Diagnosing and monitoring the progress of acute kidney damage

Abstract

The application describes a method for diagnosing and / or monitoring the progress of acute kidney damage, wherein in a urine sample the concentration of at least one G-protein-coupled receptor, selected from GPR40, GPR43, GPR91 and GPR120, is measured and compared with a normal value. It also describes the use of these G-protein-coupled receptors in this method, and these G-protein-coupled receptors for use in such a method.

Description

[0001] Diagnosis and follow-up of acute kidney injury

[0002] The invention relates to a method for diagnosing and / or monitoring the progression of acute kidney injury, the use of certain G-protein-coupled receptors in this method, and these G-protein-coupled receptors for use in such a method.

[0003] Acute kidney injury (AKI) is a common condition associated with high morbidity and mortality (Chawla et al., 2017; Sawhney et al., 2017; Wang et al., 2012). Acute kidney injury can lead to transition to chronic kidney disease (Kellum et al., 2021; Parr & Siew, 2016). The transition process can initiate a steadily progressive deterioration of kidney function (Kellum et al., 2017; Vanmassenhove et al., 2017). The risk of developing chronic kidney disease is increased by a factor of 8.8 for patients with AKI; the more severe the acute kidney injury, the more frequently it leads to chronic kidney disease (Chawla et al., 2017). Chronic kidney disease itself affects about 11% of the population in highly developed countries such as Germany (Webster et al., 2017) and, with a share of 12%, is a major factor in health expenditure in Germany (Gandjour et al., 2020) .

[0004] The rapid and reliable diagnosis of acute kidney injury as well as the monitoring of the course of acute kidney injury are therefore of great clinical relevance in order to be able to initiate appropriate therapeutic measures as quickly as possible and to minimize the risk of transition to chronic kidney disease. The diagnosis and monitoring of the course of acute kidney injury are currently based on the measurement of the accumulation of metabolic waste products in the blood (particularly creatinine); this accumulation occurs in the case of a loss of kidney function through reduced excretion of metabolic waste products in the urine (Walther et al., 2014). However, the accumulation of metabolic waste products in the blood only occurs with a significant time delay compared to kidney damage.In practice, this leads to delayed diagnosis and thus also delayed treatment of acute kidney injury (Bonventre, 2007) - often the acute kidney injury is not even recognized at all (Wilson et al., 2013; Yang et al., 2015). Current data from Germany confirm this problem (Khadzhynov et al., 2019): acute kidney injury is only correctly diagnosed in one fifth of hospital patients. Depending on the stage of kidney damage, the patients are characterized by a high mortality rate even in the hospital. It has also been shown in an international context that patients with acute kidney injury have a mortality rate that is up to 15 times higher (Glenn M. Chertow et al., 2005; Wang et al., 2012).It is obvious that the prognosis of patients could be considerably improved through faster and more sensitive diagnosis of acute kidney injury. In a highly selected cohort of surgical patients, evidence of this was found using the biomarker candidate TIMP2xIGFBP7 (Göcze et al., 2018). After cardiac surgery, early intervention following an increase in TIMP2xIGFBP7 significantly reduced the likelihood of developing kidney injury, its severity, and the length of hospital stay, which is associated with high costs. In another study, the early implementation of evidence-based measures for treating kidney injury led to a 16.6% reduction in mortality (Meersch et al., 2017).

[0005] In practice, however, biomarkers for acute kidney injury that replace the current gold standard (i.e. creatinine) have not yet been established, as they are only detectable in specific forms of kidney injury (e.g. postoperatively, see above) or, like creatinine, are only measurable with a time delay after kidney injury (e.g. KIM-1).

