Biomarker for detecting tubulointerstitial disorder and use thereof

Abstract

Provided are a novel biomarker that can simply detect a tubulointerstitial disorder or grasp a disease state of a kidney disease using a urine specimen and use thereof. This biomarker is sCD14-ST in urine.

Description

Biomarkers for detecting tubulointerstitial damage and their uses

[0001] The present invention relates to a biomarker for detecting tubulointerstitial damage and its use.

[0002] The kidney's functions can be broadly divided into fluid regulation, waste excretion, and hormone production. Excretion of waste products through urine also maintains appropriate electrolyte concentrations in body fluids and maintains a slightly alkaline state. The kidney produces the hormone erythropoietin, which produces blood, and is also responsible for activating vitamin D. Therefore, a decline in kidney function can lead to high blood pressure, edema, proteinuria, hematuria, anemia, and hypocalcemia. Chronic kidney disease (CKD) is a condition in which kidney function gradually declines, and the process is similar in many cases regardless of cause. Early kidney failure gradually progresses to renal damage, ultimately resulting in kidney failure and the need for renal replacement therapy (kidney transplantation or dialysis). Renal function was assessed by the estimated glomerular filtration rate (eGFR; calculated from serum creatinine (Cr) concentration, age, and sex), urine protein, hematuria, and renal imaging. ２CKD is diagnosed when the blood pressure drops below 100°C or abnormalities persist for more than three months. An estimated 13 million CKD patients in Japan are on dialysis, with approximately 350,000 receiving dialysis nationwide. Renal function declines with age, and currently, there is no treatment that can directly restore kidney function. While chronic glomerulonephritis was previously the most common cause of kidney function decline, diabetic nephropathy and nephrosclerosis caused by hypertension are now on the rise due to lifestyle changes. However, without a treatment that can restore kidney function, CKD treatment focuses on (1) preventing the progression of renal failure, (2) reducing the risk of cardiovascular disease, (3) treating symptoms (renal failure symptoms and complications), and (4) preparing for future renal replacement therapy (transplantation, dialysis). The primary goal of CKD treatment is to prevent end-stage renal failure. Therefore, CKD treatment involves lifestyle modification and dietary therapy. It is also important to treat hypertension and diabetes, which can worsen kidney function. Ideally, people with CKD should limit their daily salt intake to less than 6g. It is also important to be careful not to take in too much protein or potassium. Furthermore, it is important to be careful about dehydration and energy intake, and maintaining compliance in daily life is important.

[0003] The best way to understand the cause and progression of CKD is through a definitive diagnosis by renal biopsy. However, renal biopsy is limited in the patients eligible for it due to factors such as the condition of the kidneys and the patient's risk of bleeding. Furthermore, a 4-5 day hospital stay, a self-pay cost of 40,000 to 70,000 yen (30% copayment), and the risk of post-biopsy bleeding place a significant burden on patients. Therefore, less than half of patients for whom a definitive diagnosis by renal biopsy is recommended can actually undergo it. It has been reported that only approximately 5% of patients undergoing dialysis have been definitively diagnosed by renal biopsy. Maintaining renal function and slowing its decline are important to avoid the need for dialysis, and early intervention is necessary. While a renal biopsy is desirable to diagnose the cause and determine treatment options, it is currently difficult to perform. Current diagnostic methods use indicators such as proteinuria (albuminuria), hematuria, renal biopsy, serum creatinine (Cr), and eGFR levels. Indicators other than imaging diagnosis by renal biopsy are markers that reflect the overall picture of renal failure, but for patients with kidney disease who require detailed examination of the state of renal function, they do not have the diagnostic sensitivity to accurately diagnose the location and degree of damage (especially tubulointerstitial damage), and in reality, diagnoses are made comprehensively from test and clinical findings.As a result of delays in renal biopsy, or in patients who cannot undergo renal biopsy due to some risk factors, it can take too long to start effective treatment, resulting in poor treatment outcomes.

[0004] Despite rapid advances in diagnosis and treatment, approximately 35,000 people are forced to undergo dialysis treatment each year. Reducing the number of patients transitioning to dialysis is a challenge that must be overcome, both from a patient and medical economic perspective. CKD is a lifestyle-related disease, such as hypertension and diabetes, and many of these conditions require lifestyle management. Understanding the condition on a monthly basis after onset is necessary. The current situation is that there is no way to prevent dialysis without early diagnosis and early intervention for CKD. To address this issue, there has been particular emphasis in recent years on the need to stage the severity of CKD and identify the condition earlier, allowing for earlier intervention.

[0005] Currently, CKD is usually diagnosed using eGFR, which is calculated using elevated serum Cr as an indicator. However, eGFR only indicates the state of renal function and does not indicate the cause of CKD, so there is insufficient information for selecting a treatment method. Therefore, eGFR is insufficient as a diagnostic marker for CKD, and the establishment of a biomarker that can evaluate the cause of dysfunction is urgently needed. In addition to the complex patient background and diverse etiology of CKD, the lack of biomarkers that can diagnose the pathology of CKD early makes early intervention difficult.

[0006] Acute kidney injury (hereinafter referred to as "AKI") is a syndrome that presents with a rapid decline in kidney function and tissue damage over a period of several hours to several days, and is classified into three types: "prerenal renal failure," in which blood flow to the kidneys is reduced due to renal dysfunction; "renal renal failure," in which abnormalities occur in the kidneys themselves; and "postrenal renal failure," in which the kidneys are damaged by blockage of the urinary tract (renal pelvis, ureter, bladder, and urethra) through which urine produced in the kidneys flows. Symptoms include decreased urine output, swelling (edema), loss of appetite, and general fatigue, and blood tests reveal high levels of blood urea nitrogen (BUN), serum creatinine (Cr), and potassium (K). The first cause is reduced blood flow to the kidneys due to dehydration or bleeding (prerenal). The second cause is reduced renal function due to renal inflammation or tubular cell damage (renal). The third cause is obstruction of the urinary tract (postrenal). For both prerenal and postrenal disorders, renal function can often be restored by identifying the cause and providing appropriate symptomatic treatment. However, in severe cases of renal dysfunction, hemodialysis is typically performed to remove waste products and excess fluid from the body. While acute kidney injury can often be restored to normal kidney function with appropriate treatment, acute kidney injury occurring in intensive care units, such as kidney damage associated with sepsis, generally has a poor prognosis. It is necessary to quickly identify the cause and select the appropriate treatment, but rapid changes in symptoms and bleeding tendency make renal biopsy difficult, making it difficult to determine the type and extent of tissue damage. As a result, treatment options are limited, with dialysis often being the only emergency response option. In recent years, it has also been recognized that AKI is not a reversible condition, but rather a long-term risk factor for the onset and progression of CKD, and it has been known that there are issues that need to be resolved in order to improve the prognosis of AKI.

[0007] Recently, the usefulness of neutrophil gelatinase-associated lipocalin (NGAL; Non-Patent Document 1) and liver-type fatty acid-binding protein (L-FABP; Non-Patent Document 2) has been reported as urinary biomarkers capable of diagnosing AKI and CKD. However, these are insufficient for understanding renal damage and severity for timely intervention such as treatment, which is required in clinical practice. Therefore, new diagnostic biomarkers for kidney disease and biomarkers capable of evaluating the cause of kidney disease pathology have been desired.

[0008] It has been disclosed that sepsis can be diagnosed by measuring sCD14-ST (hereinafter also referred to as Presepsin (registered trademark)) in a sample (Non-Patent Document 1, Patent Document 1). It has also been disclosed that measuring sCD14-ST using its specific antibody (Patent Document 1, Patent Document 2). Furthermore, canine studies have reported that blood presepsin is excreted in urine via renal metabolism as a physiological mechanism (Patent Document 3). Furthermore, it has been reported that blood presepsin levels increase with increasing renal failure (GFR stage classification) and show a negative correlation with eGFR levels (Non-Patent Document 2). It is generally believed that excretion of renal metabolic substances is restricted in renal failure, and that excretion also decreases with decreasing urine volume, resulting in an increase in blood concentration. It has also been assumed that blood presepsin levels also increase with a decrease in eGFR. In fact, it has been reported that correcting presepsin levels with blood creatinine can correct for the effects of renal failure and does not affect the diagnosis of sepsis (Non-Patent Document 3).

[0009] Japanese Patent No. 4040666 Japanese Patent Application Laid-Open No. 2005-106694 International Publication No. 2012157751

[0010] J Infect Chemother 2005;11:234-238. PLoS One. 2015;10(6):e0129159. The Journal of Medical Investigation 2021;68(1.2):105-111

[0011] Under the above circumstances, an objective of the present invention is to provide a novel biomarker that enables simple detection of tubulointerstitial damage using a urine sample, and uses thereof. Another objective of the present invention is to provide a novel biomarker that enables simple understanding of the pathology of kidney disease using a urine sample, and uses thereof.

[0012] In light of the above-mentioned problems, the present inventors, despite prior knowledge, focused on blood presepsin and urinary presepsin (hereinafter also referred to as uP-SEP) and intensively investigated their potential as biomarkers. As described in the Examples below, the results showed that urinary presepsin was clearly associated with the degree of renal damage and changed sharply to reflect the degree of damage. Furthermore, in CKD patients, urinary presepsin showed a clear negative correlation with eGFR values ​​even after creatinine correction, reflecting the severity of renal failure, whereas blood presepsin showed no correlation with eGFR values ​​after creatinine correction. It was a surprising finding that blood presepsin did not reflect the degree of renal damage, and only urinary presepsin was significantly correlated with the degree of renal damage.

[0013] As reported in Non-Patent Document 3, it was previously believed that presepsin is produced at the site of infection, and that blood presepsin concentrations are affected by factors such as urine volume but are not directly related to kidney disease. Furthermore, because urinary presepsin was presumed to be a metabolic product of blood presepsin, changes in urinary presepsin levels were thought to reflect blood presepsin levels. The results of the inventors' studies were surprising, as they could not be predicted from known findings, and demonstrated that urinary presepsin levels can be used to detect kidney damage and understand the pathology of kidney disease, and further, can specifically detect tubulointerstitial damage.

[0014] Furthermore, we investigated the potential of urinary presepsin as a biomarker. We examined urinary presepsin concentrations in patients with various kidney diseases who underwent renal biopsy. Compared with healthy individuals, urinary presepsin concentrations were elevated in all patients, with the highest levels observed in patients with interstitial nephritis and tubulointerstitial damage. Furthermore, elevated urinary presepsin concentrations were observed in patients with kidney diseases that have been reported to have a high degree of tubulointerstitial damage (e.g., kidney disease caused by ANCA-associated vasculitis), while the lowest increase was observed in patients with IgA nephropathy, which is thought to have a relatively low degree of tubulointerstitial damage. These results suggest that urinary presepsin concentrations can be used to assess the degree of tubulointerstitial damage in various kidney diseases. Furthermore, similar results were obtained when urinary presepsin / creatinine (uP-SEP / Cr) values ​​were corrected for urinary creatinine to eliminate the influence of urine volume on urinary presepsin values, demonstrating that uP-SEP is a specific marker for tubulointerstitial damage.

