for: a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject or, the method comprising:

[0046] Determining the ratio of the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM a level of ADM-NH2 according SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; and

[0047] A) Correlating said ratio with the extent of sepsis and / or septic shock in said subject or diagnosing sepsis and / or septic shock in said subject, wherein an elevated ratio above a certain threshold is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or,

[0048] B) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock, or,

[0049] C) Correlating said ratio with need of therapy or intervention of sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is indicative for the need of therapy or intervention of sepsis and / or septic shock.

[0050] In one embodiment, the invention relates to a method for early assessing the risk of getting sepsis and / or septic shock in a subject or, the method comprising: A) Determining the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; and

[0051] B) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock.

[0052] The person skilled in the art understands that in one embodiment, the invention relates to a method for early assessing the risk of getting sepsis and / or septic shock in a subject, the method comprising:

[0053] A) Determining the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM -NHz according to SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; and

[0054] B) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock.

[0055] The same applies, mutatis mutandis, to the following corresponding embodiments of the invention.

[0056] In one embodiment, the invention relates to a method for early assessing the risk of getting sepsis and / or septic shock in a subject or, the method comprising:

[0057] A) Determining the ratio of total -ADM to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; and

[0058] B) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock.

[0059] In one embodiment, the invention relates to a method for early assessing the risk of getting sepsis and / or septic shock in a subject or, the method comprising:

[0060] A) Determining the ratio of a level of ADM-Gly according to SEQ ID No. 4 or the total-ADM to a level of ADM-NHz according to SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; and

[0061] B) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock. In some embodiments, said method is for early assessing the risk of getting sepsis and / or septic shock in a subject or, wherein the early assessing is the prediction with 48 hours, 24 hours, 12 hours, 6 hours, 4 hours, preferably 2 hours advance the occurrence of sepsis and / or septic shock. In some embodiments, said method is for early assessing the risk of getting sepsis and / or septic shock in a subject, wherein said subject has been identified of having risk of getting sepsis and / or septic shock and wherein a level of procalcitonin in plasma of said subject is less than 2.0 pg / 1, preferably less than 1.5 pg / 1, more preferably less than 1.0 pg / 1, yet more preferably less than 0.5 pg / 1.

[0062] In some embodiments said level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids or said ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids is determined by using a binder to ADM-Gly according to SEQ ID No.: 4 or fragments thereof of at least five amino acids in said method.

[0063] In some embodiments of the method, the binder is selected from the group comprising an antibody, an antibody fragment or a non-Ig-Scaffold binding to ADM-Gly according to SEQ ID No. 4 or fragments thereof of at least five amino acids.

[0064] In some embodiments of the method, the threshold for a level of ADM-Gly and fragments thereof of at least five amino acids is 20 pg / mL in a bodily fluid, preferably 25 pg / ml, more preferably 30 pg / mL, more preferably 40 pg / mL, more preferably 50 pg / mL, wherein the level of ADM-NH2 according to SEQ ID No. 5 or fragments thereof of at least five amino acids is in the range of from 5 to 30 pg / rnL.

[0065] In some embodiments of the method, the threshold for the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 or fragments thereof of at least five amino acids in a bodily fluid is 1 .5, preferably 1.6, more preferably 1.7, more preferably 1.8, more preferably 1.9, more preferably 2.

[0066] In some embodiments of the method, the fragments of ADM-Gly according to SEQ ID No. 4 of at least five amino acids comprise a fragment according to SEQ ID No. 18.

[0067] In some embodiments of the method, the fragments of ADM-NH2 according to SEQ ID No. 5 of at least five amino acids comprise a fragment according to SEQ ID No. 19.

[0068] In some embodiments of the method, the fragments are selected from the group comprising SEQ ID No.

[0069] 7, SEQ ID No. 8, SEQ ID No. 9, SEQ ID No. 10 and SEQ ID No. 11. In some embodiments of the method, the fragments are selected from the group comprising SEQ ID No. 12, SEQ ID No. 13, SEQ ID No. 14, SEQ ID No. 15 and SEQ ID No. 16.

[0070] In some embodiments of the method, the determination of the level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or total-ADM and / or the ratio of a level of said biomarker and fragments thereof of at least five amino acids or the total-ADM to a level of ADM-NH2and fragments thereof of at least five amino acids is performed more than once in said subject.

[0071] In some embodiments, the method is for stratifying the subjects into sepsis and / or septic shock grade groups.

[0072] In some embodiments of the method, the level of said biomarker or fragments thereof or said ratio is used for guidance of therapy or intervention of sepsis and / or septic shock, wherein therapy or intervention is indicated if the level of said biomarker or fragments thereof or said ratio is above a certain threshold and wherein therapy or intervention of sepsis and / or septic shock is not indicated if the level of said biomarker or fragments thereof or said ratio is below a certain threshold.

[0073] In a preferred embodiment of the method, the level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids or the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids is used for guidance of therapy or intervention of sepsis and / or septic shock, wherein therapy or intervention is indicated if the level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids or the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids is above a certain threshold and wherein therapy or intervention of sepsis and / or septic shock is not indicated if the level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids or the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids is below a certain threshold.

[0074] In some embodiments of the method, said subject is a human.

[0075] In some embodiments of the method, said therapy is selected from the group comprising administration of antibiotics, vasopressor therapy including administration of vasopressin and Angiotensin II (e.g. Giapreza), immunosuppressive therapy, renal therapy including administration of Alkaline Phosphatase. In some embodiment of the method, said therapy comprises administering anti-ADM antibodies, preferably N-terminal anti-ADM antibodies, and / or PAM (peptidylglycine alpha-amidating monooxygenase) as therapeutic agent.

[0076] In a preferred embodiment of the method, said therapy comprises administering vasopressor to said subject, and wherein in addition the level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids or the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NHj according to SEQ ID No. 5 and fragments thereof of at least five amino acids is determined and wherein treatment with said anti-ADM antibody or anti-ADM antibody fragment is initiated and / or continued when the level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids or the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids in said sample is above a certain threshold and / or wherein a treatment with an anti-ADM antibody or anti-ADM antibody fragment is withheld and / or terminated if the said level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids or the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids is below said predetermined threshold.

[0077] In some embodiments of the method, the bodily fluid is selected from whole blood, plasma, serum, urine, cerebrospinal liquid (CSF), and saliva.

[0078] It is also envisaged herein that polypeptides can be determined, which have a sequence identity to ADM- Gly, PAMP, MR-proADM, CT-proADM and ADM-NH2. For example, polypeptides can be determined in the methods and kits of the invention that have at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to ADM-Gly and ADM-NH2. In accordance with the present invention, the terms "sequence identity", "homology" or "percent homology" or "identical" or "percent identity" or "percentage identity" in the context of two or more amino acid sequences refers to two or more sequences or subsequences that are the same, or that have a specified percentage of amino acids that are the same, when compared and aligned for maximum correspondence over the window of comparison (preferably over the full length), or over a designated region as measured using a sequence comparison algorithm as known in the art, or by manual alignment and visual inspection. Sequences having, for example, 70% to 90% or greater (preferably 95% or greater) sequence identity may be considered to be substantially identical. Such a definition also applies to the complement of a test sequence. Preferably, the described identity exists over a region that is at least about 10 to about 15 amino acids in length, more preferably, over a region that is at least about 20 to about 35 amino acids in length, most preferably, over the full length. Those having skill in the art will know how to determine percent identity between / among sequences using, for example, algorithms such as those based on CLUSTALW computer program (Thompson Nucl. Acids Res. 2 (1994), 4673-4680) or FASTDB (Brutlag Comp. App. Biosci. 6 (1990), 237-245), as known in the art.

[0079] The term “biomarker” or “early biomarker” means that the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM or the ratio of a level of said biomarker or fragments thereof or total- ADM to a level of ADM-NHj and fragments thereof is elevated in a patient before the patient has developed sepsis, severe sepsis or septic shock.

[0080] The term "subject" as used herein refers to a living human or non-human organism. Preferably herein the subject is a human subject. The subject may be healthy or diseased if not stated otherwise.

[0081] As used herein, the “level” of the marker refers to the quantity of the molecular entity of the marker in the sample. In other words, the concentration of the marker is determined in the sample. For example, the concentration of ADM-Gly is determined in the sample of the subject.

[0082] A “bodily fluid” may be selected from the group comprising blood, serum, plasma, urine, cerebrospinal liquid (CSF), and saliva. A bodily fluid according to the present invention is in one particular embodiment a blood sample. A blood sample may be selected from the group comprising whole blood, serum and plasma. In a specific embodiment of the diagnostic method said sample is selected from the group comprising human citrate plasma, heparin plasma and EDTA plasma.

[0083] The term “prediction” relates to the prognosis of an outcome or a specific risk for a subject. This may also include an estimation of the chance of recovery or the chance of an adverse outcome for said subject.

[0084] The methods of the invention may also be used for monitoring, therapy monitoring, therapy guidance and / or therapy control. In the context of the present application “monitoring” relates to keeping track of a patient and potentially occurring complications, e.g., to analyze the progression of the healing process or the influence of a particular treatment or therapy on the health state of the patient.