[0006] Acute kidney injury can lead to an acute increase in serum creatinine concentration or a decrease in urine output (Walther et al., 2014). Measuring serum creatinine is inexpensive but has significant and clearly proven disadvantages: Serum creatinine concentration shows a hyperbolic relationship with excretory kidney function (i.e., glomerular filtration rate). As a result, changes in kidney function initially only minimally manifest themselves in a change in serum creatinine concentration. This is particularly the case in the GFR range between 60-90 ml / min (normal value 90-120 ml / min) (Rahn et al., 1999; Shemesh et al., 1985). As a result, around 50% of kidney function can be lost before this is noticeable in a change in serum creatinine concentration. This means that the diagnosis of kidney damage by determining creatinine is subject to great uncertainty, which is referred to as the so-calledThe term "creatinine-blind range" has been established in the literature. Furthermore, the measurement of serum creatinine concentration depends significantly on muscle mass (or its absence), since creatinine is a metabolic end product, particularly of muscle metabolism (Maciel, 2016). Due to these significant disadvantages of serum creatinine as the gold standard for diagnosing acute kidney injury, the American Society of Nephrology (ASN) has focused its research priority on the discovery of new biomarkers (American Society of Nephrology, 2005) and allocated corresponding research funds. Since then, to the best of our knowledge, 25 alternative biomarkers for diagnosing acute kidney injury have been proposed (Ostermann et al., 2020) (as of October 2022).

[0007] However, none of the alternative biomarkers described in the literature to date are used in standard clinical diagnostics for acute kidney injury or have been included in treatment guidelines of professional societies. This is due, among other things, to the following: In a meta-analysis of 2,979 patients, the sensitivity of the KIM-1 biomarker for diagnosing AKI was 74% and the specificity was 86% (Shao et al., 2014). This means that one in four patients with acute kidney injury is missed, and one in eight patients who test positive do not have kidney injury at all.For the biomarkers KIM-1 and L-FABP, the AUG (area under the curve: a measure of the quality of a biomarker, "0.5" means no predictive power, "1.0" means perfect test) was calculated to be 0.78 and 0.72, respectively, in a large multicenter cohort (TRIBE-AKI consortium) in patients undergoing cardiac surgery who have a greatly increased risk of acute kidney injury. These values ​​indicate that the two biomarkers are only marginally better at diagnosing / predicting acute kidney injury than simple clinical parameters (length of surgery, patient age, etc.).

[0008] blood pressure behavior, etc.). Both markers had no predictive power for the progression of kidney damage (Parikh et al., 2013). The markers NGAL and IL-18 had similarly inadequate test characteristics (AUG for NGAL 0.71, AUC for IL-18 0.63; (Hall et al., 2011)). The AUG for the biomarker DKK-3 in the same model of AKI was 0.78 with a sensitivity of 76% and a specificity of 79%. In contrast to KIM-1 and L-FABP, however, elevated DKK-3 values ​​at least had significant predictive power for permanent kidney damage (Schunk et al., 2019). The alternative biomarker TIMP2xIGFBP7 can only be used for an extremely narrow patient collective. Another problem with the alternative urine biomarkers proposed so far is that their measurement can be confounded by other disease processes (Nadkarni et al., 2017) .This was observed, for example, for the alternative biomarkers cystatin C and IL-18: albumin in the urine inhibits the reabsorption of cystatin C and IL-18, resulting in increased excretion of cystatin C and IL-18 in the urine (Ne at et al., 2012). It is assumed that similar problems could occur with other alternative low-molecular-weight biomarkers, such as NGAL and L-FABP (Charlton et al., 2014).

[0009] To date, the 25 biomarkers mentioned above for diagnosing acute kidney injury have been measured partly in urine and partly in blood. However, the measurement of serum creatinine concentration, i.e., the acute increase, remains the gold standard for diagnosis, with the significant limitations and problems mentioned above.

[0010] In summary, serum creatinine concentrations exhibit considerable fluctuations depending on physical constitution (muscle mass) and have significant diagnostic gaps (delayed increase, "creatinine-blind zone"; see above). The other biomarkers found in the literature do not have sufficient sensitivity and specificity to improve the diagnosis of acute kidney injury or to substantially influence resulting therapeutic decisions. None of these markers has been included in the guidelines for the diagnosis of acute kidney injury.

[0011] The present invention is therefore based on the object of enabling the diagnosis of acute kidney injury with a shorter time delay and / or higher sensitivity and specificity than the known diagnostic methods.

[0012] According to the invention, this is achieved by the subject matter of the independent claims. Preferred embodiments are defined in the dependent claims.