[0015] Furthermore, the sensitivity of uP-SEP / Cr in distinguishing tubulointerstitial nephritis (also called interstitial nephritis) was 85.7%, the specificity was 66.2%, and the AUC of the ROC curve was 0.826, demonstrating unprecedented and extremely usefulness of urinary presepsin as a single biomarker for detecting interstitial nephritis. Furthermore, based on the findings of renal damage in renal biopsy, urinary presepsin was able to significantly distinguish between groups with interstitial damage and groups without interstitial damage (including those with glomerular damage), demonstrating unprecedented and extremely usefulness of urinary presepsin as a single biomarker for detecting tubulointerstitial damage. Furthermore, the degree of increase in uP-SEP / Cr values ​​between groups in the severity of interstitial nephritis damage showed a more rapid increase with inflammatory cell infiltration, indicating that this is useful for understanding the progression of tubulointerstitial damage in the acute phase, and monitoring uP-SEP / Cr values ​​can be used to initiate treatment and assess therapeutic efficacy. Furthermore, an investigation into the relationship between the degree of tubular atrophy and the degree of inflammatory cell infiltration obtained from renal biopsy diagnosis and urinary presepsin concentrations revealed the surprising finding that urinary presepsin concentrations increased in cases with high degrees of atrophy and infiltration. Thus, it was demonstrated that tubulointerstitial damage can be easily identified at an early stage without the need for renal biopsy histology. Furthermore, the uP-SEP / Cr value can detect tubulointerstitial damage in patients with IgA nephropathy, indicating that the uP-SEP / Cr value is useful for diagnosing the severity of IgA nephropathy. Furthermore, when the correlation between the uP-SEP / Cr value and the subsequent progression of renal function was analyzed in CKD patients with diabetic nephropathy, nephrosclerosis, and other conditions who had not undergone renal biopsy, the rate of decline in eGFR indicated that patients with high uP-SEP / Cr values ​​had a higher degree of renal dysfunction, indicating that the uP-SEP / Cr value is associated with the degree of renal dysfunction in patients and can reflect prognosis. Thus, as a result of the intensive studies of the present inventors, it has become clear that urinary presepsin can be a useful indicator (biomarker) for understanding tubulointerstitial damage.

[0016] On the other hand, the results of an experiment investigating the relationship between the rate of change in eGFR values ​​and uP-SEP / Cr values ​​in patients with chronic kidney disease (CKD) revealed that the uP-SEP / Cr value is an effective indicator for understanding the severity of CKD. Furthermore, when CKD patients were observed for changes in eGFR and uP-SEP / Cr values ​​over time, a decreasing trend in eGFR was observed in patients with an increasing trend in uP-SEP / Cr values, while a decreasing trend in eGFR was observed in patients with a decreasing trend in uP-SEP / Cr values, demonstrating that the severity of CKD patients can be understood and their pathological condition can be monitored by changes in the uP-SEP / Cr value. Furthermore, in CKD patients diagnosed with drug-induced nephropathy, uP-SEP / Cr values ​​decreased and eGFR levels increased after drug discontinuation. This indicates that by observing changes in uP-SEP / Cr values, the severity of CKD patients can be assessed and treatment options, such as changing or discontinuing medication, can be selected. This demonstrates that uP-SEP / Cr values ​​can detect tubulointerstitial damage, making them useful as markers of tubulointerstitial damage in CKD patients and useful for patient monitoring. Furthermore, because uP-SEP / Cr values ​​can detect tubulointerstitial damage, they can also be useful as markers of tubulointerstitial damage in AKI patients, demonstrating their usefulness in understanding their pathological condition.

[0017] As described above, as a result of intensive studies by the present inventors, it has been found that urinary presepsin is an extremely excellent biomarker for detecting tubulointerstitial damage. Based on the above findings, the present inventors have completed the present invention.

[0018] The present invention relates to the following: [1] A method for detecting tubulointerstitial damage by measuring the urinary sCD14-ST concentration of a test subject (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level). [2] The method of [1], wherein the tubulointerstitial damage is chronic tubulointerstitial damage. [3] The method of [1] or [2], wherein the test subject is a patient suspected of having a kidney disease or a patient with such a kidney disease. [4] The method of [1] or [2], wherein the test subject is a patient suspected of having one or more kidney diseases selected from the group consisting of nephrosclerosis, IgA nephropathy, diabetic nephropathy, membranous nephropathy, ANCA-associated vasculitis, focal segmental glomerulosclerosis, minimal change nephrotic syndrome, lupus nephritis, tubulointerstitial nephritis, Sjogren's syndrome, vascular kidney disease, idiopathic TINU syndrome, and IgG4-associated nephritis or a patient with such a kidney disease. [5] The method of [1], wherein the subject is a patient suspected of having a kidney disease such as acute tubulointerstitial nephritis or chronic tubulointerstitial nephritis, or a patient with such kidney disease. [6] The method of [1] or [5], wherein the subject is a patient suspected of having a kidney disease such as acute drug-induced tubulointerstitial nephritis or chronic drug-induced tubulointerstitial nephritis, or a patient with such kidney disease. [7] A method for monitoring tubulointerstitial damage by measuring the urinary sCD14-ST concentration of a subject (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level). [8] A method for selecting a treatment for tubulointerstitial damage by measuring the urinary sCD14-ST concentration of a subject (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level). [9] A method for determining the severity of kidney disease by measuring the sCD14-ST concentration in the urine of a subject (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level).

[10] A method for detecting kidney disease by measuring the sCD14-ST concentration in the urine of a subject (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level).

[11] A method for detecting kidney disease by measuring the urinary sCD14-ST concentration of a subject (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level), wherein the kidney disease is one or more kidney diseases selected from the group consisting of nephrosclerosis, IgA nephropathy, diabetic nephropathy, membranous nephropathy, ANCA-associated vasculitis, focal segmental glomerulosclerosis, minimal change nephrotic syndrome, lupus nephritis, tubulointerstitial nephritis, Sjogren's syndrome, vascular renal disease, idiopathic TINU syndrome, and IgG4-associated nephritis.

[12] A method for detecting tubulointerstitial nephritis by measuring the urinary sCD14-ST concentration of a subject (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level).

[13] A method for detecting tubulointerstitial damage, comprising the steps of: measuring the sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) in urine collected from a patient suspected of having tubulointerstitial damage or a patient having tubulointerstitial damage; and determining that the patient has tubulointerstitial damage when the sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) is higher than that of a healthy subject.

[14] A method for detecting and treating tubulointerstitial damage, comprising the steps of: measuring the sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) in urine collected from a patient suspected of having tubulointerstitial damage or a patient who has tubulointerstitial damage; determining that there is tubulointerstitial damage when the sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) is higher than that of a healthy subject; and administering immunosuppressive therapy when it is determined that there is tubulointerstitial damage.

[15] A method for detecting kidney disease, comprising the steps of: measuring the sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) in urine collected from a patient suspected of having kidney disease or a patient with kidney disease; and determining that the patient has kidney disease when the sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) is higher than that of a healthy subject.

[16] A method for detecting and treating kidney disease, comprising the steps of: measuring the sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) in urine collected from a patient suspected of having kidney disease or a patient with kidney disease; determining that the patient has kidney disease when the sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) is higher than that of a healthy subject; and administering immunosuppressive therapy if the patient is determined to have kidney disease.

[17] Any of the methods [1] to

[16] , wherein the sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) is measured by an immunological measurement method.

[18] Use of the urinary sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) as a marker for detecting tubulointerstitial damage.

[19] A kit for detecting tubulointerstitial damage, comprising: (a) an antibody specific to sCD14-ST, (b) standard data showing the correlation between the urinary sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) and the degree of kidney damage, and (c) an instruction manual.

[20] A kit for detecting tubulointerstitial damage, comprising: (a) an antibody specific to sCD14-ST, (b) standard data showing the correlation between the urinary sCD14-ST concentration (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) and the severity of kidney disease, and (c) an instruction manual.

[0019] As used herein, "human sCD14-ST" (also known as Presepsin (registered trademark)) refers to the "soluble CD14 antigen of the first aspect" described in Japanese Patent No. 4,040,666, and more specifically, is a soluble CD14 antigen having the following properties 1) to 3): 1) a molecular weight of 13±2 kDa in SDS-PAGE under non-reducing conditions, 2) an N-terminal sequence having the amino acid sequence of SEQ ID NO: 1, and 3) specific binding to an antibody produced using the peptide consisting of 16 amino acid residues described in SEQ ID NO: 2 as an antigen. Unless otherwise specified, "sCD14-ST" used herein refers to human sCD14-ST.

[0020] SEQ ID NO: 1: Thr Thr Pro Glu Pro Cys Glu Leu Asp Asp Glu 1510 SEQ ID NO: 2: Arg Val Asp Ala Asp Ala Asp Pro Arg Gln Tyr Ala Asp Thr Val Lys 151015

[0021] "Tubular interstitial damage" refers to tubular interstitial damage that allows evaluation of the cause of the pathology of kidney disease, such as tubular interstitial damage in which inflammatory cell infiltration into the interstitium and tubular atrophy are observed based on findings of damage evaluated by kidney biopsy. The degree of tubular interstitial damage can be evaluated histologically by measuring the degree of inflammatory cell infiltration into the interstitium and tubular atrophy, by scoring based on the Banff classification, or by the Oxford classification of tubular atrophy and interstitial fibrosis (T-score). Furthermore, tubular interstitial damage also includes acute tubular interstitial damage or chronic tubular interstitial damage, as well as drug-induced tubular interstitial damage and non-drug-induced tubular interstitial damage.

[0022] "Detection of tubulointerstitial damage" is not limited to ascertaining tubulointerstitial damage for the purpose of evaluating the cause of the pathology of kidney disease, and includes, for example, detecting the presence or absence of tubulointerstitial damage and detecting the severity of the damage. Furthermore, "detection of tubulointerstitial damage" as used herein does not exclude the possibility that other kidney damage may occur when detecting tubulointerstitial damage. Depending on the type and severity of kidney disease, tubulointerstitial damage may occur concomitantly with other kidney damage.