[0085] The term “therapy monitoring” or “therapy control” in the context of the present invention refers to the monitoring and / or adjustment of a therapeutic treatment of said patient, for example by obtaining feedback on the efficacy of the therapy. As used herein, the term “therapy guidance” refers to application of certain therapies, therapeutic actions or medical interventions based on the value / level of one or more biomarkers and / or clinical parameter and / or clinical scores. This includes the adjustment of a therapy or the discontinuation of a therapy. In the present invention, the terms “risk assessment” and “risk stratification” relate to the grouping of subjects into different risk groups according to their further prognosis. Risk assessment also relates to stratification for applying preventive and / or therapeutic measures. The term “therapy stratification” in particular relates to grouping or classifying patients into different groups, such as risk groups or therapy groups that receive certain differential therapeutic measures depending on their classification. The term “therapy stratification” also relates to grouping or classifying patients with infections or having symptoms of an infectious disease into a group that are not in need to receive certain therapeutic measures.

[0086] Sepsis is defined as life-threatening organ dysfunction caused by a dysregulated host response to infection41. Organ dysfunction can be identified as an acute change in total SOFA (Sequential Organ Failure Assessment Score) score >2 points consequent to the infection.

[0087] The baseline SOFA score can be assumed to be zero in patients not known to have preexisting organ dysfunction. A SOFA score >2 reflects an overall mortality risk of approximately 10% in a general hospital population with suspected infection. Even patients presenting with modest dysfunction can deteriorate further, emphasizing the seriousness of this condition and the need for prompt and appropriate intervention, if not already being instituted. Sepsis is a life-threatening condition that arises when the body’s response to an infection injures its own tissues and organs. Patients with suspected infection who are likely to have a prolonged ICU stay or to die in the hospital can be promptly identified at the bedside with qSOFA, i.e., alteration in mental status, systolic blood pressure <100 mm Hg, or respiratory rate >22 / min.

[0088] Septic shock is a subset of sepsis in which underlying circulatory and cellular / metabolic abnormalities are profound enough to substantially increase mortality. Patients with septic shock can be identified with a clinical construct of sepsis with persisting hypotension requiring vasopressors to maintain mean arterial pressure (MAP) > 65 mm Hg and having a serum lactate level >2 mmol / L (18 mg / dL) despite adequate volume resuscitation. With these criteria, hospital mortality is in excess of 40%.

[0089] The term "sepsis" as used in the context of the present application relates to all possible stages in the development of sepsis. The term “sepsis” also includes severe sepsis or septic shock based on the SEPSIS-2 definition42. The term “sepsis” also includes subjects falling within the SEPSIS-3 definition41. Sepsis is an acutely life-threatening organ dysfunction caused by an inadequate host response to an infection. For diagnosing sepsis-associated organ dysfunction, a change in the Sequential Organ Failure Assessment (SOFA) score of > 2 points should be used. As used herein, organ dysfunction denotes a condition or a state of health where an organ does not perform its expected function. “Organ failure” denotes an organ dysfunction to such a degree that normal homeostasis cannot be maintained without external clinical intervention. Said organ failure may pertain an organ selected from the group comprising kidney, liver, heart, lung, nervous system. By contrast, organ function represents the expected function of the respective organ within physiologic ranges. The person skilled in the art is aware of the respective function of an organ during medical examination.

[0090] Organ dysfunction may be defined by the sequential organ failure assessment score (SOFA-Score) or the components thereof. The SOFA score, previously known as the sepsis-related organ failure assessment score41is used to track a person’s status during the stay in an intensive care unit (ICU) to determine the extent of a person’s organ function or rate of failure. The score is based on six different scores, one each for the respiratory, cardiovascular, hepatic, coagulation, renal and neurological systems each scored from 0 to 4 with an increasing score reflecting worsening organ dysfunction. The criteria for assessment of the SOFA score are described for example in Lamden et al. (reviewed in43). SOFA score may traditionally be calculated on admission to ICU and at each 24-h period that follows.

[0091] In particular, said organ dysfunction is selected from the group comprising renal decline, cardiac dysfunction, liver dysfunction or respiratory tract dysfunction.

[0092] The quick SOFA Score (quickSOFA or qSOFA) was introduced by the Sepsis-3 group in February 2016 as a simplified version of the SOFA Score as an initial way to identify patients at high risk for poor outcome with an infection44. The qSOFA simplifies the SOFA score drastically by only including its 3 clinical criteria and by including “any altered mentation” instead of requiring a GCS <15. qSOFA can easily and quickly be repeated serially on patients. The score ranges from 0 to 3 points. One point is given for: low blood pressure (SBP <100 mm Hg), high respiratory rate ((> 22 breaths / min) and altered mentation (GCS < 15). The presence of 2 or more qSOFA points near the onset of infection was associated with a greater risk of death or prolonged intensive care unit stay. These are outcomes that are more common in infected patients who may be septic than those with uncomplicated infection. Based upon these findings, the Third International Consensus Definitions for Sepsis recommends qSOFA as a simple prompt to identify infected patients outside the ICU who are likely to be septic45.

[0093] In preferred embodiments of the present invention the method is defined by the prediction of sepsis, severe sepsis and / or septic shock in a patient group, for whom it was previously difficult, if not impossible, to determine by routine clinical and / or molecular diagnostic means, whether a serious risk existed of serious deterioration of medical conditions, thereby requiring hospitalization. This patient group may be considered patients with mild symptoms of an infectious disease, or patients with no symptoms of serious infectious disease, e.g., without symptoms of sepsis.

[0094] In one embodiment, at the time of sample provision, the patient shows mild symptoms of an infectious disease corresponding to a qSOFA score of 0 or 1. In one embodiment, at the time of sample provision, the patient shows no clinical symptoms for a sepsis, a severe sepsis and / or septic shock.

[0095] In one embodiment, at the time of sample provision, the patient has been diagnosed of having an infectious disease of bacterial, viral, fungal, or parasitic origin.

[0096] In one embodiment, at the time of sample provision, the patient has been diagnosed with community acquired pneumonia (CAP) or urinary tract infection (UTI).

[0097] Delayed treatment in patients presenting to the emergency department (ED) with a suspected infection may result in a prolonged hospitalisation, an increased morbidity, and a greater rate of infection-related mortality.

[0098] An accurate assessment of the severity of the host response and the potential for further disease progression to sepsis, severe sepsis, septic shock and organ dysfunction is therefore crucial in order to administer a rapid and targeted therapeutic response.

[0099] One object of the invention is therefore the use of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or the total-ADM to distinguish patients who are more likely or have a high risk of developing sepsis, severe sepsis or septic shock requiring intensive treatments from patients who have a low risk of requiring such treatment.

[0100] In one embodiment of the invention, the method is used in order to stratify said patients into risk groups.

[0101] In one embodiment, a high-risk level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or the total- ADM indicates that the patient is at risk of developing sepsis, severe sepsis and / or septic shock within 48 hours, 24 hours, 12 hours, 6 hours, 4 hours, preferably 2 hours and wherein a low-risk level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or the total-ADMindicates that the patient is not at risk of developing sepsis, severe sepsis or septic shock within 48 hours, 24 hours, 12 hours, 6 hours, 4 hours, preferably 2 hours.

[0102] In one embodiment, the level a biomarker selected from the group consisting of ADM-Gly, PAMP, MR- proADM and CT-proADM or fragments thereof of at least five amino acids or the total-ADM in said sample of bodily fluid is indicative of the risk of a development of a sepsis, severe sepsis and / or septic shock in the patient that requires hospitalization. In one embodiment, a high-risk level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR-proADM and CT-proADM or fragments thereof of at least five amino acids or the total- ADMindicates that the patient is at risk of developing a sepsis, severe sepsis and / or septic shock that requires hospitalization within 48 hours, 24 hours, 12 hours, 6 hours, 4 hours, preferably 2 hours and wherein a low-risk level of a biomarker selected from the group consisting of ADM-Gly, PAMP, MR- proADM and CT-proADM or fragments thereof of at least five amino acids or the total-ADM indicates that the patient is not at risk of developing sepsis, severe sepsis and / or septic shock that requires hospitalization within 48 hours, 24 hours, 12 hours, 6 hours, 4 hours, preferably 2 hours.

[0103] In another embodiment of the invention, the method is used to stratify said patients into groups for early treatment (e.g., requirement of antibiotic administration).

[0104] The term “early” is defined as time of treatment before the patient shows clinical signs and symptoms of sepsis, severe sepsis and / or septic shock or before the patient has been diagnosed for having sepsis, severe sepsis and / or septic shock.

[0105] In another embodiment of the invention, the method is used to stratify said patients into groups for early treatment (e.g., requirement of anti-ADM antibody administration, preferably N-terminal anti-ADM antibody administration). The term “early” is defined as time of treatment before the patient shows clinical signs and symptoms of sepsis, severe sepsis and / or septic shock or before the patient has been diagnosed for having sepsis, severe sepsis and / or septic shock.

[0106] In one embodiment, the level of ADM-Gly or fragments thereof in said sample of bodily fluid is indicative of the patient requiring frequent monitoring and / or critical care treatment. In one embodiment, the patient with a high-risk level will require a medical treatment provided in a hospital setting.

[0107] Examples of these treatments include but are not limited to fluid therapy, vasopressors, intravenous antibiotics, in some embodiments essentially any treatment beyond oral antibiotics, which may be selfadministered at home.