[0013] According to the invention, a method for diagnosing and / or monitoring the progression of acute kidney injury is provided, wherein the concentration of at least one G-protein-coupled receptor selected from GPR40, GPR43, GPR91, and GPR120 is measured in a urine sample and compared with a normal value. An increase compared to the normal value indicates the presence of acute kidney injury.

[0014] A urine sample is urine that is used for subsequent laboratory testing (urine test). The urine sample was previously collected from a patient, i.e., the method according to the invention is not performed on the patient themselves. The method according to the invention is therefore an in vitro method.

[0015] The diagnosis can be used to determine acute kidney injury. Follow-up monitoring involves measuring urine samples over time. It is not necessary for both diagnosis and follow-up monitoring to be performed using the method according to the invention, although this is particularly advantageous.

[0016] In the method according to the invention, the measured concentrations of G-protein-coupled receptors are compared with a normal value, i.e., an average value that can be determined by examining a large number of non-diseased individuals. A deviation from this normal value allows a conclusion to be drawn about the presence of a disease. In acute kidney injury, the concentration of the above-mentioned G-protein-coupled receptors is increased.

[0017] The method according to the invention is based on the finding that the proteins GPR40, GPR43, GPR91, and GPR120 can be used to diagnose acute kidney injury, since their concentrations in urine are elevated in acute kidney injury. This potentially improves the prognosis of patients with acute kidney injury through early initiation of therapy and reduces healthcare costs.

[0018] In one embodiment, the concentrations of the following combinations of two G protein-coupled receptors are measured: GPR40 and GPR43; GPR40 and GPR91; GPR40 and GPR120; GPR43 and GPR91; GPR43 and GPR120; and GPR91 and GPR120.

[0019] In a further embodiment, the concentrations of the following combinations of three G-protein coupled receptors are measured: GPR40, GPR43, and GPR91; GPR40, GPR43, and GPR120; GPR40, GPR91, and GPR120; and GPR43, GPR91, and GPR120. In another embodiment, the concentration of all four G-protein coupled receptors GPR40, GPR43, GPR91, and GPR120 is measured.

[0020] The urine sample can be used in the method according to the invention without any prior treatment or processing steps. Alternatively, the urine sample can be centrifuged to separate cellular components before the concentration measurement is performed.

[0021] In one embodiment, the diagnosis and / or monitoring of acute kidney injury may include at least one of the following steps:

[0022] (1) Risk stratification of patients, for example in planned operations;

[0023] (2) Evidence of acute kidney injury, recognizable by an increase in the above-mentioned G-protein-coupled receptors

[0024] (3) Evidence of acute kidney injury in the presence of chronic kidney disease, ie evidence of acute kidney injury supersedes chronic injury;

[0025] (4) prediction of renal impairment, e.g., a reduction in excretory renal function;

[0026] (5) Diagnosis of acute kidney injury itself;

[0027] (6) quantification of the severity of acute kidney injury;

[0028] (7) Follow-up of acute kidney injury and

[0029] (8) Predicting the probability of recovery after acute kidney injury.

[0030] In another embodiment, the measured concentrations of GPR40, GPR43, GPR91, and / or GPR120 can be normalized to creatinine. This means that the concentrations of G protein-coupled receptors in urine are calculated relative to the concentration of creatinine in urine, according to the international standard. In one embodiment, the urine can come from a mammal, e.g., a human, dog, or cat.

[0031] Methods for measuring the concentration of GPR40, GPR43, GPR91, and / or GPR120 are known. For example, these concentrations can be measured using an antibody-based method, such as an ELISA.

[0032] The invention further relates to the use of at least one G-protein-coupled receptor selected from GPR40, GPR43, GPR91, and GPR120 for the diagnosis and / or monitoring of acute kidney injury, wherein the concentration of at least one of these G-protein-coupled receptors is measured in a urine sample and compared with a normal value. For the implementation of the use, reference is made in full to the above description of the method.

[0033] A further subject of the present invention is a G-protein-coupled receptor selected from GPR40, GPR43, GPR91, and GPR120 for the diagnosis and / or monitoring of acute kidney injury. Reference is also made in full to the above description of the method according to the invention.