[0023] "Severity of renal damage" refers to the degree of renal damage that can be used to evaluate the cause of the pathology of kidney disease, and is classified based on the findings of damage evaluated by kidney biopsy according to guidelines provided by the Japanese Society of Nephrology and various academic literature. For example, the degree of damage in tubulointerstitial damage is classified based on the degree of inflammatory cell infiltration into the interstitium and atrophy of the renal tubules.

[0024] "Severity" is an index for evaluating the pathology and stage of kidney disease as classified in the CKD treatment guidelines and AKI treatment guidelines set forth by the Japanese Society of Nephrology, etc. It can also be used to select ways to deal with and treat kidney disease.

[0025] Kidney disease is a condition in which the kidneys' glomeruli and tubules are damaged, causing a decline in kidney function. There are two types of kidney disease: acute kidney failure, in which symptoms appear suddenly and worsen over the course of hours or days, and chronic kidney failure, which progresses slowly and often causes no noticeable symptoms until the condition is nearing the end stage.

[0026] Chronic renal failure, characterized by chronic progressive kidney damage, has recently been referred to as chronic kidney disease (CKD). CKD has a wide range of causes, including a close association with lifestyle-related diseases (e.g., diabetes, hypertension) and metabolic syndrome, such as chronic nephritis. It is known that there are approximately 13 million CKD patients in Japan (one in eight adults aged 20 years or older). The severity of CKD is recommended to be assessed using the CGA classification based on cause (C), renal function (GFR: G), and proteinuria (albuminuria: A), as outlined in the "CKD Clinical Practice Guidelines 2012" published by the Japanese Society of Nephrology in 2012. Previously, the stage of chronic kidney disease was assessed solely by renal function, as determined by eGFR. However, the accuracy and validity of diagnosis have been further enhanced by combining the level of renal function with underlying conditions, such as diabetes and hypertension, and the state of urinary protein.

[0027] Treatments for chronic kidney disease (CKD) include lifestyle modification, dietary therapy, and drug therapy for advanced stages 2-4, while dialysis and kidney transplantation are used for end-stage stage 5 (renal failure). Regarding lifestyle modification, the Japanese Society of Nephrology (JSNE) guidelines recommend avoiding excessive exercise, avoiding fatigue, taking precautions against colds and other infections, avoiding exposure to colds, limiting consumption of recreational products, and paying attention to diet. Dietary therapy includes adequate calorie intake, limiting protein, salt, potassium, and phosphorus, and consuming an appropriate amount of fluid. Pharmacological therapy includes antihypertensive drugs, diuretics, phosphate binders, potassium binders, erythropoietin preparations, steroids / immunosuppressants, and herbal medicines, depending on the patient's condition and underlying disease. Regardless of the above, various clinical guidelines for chronic kidney disease have been published by the JSNE and other organizations, and can be used as reference for diagnosis, treatment, and prevention.

[0028] Acute renal failure is characterized by rapidly progressing kidney damage and has recently been referred to as acute kidney injury (AKI). AKI is a broad clinical syndrome defined as a "sudden decline in renal function" and is classified into three categories based on the pathology: prerenal, renal, and postrenal. Because treatment strategies for AKI vary depending on the cause, disease differentiation is important. Guidelines have been proposed with the aim of unifying the definition of AKI and promoting early diagnosis. In 2012, Kidney Disease: Improving Global Outcomes (KDIGO) published the AKI Clinical Practice Guidelines (KDIGO Guidelines), which proposed the KDIGO criteria, integrating the previous RIFLE and AKIN criteria. In Japan, the AKI Clinical Practice Guidelines 2016 published by the Japanese Society of Nephrology and others also recommends the use of the KDIGO criteria for diagnosing AKI. In the KDIGO criteria, AKI is defined by an increase in serum creatinine levels and a decrease in urine output, and the severity of AKI is classified into stages 1 to 3. It is important to measure serum creatinine and urine output at the appropriate time, and to assess the appropriate severity early on, along with medical history, physical examination, blood and urine test results, and imaging tests (ultrasound, CT, etc.), and to provide management stratified by disease stage. It is said that even after AKI has improved, continued evaluation should be conducted to check for the onset of new CKD or the worsening of existing CKD.

[0029] The treatment of AKI begins with assessing the severity and degree of complications and determining whether emergency treatment is necessary. If AKI manifests with hyperkalemia, metabolic acidosis, or severe overflow, immediate treatment, including drug therapy, is considered, regardless of the cause, and dialysis may be initiated. At the same time, the cause of AKI is identified based on the patient's medical history, physical examination, and laboratory findings, and treatment is initiated immediately. For example, in patients with prerenal acute kidney injury caused by dehydration or hypotension, treatment is initiated with fluid infusion and, if necessary, vasopressors, in parallel with treatment for the underlying cause. In patients with renal acute kidney injury caused by vasculitis (rapidly progressive glomerulonephritis), immunosuppressive therapy, such as steroids, or plasma exchange therapy, aimed at removing antibodies that cause vasculitis, may be considered. Until AKI resolves, appropriate fluid infusion and nutritional management are provided, avoiding medications that interfere with renal function, to avoid adverse effects on the kidneys. In addition, if postrenal acute kidney injury due to urinary tract obstruction is present, treatment is performed to remove the obstruction to the urinary tract. Regardless of the above, various clinical guidelines for acute kidney injury have been published by the Japanese Society of Nephrology and other organizations, and can be used as reference for diagnosis, treatment, prevention, etc.

[0030] The method of the present invention allows for easy detection of tubulointerstitial injury in its early stages, regardless of whether a renal biopsy has been performed. Unlike known biomarkers for detecting kidney injury, urinary presepsin specifically detects tubulointerstitial injury and exhibits unprecedented, highly usefulness as a single biomarker for detecting tubulointerstitial nephritis. Furthermore, the method of the present invention allows for easy detection of tubulointerstitial injury, regardless of whether a renal biopsy has been performed, assessment of the injury and its severity, and selection of a treatment method based on the injury. Furthermore, the method of the present invention allows for easy monitoring of tubulointerstitial injury, assessment of the injury trend, and selection of a treatment method based on the injury trend, regardless of whether a renal biopsy has been performed. Furthermore, the method of the present invention allows for easy assessment of the severity of chronic kidney disease, regardless of whether a renal biopsy has been performed. Furthermore, treatment methods can be selected based on the severity and the trend of the severity. Furthermore, the method of the present invention allows for easy assessment of the pathology of acute kidney injury, regardless of whether a renal biopsy has been performed. Furthermore, prognosis can be predicted based on the pathology and the trend of the pathology, and treatment methods can be selected accordingly. The biomarkers of the present invention can be used in the methods of the present invention, and the kits of the present invention can be used in the methods of the present invention.

[0031] This is a calibration curve prepared by adding a presepsin standard (manufactured by LSI Medience) to a urine sample from a healthy subject and then serially diluting the sample with the same urine for use with an automated chemiluminescent enzyme immunoassay system (PATHFAST). This is a chromatograph comparing the elution positions of urinary presepsin from a healthy subject detected by presepsin ELISA with those of a presepsin standard. This is a graph showing the results of comparing eGFR, urinary presepsin (uP-SEP / gCr), and blood presepsin (sP-SEP / Cr) in CKD patients. This is a graph showing the results of comparing urinary P-SEP concentrations (actual measured values) for various kidney diseases for which kidney biopsies were performed. AAV: ANCA-associated vasculitis. DMN: diabetic nephropathy. FSGS: focal segmental glomerulosclerosis. IgAN: IgA nephropathy. IN: interstitial nephritis. LN: lupus nephritis. MCNS: minimal change nephrotic syndrome. MN: membranous nephropathy. N: healthy individuals. A graph showing the results of ROC analysis in which the cutoff value and diagnostic ability (actual measured value) for distinguishing between four types of kidney disease were determined using AAV, DMN, lupus nephritis, interstitial nephritis, and healthy individuals as the population. A graph showing the results of an intergroup comparison between each disease using uP-SEP / Cr. A graph showing the results of ROC analysis of uP-SEP / Cr in detecting interstitial nephritis. A graph showing the results of a comparison between findings (present / absent) and uP-SEP / Cr values ​​in kidney biopsies. A graph showing the results of a comparison between patients who underwent kidney biopsy, classified into a group with interstitial damage (Mild or higher), a group without interstitial damage and without glomerular damage, and a group without interstitial damage and with glomerular damage. Graph showing the results of comparing the degree of inflammatory cell infiltration and tubular atrophy determined by renal biopsy with the uP-SEP / Cr value (vs. Minimal). Graph showing the results of comparing the degree of inflammatory cell infiltration and tubular atrophy in interstitial damage with the uP-SEP / Cr value. Images of a normal renal biopsy case (left) and an interstitial nephritis case (right) used for assessment. Graph showing the results of comparing the T-score and uP-SEP / Cr value in the Oxford classification of patients with IgA nephropathy (cutoff value: T-score>0). Graph showing the results of comparing the treatment course (eGFR value) and uP-SEP / Cr value of a CDK patient (uP-SEP / Cr value is shown in circle).

[0032] Hereinafter, embodiments of the present invention will be described in detail, mainly using a method for detecting tubulointerstitial damage or a method for detecting kidney disease as an example, but the mode of use is not limited to this. For example, the present invention includes: a method for detecting tubulointerstitial damage or a method for detecting kidney disease, which involves measuring the sCD14-ST concentration in the urine of a test subject (preferably the value obtained by dividing the sCD14-ST concentration in urine by the creatinine level in urine); a method for assisting in the detection of tubulointerstitial damage or a method for assisting in the detection of kidney disease, which involves measuring the sCD14-ST concentration in the urine of a test subject (preferably the value obtained by dividing the sCD14-ST concentration in urine by the creatinine level in urine); a method for measuring the sCD14-ST concentration in the urine of a test subject (preferably the value obtained by dividing the sCD14-ST concentration in urine by the creatinine level in urine) for detecting tubulointerstitial damage or kidney disease; an in vitro method for detecting tubulointerstitial damage or a kidney disease, which involves measuring the sCD14-ST concentration in the urine of a test subject (preferably the value obtained by dividing the sCD14-ST concentration in urine by the creatinine level in urine); Use of an antibody specific to sCD14-ST in the manufacture of a kit for detecting tubulointerstitial damage or in the manufacture of a kit for detecting kidney disease; and a method for measuring the sCD14-ST concentration in the urine of a test subject (preferably the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level) in order to provide information necessary for detecting tubulointerstitial damage or kidney disease.

[0033] The present invention also includes the use of an antibody specific to sCD14-ST in the manufacture of a kit for use in the various methods of the present invention. Furthermore, the various methods of the present invention can be carried out either in vitro or in vivo, but are preferably carried out in vitro.