[0108] Depending on the result of the method of the present invention, embodiments of the method may comprise subsequent therapeutic decisions and / or therapeutic actions. Such therapeutic decisions may include the initiation, change or modification of medical treatment. Preferably, if the method of the present invention is indicative of development to a condition requiring treatment in hospital, suitable therapeutic measures, such as initiation or change of a certain medication or fluid therapy may be initiated. Any therapy, medical treatment or therapeutic action disclosed herein can be employed in the context of the method of the invention as a subsequent therapeutic decision or therapeutic action, in particular if the therapeutic measure is specifically administered in hospital, including but not limited to intravenous fluid therapy, dialysis, management of electrolyte abnormalities (in particular potassium, calcium and phosphorus). Furthermore, a maintained intensive observation and care of the patient may be indicated potentially over extended periods of time, such as several days, weeks or even months. This may involve keeping or moving the patient to an ICU and / or prolonging the stay of the patient in an ICU.

[0109] On the other hand, if the result of the method of the present invention is indicative of the absence of risk of developing a sepsis, severe sepsis or septic shock requiring hospitalization, no specific treatment measures with respect to such complication may be required, or less serious, self- administrable treatments may be prescribed.

[0110] ADM-Gly or fragments thereof are in particular for the before -mentioned medical utilities in the patient group of Emergency Department all -comers.

[0111] Throughout the specification the term ADM-Gly includes all splice variants of ADM-Gly if not stated otherwise. Throughout the specification it should be understood that the term “total-ADM” is the sum of the levels of ADM-Gly according to SEQ ID No. 4 and ADM-NEE according SEQ ID No. 5.

[0112] The term “determining the level of ADM-Gly, its splice variants or fragments thereof of at least 5 amino acids including SEQ ID’s 2-16 means, that usually the immunoreactivity towards a region within the before mentioned molecules is determined. This means that it is not necessary that a certain fragment is measured selectively. It is understood that a binder which is used for the determination of the level of ADM-Gly or fragments thereof of at least 5 amino acids including SEQ ID’s 2-16 binds to any fragment that comprises the region of binding of said binder. Said binder may be an antibody or antibody fragment or a non-IgG scaffold.

[0113] Subject matter according to the present invention is a method, wherein the level of ADM-Gly or fragments thereof of at least 5 amino acids is determined by using a binder to ADM-Gly or fragments thereof of at least 5 amino acids.

[0114] In one embodiment of the invention said binder is selected from the group comprising an antibody, an antibody fragment or a non-Ig-scaffold binding to ADM-Gly or fragments thereof of at least 5 amino acids. As used herein, the "reference level" or “threshold” may reflect a normal level of the corresponding marker that is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or predictive for an enhanced risk of getting sepsis and / or septic shock, or indicative for the need of therapy or intervention of sepsis and / or septic shock.

[0115] In one embodiment, the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 or fragments thereof of at least five amino acids in a bodily fluid is 1.5, preferably 1.6, more preferably 1.7, more preferably 1.8, more preferably 1.9, more preferably 2. The reference level applied depends on the desired specificity and sensitivity for the prediction, i.e. a higher threshold such as about 2 will reliably identify subjects which have an enhanced risk of getting sepsis and / or septic shock, or indicative for the need of therapy or intervention of sepsis and / or septic shock.

[0116] In one embodiment, the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 or fragments thereof of at least five amino acids in a bodily fluid is 1.5, preferably 1.6, more preferably 1.7, more preferably 1.8, more preferably 1.9, more preferably 2, which is predictive for an enhanced risk of getting sepsis and / or septic shock with 48 hours, 24 hours, 12 hours, 6 hours, 4 hours, preferably 2 hours advance the occurrence of sepsis and / or septic shock.

[0117] As used herein, the term "elevated level of (the) marker " means that the level of the marker is increased, i.e. it refers to an increased level of the marker. Accordingly, the term "increase in the level of (the) marker" or “increased level” is used interchangeably herein with the term "elevated level of (the) marker". The term "elevated level” means a level above a certain threshold level. An increased level of the marker or an increase in the level of the marker of the subject means that the level of the marker is at least about 15%, preferably at least about 20%, more preferably at least about 25%, or even more preferably at least about 30% higher than the reference level of the marker.

[0118] As used herein, the term "sample" is a biological sample that is obtained from the subject. "Sample" as used herein may, e.g., refer to a sample of bodily fluid or tissue obtained for the purpose of diagnosis, prognosis, or evaluation of a subject of interest, such as a patient. Preferably herein, the sample is a sample of a bodily fluid, such as blood, serum, plasma, cerebrospinal fluid, urine, saliva, sputum, and pleural effusions. Particularly, the sample is blood, blood plasma, blood serum, or urine. The samples could be processed (pre-treated), such as by fractionation or purification procedures, for example, separation of whole blood into serum or plasma components. Such pre-treatments can also include, but are not limited to dilution, filtration, centrifugation, concentration, sedimentation, precipitation or dialysis. Pre-treatments may also include the addition of chemical or biochemical substances to the solution, such as acids, bases, buffers, salts, solvents, reactive dyes, detergents, emulsifiers, chelators. Preferably, the sample is a blood sample, more preferably a serum sample or a plasma sample. "Plasma" in the context of the present invention is the virtually cell-free supernatant of blood containing anticoagulant obtained after centrifugation. Exemplary anticoagulants include calcium ion binding compounds such as EDTA or citrate and thrombin inhibitors such as heparinates or hirudin. Cell-free plasma can be obtained by centrifugation of the anticoagulated blood (e.g. citrated, EDTA or heparinized blood), for example for at least 15 minutes at 2000 to 3000 g.

[0119] "Serum" in the context of the present invention is the liquid fraction of whole blood that is collected after the blood is allowed to clot. When coagulated blood (clotted blood) is centrifuged serum can be obtained as supernatant.

[0120] As used herein, "urine" is a liquid product of the body secreted by the kidneys through a process called urination (or micturition) and excreted through the urethra.

[0121] As used herein, the term, "antibody" refers to immunoglobulin molecules and immunologically active portions of immunoglobulin (Ig) molecules, i.e., molecules that contain an antigen binding site that specifically binds (immuno reacts with) an antigen. According to the invention, the antibodies may be monoclonal as well as polyclonal antibodies. Particularly, antibodies that bind specifically to ADM-Gly or ADM-NH2 are used. An antibody is considered to be specific, if its affinity towards the molecule of interest, e.g. ADM-Gly or ADM-NH2, or the fragment thereof is at least 50-fold higher, preferably 100- fold higher, most preferably at least 1000-fold higher than towards other molecules comprised in a sample containing the molecule of interest. It is well known in the art how to develop and to select antibodies with a given specificity. In the context of the invention, monoclonal antibodies are preferred. The antibody or the antibody binding fragment binds specifically to the herein defined markers or fragments thereof. The anti-ADM-Gly antibody specifically discriminates between the amidated and c- terminally glycine-extended forms of ADM. Therefore, the c -terminal glycine in its epitope, as specified in Seq. ID No.: 18, is mandatory for the anti-ADM-Gly antibody to react with ADM-Gly and the epitope lacking the c-terminal Glycine, as specified in Seq. ID No.: 19, is not recognized by the anti-ADM-Gly antibody. Thus, the herein defined peptides can also be epitopes to which the antibodies specifically bind to. Exemplary immunoassays can be luminescence immunoassay (LIA), radioimmunoassay (RIA), chemiluminescence- and fluorescence- immunoassays, enzyme immunoassay (EIA), Enzyme-linked immunoassays (ELISA), luminescence-based bead arrays, magnetic beads-based arrays, protein microarray assays, rapid test formats, rare cryptate assay. Further, assays suitable for point-of-care testing and rapid test formats such as for instance immune-chromatographic strip tests can be employed.

[0122] As used herein, "diagnosis" in the context of the present invention relates to the recognition and (early) detection of sepsis and / or septic shock, in a subject and may also comprise differential diagnosis. Also, the assessment of the severity of sepsis and / or septic shock may be encompassed by the term "diagnosis" . For example, the assessment of how critical the condition is and how likely the occurrence of sepsis and / or septic shock is. In addition, diagnosis means that the time can be predicted when sepsis and / or septic shock in the subject occurs.

[0123] The invention further relates to kits, the use of the kits and methods wherein such kits are used. The invention relates to kits for carrying out the herein above and below provided methods. The herein provided definitions, e.g. provided in relation to the methods, also apply to the kits of the invention. In particular, the invention relates to kits for diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, early assessing the risk of getting sepsis and / or septic shock in a subject or, predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject, wherein said kit comprises detection reagents for determining said level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or total-ADM or said ratio of a level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or total-ADM to a level of ADM-NH2 according to SEQ ID No.: 5 and fragments thereof of at least five amino acids in said sample of said subject.

[0124] The kit according to the invention may further include reference data indicating the elevated level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or total-ADM or the elevated ratio of a level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or total-ADMto a level of ADM-NH2 and fragments thereof which is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or predictive for an enhanced risk of getting sepsis and / or septic shock or indicative for the need of therapy or intervention of sepsis and / or septic shock.

