IGFBP7 Ratio Of HFpEF

Abstract

The present invention relates to a method for distinguishing between ejection fraction retention type heart failure (HFpEF) and ejection fraction reduction type heart failure (HFrEF) in a subject suffering from heart failure, the method comprising the steps of determining the amounts of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), BNP-type peptide and optionally CRP (C-Reactive Protein) in a sample from the subject, the present invention relates to a method for distinguishing ejection fraction retention type heart failure (HFpEF) from ejection fraction reduction type heart failure (HFrEF) by using a BNP peptide, calculating (i) a ratio of the amount of the IGFBP7 to the amount of the BNP peptide or (ii) a ratio of a sum of the amounts of the IGFBP7 and the CRP to the amount of the BNP type peptide, comparing the calculated ratio with a reference ratio, and distinguishing ejection fraction retention type heart failure (HFpEF) from ejection fraction reduction type heart failure (HFrEF). The invention also relates to a method for diagnosing HFpEF.

Description

[0001] This application is a divisional application of the invention patent application with the application date of May 22, 2020, application number 202080038317.8, and invention name “IGFBP7 ratio of HFpEF”. Technical Field

[0002] The present invention relates to the field of cardiac disease. Specifically, the present invention relates to a method for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in a subject suffering from heart failure. The present invention also relates to a method for diagnosing HFpEF. Background Art

[0003] The use of biomarkers in the care of patients with heart failure (HF) has expanded significantly. Natriuretic peptides (NPs), including B-type natriuretic peptide (BNP) and its amino-terminal cleavage equivalent (NT-proBNP), are now widely used in the diagnosis, prognosis, and management of affected patients. Following BNP and NT-proBNP, a wide range of new biomarkers are being investigated, each with the potential to complement the assessment of patients with the complex pathophysiology of HF. In this regard, considerable effort has been invested in better understanding the mechanistic links between cardiac biomarker concentrations and the underlying cardiovascular pathophysiological processes that trigger them.

[0004] Many prior art references describe the use of marker combinations in the assessment of heart failure, such as those associated with heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF). For example, Sanders et al. evaluated whether biomarkers reflecting pathophysiological pathways differed between HFpEF and HfrEF and whether the prognostic value of biomarkers differed in HFpEF versus HfrEF (Eur J Heart Fail. 2015 Oct;17(10):1006-14. doi:10.1002 / ejhf.414. Epub 2015 Oct 16). The study described that patients with HFpEF exhibited elevated levels of soluble interleukin-1 receptor-like 1, high-sensitivity C-reactive protein, and cystatin-C. In contrast, patients with HFrEF exhibited elevated levels of NT-proBNP, high-sensitivity troponin T, and hemoglobin.

[0005] Sinning et al. described the measurement of CRP, GDF-15, sST2, and NT-proBNP in patients with HFrEF and HFpEF (Int J Cardiol. 2017 Jan 15;227:272-277. doi:10.1016 / j.ijcard.2016.11.110. Epub 2016 Nov 9). According to Sinning, the index ((CRP+GDF-15s+sST2) / NT-proBNP) can distinguish HFpEF from HFrEF.

[0006] The insulin-like growth factor axis has previously been found to be a predictor of HF outcome (Watanabe et al., Insulin-like growth factor axis (insulin-like growth factor-i / insulin-like growth factor-binding protein-3) as a prognostic predictor of heart failure: Association with adiponectin, Eur J Heart Fail. 2010; 12: 1214-1222). IGFBP-7 (insulin-like growth factor binding protein 7) is a 30-kDa modular glycoprotein known to be secreted by endothelial cells, vascular smooth muscle cells, fibroblasts, and epithelial cells (Ono et al., Biochem Biophys Res Comm 202 (1994) 1490). IGFBP-7 has been described as being associated with cellular senescence, tissue aging, and obesity. IGFBP-7 has been proposed as a marker for diastolic dysfunction and HFpEF, a disease of the elderly and obese. A mechanistic link between IGFBP-7 and impaired diastolic function has been demonstrated in a mouse model (US 2018 / 0127752). Elevated circulating levels of IGFBP-7 have been described in the PARAMOUNT study to be associated with diastolic dysfunction (Januzzi et al., Circ Heart Fail. (2018) 11).

[0007] WO 2008 / 089994 discloses the use of IGFBP7 in the assessment of heart failure. In addition to IGFBP7, NTproBNP and CRP can also be measured.

[0008] WO 2015 / 144767 describes IGFBP7 as a marker for diastolic dysfunction. In Example 2.2, patients with HFpEF were analyzed. In this example, IGFBP7 was combined with, for example, osteopontin, troponin T, and NT-proBNP.

[0009] WO 2014 / 086833 describes IGFBP7 as a marker for selecting heart failure therapies.

[0010] The relationship between IGFBP7 and HFpEF and HFrEF has also been analyzed. For example, Hage et al. described that IGFBP-7 is lower in HFpEF than in HFrEF (Am J Cardiol. 2018 Jun 15;121(12):1558-1566. doi:10.1016 / j.amjcard.2018.02.041. Epub 2018 Mar 14).

[0011] The diagnosis of HFpEF is challenging due to the preserved ejection fraction. It is mainly based on symptoms that can be vague and excludes other causes, such as lung disease. In addition, biomarker-based diagnosis of HFpEF is cumbersome. To date, no biomarkers have been established in clinical routine to aid in the diagnosis of HFpEF. NTproBNP has shown a stronger association with systolic dysfunction than with diastolic dysfunction. It is only modestly increased in HFpEF. Although IGFBP-7 has shown a mechanistic link with diastolic function, IGFBP-7 has not been described to help identify patients with preserved ejection fraction.

[0012] As As reviewed, the presentation and pathophysiology of HFpEF are diverse, and its management remains challenging (Clin Res Cardiol. 2018 Jan;107(1):1-19. doi:10.1007 / s00392-017-1170-6. Epub 2017 Oct 10). Until now, no therapy has improved survival in patients with HFpEF. Therefore, the goals of treatment are to alleviate symptoms, improve quality of life, and reduce cardiac decompensation by controlling fluid retention and managing risk factors and comorbidities. For example, angiotensin-aldosterone inhibitors, diuretics, calcium channel blockers (CBBs), and beta-blockers are currently used to treat HFpEF. However, these drugs have not been shown to reduce mortality in large randomized controlled trials. Recently, new targets for the treatment of HFpEF have been identified, such as soluble guanylate cyclase stimulators, inorganic nitrates, the angiotensin receptor neprilysin inhibitor LCZ 696, and SGLT2 inhibitors. Patients with HFpEF may particularly benefit from these new treatments.

[0013] There is a great need for biomarker-based methods that can be used to reliably assess HFpEF and to differentiate between HFrEF and HFpEF.The technical problem of the present invention can be seen as providing means and methods that meet the above needs.

[0014] This technical problem is solved by the embodiments characterized in the claims and below. Summary of the Invention

[0015] Advantageously, in the present study, it was found that the ratio of IGFBP-7 to a BNP-type peptide (such as NT-proBNP) and the ratio of the sum of the amounts of IGFBP-7 + CRP to the amount of a BNP-type peptide improved the unique identification of HFpEF in heart failure patients compared to each marker alone. Thus, the above ratios allow for reliable differentiation between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in subjects with heart failure.

[0016] Therefore, the present invention relates to a method for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in a subject suffering from heart failure, the method comprising the steps of:

[0017] (a) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), a BNP-type peptide and optionally CRP (C-Reactive Protein) in a sample from said subject,

[0018] (b) calculating (i) the ratio of the amount of IGFBP7 to the amount of the BNP peptide or (ii) the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide,

[0019] (c) comparing the ratio calculated in step (b) with a reference ratio, and

[0020] (d) Differentiate between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF).

[0021] The present invention also relates to a method for diagnosing heart failure with preserved ejection fraction (HFpEF) in a subject suspected of having HFpEF, the method comprising the steps of:

[0022] (a) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), a BNP-type peptide and optionally CRP (C-Reactive Protein) in a sample from said subject,

[0023] (b) calculating (i) the ratio of the amount of IGFBP7 to the amount of the BNP peptide or (ii) the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide,

[0024] (c) comparing the ratio calculated in step (b) with a reference ratio, and

[0025] (d) Diagnosis of heart failure with preserved ejection fraction.

[0026] In one embodiment of the method of the present invention, the amounts of IGFBP7, the BNP-type peptide, and the CRP are determined in step (a), and the ratio of the sum of the amounts of IGFBP7 and the CRP to the amount of the BNP-type peptide is calculated in step (b). Thus, steps (a) and (b) are as follows:

[0027] (a) determining the amount of IGFBP7, BNP-type peptide and CRP in a sample from said subject,

[0028] (b) Calculating the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide.

[0029] Method of the present invention is preferably an isolated method, or especially an in vitro method. In addition, it can also include the step except the step clearly mentioned above. For example, other steps may relate to sample pretreatment or the result obtained by the method is evaluated. The method can be performed manually or assisted by automation. Preferably, steps (a), (b), (c) and / or (d) can be assisted in whole or in part by automation, for example, by the measurement in step (a) or the computer-implemented calculation in step (b) or the computer-implemented comparison in step (c) and / or based on the computer-implemented differentiation / diagnosis of the comparison in step (c) assisted by suitable robots and sensing devices.

[0030] As used herein, the term "diagnose" means assessing whether a subject referred to herein has HFpEF. In one embodiment, the subject is diagnosed as having HFpEF. In an alternative embodiment, the subject is diagnosed as not having HFpEF.

[0031] The diagnosis in step (d) or the distinction in step (d) is based on the results of the comparison step (c). Preferably, the diagnosis in step (d) is based on the results of the comparison step (c). However, it will be appreciated that the actual diagnosis of whether a subject suffers from AF may include further steps, such as confirmation of the diagnosis. Thus, the diagnosis referred to herein should allow for an assessment of the likelihood that a subject suffers from HFpEF. As can be seen from the above, the diagnosis of HFpEF is understood to be helpful for the diagnosis of HFpEF. Thus, the term "diagnosis" in the context of the present invention also includes helping a physician to assess whether a subject suffers from HFpEF.

[0032] The term "differentiation" as used herein means differentiating between HFpEF and HFrEF in a subject with the disease. The term as used herein preferably includes differentiating between HFpEF and HFrEF in a subject diagnosed with heart failure. Preferably, the differentiation in step (d) is based on the results of comparing step (c). In addition, the method of the present invention allows for the assessment of whether a subject with atrial fibrillation has paroxysmal atrial fibrillation or persistent atrial fibrillation. The actual differentiation may include further steps, such as confirmation of the differentiation. Therefore, the term "differentiation" in the context of the present invention also includes helping the physician to distinguish between HFpEF and HFrEF.

[0033] As will be understood by those skilled in the art, the assessments described herein (i.e., differentiation or diagnosis) are generally not intended to be correct for all (i.e., 100%) patients to be diagnosed / graded. In embodiments of the present invention, a statistically significant portion of patients (e.g., a cohort in a cohort study) can be identified. Those skilled in the art can readily determine whether a portion is statistically significant using various well-known statistical evaluation tools (e.g., determination of confidence intervals, p-value determination, Student t-test, Mann-Whitney test, etc.). See Dowdy and Wearden, Statistics for Research, John Wiley & Sons, New York 1983. Preferred confidence intervals are at least 90%, at least 95%, at least 97%, at least 98%, or at least 99%. The p-value is preferably 0.1, 0.05, 0.01, 0.005, or 0.0001. More preferably, at least 60%, at least 70%, at least 80%, or at least 90% of patients in a population can be correctly diagnosed / differentiated by the methods of the present invention.

[0034] As used herein, the term "subject" is preferably a mammal. Mammals include, but are not limited to, domesticated animals (e.g., cows, sheep, cats, dogs, and horses), primates (e.g., humans and non-human primates such as monkeys), rabbits, and rodents (e.g., mice and rats). Preferably, the subject is a human subject. The terms "subject," "patient," and "individual" are used interchangeably herein.

[0035] According to the method for distinguishing HFpEF from HFrEF, the patient to be tested should suffer from heart failure. The term "heart failure" is well known in the art. As used herein, the term preferably relates to impaired cardiac function accompanied by symptoms of heart failure as known to those skilled in the art. Thus, the patient preferably suffers from symptomatic heart failure.

[0036] The ACC / AHA classification is a classification of heart failure developed by the American College of Cardiology and the American Heart Association (see J. Am. Coll. Cardiol. 2001; 38; 2101-2113, updated in 2005, see J. Am. Coll. Cardiol. 2005; 46; e1-e82). Four stages are defined: A, B, C, and D. Stages A and B are not HF (heart failure), but are considered to help identify patients early before they develop "true" HF. Stage A and B patients are best defined as those who have risk factors for developing HF. For example, a patient with coronary artery disease, hypertension, or diabetes who has not yet demonstrated impaired left ventricular (LV) function, hypertrophy, or geometric chamber distortion would be considered stage A, while an asymptomatic patient who demonstrates LV hypertrophy (LVH, a phenomenon of thickening of the ventricular wall) and / or impaired LV function would be designated stage B. Stage C denotes patients with current or past symptoms of HF related to underlying structural heart disease (the majority of patients with HF), while stage D designates patients with true refractory HF.

[0037] In one embodiment of the method of the present invention, the patient to be tested should preferably be suffering from heart failure stage C or D according to the ACC / AHA classification (see above reference). At these stages, the patient exhibits symptoms of heart failure. Symptoms of heart failure are well known in the art and include dyspnea, fatigue, and fluid retention. Fluid retention can lead to pulmonary congestion and peripheral edema, with typical signs on physical examination being edema and rales. Therefore, the patient to be tested preferably exhibits symptoms of heart failure.

[0038] According to the present invention, a distinction should be made between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in subjects suffering from heart failure.

[0039] Subjects with HFrEF have heart failure with a reduced left ventricular ejection fraction (LVEF). The term "left ventricular ejection fraction" is well known in the art. Patients with a reduced LVEF preferably have an LVEF of less than 50%, more preferably less than 45%, and most preferably less than 40%. In addition, patients with an LVEF of less than 30% are envisioned.

[0040] A subject with HFpEF has heart failure with preserved LVEF. Thus, the term HFpEF preferably refers to heart failure with an LVEF equal to or greater than 50%. A patient with preserved LVEF may also have an LVEF greater than 55% or greater than 60%.

[0041] How to assess LVEF is well known in the art. In one embodiment, LVEF may be determined as described, for example, in McMurray et al. (European Heart Journal (2012) 33, 1787-1847, see, for example, pages 1800 et seq.).

[0042] In one embodiment of the method for diagnosing HFpEF, the patient to be tested is suspected of having heart failure, in particular HFpEF. The subject suspected of having heart failure preferably exhibits symptoms of heart failure. In an alternative embodiment of the method for diagnosing HFpEF, the patient to be tested is suspected of having heart failure. Thus, the patient should be known to have heart failure.

[0043] A patient to be tested according to the method for diagnosing HFpEF may have a normal ejection fraction, ie, a preserved LVEF.

[0044] According to the present invention, the subject to be tested may be elderly and / or overweight.

[0045] Preferably, the subject is older than 50 years, more preferably older than 60 years or most preferably older than 75 years. Alternatively or additionally, the subject has a 2 , especially more than 27.5kg / m 2 In addition, the subject may have a body mass index (BMI) greater than 30.0 kg / m 2 BMI.

[0046] The term "sample" refers to a sample of body fluid, a sample of isolated cells, or a sample from a tissue or organ. The sample of body fluid can be obtained by well-known technology and includes blood, blood plasma, serum, urine, lymph, sputum, ascites, or any other body secretions or samples thereof. Preferred body fluid samples are urine, blood, serum, or blood plasma. Tissue or organ samples can be obtained from any tissue or organ by, for example, biopsy. Isolated cells can be obtained from body fluid or tissue or organ by separation techniques such as centrifugation or cell sorting. For example, cells, tissues, or organ samples can be obtained from those cells, tissues, or organs that express or produce biomarkers. Samples can be freezing, fresh, fixed (for example, formalin fixed), centrifuged, and / or embedded (for example, paraffin embedded) samples, etc. Of course, various well-known collections can be performed on cell samples before the amount of markers in the assessment sample, followed by preparation and storage techniques (for example, nucleic acid and / or protein extraction, fixing, storage, freezing, ultrafiltration, concentration, evaporation, centrifugation, etc.) processing.

[0047] Furthermore, it is contemplated that the blood sample is a dried blood spot sample. A dried blood spot sample can be obtained by applying a drop of blood to absorbent filter paper. The blood is allowed to completely saturate the paper and air dry for several hours. The blood can be drawn from the subject to be tested (e.g., from a finger) using a lancet.

[0048] In preferred embodiments, the sample is a blood (i.e., whole blood), serum, or plasma sample. Serum is the liquid fraction of whole blood obtained after blood coagulation. To obtain serum, the clot is removed by centrifugation and the supernatant is collected. Plasma is the cell-free liquid portion of blood. To obtain a plasma sample, whole blood is collected in a test tube treated with an anticoagulant (e.g., a citrate-treated test tube or an EDTA-treated test tube). Cells are removed from the sample by centrifugation and a supernatant (i.e., a plasma sample) is obtained.

[0049] According to the present invention, the amount of insulin-like growth factor binding protein 7 (IGFBP-7) is determined. Preferably, the amount of IGFBP-7 polypeptide is determined. IGFBP-7 is a 30-kDa modular glycoprotein known to be secreted by endothelial cells, vascular smooth muscle cells, fibroblasts, and epithelial cells (Ono, Y. et al., Biochem Biophys Res Comm 202 (1994) 1490-1496). Preferably, the term "IGFBP-7" refers to human IGFBP-7. The sequence of this protein is well known in the art and can be obtained, for example, via Uni-Prot (Q16270, IBP7_HUMAN) or via GenBank (NP_001240764.1). A detailed definition of the biomarker IGFBP-7 is provided, for example, in WO 2008 / 089994, the entire contents of which are incorporated herein by reference. IGFBP-7 has two isoforms: isoform 1 and isoform 2, which are produced by alternative splicing. In an embodiment of the present invention, the total amount of both isoforms was determined (for sequences, see UniProt database entries (Q16270-1 and Q16270-2)).

[0050] IGF binding protein 7 (=IGFBP7) is a 30-kDa modular glycoprotein known to be secreted by endothelial cells, vascular smooth muscle cells, fibroblasts, and epithelial cells (Ono, Y. et al., Biochem Biophys Res Comm 202 (1994) 1490-1496). In this literature, this molecule is also designated as FSTL2; IBP 7; IGF binding protein-related protein 1; IGFBP 7; IGFBP 7v; IGFBP rP1; IGFBP7; IGFBPRP1; insulin-like growth factor binding protein 7; insulin-like growth factor binding protein 7 precursor; MAC25; MAC25 protein; PGI2 stimulating factor; and PSF or prostacyclin stimulating factor. Northern blot studies revealed that this gene is widely expressed in human tissues, including heart, brain, placenta, liver, skeletal muscle, and pancreas (Oh, Y. et al., J. Biol. Chem. 271 (1996) 30322-30325).

[0051] IGFBP7 was initially identified as a gene differentially expressed in normal leptomeningeal and mammary epithelial cells compared to their corresponding tumor cells and was named meningioma-associated cDNA (MAC25) (Burger, AM et al., Oncogene 16 (1998) 2459-2467). The expressed protein was independently purified as a tumor-derived adhesion factor (later renamed angiomodulin) (Sprenger, CC et al., Cancer Res 59 (1999) 2370-2375) and as a prostacyclin stimulator (Akaogi, K., et al., Proc Natl Acad Sci USA 93 (1996) 8384-8389). It was also reported as T1A12, a gene downregulated in breast cancer (St Croix, B. et al., Science 289 (2000) 1197-1202).

[0052] Preferably, the term "IGFBP7" refers to human IGFBP7. The sequence of this protein is well known in the art and is available, for example, via GenBank (NP_001240764.1). As used herein, IGFBP7 preferably also includes variants of specific IGFBP7 polypeptides.

[0053] As used herein, the term "BNP-type peptide" includes pre-proBNP, proBNP, NT-proBNP, and BNP. The precursor propeptide (134 amino acids in the case of pre-proBNP) contains a short signal peptide that is enzymatically cleaved to release the leader peptide (108 amino acids in the case of proBNP). The propeptide is further cleaved into the N-terminal propeptide (NT-propeptide, 76 amino acids in the case of NT-proBNP) and the active hormone (32 amino acids in the case of BNP). Preferably, the BNP-type peptide according to the present invention is NT-proBNP, BNP, and variants thereof. BNP is the active hormone and has a shorter half-life than its inactive counterpart, NT-proBNP. BNP is metabolized in the blood, while NT-proBNP circulates in the blood as an intact molecule and is therefore cleared by the kidneys. NT-proBNP has a 120-minute longer in vivo half-life than BNP, which has a half-life of 20 minutes (Smith 2000, J Endocrinol. 167:239-46). Pre-analysis of NT-proBNP is more reliable, and samples can be easily shipped to a central laboratory (Mueller 2004, Clin Chem Lab Med 42:942-4). Blood samples can be stored at room temperature for several days or mailed or shipped without recovery losses. In contrast, BNP storage at room temperature or 4°C for 48 hours results in a concentration loss of at least 20% (Mueller loc. cit.; Wu 2004, Clin Chem 50:867-73). Therefore, depending on the time course or property of interest, measuring either the active or inactive form of the natriuretic peptide may be advantageous. The most preferred natriuretic peptide according to the present invention is NT-proBNP or a variant thereof. As briefly discussed above, the human NT-proBNP referred to in accordance with the present invention is a polypeptide preferably comprising 76 amino acids corresponding to the N-terminal portion of the human NT-proBNP molecule. The structures of human BNP and NT-proBNP are described in detail in the prior art (e.g., WO 02 / 089657, WO 02 / 083913, or Bonow loc.cit). Preferably, the human NT-proBNP used herein is the human NT-proBNP disclosed in EP 0648228 B1. These prior art documents are incorporated herein by reference with respect to the specific sequences of NT-proBNP and variants thereof disclosed therein.

[0054] CRP (C-reactive protein) is an acute phase protein that was discovered more than 75 years ago to be a blood protein that binds to the C polysaccharide of pneumococcus. CRP is known as a reactive inflammatory marker and is produced by a distal organ (i.e., the liver) in response to or reaction to chemokines or interleukins originating from the primary lesion site. It is known that CRP consists of five single subunits that are non-covalently linked and assembled into a cyclic pentamer with a molecular weight of approximately 110-140 kDa. Preferably, CRP as used herein relates to human CRP. The sequence of human CRP is well known and disclosed, for example, by Woo et al. (J. Biol. Chem. 1985. 260 (24), 13384-13388). CRP levels are generally low in normal individuals, but may rise 100 to 200 times or more due to inflammation, infection, or injury (Yeh (2004) Circulation. 2004; 109: 11-11-11-14). CRP is known to be an independent factor in predicting cardiovascular risk. In particular, it has been shown that CRP is suitable as a predictor of myocardial infarction, stroke, peripheral arterial disease, and sudden cardiac death. In addition, elevated CRP levels can also predict recurrent ischemia and death in subjects with acute coronary syndrome (ACS) and those undergoing coronary intervention.

[0055] As used herein, the term "amount" includes the absolute amount of the biomarkers mentioned herein, the relative amount or concentration of the biomarkers, and any value or parameter related thereto or derivable therefrom. Such values ​​or parameters include intensity signal values ​​of all specific physical or chemical properties obtained from the peptide by direct measurement, such as intensity values ​​in a mass spectrum or NMR spectrum. In addition, included are all values ​​or parameters obtained by indirect measurement specified elsewhere in this specification, such as the response amount measured from a biological readout system in response to the intensity signal obtained from the peptide or from a specifically bound ligand. It should be understood that values ​​related to the above amounts or parameters can also be obtained by all standard mathematical operations.

[0056] In some embodiments of the invention, the term "amount" refers to "mass concentration," which is defined as the mass of the biomarker divided by the sample volume. The SI unit for mass concentration is kg / m 3 (kilograms per cubic meter), which is equivalent to mg / mL and g / L. For the concentration of a biomarker, other units such as "mg / ml" or "pg / ml" are often used. Therefore, the amount of a biomarker can be the mass of the biomarker in 1 ml of test sample.

[0057] As used herein, the term "determining" the amount of a biomarker refers to quantification of the biomarker, for example, determining the level of the biomarker in a sample using a suitable detection method as described elsewhere herein.

[0058] In one embodiment, the amount of a biomarker is determined by contacting the sample with a reagent that specifically binds to the biomarker, thereby forming a complex between the reagent and the biomarker, detecting the amount of the complex formed, and thereby determining the amount of the biomarker.

[0059] The biomarkers mentioned herein can be detected using methods generally known in the art. Detection methods generally include methods for quantifying the amount of biomarkers in a sample (quantitative methods). Those skilled in the art generally know which of the following methods are suitable for qualitative and / or quantitative detection of biomarkers. Commercially available Western and immunoassays (such as ELISA, RIA, fluorescence and luminescence-based immunoassays) can be used to conveniently measure, for example, proteins in a sample. Other suitable methods for detecting biomarkers include determining the specific physical or chemical properties of a peptide or polypeptide, such as its accurate molecular mass or NMR spectrum. The methods include, for example, biosensors, optical devices coupled to immunoassays, biochips, analytical devices (such as mass spectrometers, NMR analyzers or chromatographic devices). In addition, methods include microplate ELISA-based methods, fully automated or robotic immunoassays (e.g., on an Elecsys TM available on analyzers), CBA (e.g., at Roche-Hitachi TM Enzymatic cobalt binding assays available on analyzers) and latex agglutination assays (e.g., on Roche-Hitachi TM available on the analyzer).

[0060] For the detection of biomarker proteins mentioned herein, various immunoassay techniques using this assay format are available, see, for example, U.S. Patent Nos. 4,016,043, 4,424,279, and 4,018,653. These techniques include single-site and two-site or "sandwich" assays of the non-competitive type, as well as traditional competitive binding assays. These assays also include direct binding of labeled antibodies to target biomarkers. Sandwich assays are one of the most useful immunoassays.

[0061] Methods employing electrochemiluminescent labels are well known. Such methods exploit the ability of specific metal complexes to achieve an excited state upon oxidation, from which they decay to a ground state, thereby emitting electrochemiluminescence. For a review, see Richter, MM, Chem. Rev. 104 (2004) 3003-3036.

[0062] In one embodiment, the detection antibody (or antigen-binding fragment thereof) to be used to determine the amount of the biomarker is ruthenated or iridium-containing. Thus, the antibody (or antigen-binding fragment thereof) should contain a ruthenium tag. In one embodiment, the ruthenium tag is a bipyridine-ruthenium (II) complex. Alternatively, the antibody (or antigen-binding fragment thereof) should contain an iridium tag. In one embodiment, the iridium tag is a complex as disclosed in WO 2012 / 107419.

[0063] Determining the amount of a polypeptide (such as IGFBP-7) may preferably comprise the steps of: (a) contacting the polypeptide with a reagent that specifically binds to the polypeptide, (b) (optionally) removing unbound reagent, and (c) determining the amount of bound binding agent, i.e., the complex of reagent formed in step (a). According to a preferred embodiment, the contacting, optionally removing, and determining steps may be performed by an analyzer unit. According to some embodiments, the steps may be performed by a single analyzer unit of the system or by more than one analyzer unit that are in operable communication with each other. For example, according to a specific embodiment, the system disclosed herein may comprise a first analyzer unit for performing the contacting and optionally removing steps; and a second analyzer unit for performing the determining step, the second analyzer unit being operably connected to the first analyzer unit via a transfer unit (e.g., a robotic arm).

[0064] The reagent that specifically binds to the biomarker (also referred to herein as a "binding agent") can be covalently or non-covalently coupled to a label, thereby allowing detection and measurement of the bound reagent. Labeling can be performed by direct or indirect methods. Direct labeling involves direct (covalent or non-covalent) coupling of a label to a binding agent. Indirect labeling involves binding of a secondary binding agent to a first binding agent (covalently or non-covalently). The secondary binding agent should specifically bind to the first binding agent. The secondary binding agent can be coupled to a suitable label and / or be the target (receptor) of a tertiary binding agent to which the secondary binding agent binds. Suitable secondary and higher-level binding agents can include antibodies, secondary antibodies, and well-known binding systems, such as the streptavidin-biotin system (Vector Laboratories, Inc.). The binding agent or substrate can also be "labeled" with one or more labels known in the art. Such labels can then be the target of a higher-level binding agent. Suitable labels include biotin, digoxigenin, His tags, glutathione-S-transferase, FLAG, GFP, myc tags, influenza A virus hemagglutinin (HA), maltose binding protein, and the like. In the case of peptides or polypeptides, the tag is preferably located at the N-terminus and / or the C-terminus. Suitable labels are any tags that can be detected by an appropriate detection method. Typical labels include gold particles, latex beads, dihydroacridinium esters, luminol, ruthenium complexes, iridium complexes, enzyme activity tags, radioactive tags, magnetic tags (e.g., magnetic beads, including paramagnetic tags and superparamagnetic tags), and fluorescent tags. Enzyme activity tags include, for example, horseradish peroxidase, alkaline phosphatase, β-galactosidase, luciferase, and derivatives thereof. Suitable substrates for detection include diaminobenzidine (DAB), 3,3'-5,5'-tetramethylbenzidine, NBT-BCIP (4-nitro blue tetrazolium chloride and 5-bromo-4-chloro-3-indolyl phosphate, available as a ready-to-use stock solution from Roche Diagnostics), CDP-Star TM (Amersham Bio-sciences), ECF TM (Amersham Biosciences). Suitable enzyme-substrate combinations can produce colored reaction products, fluorescence or chemiluminescence, which can be measured according to methods known in the art (e.g., using photographic film or a suitable camera system). For measuring enzyme reactions, the above-given criteria are similarly applicable. Typical fluorescent labels include fluorescent proteins (such as GFP and its derivatives), Cy3, Cy5, Texas Red, fluorescein, and Alexa dyes (e.g., Alexa 568). Additional fluorescent labels are available, for example, from Molecular Probes (Oregon). Similarly, quantum dots are also contemplated as fluorescent labels. Radioactive labels can be detected by any known and appropriate method (e.g., photographic film or phosphorimager).

[0065] The amount of the polypeptide can also preferably be determined by (a) contacting a solid support comprising a binding agent for the polypeptide as described elsewhere herein with a sample comprising the peptide or polypeptide, and (b) determining the amount of the peptide or polypeptide bound to the support. Materials for making supports are well known in the art and include, among others, commercially available column materials, polystyrene beads, latex beads, magnetic beads, colloidal metal particles, glass and / or silicon wafers and surfaces, nitrocellulose tapes, membranes, sheets, duracytes, wells and walls of reaction trays, plastic tubing, and the like.

[0066] In another aspect, the sample is removed from the complex formed between the binding agent and a marker prior to measuring the amount of the complex formed. Thus, in one aspect, the binding agent can be immobilized on a solid support. In another aspect, the sample can be removed from the complex formed on the solid support by applying a washing solution.

[0067] "Sandwich assay" is one of the most useful and most commonly used assays, including many variations of sandwich assay technology. Simply put, in a typical assay, an unlabeled (capture) binding agent is fixed or can be fixed on a solid substrate, and the sample to be tested is contacted with the capture binding agent. After a suitable incubation period, after a period of time sufficient to allow the formation of a binding agent-biomarker complex, a second (detection) binding agent labeled with a reporter molecule capable of producing a detectable signal is then added, and incubated for a time sufficient to allow the formation of another complex of the binding agent-biomarker-labeled binding agent. Optionally, any unreacted material can be washed away. The presence of a biomarker is determined by observing the signal generated by the reporter molecule bound to the detection binding agent. The results can be qualitatively characterized by simply observing the visible signal, or can be quantitatively characterized by comparison with a control sample containing a known amount of biomarkers.

[0068] The incubation steps of a typical sandwich assay can be varied as needed and as appropriate. Such variations include, for example, simultaneous incubations in which two or more binding agents and biomarkers are co-incubated. For example, both the sample to be analyzed and the labeled binding agent are added to the immobilized capture binding agent simultaneously. Alternatively, the sample to be analyzed and the labeled binding agent can be incubated first, followed by addition of the antibody that binds to or is capable of binding to the solid.

[0069] The complex formed between a specific binding agent and a biomarker should be proportional to the amount of the biomarker present in the sample. It should be understood that the specificity and / or sensitivity of the binding agent to be used defines the degree of proportion of at least one marker contained in the sample that can be specifically bound. Further details on how the measurement can be performed can also be found elsewhere herein. The amount of the complex formed should be converted to the amount of the biomarker, thereby reflecting the actual amount present in the sample.

[0070] The terms "binding agent", "specific binding agent", "analyte-specific binding agent", "detection agent" and "agent that specifically binds to a biomarker" are used interchangeably herein. Preferably, they relate to an agent that comprises a binding portion that specifically binds to the corresponding biomarker. Examples of "binding agents" or "agents" are nucleic acid probes, nucleic acid primers, DNA molecules, RNA molecules, aptamers, antibodies, antibody fragments, peptides, peptide nucleic acids (PNA) or compounds. Preferred agents are antibodies or antigen-binding fragments thereof that specifically bind to the biomarker to be measured. The term "antibody" is used herein in the broadest sense and includes various antibody structures, including but not limited to monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies) and antibody fragments, as long as they exhibit the desired antigen-binding activity (i.e., antigen-binding fragments thereof). Preferably, the antibody is a polyclonal antibody. More preferably, the antibody is a monoclonal antibody.

[0071] The term "specific binding" or "specific binding" refers to a binding reaction in which the binding pair molecules exhibit binding to each other under conditions in which they do not significantly bind to other molecules. The term "specific binding" or "specific binding" when referring to a protein or peptide as a biomarker refers to a binding reaction in which the binding agent binds to the protein or peptide with a specific binding affinity of at least 10 -7 The term "specific binding" or "specific binding" preferably refers to an affinity of at least 10 for its target molecule. -8 M or even more preferably at least 10 -9 M. The terms "specific" or "specific" are used to indicate that other molecules present in a sample do not significantly bind to a binding agent specific for a target molecule.

[0072] In step (b) of the present invention, (i) the ratio of the amount of IGFBP7 to the amount of BNP peptide. Alternatively, (ii) the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of BNP-type peptide.

[0073] The term "calculating a ratio" as referred to herein relates to calculating the ratio of i) the amount of IGFBP7 or ii) the sum of the amounts of IGFBP7 and CRP by dividing the amount of BNP-type peptide or by performing any other comparable mathematical calculation that relates the amount in i) or ii) to the amount of BNP-type peptide. Preferably, the amount in i) or ii) is divided by the amount of BNP-type peptide to calculate the ratio (thus, the ratio of the amount in i) or ii) to the amount of BNP-type peptide is calculated).

[0074] In some embodiments, the sum of the amounts of IGFBP7 and CRP is determined by adding the amounts of IGFBP7 and CRP (e.g., by adding the mass concentration of IGFBP7 and the mass concentration of CRP). Typically, the same unit is used for the amounts of CRP and IGFBP7, such as "ng / mL." The sum of the determined amounts of IGFBP7 and CRP is then divided by the amount of the BNP-type peptide. For the BNP-type peptide, the same unit as for the amounts of CRP and IGFBP7 can be used. However, it is also conceivable that amounts expressed in pg / ml will be used for the calculations. In some embodiments, amounts expressed in ng / ml are used for CRP and IGFBP7, while amounts expressed in pg / ml are used for the BNP-type peptide.

[0075] If the amount in i) or ii) is divided by the amount of BNP-type peptide to calculate the ratio, the following ratio is calculated:

[0076] 1. IGFBP7 / BNP-type peptide (i.e., the ratio of the amount of IGFBP7 to the amount of BNP-type peptide), or

[0077] 2. (IGFBP7+CRP) / BNP-type peptide (i.e., the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of BNP-type peptide)

[0078] If the above ratio (1. or 2.) is determined, the following applies to the diagnostic algorithm in the method of differentiating HFpEF from HFrEF.

[0079] Preferably, a ratio above the reference ratio is indicative of heart failure with preserved ejection fraction (HFpEF), whereas a ratio below the reference ratio is indicative of heart failure with reduced ejection fraction (HFrEF).

[0080] If the above ratio (1. or 2.) is determined, the following applies to the diagnostic algorithm in the method for diagnosing HFpEF.

[0081] Preferably, a ratio above the reference ratio is indicative of a subject with heart failure with preserved ejection fraction (HFpEF), whereas a ratio below the reference ratio is indicative of a subject without HFpEF, ie a subject with HF or suspected of having HF but not with HFpEF.

[0082] Also preferably, the amount of the BNP-type peptide is divided by the amount of i) or ii) to calculate the ratio (thus, the ratio of the amount of the BNP-type peptide to the amount of i) or ii) is calculated).

[0083] Therefore, the following ratio is calculated

[0084] 3. BNP-type peptide / IGFBP7, or

[0085] 4. BNP-type peptide / (IGFBP7+CRP)

[0086] If the above ratio (3. or 4.) is determined, the following applies to the diagnostic algorithm in the method of differentiating HFpEF from HFrEF.

[0087] Preferably, a ratio below the reference ratio is indicative of heart failure with preserved ejection fraction (HFpEF), whereas a ratio above the reference ratio is indicative of heart failure with reduced ejection fraction (HFrEF).

[0088] If the above ratio (3. or 4.) is determined, the following applies to the diagnostic algorithm in the method for diagnosing HFpEF.

[0089] Preferably, a ratio below the reference ratio is indicative of a subject with heart failure with preserved ejection fraction (HFpEF), whereas a ratio above the reference ratio is indicative of a subject without HFpEF, ie a subject with HF or suspected of having HF but not with HFpEF.

[0090] In step c) of the method of the invention, the ratio calculated in step (b) is compared with a reference ratio.

[0091] The term "comparison" as used herein preferably refers to comparing the ratio calculated according to the method of the present invention with a reference ratio (i.e., a suitable reference ratio). It should be understood that comparison as used herein generally refers to a comparison of values. The comparison can be performed manually or in a computer-assisted manner. Thus, the comparison can be performed by a computing device. The values ​​of the calculated ratio and the reference ratio can be compared, for example, to each other, and the comparison can be performed automatically by a computer program that executes an algorithm for the comparison. The computer program that performs the evaluation will provide the required evaluation in a suitable output format. For computer-assisted comparison, the value of the calculated ratio can be compared with a value corresponding to a suitable reference ratio, which is stored by the computer program in a database. The computer program can further evaluate the result of the comparison, i.e., automatically provide the required evaluation in a suitable output format. The result of the comparison can preferably be used as an aid in diagnosing HFpEF or distinguishing HFpEF from HFrEF.

[0092] In the method for distinguishing HFpEF from HFrEF, HFpEF and HfrEF in a heart failure subject can be distinguished based on a comparison of the calculated ratio with a reference ratio. Therefore, the reference ratio in the method for distinguishing HFpEF from HFrEF is selected such that the difference or similarity in the comparison values ​​allows for the distinction between HFpEF and HFrEF. Therefore, the term "reference ratio" refers to a value / ratio that allows for the distinction between HFpEF and HFrEF, based on the difference between HFpEF and HfrEF. Thus, the reference ratio can be a threshold value that separates these groups from each other.

[0093] In the method for diagnosing HFpEF, a subject with HFpEF can be distinguished from a subject without HFpEF based on a comparison of the calculated ratio with a reference ratio. Thus, the reference ratio is selected such that the difference or similarity in the compared values ​​allows for the distinction between HFpEF and HFrEF. Thus, in the context of the diagnosis of HFpEF, the term "reference ratio" refers to a value / ratio that allows for the distinction between a subject with HFpEF and a subject without HFpEF.

[0094] The reference ratio applicable to individual subjects may vary according to various physiological parameters such as age, sex or subpopulation and the means for determining the polypeptide or peptide mentioned herein. Suitable reference ratios may be determined from a reference sample to be analyzed together with the test sample (i.e., simultaneously or subsequently).

[0095] In principle, the reference ratio of the subject queue can be calculated based on the average ratio by applying standard statistical methods. In particular, the accuracy of the test (such as the method for diagnosing the occurrence or non-occurrence of an event) is best described by its receiver operating characteristic (ROC) (see in particular Zweig 1993, Clin. Chem. 39: 561-577). The ROC curve is a curve of all sensitivity and specificity pairs produced by continuously changing the decision threshold value within the entire data range observed. The clinical performance of a diagnostic method depends on its accuracy, that is, it can correctly assign a subject to a certain ability to distinguish or diagnose. The ROC curve shows the overlap between the two distributions by plotting the sensitivity and 1-specificity of the entire threshold range applicable to the distinction into a curve. On the y-axis is sensitivity, or true positive score, which is defined as the ratio of the product of the number of true positive test results and the number of true positive test results and the number of false negative test results. This is also referred to as positive when there is a disease or illness. It is calculated only from the affected subgroup. On the x-axis is false positive score, or 1-specificity, which is defined as the ratio of the product of the number of false positive results and the number of true negative results and the number of false positive results. It is an index of specificity and is calculated entirely from unaffected subgroups. Because true positive fraction and false positive fraction are calculated completely separately, by using the test results from two different subgroups, the ROC curve is independent of the prevalence of events in the cohort. Each point on the ROC curve represents the sensitivity / -specificity pair corresponding to a specific decision threshold. A test with perfect discrimination (two result distributions do not overlap) has an ROC curve that passes through the upper left corner, where the true positive fraction is 1.0 or 100% (perfect sensitivity), and the false positive fraction is 0 (perfect specificity). The theoretical curve for a test without discrimination (the result distributions of the two groups are identical) is a 45-degree diagonal line from the lower left corner to the upper right corner. Most curves fall between these two extremes. If the ROC curve is completely below the 45-degree diagonal, it is easily corrected by reversing the standard of "positive" from "greater than" to "less than", and vice versa. Qualitatively, the closer the curve is to the upper left corner, the higher the overall accuracy of the test. Depending on the desired confidence interval, a threshold value can be derived from the ROC curve, allowing the distinction or diagnosis mentioned herein to be performed at an appropriate balance of sensitivity and specificity, respectively. Therefore, a reference for the method of the present invention (i.e., a threshold value allowing the distinction between HFpEF and HFrEF or the diagnosis of HFpEF) can preferably be generated by establishing an ROC for the cohort as described above and deriving a threshold ratio therefrom. Depending on the sensitivity and specificity required for the diagnostic method, the ROC curve allows the derivation of a suitable threshold value.

[0096] The definitions and explanations given above apply mutatis mutandis to the following methods of the present invention.

[0097] The present invention also relates to a computer-implemented method for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in a subject suffering from heart failure, the method comprising the steps of:

[0098] (a) receiving at a processing unit a value for the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), a value for the amount of a BNP-type peptide and optionally a value for the amount of CRP (C-reactive Protein) in a sample from said subject,

[0099] (b) calculating, by the processing unit, (i) the ratio of the amount of the IGFBP7 to the amount of the BNP peptide or (ii) the ratio of the sum of the amounts of the IGFBP7 and the CRP to the amount of the BNP-type peptide,

[0100] (c) comparing, by the processing unit, the ratio calculated in step (b) with a reference ratio, and

[0101] (d) Differentiate between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF).

[0102] The present invention also relates to a computer-implemented method for diagnosing heart failure with preserved ejection fraction (HFpEF) in a subject suspected of having HFpEF, the method comprising the steps of:

[0103] (a) receiving at a processing unit a value for the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), a value for the amount of a BNP-type peptide and optionally a value for the amount of CRP (C-reactive Protein) in a sample from said subject,

[0104] (b) calculating, by the processing unit, (i) the ratio of the amount of the IGFBP7 to the amount of the BNP peptide or (ii) the ratio of the sum of the amounts of the IGFBP7 and the CRP to the amount of the BNP-type peptide,

[0105] (c) comparing, by the processing unit, the ratio calculated in step (b) with a reference ratio, and

[0106] (d) Diagnosis of heart failure with preserved ejection fraction.

[0107] The above method is a computer-implemented method. Preferably, all steps of the computer-implemented method are performed by one or more processing units of a computer (or computer network). Therefore, the assessment (i.e., diagnosis or differentiation) in step (d) is performed by the processing unit. Preferably, the assessment is based on the results of step (c).

[0108] The values ​​received in step (a) will be derived from the determination of the amount of IGFBP-7, BNP-type peptides (and optionally CRP) in the sample from the experimenter as described elsewhere herein. Preferably, these values ​​are the values ​​of the amount of the marker. These values ​​will typically be received by the processing unit by uploading or sending the value to the processing unit. Alternatively, the value can be received by the processing unit by inputting the value via a user interface.

[0109] In an embodiment of the above method, the reference ratio as described in step (c), ie the value of the reference ratio, is established from a memory.

[0110] In an embodiment of the above-described computer-implemented method of the present invention, the results of the evaluation performed in step d) are provided via a display configured for presenting the results.

[0111] In an embodiment of the above-described computer-implemented method of the present invention, the method may comprise the further step of transmitting information about the evaluation performed in step d) to the personal electronic medical record.

[0112] The present invention also relates to a computer program comprising computer-executable instructions for performing the steps of the computer-implemented method for differentiating between HFpEF and HfrEF or for diagnosing HFpEF according to the present invention, when the program is executed on a computer or a computer network. In general, the computer program may specifically comprise computer-executable instructions for performing the steps of the method as disclosed herein. In particular, the computer program may be stored on a computer-readable data carrier.

[0113] The present invention also relates to a method for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in a subject suffering from heart failure, the method comprising the steps of:

[0114] (a) receiving a sample from the subject,

[0115] (b) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), BNP-type peptides and optionally CRP (C-Reactive Protein) in said sample, and

[0116] (c) providing the values ​​of the amounts of said IGFBP7, said BNP-type peptide and optionally said CRP to the attending physician, thereby allowing differentiation between HFpEF and HfrEF.

[0117] The present invention also relates to a method for diagnosing heart failure with preserved ejection fraction (HFpEF) in a subject suspected of having HFpEF, the method comprising the steps of:

[0118] (a) receiving a sample from the subject,

[0119] (b) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), BNP-type peptides and optionally CRP (C-Reactive Protein) in said sample, and

[0120] (c) providing the values ​​of the amounts of said IGFBP7, said BNP-type peptide and optionally said CRP to the attending physician, thereby allowing the diagnosis of HFpEF.

[0121] The physician using the aforementioned method should be the physician who is required to determine the biomarkers IGFBP-7, BNP-type peptides, and optionally CRP for diagnosis or differentiation, i.e., the physician is the attending physician. The physician should be treating the subject to be tested. The aforementioned methods should assist the attending physician in making differentiation and diagnosis, respectively.

[0122] The step a) of receiving a sample does not include extracting a sample from the subject. Instead, a sample obtained from the subject is provided (e.g., under the supervision of an attending physician). For example, the sample can be provided by delivering the sample to a laboratory that determines the amount of biomarkers in the sample.

[0123] After determining these quantities, information about the values ​​of the determined quantities or information about the calculated ratios is provided to the physician. In addition, information about the value of the reference ratio can be provided. The information provided can also include an indication of whether the subject has HFrEF or HFpEF.

[0124] Instead of providing the attending physician with values ​​for the amounts of IGFBP7, BNP-type peptides and optionally CRP (as described in step c of the above method), the attending physician may be provided with values ​​for the ratios as described herein, thereby allowing differentiation between HFpEF and HfrEF (or diagnosis of HFpEF).

[0125] The method for diagnosing HFpEF of the present invention may further comprise the step of treating the subject based on the results of the diagnosis. Preferably, the subject diagnosed with HFpEF by the method of the present invention is treated. Thus, the method may comprise the step of selecting a subject with HFpEF. The selection should be based on the results of the comparison step.

[0126] Treatment can include any treatment that allows for the treatment of HFpEF. The term includes lifestyle changes, dietary regimens, interventions on the body, and administration of appropriate medications.

[0127] In some embodiments, treatment comprises administering at least one drug that allows for the treatment of HFpEF. In some embodiments, treatment comprises administering at least one drug selected from angiotensin-converting enzyme inhibitors (ACE inhibitors), angiotensin II receptor blockers (also commonly referred to as angiotensin II receptor antagonists), beta-adrenergic blocking agents (also referred to herein as beta blockers), aldosterone antagonists, and diuretics.

[0128] In one embodiment, an ACE inhibitor is administered, such as benazepril, captopril, cilazapril, enalapril, fosinopril, lisinopril, moexipril, perindopril, quinapril, ramipril, spirapril, or trandolapril.

[0129] In one embodiment, a beta blocker is administered, such as acebutolol, alprenolol, atenolol, betaxolol, bisoprolol, bupranolol, carazolol, carteolol, carvedilol, celiprolol, metipranolol, metoprolol, nadolol, nebivolol, oxprenolol, penbutolol, pindolol, propranolol, sotalol, talinolol, or timolol.

[0130] In one embodiment, an angiotensin II receptor antagonist is administered, such as losartan, valsartan, irbesartan, candesartan, telmisartan, or eprosartan.

[0131] In one embodiment, a diuretic is administered, such as thiazide and thiazide-like diuretics or potassium-sparing diuretics.

[0132] In one embodiment, an aldosterone antagonist is administered, such as Eplerone, Spironolactone, Canrenone, Mexrenone, or Prorenone.

[0133] Other therapies used to treat HFpEF are et al. (Clin Res Cardiol. 2018 Jan; 107(1): 1-19. doi: 10.1007 / s00392-017-1170-6. Epub 2017 Oct 10), the entire contents of which are incorporated herein by reference. Preferably, the treatment comprises administering at least one drug selected from the group consisting of SGLT2 (sodium-glucose cotransporter-2) inhibitors, soluble guanylate cyclase stimulators, inorganic nitrates, and angiotensin receptor-neprilysin inhibitors (ARNi).

[0134] In one embodiment, the drug is an SGLT2 (sodium-glucose cotransporter-2) inhibitor. In another embodiment, the drug is a soluble guanylate cyclase stimulator. In another embodiment, the drug is an inorganic nitrate. In another embodiment, the drug is an angiotensin receptor-neprilysin inhibitor (ARNi), i.e., a combination drug comprising a neprilysin inhibitor (e.g., sacubitril) and an angiotensin receptor blocker (e.g., valsartan).

[0135] Furthermore, the present invention relates to the use of IGFBP7, a BNP-type peptide and optionally CRP in a sample from a subject for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) or for diagnosing HFpEF.

[0136] The method of the present invention for distinguishing HFpEF from HFrEF may also include a step of treating the subject based on the differentiation result. Typically, the subject diagnosed with HFpEF and the subject diagnosed with HFrEF are treated. The subject diagnosed with HFpEF is generally treated as described above in conjunction with the method for diagnosing HFpEF. In some embodiments, treating the subject comprises administering at least one drug selected from SGLT2 (sodium-glucose cotransporter-2) inhibitors, soluble guanylate cyclase stimulators, inorganic nitrates, and angiotensin receptor-neprilysin inhibitors (ARNi).

[0137] Subjects who have been determined to have HFrEF are preferably treated according to guidelines. In some embodiments, treating HFrEF comprises administering at least one drug selected from angiotensin-converting enzyme inhibitors (ACE inhibitors), angiotensin II receptor blockers (also commonly referred to as angiotensin II receptor antagonists), beta-adrenergic blocking agents (also referred to herein as beta blockers), aldosterone antagonists, and diuretics.

[0138] Finally, the present invention relates to the use of at least one detection agent that binds to IGFBP7 in a sample from a subject, at least one detection agent that binds to a BNP-type peptide in a sample from a subject, and optionally at least one detection agent that binds to CRP in a sample from a subject for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) or for diagnosing HFpEF.

[0139] In one embodiment, the detection agent is an antibody or an antigen-binding fragment thereof.

[0140] Embodiments of the present invention

[0141] The following embodiments of the present invention may be used in combination with any other embodiments described herein.The definitions and explanations provided above apply mutatis mutandis to the following.

[0142] 1. A method for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in a subject with heart failure, the method comprising the steps of:

[0143] (a) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), a BNP-type peptide and optionally CRP (C-Reactive Protein) in a sample from said subject,

[0144] (b) calculating (i) the ratio of the amount of IGFBP7 to the amount of the BNP peptide or (ii) the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide,

[0145] (c) comparing the ratio calculated in step (b) with a reference ratio, and

[0146] (d) Differentiate between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF).

[0147] 2. The method of embodiment 1, wherein the subject is a human.

[0148] 3. The method of embodiments 1 and 2, wherein the sample is a blood, serum or plasma sample.

[0149] 4. The method of any one of embodiments 1 to 3, wherein the BNP-type peptide is BNP or NT-proBNP.

[0150] 5. The method of any one of embodiments 1 to 4, wherein the ratio of the amount of IGFBP7 to the amount of the BNP-type peptide is calculated.

[0151] 6. The method according to any one of embodiments 1 to 4, wherein the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide is calculated.

[0152] 7. The method of embodiment 5 or 6, wherein a ratio above the reference ratio is indicative of heart failure with preserved ejection fraction (HFpEF), and / or wherein a ratio below the reference ratio is indicative of heart failure with reduced ejection fraction (HFrEF).

[0153] 8. A method for diagnosing heart failure with preserved ejection fraction (HFpEF) in a subject suspected of having HFpEF, the method comprising the steps of:

[0154] (a) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), a BNP-type peptide and optionally CRP (C-Reactive Protein) in a sample from said subject,

[0155] (b) calculating (i) the ratio of the amount of IGFBP7 to the amount of the BNP peptide or (ii) the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide,

[0156] (c) comparing the ratio calculated in step (b) with a reference ratio, and

[0157] (d) Diagnosis of heart failure with preserved ejection fraction.

[0158] 9. A computer-implemented method for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in a subject with heart failure, the method comprising the steps of:

[0159] (a) receiving at a processing unit a value for the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), a value for the amount of a BNP-type peptide and optionally a value for the amount of CRP (C-reactive Protein) in a sample from said subject,

[0160] (b) calculating, by the processing unit, (i) the ratio of the amount of the IGFBP7 to the amount of the BNP peptide or (ii) the ratio of the sum of the amounts of the IGFBP7 and the CRP to the amount of the BNP-type peptide,

[0161] (c) comparing, by the processing unit, the ratio calculated in step (b) with a reference ratio, and

[0162] (d) Differentiate between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF).

[0163] 10. A computer-implemented method for diagnosing heart failure with preserved ejection fraction (HFpEF) in a subject suspected of having HFpEF, the method comprising the steps of:

[0164] (a) receiving at a processing unit a value for the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), a value for the amount of a BNP-type peptide and optionally a value for the amount of CRP (C-reactive Protein) in a sample from said subject,

[0165] (b) calculating, by the processing unit, (i) the ratio of the amount of the IGFBP7 to the amount of the BNP peptide or (ii) the ratio of the sum of the amounts of the IGFBP7 and the CRP to the amount of the BNP-type peptide,

[0166] (c) comparing, by the processing unit, the ratio calculated in step (b) with a reference ratio, and

[0167] (d) Diagnosis of heart failure with preserved ejection fraction.

[0168] 11. A method as described in embodiments 9 and 10, wherein the reference ratio is established from a memory.

[0169] 12. A method for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in a subject with heart failure, the method comprising the steps of:

[0170] (a) receiving a sample from the subject,

[0171] (b) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), BNP-type peptides and optionally CRP (C-Reactive Protein) in said sample, and

[0172] (c) providing the values ​​of the amounts of said IGFBP7, said BNP-type peptide and optionally said CRP to the attending physician, thereby allowing differentiation between HFpEF and HfrEF.

[0173] 13. A method for diagnosing heart failure with preserved ejection fraction (HFpEF) in a subject suspected of having HFpEF, the method comprising the steps of:

[0174] (a) receiving a sample from the subject,

[0175] (b) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), BNP-type peptides and optionally CRP (C-Reactive Protein) in said sample, and

[0176] (c) providing the values ​​of the amounts of said IGFBP7, said BNP-type peptide and optionally said CRP to the attending physician, thereby allowing the diagnosis of HFpEF.

[0177] 14. Use of IGFBP7, a BNP-type peptide and optionally CRP as biomarkers in a sample from a subject for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) or for diagnosing HFpEF.

[0178] 15. Use of at least one detection agent that binds to IGFBP7 in a sample from a subject, at least one detection agent that binds to a BNP-type peptide in a sample from a subject, and optionally at least one detection agent that binds to CRP in a sample from a subject for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) or for diagnosing HFpEF.

[0179] 16. The use according to embodiment 15, wherein the detection agent is an antibody or an antigen-binding fragment thereof.

[0180] 17. The method of any one of embodiments 8 to 13 or the use of embodiment 14 or 16, wherein the sample is a blood, serum or plasma sample.

[0181] 18. The method of embodiments 8 to 13 or the use of embodiment 14 or 16, wherein the subject is a human.

[0182] 19. The method of embodiments 8 to 13 or the use of embodiment 14 or 16, wherein the BNP-type peptide is BNP or NT-proBNP. BRIEF DESCRIPTION OF THE DRAWINGS

[0183] Figure 1 Shown: Biomarker measurements for diagnosing HFpEF

[0184] Figure 1a : Measurement of NT-proBNP in 499 HFrEF patient samples and 123 HFpEF patient samples for the diagnosis of HFpEF

[0185] Figure 1b : Measurement of CRP in 411 HFrEF patient samples and 107 HFpEF patient samples for the diagnosis of HFpEF

[0186] Figure 1c : Measurement of GDF15 in 451 HFrEF patient samples and 110 HFpEF patient samples for the diagnosis of HFpEF

[0187] Figure 1d Measurement of IGFBP-7 in 366 HFrEF heart failure patient samples and 96 HFpEF patient samples for the diagnosis of HFpEF

[0188] Figure 2 Shown: GDF15 biomarker ratio for diagnosing HFpEF

[0189] Figure 2a: Diagnosis of HFpEF and HFrEF in heart failure patients using the ratio CRP+GDF15 (elevated in HFrEF vs. not elevated in HFpEF) / NTproBNP (elevated in HFrEF vs. HFpEF) in samples from 410 patients with HFrEF and 107 patients with HFpEF

[0190] Figure 2b : Diagnosis of HFpEF and HFrEF in heart failure patients using the ratio CRP+GDF15+ST2 (elevated in HFrEF vs. not elevated in HFpEF) / NTproBNP (elevated in HFrEF vs. HFpEF) in samples from 411 patients with HFrEF and 107 patients with HFpEF

[0191] Figure 3 Shown: IGFBP-7 biomarker ratio for diagnosing HFpEF

[0192] Figure 3a : Diagnosis of HFpEF and HFrEF in heart failure patients using the ratio CRP+IGFBP-7 (elevated in HFrEF vs. not elevated in HFpEF) / NTproBNP (elevated in HFrEF vs. HFpEF) in samples from 359 patients with HFrEF and 96 patients with HFpEF

[0193] Figure 3b : Diagnosis of HFpEF and HFrEF in heart failure patients using the ratio CRP+IGFBP-7+ST2 (elevated in HFrEF vs. not elevated in HFpEF) / NTproBNP (elevated in HFrEF vs. HFpEF) in samples from 359 patients with HFrEF and 96 patients with HFpEF

[0194] Figure 3c Diagnosis of HFpEF and HFrEF in heart failure patients using the ratio GDF-15+IGFBP-7 (unelevated in HFrEF vs. unelevated in HFpEF) / NTproBNP (elevated in HFrEF vs. HFpEF)

[0195] All references cited herein are hereby incorporated by reference either for their general disclosure or for their specific disclosure above. DETAILED DESCRIPTION

[0196] The invention will now be illustrated by the following examples, which are not intended to limit or define the scope of the invention.

[0197] Example

[0198] Example 1: Assessment of HFpEF using circulating biomarkers

[0199] NTproBNP, GDF-15, ST2, CRP (hs CRP), and IGFBP-7 levels were determined in HF patients with and without reduced ejection fraction.

[0200] The results are shown in Figure 1. NTproBNP was significantly elevated in patients with heart failure with reduced ejection fraction (HFrEF) compared to patients with heart failure with preserved ejection fraction (HFpEF) (AUC 0.659), see Figure 1a . Figure 1b showed that CRP was not significantly elevated in patients with heart failure with reduced ejection fraction (HFrEF) compared with patients with heart failure with preserved ejection fraction (HFpEF) (AUC 0.575). Figure 1c The results showed that GDF15 was not significantly elevated in patients with heart failure with reduced ejection fraction (HFrEF) compared with patients with heart failure with preserved ejection fraction (HFpEF) (AUC 0.554). Figure 1d showed that IGFBP-7 was not significantly elevated in patients with heart failure with reduced ejection fraction (HFrEF) compared with patients with heart failure with preserved ejection fraction (HFpEF) (AUC 0.494).

[0201] Table 1 shows the titers of circulating biomarkers in a subgroup of patients with heart failure with reduced ejection fraction (HFrEF) and a subgroup of patients with heart failure with preserved ejection fraction (HFpEF). Biomarker data for the full set of biomarkers (NTproBNP, GDF15, ST2, CRPhs, and IGFBP7) were available for 366 HFrEF patient samples and 96 HFpEF patient samples.

[0202] Table 1: Titers of circulating biomarkers in patients with HFpEF and HFrEF.

[0203]

[0204] Table 1 clearly shows that in univariate analysis, NTproBNP was detected in both subgroups (HFrEF population compared to HFrEF population) with significantly different marker concentrations.

[0205] Patients with HFrEF exhibited higher NT-proBNP titers compared with those with HFpEF (median values ​​of 4410 vs. 2189 pg / mL, p < 0.001). In contrast, for GDF15, ST2, CRP, and IGFBP7, only slightly elevated marker concentrations were detected in the HFpEF patient subgroup compared with the HFrEF patient subgroup. However, in univariate analysis, the observed differences in marker concentrations between HFpEF and HFrEF were not significant for GDF15, ST2, CRPhs, and IGFBP-7. HFpEF patients exhibited slightly higher titers of GDF15, ST2, CRPhs, and IGFBP-7 compared with those with HFrEF (median values ​​of 4304 vs. 3939 pg / mL, 37.6 vs. 35.7 pg / mL; 8.54 vs. 6.66 ng / mL; 237.1 vs. 242.1 ng / mL; p > 0.001).

[0206] As can be seen above, the differential diagnosis and identification of patients with HFpEF is challenging. Significant increases in circulating titers of NTproBNP were observed in patients with HFrEF and HFpEF. In contrast, no significant differences in marker levels were detected between HFpEF and HFrEF patients for CRP, GDF15, ST2, or IGFBP-7. Therefore, there is no clear indication that the use of a marker panel improves the diagnosis of HFpEF compared with measuring NTproBNP.

[0207] Example 2: GDF15 ratio for diagnosis of HFpEF

[0208] As shown in Table 1, elevated circulating NTproBNP levels were detected in patients with HFrEF compared with those with HFpEF. In contrast, for CRP and GDF15, no significant differences in circulating levels were detected between patients with HF with or without reduced ejection fraction.

[0209] The ratio CRP+GDF15 (unelevated in HFrEF vs. not elevated in HFpEF) / NTproBNP (elevated in HFrEF vs. HFpEF) was evaluated to improve the diagnosis of HFpEF.

[0210] like Figure 2a As shown in Figure 2, the observed AUC for the ratio (CRP+GDF15) / NTproBNP for detecting HFpEF was 0.671. Figure 2bAs shown, the ratio (CRP+GDF15+ST2) / NTproBNP did not further improve the ratio (CRP+GDF15) / NTproBNP. The observed AUC for the ratio (CRP+GDF15+ST2) / NTproBNP for detecting HFpEF was 0.671.

[0211] Based on these results, it can be concluded that the ratio (CRP+GDF15) / NTproBNP improves the diagnosis of HFpEF compared with single biomarker determination of NTproBNP or GDF15 or CRP.

[0212] Example 3: IGFBP-7 Biomarker Ratio for Diagnosis of HFpEF

[0213] As shown in Table 1, elevated circulating NTproBNP levels were detected in patients with HFrEF compared with those with HFpEF. In contrast, for CRP and IGFBP-7, no significant differences in circulating levels were detected between HF patients with or without reduced ejection fraction. The ratio CRP+IGFBP-7 (elevated in HFrEF versus not elevated in HFpEF) / NTproBNP (elevated in HFrEF versus HFpEF) was evaluated to improve the diagnosis of HFpEF.

[0214] like Figure 3a As shown in Figure 2, the AUC for the calculated index of the sum of the biomarkers (CRP + IGFBP-7) / NTproBNP for detecting HFpEF was observed to be 0.699. In comparison, the AUC for the ratio IGFBP-7 / NTproBNP for detecting HFpEF was observed to be 0.680. Figure 3c As shown in Figure 2, the calculated index (GDF-15+IGFBP-7) / NTproBNP for the sum of biomarkers for detecting HF was observed to have an AUC of 0.693. Figure 3bAs shown, the ratio (CRP+IGFBP-7+ST2) / NTproBNP was observed to have comparable performance compared to the ratio (CRP+IGFBP-7) / NTproBNP. The observed AUC for the ratio (CRP+IGFBP-7+ST2):NTproBNP for detecting HFpEF was 0.703. Surprisingly, the calculated index of the sum of the biomarkers (CRP+IGFBP-7):NTproBNP even showed superior performance for diagnosing HFpEF than the ratio (CRP+GDF15) / NTproBNP. The ratio (CRP+IGFBP-7) / NTproBNP even showed superior performance than the ratio (CRP+GDF15+ST2) / NTproBNP, which has been described by Sinning et al. (AUC 0.699 vs. AUC 0.671).

[0215] Table 2 shows the fold changes in the calculated indices in patients with HFpEF and HFrEF.

[0216] algorithm Fold change between HFpEF and HFrEF (IGFBP-7) / NTproBNP 74,97％ (IGFBP-7+CRP) / NTproBNP 87,81％ (GDF15+CRP) / NTproBNP 66,49％ (GDF15+CRP+ST2) / NTproBNP 65,34％

[0217] The fold change was calculated as ((median HFpEF) / (median HFrEF))-1*100.

[0218] Table 2 clearly shows that the calculated index of the sum of (IGFBP-7 + CRP) / NTproBNP has improved performance in the differential diagnosis of HFpEF from HFrEF compared to the calculated index of the sum of (GDF15 + CRP + ST2) / NTproBNP (fold change of 87.81% vs. 65.34%). Compared to the ratio IGFBP-7 / NTproBNP, a fold change of 74.97% was observed.

[0219] In conclusion, IGFBP-7 improved the identification of HFpEF compared with a score based on NTproBNP, GDF15, and CRP without IGFBP-7.

[0220] The current score for IGFBP-7 showed a better effect size compared with the score published by Sinning et al.

[0221] Conclusions: An algorithm including the ratio (CRP+IGFBP-7) / NTproBNP is helpful in diagnosing patients with HFpEF. This algorithm can be used to diagnose HFpEF in patients with heart failure and to identify HFpEF in individuals suspected of having heart failure.

[0222] Example 4: Case Study

[0223] A 76-year-old obese hypertensive woman presented with shortness of breath. NTproBNP, IGFBP-7, and CRP were measured in a serum sample obtained from the patient (using Elecsys NTproBNP, Elecsys IGFBP-7, and Cobas CRP). Due to obesity, the NTproBNP value may be in the gray area, so a differential diagnosis of heart failure was made. The NTproBNP value was 476 pg / mL, the IGFBP-7 value was 190.8 ng / mL, and the CRP value was 5.8 ng / mL. The calculated index (CRP+IGFBP-7):NTproBNP was 0.41, which was elevated relative to the calculated reference ratio (0.09). The reference ratio was calculated as the median ratio in the reference cohort. The obtained ratio (0.41) indicates heart failure with preserved ejection fraction, and it has been determined that the patient would benefit more from treatment with a HFpEF-modification strategy (e.g., treatment with an SGLT2 inhibitor). The patient's therapy was adjusted accordingly.

[0224] An 81-year-old hypertensive male patient presented with a history of atrial fibrillation and shortness of breath. NTproBNP, IGFBP-7, and CRP were measured in a serum sample obtained from the patient (using the aforementioned kits). The observed ejection fraction was in the gray zone (50%). Given the presence of atrial fibrillation, the NTproBNP value was also likely elevated, leading to a differential diagnosis of heart failure. The NTproBNP value was 3289 pg / mL, the IGFBP-7 value was 250.7 ng / mL, and the CRP value was 10.8 ng / mL. The calculated index (CRP+IGFBP-7):NTproBNP was 0.79, which was elevated relative to the calculated reference ratio (0.09). The reference ratio was calculated as the median ratio in the reference cohort. The obtained ratio (0.79) indicated heart failure with preserved ejection fraction, and it was determined that the patient would benefit more from treatment with a HFpEF-modification strategy (e.g., treatment with an SGLT2 inhibitor). The patient's therapy was adjusted accordingly.

[0225] A 68-year-old obese woman with diabetes and hypertension presented with symptoms of exercise intolerance. Because the symptoms of obesity are similar to those of heart failure with preserved ejection fraction (e.g., exercise intolerance), a differential diagnosis of heart failure was performed. NTproBNP, IGFBP-7, and CRP were measured in serum samples obtained from the patient (using the above-mentioned kits). The NTproBNP value was 4643 pg / mL, the IGFBP-7 value was 245.5 ng / mL, and the CRP value was 5.6 ng / mL. The calculated index (CRP+IGFBP-7): NTproBNP was 0.05, which was reduced relative to the reference ratio (0.09). This result indicates heart failure with reduced ejection fraction, and it has been determined that the patient will not benefit further from treatment with HFpEF regulation strategies (e.g., treatment with SGLT2 inhibitors). Therefore, the patient's therapy was not adjusted accordingly.

Claims

1. Use of at least one detection agent that binds to IGFBP7 (insulin-like growth factor binding protein 7), at least one detection agent that binds to a BNP-type peptide, and at least one detection agent that binds to CRP (C-reactive protein) in the preparation of a medicament for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in a subject suffering from heart failure by the following method, the method comprising the following steps: (a) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), BNP-type peptides and CRP (C-reactive protein) in a blood, serum or plasma sample from said subject, (b) calculating the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide, (c) comparing the ratio calculated in step (b) with a reference ratio, and (d) Differentiate between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF).

2. The method of claim 1, wherein the subject is a human.

3. Use according to claims 1 and 2, wherein the BNP-type peptide is BNP or NT-proBNP. 4 . The method according to claim 1 , wherein the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide is calculated.

5. Use according to claim 4, wherein a ratio above the reference ratio is indicative of heart failure with preserved ejection fraction (HFpEF) and / or wherein a ratio below the reference ratio is indicative of heart failure with reduced ejection fraction (HFrEF).

6. Use of at least one detection agent that binds to IGFBP7 (insulin-like growth factor binding protein 7), at least one detection agent that binds to a BNP-type peptide, and at least one detection agent that binds to CRP (C-reactive protein) in the preparation of a medicament for diagnosing heart failure with preserved ejection fraction (HFpEF) in a subject suspected of having HFpEF by the following method, the method comprising the steps of: (a) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), BNP-type peptides and CRP (C-reactive protein) in a blood, serum or plasma sample from said subject, (b) calculating the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide, (c) comparing the ratio calculated in step (b) with a reference ratio, and (d) Diagnosis of heart failure with preserved ejection fraction.

7. Use of at least one detection agent that binds to IGFBP7 (insulin-like growth factor binding protein 7), at least one detection agent that binds to a BNP-type peptide, and at least one detection agent that binds to CRP (C-reactive protein) in the preparation of a medicament for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) in a subject suffering from heart failure by the following method, the method comprising the steps of: (a) receiving a blood, serum or plasma sample from the subject, (b) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), BNP-type peptides and CRP (C-reactive protein) in the sample, and (c) providing the attending physician with the value of the ratio of the sum of the amounts of said IGFBP7 and said CRP to the amount of said BNP-type peptide, thereby allowing differentiation between HFpEF and HfrEF.

8. Use of at least one detection agent that binds to IGFBP7 (insulin-like growth factor binding protein 7), at least one detection agent that binds to a BNP-type peptide, and at least one detection agent that binds to CRP (C-reactive protein) in the preparation of a medicament for diagnosing heart failure with preserved ejection fraction (HFpEF) in a subject suspected of having HFpEF by the following method, the method comprising the steps of: (a) receiving a blood, serum or plasma sample from the subject, (b) determining the amount of IGFBP7 (Insulin-like Growth Factor Binding Protein 7), BNP-type peptides and CRP (C-reactive protein) in the sample, and (c) providing the value of the ratio of the sum of the amounts of said IGFBP7 and said CRP to the amount of said BNP-type peptide to the attending physician, thereby allowing diagnosis of HFpEF.

9. Use of IGFBP7, BNP-type peptides and CRP as biomarkers in a blood, serum or plasma sample from a subject in the preparation of a kit for distinguishing heart failure with preserved ejection fraction (HFpEF) from heart failure with reduced ejection fraction (HFrEF) or for diagnosing HFpEF.

10. The use according to claim 9, wherein the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide is calculated.

11. Use of at least one detection agent that binds to IGFBP7 in a blood, serum or plasma sample from a subject, at least one detection agent that binds to a BNP-type peptide in a blood, serum or plasma sample from a subject, and at least one detection agent that binds to CRP in a blood, serum or plasma sample from a subject in the preparation of a kit for distinguishing between heart failure with preserved ejection fraction (HFpEF) and heart failure with reduced ejection fraction (HFrEF) or for diagnosing HFpEF. 12 . The method according to claim 11 , wherein the ratio of the sum of the amounts of IGFBP7 and CRP to the amount of the BNP-type peptide is calculated.

13. The use according to claim 11, wherein the detection agent is an antibody or an antigen-binding fragment thereof.

14. The use according to any one of claims 6 to 13, wherein the subject is a human.

15. Use according to any one of claims 6 to 13, wherein the BNP-type peptide is BNP or NT-proBNP.

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