[0034] The detection method of the present invention (hereinafter referred to as the "method of the present invention") involves measuring urinary presepsin collected from a subject. Examples of subjects of the present invention include patients suspected of having or having tubulointerstitial damage. Specifically, examples include patients suspected of having or having acute tubulointerstitial damage, patients suspected of having or having chronic tubulointerstitial damage, patients suspected of having or having non-drug-induced tubulointerstitial damage, and patients suspected of having or having drug-induced tubulointerstitial damage. For example, drug-induced tubulointerstitial damage is known to be caused by immunosuppressants such as steroids, NSAIDs such as aspirin and anti-inflammatory analgesics, antibiotics, proton pump inhibitors (PPIs), and the like. Drug-induced tubulointerstitial damage is classified into acute drug-induced tubulointerstitial damage, which occurs acutely, and chronic drug-induced tubulointerstitial damage, which occurs after the transition from acute to chronic conditions. The method of the present invention can detect any type of tubulointerstitial damage.

[0035] Furthermore, chronic tubulointerstitial damage often progresses without exhibiting clinical symptoms, and the ability of the method of the present invention to detect tubulointerstitial damage, regardless of whether or not a renal biopsy is performed, is a very useful effect. Kidney diseases associated with chronic tubulointerstitial damage include chronic kidney disease (CKD), specifically nephrosclerosis, IgA nephropathy, diabetic nephropathy, membranous nephropathy, ANCA-associated vasculitis, focal segmental glomerulosclerosis, minimal change nephrotic syndrome, lupus nephritis, tubulointerstitial nephritis (also referred to as interstitial nephritis), Sjögren's syndrome, vascular renal disease, idiopathic TINU syndrome, IgG4-associated nephritis, etc. In particular, the method is preferred because it can differentiate between tubulointerstitial nephritis, nephrosclerosis, diabetic nephropathy, membranous nephropathy, ANCA-associated vasculitis, minimal change nephrotic syndrome, lupus nephritis, etc., which have severe damage to the tubulointerstitial structure. In particular, tubulointerstitial nephritis is highly preferable because it can be differentiated based on the urinary presepsin concentration alone. Note that the "urinary presepsin concentration" includes not only the urinary presepsin concentration before correction with urinary creatinine, but also, for example, the urinary presepsin concentration / urinary creatinine (uP-SEP / Cr) value, which is a value corrected with urinary creatinine.

[0036] Furthermore, kidney diseases with acute tubulointerstitial damage include acute kidney injury (AKI), particularly prerenal and renal pathologies with damage to the renal tubules or interstitium.

[0037] The method of the present invention is also preferred because it can detect tubulointerstitial damage regardless of the type of kidney disease and determine the type and severity of the damage. Furthermore, it is also preferred because it allows for the selection of a treatment based on the type and severity of the damage. Such subjects include, for example, patients with kidney disease requiring treatment with immunosuppressants such as steroids, patients with severe kidney disease (such as renal failure), and patients who have undergone kidney transplantation. Treatments for kidney disease include lifestyle modification, dietary therapy, and drug therapy, which can be appropriately selected and combined depending on the pathology. However, the method of the present invention enables the easy and early detection of the progression of tubulointerstitial damage and the worsening of kidney disease, thereby allowing for the selection of a treatment more tailored to the patient's pathology. Furthermore, it is also preferred because it allows for the selection of a treatment based on the use of steroids, which may have adverse side effects, regardless of whether or not a kidney biopsy is performed. Furthermore, monitoring urinary presepsin concentrations is also preferred because it allows for the identification of the progression of tubulointerstitial damage and the worsening of kidney disease, allowing for the selection of a treatment based on these trends. Those skilled in the art can appropriately design and use methods for detecting and determining tubulointerstitial damage and kidney disease based on publicly known information.

[0038] Methods for measuring presepsin are known and can be performed using various known protein analytical methods, such as immunological analytical methods using antibodies and biochemical analytical methods such as electrophoresis, or automated analyzers for clinical testing. Analytical methods using substances with properties similar to antibodies, such as RNA aptamers, are also included in the present invention.

[0039] For example, Japanese Patent No. 4,040,666 discloses a method for measuring human sCD14-ST, more specifically, a sandwich EIA system [Example 7-(1) of Japanese Patent No. 4,040,666] using a combination of a polyclonal antibody (S68 antibody) prepared using a peptide consisting of 16 amino acid residues set forth in SEQ ID NO: 2 (the S68 peptide described in Japanese Patent No. 4,040,666) as an antigen, a monoclonal antibody (F1146-17-2 antibody), and an anti-CD14 antigen monoclonal antibody (e.g., F1031-8-3 antibody, F1106-13-3 antibody, etc.), which can be applied to the method of the present invention. Furthermore, as shown in the Examples below, presepsin can also be measured by a chemiluminescent enzyme immunoassay using magnetic particles, using an automated chemiluminescent immunoassay system (PATHFAST; LSI Medience Corporation).

[0040] The sample used in the method of the present invention is not particularly limited as long as it allows measurement of urinary presepsin, and for example, urine collected from a subject can be used.

[0041] In the method of the present invention, changes in urinary presepsin concentration can be used as an indicator of tubulointerstitial damage. Alternatively, instead of urinary presepsin concentration, the urinary presepsin concentration / urinary creatinine (uP-SEP / Cr) value, which is the value obtained by correcting the urinary presepsin concentration with urinary creatinine, can be used. Correcting with urinary creatinine is preferable because it eliminates the influence of urine volume, etc. Unless otherwise specified, urinary presepsin concentration or the urinary presepsin concentration / urinary creatinine value will be referred to as the urinary presepsin value.

[0042] For example, as shown in Examples 5 and 6 described below, urinary presepsin levels were high in patients with diseases characterized by severe tubulointerstitial damage, whereas low levels were found in patients with mild tubulointerstitial damage. Thus, the method of the present invention can determine whether a high urinary presepsin level indicates severe tubulointerstitial damage or progression of the damage. Furthermore, the method of the present invention can determine whether a high urinary presepsin level indicates high kidney disease severity or progression of the disease. On the other hand, a low urinary presepsin level can indicate mild tubulointerstitial damage. On the other hand, a low urinary presepsin level can indicate low kidney disease severity or improvement of the disease. For example, if the urinary presepsin level in a sample is higher than the quantile (e.g., median) of healthy individuals, it can be determined whether tubulointerstitial damage is severe or whether the kidney disease is severe. Statistical methods such as Cox regression and logistic regression can also be used. Furthermore, the level of urinary presepsin can be used to determine the severity of tubulointerstitial damage or the severity of kidney disease. A previously prepared "threshold" can be used as a criterion for assessment.

[0043] For example, when the urinary presepsin concentration is higher than about 990 pg / mL, it can be determined that the patient has severe tubulointerstitial damage, or that the patient has ANCA-associated vasculitis (AAV), diabetic nephropathy (DMN), lupus nephritis, or interstitial nephritis. Alternatively, when the uP-SEP / Cr value is higher than about 1000 ng / gCr, it can be determined that the patient has severe tubulointerstitial damage, or that the patient has minimal change nephrotic syndrome (MCMS), membranous nephropathy (MN), ANCA-associated vasculitis (AAV), diabetic nephropathy (DMN), lupus nephritis, or interstitial nephritis. Furthermore, if the uP-SEP / Cr value is higher than approximately 900 ng / gCr, it can be determined that tubulointerstitial damage has occurred to a mild or severe level; if it is higher than approximately 1200 ng / gCr, it can be determined that tubulointerstitial damage has occurred to a moderate or severe level; and if it is higher than approximately 1800 ng / gCr, it can be determined that tubulointerstitial damage has occurred to a severe level.

[0044] In the method of the present invention, the threshold value of urinary presepsin levels for determining the severity of tubulointerstitial damage or the degree of severity of kidney disease, or for differentiating between tubulointerstitial damage and kidney disease, is expected to vary depending on various conditions, such as gender and age. However, a person skilled in the art can determine the threshold value for judgment by appropriately selecting an appropriate population corresponding to the subject and statistically processing the data obtained from that population. The population may be selected from a group of healthy individuals, a non-tubulointerstitial disorder group, a tubulointerstitial disorder group, a group of severity of each non-tubulointerstitial disorder, a group of severity of each tubulointerstitial disorder, a group of types of non-tubulointerstitial disorders, a group of types of tubulointerstitial disorders, a non-renal disease group, a kidney disease group, a group of severity (scale, etc.) of each non-renal disease condition, a group of severity (scale, etc.) of each kidney disease condition, a group of types (scale, etc.) of each non-renal disease condition, a group of types (scale, etc.) of each kidney disease condition, an acute group, a chronic group, a non-drug-induced group, a drug-induced group, etc.

[0045] In Example 4 described below, the threshold for detecting pathologies and diseases causing tubulointerstitial damage was determined by performing the receiver operating characteristic (ROC) analysis shown in Figure 5, resulting in an optimal cutoff value of 982 pg / mL (sensitivity = 73%, specificity = 90%, AUC: 0.869 (95% CI = 0.785-0.954)). In Example 6 described below, the threshold for detecting interstitial nephritis was determined by performing the ROC analysis shown in Figure 7, resulting in an optimal cutoff value of 1525.689 (ng / gCr) (sensitivity = 85.7%, specificity = 66.2%, AUC: 0.826 (95% CI: 0.752-0.9)). In Example 7 described below, an optimal cutoff value of 641.461 (ng / gCr) (sensitivity=66.7%, specificity=63.7%, AUC: 0.707 (95% CI: 0.563-0.852)) was determined as the threshold for detecting renal damage by performing the ROC analysis shown in Figure 8. In Example 8 described below, a threshold for detecting tubulointerstitial damage (inflammatory cell infiltration or tubular atrophy) was determined as follows by performing the ROC analysis shown in Table 4. For inflammatory cell infiltration, the optimal cutoff value for detecting Mild or higher was 922.196 (ng / gCr) (sensitivity = 64.1%, specificity = 65.5%, AUC: 0.702 (95% CI: 0.653-0.750)), and the optimal cutoff value for detecting Moderate or higher was 1210.588 (ng / gCr). (Sensitivity = 69.8%, Specificity = 66.5%, AUC: 0.665 (95% CI: 0.665-0.768)), and the optimal cutoff value of 1954.418 (ng / gCr) (Sensitivity = 80.4%, Specificity = 73.8%, AUC: 0.811 (95% CI: 0.751-0.871)) was determined as the threshold for detecting Severe or higher.Regarding tubular atrophy, the optimal cutoff value for detecting Mild or higher was 992.225 (ng / gCr) (sensitivity = 57.5%, specificity = 63.8%, AUC: 0.649 (95% CI: 0.595-0.704)), and the optimal cutoff value for detecting Moderate or higher was 1210.588 (ng / gCr) ( Sensitivity = 65.2%, specificity = 65.8%, AUC: 0.706 (95% CI: 0.656-0.756)), and the threshold for detecting severe or higher was 1776.823 (ng / gCr) as the optimal cutoff value (sensitivity = 70.3%, specificity = 71.4%, AUC: 0.760 (95% CI: 0.700-0.820)). In Example 9 described below, the threshold for detecting tubulointerstitial damage in patients with IgA nephropathy was determined by performing the ROC analysis shown in Figure 9, and the optimal cutoff value was 543.082 (ng / gCr) (sensitivity = 72.2%, specificity = 60.5%, AUC: 0.699 (95% CI: 0.575-0.823)). In the method of the present invention, a threshold value for determination is determined and the measured urinary presepsin level in a sample is compared with the threshold value for determination, thereby making it possible to detect tubulointerstitial damage regardless of whether or not a renal biopsy is performed.