[0125] In particular, the invention relates to kits for early assessing the risk of getting sepsis and / or septic shock in a subject, wherein said kit comprises detection reagents for determining said level of ADM-Gly according to SEQ ID No.: 4 and fragments thereof of at least five amino acids or said ratio of a level of ADM-Gly according to SEQ ID No.: 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No.: 5 and fragments thereof of at least five amino acids in said sample of said subject, and reference data indicating the elevated level of ADM-Gly and fragments thereof or the elevated ratio of a level of ADM-Gly and fragments thereof to a level of ADM-NH2 and fragments thereof, which is predictive for an enhanced risk of getting sepsis and / or septic shock with 48 hours, 24 hours, 12 hours, 6 hours, 4 hours, preferably 2 hours advance the occurrence of sepsis and / or septic shock.

[0126] The invention also relates to a kit for and its use in the method according to the invention.

[0127] In particular, the invention relates to a use of the kit for a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject.

[0128] In particular, the invention relates to a use of the kit for a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject in a method comprising the following steps:

[0129] Determining the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or total- ADM in a bodily fluid obtained from said subject; and

[0130] A) Correlating said level of said biomarker and fragments thereof of at least five amino acids with the extent of sepsis and / or septic shock in said subject or diagnosing sepsis and / or septic shock in said subject, wherein an elevated level above a certain threshold is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or, B) Correlating said level of said biomarker and fragments thereof of at least five amino acids with risk for getting sepsis and / or septic shock, wherein an elevated level above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock, or

[0131] C) Correlating said level of said biomarker and fragments thereof of at least five amino acids with need of therapy or intervention of sepsis and / or septic shock, wherein an elevated level above a certain threshold is indicative for the need of therapy or intervention of sepsis and / or septic shock.

[0132] As used herein, the "detection reagent" or the like are reagents that are suitable to determine the herein described marker(s), e.g. ADM-Gly or ADM-NH2 or fragments thereof. Such exemplary detection reagents are, for example, ligands, e.g. antibodies or fragments thereof, which specifically bind to the peptide or epitopes of the herein described marker(s). Such ligands might be used in immunoassays as described above. Further reagents that are employed in the immunoassays to determine the level of the marker(s) may also be comprised in the kit and are herein considered as detection reagents.

[0133] Detection reagents can also relate to reagents that are employed to detect the markers or fragments thereof by MS based methods. Such detection reagent can thus also be reagents, e.g. enzymes, chemicals, buffers, etc, that are used to prepare the sample for the MS analysis. A mass spectrometer can also be considered as a detection reagent. Detection reagents according to the invention can also be calibration solution(s), e.g. that can be employed to determine and compare the level of the marker(s).

[0134] The given definitions and explanations also apply mutatis mutandis to the following items. The present invention also relates to the following items:

[0135] 1. A method for a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject or, the method comprising:

[0136] Determining the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the sum of the levels of ADM-Gly according to SEQ ID No. 4 and ADM-NH2 according SEQ ID No. 5 ("total-ADM") in a bodily fluid obtained from said subject; and

[0137] A) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with the extent of sepsis and / or septic shock in said subject or diagnosing sepsis and / or septic shock in said subject, wherein an elevated level above a certain threshold is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or,

[0138] B) Correlating said level of said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with risk for getting sepsis and / or septic shock, wherein an elevated level above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock, or

[0139] C) Correlating said level of said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with need of therapy or intervention of sepsis and / or septic shock, wherein an elevated level above a certain threshold is indicative for the need of therapy or intervention of sepsis and / or septic shock.

[0140] The person skilled in the art is aware that item 1 comprises the following steps:

[0141] A method for a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject, the method comprising:

[0142] Determining the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the sum of the levels of ADM-Gly according to SEQ ID No. 4 and ADM-NH2 according SEQ ID No. 5 ("total-ADM") in a bodily fluid obtained from said subject; and

[0143] A) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total- ADM with the extent of sepsis and / or septic shock in said subject or diagnosing sepsis and / or septic shock in said subject, wherein an elevated level above a certain threshold is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or,

[0144] B) Correlating said level of said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with risk for getting sepsis and / or septic shock, wherein an elevated level above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock, or

[0145] C) Correlating said level of said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with need of therapy or intervention of sepsis and / or septic shock, wherein an elevated level above a certain threshold is indicative for the need of therapy or intervention of sepsis and / or septic shock.

[0146] This also applies, mutatis mutandis, to the following items.

[0147] 2. The method of item 1 for a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject or, the method comprising:

[0148] Determining the ratio of of the level of a biomarker selected from the group consisting of ADM- Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM to a level of ADM-NEE according SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; and A) Correlating said ratio with the extent of sepsis and / or septic shock in said subject or diagnosing sepsis and / or septic shock in said subject, wherein an elevated ratio above a certain threshold is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or,

[0149] B) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock, or

[0150] C) Correlating said ratio with need of therapy or intervention of sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is indicative for the need of therapy or intervention of sepsis and / or septic shock. . The method of item 1 or 2 for early assessing the risk of getting sepsis and / or septic shock in a subject or, the method comprising:

[0151] Determining the ratio of a level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; and

[0152] B) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock.

[0153] The person skilled in the art is aware that item 3 comprises method of item 1 or 2 for early assessing the risk of getting sepsis and / or septic shock in a subject, the method comprising:

[0154] Determining the ratio of a level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; and

[0155] B) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock.

[0156] This also applies, mutatis mutandis, to the following items. . The method according to any of items 1 to 3, wherein the early predicting is the prediction with 48 hours, 24 hours, 12 hours, 6 hours, 4 hours, preferably 2 hours advance the occurrence of sepsis and / or septic shock. . The method according to any of items 1 to 4 , wherein said biomarker is ADM-Gly according to SEQ ID No. 4 and said level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids or said ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NI I2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids is determined by using a binder to ADM-Gly according to SEQ ID No.: 4 or fragments thereof of at least five amino acids.

[0157] 6. The method according to item 5, wherein the binder is selected from the group comprising an antibody, an antibody fragment or a non-Ig-Scaffold binding to ADM-Gly according to SEQ ID No. 4 or fragments thereof of at least five amino acids.

[0158] 7. The method according to any of items 1 to 6, wherein said threshold for a level of ADM-Gly and fragments thereof of at least five amino acids is 20 pg / mL in a bodily fluid, preferably 25 pg / ml, more preferably 30 pg / mL, more preferably 40 pg / mL, more preferably 50 pg / mL, wherein the level of ADM-NH2 according to SEQ ID No. 5 or fragments thereof of at least five amino acids is in the range of from 5 to 30 pg / mL.

[0159] 8. The method according to any of items 2 to 7 , wherein said threshold for the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 or fragments thereof of at least five amino acids in a bodily fluid is 1 .5, preferably 1.6, more preferably 1.7, more preferably 1.8, more preferably 1.9, more preferably 2.

[0160] 9. The method according to any of items 1 to 8, wherein said fragments of ADM-Gly according to SEQ ID No. 4 of at least five amino acids comprise a fragment according to SEQ ID No. 18. 10. The method according to any of items 2 to 9, wherein said fragments of ADM-NH2 according to SEQ ID No. 5 of at least five amino acids comprise a fragment according to SEQ ID No. 19. 11. The method according to item 9, wherein said fragments are selected from the group comprising SEQ ID No. 7, SEQ ID No. 8, SEQ ID No. 9, SEQ ID No. 10 and SEQ ID No. 11. 12. The method according to item 10, wherein said fragments are selected from the group comprising SEQ ID No. 12, SEQ ID No. 13, SEQ ID No. 14, SEQ ID No. 15 and SEQ ID No. 16. 13. The method according to any of items 1 to 12 wherein said determination of said level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT- proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM or said ratio is performed more than once in said subject. 14. The method according to any of items 1 to 13 in order to stratify said subjects into sepsis and / or septic shock grade groups. 15. The method according to any of items 1 to 14 wherein said level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM or said ratio is used for guidance of therapy or intervention of sepsis and / or septic shock, wherein therapy or intervention is indicated if the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM or said ratio is above a certain threshold and wherein therapy or intervention of sepsis and / or septic shock is not indicated if said level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR- proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM or said ratio is below a certain threshold. 16. The method according to any of items 1 to 15 wherein said subject is a human. 17. The method according to any of items 1 to 16 wherein said therapy is selected from the group comprising administration of antibiotics, vasopressor therapy, immunosuppressive therapy.

[0161] 18. The method according to any of items 1 to 17, wherein said therapy comprises administering anti-ADM antibodies, preferably N-terminal anti-ADM antibodies, and / or PAM as therapeutic agent.

[0162] 19. The method according to any of items 1 to 18, wherein said bodily fluid is selected from whole blood, plasma, serum, urine, cerebrospinal liquid (CSF), and saliva.

[0163] 20. A kit for carrying out the method according to any one of items 1 to 19, wherein said kit comprises detection reagents for determining said level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR- proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM or said ratio in said sample of said subject.

[0164] 21. Use of the kit according to item 20 in the method of any one of the claims 1 to 19.

[0165] 22. Use of the kit according to item 20 or 21 for a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject.

[0166] As used herein, the terms "comprising" and "including", or grammatical variants thereof are to be taken as specifying at least the stated features, integers, steps or components but do not preclude the addition of one or more additional features, integers, steps, components or groups thereof. This term encompasses the terms "consisting of and "consisting essentially of that are understood to specify only the stated feature, integers, steps or components to the exclusion of any additional features.

[0167] Thus, the terms "comprising / including / having" mean that any further component (or likewise features, integers, steps and the like) can / may be present. The term "consisting” of means that no further component (or likewise features, integers, steps and the like) is present.

