Alkaline phosphatase for use in treating acute exacerbations of chronic kidney disease

Abstract

This disclosure relates to the use of alkaline phosphatases, particularly improved alkaline phosphatases such as RecAP, for the prevention, treatment, cure, or improvement of symptoms of acute kidney injury in addition to chronic kidney injury. This application relates to alkaline phosphatases for use in a method to preserve renal function, shorten the duration of renal replacement therapy, preserve creatinine clearance, and reduce the risk of death in subjects having acute kidney injury in addition to chronic kidney injury.

Description

[Background technology]

[0001] The kidneys perform several functions in the animal body, including waste elimination, acid-base homeostasis, osmolality regulation, blood pressure regulation, and hormone secretion. To enable the kidneys to perform these tasks, they receive approximately 20% of cardiac output, despite their relatively small size. Consequently, interruptions in blood flow to the kidneys (renal blood flow, RBF) have a direct impact on many aspects of kidney function. For example, this can lead to reduced elimination of nitrogenous waste products and disruptions in fluid and electrolyte balance. In the long term, not only reduced RBF but also other toxic events, such as ischemia (reperfusion injury), contrast agents, or (other) nephrotoxic drugs, such as antibiotics, can also cause acute kidney injury (AKI) because they are very stressful on the kidneys.

[0002] Acute kidney injury (AKI) is observed in up to 60% of patients in the intensive care unit (ICU), and its incidence is increasing (Hoste et al. Intensive Care Med 2015;41:1411-23, Hoste et al. Critical Care 2006;10:R73, Nisula et al. Intensive Care Med 2013;39:420-8, Vincent et al. Critical Care Medicine 2006;34:344-53). The development of acute kidney injury (AKI) in patients with sepsis is associated with increased mortality (Kellum et al. Critical Care Medicine 2016;193:281-7), while survivors are at risk of developing chronic kidney disease (CKI, also known as chronic kidney disease (CKD)) (Chawla et al. Kidney Int 2011;79:1361-9, Oppert et al. Nephrol Dial Transplant 2008;23:904-9, Vaara et al. Crit Care 2012;16:R197), which places a significant burden on both patients and society.

[0003] AKI itself is a common disorder with an incidence of approximately 2,000 per million people (pmp). Patients with chronic kidney disease (CKI) are at higher risk of developing AKI, as evidenced by a low estimated glomerular filtration rate (eGFR) or the presence of proteinuria for more than three months. CKI is a strong risk factor for cardiovascular events, and patients with CKI have a specific mortality risk if they develop AKI in addition to CKI, known as acute exacerbation of chronic kidney disease (AoCKI).

[0004] Firstly, AKI is associated with high costs, morbidity, and mortality in CKI patients; and secondly, AoCKI can cause further decline in renal function in these already impaired patients, potentially leading to irreversible loss of renal function below a critical threshold; therefore, preventing and / or appropriately treating AoCKI is extremely important. [Overview of the project]

[0005] The present invention provides alkaline phosphatase (AP) for use in a method of treating acute kidney injury (AKI) in a subject requiring treatment, wherein the method comprises administering an effective amount of AP to the subject, the subject having an acute exacerbation of chronic kidney injury (AoCKI), the chronic kidney injury (CKI) being mild, moderate, or severe, preferably moderate to severe. Preferably, the estimated glomerular filtration rate (eGFR) before the onset of AKI is ≤75 ml / min / 1.73 m². 2 For example, ≤60 ml / min / 1.73 m 2 More preferably ≤ 45 ml / min / 1.73 m 2 The pre-AKI eGFR is preferably ≥ 15 ml / min / 1.73 m². 2 For example, ≥25 ml / min / 1.73 m 2 That is the case.

[0006] In a preferred embodiment, AP for use according to the present invention is provided, and administration of AP results in a reduced risk of developing a major adverse renal event (MAKE90) by day 90. Preferably, the reduction in the risk of MAKE90 includes a reduction in the risk of MAKE90 compared to the risk of MAKE90 in the absence of treatment. Preferably, MAKE90 includes one or more of the following events: (i) death before or at day 90, (ii) renal replacement therapy before or at day 90, (iii) a ≥25% decrease in eGFR at day 90 compared to eGFR before AKI, and (iv) readmission before or at day 90.

[0007] In a preferred embodiment, the present invention provides AP for use according to the present invention, wherein AP is administered in at least one dose of 500 U / kg to 2,000 U / kg.

[0008] In a preferred embodiment, the present invention provides an AP for use according to the present invention, wherein the AP is a human AP.

[0009] In a preferred embodiment, the present invention provides an AP for use according to the present invention, wherein the AP is a recombinant AP, preferably a chimeric AP, and more preferably a chimeric AP having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with respect to the amino acid sequence of RecAP (SEQ ID NO: 1).

[0010] In a preferred embodiment, the present invention provides AP for use according to the present invention, the AP being administered in once-daily or three-times-daily doses.

[0011] In a preferred embodiment, the present invention provides AP for use according to the present invention, wherein AP is administered intravenously.

[0012] In a preferred embodiment, the present invention provides an AP for use according to the present invention, and the AP dosage is about 1.6 mg / kg of RecAP and / or about 1000 U / kg of RecAP.

[0013] In a preferred embodiment, the present invention provides an AP for use according to the present invention, and administration of at least one dose of AP results in preservation of glomerular filtration rate (GFR) or estimated GFR (eGFR) on day 90 in the subject relative to the pre-AKI onset eGFR.

[0014] In a preferred embodiment, the present invention provides an AP for use according to the present invention, and the AKI is caused by or associated with sepsis. This is referred to as sepsis-associated AKI (SA-AKI).

[0015] In a preferred embodiment, the present invention provides an AP for use according to the present invention, and the treatment is initiated within 24 hours after sepsis is detected and / or within 48 hours after acute kidney injury (AKI) is detected in the subject.

[0016] In a preferred embodiment, the present invention provides an AP for use according to the present invention, and the decreased eGFR due to chronic kidney disease is determined at least 15 days before, preferably at least 30 days before, more preferably at least 60 days before, and most preferably at least 90 days before administration of the AP.

Brief Description of the Drawings

[0017] [Figure 1] Provide monitoring of the individual test visit flow of the subject, including a screening and baseline period (before day 1), a treatment period (day 1 to day 3), and a follow-up period (day 4 to day 180). [Figure 2] Shows the RecAP amino acid sequence (SEQ ID NO: 1). [Figure 3]This flowchart shows registration, randomization, untreated patients, efficacy analysis according to the protocol, safety analysis population, and combined population. Abbreviations: ICF (Informed Consent Form), ITT (Intention to Treat), mCKD (Moderate to Severe Chronic Kidney Disease). [Figure 4] This is a Kaplan-Meier plot showing survival rates for patients treated with ilophotase alfa versus those treated with placebo. The 90-day survival rates are compared to a Wald test for the difference in proportions, and the respective p-values ​​are given. Mortality can be calculated as 1 - survival rate. Proportion of patients surviving at 90 days: ilophotase alfa: 0.6554; placebo: 0.6397. P-value based on Wald test for the difference in the proportion of patients surviving at 90 days (ilophotase alfa - placebo): 0.3404. Patients who died after 90 days were censored at 90 days. [Figure 5] This is a Kaplan-Meier plot showing the proportion of subjects treated with ilofotase alfa versus patients treated with placebo who did not experience a MAKE90 event. The proportion of subjects without a MAKE90 event at day 90 is compared to a Wald test for the difference in proportions, and the respective p-values ​​are given. The proportion of subjects with a MAKE90 event up to day 90 can be calculated as 1 minus the proportion of subjects without a MAKE90 event. Proportion of patients without a MAKE90 event at day 90: ilofotase alfa: 0.4277; placebo: 0.3533. P-value for the difference (ilofotase alfa-placebo: 0.030705). Patients who discontinued before day 90 without a MAKE90 event were censored on the day of discontinuation. Patients who discontinued after day 90, completed the trial, died, or were hospitalized were censored on day 90. MAKE90 events (receiving RRT at day 90 / <25% decrease in eGFR) were recorded at visits before or 12 days after day 90. Abbreviations: MAKE90 (major adverse renal event up to day 90), RRT (renal replacement therapy). [Figure 6]This plot shows the predicted probability of experiencing a Make90 event, including a 95% confidence interval. The predictions are calculated based on the parameter estimations of the logistic regression model described in Table 7. Predictability is taken from the results of the logistic regression model including MAKE90 as the outcome of interest, as well as pre-AKI reference eGFR with respect to treatment, pre-AKI reference eGFR with respect to treatment interactions (p=0.0235). Patients who did not meet the criteria for a confirmed MAKE90 event are assumed not to have a MAKE90 event in the logistic regression model. [Figure 7] This is a Kaplan-Meier plot showing the proportion of patients with pre-AKI reference eGFR < 60 who did not experience a MAKE90 event, compared to patients treated with ilofotase alfa and those treated with placebo. The proportion of patients without a MAKE90 event at day 90 is compared to a Wald test for the difference in proportions, and the respective p-values ​​are given. The proportion of patients with a MAKE90 event up to day 90 can be calculated as 1 minus the proportion of patients without a MAKE90 event. Proportion of patients without a MAKE90 at day 100: ilofotase alfa: 0.4032; placebo: 0.2092. P-value for difference (ilofotase alfa - placebo): 0.004357. Abbreviation: MAKE90 (Major adverse renal event up to day 90). [Figure 8]This is a Kaplan-Meier plot showing the proportion of subjects with pre-AKI reference eGFR < 75 who did not experience a MAKE90 event, compared to patients treated with ilofotase alfa and those treated with placebo. The proportion of subjects without a MAKE90 event at day 90 is compared to a Wald test for the difference in proportions, and the respective p-values ​​are given. The proportion of subjects with a MAKE90 event up to day 90 can be calculated as 1 minus the proportion of subjects without a MAKE90 event. Proportion of patients without a MAKE90 event at day 90: ilofotase alfa: 0.4361; placebo: 0.2971. P-value for the difference (ilofotase alfa-placebo: 0.0047). Patients who discontinued before day 90 without a MAKE90 event were censored on the day of discontinuation. Patients who discontinued after day 90, completed the trial, died, or were hospitalized were censored on day 90. MAKE90 events (receiving RRT at day 90 / <25% decrease in eGFR) were recorded at visits within 12 days before or after day 90. Abbreviations: MAKE90 (major adverse renal event up to day 90), RRT (renal replacement therapy). [Modes for carrying out the invention]

[0018] I General Sepsis is the leading cause of acute kidney injury (AKI) and a major cause of death. Patients with sepsis-associated AKI (SA-AKI) have high mortality and morbidity rates and are at risk of developing or worsening pre-existing chronic kidney disease (CKI), also known as chronic kidney disease (CKD). Alkaline phosphatase (AP) is a homodimeric endogenous enzyme present in many cells and organs, such as the intestines, placenta, liver, bones, kidneys, and granulocytes. It exerts detoxification through the dephosphorylation of endotoxins, pathogen-associated molecular pattern molecules (PAMPS, e.g., lipopolysaccharide [LPS]), and injury-associated molecular pattern molecules (DAMPS, e.g., adenosine triphosphate and diphosphate [ATP and ADP]). In animal models of sepsis and AKI, AP administration reduces the inflammatory response, improves renal function, and / or reduces mortality.

[0019] Sepsis-associated acute kidney injury (SA-AKI) is a multifactorial syndrome involving inflammatory, nephrotoxic, and ischemic injury occurring simultaneously with other pathophysiological responses that rapidly impair renal function (Bonventre et al. J Clin Invest 2011;121:4210-21, Gomez et al. Shock 2014;41:3-11). Currently, there are no approved pharmacological interventions available to prevent or treat AKI (Joannidis et al. Intensive Care Medicine 2017;43:730-49).