[0046] In the method of the present invention, sample collection times include when tubulointerstitial damage is suspected, when the progression of tubulointerstitial damage is suspected, and after treatment. In particular, samples may be collected over time after a kidney disease accompanied by tubulointerstitial damage is suspected, before determining whether or not to perform a renal biopsy, or after a renal biopsy. For example, samples may be collected monthly, every other month, or every year after a renal biopsy. Furthermore, because sample collection can be performed as a simple alternative to a renal biopsy to evaluate the pathology, the timing of sample collection can be appropriately selected, such as daily, weekly, monthly, every other month, or every year, without placing a burden on the patient.

[0047] In a first aspect of the method of the present invention, the presence or absence and severity of tubulointerstitial damage can be determined in the early stages, regardless of whether a renal biopsy has been performed. The early stages of tubulointerstitial damage also include the stage at which tubulointerstitial damage is suspected. Detecting tubulointerstitial kidney damage is preferable because it allows for understanding the renal damage in kidney diseases with tubulointerstitial damage and therefore the severity of the disease. Furthermore, chronic tubulointerstitial damage often progresses without exhibiting clinical symptoms, and the ability of the method of the present invention to detect tubulointerstitial damage, regardless of whether a renal biopsy has been performed, is a very useful effect.

[0048] The stage at which tubulointerstitial damage is suspected refers to, for example, a stage at which clinical symptoms are not clearly manifested.Chronic kidney disease (CKD) progresses before symptoms appear, and by the time symptoms appear, it is often severe, so it is very useful to be able to detect tubulointerstitial damage early.Since the method of the present invention can be used to detect tubulointerstitial damage before the time when chronic kidney disease is recognized using conventional diagnostic guidelines, in certain embodiments, the disease state of a patient in early chronic kidney disease is confirmed earlier than when the signs of chronic kidney disease are more clinically obvious.In other words, it can also be called the stage at which a doctor has a clinical suspicion of chronic kidney disease due to clinical symptoms.

[0049] The method also includes a step of determining whether or not to perform a renal biopsy to confirm tubulointerstitial damage, etc. Conventionally, renal biopsy has been performed to evaluate renal damage in order to confirm chronic kidney disease, but in some cases, renal biopsy may pose a higher risk to the patient than the benefit, so it is necessary to carefully determine whether or not to perform a renal biopsy. According to the method of the present invention, tubulointerstitial damage can be detected using a urine sample, which is preferable because it allows the appropriate timing for renal biopsy to be selected easily and early.

[0050] Furthermore, because acute tubulointerstitial damage occurs as acute kidney injury (AKI), it progresses rapidly, and treatment of the underlying disease is generally the primary treatment. Therefore, tests that accurately assess tissue damage, such as renal biopsy performed in a resting state, cannot be performed on AKI patients. Since the method of the present invention can be used to detect tubulointerstitial damage before AKI is recognized without performing a renal biopsy, in certain embodiments, the disease state of a patient in early acute kidney injury can be confirmed retrospectively, rather than when the signs of acute kidney injury are more clinically evident. In other words, it can also be said to be the stage at which a doctor has a clinical suspicion of acute kidney injury based on clinical symptoms.

[0051] It also includes understanding the pathology of acute kidney injury from acute tubulointerstitial damage, determining the severity, and selecting an optimal treatment. In acute kidney injury, it is difficult to accurately determine tissue damage using a renal biopsy or the like, and it is therefore highly preferable to be able to detect tubulointerstitial damage using a single biomarker. According to the method of the present invention, tubulointerstitial damage can be detected using a urine sample, which is preferable because it allows for easy and early selection of a treatment.

[0052] In a second aspect of the method of the present invention, urinary presepsin can be measured after tubulointerstitial damage has been confirmed, regardless of whether a renal biopsy has been performed, to detect tubulointerstitial damage and determine the presence or absence and severity of the damage. Detecting tubulointerstitial damage is preferable because it allows for the determination of renal damage in chronic kidney disease patients with tubulointerstitial damage, thereby determining the severity of the damage. As in the first aspect, detection of tubulointerstitial damage using a urine sample is preferable because it allows for the easy and early selection of an appropriate time for renal biopsy.

[0053] Another method includes measuring urinary presepsin levels over time after performing a kidney biopsy, which is preferable because it can reduce or eliminate the need for repeated kidney biopsies.

[0054] This also includes detecting renal damage by measuring urinary presepsin levels over time in patients who do not undergo renal biopsy. Currently, renal biopsy cannot be performed when kidney disease progresses to a severe stage (e.g., stage 3, 4, 5, etc.), when renal fibrosis and sclerosis occur, when the kidney shrinks, or when a renal biopsy sample cannot be obtained due to a single kidney, or when the patient is at high risk. As a result, the inability to perform renal biopsy results in a loss of opportunity to select a treatment method tailored to the patient's condition. The method of the present invention enables simple and early detection of progression of the disease, thereby enabling the selection of a treatment method tailored to the patient's condition. Furthermore, regardless of whether a renal biopsy is performed, this method is preferable because it provides information for selecting a treatment method using immunosuppressants such as steroids based on an assessment of the severity of tubulointerstitial damage.

[0055] This is preferable because it allows the degree of kidney damage to be diagnosed as a prognosis for AKI, and therefore makes it possible to easily and early grasp the progression to CKD.

[0056] In a third aspect of the method of the present invention, the severity of kidney disease resulting from the therapeutic effect can be determined. The term "determining the severity resulting from the therapeutic effect" is not particularly limited, but can be used to monitor the therapeutic effect as follows. For example, samples are collected after a patient visit, before treatment, after treatment, and at the time when the therapeutic effect appears. If the level exceeds a set threshold despite treatment, the lack of therapeutic effect and progression of severity can be determined. Furthermore, if the level drops below the threshold after treatment, from a pathological condition that already exceeded the threshold at the patient visit, the therapeutic effect and improvement of severity can be determined.

[0057] It is preferable that the method of the present invention can grasp the severity of the condition regardless of whether or not a renal biopsy is performed. In particular, in the case of acute kidney injury, current diagnostic guidelines do not allow differentiation of tissue damage by renal biopsy or the like, and therefore the choice of treatment is limited. Therefore, it is particularly preferable that the method of the present invention can easily grasp the pathology of acute kidney injury.

[0058] The kit of the present invention can be used to carry out the method of the present invention and comprises: (a) an antibody specific to presepsin; (b) standard data showing the correlation between urinary presepsin levels and the degree of renal damage; and (c) an instruction manual.

[0059] The kit of the present invention can be used to carry out the method of the present invention and comprises: (a) an antibody specific to presepsin; (b) standard data showing the correlation between urinary presepsin levels and the severity of kidney disease; and (c) an instruction manual.

[0060] The antibody used in the kit of the present invention may be either a monoclonal antibody or a polyclonal antibody. In addition, antibody fragments that retain the ability to specifically bind to presepsin, such as Fab, Fab', and F(ab'), may also be used. ２ , or Fv, can also be used in the kit.

[0061] Furthermore, the antibody can be used in the kit as it is, or in a form appropriate for the immunological technique to be used, for example, in a state immobilized on a latex carrier if latex agglutination immunoassay is used; in a state immobilized on magnetic particles if a highly sensitive measurement method using magnetic particles or the like is used; in a state immobilized on a substrate if a method using a substrate such as immunochromatography is used; or in a labeled state if labeling with a labeling substance (e.g., an enzyme, a fluorescent substance, a chemiluminescent substance, a radioisotope, biotin, avidin) is required.

[0062] The standard data included in the kit of the present invention is not particularly limited as long as it indicates a correlation between urinary presepsin levels and the degree of renal damage or the severity of renal disease, and examples thereof include a threshold value for determination, original data for calculating the threshold value, or statistically processed data. The standard data may be included in the instruction manual or may be attached as a separate data sheet. The attached document may be in the form of paper, an electronic medium such as a CD-ROM, or a document downloaded from a website.

[0063] Furthermore, the instruction manual included in the kit of the present invention is not particularly limited as long as it at least mentions the relationship between urinary presepsin levels and the degree of renal damage or the severity of renal disease, and in addition to the above-mentioned mentions, it may also include, for example, an explanation of the procedure for performing immunological measurements using the kit of the present invention, an explanation of the procedure for predicting prognosis based on the obtained measurement values, and precautions regarding the storage and handling of the kit itself.

[0064] The present invention is not limited to the following embodiments and examples. Various modifications within the scope of the claims and within the scope that can be easily conceived by a person skilled in the art are also included in the present invention.

[0065] Example 1: Measurement of Urinary Presepsin by CLEIA Using Anti-Presepsin Antibody Urinary presepsin was measured by modifying Example 7-(1) of Japanese Patent No. 4040666, which is capable of measuring blood presepsin. Specifically, alkaline phosphatase (ALP)-labeled polyclonal antibody (S68 antibody) and monoclonal antibody (F1031-8-3 antibody) immobilized on magnetic particles (JSR Corporation) were used, and measurements were performed using an automated immunoluminescence analyzer, PATHFAST (LSI Medience Corporation). The ALP-labeled polyclonal antibody (S68 antibody) was prepared by preparing a Fab' fraction of the polyclonal antibody (S68 antibody) and conjugating it to ALP using the maleimide method. CDP-star (Applied Biosystems) was used as the luminescent substrate. In the following examples, the kit is sometimes referred to as the Presepsin PATHFAST Kit.

[0066] Measurements were performed according to the following procedure. First, a sample was reacted with a magnetic particle-immobilized antibody and an ALP-labeled antibody to form a complex between the presepsin in the sample and the two antibodies. The complex was then collected using a magnetic field, and unbound ALP-labeled antibody was removed. A luminescent substrate was added, and the amount of luminescence was detected as the amount of presepsin. Using the above measurement method, a presepsin standard (manufactured by LSI Medience Corporation) was added to a urine sample from a healthy subject and serially diluted with the same urine. A calibration curve was created using the above method, as shown in Figure 1. This demonstrates that urinary presepsin can be similarly measured using the blood presepsin measurement method.