[0168] The term "method" refers to manners, means, techniques and procedures for accomplishing a given task including, but not limited to, those manners, means, techniques and procedures either known to, or readily developed from known manners, means, techniques and procedures by practitioners of the chemical, biological and biophysical arts.

[0169] As used herein, the term "about" refers to ±10% of the indicated numerical value, and in particular to ±5% of the indicated numerical value. Whenever the term "about" is used, a specific reference to the exact numerical value indicated is also included. If the term "about" is used in connection with a parameter that is quantified in integers, such as the number of nucleotides in a given nucleic acid, the numbers corresponding to ±10% or ±5% of the indicated numerical value are to be rounded to the nearest integer. For example, the expression "about 25 amino acids" refers to the range of 23 to 28 amino acids, in particular the range of 24 to 26 amino acids, and preferably refers to the specific value of 25 amino acids.

[0170] The present invention is further described by reference to the following non-limiting figures.

[0171] Figure 1 A shows a typical ADM-Gly dose / signal curve.

[0172] Figure IB shows a normal distribution of ADM-Gly from n=157 self-reported healthy individuals from Example 1.

[0173] Figure 2 shows analyses of bio-ADM (black bars) and ADM-Gly (grey bars) from human endotoxaemia model samples as a reaction to EPS induced systemic inflammation from Example 2.

[0174] Figure 3 shows analyses of ADM-Gly / bio-ADM ratio (ADM ratio) from human endotoxemia model samples as a reaction to LPS induced systemic inflammation from Example 2.

[0175] Figure 4 shows analyses of total ADM (sum of molar concentrations of ADM-Gly and bio-ADM) from human endotoxaemia model samples as a reaction to LPS induced systemic inflammation from Example 2.

[0176] Figure 5 shows analyses of ADM-Gly / total-ADM ratio (total-ADM ratio) from human endotoxemia model samples as a reaction to LPS induced systemic inflammation from Example 2.

[0177] Figure 6 shows analyses of bio-ADM (black bars) and ADM-Gly (grey bars) from an induced swine model of sepsis from Example 3.

[0178] Figure 7 shows analyses of ADM-Gly / bio-ADM ratio (ADM ratio) from an induced swine model of sepsis from Example 3. Figure 8 shows analyses of total ADM (sum of molar concentrations of ADM-Gly and bio-ADM) and from an induced swine model of sepsis from Example 3.

[0179] Figure 9 shows analyses of ADM-Gly / total-ADM ratio (total-ADM ratio) from an induced swine model of sepsis from Example 3.

[0180] The present invention is further described by reference to the following non-limiting examples.

[0181] Examples

[0182] Example 1: ADM Measurements

[0183] Quantification of bio-ADM was conducted using the sphingotest bio-ADM assay as described elsewhere46. ADM-Gly was quantified as based on Weber et al.46for bioactive ADM with the following modifications: the tracer-antibody used for ADM-Gly detection, labelled with MACN-acridinium-NHS, was directed to the C -terminal glycine of ADM-Gly. The assay was calibrated with synthetic ADM-Gly. The limit of detection (LOD) was 9 pg / mL of ADM-Gly. The antibody directed to the C -terminal glycine of ADM had no cross-reactivity with bio-ADM. A typical standard curve is shown in Figure 1A. A normal distribution of ADM-Gly from n=157 self-reported healthy individuals is shown in Figure IB (Median: 21.1 pg / mL; 25% Percentile: 14.1 pg / mL; 75% Percentile: 28.15 pg / mL. The data did not pass the D’Agostino & Pearson normality test). Using ADM-Gly values and bio-ADM values, the ratio of both peptides was calculated using formula (1). ADM-Gly frequency distribution from n=157 selfreported healthy subjects.

[0184] Formula 1 :

[0185] [ADMGly]: Concentration of ADM-Gly in pg / mL; MWbio: the molecular weight of bio-ADM (6028.9 pg / pmol); MWGly: the molecular weight of ADM-Gly (6086.9 pg / pmol); [bio ADM]: Concentration of bio-ADM in pg / mL.

[0186] Example 2: ADM A nalyses in a human endotoxaemia model.

[0187] In the present invention ADM-Gly and bio-ADM levels were determined using human EDTA plasma samples (n=28 per timepoint) from a human endotoxaemia model. Therefore, LPS (lipopolysaccharide, E. coli type 0113) was infused for 3h hours in healthy volunteers at an infusion rate of lng*kg-l*h-l to induce systemic inflammation. Blood samples were drawn directly before LPS Infusion (t=0) and at several timepoints during the infusion. The LPS induced model is a well-established and characterized model of systemic inflammation which is well documented in literature47,48.

[0188] As expected, and previously reported33, a slight elevation of bio-ADM was observed 180 and 240 minutes after begin of infusion. A relatively higher elevation of ADM-Gly (by factor of approx. 2.5) when compared to bio-ADM was observed at t=l 80 min and t=250 min after begin of LPS infusion (Figure 2).

[0189] Surprisingly and unexpectedly, ADM-Gly elevation as a reaction to LPS induced systemic inflammation precedes elevation of bio-ADM by several hours. As shown in Figure 2, a significant elevation of ADM- Gly was surprisingly observed already 90 minutes after begin of LPS infusion, further increasing at t=120 min and reaching its peak at t=l 80 and t=240 minutes.

[0190] The data clearly show that elevation of ADM-Gly precedes elevation of bio-ADM by at least 90 Minutes. Thus, ADM-Gly, contrary to bio-ADM, is clearly an early marker suitable for prediction of systemic inflammation.

[0191] The ADM ratio, as calculated using formula (1), clearly shows that at t=90 min after LPS infusion, the ratio is significantly elevated indicating a shift in the ADM homeostasis towards ADM-Gly (Figure 3). Thus, not only ADM-Gly, but also the ADM ratio is an early marker suitable for prediction of systemic inflammation. The ADM ratio further increases after 120 and 180 minutes as a result of the fast ADM- Gly and a slow bio-ADM-increase.

[0192] Formula 2:

[0193] [ADMGly]: Concentration of ADM-Gly in pg / mL; MWbio: the molecular weight of bio-ADM (6028.9 pg / pmol); MWGly: the molecular weight of ADM-Gly (6086.9 pg / pmol); [bioADM]: Concentration of bio-ADM in pg / mL.

[0194] Surprisingly and unexpectedly, the elevation of total ADM irrespective of its C -terminal amidation status as a reaction to LPS induced systemic inflammation, calculated as the molar sum of ADM-Gly and bio-ADM using formula 2, showed a significant increase after 180 minutes as shown in Figure 4. The data clearly show that analysis of total ADM irrespective of its C-terminal amidation status may be an early marker of systemic inflammation, while ADM-Gly alone as depicted in figure 2 is superior in comparison to total ADM as a biomarker of systemic inflammation. The ratio of ADM-Gly to total ADM as shown in figure 5 is elevated already after t=90 minutes after LPS infusion, this elevation is statistically not significant, while elevation of the total ADM Ratio becomes significant 120 minutes after LPS infusion. The data show that the total ADM ratio may be an early marker of systemic inflammation, while the ADM-Gly / bio-ADM ratio as shown in Figure 3 is superior to the ratio total ADM ratio.

[0195] Figure 2 shows analyses of bio-ADM (black bars) and ADM-Gly (grey bars) from human endotoxaemia model samples as a reaction to LPS induced systemic inflammation. The data for bio-ADM and ADM- Gly was normalized to baseline levels (t=0, set as 100%, indicated by dotted line), respectively. Significance was tested with non-normalized raw-data using non-parametric, two-tailed Mann- Whitney test. ****: p<0.0001; **: p=0.0075 - 0.0023; *: p=0.046 - 0.024

[0196] Figure 3 shows analyses of ADM-Gly / bio-ADM ratio (ADM ratio) from human endotoxemia model samples as a reaction to LPS induced systemic inflammation. The data for ADM ratio was normalized to baseline levels (t=0, set as 100%, indicated by dotted line). Significance was tested with nonnormalized raw-data using non-parametric, two-tailed Mann- Whitney test. ***: p=0.0001; **: p=0.0042; *: p=0.028

[0197] Figure 4 shows analyses of total ADM calculated using Formula 2, from human endotoxaemia model samples as a reaction to LPS induced systemic inflammation. The data for total ADM was normalized to baseline levels (t=0, set as 100%, indicated by dotted line). Significance was tested with nonnormalized raw-data using non-parametric, two-tailed Mann-Whitney test. **: p=0.008 - 0.0041; n.s.: not significant

[0198] Figure 5 shows analyses of ADM-Gly / total-ADM ratio (total ADM ratio) from human endotoxemia model samples as a reaction to LPS induced systemic inflammation. The data for total ADM ratio was normalized to baseline levels (t=0, set as 100%, indicated by dotted line). Significance was tested with non-normalized raw-data using non-parametric, two-tailed Mann- Whitney test. ****; pcO.OOOl; ***: p=0.0001; **: p=0.0012; n.s.: not significant

[0199] Example 3; ADM Analyses in a porcine model of septic shock

[0200] In the present invention ADM-Gly and bio-ADM levels were determined using EDTA plasma samples from a porcine model of septic shock. The beneficiaries of this animal model are as follows:

[0201] This model allows the analysis of blood samples directly after induction of the model and prior to the development of sepsis and shock, what is difficult to achieve with human clinical samples, since ICU patients usually are septic at admission to ICU. The porcine animal model shows high similarity to humans in terms of cardio-physiology, immunology and the gastrointestinal system49’50.