[0020] Alkaline phosphatase (AP) was originally considered a novel treatment for sepsis in general (Poelstra et al. American Journal of Pathology 1997;151:1163-69., Verweij et al. Shock 2004;22:174-9). In two small clinical trials, bovine AP administration improved renal function in patients with sepsis (Heemskerk et al. Crit Care Med 2009;37:417-23,e1, Pickkers et al. Crit Care 2012;16:R14).

[0021] A novel, recombinant chimeric human AP medical product called recAP was used as an intravenous (IV) infusion for the treatment of SA-AKI in the Phase 2 trial STOP-AKI. Herein, survival benefits were observed in the two highest dose groups, 0.8 mg / kg and 1.6 mg / kg, compared to the placebo group. No safety or tolerability concerns were observed for any of the tested doses (0.4, 0.8, and 1.6 mg / kg). The 1.6 mg / kg recAP dose was selected for the REVIVAL Phase 3 trial based on the observed significant survival benefit.

[0022] In the STOP-AKI trial, the subject of patent publication WO2019 / 172766, a history of CKI was one of the exclusion criteria. However, in the REVIVAL trial, subjects with mild to moderate CKI prior to the onset of AKI were not excluded. Therefore, the REVIVAL trial included subjects with normal renal function or mild renal impairment (i.e., eGFR > 60 ml / min / 1.73 m² before admission). 2 This includes not only those with the condition described above, but also those with reduced renal function due to moderate to severe chronic kidney disease prior to hospitalization (i.e., eGFR ≤ 60 ml / min / 1.73 m²). 2 Subjects who already had CKI were also treated. The latter are more susceptible to further acute renal decline, known as acute exacerbation of chronic kidney disease (AoCKI), and leave subjects with even lower residual renal function after the acute portion of the renal impairment is treated. Surprisingly, this invention shows that treatment of this selected patient population, which was not included in the STOP-AKI trial, yielded greater benefits from AP treatment than patients who had not had CKI before developing AKI.

[0023] Acute kidney injury (AKI) itself is a common disorder with an incidence of approximately 2,000 cases per million people (pmp). Patients with chronic kidney disease (CKI) are at higher risk of developing AKI, as evidenced by a low eGFR or the presence of proteinuria for more than three months. CKI is a strong risk factor for cardiovascular events, and patients with CKI are at particularly high risk of death if they develop advanced acute kidney injury (AoCKI).

[0024] RecAP is a chimeric AP that combines the characteristics of two human isoenzymes, intestinal and placental APs (Kiffer-Moreira et al. PLoS One 2014;9:e89374). Replacing the crown domain of the intestinal AP (the most biologically active isoenzyme) with the crown domain of the placental AP (which has the longest half-life) creates a highly stable, biologically active enzyme (Kiffer-Moreira et al. PLoS One 2014;9:e89374). See, for example, U.S. Patent Nos. 8586032 and 8557545, and U.S. Patent Application Publications 2017 / 0009216 and 2016 / 0250299.

[0025] Accordingly, the present invention relates to an AP for use in a method of treating acute kidney injury (AKI) in a subject requiring treatment, wherein the method comprises administering an effective amount of AP to the subject, the subject having an acute exacerbation of chronic kidney injury (AoCKI), the chronic kidney injury (CKI) being mild, moderate, or severe, preferably moderate to severe.

[0026] definition In this specification and the appended claims, the singular forms "a," "an," and "the" refer to multiple subjects unless the context clearly indicates otherwise. The terms "a" (or "an"), as well as "one or more" and "at least one," may be used interchangeably in this specification.

[0027] Furthermore, when used herein, “and / or” should be interpreted as a specific disclosure of each of the two designated features or components, with or without the other. Thus, in this specification, the term “and / or” as used in phrases such as “A and / or B” is intended to include “A and B,” “A or B,” “A” (alone), and “B” (alone). Similarly, the term “and / or” as used in phrases such as “A, B, and / or C” is intended to include each of the following embodiments: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0028] Wherever an aspect is described in this specification in the language “including,” other similar aspects are also provided, which would otherwise be described in terms of “consisting of” and / or “essentially consisting of.”

[0029] The term “about” as used in relation to numbers throughout this specification and the claims indicates an acceptable interval of precision, which is well known to those skilled in the art. Generally, such an interval of precision is ±15%.

[0030] Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art to which this disclosure relates.

[0031] Units, prefixes, and symbols are shown in their Systeme International de Unites (SI) approved forms.

[0032] Numerical ranges include the numbers that define the range. When a range of values ​​is enumerated, it should be understood that each intervening integer value between the enumerated upper and lower bounds of that range, and each fraction thereof, along with each subrange between such values, are also specifically disclosed. The upper and lower bounds of any range may, independently, be included in or excluded from the range, and each range that includes either limit, neither limit, or both limits is also encompassed within the Invention. When values ​​are explicitly enumerated, it should be understood that values ​​that are substantially the same quantity or amount as the enumerated values ​​are also within the scope of the Invention. When a combination is disclosed, each subcombination of the elements of that combination is also specifically disclosed and within the scope of the Invention. Conversely, when different elements or groups of elements are disclosed individually, their combinations are also disclosed. When any element of the Invention is disclosed as having multiple substitutes, examples of the Invention in which each substitute is excluded, either alone or in any combination with other substitutes, are also disclosed herein, and two or more elements of the Invention may have such exclusions, and all combinations of elements having such exclusions are disclosed herein.

[0033] As used herein, the terms “to treat,” “to treat,” or “to treat ~” mean (i) reducing the potential or risk of a disease or disorder, e.g., AKI (e.g., SA-AKI and / or AoCKI); (ii) reducing the incidence of a disease or disorder, e.g., AKI (e.g., SA-AKI and / or AoCKI); (iii) reducing the severity of a disease or disorder, e.g., AKI (e.g., SA-AKI and / or AoCKI) (e.g., improving symptoms); (iv) reducing the risk of a major adverse renal event (MAKE90) up to day 90; or (v) a combination thereof. For example, “to treat” may mean the ability of a therapy, when administered to a subject, to prevent or reduce the risk of developing renal impairment, e.g., SA-AKI and / or AoCKI (e.g., in a subject diagnosed with sepsis or at risk of sepsis), and / or to cure or alleviate the symptoms, signs, or causes of renal impairment, e.g., SA-AKI and / or AoCKI. The term "treating" also means, when compared to a non-treated (or placebo-treated) group, reducing or decreasing at least one clinical symptom, and / or inhibiting or delaying the progression of a condition, and / or preventing or delaying the onset of a disease or illness. Thus, the terms "treat," "treating," or "treatment of ~" (or grammatically equivalent terms) refer to both prophylactic and therapeutic treatment regimes. In the case of kidney disease, it is preferable that subjects, after being treated with AP in the manner disclosed herein, show improvement in renal function or a lesser decline in renal function compared to subjects not treated with AP, particularly at 90 days after the start of treatment.

[0034] As used herein, the term “day” means, in a numerical context and in relation to treatment, the number of days relative to the day (which is day 1) of administration of the first dose of either AP or placebo, such as “day 90” or “day 28” (see also Figure 1).

[0035] As used herein, the term “preserve” includes preventing a decline, slowing a decline, stopping a decline, and / or reversing a decline in renal function at least partially.

[0036] The term "increase" is not necessarily limited to increasing kidney function to a level equal to or greater than that before the treatment was administered. It includes partially restoring kidney function.

[0037] The term "treatment with a risk of reducing kidney function" is typically used to refer to a treatment that carries the risk of reducing kidney function when comparing the value of at least one kidney-related parameter to the recognized or mean (laboratory) value of that parameter, or when comparing that parameter to the value before the treatment was performed. For example, if the amount of protein in the urine of a subject, preferably a human, is significantly higher than the recognized or mean (laboratory) value, then the kidney function is said to be "reduced." The corresponding analysis can be performed in a laboratory, but can also be performed in a home setting. For example, since September 2006, the Dutch "Nierstichting" has introduced a simple test that can be performed at home to test whether the kidneys are functioning properly (called a kidney check ("Niercheck")). This test, for example, measures the amount of protein in the urine.

[0038] As used herein, the terms “subject” or “patient” refer to any subject, particularly mammalian subjects, for whom therapy or prognosis is desired for renal impairment, e.g., SA-AKI and / or AoCKI. As used herein, the terms “subject” or “patient” include any human or non-human animal. As used herein, phrases such as “patient with AKI, preferably SA-AKI, and / or AoCKI” or “patient with sepsis” include subjects, e.g., mammalian subjects, who would benefit from the administration of therapy in AP as disclosed herein.

[0039] In some aspects of this disclosure, the subject is a naive subject. A naive subject is a subject that has not received any therapy, such as a renal function-related therapeutic agent. In some aspects, the naive subject has not been treated with a therapeutic agent before being diagnosed with renal impairment, such as SA-AKI and / or AoCKI, or a disease or condition that may cause renal impairment (e.g., sepsis).

[0040] In another embodiment, the subject is receiving therapy and / or one or more doses of therapeutic agents prior to being diagnosed with renal impairment, e.g., SA-AKI and / or AoCKI, or a disease or condition that may cause renal impairment (e.g., sepsis).

[0041] In some embodiments, if the subject's pre-AKI eGFR is below a predetermined threshold level or within a predetermined range, at least one therapeutically effective dose of AP, such as RecAP, may be administered.

[0042] As used herein, the terms “therapeutic agent” and “drug” also refer to any therapeutic active substance that, when administered to a subject having a disease or disorder, such as renal impairment, e.g., SA-AKI and / or AoCKI, or a disease or condition that may cause renal impairment (e.g., sepsis), produces a desired, usually beneficial, effect. A therapeutic agent may also be a prodrug, which, when administered to a subject, is metabolized to the desired therapeutic active substance. In some embodiments, a therapeutic agent is a prophylactic agent. In addition, a therapeutic agent may be pharmaceutically formulated. A therapeutic agent may also be, or may include, a radioisotope or drug that is activated by some other form of energy, such as light or ultrasonic energy, or by other circulating molecules that can be administered systemically.

[0043] In some aspects of this disclosure, therapeutic agents for use in methods of treating, preventing, or improving symptoms or long-term adverse effects of renal impairment, e.g., SA-AKI and / or AoCKI, or diseases or conditions that may cause renal impairment (e.g., sepsis), e.g., MAKE90, may include AP, e.g., RecAP, alone, or in combination with one or more standard therapeutic agents commonly used for the treatment of renal impairment, e.g., SA-AKI and / or AoCKI, or diseases or conditions that may cause renal impairment (e.g., sepsis).

[0044] As used herein, “therapeutic effective” is the amount of a therapeutic agent that provides some improvement or benefit to a subject having a disease or disorder, e.g., renal impairment, e.g., SA-AKI and / or AoCKI, or a disease or condition that may cause renal impairment (e.g., sepsis). Thus, “therapeutic effective” is the amount that results in some relief, reduction, and / or decrease of at least one clinical symptom or adverse event, e.g., MAKE90, of a disease or disorder, e.g., renal impairment, e.g., SA-AKI and / or AoCKI, or a disease or condition that may cause renal impairment (e.g., sepsis).

[0045] The clinical symptoms associated with diseases or conditions that can cause renal impairment (e.g., SA-AKI and / or AoCKI), or diseases or conditions that can cause renal impairment (e.g., sepsis), that can be treated by the compositions, methods, as specific dosage regimens of this disclosure are well known to those skilled in the art. Furthermore, those skilled in the art will understand that the therapeutic effect does not need to be complete or curative, as long as some benefit is provided to the target. In some embodiments, the term “therapeutic effect” refers to the amount of therapeutic agent that can modify biomarker levels, e.g., endogenous creatinine clearance (ECC) or eGFR, in patients who need it.

[0046] As used herein, “sufficient amount” or “sufficient amount to achieve a particular outcome” in patients with a disease or disorder, e.g., renal impairment, e.g., SA-AKI and / or AoCKI, or a disease or condition that may cause renal impairment (e.g., sepsis) means an amount of an effective therapeutic agent (e.g., an AP such as RecAP) that produces the desired effect, which is optionally a therapeutic effect (i.e., by administering a therapeutically effective dose). In some embodiments, such particular outcome is an improvement in renal function, an increase in survival, and / or a reduction in the risk of experiencing adverse effects, such as MAKE90.

[0047] As used herein, the term “healthcare provider” refers to an individual or institution that directly interacts with and administers to a living subject, e.g., a human patient. Non-exclusive examples of healthcare providers include physicians, nurses, technicians, therapists, pharmacists, counselors, alternative medicine practitioners, medical facilities, clinics, hospitals, emergency rooms, clinics, emergency treatment centers, alternative medicine clinics / facilities, and any other entity that provides advice relating to all or any part of a patient’s health condition, including general and / or specialized treatment, assessment, maintenance, therapy, medicines, and / or advice, but not limited to general medicine, specialized medicine, surgery, and / or any other type of treatment, assessment, maintenance, therapy, medicines, and / or advice.

[0048] As used herein, the term “clinical laboratory” refers to a facility for the testing or processing of materials of living subjects, e.g., human. Non-limiting examples of processing include, for example, biological, biochemical, serological, chemical, immunohematological, hematological, biophysical, cytological, pathological, genetic, or other testing of materials of human bodies for the purpose of providing information for the diagnosis, prevention, or treatment of any disease or disorder of a living subject, e.g., a human, or for the assessment of health. These tests may also include procedures for collecting or otherwise obtaining samples and preparing, determining, measuring, or otherwise describing the presence or absence of various substances in a living subject, e.g., a human body, or a sample obtained from a living subject, e.g., a human body.

[0049] II. Treatment of acute kidney injury in AP In certain aspects, the disclosure relates to alkaline phosphatase (AP) for use in methods for preserving or improving renal function in a population of patients suffering from ckinergic kidney disease (CKI) who have been determined to respond particularly well to treatment with AP.

[0050] The REVIVAL trial identified a statistically significant correlation between CKI severity and thresholds corresponding to clinical outcomes of AP treatment. As discussed above, stratifying patients in the REVIVAL clinical trial according to pre-AKI eGFR identifies specific effects of AP administration on particular subgroups. Parameters(s) defining each of these subgroups can be used, for example, to individualize AP therapy for a particular subgroup, to select patients for treatment, to make decisions related to AP treatment (e.g., modifying dosing or dosage schedules), and / or to assess the likelihood of positive outcomes.

[0051] Thus, in some embodiments, an AP for use in the methods disclosed herein includes administering an AP, such as a RecAP, to a subject for treating AKI in a subject who has been determined to already have mild, moderate, or severe CKI.

[0052] As used herein, the term "moderate chronic kidney disease" refers to chronic kidney injury or renal dysfunction that results in a creatinine clearance rate of 45-60 ml / min / 1.73m 2 of body surface area.

[0053] As used herein, the term "severe chronic kidney disease" refers to chronic kidney injury or renal dysfunction that results in a creatinine clearance rate lower than 45 ml / min / 1.73m 2 of body surface area.

[0054] As used herein, the term "moderate to severe chronic kidney disease" refers to chronic kidney injury or renal dysfunction that results in an eGFR of 15 ml / min / 1.73m 2 to 60 ml / min / 1.73m 2 of body surface area, preferably an eGFR of 25 ml / min / 1.73m 2 to 60 ml / min / 1.73m 2 of body surface area.

[0055] As used herein, the term "mild kidney disease" refers to kidney injury or renal dysfunction that results in a creatinine clearance rate higher than 60 ml / min / 1.73m 2 of body surface area, preferably 60 ml / min / 1.73m 2 to 75 ml / min / 1.73m 2 of body surface area.

[0056] As used herein, the term “pre-AKI eGFR” refers to the estimated glomerular filtration rate prior to the onset of AKI, which is already reduced due to chronic kidney injury (CKI). Typically, the eGFR of a CKI patient is known and regularly determined. In some embodiments, the reduced eGFR due to chronic kidney injury, also referred to herein as pre-AKI eGFR, is determined at least 15 days prior, preferably at least 30 days prior, more preferably at least 60 days prior, and most preferably at least 90 days prior to AP administration. eGFR is an estimate of the actual glomerular filtration rate and is widely used as a measure of renal function.

[0057] In some embodiments, the Disclosure relates to alkaline phosphatase (AP) for use in a method of treating acute kidney injury in a subject requiring treatment, the method comprising administering an effective amount of alkaline phosphatase (AP) to the subject, wherein the pre-AKI eGFR is ≤75 ml / min / 1.73 m². 2 Preferably ≤60 ml / min / 1.73 m 2 The present invention provides alkaline phosphatase (AP). In some embodiments, the pre-AKI eGFR is ≤45 ml / min / 1.73m². 2 In some cases, the pre-AKI eGFR is ≥ 15 ml / min / 1.73 m². 2 Or ≥25ml / min / 1.73m 2 In some cases, before the onset of AKI, the blood flow rate was 15-60 ml / min / 1.73 m². 2 Alternatively, 25-60 ml / min / 1.73 m 2 In some cases, before the onset of AKI, the blood flow rate was 15-45 ml / min / 1.73 m². 2 Alternatively, 25-45 ml / min / 1.73 m 2 That is the case.

[0058] In some embodiments, AP (e.g., RecAP) is administered in doses of at least about 500 U / kg, at least about 600 U / kg, at least about 700 U / kg, at least about 800 U / kg, at least about 900 U / kg, at least about 1000 U / kg, at least about 1100 U / kg, at least about 1200 U / kg, at least about 1300 U / kg, at least about 1400 U / kg, at least about 1500 U / kg, at least about 1600 U / kg, at least about 1700 U / kg, at least about 1800 U / kg, at least about 1900 U / kg, or at least about 2000 U / kg. In some embodiments, AP (e.g., RecAP) is administered in doses exceeding 2000 U / kg. In some embodiments, AP (e.g., RecAP) is administered in doses below 500 U / kg.

[0059] In some embodiments, AP (e.g., RecAP) is administered in doses of approximately 500 U / kg to 1500 U / kg, 600 U / kg to 1400 U / kg, 700 U / kg to 1300 U / kg, 800 U / kg to 1200 U / kg, or 900 U / kg to 1100 U / kg. In some specific embodiments, AP is administered in a dose of approximately 1000 U / kg.

[0060] In some embodiments, AP is human AP. In some embodiments, AP is recombinant AP. In some embodiments, AP is chimeric AP. In certain embodiments, chimeric AP is RecAP (SEQ ID NO: 1). In some embodiments, AP provided herein has at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% sequence identity with respect to the full-length amino acid sequence of SEQ ID NO: 1. In some embodiments, AP is a functional fragment (i.e., a fragment of AP, e.g., an AP that preserves at least about 10%, at least about 20%, at least about 30%, at least 40%, at least about 50%, at least about 60%, at least 70%, at least about 80%, or at least about 90% of the AP activity of the corresponding full-length AP). In some embodiments, AP is a variant or derivative of AP disclosed herein. Other APs that can be used are discussed in detail below.

[0061] In some embodiments, AP is RecAP (e.g., clinical-grade RecAP as used in this disclosure) and is administered in doses of at least about 0.1 mg / kg, at least about 0.2 mg / kg, at least about 0.3 mg / kg, at least about 0.4 mg / kg, at least about 0.5 mg / kg, at least about 0.6 mg / kg, at least about 0.7 mg / kg, at least about 0.8 mg / kg, at least about 0.9 mg / kg, at least about 1 mg / kg, at least about 1.1 mg / kg, at least about 1.3 mg / kg, at least about 1.4 mg / kg, at least about 1.5 mg / kg, at least about 1.6 mg / kg, at least about 1.7 mg / kg, at least about 1.8 mg / kg, at least about 1.9 mg / kg, at least about 2 mg / kg, at least about 2.1 mg / kg, at least about 2.2 mg / kg, at least about 2.3 mg / kg, or at least about 2.4 mg / kg. In some embodiments, AP is administered at doses exceeding 2.4 mg / kg per dose.

[0062] In some embodiments, AP is RecAP (e.g., clinical-grade RecAP as used in this disclosure) and is administered in doses of at least about 100 U / kg, at least about 200 U / kg, at least about 300 U / kg, at least about 400 U / kg, at least about 500 U / kg, at least about 600 U / kg, at least about 700 U / kg, at least about 800 U / kg, at least about 900 U / kg, at least about 1000 U / kg, at least about 1100 U / kg, at least about 1200 U / kg, at least about 1300 U / kg, at least about 1400 U / kg, at least about 1500 U / kg, at least about 1600 U / kg, at least about 1700 U / kg, at least about 1800 U / kg, at least about 1900 U / kg, or at least about 2000 U / kg. In some specific embodiments, AP is RecAP, administered at doses of 500 U / kg to 2000 U / kg.

[0063] In some embodiments, AP is RecAP (e.g., clinical-grade RecAP as used in this disclosure) and is administered in doses of approximately 0.8 mg / kg to approximately 2.4 mg / kg, approximately 0.9 mg / kg to approximately 2.3 mg / kg, approximately 1 mg / kg to approximately 2.2 mg / kg, approximately 1.1 mg / kg to approximately 2.1 mg / kg, approximately 1.2 mg / kg to approximately 2 mg / kg, approximately 1.3 mg / kg to approximately 1.9 mg / kg, approximately 1.4 mg / kg to approximately 1.8 mg / kg, or approximately 1.5 mg / kg to approximately 1.7 mg / kg. In some specific embodiments, AP is administered in a dose of approximately 1.6 mg / kg.

[0064] In some embodiments, AP is RecAP (e.g., clinical-grade RecAP as used in this disclosure) and has a specific activity of at least about 100 U / mg, at least about 200 U / mg, at least about 300 U / mg, at least about 400 U / mg, at least about 500 U / mg, at least about 600 U / mg, at least about 700 U / mg, at least about 800 U / mg, at least about 900 U / mg, at least about 1000 U / mg, at least about 1100 U / mg, at least about 1200 U / mg, at least about 1300 U / mg, at least about 1400 U / mg, at least about 1500 U / mg, at least about 1600 U / mg, at least about 1700 U / mg, at least about 1800 U / mg, at least about 1900 U / mg, or at least about 2000 U / mg.

[0065] In some embodiments, AP is RecAP (e.g., clinical-grade RecAP as used in this disclosure) and has a specific activity of about 1000 U per 1.6 mg.

[0066] In some embodiments, AP is RecAP and has a specific activity of about 600 U / mg to about 700 U / mg, or about 500 U / mg to about 800 U / mg, or about 400 U / mg to about 900 U / mg, or about 300 U / mg to about 1000 U / mg, or about 200 U / mg to about 1100 U / mg, or 100 U / mg to about 1200 U / mg. In some embodiments, AP is RecAP and has a specific activity of less than 100 U / mg. In some embodiments, AP is RecAP and has a specific activity of more than 1200 U / mg.

[0067] In some embodiments, only one dose of AP (e.g., RecAP) is administered per treatment (e.g., one dose per day over 1 to 7 days). In other embodiments, two or more doses of AP are administered. In some embodiments, two, three, four, five, six, seven, eight, nine, ten, eleven, twelfth, thirteenth, twelve, thirteenth, fourteenth, fifteenth, sixteenth, seventeenth, eighteenth, nineteenth, twenty-one doses of AP are administered (e.g., at least two doses per day over 1 to 7 days).

[0068] In some embodiments, the AP dose is administered daily. In other embodiments, the AP dose is administered every 2, 3, 4, 5, 6, or 7 days.

[0069] In some embodiments, a single dose is administered daily. In some embodiments, two, three, or more doses are administered daily.

[0070] In some embodiments, treatment with AP lasts less than approximately 4 days. In some embodiments, treatment with AP lasts less than 3 days, for example, less than 2 days, or less than 1 day.

[0071] In certain embodiments, AP is administered as a daily dose of approximately 1000 U / kg over three consecutive days. In some specific embodiments, when AP is RecAP, AP is administered as a daily dose of 1.6 mg / kg over three consecutive days.

[0072] In some embodiments, each AP dose, for example, RecAP (e.g., clinical-grade RecAP as used in this disclosure) dose is approximately 0.10 mg / kg to approximately 3 mg / kg, or approximately 0.20 mg / kg to approximately 2.9 mg / kg, or approximately 0.3 mg / kg to approximately 2.8 mg / kg, or approximately 0.4 mg / kg to approximately 2.7 mg / kg, or approximately 0.5 mg / kg to approximately 2.6 mg / kg, or approximately 0.6 mg / kg to approximately 2. The dosage is 5 mg / kg, or approximately 0.7 mg / kg to approximately 2.4 mg / kg, or approximately 0.8 mg / kg to approximately 2.3 mg / kg, or approximately 0.9 mg / kg to approximately 2.2 mg / kg, or approximately 1 mg to approximately 2.1 mg / kg, or approximately 1.1 mg / kg to approximately 2 mg / kg, or approximately 1.2 mg / kg to approximately 1.9 mg / kg, or approximately 1.3 mg / kg to approximately 1.8 mg / kg, or approximately 1.4 mg / kg to approximately 1.7 mg / kg.

[0073] In some embodiments, each AP dose, for example, the RecAP dose, is at least about 0.1 mg AP / kg, at least about 0.2 mg AP / kg, at least about 0.3 mg AP / kg, at least about 0.4 mg AP / kg, at least about 0.5 mg AP / kg, at least about 0.6 mg AP / kg, at least about 0.7 mg AP / kg, at least about 0.8 mg AP / kg, at least about 0.9 mg AP / kg, at least about 1 mg AP / kg, at least about 1.1 mg AP / kg, at least about 1.2 mg AP / kg, at least about 1.3 mg AP / kg, at least about 1.4 mg AP / kg, at least about 1.5 mg AP / kg, at least about 1.6 mg AP / kg, at least about 1.7 mg AP / kg, at least about 1.8 mg AP / kg, at least about 1.9 mg AP / kg, at least about 2 mg AP / kg, at least about 2.1 mg AP / kg, at least about 2.2 mg AP / kg, at least about 2.3 mg It contains AP / kg, at least about 2.4 mg AP / kg, at least about 2.5 mg AP / kg, at least about 2.6 mg AP / kg, at least about 2.6 mg AP / kg, at least about 2.7 mg AP / kg, at least about 2.8 mg AP / kg, at least about 2.9 mg AP / kg, or at least about 3 mg AP / kg.

[0074] AP can be administered via different routes, for example, intravenously, rectally, bronchially, or orally. In preferred embodiments, AP is administered intravenously, for example, by intravenous injection or infusion.

[0075] While short-term preservation of renal function can lead to immediate life-saving results, it is preferable for the effects of AP on renal function to be prolonged.

[0076] eGFR can be determined by methods known in the art. Either the Modification of Diet in Renal Disease (MDRD) test equation or the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equation, preferably the CKD-EPI equation, can be used. Those skilled in the art know how to estimate eGFR according to either method, for example by using the website for eGFR calculation provided by the National Kidney Foundation (kidney.org).

[0077] In a preferred embodiment, administration of AP results in a reduced risk of developing a major adverse renal event (MAKE90) by day 90, preferably the reduction in the risk of MAKE90 includes a reduction in the risk of MAKE90 compared to the risk of MAKE90 in the absence of treatment.

[0078] MAKE90 is a clinically meaningful endpoint for patients with kidney disease to predict poor long-term outcomes, including the development of end-stage renal disease (ESRD). The MAKE90 complex includes death, need for dialysis, substantial deterioration of renal function by day 90 (≥25% decrease in estimated glomerular filtration rate (eGFR)), and readmission.

[0079] Preferably, MAKE90 includes one or more of the following events: (i) death before or on day 90, (ii) renal replacement therapy up to day 28, including day 28, or on day 90, (iii) a ≥25% decrease in eGFR on day 90 after treatment with AP initiated against pre-AKI eGFR, and (iv) readmission before or on day 90.

[0080] One or more of these events are: (i) death before or on day 90, (ii) renal replacement therapy before or on day 90, (iii) a ≥25% decrease in eGFR on day 90 after treatment with AP initiated for pre-AKI eGFR, (iv) readmission before or on day 90, (i) death before or on day 90, and (ii) renal replacement therapy before or on day 90, (i) death before or on day 90. (iii) death, and (iii) a ≥25% decrease in eGFR at 90 days after treatment with AP initiated for pre-AKI eGFR, (i) death before or at 90 days, and (iv) readmission before or at 90 days, (ii) renal replacement therapy before or at 90 days, and (iii) a ≥25% decrease in eGFR at 90 days after treatment with AP initiated for pre-AKI eGFR, (ii) before or at 90 days This may include (iv) rehospitalization on day 90, and (iii) a ≥25% decrease in eGFR on day 90 after treatment with AP initiated for pre-AKI eGFR, and (iv) rehospitalization on day 90, (i) death on day 90, (ii) rehospitalization on day 90, and (iii) a ≥25% decrease in eGFR on day 90 after treatment with AP initiated for pre-AKI eGFR, (i) death on day 90, (ii) rehospitalization on day 90, (ii) rehospitalization on day 90, and (iv) rehospitalization on day 90, or (ii) rehospitalization on day 90, and (iii) a ≥25% decrease in eGFR on day 90 after treatment with AP initiated for pre-AKI eGFR.

[0081] In some embodiments, AP (e.g., RecAP) is administered to the subject only if sepsis is detected less than 96 hours before treatment is initiated. In other embodiments, AP is administered only if sepsis is detected less than 72 hours before AKI detection.

[0082] In some aspects of this disclosure, treatment with AP is initiated within 48 hours or 24 hours after sepsis is detected. The presence of sepsis can be detected, for example, as disclosed in the Examples section of this application.

[0083] In some embodiments, treatment is initiated within 48 hours, preferably within 24 hours, of the detection of AKI in the subject.

[0084] In some embodiments, administration of at least one dose of AP results in a reduction or discontinuation of renal replacement therapy (RRT) in subjects receiving RRT.

[0085] In some embodiments, administration of at least one dose of AP results in preservation or increase of glomerular filtration rate (GFR) in a subject. GFR can be assessed in several ways, for example, by inulin or chromium EDTA clearance, or an approximation of GFR can be made, for example, by calculating endogenous creatinine clearance (ECC). This is calculated from measured 24-hour urine volume, urinary creatinine levels, and serum creatinine levels. GFR can also be estimated based on serum creatinine (eGFR).

[0086] In some embodiments disclosed herein, administration of AP prevents a reduction in renal function below a critical threshold, which would result in an increase in renal function or eliminate the need for administration of certain treatments, such as antibiotics to treat sepsis. Thus, in some embodiments, AP administration can prevent a reduction in renal function below a critical threshold, thereby enabling such a person to receive treatment. Thus, in some embodiments, an indicator of renal function (e.g., ECC or eGFR) is determined before administering AP to preserve renal function in order to determine the risk that the person's renal function will be reduced below a certain threshold level.

[0087] In some embodiments, the present invention includes detecting changes in markers of renal function, such as ECC, eGFR, or blood urea nitrogen (BUN) clearance, either alone or in combination with the detection of changes at the levels of one, two, three, or more biomarkers.

[0088] In some embodiments, the present invention includes predicting an increased clinical response to therapy with AP, e.g., RecAP, based on detected renal function parameters (e.g., pre-AKI eGFR). In some embodiments, the disclosure includes evaluating whether a renal function parameter (e.g., pre-AKI eGFR) is within a certain range or above or below a certain threshold (e.g., an eGFR threshold for the severity of chronic kidney disease). Thus, for example, if a renal function parameter (e.g., pre-AKI eGFR), alone or in combination with other biomarkers, indicates that a patient will benefit from therapy with AP, the therapy may be initiated, maintained, or modified (e.g., increasing or decreasing the dose, or increasing or decreasing the dose frequency).

[0089] Conversely, if, for example, renal function parameters (e.g., pre-AKI eGFR), either alone or in combination with other biomarkers, indicate that the patient does not benefit from AP therapy, the therapy may be discontinued, temporarily interrupted, or modified (e.g., by increasing or decreasing the dose, or increasing or decreasing the dose frequency).

[0090] In other words, specific levels of renal function parameters (e.g., pre-AKI eGFR), either alone or in combination with other molecules or clinical biomarkers, correlate with the clinical efficacy of AP therapy and are useful in predicting clinical outcomes in specific populations of patients with CKI in addition to sepsis and / or AKI.

[0091] The present invention thus provides alkaline phosphatase (AP) for use in a method for treating acute kidney injury (AKI) in a subject requiring treatment, wherein the method comprises administering an effective amount of AP to the subject, the subject having an acute exacerbation of chronic kidney injury (CKI) which is moderate to severe. In a preferred embodiment, the subject receives 15-60 ml / min / 1.73m². 2 The patient has a pre-AKI eGFR. In a more preferred embodiment, the patient receives 25-60 ml / min / 1.73 m 2 The patient has a pre-AKI eGFR. Pre-AKI eGFR, as used herein, means that the eGFR is reduced due to chronic kidney disease and is determined at least 15 days, preferably at least 30 days, more preferably at least 60 days, and most preferably at least 90 days before administration of AP.

[0092] In a preferred embodiment, an AP for use according to the present invention is provided, wherein the AP is a human AP.

[0093] In a preferred embodiment, AP is recombinant AP, preferably a chimeric AP, and more preferably has at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100% sequence identity with respect to the amino acid sequence of RecAP (SEQ ID NO: 1). This sequence identity is preferably determined across the full-length protein as shown in SEQ ID NO: 1.

[0094] In a preferred embodiment, AP is provided for use according to the present invention, and administration of AP results in a reduced risk of developing a major adverse kidney event (MAKE90) by day 90, preferably the reduction in the risk of MAKE90 includes the reduction in the risk of MAKE90 compared to the risk of MAKE90 in the absence of treatment.

[0095] In a preferred embodiment, an AP for use according to the present invention is provided, and sepsis is detected less than 96 hours before AP administration.

[0096] In preferred embodiments, an AP for use according to the present invention is provided, in which sepsis is detected less than 72 hours before AKI detection. Preferably, treatment is initiated within 24 hours after sepsis is detected and / or after AKI is detected. In preferred embodiments of the present invention, AKI is sepsis-associated AKI (SA-AKI). SA-AKI is used to describe AKI that is caused by or associated with sepsis.

[0097] In a preferred embodiment, AP for use according to the present invention is provided, and AP is administered once daily. In a preferred embodiment, AP is administered intravenously. In a preferred embodiment, AP is administered in three daily doses.

[0098] In a preferred embodiment, an AP for use according to the present invention is provided, where the AP is RecAP, and the dose is 0.06 mg / kg to 3.2 mg / kg, or 375 U / kg to 2,000 U / kg, preferably 0.08 mg / kg to 3.2 mg / kg or 500 U / kg to 2,000 U / kg.

[0099] In a preferred embodiment, AP for use according to the present invention is provided, in which administration of at least one dose of AP results in a reduction or discontinuation of the duration of renal replacement therapy (RRT) in a subject receiving RRT.

[0100] In one preferred embodiment, AP is provided for use according to the present invention, wherein administration of at least one dose of AP results in preservation or increase of glomerular filtration rate (GFR) in a subject.

[0101] As used herein, the term “biomarker” refers to a factor that is a characteristic indicator of a biological process, biological event, and / or pathological condition, e.g., a predictor of the clinical response to treatment in AP, e.g., RecAP. As used herein, the term biomarker encompasses both clinical markers and molecular biomarkers (biological markers). Thus, in the context of this disclosure, the term “biomarker” encompasses, for example, “biological biomarker” or “molecular biomarker.” In some embodiments, biological or molecular biomarkers used to assess renal function include markers of liver function (e.g., alanine aminotransferase, aspartate aminotransferase, gamma-glutamyltransferase, bilirubin, or LDH), C-reactive protein (CRP), interleukin-6 (IL-6), interleukin-18 (IL-18), lipopolysaccharide-binding protein, kidney injury molecule 1 (KIM-1), or a combination thereof.

[0102] As disclosed above, the term “biomarker” also encompasses “clinical biomarkers,” also called “clinical status markers,” which can predict a response to biological therapy, such as sex, age, concomitant medications, smoking status, and body mass index (BMI).

[0103] As discussed above, eGFR that decreases due to chronic kidney disease (referred to as pre-AKI eGFR) is used for the cutoff approach. Therefore, the target pre-AKI eGFR is 60 ml / min / 1.73m². 2 If the threshold is below this, the individual may be a candidate for treatment with a specific AP therapy, such as a specific AP regimen, such as one or more doses of RecAP.

[0104] AP for use in the methods disclosed herein includes prescribing, initiating, and / or modifying, for example, prevention and / or treatment for AKI, either alone or in combination with one or more additional biomarkers, based at least partially on the subject's past eGFR (or another renal function parameter). Past means that the eGFR was determined before the patient was hospitalized, for example, at least 15, 30, 60, or 90 days before the administration of AP was initiated.

[0105] This disclosure provides a method for determining whether to treat a patient with AoCKI with a treatment regimen including the administration of AP, the method comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) if it is determined that the patient has a higher or lower pre-AKI eGFR compared to a predetermined threshold level(s), treating the patient with a treatment regimen including the administration of AP, for example, RecAP, or instructing a healthcare provider to treat the patient, or discontinuing treatment, not initiating treatment, refusing treatment, or instructing a healthcare provider to discontinue, not initiate, or refuse treatment. A method is also provided for determining whether a patient with AKI is likely to respond to a treatment regimen including the administration of AP, comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) if the patient is determined to have a higher or lower pre-AKI eGFR compared to a predetermined threshold level(s), determining whether the patient is likely or unlikely to respond to treatment with a treatment regimen including the administration of AP, for example, RecAP.

[0106] In one embodiment, the Disclosure provides a method for determining whether to treat a patient with AoCKI with a therapeutic regimen including the administration of AP, the method comprising: (a) measuring the levels of eGFR and, optionally, additional biomarkers such as kidney injury molecule 1 (KIM-1) in a sample taken from the patient, or instructing a clinical laboratory to do so; and (b) if it is determined that the patient has lower or reduced eGFR and higher or increased levels of at least one optional additional biomarker, such as KIM-1, in the sample compared to a predetermined biomarker threshold level or compared to biomarker levels in one or more controls, or instructing a healthcare provider to do so.

[0107] A method is also provided for determining whether a patient having AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, is likely to respond to a treatment regimen including administration of AP, comprising: (a) measuring the level of eGFR and, optionally, additional biomarkers such as KIM-1 in a sample taken from the patient, or instructing a clinical laboratory to do so; and (b) determining that the patient is likely to respond to a treatment regimen including administration of AP if it is determined that the patient has a lower or reduced eGFR and a higher or increased level of at least one optional additional biomarker, such as KIM-1, in the sample compared to a predetermined biomarker threshold level or compared to biomarker levels in one or more controls.

[0108] In one embodiment, the Disclosure provides a method for determining whether to treat a patient having AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, with a treatment regimen including the administration of AP, the method comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) if it is determined that the patient has a higher or increased pre-AKI eGFR compared to a predetermined biomarker threshold level, discontinuing treatment of the patient with a treatment regimen including the administration of AP, e.g., RecAP, not initiating treatment, refusing treatment, or instructing a healthcare provider to discontinue, not initiate, or refuse treatment.

[0109] A method is also provided for determining whether to treat a patient having AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, with a treatment regimen including the administration of AP, comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) determining that if the patient is determined to have a higher or increased pre-AKI eGFR compared to a predetermined biomarker threshold level, the patient is likely to respond to a treatment regimen including the administration of AP, e.g., RecAP, to the patient.

[0110] A method is also provided for selecting a patient diagnosed with AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, as a candidate for treatment with AP, comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) treating the patient with AP, or instructing a healthcare provider to treat the patient with AP, if it is determined that the patient has a lower or reduced pre-AKI eGFR compared to a predetermined threshold level(s).

[0111] A method is also provided for selecting patients diagnosed with AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, as candidates for treatment with AP, comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) determining that if the patient is determined to have a lower or reduced pre-AKI eGFR compared to a predetermined threshold level(s), the patient is likely to respond to treatment with AP.

[0112] A method is also provided for selecting a patient diagnosed with AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, as a candidate for treatment with AP, comprising (a) obtaining a pre-AKI eGFR value from the patient, and (b) if it is determined that the patient has a higher or increased pre-AKI eGFR compared to a predetermined threshold level(s), discontinuing treatment of the patient with AP, e.g., RecAP, not initiating treatment, refusing treatment, or instructing the healthcare provider to discontinue, not initiate, or refuse treatment.

[0113] A method is also provided for selecting patients diagnosed with AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, as candidates for treatment with AP, comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) determining that if the patient is determined to have a higher or increased pre-AKI eGFR compared to a predetermined threshold level, the patient is less likely to respond to treatment with AP, e.g., RecAP. The predetermined eGFR threshold level is approximately 60 ml / min / 1.73m2.

[0114] Pre-AKI eGFR values ​​can be obtained, for example, from the patient's medical records. The term “medical records” or “patient medical records” typically refers to a description of a patient’s examinations and / or treatments, including one or more of the following: the patient’s medical history and complaints, the physician’s physical findings, the results of diagnostic tests and procedures, and the patient’s medications and treatment procedures. Medical records are typically prepared by one or more physicians and / or physicians’ assistants and are written, transcribed, or otherwise recorded records and / or histories of various illnesses or disorders that require frequent health information about medical treatment, and / or vaccinations, and / or allergies, and / or treatments, and / or prognosis, and / or parents, siblings, and / or occupations. Records may be reviewed by the physician in diagnosing the condition.

[0115] Medical records may be in paper form and / or maintained on computer-readable media. Medical records may be maintained by laboratories, clinics, hospitals, health maintenance organizations, insurance companies, and / or personal medical record websites.

[0116] In some embodiments, a diagnosis based at least in part on the measured eGFR is recorded on or in a medical warning item such as a card, wearable item, and / or radio frequency identification (RFID) tag. As used herein, the term “wearable item” means any item that can be worn on the subject’s body, including but not limited to tags, bracelets, necklaces, armbands, or headbands.

[0117] As used herein, the term “diagnosis” means detecting a disease or determining the stage or severity of a disease. Typically, a diagnosis of a disease is based on an assessment of one or more factors and / or symptoms that indicate the disease. That is, a diagnosis can be made based on the presence, absence, or amount of factors that indicate the presence or absence of a disease or disorder. Each factor or symptom that is considered to indicate a diagnosis of a particular disease does not have to be exclusively related to that particular disease; for example, there may be differential diagnoses that can be inferred from the diagnostic factors or symptoms. Similarly, there may be cases where factors or symptoms that indicate a particular disease are present in an individual who does not have that particular disease.

[0118] The term “diagnosis” also encompasses determining the therapeutic effect of drug therapy, such as AP therapy, or predicting patterns of response to drug therapy. Diagnostic methods may be used independently or in combination with other diagnostic and / or staging methods known in the medical field for a particular disease.

[0119] As used herein, the term “differential diagnosis” refers to the determination, based on the analysis of clinical data, of which of two or more diseases with similar symptoms is most likely to be the cause of the symptoms in question. The term is also used to refer to the determination of whether a patient is susceptible to treatment with AP, depending on whether the measured eGFR in a patient sample is above or below a predetermined threshold level, or whether it is elevated or decreased compared to the level in one or more controls.

[0120] As used herein, the term “prognosis” refers to the predicted course and outcome of a clinical condition or disease, such as sepsis or SA-AKI. Prognosis is usually diagnosed by evaluating disease factors or symptoms that indicate a favorable or unfavorable course or outcome of the disease. As used herein, the phrase “determining the prognosis” refers to the process by which a person skilled in the art can predict the course or outcome of a condition in a patient. The term “prognosis” does not imply the ability to predict the course or outcome of a condition with 100% accuracy. Rather, a person skilled in the art will understand that the term “prognosis” refers to the increased probability that a particular course or outcome will occur, i.e., that the course or outcome is more likely to occur in a patient exhibiting a given condition compared to an individual not exhibiting the condition.

[0121] As used herein, the terms “favorable prognosis” and “positive prognosis,” or “unfavorable prognosis” and “negative prognosis,” are relative terms for predicting the likely course and / or likely outcome of a condition or disease, such as sepsis or SA-AKI. A favorable or positive prognosis predicts a better outcome for the condition than an unfavorable or negative prognosis. In a general sense, a “favorable prognosis” is a relatively better outcome than many other possible prognoses that may be associated with a particular condition, while an unfavorable prognosis predicts a relatively worse outcome than many other possible prognoses that may be associated with a particular condition. Typical examples of a favorable or positive prognosis include increased renal function, preservation of renal function, increases in ECC or eGFR (or another renal function parameter), and reduced risk of adverse events, such as MAKE90.

[0122] This disclosure provides a method for treating a patient having AoCKI or AP, for use in a method for treating AoCKI in a subject, comprising administering AP to the patient when it is determined that the patient has a pre-AKI eGFR that is lower or reduced compared to a predetermined eGFR threshold level.

[0123] The Disclosure also provides an AP for use in a method of treating a patient with AoCKI, the method comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) administering the AP to the patient if the patient has a pre-AKI eGFR that is lower or reduced compared to a predetermined eGFR threshold level.

[0124] Also provided is an AP for use in a method of treating a patient with AoCKI, the AP comprising (a) obtaining a pre-AKI eGFR value from the patient, and (b) if the patient has a higher or increased pre-AKI eGFR compared to a predetermined eGFR threshold level, discontinuing or not initiating administration of the AP, e.g., RecAP, to the patient.

[0125] The Disclosure also provides an AP for use in a method of treating a patient having AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, the AP comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) determining whether the pre-AKI eGFR value is higher or increasing, or lower or decreasing, compared to a predetermined eGFR threshold level.

[0126] In some embodiments, the method further includes administering or advising a healthcare provider to administer an AP, such as RecAP, to a patient if it is determined that the patient has a lower or reduced pre-AKI eGFR compared to a predetermined eGFR threshold level, or discontinuing or refusing to administer an AP if it is determined that the patient has a higher or increased pre-AKI eGFR level compared to a predetermined eGFR threshold level.

[0127] AP for use in methods of treating patients having AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, the AP comprising: (a) obtaining a pre-AKI eGFR value from the patient; and (b) administering the AP to the patient if it is determined that the patient has a lower or reduced pre-AKI eGFR compared to a predetermined eGFR threshold level, or discontinuing, not initiating, or refusing to administer the AP to the patient if it is determined that the patient has a higher or increased pre-AKI eGFR in the sample compared to a predetermined eGFR threshold level.

[0128] The disclosure also provides a method for determining the efficacy or pharmacodynamics of an AP in a patient diagnosed with AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, comprising: (a) performing a first measurement of the patient's eGFR in a first sample taken from the patient; (b) administering an AP, e.g., RecAP; and (c) performing a second measurement of the patient's eGFR in a second sample taken from the patient, wherein an increase or preservation of the eGFR in the second measurement compared to the patient's eGFR in the first measurement indicates that the patient is responding to treatment with an AP, e.g., RecAP.

[0129] The disclosure also provides a method for determining the efficacy or pharmacodynamics of an AP in a patient diagnosed with AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, comprising: (a) performing a first measurement of the patient's eGFR in a first sample taken from the patient; and (b) performing a second measurement of the patient's eGFR in a second sample taken from the patient after the patient has been given an AP, e.g., RecAP, wherein an increase or preservation of the eGFR in the second measurement compared to the patient's eGFR in the first measurement indicates that the patient is responding to treatment with an AP, e.g., RecAP.

[0130] The disclosure also provides a method for measuring the efficacy or pharmacodynamics of an AP in a patient diagnosed with AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, comprising: (a) performing a first measurement of the patient's eGFR in a first sample taken from the patient; (b) administering an AP, e.g., RecAP; and (c) performing a second measurement of the patient's eGFR in a second sample taken from the patient, wherein a decrease in the eGFR in the second measurement compared to the patient's eGFR in the first measurement indicates that the patient is not responding to treatment with an AP, e.g., RecAP.

[0131] The disclosure also provides a method for measuring the efficacy or pharmacodynamics of an AP in a patient diagnosed with AKI, preferably sepsis-associated AKI (SA-AKI) and / or AoCKI, comprising: (a) performing a first measurement of the patient's eGFR in a first sample taken from the patient; and (b) performing a second measurement of the patient's eGFR in a second sample taken from the patient after the patient has been given an AP, e.g., RecAP, wherein a decrease in the eGFR in the second measurement compared to the patient's eGFR in the first measurement indicates that the patient is not responding to treatment with an AP, e.g., RecAP.

[0132] In some embodiments, the second measurement is performed at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, or 90 days after administration of AP, e.g., RecAP, or at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16 weeks, or at an intervening time.

[0133] In certain embodiments, an AP "loading" dose is administered to achieve a desired level of renal function in the patient. If the AP loading dose does not significantly affect the patient's renal function, a decision may be made to discontinue treatment, for example, to switch to an alternative therapy.

[0134] If the loading dose results in increased renal function in the patient, a decision may be made to reduce the AP dose size or frequency to a “maintenance” dose. It is important to note that the methods provided herein are guidelines for administering treatment for healthcare providers, and the final treatment decision will be based on the sound judgment of the healthcare provider.

[0135] The formulations, dosage regimens, and routes of administration of AP, such as RecAP, can be adjusted to provide an effective dose for an optimal therapeutic response according to the methods disclosed herein. With regard to the administration of AP, AP may be administered through any suitable means, compositions, and routes known in the art. With regard to dosage regimens, a single bolus may be administered, several divided doses may be administered over time, or the dose may be proportionally reduced or increased as indicated by the urgency of the treatment situation.

[0136] III. Alkaline phosphatase (AP) Alkaline phosphatase (AP, EC 3.1.3.1 according to IUBMB enzyme nomenclature) is an enzyme that catalyzes the reaction of phosphatase monoesters and H2O to alcohols and phosphates. Other names for AP include alkaline phosphomonoesterase, phosphomonoesterase, glycerophosphatase, alkaline phosphohydrolase, alkaline phenylphosphatase, and orthophosphophosphate monoester phosphohydrolase (alkaline optimal). The systematic name for AP is phosphate monoester phosphohydrolase (alkaline optimal).

[0137] APs are enzymes of broad specificity and also catalyze transphosphorylation. In humans and other mammals, at least four different but related APs are known. These are intestinal, placental, placental-like, and liver / bone / kidney (or tissue-nonspecific) APs. The first three are located together on chromosome 2, while the tissue-nonspecific forms are located on chromosome 1.

[0138] The term "AP as used in this disclosure" refers to isolated alkaline phosphatases, including their splice variants, isoforms, and polymorphic forms. Recombinant APs and chimeric APs are also included. In certain embodiments, AP is RecAP. The amino acid sequence of RecAP is shown in Figure 1.

[0139] In some embodiments, the APs disclosed herein have sequence identity of at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% with respect to the amino acid sequence of SEQ ID NO: 1.

[0140] In some embodiments, AP is a functional fragment (i.e., a fragment of AP, e.g., AP that preserves at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least 70%, at least about 80%, or at least about 90% of the AP activity of the corresponding full-length AP). In some embodiments, AP is a variant or derivative of AP disclosed herein.

[0141] AP for use as disclosed herein may be a commercially available AP enzyme, or any composition comprising an AP enzyme and any means capable of producing a functional AP enzyme in the context of the present invention, such as DNA or RNA nucleic acid encoding the AP protein.

[0142] Nucleic acids encoding AP can be embedded in suitable vectors such as plasmids, phagemids, phages, (retro)viruses, transposons, gene therapy vectors, and other vectors capable of inducing or conferring AP production. Natural or recombinant microorganisms, such as bacteria, fungi, protozoa, and yeasts, can also be used as sources of AP in the context of this disclosure.

[0143] AP-containing compositions for use in accordance with this disclosure may comprise eukaryotic APs, e.g., mammalian APs, which may be tissue-nonspecific APs, such as liver-bone or kidney types, or tissue-specific types, such as placental APs, intestinal APs, and placental-like APs. The latter, also known as germ cell APs, are localized in the testes, thymus, and certain germ cell tumors and are closely associated with both the placental and intestinal forms of APs.

[0144] In some embodiments, mammalian APs are human or bovine APs. Non-restrictive examples of human AP sequences can be found in the NCBI(Genpept) collection and include NP_001622 (intestinal AP), NP_001623 (placental AP), NP_112603 (placental-like AP), or NP_000469 (tissue-nonspecific AP). In some embodiments, APs include polymorphisms. In some embodiments, APs are placental APs, placental-like APs, intestinal APs, liver / bone / kidney APs, or combinations thereof. In some embodiments, APs are recombinant APs.

[0145] From a structural standpoint, AP consists of approximately two domains: a crown domain and an active site domain. The active site domain can be divided into separate parts, such as a catalytic residue and three metal ion sites (Zn1, Zn2, and Mg3). From a primary structural standpoint, the crown domain is adjacent to the amino acids that form the active site domain. The amino acid sequence of AP, as well as the relative positions of the catalytic domain and the crown domain, are known to those skilled in the art.

[0146] In some aspects of this disclosure, AP is an isolated or recombinant AP comprising a crown domain and a catalytic domain, wherein the crown domain and the catalytic domain are obtained from different APs, and at least one of the different phosphatases is a human phosphatase. In some aspects, AP is, for example, ECAP (Escherichia coli AP) or one of the seven known BIAPs (bovine intestinal APs).

[0147] In some embodiments, the AP is an isolated or recombinant AP comprising a crown domain and a catalytic domain, wherein the crown domain and the catalytic domain are obtained from a different AP, and the different AP is a human AP. This is particularly useful when the modified phosphatase is subsequently used in human therapy. The AP for use in the disclosed methods may be a human-derived modification, such as a genetically modified AP, which is not immunogenic or is only weakly immunogenic.

[0148] The modified APs disclosed herein can be used, for example, in "in vitro" or "ex vivo" diagnostic or therapeutic applications. Such modified phosphatases may include, for example, human and E. coli APs, or may consist of bovine and E. coli APs.

[0149] In some embodiments of this disclosure, AP is an isolated or recombinant AP comprising a crown domain and a catalytic domain, wherein the crown domain and the catalytic domain are obtained from different APs, the crown domain being the crown domain of placental AP (ALPP) and the catalytic domain being the catalytic domain of intestinal AP (ALPI). In some embodiments, at least one of the different APs is a human phosphatase. In other embodiments, both different APs are human phosphatases.

[0150] Domain-exchange variants suitable for the methods disclosed herein, which are based on human AP, are listed in Table 1. [Table 1]

[0151] In some embodiments, AP is a combination of the catalytic domain of ECAP or a human form (ALPI, ALPP, GCAP, or TNAP) with the crown domain of BIAP. Furthermore, a combination of the crown domain of BIAP with either the catalytic domain or a human form of ECAP can also be produced.

[0152] In some embodiments, modified APs are APs that, under natural conditions, bind to the cell membrane via a glycosylphosphatidylinositol (GPI) anchor, but are modified to no longer bind to the cell membrane. All isoenzymes are functionally active at the cell membrane, and GPI-anchor-deficient forms do not naturally exist at detectable levels. Serum AP activity has been shown, but the enzyme is generally still found in the decidual fraction or membrane vesicles. Milk AP activity is also found in fractions containing membrane vesicles. GPI anchors are stored as precursor molecules in cells, binding to the binding site via transamidases. The GPI-anchor skeleton is identical in mammals, but cell-type-dependent modifications are known.

[0153] In some embodiments, for the treatment of human subjects, AP is human. This is primarily due to the fact that AP forms derived from other species may be immunogenic in human subjects, and treatment may induce immunological reactions and pathological side effects. In some subjects, lethal side effects, namely anaphylactic shock, may occur, and therefore, it is preferable to minimize the risk of immunological side effects by using human AP forms.

[0154] Because isolating AP from humans is impractical, recombinant human AP proteins can be routinely produced on different recombinant expression platforms. However, the expression and purification of GPI-modified and membrane-anchored proteins are notoriously difficult, as GPI proteins are difficult to separate from membranes and are difficult to isolate and purify. Therefore, in some embodiments, recombinant AP involves modifications in the GPI signaling sequence, which result in secreted AP, i.e., AP that is not bound to the cell membrane.

[0155] There is no common sequence that causes GPI anchor binding, but there are some specific consensus features: Hydrophobic stretch of C-terminal amino acids (at least 11 amino acids, but preferably more than 11 amino acids), Spacers of hydrophilic amino acids (5-12 amino acids) located upstream of the hydrophobic region, GPIs are bound to small amino acids: glycine, aspartic acid, asparagine, alanine, serine, or cysteine, and The two subsequent amino acids downstream of the GPI binding site must be small amino acids, and in most cases they are selected from glycine, aspartic acid, asparagine, alanine, serine, or cysteine.

[0156] In some embodiments, recombinant AP comprises modifications in the GPI signaling sequence, such modifications result in secreted AP that is biologically active, i.e., exhibits activity toward a biologically relevant substrate.

[0157] In some embodiments, secreted AP is human AP. In some embodiments, secreted human AP is human hepatic-renal-bone phosphatase, human intestinal AP, or human placental-like alkaline phosphatase.

[0158] Based on the consensus features described above, those skilled in the art can introduce modifications that disrupt some of the consensus results in APs that cannot bind to the GPI anchor, for example, by inserting one or more amino acids. Thus, in some embodiments, recombinant APs include modifications in the GPI signaling sequence that result in secreted APs, the modifications including mutations or deletions of at least one amino acid in the sequence encompassing the consensus GPI signaling sequence.

[0159] In some embodiments, AP is the AP disclosed in U.S. Patent No. 8,557,545. In some embodiments, AP is a chimeric AP or chimeric AP-like protein, such as those described in U.S. Patent Application Publications 2017 / 0009216 and 2014 / 0193388. In some specific embodiments of this disclosure, AP is a recombinant alkaline phosphatase containing the catalytic domain of ALPI (enteric alkaline phosphatase) and the crown domain of ALPP (placental alkaline phosphatase), e.g., RecAP (SEQ ID NO: 1). RecAP is also known as catALPI / crownALPP, Xinplap, and sALPI-ALPP-CD. In some embodiments, AP is an improved RecAP, e.g., LVL-RecAP (corresponding to SEQ ID NO: 1 in U.S. Patent Application Publication 2017 / 0009216).

[0160] In some embodiments, the AP of the present disclosure comprises (i) a sequence having at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least 99% sequence identity with the crown domain of human ALPP, and (ii) a sequence having at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least 99% sequence identity with the catalytic domain of human ALPI.

[0161] In some embodiments, the sequence having sequence identity with the crown domain of ALPP is located in the protein according to the present invention at approximately the same position as the crown domain of ALPP in the natural ALPP protein.

[0162] The percentage of amino acid or nucleic acid sequence identity, or the term "sequence identity %", is defined herein as the percentage of residues in a candidate amino acid or nucleic acid sequence that are identical to residues in a reference sequence after aligning two sequences and introducing gaps as necessary to achieve the maximum identity percentage. In preferred embodiments, the calculation of at least this percentage of sequence identity is performed without introducing gaps. Methods and computer programs for alignment, such as "Align 2" or the BLAST service of the National Center for Biotechnology Information (NCBI), are well known in the art. [Examples]

[0163] Example 1 - REVIVAL Human recombinant alkaline phosphatase for sepsis-associated acute kidney injury method Overall design The REVIVAL trial was a phase 3, multicenter, randomized, double-blind, placebo-controlled, two-arm parallel-group, sequentially designed pivotal trial in which patients with SA-AKI were randomly assigned in a 1:1 ratio to receive either placebo or 1.6 mg / kg of RecAP.

[0164] Randomization was stratified by the following: ●Pre-AKI reference eGFR ≥ 25 and < 45 mL / min / 1.73m² 2 "Moderate" chronic kidney disease (CKD) is defined as ●Baseline-modified continuous organ failure assessment (mSOFA) score, i.e., excluding the Glasgow Coma Scale (GCS) portion (≤9, >9) ●Clinical facilities

[0165] Based on this, we defined three different SA-AKI trial populations: 1. Primary study population: Pre-AKI reference eGFR ≥ 45 mL / min / 1.73 m² 2 Patients who have the following characteristics and do not have confirmed or suspected COVID-19 at the time of randomization, 2. "Moderate" CKD population: Pre-AKI reference eGFR ≥ 25 and < 45 mL / min / 1.73m² 2 Patients who have the following characteristics and do not have confirmed or suspected COVID-19 at the time of randomization. 3. COVID-19 population: Patients with confirmed or suspected COVID-19 at the time of randomization, regardless of whether they had "moderate" CKD. For patients in this population, COVID-19 must have been the primary cause of SA-AKI.

[0166] An independent randomized list has been created for each of the three groups.

[0167] Patients were enrolled at approximately 100 sites, primarily across Europe, North America, Japan, and Australia. The final number of patients to be enrolled was set to depend on the recommendations of the Data Monitoring Committee (DMC) based on safety data review and interim analysis of futility / success. Individual trial flows for patients are shown in Figure 1.

[0168] Inclusion criteria To be eligible for this trial, patients must meet all five of the following inclusion criteria. 1. 18 years of age or older. 2. The patient is in the ICU or intermediate care unit for clinical reasons. 3. Having sepsis requiring vasopressor therapy (norepinephrine, epinephrine, dopamine, phenylephrine, vasopressin, or angiotensin II), i.e.: a. Suspected or confirmed bacterial or viral infection, and b. Despite appropriate fluid resuscitation in accordance with clinical judgment, the patient receives vasopressor therapy (≥0.1 μg / kg / min norepinephrine or equivalent) for at least one hour for sepsis-induced hypotension. Any dose of vasopressor is counted as vasopressor therapy after at least one hour following the first dose of 0.1 μg / kg / min norepinephrine or equivalent. The combination of a) and b) automatically ensures that the patient meets the sepsis criterion 3, as 0.1 μg / kg / min of norepinephrine corresponds to a score of +4 in the cardiovascular subscore of the SOFA score. 4. The following Kidney Diseases: Having an AKI that meets at least one of the Improving Global Outcomes (KDIGO) criteria a-d: a. An absolute increase in serum or plasma creatinine (CR) of ≥0.3 mg / dL (≥26.5 μmol / L) within 48 hours. b. A relative increase in complete criterion (CR) up to ≥1.5 times the pre-AKI reference CR value, which is known or presumed to have occurred within the past 7 days. c. A reduction in urine output to <0.5 mL / kg / hour for at least 6 hours after appropriate fluid resuscitation. d. If the patient has no known history of CKD and there is no pre-AKI reference CR value available from the past 12 months: A CR value greater than or equal to the levels shown in Table 2, with an increase in CR estimated to have occurred within the last 7 days. 5. Provision of a signed and dated Informed Consent Form (ICF) in accordance with local regulations. [Table 2]

[0169] Exclusion criteria Patients who meet any of the following criteria will be excluded from participation in this study: 1. Recorded CKD specified below: a) In selected facilities where enrollment of “moderate” CKD patients is permitted: Pre-AKI reference eGFR < 25 mL / min / 1.73 m² 2 "Severe" CKD is defined as such. ●For patients with known chronic kidney disease, the most recent eGFR prior to the initial hospitalization should be ≥25 mL / min / 1.73 m². 2 It needs to be recorded as such. ● For patients with a known history of chronic kidney disease (CKD) prior to hospitalization but without a known eGFR, the recommended blood flow rate was 25-60 mL / min / 1.73 m². 2 Using the presented eGFR, it is also possible to exclude "severe" CKD. b) At all other facilities: "Moderate" and "severe" chronic kidney disease (CKD) are defined as having a pre-AKI reference eGFR < 45 mL / min / 1.73 m2. ●For patients with known chronic kidney disease, the most recent eGFR prior to the initial hospitalization should be ≥45 mL / min / 1.73 m². 2 It needs to be recorded as such. ● For patients with a known history of chronic kidney disease (CKD) prior to hospitalization but without a known eGFR, the recommended blood flow rate was 45-60 mL / min / 1.73 m². 2 Using the presented eGFR, it is also possible to exclude "moderate" and "severe" CKD. 2. A progressive chronic liver disease defined as a Childpues score (Class C) of 10-15. 3. Acute pancreatitis without an established source of infection. 4. Urinary tract sepsis associated with suspected or confirmed urinary tract obstruction. 5. Major causes of AKI that are not sepsis. 6. Confirmed or suspected SARS-CoV-2 infection. This exclusion criterion does not apply to patients in the COVID-19 population. 7. Severe burns requiring ICU treatment. 8. For example, severe immunosuppression due to: ● Hematopoietic stem cell transplantation or acute or chronic graft-versus-host disease within the past 6 months prior to screening, ●Solid organ transplantation, ● Sepsis, i.e., leukopenia unrelated to preceding sepsis, ● Human immunodeficiency virus (HIV) / Acquired immunodeficiency syndrome (AIDS) ● The individual has received chemotherapy within 30 days prior to screening. 9. For example, they are at high risk of becoming untraceable (LTFU) because they are known to have a known current or recent (within the past 6 months) IV substance abuse or are homeless. 10. Restrictions on the use of mechanical ventilation (MV), RRT, or vasopressors and inotropes (Note: Restrictions on cardiopulmonary resuscitation (CPR) alone are not an exclusion criterion). 11. Previous administration of recAP. 12. Use of a non-marketed drug within the last month, or concurrent or planned participation in a clinical trial for a non-marketed drug or device. (Note: Concurrent enrollment or participation in observational, non-interventional trials that do not use protocolized treatments or procedures is always permitted. Concurrent enrollment or participation in trials using protocolized treatments or procedures, such as blood sampling, requires prior approval from the TSC.) 13. Current or planned extracorporeal membrane oxygenation (ECMO). 14. The patient received RRT > 24 hours prior to the start of the investigational drug treatment. 15. The patient was no longer receiving vasopressor therapy at the time of randomization. 16. The patient was receiving continuous vasopressor therapy > 72 hours prior to the start of the investigational drug. 17. Estimated glomerular filtration rate (eGFR) > 60 mL / min / 1.73 m² based on the most recent available CR sample at the time of screening. 2 (Note: This is often the sample used to diagnose AKI). eGFR should be calculated using the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) formula. 18. It is impossible to start the investigational drug within the following timeframe: a) When the AKI diagnosis precedes the initiation of vasopressor therapy, 48 hours after the AKI diagnosis, or b) If AKI is diagnosed after the initiation of vasopressor therapy, within 24 hours of the AKI diagnosis. 19. Pregnant or breastfeeding women.

[0170] Investigational drug, dosage, and route of administration The investigational drug (recAP or placebo) will be provided in a glass vial as an infusion concentrate (aqueous buffer solution at pH 7.0). Prior to administration, the investigational drug will be diluted to a final volume of 50 mL with 0.9% sterile sodium chloride for injection (isotonic saline), USP / EP, or equivalent, and administered as an IV infusion using an administration syringe or infusion bag. The intended recAP dose is 1.6 mg (1,000 U) per kg of patient body weight. Patients with a body weight > 120 kg will be administered a fixed dose of 192 mg.

[0171] The investigational drug (recAP or placebo) was administered by qualified staff in the ICU or intermediate care unit as a 1-hour continuous IV infusion on days 1, 2, and 3. The first infusion should be initiated as soon as possible after patient randomization on day 1. On days 2 and 3, investigational drug administration should be initiated 24+ / - 2 hours after the previous investigational drug administration. If a patient is discharged from the ICU or intermediate care unit to a hospital ward before completing the final investigational drug administration on day 3, the investigational drug should be administered in the ward by a qualified person according to instructions provided by the study team.

[0172] The preferred route for administering the investigational drug is through a central catheter; if this is not feasible, a peripheral line is acceptable. The investigational drug will be administered separately from any other concomitant medications using a dedicated lumen of the catheter.

[0173] Randomized and blinded Eligible patients are assigned a unique patient identification number via Interactive Response Technology (IRT), which randomly assigns each patient to either active or placebo according to a randomization schedule generated by a validated computer program. Detailed procedural information is provided in the IRT manual, which is available to all facilities.

[0174] Patients are randomly assigned in a 1:1 ratio to receive either recAP or placebo.

[0175] Randomization for the primary study population and the "moderate to severe" CKD population will be stratified as follows: Pre-AKI reference eGFR ≥ 25 and < 45 mL / min / 1.73m² 2 "Moderate to severe" CKD is defined as such. Yes, No Baseline mSOFA score, i.e., excluding the GCS portion. mSOFA score ≤ 9 mSOFA score > 9 Clinical facilities

[0176] Randomization of the COVID-19 population will be stratified by the following: Baseline mSOFA score, i.e., excluding the GCS portion. mSOFA score ≤ 9 mSOFA score > 9 Clinical facilities

[0177] Related Endpoints The following endpoints were measured: 90-day all-cause mortality; MAKE 90: Death or RRT up to D28 and including D28 or RRT on D90, a ≥25% decrease in estimated glomerular filtration rate (eGFR) at day 90 relative to known or estimated pre-AKI reference level, and / or readmission up to D90. Adverse events.

[0178] analysis Analysis group: Overall efficacy population: All patients who were randomly assigned to the investigational drug and initiated treatment with the investigational drug. This constitutes one overall population consisting of the primary study population, the "moderate to severe" CKD population, and the COVID-19 population. All patients were analyzed according to the treatment they received, regardless of what their planned treatment was. One treated subject with no efficacy data was excluded from the efficacy analysis.

[0179] Overall safety population: All patients who were randomly assigned to the investigational drug and initiated treatment with the investigational drug. One overall population consisting of the primary study population, the "moderate to severe" CKD population, and the COVID-19 population. All patients were analyzed according to the treatment they received, regardless of what their planned treatment was.

[0180] Effectiveness analysis Mortality at 90 days; The absolute and relative frequencies of patients who died before day 90 were analyzed. All available data for patients who were lost to follow-up before day 90 were included in the event proportion analysis.

[0181] Kaplan-Meier estimates of time-versus-mortality up to day 90 (including day 90) have been generated. Differences in proportions within survival rates have been tested using Wald tests for differences in proportions. Patients who dropped out before day 90 were censored.

[0182] Make90 was defined as death, RRT up to and including D28, or RRT at D90, or a ≥25% decrease in estimated glomerular filtration rate (eGFR) at day 90 relative to a known or estimated pre-AKI reference level, or readmission up to D90.

[0183] The absolute and relative frequencies of patients with a MAKE90 event were analyzed. All available data for patients who were lost to follow-up before day 90 were included in the event proportion analysis. The analysis was performed on the overall population and differentiated with respect to a 15 ml increment in the pre-AKI reference eGFR value of the subjects.

[0184] Kaplan-Meier estimates for the first MAKE90 event (including day 90) are generated. Differences in proportions in survival rates are tested using Wald's test for differences in proportions. For evaluation, information from visit records is used even if the visit was not exactly on day 90, regardless of whether the subject received RRT on day 90 or whether a 25% decrease in eGFR was reported on day 90. Patients who dropped out before day 90 were censored.

[0185] Logistic regression of the MAKE90 event was performed to analyze the interaction between pre-AKI reference eGFR and the treatment received. The model included MAKE90 as the outcome variable of interest, as well as treatment, pre-AKI reference eGFR, and pre-AKI reference eGFR due to interaction as factors.

[0186] Safety Analysis: Adverse events, AE incidence, and serious AEs were summarized by investigational drug group. Event proportions were compared using the chi-square test.

[0187] result participants Of the 676 enrolled patients, 21 (3.1%) failed screening, the majority of which were due to study protocol deviations (i.e., failure to meet inclusion / exclusion criteria) reported in 16 patients (2.4%) (see Table 3). 655 and 650 patients, when randomized, were treated with either ilofotase alfa 1.6 mg / kg (recAP 1.6 mg / kg) or placebo.

[0188] The overall efficacy population included 649 patients, 330 subjects in the ilophotase alpha arm, and 319 subjects in the placebo arm (Figure 3). One subject was excluded from the efficacy analysis due to missing efficacy data.

[0189] The overall safety population included 650 patients, 330 subjects in the ilophotase alpha arm, and 320 subjects in the placebo arm (Figure 3).

[0190] Efficacy Outcome 90-day all-cause mortality Regarding mortality up to day 90, including day 90, the proportion of patients who died in the placebo group was 34.8%, compared to 33.9% in the ilophotase alfa group (Table 4). The estimated Kaplan-Meier survival rate at day 90 was 65.54% for the ilophotase alfa group and 63.97% for the placebo group. The estimated mortality rate was 34.46% (1-0.6554) in the ilophotase alfa group and 36.03% (1-0.6397) in the placebo group. No statistically significant difference in the proportion of patients surviving at day 90 could be detected using the Wald test (p=0.3404) (Figure 4).

[0191] MAKE90 Regarding MAKE90, the proportion of patients experiencing a MAKE90 event was 56.7% in the ilophotase alfa group compared to 64.6% in the placebo group (Table 5).

[0192] The Kaplan-Meier survival rate, i.e., the proportion of patients without a MAKE90 event, was estimated to be 42.77% in the ilophotase alfa group and 35.33% in the placebo group. The estimated proportion of patients with a MAKE90 event was 57.23% (1-0.4277) in the ilophotase alfa group and 64.7% (1-0.3533) in the placebo group. A Wald test was used to detect a significant difference favoring ilophotase alfa in the proportion of patients without a MAKE90 event (p=0.031) (Figure 5).

[0193] To further define the potential benefit for patients with lower pre-AKI reference eGFR than those receiving ilophotase alfa, MAKE90 was also investigated using subgroups defined by increments of pre-AKI reference eGFR equal to 15 mL / min / 1.73 m2. The following subgroups were analyzed: <=30, 30~<=45, 45~<=60, 60~<=75, 75~<=90, and >90. The observed number and percentage of patients with a MAKE90 event due to the treatment received are shown in Table 6. From this table, pre-AKI reference eGFR <=90 mL / min / 1.73 m2 2 In all subgroups, a lower percentage of patients receiving ilophotase alfa experienced MAKE90 events compared to patients receiving placebo. However, the difference was >75 to <=90 mL / min / m². 2 It was less in the subgroup.

[0194] This effect was further demonstrated by performing logistic regression on MAKE90. The model included MAKE90 as the outcome variable of interest, as well as treatment, pre-AKI reference eGFR, and pre-AKI reference eGFR due to interaction as factors. The therapeutic effect on MAKE90 was observed to be significantly more favorable to ilophotase alfa (p=0.068). The interactions between pre-AKI reference eGFR (p=0.0174) and treatment and pre-AKI reference eGFR (0.0235) were also significant, suggesting a greater benefit for subjects with lower pre-AKI reference eGFR when comparing ilophotase alfa versus placebo (Table 7). This effect can be visualized by plotting the predicted probability of the Make 90 event against pre-AKI reference eGFR (Figure 6).

[0195] Make90 in a subset of pre-AKI reference eGFR Pre-AKI reference eGFR < 60 Regarding MAKE90, the proportion of patients experiencing a Make 90 event was 59.3% in the ilophotase alfa group compared to 81.3% in the placebo group (Table 8).

[0196] The Kaplan-Meier survival rate, i.e., the proportion of patients without a MAKE90 event, was estimated to be 42.15% in the ilophotase alfa group and 21.03% in the placebo group. The estimated proportion of patients with a MAKE90 event was 57.85% (1-0.4215) in the ilophotase alfa group and 78.97% (1-0.2103) in the placebo group. A Wald test was used to detect a significant difference favoring ilophotase alfa in the proportion of patients without a MAKE90 event (p=0.002) (Figure 7).

[0197] Reference eGFR < 75 before AKI onset Regarding MAKE90, the proportion of patients experiencing a Make 90 event was 56.6% in the ilophotase alfa group compared to 71.3% in the placebo group (Table 9).

[0198] The Kaplan-Meier survival rate, i.e., the proportion of patients without a MAKE90 event, was estimated to be 43.61% in the ilophotase alfa group and 29.71% in the placebo group. The estimated proportion of patients with a MAKE90 event was 56.39% (1-0, 0, 4361) in the ilophotase alfa group and 70.29% (1-0, 2971) in the placebo group. A Wald test was used to detect a significant difference favoring ilophotase alfa in the proportion of patients without a MAKE90 event (p=0.005) (Figure 8).

[0199] Safety Outcomes Adverse events A brief summary of adverse events (AEs) within the safety set for the total population is presented in Table 10. The proportion of patients experiencing AEs (67.9% for ilophotase alfa and 75.0% for placebo) was statistically significant (p=0.0446).

[0200] Conclusion: Compared to patients receiving placebo, a significantly lower percentage of patients receiving ilophotase alfa were estimated to experience a MAKE90 event after initiation of the investigational drug (57.2% for ilophotase alfa and 64.7% for placebo; p-value for ilophotase alfa-placebo difference = 0.031). This effect was most pronounced in patients with pre-existing renal repair.

[0201] Overall, ilophotase was well-tolerated, and no safety concerns were identified. [Table 3] [Table 4] [Table 5] [Table 6] [Table 7] [Table 8] [Table 9] [Table 10]

Claims

1. Alkaline phosphatase (AP) for use in a method of treating acute kidney injury (AKI) in a subject requiring treatment, wherein the method comprises administering an effective amount of AP to the subject, the subject having an acute exacerbation of chronic kidney injury, the chronic kidney injury (CKI) being mild, moderate, or severe.

2. The estimated glomerular filtration rate (eGFR) before the onset of AKI was ≤75 ml / min / 1.73 m². ２ Preferably ≤60 ml / min / 1.73 m ２ AP for use as described in claim 1.

3. eGFR before AKI onset: ≥ 15 ml / min / 1.73 m ２ Preferably ≥ 25 ml / min / 1.73 m ２ AP for use according to claim 1 or 2.

4. AP for use according to any one of claims 1 to 3, wherein administration of AP results in a reduced risk of developing a major adverse renal event (MAKE 90) by day 90, compared to the risk of MAKE 90 in the absence of treatment.

5. AP for use according to claim 4, wherein the MAKE90 includes one or more of the following events: (i) death before or on day 90, (ii) renal replacement therapy before or on day 90, (iii) a ≥25% decrease in eGFR on day 90 compared to eGFR before the onset of AKI, and (iv) readmission before or on day 90.

6. AP for use according to any one of claims 1 to 5, wherein AP is administered in at least one dose of 500 U / kg to 2,000 U / kg.

7. AP for use according to any one of claims 1 to 6, wherein the AP is human AP.

8. AP for use according to any one of claims 1 to 7, wherein the AP is recombinant AP, preferably the recombinant AP is a chimeric AP, and more preferably the chimeric AP has sequence identity of at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% with respect to the amino acid sequence of RecAP (SEQ ID NO: 1).

9. AP for use according to any one of claims 1 to 8, wherein AP is administered in a once-daily or three-times-daily dose.

10. AP for use according to any one of claims 1 to 9, wherein AP is administered intravenously.

11. AP for use according to any one of claims 1 to 10, wherein the AP dose is approximately 1.6 mg / kg of RecAP and / or approximately 1000 U / kg of RecAP.

12. AP for use according to any one of claims 1 to 11, wherein the administration of at least one dose of AP results in preservation of glomerular filtration rate (GFR) or estimated GFR (eGFR) at day 90 in the subject, with respect to pre-AKI GFR or eGFR.

13. AP for use according to any one of claims 1 to 12, wherein AKI is caused by or associated with sepsis.

14. AP for use according to any one of claims 1 to 13, wherein treatment is initiated within 96 hours after sepsis is detected and / or within 48 hours after acute kidney injury (AKI) is detected in the subject.

15. AP for use according to any one of claims 1 to 14, wherein the reduced eGFR due to chronic kidney disease is determined at least 15 days prior, preferably at least 30 days prior, more preferably at least 60 days prior, and most preferably at least 90 days prior to the administration of AP.

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