[0067] Example 2: Specific detection of urinary presepsin To confirm whether presepsin present in human urine was actually detected, urine was concentrated, the sample was fractionated by gel filtration, and each fraction was assayed by ELISA (enzyme-linked immunosorbent assay) using an anti-presepsin antibody.

[0068] First, urine (5 mL) from two healthy volunteers was concentrated approximately 10-fold using a centrifugal filter (Amicon Ultra-15, 3000MNWL, Merck). 0.4 mL of the concentrated sample was applied to a gel filtration column (Superdex 75 Increase 10 / 300; GE Healthcare Japan) attached to a high-performance liquid chromatography system (SHIMADZU LC-10AT / SPD-6A / UV; Shimadzu Corporation), and eluted with Tris-buffered saline pH 7.6 (hereinafter referred to as T-TBS) containing 0.05% Tween-20 (Sigma-Aldrich). At 0.4 mL per fraction, 40 fractions were obtained per sample. Next, 0.4 mL of a presepsin standard (manufactured by LSI Medience Corporation) was applied to the same device and fractionated in the same manner.

[0069] Each of the obtained fractions was measured by ELISA (presepsin ELISA) prepared using the anti-presepsin antibody described in Example 1. The sandwich ELISA constructed using the anti-presepsin antibody is shown below. Specifically, F1106-13-3 antibody was diluted to 5 μg / mL in 0.05 mol / L Tris buffer solution (TBS, pH 7.6) and added to an immunoplate (Maxisorp; NUNC) at 100 μL / well. After overnight incubation at 4°C, the plate was washed three times with Tris-buffered saline (T-TBS) containing 0.05% Tween-20, and 200 μL of TBS containing 0.1% Stabiligard (Surmodics) and 0.1% Tween-20 was added to each well for blocking. Next, a dilution series of the human presepsin standard (manufactured by LSI Medience) was prepared using 0.1% bovine serum albumin (manufactured by Sigma-Aldrich) / TBS. Each fraction was measured in its original form. 100 μL of each fraction was added to each well, incubated at room temperature for 1 hour, and then washed three times in the same manner. Next, the prepared biotin-labeled S68 antibody (prepared using EZ-Link NHS-LC-Biotin (manufactured by Thermo Fisher) reagent according to the manufacturer's instructions) was diluted to 1 μg / mL with T-TBS and added to each well at 50 μL / well. After incubation at room temperature for 1 hour, the plate was washed three times in the same manner. Next, AMDEX streptavidin-conjugated horseradish peroxidase (GE Healthcare) was diluted 7000-fold with T-TBS and added to each well at 50 μL / well. After incubation at room temperature for 30 minutes, the plate was washed four times in the same manner, and 3,3',5,5'-tetramethylbenzidine (TMB) solution (Sigma-Aldrich) was added to each well at 50 μL / well. After incubation at room temperature for approximately 20 minutes, the reaction was stopped with 1 mol / L sulfuric acid solution (Fujifilm Wako). The absorbance at 450 nm (sub wavelength 630 nm) was measured using a plate spectrophotometer (EL312e; manufactured by BIO-TEK INSTRUMENTS).

[0070] In Figure 2, the presepsin concentration obtained by presepsin ELISA in urine samples from two healthy subjects is plotted on the vertical axis, and each fraction number is plotted on the horizontal axis. The positions where the presepsin standard was detected are indicated by arrows. A fraction with a molecular weight equivalent to that of the presepsin standard was detected in the urine samples from healthy subjects, confirming that the ELISA prepared with anti-presepsin antibodies specifically detected presepsin in urine.

[0071] Example 3: Urine sample collection and renal biopsy Based on the results of serum creatinine, eGFR values, urinary protein, urinary trace albumin, urinary occult blood, etc., it was determined that a renal biopsy was necessary. Patients who underwent renal biopsy between March and December 2020 were given consent for the Nagoya University Kidney Disease Registry (N-KDR) study (Nagoya University Ethics Committee Approval Number: 2010-1135-7), and random urine samples were collected from the patients on the day before the renal biopsy. In the case of affiliated hospitals, urine was collected within 24 hours of refrigeration, aliquoted, and frozen at -80°C. Urine samples collected at Nagoya University Hospital were refrigerated and aliquoted within 6 hours, then frozen at -80°C.

[0072] Renal biopsies were performed as follows. The patient was placed in the prone position, and the puncture site was disinfected with an iodine-based disinfectant. Next, to avoid complications of renal biopsy (such as perinephric bleeding), the puncture site was primarily selected in the inferolateral region of the lower pole of the kidney, where sufficient renal cortical tissue could be obtained. The ultrasound probe (3.0-3.5 MHz) was placed on the left back, and the skin at the planned needle insertion site was marked with a marker. Local anesthetic was administered along the needle insertion path and the skin at the planned needle insertion site. Before inserting the biopsy needle, an incision was made in the skin, and the automated biopsy device was operated in conjunction with a 14-18G biopsy device to obtain renal parenchymal tissue. The collected renal tissue was transferred to saline-soaked gauze to prevent drying until each fixation. Tissue samples were collected two to three times in this manner. After the puncture, manual pressure was applied to the patient's back to stop bleeding.

[0073] The collected tissue was cross-cut using a feather double-edged razor. Next, for renal biopsy pathological diagnosis, four types of staining were performed, and one section of each was prepared. Specimens for optical microscopy were fixed in masked formaldehyde fixative for 3 hours to overnight, then dehydrated, delipidated, and infiltrated with paraffin to prepare paraffin-embedded blocks. Staining was performed using a combination of various staining methods, including HE staining, PAS staining, PAM staining, and Elastica-Masson staining, according to standard procedures. Optical microscopy allowed for overall observation of glomeruli, tubules, and blood vessels, providing basic tissue information. Specimens for electron microscopy were fixed in 2.5% glutaraldehyde for 2 hours to overnight (4°C) and washed three times for 10 minutes in 0.1 mol / L PB buffer. Next, specimens were fixed in 2% osmic acid fixative for 2 hours and dehydrated in ethanol concentrations ranging from 50% to 100%. The obtained sections were magnified approximately 20,000 times under an electron microscope to confirm the structure of glomeruli and tubules, the internal structure of the cells that make them up, and deposits that cause nephritis. Furthermore, frozen tissue sections were prepared. Immunofluorescence assays were used to observe the presence and location of immunoglobulins (IgG, IgA, IgM) and complements (C3, C1q). Specifically, fluorescently labeled antibodies that specifically react with target antigens were reacted. After the reaction, the molecules to which the antibodies bind were observed using a fluorescence microscope. The three tests (i.e., light microscopy, electron microscopy, and immunofluorescence assay) were combined to diagnose the cause of kidney disease, while also calculating the degree of inflammatory cell infiltration and the degree of tubular atrophy. The calculation method was to visually inspect the area of ​​each lesion using Elastica-Masson staining and classify it into four levels in 25% increments: Minimal (<5%), Mild (5-25%), Moderate (26-50%), and Severe (>50%). The severity of each lesion was evaluated by consensus of three or more nephrologists.

[0074] Example 4: Comparison of Urinary and Plasma Presepsin Levels Presepsin levels are elevated in patients with sepsis and are therefore used as a marker for diagnosing sepsis. It has been reported that the effects of renal failure are not observed up to G2 in the GFR classification of CKD patients, but gradually increase from G3 onward. Studies in dogs have revealed that presepsin is excreted in urine via renal metabolism (Patent Document 3), and it has been reported that decreased renal function (increased GFR stage classification) leads to increased blood presepsin levels (Non-Patent Document 2). Miyoshi et al. (Non-Patent Document 3) reported that correcting blood presepsin levels with blood creatinine levels can eliminate the effects of decreased renal function. Therefore, the relationship between blood and urinary presepsin levels in patients with renal failure without infection was analyzed after correcting for creatinine. We compared blood presepsin (per mg / dL of creatinine) and urinary presepsin (uP-SEP / gCr) in five CKD patients whose plasma and urine samples were preserved. Similar to blood presepsin, urinary presepsin also showed a negative correlation with GFR stage classification. Although blood presepsin tended to increase gradually with a decrease in eGFR, the increase was small and not sufficient to assess the pathological condition. Furthermore, this finding merely supports the findings of Non-Patent Document 3. On the other hand, urinary presepsin was found to respond sensitively to a decrease in eGFR, with a high rate of increase. These results suggest that urinary presepsin may more sensitively reflect renal status than blood presepsin, demonstrating that urinary presepsin is an excellent marker for assessing renal status (Figure 3).

[0075] Immunohistological examination of renal biopsies from patients with nephropathy reveals a significant increase in the number of macrophages in the glomeruli and interstitium compared with normal renal tissue. Macrophages have diverse functions, including antigen presentation, phagocytosis, and production of cytokines and growth factors, and play an important role in host defense, including immune responses. However, activated macrophages are also important factors in the progression of tissue injury by releasing inflammatory cytokines, proteases, and nitric oxide (NO) locally (Kenichi Shikata, Journal of the Japanese Society of Nephrology 2007; 49(5): 474-480). This indicates that an increase in inflammatory cells is closely related to renal injury. The more sensitive increase in presepsin levels in urine than in blood may reflect presepsin production at the site of injury and its subsequent transfer into the urine through damaged, leaky renal tubules. It was previously known that presepsin levels increase with the phagocytosis of microorganisms during infectious diseases, but it was not known that presepsin is also produced in tissue disorders such as kidney damage, and its production outside the site of infection was a surprising discovery.

[0076] Example 5: Urinary presepsin (urinary P-SEP) concentrations in various kidney diseases. We conducted extensive research to determine whether urinary presepsin better reflects the state of the kidneys and discovered that urinary presepsin levels are elevated in kidney diseases, particularly interstitial nephritis. To clarify the association between urinary presepsin and kidney diseases, we investigated the association between urinary presepsin levels and chronic kidney disease (CKD) in 463 patient urine samples collected in a registry study. All 463 patients underwent kidney biopsies to confirm the diagnosis of each kidney disease. The breakdown of the 463 cases was as follows: 38 cases of ANCA-associated vasculitis (AAV), 26 cases of diabetic nephropathy (DMN), 23 cases of glomerular sclerosis (FSGS), 106 cases of IgA nephropathy, 53 cases of nephrotic syndrome (MCNS), 41 cases of membranous nephropathy (MN), 12 cases of lupus nephritis, 28 cases of interstitial nephritis, and others. Patient background information is shown in Table 1. Urine samples were also collected from 10 healthy volunteers.