[0202] The laboratory model was induced in anesthetized animals (n=8) after drawing of baseline samples by intraperitoneal application of 3g / kg of autologous feces. During the study period several parameters, such as heart rate, cardiac output, change of Lactate, pulse pressure variation, venous oxygen saturation, central venous-to-arterial CO2 gap, changes in creatinine and haemoglobin as well as blood-pressure and mean arterial pressure were monitored. The animals were considered as septic when: lactate increased over the level of 2 mmol / L, heart rate increased to over 120 bpm, cardiac output decreased below 6 L / min, venous oxygen saturation decreased below 60%. The animals were considered to be in the state of septic shock when mean arterial pressure dropped below 55 mmHg, which occurred on average 1.1 hours after diagnosis of sepsis.

[0203] As expected, levels of bio-ADM and ADM-Gly were elevated at timepoint of sepsis by the factor 3.5 and 6 for bio-ADM and ADM-Gly, respectively, when compared to baseline. A further increase of both markers was observed at the timepoint of septic shock (Figure 6).

[0204] Surprisingly and unexpectedly, ADM-Gly elevation as a reaction to feces induced peritonitis precedes elevation of bio-ADM by several hours. As shown in Figure 6, a significant elevation of plasma ADM- Gly concentration was observed already up to 4 hours before animals entered the state of sepsis. ADM- Gly concentration were further elevated 1-2 hours before diagnosis of sepsis, reaching its peak at timepoint of septic shock, while bio-ADM levels remained unchanged before the diagnosis of sepsis. Plasma levels of ADM-Gly clearly show a significant and surprisingly early elevation at least four hours before diagnosis of sepsis. Therefore, the early increase of ADM-Gly is clearly a predictive marker for the entry into sepsis and septic shock at least 4 hours up front.

[0205] Furthermore, the ADM ratio, as calculated using formula (1), clearly shows a significant change 4 hours before sepsis (Figure 7). The ratio is significantly elevated indicating a shift in the ADM homeostasis towards ADM-Gly (Figure 7). Thus, not only ADM-Gly, but also the ADM ratio is an early marker suitable for prediction of sepsis and septic shock. The ADM ratio reached its peak 1 -2 hours before sepsis, and is further declining in sepsis and shock as a result of fast increase of bio-ADM.

[0206] Unexpectedly, the elevation of total ADM irrespective of its C -terminal amidation status as a reaction to feces induced peritonitis, calculated as the molar sum of ADM-Gly and bio-ADM using formula 2, showed a significant increase comparable to the increase of ADM-Gly after 180 minutes as shown in Figure 8. The data clearly show that analysis of total ADM irrespective of its C -terminal amidation status may be an early marker of systemic inflammation. The ratio of ADM-Gly / total ADM as shown in figure 9 is elevated 4 hours prior to sepsis indicating a shift in the total produced ADM towards ADM- Gly. The data show that the total ADM ratio may be an early marker of systemic inflammation, comparable to the ADM-Gly / bio-ADM ratio as shown in Figure 3.

[0207] Figure 6 shows analyses of bio-ADM (black bars) and ADM-Gly (grey bars) from an induced swine model of sepsis. The data for bio-ADM and ADM-Gly was normalized to baseline levels (baseline set as 100%), respectively. Significance was tested with non-normalized raw-data using non-parametric, two-tailed Mann-Whitney test. ***: p=0.0002; **: p=0.004; *: p=0.014.

[0208] Figure 7 shows analyses of ADM-Gly / bio-ADM ratio (ADM ratio) from an induced swine model of sepsis. The data for ADM ratio was normalized to baseline levels (baseline set as 100%). Significance was tested with non-normalized raw-data using non-parametric, two-tailed Mann- Whitney test. **: p=0.0047; *: p=0.0016 - 0.027.

[0209] Figure 8 shows analyses of total ADM from an induced swine model of sepsis. The data for total ADM was normalized to baseline levels (baseline set as 100%). Significance was tested with non-normalized raw-data using non-parametric, two-tailed Mann- Whitney test. ***: p=0.0002; **: p=0.004; *: p=0.014.

[0210] Figure 9 shows analyses of ADM-Gly / total-ADM ratio (total ADM ratio) from an induced swine model of sepsis. The data for total ADM ratio was normalized to baseline levels (baseline set as 100%). Significance was tested with non-normalized raw-data using non-parametric, two-tailed Mann- Whitney test. **: p=0.0030; *: p=0.0140 - 0.0162.

[0211] All references cited herein are fully incorporated by reference.

[0212] 1. De La Rica AS, Gilsanz F, Maseda E. Epidemiologic trends of sepsis in western countries. Ann Tr ansi Med. 2016;4(17):4-9. doi: 10.21037 / atm.2016.08.59

[0213] 2. Kaukonen KM, Bailey M, Suzuki S, Pilcher D, Bellomo R. Mortality related to severe sepsis and septic shock among critically ill patients in Australia and New Zealand, 2000-2012. JAMA - J Am Med Assoc. 2014;311(13): 1308-1316. doi:10.1001 / jama.2014.2637

[0214] 3. Sugo S, Minamino N, Kangawa K, et al. Endothelial Cells Actively Synthesize and Secrete Adrenomedullin. Biochem Biophys Res Cornmun. 1994;201(3):l 160-1166. doi: 10.1006 / bbrc.1994.1827

[0215] 4. Eto T. A review of the biological properties and clinical implications of adrenomedullin and proadrenomedullin N-terminal 20 peptide (PAMP), hypotensive and vasodilating peptides. Peptides. 2001;22:1693-1711.

[0216] 5. Eto T, Kato J, Kitamura K. Regulation of production and secretion of adrenomedullin in the cardiovascular system. RegulPept. 2003;l 12(l-3):61-69. doi: 10.1016 / S0167-0115(03)00023-5

[0217] 6. Lan Z, Zongyin QIU, Chi YE, et al. Vasoactive effects of adrenotensin and its interactions with adrenomedullin. Chin Med J (Engl). 2023;l 13(03):269-271. doi:10.3760 / cma.j.issn.0366- 6999.2000.03.115 7. Hashida S, Kitamura K, Nagatomo Y, et al. Development of an ultrasensitive enzyme immunoassay for human proadrenomedullin N -terminal 20 peptide and direct measurement of two molecular forms of P AMP in plasma from healthy subjects and patients with cardiovascular disease. Clin Biochem. 2004;37(l): 14-21. doi: 10.1016 / J.CLINBIOCHEM.2003.09.007

[0218] 8. Hinson IP, Kapas S, Smith DM. Adrenomedullin, a multifunctional regulatory peptide. Endocr Rev. 2000;21(2):138-167. doi: 10.1210 / er.21.2.138

[0219] 9. Kuwasako K, Kitamura K, Kangawa K, Ishiyama Y, Kato J, Eto T. Increased plasma proadrenomedullin N-terminal 20 peptide in patients with essential hypertension. Ann Clin Biochem. 1999;36(5):622-628. doi:10.1177 / 000456329903600510

[0220] 10. Tsuruda T, Kato J, Kitamura K, et al. Secretion of proadrenomedullin N-terminal 20 peptide from cultured neonatal rat cardiac cells. Life Sci. 2001;69(2):239-245. doi:10.1016 / S0024- 3205(01)01119-5

[0221] 11. Matsui E, Kitamura K, Yoshida M, et al. Biosynthesis and secretion of adrenomedullin and proadrenomedullin N-terminal 20 peptide in a rat model of endotoxin shock. Hypertens Res. 2001;24(5):543-549. doi: 10.1291 / hypres.24.543

[0222] 12. Eto T, Washimine H, Kato J, Kitamura K, Yamamoto Y. Adrenomedullin and proadrenomedullin N-terminal 20 peptide in impaired renal function. Kidney Int. 1996;49.

[0223] 13. Etoh T, Kato J, Takenaga M, et al. Differential hormonal profiles of adrenomedullin and proadrenomedullin N-terminal 20 peptide in patients with heart failure and effect of treatment on their plasma levels. Clin Cardiol. 1999;22(2):113-117. doi:10.1002 / clc.4960220211

[0224] 14. Kinoshita H, Fujimoto S, Kitamura K, et al. Measurements of Plasma and Urine. 1999;34( 1 ) : 114- 119.

[0225] 15. Kiriyama Y, Nochi H. Procalcitonin and Adrenomedullin in Infectious Diseases. Microbiol Res (Pavia). 2023;14(l):190-204. doi:10.3390 / microbiolresl4010016

[0226] 16. Rehfeld JF. The long goodbye to glycine-extended peptide hormones. Int J Endocr Oncol. 2017;4(4): 147-149. doi: 10.2217 / ij e-2017-0015

[0227] 17. Kitamura K, Kato J, Kawamoto M, et al. The intermediate form of glycine -extended adrenomedullin is the major circulating molecular form in human plasma. Biochem Biophys Res Commun. 1998;244(2):551-555. doi: 10.1006 / bbrc.1998.8310

[0228] 18. Yamaga J, Hashida S, Kitamura K, et al. Direct measurement of glycine -extended adrenomedullin in plasma and tissue using an ultrasensitive immune complex transfer enzyme immunoassay in rats. Hypertens Res. 2003;26(SUPPL.):45-53. doi:10.1291 / hypres.26.S45

[0229] 19. Otao G, Maruta T, Yonaha T, et al. The usefulness of plasma levels of mature and total adrenomedullin as biomarkers indicating the magnitude of surgical stress responses: A singlecenter, prospective, observational study. J Clin Transl Res. 2021;7(3):302.