[0034] The invention is based on the finding that determining the concentration of the above-mentioned proteins from the family of G-protein-coupled receptors in urine, e.g. using standard laboratory methods (ELISA), improves the current problem of insufficient sensitivity and specificity and / or time delay in the diagnosis of acute kidney injury. The receptors are GPR40, GPR43, GPR91 and GPR120. None of these receptors has previously been described in the literature in connection with the diagnosis or follow-up of acute kidney injury. In particular, there are no publications reporting on the measurement of these receptors for the diagnosis or follow-up of acute kidney injury. The receptors can (cf.above) are used for: (1) risk stratification of patients, for example in planned operations, (2) detection of acute kidney injury, (3) detection of acute kidney injury in the presence of existing chronic kidney disease, i.e. detection of acute kidney injury superimposed on chronic damage, (4) prediction of impaired renal function, e.g. a reduction in excretory kidney function, (5) diagnosis of acute kidney injury, (6) quantification of the severity of kidney damage, (7) monitoring the course of kidney damage, and (8) prediction of the probability of recovery from kidney damage. With the present invention, kidney damage is defined directly via its pathological correlate, acute tubular damage, and not indirectly via the glomerular filtration rate or its surrogate parameter, serum creatinine concentration.

[0035] Acute kidney injury primarily affects proximal tubular epithelial cells (PTECs), which reabsorb the majority of the glomerular primary filtrate. PTECs are highly metabolically active, as they require large amounts of energy for their enormous reabsorption capacity. They are particularly dependent on mitochondria, where fatty acids are broken down to generate energy ("ß-oxidation"). It is therefore not surprising that fatty acid metabolism plays a crucial role in the energy balance of PTECs (Khan et al., 2018, 2020; Mikami et al., 2020; Noels et al., 2021; Yamamoto et al., 2007). It has now been found that the aforementioned G protein-coupled receptors (GPCRs) for fatty acids and fatty acid metabolites are highly expressed in PTEC (see also Fig. 2 below). These receptors are:

[0036] FFAR1 (=GPR40), which binds medium-chain and long-chain saturated and unsaturated fatty acids, FFAR2 (=GPR43), which binds short-chain fatty acids, FFAR4 (=GPR120), which binds long-chain fatty acids, and SUCNR1 (=GPR91), which binds succinate (a metabolite of fatty acids, with a crucial role in the citric acid cycle)

[0037] Acute kidney injury is a very common condition. It affects approximately 8–22% of all hospitalized patients and is associated with a 1.4–15.4-fold increase in mortality (GM Chertow et al., 2006; Glenn M. Chertow et al., 2005; Liangos et al., 2006; Uchino et al., 2006; Wang et al., 2012). Chronic kidney disease, as a common consequence and independent driver of acute kidney injury, affects more than one in ten people in Germany (Brück et al., 2016; Girndt et al., 2016; Levey et al., 2011; Stel et al., 2017) , and about 13% of the world's population is affected by chronic kidney disease, with an increasing trend due to demographic developments (Hill et al., 2016; Xie et al., 2018) .One study identified chronic kidney disease, beyond its own disease-specific complications and risks, as a significant driver of mortality, cardiovascular events (heart attacks), and hospitalization (admission to hospitals) (Go et al., 2004). These findings have since been confirmed and clearly established (Chronic Kidney Disease Prognosis Consortium et al., 2010; Eckardt et al., 2013). Chronic kidney disease is therefore a significant factor in costs and, of course, healthcare expenditures in the insurance system. There is therefore significant public interest in improving healthcare for people with kidney disease, as it is a significant cost factor in the healthcare system.

[0038] Herein, a G protein-coupled receptor (G protein-coupled receptor; abbreviation "GPCR" or "GPR") is understood to mean, in particular, a biological receptor that can transmit signals via GTP-binding proteins ("G proteins" for short) or via other signal transduction pathways. The claims describe that the concentration of at least one G protein-coupled receptor is measured in a urine sample. In principle, the described method for diagnosing and / or monitoring the progression of acute kidney injury using the described G protein-coupled receptors also allows the use of another biological sample, e.g., a blood sample.

[0039] The invention is explained in more detail below with reference to the examples and figures. It is expressly pointed out that the figures and examples should not be understood as limiting the invention to them.

[0040] Fig. 1 is a schematic representation of the inventive method and potential benefits.

[0041] Fig. 2 shows the increased expression of GPR91 (SUCNR1), GPR40 (FFAR1), GPR43 (FFAR2), and GPR120 (FFAR4) in proximal tubular epithelial cells (PTEC) in acute kidney injury, which is independent of the etiology of the acute kidney injury (PAS staining; the attached image is in grayscale for technical reasons only). Ak. Tub. -Necrosis: acute tubular necrosis; inter. Nephr.: interstitial nephritis; ANCA-Vask.: ANCA vasculitis; diab. Nephr.: diabetic nephropathy.

[0042] Figure 3 shows a histological section (PAS staining; the attached image is in grayscale for technical reasons only) of acute kidney injury. It shows that kidney damage is generally not homogeneous throughout the entire organ, but rather focally accentuated.

[0043] Fig. 4 shows an immunohistochemical staining for GPR43. This allows a direct internal comparison between damaged and undamaged renal tubules, thus allowing a quantification of expression changes. It shows that GPR43 is significantly more highly expressed (red staining or darker staining in grayscale) in a damaged tubule (left half of the image, coarsely dashed line) than in an undamaged tubule (right half, finely dashed line).

[0044] Fig. 5 shows a semi-quantitative analysis of 24 patients with acute kidney injury characterized by a significant increase in the expression of GPR91, GPR40, GPR43, and GPR120 in damaged PTEC. This is independent of the etiology of acute kidney injury. ATN: acute tubular necrosis; int. nephr.: interstitial nephritis; ANCA-vask.: ANCA vasculitis; diab. nephr.: diabetic nephropathy.

[0045] Fig. 6 shows schematically the possible debriding process of GPCR in acute kidney injury.

[0046] Fig. 7 shows the results of carrying out the method according to the invention according to Example 4. Fig. 8 shows a schematic representation of an ELISA test method.

[0047] Fig. 1 shows a schematic representation of the implementation of the method according to the invention. The concentrations of GPR40, GPR43, GPR91 and GPR120 are determined in a patient’s urine sample. This can be done using known ELISA tests, which may be commercially available. If the measured values ​​are above a previously determined normal value, this means that acute kidney injury is present. However, if the values ​​are within the normal range or below, this indicates that no acute kidney injury is present. The method according to the invention enables a simple, reliable and early diagnosis of acute kidney injury. Furthermore, statements can be made about the severity of the acute kidney injury and / or the transition to or progression of CKD.

[0048] An ELISA test procedure for determining the concentration of G-protein-coupled receptors is shown schematically in Fig. 8. In the first step, a urine sample is applied to an ELISA plate coated with an antibody against the G-protein-coupled receptor. Subsequently, a second specific antibody binds to the G-protein-coupled receptor. After this, a biotin-labeled third antibody binds to the second antibody. Finally, horseradish peroxidase (HRP)-coupled streptavidin binds to the biotin. After the addition of a substrate, a catalytic reaction takes place which leads to a measurable color change of the liquid depending on the amount of the target protein.

[0049] Example 1: As shown in Fig. 2, acute kidney injury leads to significantly increased expression of GPR91 (SUCNR1), GPR40 (FFAR1), GPR43 (FFAR2), and GPR120 (FFAR4) in proximal tubular epithelial cells (PTEC). The image shows immunohistochemical staining (red) of GPCRs in human kidney tissue (primarily proximal tubule segments) in the normal state (left column) and in acute kidney injury in various entities (acute tubular necrosis, interstitial nephritis, ANCA vasculitis, and diabetic nephropathy / other columns). The fatty acid receptors (GPR40=FFAR1, GPR43=FFAR2, and GPR120=FFAR4) are constitutively detected in the brush border of the tubules. This receptor is debrided in acute kidney injury. In acute kidney injury, there is also strong cytoplasmic expression of these fatty acid receptors. GPR 91 (=SUCNR1) is only very weakly detectable in healthy kidney tissue and is strongly induced in acute kidney injury.GPR 35 expression is significantly less induced by acute kidney injury. The negative control shows no staining (not shown).

[0050] Example 2:

[0051] In a next step, the changes in the expression levels of these receptors in kidney tissue during acute kidney injury were quantified. This took advantage of the fact that kidney injury is usually not homogeneous throughout the organ, but rather focally accentuated (Fig. 3). This allows for a direct internal comparison between damaged and undamaged kidney sections, and thus a quantification of expression changes (Fig. 4).

[0052] Acute kidney injury is often focal (Fig. 3). PAS staining of kidney tissue with intact proximal tubular epithelium (within the finely stippled

[0053] area (top left) and tubular epithelium with acute damage (within the coarsely dashed area, bottom right). This allows for an internal comparison between the expression levels and patterns of normal and damaged tubular segments.

[0054] Fig. 4 shows an immunohistochemical staining of GPR 43. The left section of the image shows distinct cytoplasmic staining of acutely damaged proximal tubular epithelium. This tubule shows cytological signs of acute injury: cytoplasmic vacuolization, impaired lateral cell-cell adhesion, and disruption of the brush border. The adjacent, healthy tubular section to the right shows a GPR 43-immunopositive brush border with intact epithelium, no vacuolization, and no cytoplasmic staining for GPR 43.

[0055] Example 3:

[0056] As shown in Fig. 5, semi-quantitative analyses of 24 patients with acute kidney injury confirm a significant increase in the expression of GPR91, GPR40, GPR43, and GPR120 in injured PTEC. Interestingly, this effect appears to be specific to receptors of fatty acids and metabolites of fatty acid metabolism: For example, the expression of GPR35, a receptor for kynurenic acid (a metabolite of the amino acid tryptophan), is barely altered (Fig. 5; see also Fig. 2).The available data also show that the increased expression of GPR91, GPR40, GPR43 and GPR120 is essentially independent of the context (specific genesis) of the acute kidney injury and of the underlying kidney disease, and therefore proceeds similarly across different injury mechanisms and different kidney diseases: Very similar results are found for acute kidney injury in the context of acute tubular necrosis (for example, in the context of cardiac surgery or kidney injury in the context of severe sepsis), interstitial nephritis, which is often induced by drug-toxic drugs, autoimmune ANCA-associated vasculitis and diabetic kidney disease (Fig. 5). These data suggest that the observed effects represent fundamental mechanisms of acute kidney injury.Thus, the measurement of the GPCRs investigated here in urine is likely to be meaningful for the diagnosis and follow-up of all forms of acute kidney injury.

[0057] Fig. 5 shows that the expression of GPR91, GPR40, GPR43 and GPR120 is increased in acutely injured PTEC regardless of the genesis of the acute kidney injury. Shown are the quantifications of immunohistochemical staining for the mentioned GPCRs in kidney tissue from six patients each (numbered 1 - 6) with acute kidney injury of different genesis: acute tubular necrosis (ATN), interstitial nephritis, ANCA vasculitis and diabetic nephropathy. The cytoplasmic expression levels in normal and cytologically damaged proximal tubular epithelium are compared. The intensity of the staining was evaluated semiquantitatively (scale with increasing severity from 0 to 3). A total of 24 patient samples were examined.

[0058] Test procedure:

[0059] In acute kidney injury, the brush border (apical membrane) of PTEC is debrided into the urine; PTEC can also die in acute kidney injury. Proteins, including receptors such as GPR91, GPR40, GPR43 and GPR120, which are highly expressed in damaged PTEC, are therefore released from PTEC in acute kidney injury and can then be detected in the urine by ELISA (enzyme-linked immunosorbent assay), rapid tests or other tests (e.g. POGT, point-of-care tests) (Fig. 6). These tests can be performed quickly and safely not only in the hospital but also in the outpatient setting (including doctor's offices).

[0060] Example 4 :

[0061] Proceed :

[0062] Urine samples were collected from the subjects (patients and healthy controls) over a period of 24 hours, and approximately 10 ml of these samples were frozen at -80°C. Prior to ELISA analysis, the urine samples were centrifuged at 1,500 g for 10 minutes.

[0063] The concentration was determined using an ELISA kit. The kit was based on sandwich enzyme-linked immunosorbent assay (ELISA) technology. For this purpose, a 96-well plate was precoated with an antibody (first GPCR-specific capture antibody) against the protein to be measured. Standards, test samples, and biotin-conjugated antibody (second GPCR-specific detection antibody) were added to the wells and incubated. The respective (intermediate) incubation times varied between kits. Subsequently, the HRP (horse radish peroxidase)-conjugated reagent was added, and the entire plate was incubated again. Unbound conjugates were removed in each step with wash buffer. TMB substrate was used to quantify the HRP enzyme reaction.After adding the TMB substrate, a blue-colored product was obtained only in wells containing sufficient amounts of the respective measured protein (GPCR). This product turned yellow upon addition of the acidic stop solution. The intensity of the yellow color is proportional to the amount of GPCR bound to the plate. The optical density (OD) was measured spectrophotometrically at 450 nm in a microplate reader, from which the concentration of GPCRs can be calculated.

[0064] The results shown in Fig. 7 correspond to the derivations of the concentrations from the spectroscopically measured optical density in the microplate reader at 450 nm (top row).

[0065] In the lower row of Fig. 7, the measured values ​​shown in the upper row were divided by the urine creatinine concentration of the respective subject measured in the routine laboratory.

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Claims

Patent claims 1. A method for the diagnosis and / or monitoring of acute kidney injury, wherein the concentration of at least one G protein-coupled receptor selected from GPR40, GPR43, GPR91 and GPR120 is measured in a urine sample and compared with a normal value.

2. The method according to claim 1, wherein the respective concentration of the following combinations of two G protein-coupled receptors is measured: GPR40 and GPR43; GPR40 and GPR91; GPR40 and GPR120; GPR43 and GPR91; GPR43 and GPR120; and GPR91 and GPR120.

3. The method according to claim 1, wherein the respective concentrations of the following combinations of three G protein-coupled receptors are measured: GPR40, GPR43, and GPR91; GPR40, GPR43, and GPR120; GPR40, GPR91, and GPR120; and GPR43, GPR91, and GPR120.

4. The method according to claim 1, wherein the concentration of all G protein-coupled receptors GPR40, GPR43, GPR91 and GPR120 is measured.

5. A method according to any one of the preceding claims, wherein the urine sample is centrifuged prior to the concentration measurement in order to separate cellular components.

6. Method according to one of the preceding claims, wherein the diagnosis and / or monitoring of the acute kidney injury comprises at least one of the following steps: (1) Risk stratification of patients, for example in planned operations; (2) Evidence of acute kidney injury; (3) Evidence of acute kidney injury in the presence of chronic kidney disease, ie evidence of acute kidney injury supersedes chronic injury; (4) prediction of renal impairment, e.g., a reduction in excretory renal function; (5) Diagnosis of kidney damage; (6) quantification of the severity of kidney injury; (7) Monitoring of kidney damage and (8) Predicting the probability of recovery from kidney injury .

7. Method according to one of the preceding claims, wherein the measured concentrations of GPR40, GPR43, GPR91 and / or GPR120 are normalized to creatinine (concentration in urine).

8. A method according to any one of the preceding claims, wherein the urine originates from a mammal.

9. Method according to one of the preceding claims, wherein the concentrations of GPR40, GPR43, GPR91 and / or GPR120 are measured using an antibody-based method, for example ELISA.

10. Use of at least one G-protein-coupled receptor selected from GPR40, GPR43, GPR91 and GPR120 for the diagnosis and / or monitoring of acute kidney injury, wherein the concentration of at least one of these G-protein-coupled receptors is measured in a urine sample and compared with a normal value.

11. G protein-coupled receptor selected from GPR40, GPR43, GPR91 and GPR120 for the diagnosis and / or monitoring of acute kidney injury.

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