[0077]

[0078] Urinary presepsin concentrations were measured using an automated immunoluminescence analyzer (PATHFAST). Specifically, the presepsin PATHFAST kit of Example 1 was prepared, and 100 μL of urine sample was dispensed into the sample port of the reagent cartridge and placed in the instrument. The instrument was set up according to the manual, and urinary presepsin concentrations were measured. The measurement results (actual values) of urinary presepsin concentrations for each disease are shown in Figure 4. The mean ± standard deviation and median (25th and 75th quartiles) for each disease are also shown in Table 2.

[0079]

[0080] Urinary presepsin concentrations were confirmed to be low in healthy subjects (control group) and significantly elevated in patients with various renal disorders (Mann-Whitney U test, *; P value < 0.05). Furthermore, urinary presepsin concentrations were low in IgA nephropathy, FSGS, MN, and MCNS, which are glomerular diseases presumed to involve relatively low levels of interstitial and tubular damage, but were significantly elevated in patients with interstitial nephritis, AAV, DMN, and lupus nephritis, which involve high levels of interstitial and tubular damage, compared with healthy subjects or IgA nephropathy. These results demonstrate that urinary presepsin concentrations can be used as a marker for detecting the degree of interstitial and tubular damage (tubulointerstitial damage), in particular. Figure 5 shows the results of receiver-operating characteristic curve (ROC) analysis of these four diseases, using a population consisting of patients with AAV, DMN, lupus nephritis, and interstitial nephritis, which showed significant differences compared to healthy controls, plus healthy controls. The results showed that a cutoff value of 982 pg / mL yielded a sensitivity of 73.1%, a specificity of 90.0%, and an AUC of 0.869 (95% CI 0.785-0.954), demonstrating that this analysis can clearly differentiate diseases with a high likelihood of renal damage (particularly tubulointerstitial damage). It has been known that presepsin, which is induced and produced by macrophages following bacterial infection, can be used as an indicator of infectious diseases (particularly sepsis). As shown in this example, it was surprising that presepsin can reflect inflammation independently of infection, and furthermore, that it correlates with inflammation occurring in the renal interstitium, rather than glomerular inflammation.

[0081] Example 6: Correction of Urinary Presepsin Levels with Urinary Creatinine When measuring markers in random urine samples, correction with urinary creatinine (Cr) is used to eliminate the effect of variations in concentration in the sample due to urine volume. Therefore, in this study, evaluation was performed using the value (uP-SEP / Cr) obtained by dividing the urinary presepsin concentration (uP-SEP) by the urinary creatinine (Cr). Figure 6 shows the uP-SEP / Cr values ​​for various kidney diseases as Log10 uP-SEP / Cr (ng / gCr). Comparisons between disease groups were performed using Bonferroni's multiple comparisons method, and the results are shown using the Kruskal-Wallis test (*: P value < 0.05).

[0082] These results revealed that uP-SEP / Cr values ​​were significantly higher in interstitial nephritis, which is characterized by a high degree of interstitial and tubular damage, and that significant differences were observed among diseases. The increase in uP-SEP / Cr values ​​was low in focal glomerular sclerosis and IgA nephropathy, which are glomerular diseases and are presumed to involve relatively low degrees of interstitial and tubular damage, and consistent with interstitial damage regardless of disease. Furthermore, it was demonstrated that uP-SEP / Cr does not reflect the concentration of urinary presepsin due to a decrease in urine volume associated with the pathological condition, but is a specific marker for the degree of kidney damage (particularly tubulointerstitial damage).

[0083] Next, the sensitivity and specificity of the uP-SEP / Cr value in detecting interstitial nephritis were calculated. The results of receiver-operating characteristic curve (ROC) analysis are shown in Figure 7. The cutoff value was set as follows using ROC curve analysis. That is, because renal failure resulting from interstitial damage is severe, this cutoff value was used to distinguish whether damage has progressed to the interstitium. Compared to the uP-SEP / Cr values ​​of patients with renal disorders other than interstitial nephritis, the cutoff value for detecting interstitial nephritis was calculated to be 1525.689 (ng / gCr). The sensitivity of the urinary presepsin marker was 85.7%, and the specificity was 66.2%. The AUC of the ROC curve was 0.826, demonstrating unprecedented and extremely high usefulness as a single biomarker for detecting interstitial nephritis. In the examples described later, the results of uP-SEP / Cr values ​​will be shown, but similar results were also obtained for uP-SEP values.

[0084] Example 7 Renal Biopsy Findings and uP-SEP / Cr Values ​​For all patients, the results of renal biopsies were compared between patients with and without findings of renal impairment. Figure 8 shows the uP-SEP / Cr values ​​for each group as Log10 uP-SEP / Cr (ng / gCr). Comparisons between groups were performed using Bonferroni's multiple comparisons method and the results are shown using the Kruskal-Wallis test (*: P value < 0.05). As a result, patients with findings of renal impairment, i.e., patients with renal impairment such as interstitial damage or glomerular damage, had significantly higher uP-SEP / Cr values ​​(Figure 8, P = 0.0332). Furthermore, ROC analysis revealed a cutoff value of 641.461 (ng / gCr), with a sensitivity of 66.7%, a specificity of 63.7%, and an AUC of 0.707 (95% CI: 0.563-0.852). These results indicated a correlation between patients with renal biopsy findings and urinary presepsin levels. Furthermore, patients who underwent renal biopsy were analyzed by dividing them into those with interstitial damage and those with glomerular damage. Patients were classified into three groups: those with interstitial damage (Mild or higher), those without interstitial damage and those without glomerular damage, and those without interstitial damage and those with glomerular damage, and the uP-SEP / Cr values ​​were compared. As shown in Figure 9, a Bonferroni multiple comparison was performed to compare groups, followed by a Kruskal-Wallis test (P value < 0.05), and it was found that the group with interstitial damage and the group without interstitial damage (including those with glomerular damage) were significantly distinguishable. The uP-SEP / Cr values ​​for each group were expressed as Log10 uP-SEP / Cr (ng / gCr). Furthermore, within the group without interstitial damage, there was no significant difference between the group with glomerular damage and the group without glomerular damage. These results demonstrate that urinary presepsin can specifically diagnose interstitial damage and can also be used to differentiate it from glomerular damage.

[0085] Example 8: Differentiation of the degree of renal damage based on renal biopsy findings and uP-SEP / Cr values. The tissue surrounding the glomeruli and tubules in the kidney is called the interstitium, and interstitial nephritis is a disease in which inflammation in this area leads to impaired renal function. Furthermore, inflammation often occurs not only in the interstitium but also in the tubules, and the infiltration of inflammatory cells into the interstitium and atrophy of tubules observed in renal findings are important findings indicative of the degree of renal damage in interstitial nephritis. Based on the imaging diagnosis of renal biopsies, the degree of damage was classified into minimal (<5%), mild (<5-25%), moderate (26-50%), and severe (>50%). A significance test for minimal between-group differences was performed using Bonferroni's multiple comparison test, and a significant difference was calculated for Kruskal-Wallis samples (P value <0.001).

[0086] The results showed that increases in uP-SEP / Cr levels correlated with increases in the degree of damage for both inflammatory cell infiltration and tubular atrophy, with significant differences between the groups (Figure 10). Regarding the degree of inflammatory cell infiltration and tubular atrophy shown in Figure 10, there was a significant difference between Minimal and Mild for inflammatory cell infiltration, but no significant difference for tubular atrophy. Inflammatory cell infiltration occurs before tubular atrophy. Thus, the degree of increase in uP-SEP / Cr levels between the groups indicated a more rapid increase with inflammatory cell infiltration, making this useful for understanding the progression of tubulointerstitial damage in the acute phase. Monitoring uP-SEP / Cr levels can be used to determine the initiation of treatment and assess therapeutic efficacy. To more clearly confirm the relationship between the degree of inflammatory cell infiltration and tubular atrophy, which indicate interstitial damage, and the uP-SEP / Cr value, a significance test was performed between a patient group in which either inflammatory cell infiltration or tubular atrophy, or both, were Mild or greater and a group in which both inflammatory cell infiltration and tubular atrophy were Minimal (Figure 11). The results showed that when either inflammatory cell infiltration or tubular atrophy was Mild or greater, the uP-SEP / Cr value was elevated (p<0.001), demonstrating that urinary presepsin can be used to determine the presence or absence of interstitial renal damage. Next, Table 3 shows the results of a Bonferroni multiple comparison test for significance between groups in the Minimal group, and the significant difference was calculated for Kruskal-Wallis samples (P<0.001). The results show that the degree of inflammatory cell infiltration and tubular atrophy (from mild to severe) increases with every 100 ng / gCr increase in the uP-SEP / Cr value, and these differences are significantly distinguishable.

[0087]

[0088] The cutoff values, sensitivity, specificity, and AUC obtained from the ROC analysis are shown in Table 4. These results indicate that there was no difference in the cutoff values ​​between the degree of inflammatory cell infiltration and the degree of tubular atrophy, and that although the degree of inflammatory cell infiltration and the degree of tubular atrophy cannot be distinguished, the progression of tubulointerstitial damage can be determined by the degree of increase in the acute phase. Furthermore, it was found that the cutoff value reflected the sum of tubular atrophy (chronic symptoms) and inflammatory cell infiltration (acute symptoms).

[0089]

[0090] As described above, the uP-SEP / Cr value is a sensitive biomarker for the progression of tubulointerstitial damage, and is also useful as a marker reflecting the overall degree of tubulointerstitial damage, demonstrating its usefulness as a complement to renal biopsy.

[0091] The images of the kidney biopsy used for the assessment are shown on the left in Figure 12 (normal case: minimal inflammatory cell infiltration, minimal tubular atrophy) and on the right in Figure 12 (interstitial nephritis case: severe inflammatory cell infiltration, severe tubular atrophy). In normal cases, the spaces between the tubules are very small, and the tubules are adjacent to each other. Furthermore, even if fibrosis is observed, it is minimal. However, in cases of interstitial nephritis, inflammatory cell infiltration is observed in the interstitium, and the spaces are larger due to fibrosis, tubular atrophy, etc.