[0230] 20. Yonaha T, Maruta T, Otao G, et al. The diagnostic and prognostic value of mature and total adrenomedullin for sepsis: a prospective observational study. Anaesthesiol Intensive Ther. 2021;53(5):411-417.

[0231] 21. Nishikimi T, Shibasaki I, lida H, et al. Molecular forms of adrenomedullin in pericardial fluid and plasma in patients with ischaemic heart disease. Clin Set. 2002;102(6):669-677. doi:10.1042 / CS20010236

[0232] 22. Nishikimi T, Horio T, Kohmoto Y, et al. Molecular forms of plasma and urinary adrenomedullin in normal, essential hypertension and chronic renal failure. J Hypertens. 2001;19(4):765-773. doi : 10.1097 / 00004872-200104000-00014

[0233] 23. Nomura I, Kato J, Tokashiki M, Kitamura K. Increased plasma levels of the mature and intermediate forms of adrenomedullin in obesity. Regul Pept. 2009; 158(1-3): 127-131. doi:10.1016 / J.REGPEP.2009.08.003

[0234] 24. Sugo S, Minamino N, Shoji H, et al. Interleukin- 1, Tumor Necrosis Factor and Lipopolysaccharide Additively Stimulate Production of Adrenomedullin in Vascular Smooth Muscle Cells. Biochem Biophys Res Commun. 1995;207(l):25-32. doi:10.1006 / BBRC.1995.1148

[0235] 25. Fischer JP, Els-Heindl S, Beck-Sickinger AG. Adrenomedullin - Current perspective on a peptide hormone with significant therapeutic potential. Peptides. 2020; 131 :170347. doi : 10.1016 / J.PEPTIDES .2020.170347

[0236] 26. Hirayama N. Study patients , cardiac catheterization and blood sampling of antisera for AM for mAM and total AM ( MM ). Life Sci 1999;64(26)2505-9. 1999;64(26):0-4.

[0237] 27. Yu CM, Cheung BMY, Leung R, Wang Q, Lai WH, Lau CP. Increase in plasma adrenomedullin in patients with heart failure characterised by diastolic dysfunction. Heart. 2001 ;86(2): 155- 160.

[0238] 28. Ehlenz K, Koch B, Preuss P, Simon B, Koop I, Lang RE. High levels of circulating adrenomedullin in severe illness: correlation with C-reactive protein and evidence against the adrenal medulla as site of origin. Exp Clin Endocrinol diabetes. 1997;105(03): 156-162.

[0239] 29. Tomoda Y, Isumi Y, Katafuchi T, Minamino N. Regulation of adrenomedullin secretion from cultured cells. Peptides. 2001;22(l 1):1783-1794. doi: 10.1016 / S0196-9781(01)00519-8

[0240] 30. Ueda S, Nishio K, Minamino N, et al. Increased plasma levels of adrenomedullin in patients with systemic inflammatory response syndrome. Am J Respir Crit Care Med. 1999; 160(1 ): 132- 136. doi: 10.1164 / ajrccm.160.1.9810006

[0241] 31. Wang P. Andrenomedullin and cardiovascular responses in sepsis. Peptides. 2001 ;22(11): 1835- 1840.

[0242] 32. Vila G, Resl M, Stelzeneder D, et al. Plasma NT -proBNP increases in response to LPS administration in healthy mmeenn.. J Appl Physiol. 2008; 105(6): 1741-1745. doi:10.1152 / japplphysiol.90442.2008

[0243] 33. Geven C, van Lier D, Blet A, et al. Safety, tolerability and pharmacokinetics / pharmacodynamics of the adrenomedullin antibody adrecizumab in a first-in-human study and during experimental human endotoxaemia in healthy subjects. Br J Clin Pharmacol. 2018;84(9):2129-2141. doi:10.1111 / bcp.13655

[0244] 34. Schuetz P, Hausfater P, Amin D, et al. Biomarkers from distinct biological pathways improve early risk stratification in medical emergency patients: The multinational, prospective, observational TRIAGE study. Crit Care. 2015; 19(1): 1-11. doi:10.1186 / sl3054-015-1098-z

[0245] 35. Enguix-Armada A, Escobar-Conesa R, La Torre AG-D, De La Torre-Prados MV. Usefulness of several biomarkers in the management of septic patients: C -reactive protein, procalcitonin, presepsin and mid-regional pro-adrenomedullin. 2016;54(l): 163-168. doi:doi:10.1515 / cclm- 2015-0243

[0246] 36. Andaluz-Ojeda D, Nguyen HB, Meunier-Beillard N, et al. Superior accuracy of mid-regional proadrenomedullin for mortality prediction in sepsis with varying levels of illness severity. Ann Intensive Care. 2017;7(l): 1-8.

[0247] 37. Andres C, Andaluz-Ojeda D, Cicuendez R, et al. MR- proADM to detect specific types of organ failure in infection. Eur J Clin Invest. 2020;50(6):0-2. doi: 10.1111 / eci.13246

[0248] 38. Maisel A, Mueller C, Nowak R, et al. Mid-Region Pro-Hormone Markers for Diagnosis and Prognosis in Acute Dyspnea. Results From the BACH (Biomarkers in Acute Heart Failure) Trial. J Am Coll Cardiol. 2010;55(19):2062-2076. doi: 10.1016 / j.jacc.2010.02.025

[0249] 39. Von Haehling S, Filippatos GS, Papassotiriou J, et al. Mid-regional pro-adrenomedullin as a novel predictor of mortality in patients with chronic heart failure. Eur J Heart Fail. 2010;12(5):484-491.

[0250] 40. Adlbrecht C, Hulsmann M, Strunk G, et al. Prognostic value of plasma midregional proadrenomedullin and C-terminal-pro-endothelin- 1 in chronic heart failure outpatients. Eur J Heart Fail. 2009; 11(4):361-366.

[0251] 41. Singer M, Deutschman CS, Seymour CW, et al. The Third International Consensus Definitions for Sepsis and Septic Shock (Sepsis-3). JAMA. 2016;315(8):801 -810. doi:10.1001 / jama.2016.0287

[0252] 42. Bone RC, Balk RA, Cerra FB, et al. Definitions for Sepsis and Organ Failure and Guidelines for the Use of Innovative Therapies i inn Sepsis. Chest. 1992;101(6): 1644-1655. doi:10.1378 / chest.l01.6.1644

[0253] 43. Lambden S, Laterre PF, Levy MM, Francois B. The SOFA score — development, utility and challenges of accurate assessment in clinical trials. Crit Care. 2019;23(l):374. doi:10.1186 / sl3054-019-2663-7

[0254] 44. Angus DC, Seymour CW, Coopersmith CM, et al. A Framework for the Development and Interpretation of Different Sepsis Definitions and Clinical Criteria. Crit Care Med. 2016;44(3). https: / / joumals.lww.eom / ccmjoumal / Fulltext / 2016 / 03000 / A_Framework_for_the_Developmen t and lnterpretation.35. aspx.

[0255] 45. Seymour CW, Liu VX, Iwashyna TJ, et al. Assessment of clinical criteria for sepsis for the third international consensus definitions for sepsis and septic shock (sepsis-3). JAMA - J Am Med Assoc. 2016;315(8):762-774. doi: 10.1001 / jama.2016.0288

[0256] 46. Weber, Julia et al. 2017. “Sandwich Immunoassay for Bioactive Plasma Adrenomedullin.” The

[0257] Journal of Applied Laboratory Medicine: An AACC Publication 2(2): 222-33.

[0258] 47. Andreasen, A. S. et al. 2008. “Experimental Human Endotoxemia as a Model of Systemic

[0259] Inflammation.” Biochimie 159: 99 106.

[0260] 48. Lier, Dirk Van, Christopher Geven, Guus P Leijte, and Peter Pickkers. 2018. “Biochimie

[0261] Experimental Human Endotoxemia as a Model of Systemic in Fl Animation.” Biochimie: 1-8. https: / / doi.Org / 10.1016 / j.biochi.2018.06.014.

[0262] 49. Goldfarb, Roy D et al. 1996. “A PORCINE MODEL OF PERITONITIS AND BACTEREMIA

[0263] SIMULATES HUMAN SEPTIC SHOCK.” Shock 6(6). https: / / joumals.lww.eom / shockjoumal / Fulltext / 1996 / 12000 / A_PORCINE_MODEL_OF_PERI TONITIS_AND_BACTEREMIA.9.aspx.

[0264] 50. Patterson, Jannine K, Xin Gen Lei, and Dennis D Miller. 2008. “The Pig as an Experimental Model for Elucidating the Mechanisms Governing Dietary Influence on Mineral Absorption.” Experimental Biology and Medicine 233(6): 651 64. https: / / doi.org / 10.3181 / 0709-MR-262.

[0265] SEQUENCES

[0266] SEQ ID No: 1 - pre-proADM (aa 1-185)

[0267] 10 20 30 40 50

[0268] MKLVSVALMY LGSLAFLGAD TARLDVASEF RKKWNKWALS RGKRELRMSS

[0269] 60 70 80 90 100

[0270] SYPTGLADVK AGPAQTLIRP QDMKGASRSP EDSSPDAARI RVKRYRQSMN

[0271] 110 120 130 140 150

[0272] NFQGLRSFGC RFGTCTVQKL AHQIYQFTDK DKDNVAPRSK ISPQGYGRRR

[0273] 160 170 180

[0274] RRSLPEAGPG RTLVSSKPQA HGAPAPPSGS APHFL

[0275] SEQ ID No: 2 - PAMP-Gly (aa 22-42 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0276] ARLDVASEFR KKWNKWALSR G

[0277] SEQ ID No: 3 - CT-proADM (aa 148-185 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0278] RRRRRSLPEA GPGRTLVSSK PQAHGAPAPP SGSAPHFL

[0279] SEQ ID No: 4 - ADM-Gly (aa 95-147 of pre-proADM SEQ ID No. 1)

[0280] 10 20 30 40 50

[0281] YRQSMNNFQG LRSFGCRFGT CTVQKLAHQI YQFTDKDKDN VAPRSKISPQ GYG

[0282] SEQ ID No: 5 - bio-ADM (aa 95-146 of pre-proADM SEQ ID No. 1)

[0283] 10 20 30 40 50

[0284] YRQSMNNFQG LRSFGCRFGT CTVQKLAHQI YQFTDKDKDN VAPRSKISPQ GY SEQ ID No: 6 - MR-proADM (aa 45-92 of pre-proADM SEQ ID No. 1)

[0285] 10 20 30 40 50

[0286] ELRMSSSYPT GLADVKAGPA QTLIRPQDMK GASRSPEDSS PDAARIRV

[0287] SEQ ID No: 7 - ADM-Gly 2-53 (aa 96-147 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0288] RQSMNNFQG LRSFGCRFGT CTVQKLAHQI YQFTDKDKDN VAPRSKISPQ GYG

[0289] SEQ ID No: 8 - ADM-Gly 8-53 (aa 102-147 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0290] FQG LRSFGCRFGT CTVQKLAHQI YQFTDKDKDN VAPRSKISPQ GYG

[0291] SEQ ID No: 9 - ADM-Gly 22-53 (aa 115-147 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0292] TVQKLAHQI YQFTDKDKDN VAPRSKISPQ GYG

[0293] SEQ ID No: 10 - ADM-Gly 27-53 (aa 120-147 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0294] AHQI YQFTDKDKDN VAPRSKISPQ GYG

[0295] SEQ ID No: 11 - ADM-Gly 33-53 (aa 126-147 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0296] FTDKDKDN VAPRSKI SPQ GYG

[0297] SEQ ID No: 12 - bio-ADM 2-52 (aa 96-146 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0298] RQSMNNFQG LRSFGCRFGT CTVQKLAHQI YQFTDKDKDN VAPRSKISPQ GY

[0299] SEQ ID No: 13 - bio-ADM 8-52 (aa 102-146 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0300] FQG LRSFGCRFGT CTVQKLAHQI YQFTDKDKDN VAPRSKISPQ GY

[0301] SEQ ID No: 14 - bio-ADM 22-52 (aa 115-146 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0302] TVQKLAHQI YQFTDKDKDN VAPRSKISPQ GY

[0303] SEQ ID No: 15 - bio-ADM 27-52 (aa 120-146 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0304] AHQI YQFTDKDKDN VAPRSKISPQ GY

[0305] SEQ ID No: 16 - bio-ADM 33-52 (aa 126-146 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0306] FTDKDKDN VAPRSKI SPQ GY

[0307] SEQ ID No: 18 - (aa 143-147 of pre-proADM SEQ ID No. 1) 10 20 30 40 50

[0308] PQGYG

[0309] SEQ ID No: 19 - (aa 143-146 of pre-proADM SEQ ID No. 1)

[0310] 10 20 30 40 50

[0311] PQGY

[0312] SEQ ID No: 20 - PAMP-NH2 (aa 22-41 of pre-proADM SEQ ID No. 1)

[0313] 10 20 30 40 50

[0314] ARLDVASEFR KKWNKWALSR

Claims

Claims1. A method for a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or, b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject, the method comprising:Determining the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the sum of the levels of ADM-Gly according to SEQ ID No. 4 and ADM-NH2 according SEQ ID No. 5 ("total-ADM") in a bodily fluid obtained from said subject; andA) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with the extent of sepsis and / or septic shock in said subject or diagnosing sepsis and / or septic shock in said subject, wherein an elevated level above a certain threshold is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or,B) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with risk for getting sepsis and / or septic shock, wherein an elevated level above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock, orC) Correlating said level of said biomarker or fragments thereof of at least five amino acids or said total-ADM with need of therapy or intervention of sepsis and / or septic shock, wherein an elevated level above a certain threshold is indicative for the need of therapy or intervention of sepsis and / or septic shock.

2. The method of claim 1 for a) diagnosing sepsis and / or septic shock or assessing or monitoring the extent of sepsis and / or septic shock in a subject or,b) early assessing the risk of getting sepsis and / or septic shock in a subject or, c) predicting or determining or monitoring need of therapy or intervention of sepsis and / or septic shock in a subject, the method comprising:Determining the ratio of the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM to a level of ADM-NH2according SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; andA) Correlating said ratio with the extent of sepsis and / or septic shock in said subject or diagnosing sepsis and / or septic shock in said subject, wherein an elevated ratio above a certain threshold is indicative of sepsis and / or septic shock or the extent of sepsis and / or septic shock or,B) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock, orC) Correlating said ratio with need of therapy or intervention of sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is indicative for the need of therapy or intervention of sepsis and / or septic shock.

3. The method of claim 1 or 2 for early assessing the risk of getting sepsis and / or septic shock in a subject, the method comprising:Determining the ratio of the level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids in a bodily fluid obtained from said subject; andB) Correlating said ratio with risk for getting sepsis and / or septic shock, wherein an elevated ratio above a certain threshold is predictive for an enhanced risk of getting sepsis and / or septic shock.

4. The method according to any of claims 1 to 3, wherein the early predicting is the prediction with 48 hours, 24 hours, 12 hours, 6 hours, 4 hours, preferably 2 hours advance the occurrence of sepsis and / or septic shock.

5. The method according to any of claim 1 to 4 , wherein said biomarker is ADM-Gly according to SEQ ID No. 4 and said level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids or said ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 and fragments thereof of at least five amino acids is determined by using a binder to ADM-Gly according to SEQ ID No.: 4 or fragments thereof of at least five amino acids.

6. The method according to claim 5, wherein the binder is selected from the group comprising an antibody, an antibody fragment or a non-Ig-Scaffold binding to ADM-Gly according to SEQ ID No. 4 or fragments thereof of at least five amino acids.

7. The method according to claim 5 or 6, wherein said threshold for a level of ADM-Gly and fragments thereof of at least five amino acids is 20 pg / mL in a bodily fluid, preferably 25 pg / ml, more preferably 30 pg / mL, more preferably 40 pg / mL, more preferably 50 pg / mL, whereinthe level of ADM-NH2 according to SEQ ID No. 5 or fragments thereof of at least five amino acids is in the range of from 5 to 30 pg / mL.

8. The method according to any of claims 5 to 7, wherein said threshold for the ratio of a level of ADM-Gly according to SEQ ID No. 4 and fragments thereof of at least five amino acids to a level of ADM-NH2 according to SEQ ID No. 5 or fragments thereof of at least five amino acids in a bodily fluid is 1.5, preferably 1.6, more preferably 1.7, more preferably 1.8, more preferably 1.9, more preferably 2.

9. The method according to any of claims 1 to 8, wherein said fragments of ADM-Gly according to SEQ ID No. 4 of at least five amino acids comprise a fragment according to SEQ ID No. 18.

10. The method according to any of claims 2 to 9, wherein said fragments of ADM-NH2 according to SEQ ID No. 5 of at least five amino acids comprise a fragment according to SEQ ID No. 19.

11. The method according to any of claims 1 to 10, wherein said determination of said level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No.4, PAMP according to SEQ ID No. 2, MR-proADM according to SEQ ID No. 6 and CT-proADMaccording to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total-ADM or said ratio is performed more than once in said subject.

12. The method according to any of claims 1 to 11 in order to stratify said subjects into sepsis and / or septic shock grade groups.

13. The method according to any of claims 1 to 12, wherein said therapy is selected from the group comprising administration of antibiotics, vasopressor therapy, immunosuppressive therapy, renal therapy including administration of Alkaline Phosphatase.

14. The method according to any of claims 1 to 13, wherein said therapy comprises administering anti-ADM antibodies, preferably N-terminal anti-ADM antibodies, and / or PAM (peptidylglycine alpha-amidating monooxygenase) as therapeutic agent.

15. A kit for carrying out the method according to any one of claims 1 to 14, wherein said kit comprises detection reagents for determining said level of a biomarker selected from the group consisting of ADM-Gly according to SEQ ID No. 4, PAMP according to SEQ ID No. 2, MR- proADM according to SEQ ID No. 6 and CT-proADM according to SEQ ID No. 3 or fragments thereof of at least five amino acids or the total ADM or said ratio in said sample of said subject.

Images