[0092] Example 9: Relationship between tubular atrophy and interstitial fibrosis (T-score) in the Oxford classification of IgA nephropathy and uP-SEP / Cr values. Renal biopsy is required for a definitive diagnosis of IgA nephropathy. Renal biopsy is useful not only for definitive diagnosis but also for predicting prognosis and selecting treatment based on tissue activity and severity. Renal biopsy reveals cellular proliferation in the mesangial region of the glomerulus and deposition of IgA and complement C3, a type of immune component, in the mesangial region of the glomerulus. Therefore, to suppress abnormal IgA production, tonsillectomy is performed, followed by steroid pulse therapy to suppress glomerular inflammation (tonsillectomy pulse therapy). However, it has been reported that approximately 40% of IgA nephropathy cases progress to renal failure within 20 years, which is thought to be due to chronic glomerular inflammation and gradual glomerular sclerosis, leading to a decline in renal function. The Oxford classification, T-score, is based on the percentage of tubular atrophy or interstitial fibrosis in the cortex. It is expected to accurately predict the progression of IgA nephropathy and contribute to improving prognosis. Therefore, we investigated whether the uP-SEP / Cr value correlates with T-score. 106 patients with IgA nephropathy whose T-scores were calculated were classified using a T-score > 0 cutoff value. A Bonferroni multiple comparison test was performed to determine significance. Significance was calculated for Kruskal-Wallis samples (P < 0.001), and an increase in uP-SEP / Cr was confirmed in patients with high T-scores (Figure 13, P = 0.00908). Furthermore, ROC analysis revealed that the cutoff value was 543.082 (ng / gCr), with a sensitivity of 72.2%, a specificity of 60.5%, and an AUC of 0.699 (95% CI: 0.575-0.823). These results demonstrate that the uP-SEP / Cr value can be used to detect tubulointerstitial damage in patients with IgA nephropathy, and that the uP-SEP / Cr value is useful for diagnosing the severity of IgA nephropathy.

[0093] Example 10: Evaluation of uP-SEP / Cr values ​​in patients with chronic kidney disease (CKD) The correlation between the uP-SEP / Cr values ​​of five patients with CKD, such as diabetic nephropathy and nephrosclerosis, and the subsequent changes in renal function was analyzed. Table 5 shows the patient background, uP-SEP / Cr values, and the subsequent rate of decline in eGFR values ​​for the five patients.

[0094]

[0095] Looking at five CKD patients, it was confirmed that patients with a high rate of decline in eGFR values ​​had high uP-SEP / Cr values, while patients with a low rate of decline tended to have low uP-SEP / Cr values. These results indicate that the uP-SEP / Cr value reflects the degree of renal dysfunction in CKD patients, and that the uP-SEP / Cr value reflects the degree of renal dysfunction even in stages where renal biopsy cannot or is not performed. Furthermore, the rate of decline in eGFR indicated that patients with high uP-SEP / Cr values ​​had a high degree of renal dysfunction. These results indicated that the uP-SEP / Cr value is related to the degree of renal dysfunction in patients and can reflect prognosis. Therefore, it was demonstrated that the uP-SEP / Cr value is useful for diagnosing the worsening of CKD patients.

[0096] Example 11: Treatment Progress of Chronic Kidney Disease (CKD) Patients and Patient Monitoring Using uP-SEP / Cr Values ​​Renal biopsy is a crucial test for identifying the cause of CKD and formulating a treatment plan. However, many patients cannot undergo renal biopsy, such as those with a strong tendency to bleed, those with one kidney, those with renal atrophy, or those who do not wish to undergo renal biopsy. In such cases, the cause must be estimated and treatment initiated, but there are disadvantages, such as the inability to select potent steroid therapy if a definitive diagnosis cannot be made. The results described above demonstrate that the uP-SEP / Cr value is a marker of tubulointerstitial damage, and it is expected that the uP-SEP / Cr value will reflect changes in the patient's condition when monitoring CKD patients. Therefore, we observed changes in eGFR and uP-SEP / Cr values ​​over a period of approximately nine years, from 2010 to 2019, in four CKD patients who did not undergo renal biopsy.

[0097] The results are shown in Figure 14. Case 1 (Pt1) and Case 2 (Pt2) showed a correlation between a decrease in eGFR and an increase in uP-SEP / Cr values. Case 4 (Pt4) showed a high uP-SEP / Cr value of 1489.3 in 2011, but in 2013 and 2016, the uP-SEP / Cr values ​​decreased to 238.0 and 101.3, respectively. The eGFR also increased above 30, the index for GFR stage 5, improving to stage 4. This case was subsequently diagnosed with drug-induced kidney injury (DKI), and a change in medication was confirmed to have improved the renal damage. These results indicate that the uP-SEP / Cr value is a useful marker of tubulointerstitial damage in CKD patients and can be used to monitor patients. These results suggest that elevated urinary presepsin concentrations are highly likely to indicate tubulointerstitial damage. In other words, the use of urinary presepsin as a marker makes it possible to easily and early detect tubulointerstitial damage in CKD patients.

[0098] According to the present invention, it is possible to detect and monitor tubulointerstitial damage using urinary presepsin concentration as an indicator. The present invention allows for a simple and early diagnosis, replacing renal biopsy, to determine the degree of tubulointerstitial damage in patients who cannot undergo renal biopsy, and to determine the degree of tubulointerstitial damage after treatment in patients whose treatment plan has been determined by renal biopsy. Early determination of the degree of tubulointerstitial damage allows for proactive therapeutic intervention at an early stage, leading to improved prognosis. Urinary presepsin concentration is also useful for estimating the prognosis of renal function.

[0099] The present invention enables the detection of the degree of renal damage in patients suspected of or already suffering from tubulointerstitial damage using urinary presepsin levels as an indicator. Furthermore, the present invention allows for a simple and early diagnosis, replacing renal biopsy, for determining the degree of renal damage in patients for whom renal biopsy cannot be performed, or for determining the degree of renal damage after treatment in patients for whom a treatment plan has been determined by renal biopsy. Early determination of the degree of renal damage allows for proactive therapeutic intervention at an early stage, leading to improved prognosis. Urinary presepsin levels are also useful for determining the severity of kidney disease. While the present invention has been described above in relation to specific embodiments, modifications and improvements obvious to those skilled in the art are within the scope of the present invention.

Claims

1. A method for detecting tubulointerstitial damage, comprising measuring the urinary sCD14-ST concentration of a test subject, or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level.

2. The method of claim 1, wherein the tubulointerstitial disorder is a chronic tubulointerstitial disorder.

3. The method according to claim 1 or 2, wherein the subject is a patient suspected of having kidney disease or a patient with kidney disease.

4. The method according to claim 1 or 2, wherein the subject is a patient suspected of having one or more kidney diseases selected from the group consisting of nephrosclerosis, IgA nephropathy, diabetic nephropathy, membranous nephropathy, ANCA-associated vasculitis, focal segmental glomerulosclerosis, minimal change nephrotic syndrome, lupus nephritis, tubulointerstitial nephritis, Sjogren's syndrome, vascular renal disease, idiopathic TINU syndrome, and IgG4-associated nephritis, as well as a patient with such kidney disease.

5. The method according to claim 1, wherein the subject is a patient suspected of having a kidney disease, which is acute tubulointerstitial nephritis or chronic tubulointerstitial nephritis, or a patient with said kidney disease.

6. The method according to claim 1 or 5, wherein the subject is a patient suspected of having a kidney disease, which is acute drug-induced tubulointerstitial nephritis or chronic drug-induced tubulointerstitial nephritis, or a patient with said kidney disease.

7. A method for monitoring tubulointerstitial damage by measuring the urinary sCD14-ST concentration of a test subject, or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value.

8. A method for selecting a treatment for tubulointerstitial damage by measuring the urinary sCD14-ST concentration of a subject or a value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value.

9. A method for determining the severity of kidney disease by measuring the sCD14-ST concentration in the urine of a subject, or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value.

10. A method for detecting kidney disease, comprising measuring the urinary sCD14-ST concentration of a test subject, or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value.

11. A method for detecting kidney disease by measuring the urinary sCD14-ST concentration of a test subject, or a value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value, wherein the kidney disease is one or more kidney diseases selected from the group consisting of nephrosclerosis, IgA nephropathy, diabetic nephropathy, membranous nephropathy, ANCA-associated vasculitis, focal segmental glomerulosclerosis, minimal change nephrotic syndrome, lupus nephritis, tubulointerstitial nephritis, Sjogren's syndrome, vascular kidney disease, idiopathic TINU syndrome, and IgG4-associated nephritis.

12. A method for detecting tubulointerstitial nephritis, which comprises measuring the sCD14-ST concentration in the urine of a subject, or a value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value.

13. Measuring the sCD14-ST concentration in urine collected from a patient suspected of having tubulointerstitial damage or a patient having tubulointerstitial damage, or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value; and A step of determining that there is tubulointerstitial damage when the sCD14-ST concentration or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value is higher than that of a healthy subject. A method for detecting tubulointerstitial damage, comprising:

14. measuring the sCD14-ST concentration in urine collected from a patient suspected of having tubulointerstitial damage or a patient having tubulointerstitial damage, or a value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value; determining that there is tubulointerstitial damage when the sCD14-ST concentration or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value is higher than that of a healthy subject; and A process of administering immunosuppressive therapy when it is determined that there is tubulointerstitial damage. A method for detecting and treating tubulointerstitial damage comprising:

15. Measuring the sCD14-ST concentration in urine collected from a patient suspected of having kidney disease or a patient with kidney disease, or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value; and A step of determining that the subject has kidney disease when the sCD14-ST concentration or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value is higher than that of a healthy subject.

23. A method for detecting kidney disease comprising:

16. measuring the sCD14-ST concentration in urine collected from a patient suspected of having kidney disease or a patient with kidney disease, or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine level; determining that the subject has kidney disease when the sCD14-ST concentration or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value is higher than that of a healthy subject; and A process of administering immunosuppressant therapy if kidney disease is determined.

23. A method for detecting and treating kidney disease comprising:

17. A method for determining whether or not to perform a renal biopsy by measuring the sCD14-ST concentration in the urine of a subject prior to the renal biopsy, or the value obtained by dividing the sCD14-ST concentration in the urine by the urinary creatinine value.

18. The method according to any one of claims 1, 2, 5, and 7 to 17, wherein the sCD14-ST concentration or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value is measured by an immunological measurement method.

19. Use of the urinary sCD14-ST concentration, or a value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value, as a marker for detecting tubulointerstitial damage.

20. A kit for detecting tubulointerstitial damage, comprising: (a) an antibody specific for sCD14-ST; (b) standard data showing the correlation between the urinary sCD14-ST concentration or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value and the degree of renal damage; (c) Instructions for use The kit comprises:

21. A kit for detecting tubulointerstitial damage, comprising: (a) an antibody specific for sCD14-ST; (b) standard data showing the correlation between the urinary sCD14-ST concentration or the value obtained by dividing the urinary sCD14-ST concentration by the urinary creatinine value and the severity of kidney disease; (c) Instructions for use The kit comprises: