Recombinant alkaline phosphatase for use in the treatment of acute respiratory distress syndrome - Patent Application 20070122999

Abstract

The present disclosure relates to the use of alkaline phosphatases, particularly improved alkaline phosphatases such as RecAP, for the prevention, treatment, cure, or amelioration of symptoms of acute respiratory distress syndrome (ARDS). The disclosure also relates to methods for maintaining pulmonary function, shortening the duration of mechanical ventilation, and increasing the P / F ratio.

Description

[Technical Field]

[0001] The present invention relates to the medical field. In particular, the present invention relates to the use of alkaline phosphatase, particularly an improved alkaline phosphatase such as RecAP, for the prevention, treatment, cure, or amelioration of symptoms of acute respiratory distress syndrome (ARDS). The present invention also relates to methods for maintaining pulmonary function, shortening the duration of mechanical ventilation, and increasing the P / F ratio. [Background technology]

[0002] Acute respiratory distress syndrome (ARDS) is a respiratory dysfunction characterized by widespread lung inflammation. Symptoms include shortness of breath, rapid breathing, and pale skin (Fan, E; Brodie, D; Slutsky, AS (20 February 2018). "Acute Respiratory Distress Syndrome: Advances in Diagnosis and Treatment". JAMA. 319 (7): 698-710). The overall prognosis for ARDS is poor, with a mortality rate of approximately 40% (Lewis, Sharon R.; Pritchard, Michael W.; Thomas, Carmel M.; Smith, Andrew F. (July 23, 2019). "Pharmacological agents for adults with acute respiratory distress syndrome". The Cochrane Database of Systematic Reviews. 7: CD004477). For those who survive ARDS, reduced quality of life is common (Matthay, MA; Zemans, RL; Zimmerman, GA; Arabi, YM; Beitler, JR; Mercat, A; Herridge, M; Randolph, AG; Calfee, CS (14 March 2019). "Acute respiratory distress syndrome". Nature Reviews. Disease Primers. 5 (1): 18).

[0003] Worldwide, ARDS affects more than 3 million people annually (Fan E et al. (2018)). While the term "adult respiratory distress syndrome" is sometimes used to distinguish ARDS from "infant respiratory distress syndrome" in newborns, international consensus is that "acute respiratory distress syndrome" is the most appropriate term because ARDS can affect people of all ages (Bernard G, Artigas A, Brigham K, Carlet J, Falke K, Hudson L, Lamy M, Legall J, Morris A, Spragg R (1994). "The American-European Consensus Conference on ARDS. Definitions, mechanisms, relevant outcomes, and clinical trial coordination". Am J Respir Crit Care Med. 149 (3 Pt 1): 818-24). Separate diagnostic criteria exist for children and those in resource-poor areas (Matthay MA et al. (2019)).

[0004] The annual incidence of ARDS is typically 13-23 cases per 100,000 in the general population. ARDS is common in people with acute lung injury who are receiving mechanical ventilation (Lewandowski K, Lewandowski M (2006). "Epidemiology of ARDS". Minerva Anestesiol. 72 (6): 473-7). COVID-19 increased ARDS incidence in 2020 (Guo, YR; Cao, QD; Hong, ZS; Tan, YY; Chen, SD; Jin, HJ; Tan, KS; Wang, DY; Yan, Y (13 March 2020). "The origin, transmission and clinical therapies on coronavirus disease 2019 (COVID-19) outbreak - an update on the status". Military Medical Research. 7 (1): 11; Solaimanzadeh, I (20 March 2020). "Acetazolamide, Nifedipine and Phosphodiesterase Inhibitors: Rationale for Their Utilization as Adjunctive Countermeasures in the Treatment of Coronavirus Disease 2019 (COVID-19)". Cureus. 12 (3): e7343).

[0005] Worldwide, in the absence of a viral pandemic such as COVID-19, severe sepsis is the most common cause of ARDS (Goldman, Lee (2011). Goldman's Cecil Medicine (24th ed.). Philadelphia: Elsevier Saunders. p. 635. ISBN 978-1437727883). Other contributing factors include mechanical ventilation, pneumonia, Gilchrist disease, drowning, circulatory shock, aspiration, trauma (especially pulmonary contusion), major surgery, massive blood transfusion (Laar, Alexander PJ; Binnekade, Jan M.; Prins, David; van Stein, Danielle; Hofstra, Jorrit J.; Schultz, Marcus J.; Juffermans, Nicole P. (March 2010). "Risk factors and outcome of transfusion-related acute lung injury in the critically ill: A nested case-control study*". Critical Care Medicine. 38 (3): 771-778), smoke inhalation, drug reaction or overdose, fat embolism after lung transplantation or pulmonary embolectomy, and reperfusion pulmonary edema.

[0006] Following treatment of the underlying cause, such as antibiotics, antivirals, and anti-inflammatory drugs, ARDS is typically treated with mechanical ventilation in the intensive care unit (ICU). Mechanical ventilation is typically performed through a rigid tube inserted through the mouth and secured to the airway (endotracheal intubation) or via tracheotomy if prolonged ventilation (longer than two weeks) is required. The role of noninvasive ventilation is limited to the very early stages of the disease or to prevent worsening respiratory distress in patients with atypical pneumonia, pulmonary contusion, or major surgery who are at risk for developing ARDS. No specific mechanical ventilation mode is known to improve the mortality rate of ARDS, which is between 35% and 50% (Fan E et al. (2018)). Therefore, a treatment for ARDS is needed. Summary of the Invention [Means for solving the problem]

[0007] The present disclosure provides a method for treating acute respiratory distress syndrome (ARDS) in a subject in need thereof, comprising administering to the subject an effective amount of alkaline phosphatase (AP). In a preferred embodiment, the AP is administered in at least one dose of 500 U / kg to 2,000 U / kg.

[0008] The severity of ARDS is usually classified as "mild," "moderate," and "severe." Mild, moderate, and severe ARDS are defined by the following criteria: A decrease in the PaO2 / FiO2 ratio (the PaO2 / FiO2 ratio is the ratio between the arterial oxygen partial pressure (PaO2 in mmHg) and the inspired oxygen partial pressure (FiO2 expressed as a fraction, not a percentage), also called the P / F ratio): >200mmHg and ≤300mmHg (>26.66kPa and ≤40.00kPa): mild ARDS; >100mmHg and ≤200mmHg (>13.33kPa and ≤26.66kPa): moderate ARDS; ≦100mmHg (≦13.33kPa): Severe ARDS; Here, the P / F ratio is measured using a minimum positive end-expiratory pressure (PEEP) of 5 cmH2O.

[0009] In some embodiments, the subject has severe or moderate ARDS as defined above (i.e., a P / F ratio of ≦200 mmHg with a minimum PEEP of 5 cmH2O) prior to treatment with AP. In some embodiments, the subject has severe ARDS (i.e., a P / F ratio of ≦100 mmHg with a minimum PEEP of 5 cmH2O) prior to treatment with AP. In some embodiments, the subject has moderate ARDS (i.e., a P / F ratio of 100-200 mmHg with a minimum PEEP of 5 cmH2O) prior to treatment with AP. In some embodiments, the ARDS is associated with and / or caused by sepsis. In other embodiments, the ARDS is associated with and / or caused by a viral infection, preferably an infection by a coronavirus, more preferably an infection by a severe acute respiratory syndrome (SARS)-associated coronavirus. In some embodiments, the AP is a human AP. In some embodiments, the AP is a recombinant AP. In some embodiments, the recombinant AP is chimeric. In some embodiments, the chimeric AP has 70% or more, 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 100% sequence identity to RecAP (SEQ ID NO: 1).

[0010] In some embodiments, the increased respiratory function comprises an increase in the subject's P / F ratio compared to the subject's P / F ratio in the absence of treatment. In some embodiments, the subject's sepsis is detected within 96 hours prior to the administration of AP. In some embodiments, the sepsis is detected within 72 hours prior to the detection of ARDS.

[0011] In some embodiments, treatment according to the present invention is initiated within 24 hours of detecting sepsis in a subject. In some embodiments, treatment according to the present invention is initiated within 24 hours of detecting ARDS in a subject. In some embodiments, AP is administered once daily. In some embodiments, AP is administered intravenously. In some embodiments, AP is administered in three doses daily. In some embodiments, the AP dose is 0.8 mg / kg or 1.6 mg / kg of RecAP, e.g., clinical-grade RecAP used in the present disclosure. In some embodiments, the AP dose is 500 U / kg or 1000 U / kg of RecAP, e.g., clinical-grade RecAP used in the present disclosure.

[0012] In some embodiments, administration of at least one dose of AP results in a reduction in or cessation of mechanical ventilation in a subject undergoing mechanical ventilation. In some embodiments, administration of at least one dose of AP results in a maintenance or increase in the P / F ratio in a subject. [Brief explanation of the drawings]

[0013] [Figure 1] FIG. 1 shows a table showing proposed RecAP dose groups. [Figure 2] FIG. 1 shows the RecAP amino acid sequence (SEQ ID NO: 1). [Figure 3] FIG. 1 shows the timeline of the STOP-AKI study, including the dosing regimen and time frame for determining AUC1-7, RRT incidence, and mortality and MAKE. [Figure 4] Figure 1 shows a flow chart illustrating patient enrollment, randomization, and follow-up in the clinical trial. Abbreviations: AKI (acute kidney injury), AUC (area under the time-corrected curve), ECC (endogenous creatinine clearance), ICF (informed consent form), ITT (intent to treat), IV (intravenous), MAKE (major adverse renal event), NS (not specified), RecAP (human recombinant alkaline phosphatase having the sequence of SEQ ID NO: 1), RRT (renal replacement therapy), SA-AKI (sepsis-associated acute kidney injury), SCr (serum creatinine). [Figure 5] Figure 1 shows a table displaying demographic and baseline characteristics for all groups in the trial, demonstrating that there were no significant differences between groups in any baseline characteristics. For variables with missing data, summary data are based on adjusted values. Body mass index (BMI) is weight (kg) divided by height (m) squared. Acute Physiology and Chronic Health Evaluation II (APACHE II) scores range from 0 to 71, with higher scores indicating greater disease severity. Scores on the Sequential Organ Failure Assessment (SOFA) scores range from 0 to 24, with higher scores indicating greater functional impairment. Simplified Acute Physiology Score (SAPS) scores range from 0 to 163, with higher scores indicating greater disease severity. Estimated glomerular filtration rate (eGFR) was calculated according to the CKD-EPI (Chronic Kidney Disease Epidemiology Collaboration) equation: 141 * min(Scr / κ,1) * α * max(Scr / κ,1) -1.209 * 0.993 age * 1.018 [for women] * 1.159 [for blacks]. where Scr is serum creatinine (mg / dL), κ is 0.7 for women and 0.9 for men, α is -0.329 for women and -0.411 for men, min is the minimum value of Scr / κ or 1, and max is the maximum value of Scr / κ or 1 (www.kidney.org / content / ckd-epi-creatinine-equation-2009). Acute kidney injury (AKI) stages were stratified according to the definitions of the AKI-Network (www.akinet.org / akinstudies.php). Medians and quartiles, or numbers and percentages, are shown. [Figure 6] Figure 1 shows an AUC vs. dose graph demonstrating a linear increase in RecAP concentrations. Patient exposure in the STOP-AKI clinical trial was slightly higher compared to healthy subjects. Dose linearity and proportionality were observed across the dose range (0.4-1.6 mg / kg). [Figure 7] Table showing primary and secondary endpoints (placebo and 0.6 mg / kg RecAP groups). AUC1-7 indicates time-corrected clearance (mL / min), provided by dividing the area under the ECC curve from days 1 to 7 by 7. Abbreviations: CI (confidence interval), CKD-EPI (Chronic Kidney Disease Epidemiology Collaboration), ECC (intrinsic creatinine clearance), eGFR (estimated glomerular filtration rate), ICU (intensive care unit), IQR (interquartile range), MAKE (major adverse renal event), RRT (renal replacement therapy), SOFA (Sequential Organ Failure Assessment). § RRT incidence indicates the proportion of patients who required RRT after randomization. || Last observation advanced. * MAKE28: received RRT before or on day 28, or died before or on day 28. Proportion who met at least one of the criteria. †MAKE60: eGFR at 60 days estimated by the CKD-EPI formula based on serum creatinine before or at 60 days or required chronic RRT <60 mL / min, or death before or at 60 days. Proportion of patients who met at least one of the criteria. ‡MAKE90: eGFR at 90 days estimated by the CKD-EPI formula based on serum creatinine before or at 90 days of RRT or required chronic RRT <60 mL / min, hospitalization due to a new AKI episode before or at 90 days, or death before or at 90 days. Proportion of patients who met at least one of the criteria. §§ Represents absolute difference (for continuous variables). †† Represents odds ratio (for categorical variables). ‡‡ Represents hazard ratio (for events). [Figure 8] FIG. 1 shows the primary and secondary endpoints for all treatment groups (RecAP doses of 0.4 mg / kg, 0.8 mg / kg, or 1.6 mg / kg or placebo). [Figure 9A] Quality of life (QoL) assessment measured by EuroQoL (EQ5D) score in the ICU (left panel) and at 90 days (right panel). [Figure 9B] Median pulmonary function parameters (P / F ratio, tidal volume, and PEEP) are shown for the 1.6 mg / kg RecAP dose group and the placebo group. Values ​​are displayed below the graph. [Figure 9C] 1 shows median liver function parameters (alanine aminotransferase activity and aspartate aminotransferase activity) for the 1.6 mg / kg RecAP dose group and the placebo group. Values ​​are displayed below the graph. [Figure 10] FIG. 1 shows the results of post-hoc multivariate analysis. [Figure 11] FIG. 1 shows treatment-emergent adverse events observed during the STOP-AKI trial. [Figure 12] Figure 1 shows P / F ratio values ​​in STOP-AKI for subjects with measurable screening or day 1 values ​​who were randomized to either placebo or high-dose recAP by day 29 (including day 29). Numbers indicate the number of patients at a particular time point per treatment arm. Mean values ​​are displayed with bootstrapped 95% confidence intervals (CL). A small time stagger was used for each treatment group to increase the discriminatory power of the CL. [Figure 13] Figure 1 shows the fold change in P / F ratio values ​​in STOP-AKI in subjects with measurable screening or day 1 values ​​who were randomized to either placebo or high-dose recAP by day 29 (including day 29). Numbers indicate the number of patients at a particular time point per treatment arm. Mean values ​​are displayed with bootstrapped 95% confidence intervals. A slight time stagger was used for each treatment group to increase the discriminatory power of CL. [Figure 14]Figure 1 shows the fold change in P / F ratio values ​​in STOP-AKI in subjects with measurable screening or day 1 values ​​who were randomized to either placebo or high-dose recAP by day 29 (including day 29). Numbers indicate the number of patients at a particular time point per treatment arm. Mean values ​​are displayed with bootstrapped 95% confidence intervals. A slight time stagger was used for each treatment group to improve discrimination of CL. Data for MeanBase values ​​are displayed in separate panels according to grouped ARDS criteria. [Figure 15] Figure 1 shows survival rates for patients in STOP-AKI randomized to either placebo or high-dose recAP through day 29 (including day 29). The proportion of patients who were alive is shown for the placebo or high-dose AP groups in patients with severe-to-moderate ARDS at baseline compared to day 0. On day 0, there were 40 patients in the placebo group and 42 in the high-dose recAP group. DETAILED DESCRIPTION OF THE INVENTION

[0014] It is an object of the present disclosure to provide a method for maintaining or improving respiratory function in a subject, particularly when the subject is suffering from or at risk of acute respiratory distress syndrome (ARDS). Another object of the present disclosure is to prevent or shorten the duration of mechanical ventilation, maintain or increase the P / F ratio in a subject, and / or increase survival rate.

[0015] Therefore, the present disclosure relates to the use of alkaline phosphatase (AP), such as RecAP, to maintain or improve respiratory function in subjects with ARDS or at risk of ARDS. The present disclosure provides a method for treating a subject with severe to moderate ARDS, comprising, for example, administering AP, such as RecAP, to the subject. In some embodiments, each AP dose administered to the subject contains 500 U / kg or more (0.8 mg / kg for the clinical grade RecAP used). In some specific embodiments, each AP dose is 1,000 U / g or more (1.6 mg / kg for the clinical grade RecAP used). In some embodiments, the ARDS is sepsis-related ARDS. In other aspects, the ARDS is caused or exacerbated by a viral infection, preferably a coronavirus or orthomyxovirus infection, more preferably a severe acute respiratory syndrome (SARS)-related coronavirus infection (e.g., severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the coronavirus strain that causes coronavirus disease 2019 (COVID-19), the respiratory disease responsible for the COVID-19 pandemic of 2020). In the case of an orthomyxovirus infection, the infection is preferably caused by an influenza virus, such as influenza A, B, C, or D, more preferably influenza A or B.

[0016] RecAP is a chimeric AP that combines the properties of two human isozymes, intestinal AP and placental AP (Kiffer-Moreira et al. PLoS One 2014;9:e89374). Replacing the crown domain of intestinal AP (the most biologically active isozyme of the two) with the crown domain of placental AP (which has the longest half-life) generates a highly stable and biologically active enzyme (Kiffer-Moreira et al. PLoS One 2014;9:e89374). See, e.g., U.S. Patent Application Publication Nos. 20170009216 and 20160250299, the disclosures of which are incorporated herein by reference in their entireties.

[0017] In some embodiments, when ARDS is observed and the ARDS is determined to be moderate or severe, an AP (e.g., RecAP) is administered to the subject. Thus, in some embodiments of the present disclosure, the methods disclosed herein include measuring the severity of ARDS before treatment with AP, for example, by determining the P / F ratio, to determine whether the subject is suffering from moderate ARDS or severe ARDS.

[0018] Diagnostic criteria for ARDS have changed over time as our understanding of its pathophysiology has improved. The international consensus criteria for ARDS were updated in 2012 and are known as the "2012 Berlin definition" (Ranieri VM, Rubenfeld GD, Thompson BT, Ferguson ND, Caldwell E, Fan E, Camporota L, Slutsky AS (Jun 2012). "Acute respiratory distress syndrome: the Berlin Definition. ARDS Definition Task Force". JAMA. 307 (23): 2526-33; Ferguson ND, Fan E, Camporota L, Antonelli M, Anzueto A, Beale R, Brochard L, Brower R, Esteban A, et al. (Oct 2012). "The Berlin definition of ARDS: an expanded rationale, justification, and supplementary material". Intensive Care Med. 38 (10): 1573-82).In addition to broadening the diagnostic threshold overall, other notable changes from the previous 1994 consensus criteria (Bernard G, Artigas A, Brigham K, Carlet J, Falke K, Hudson L, Lamy M, Legall J, Morris A, Spragg R (1994). "The American-European Consensus Conference on ARDS. Definitions, mechanisms, relevant outcomes, and clinical trial coordination". Am J Respir Crit Care Med. 149 (3 Pt 1): 818-24) include refraining from using the term "acute lung injury" and defining grades of ARDS severity according to the degree of reduction in oxygen content in the blood.

[0019] According to the 2012 Berlin definition, adult ARDS is characterized by: Acute-onset lung injury with progression of respiratory symptoms within one week of an apparent clinical attack Bilateral opacities on chest imaging (chest radiograph or CT) that are not explained by other pulmonary pathology (e.g., pleural effusion, lobar / lung collapse, or nodules) Respiratory failure not explained by heart failure or volume overload Decrease in PaO2 / FiO2 ratio (a decrease in PaO2 / FiO2 ratio indicates a decrease in arterial oxygen delivery from available inspired gas): Mild ARDS: >200mmHg and ≤300mmHg (>26.66kPa and ≤40.00kPa); Moderate ARDS: >100mmHg and ≤200mmHg (>13.33kPa and ≤26.66kPa); ·Severe ARDS: ≦100mmHg (≦13.33kPa). The Berlin definition requires a minimum positive end-expiratory pressure (PEEP) of 5 cmH2O to account for the PaO2 / FiO2 ratio. This level of PEEP may be provided noninvasively using continuous positive airway pressure (CPAP) to diagnose mild ARDS.

[0020] In this regard, it should be noted that, as used herein, ARDS and acute lung injury (ALI) can be used interchangeably within the boundaries of the above definitions, as known in the art (Hernu, R., Wallet, F., Thiolliere, F. et al. An attempt to validate the modification of the American-European consensus definition of acute lung injury / acute respiratory distress syndrome by the Berlin definition in a university hospital. Intensive Care Med 39, 2161-2170 (2013)), i.e., using the P / F boundary and further criteria of the 2012 Berlin definition to distinguish between mild, moderate, and severe disease.

[0021] In some embodiments of the present disclosure, ARDS is classified as severe if the baseline P / F ratio is ≦100 mmHg. In some embodiments, ARDS is classified as moderate if the baseline P / F ratio is >100 mmHg and ≦200 mmHg. In other embodiments, other diagnostic measurements can be used to determine the severity of ARDS. In some embodiments of the present disclosure, if ARDS is determined to be mild or nonexistent (e.g., if the baseline P / F ratio is >200 mmHg), AP is not administered to the subject.

[0022] In some embodiments, the AP, e.g., RecAP, is administered prophylactically. In some embodiments, the methods disclosed herein include administering at least one dose of AP, e.g., RecAP, to a subject, each dose being at least 500 U / kg (0.8 mg / kg for the clinical-grade RecAP used) or 1000 U / kg (1.6 mg / kg for the clinical-grade RecAP used). In some specific embodiments, the dosing regimen includes administering 0.5 mg / kg to 2 mg / kg of AP, e.g., RecAP, by intravenous infusion in a daily dose for at least three days. In some specific embodiments, the dosing regimen includes administering 0.5 mg / kg to 2 mg / kg of AP, e.g., RecAP, by intravenous infusion in a daily dose for at least three days.

[0023] In order that this disclosure may be more readily understood, certain terms are first defined. Additional definitions are set forth throughout the Detailed Description.

[0024] I. Definition As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. The terms "a" (or "an"), as well as "one or more" and "at least one," can be used interchangeably herein.

[0025] Furthermore, as used herein, "and / or" should be considered as each specific disclosure of one or the other of the two specified features or components. Thus, the term "and / or" used herein 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" used in phrases such as "A, B, and / or C" is intended to encompass each of the following aspects: 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).

[0026] Whenever an embodiment is described herein with the word "comprising," analogous embodiments described in terms of "consisting of" and / or "consisting essentially of" are also provided.

[0027] The term "about" as used throughout this specification and claims in connection with numerical values ​​is familiar to those skilled in the art and indicates an acceptable range of accuracy. Generally, such an interval of accuracy is ±15%.

[0028] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains.

[0029] Units, prefixes, and symbols are shown in the form accepted by the International System of Units (SI).

[0030] Numerical ranges are inclusive of the numbers defining the range. When a range of values ​​is described, it is understood that each intervening integer and fraction between the stated upper and lower limits of that range is also specifically disclosed, along with each subrange between such values. The upper and lower limits of any range are independently included or excluded from the range, and ranges not including either or both limits are also encompassed within the invention. If the lower limit is preceded by ">", the lower limit of the range is excluded from the range. If the upper limit is preceded by "<", the upper limit of the range is excluded from the range. If the lower limit is preceded by "≥", the lower limit of the range is included. If the upper limit is preceded by "≤", the upper limit of the range is included. When values ​​are explicitly recited, it is understood that values ​​approximately the same quantity or amount as the recited value 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 is within the scope of the invention. Conversely, when different elements or groups of elements are individually disclosed, combinations of those elements are also disclosed. Where any element of the invention is disclosed as having multiple alternatives, examples of the invention in which each alternative is excluded alone or in any combination with the other alternatives are also disclosed herein, and more than one element of the invention may have such an exclusion, and all combinations of elements with such exclusions are disclosed herein.

[0031] As used herein, the terms "treat," "treatment," or "treatment of" mean (i) reducing the likelihood or risk of a disease or disorder, such as ARDS; (ii) reducing the occurrence of a disease or disorder, such as ARDS; (iii) reducing the severity (e.g., ameliorating symptoms) of a disease or disorder, such as ARDS; or (iv) a combination thereof.

[0032] For example, treating can refer to the ability of a treatment, when administered to a subject, to prevent or reduce the risk of lung damage caused by or associated with ARDS and / or cure or alleviate the symptoms, signs, or causes of lung damage, e.g., ARDS. Treating can also mean alleviating or reducing at least one clinical symptom and / or inhibiting or delaying the progression of symptoms and / or preventing or delaying the onset of a disease or condition when compared to an untreated (or placebo-treated) group. Thus, the terms "treat," "treating," or "treatment of" (or grammatically equivalent terms) refer to a therapeutic treatment regimen for ARDS that may (further) prevent lung damage caused by or associated with ARDS. In the case of respiratory diseases, a subject preferably experiences improved or less decline in respiratory function after being treated with AP according to the methods disclosed herein compared to a subject not treated with AP.

[0033] As used herein, the term "maintain" includes preventing, slowing, halting, and / or at least partially reversing the decline in lung function. The term "increase" is not necessarily limited to increasing respiratory function to a value equal to or greater than the value before treatment was administered. This includes partial restoration of respiratory function.

[0034] As used herein, the term "subject" or "patient" refers to any subject, particularly a mammalian subject, for whom treatment or prognosis of lung injury (e.g., ARDS) is desired. As used herein, the term "subject" or "patient" includes any human or non-human animal. As used herein, phrases such as "patient with ARDS" or "patient with sepsis" include subjects, such as mammalian subjects, who would benefit from the administration of treatment with an AP, as disclosed herein.

[0035] In some embodiments of the present disclosure, the subject is a naive subject. A naive subject is a subject that has not been administered a treatment, for example, a therapeutic agent. In some embodiments, a naive subject has not been treated with a therapeutic agent before being diagnosed with lung injury, such as ARDS, or a disease or condition that may lead to lung injury (e.g., sepsis).

[0036] In another aspect, the subject has received a treatment and / or one or more doses of a therapeutic agent before being diagnosed with lung injury (e.g., ARDS) or a disease or condition that may lead to lung injury and / or ARDS (e.g., sepsis).

[0037] In some embodiments, a subject may be administered at least one therapeutically effective dose of AP (e.g., RecAP) if the subject's baseline P / F ratio is below a predetermined P / F ratio threshold level or if the baseline P / F ratio is within a predetermined range.

[0038] As used herein, the terms "therapeutic agent" and "drug" also refer to any therapeutically active substance that is administered to a subject having a disease or disorder, such as lung injury (e.g., ARDS), or a disease or condition that may lead to lung injury (e.g., sepsis), to produce a desired, usually beneficial, effect. A therapeutic agent may also be a prodrug that metabolizes into a desired therapeutically active substance when administered to a subject. In some embodiments, a therapeutic agent is a prophylactic agent. A therapeutic agent may also be pharmaceutically formulated. A therapeutic agent may also include radioisotopes or drugs that are activated by other forms of energy, such as light or ultrasound energy, or by other circulating molecules that can be administered systemically.

[0039] In some aspects of the present disclosure, a therapeutic agent for the treatment, prevention, or amelioration of symptoms of lung injury (e.g., ARDS) or a disease or condition that may lead to lung injury (e.g., sepsis) can include an AP (e.g., RecAP) alone or in combination with one or more standard therapeutic agents typically used to treat symptoms of lung injury (e.g., ARDS) or a disease or condition that may lead to lung injury (e.g., sepsis).

[0040] As used herein, a "therapeutically effective" amount is an amount of a therapeutic agent that provides some improvement or benefit to a subject having a disease or disorder, such as lung injury (e.g., ARDS), or a disease or condition that may lead to lung injury (e.g., sepsis). Thus, a "therapeutically effective" amount is an amount that provides some relief, alleviation, and / or reduction of at least one clinical symptom of a disease or disorder, such as lung injury (e.g., ARDS), or a disease or condition that may lead to lung injury (e.g., sepsis).

[0041] Clinical symptoms associated with lung injury (e.g., ARDS) or diseases or conditions that may lead to lung injury (e.g., sepsis) that can be treated by the compositions and methods, such as particular dosing regimens, of the present disclosure are well known to those of skill in the art. Furthermore, those skilled in the art will understand that the therapeutic effect need not be complete or curative, as long as some benefit is provided to the subject. In some embodiments, the term "therapeutically effective" refers to an amount of a therapeutic agent that can alter biomarker levels, such as the P / F ratio, in a patient in need thereof.

[0042] As used herein, an "amount sufficient" or "an amount sufficient to" achieve a particular result in a patient with a disease or disorder, such as lung injury (e.g., ARDS), or a disease or condition that may lead to lung injury (e.g., sepsis), refers to an amount of a therapeutic agent (e.g., an AP such as RecAP) effective to produce a desired effect, which may be a therapeutic effect (i.e., by administration of a therapeutically effective amount). In some embodiments, such a particular result is improved lung function.

[0043] As used herein, the term "healthcare provider" means an individual or institution that directly interacts with and administers to a living subject (e.g., a human patient). Non-limiting examples of healthcare providers include doctors, nurses, technicians, therapists, pharmacists, counselors, alternative medicine practitioners, medical facilities, clinics, hospitals, emergency rooms, medical offices, urgent care centers, alternative medicine clinics / facilities, and any other organization that provides general and / or specialized treatment, evaluation, maintenance, therapy, medication, and / or advice related to all or any portion of a patient's health condition, including, but not limited to, general medical, specialty medical, surgical, and / or other types of treatment, evaluation, maintenance, therapy, medication, and / or advice.

[0044] As used herein, the term "clinical laboratory" refers to a facility for testing or processing materials from living subjects (e.g., humans). Non-limiting examples of processing include biological, biochemical, serological, chemical, immunohematological, hematological, biophysical, cytological, pathological, genetic, or other testing of materials from the human body for the purpose of providing information for, e.g., the diagnosis, prevention, treatment, or evaluation of the health status of any disease or disorder in a living subject (e.g., human). These tests can also include procedures for taking or otherwise obtaining samples, preparing, determining, measuring, or otherwise describing the presence or absence of various substances in or from a living subject (e.g., human).

[0045] As used herein, the term "healthcare benefits provider" encompasses any individual party, organization, or group involved in providing, offering, proposing, paying for, or providing access to a patient for one or more healthcare benefits, benefit plans, health insurance, and / or healthcare account programs, in whole or in part.

[0046] In some aspects, a healthcare provider can administer, or instruct another healthcare provider to administer, a treatment to prevent, treat, or ameliorate the symptoms of a disease or disorder, such as lung injury (e.g., ARDS), or a disease or condition that can lead to lung injury (e.g., sepsis). A healthcare provider may take, or instruct another healthcare provider or patient to take, the following actions: obtain a sample, process a sample, submit a sample, receive a sample, transfer a sample, analyze or measure a sample, quantify a sample, provide results after analyzing / measuring / quantifying a sample, receive results after analyzing / measuring / quantifying a sample, compare / score results after analyzing / measuring / quantifying one or more samples, provide a comparison / score from one or more samples, obtain a comparison / score from one or more samples, administer a treatment (e.g., an AP such as RecAP), begin administering a treatment, stop administering a treatment, continue administering a treatment, temporarily suspend administering a treatment, increase the amount of a therapeutic agent administered, decrease the amount of a therapeutic agent administered, continue administering an amount of a therapeutic agent, increase the frequency of administration of a therapeutic agent, decrease the frequency of administration of a therapeutic agent, maintain the same frequency of administration of a therapeutic agent, replace a treatment or therapeutic agent with at least another treatment or therapeutic agent, combine a treatment or therapeutic agent with at least another treatment or additional therapeutic agent.

[0047] In some aspects, the health care benefit provider may allow or deny, for example, sample collection, sample processing, sample submission, sample receipt, sample movement, sample analysis or measurement, sample quantification, providing results obtained after analyzing / measuring / quantifying samples, transferring results obtained after analyzing / measuring / quantifying samples, comparing / scoring results obtained after analyzing / measuring / quantifying one or more samples, transferring comparisons / scores from one or more samples, administering / administering a treatment or therapeutic agent, initiating administration / administration of a treatment or therapeutic agent, stopping administration / administration of a treatment or therapeutic agent, continuing administration / administration of a treatment or therapeutic agent, temporarily interrupting administration / administration of a treatment or therapeutic agent, increasing the dosage of a therapeutic agent, decreasing the dosage of a therapeutic agent, continuing administration of an amount of a therapeutic agent, increasing the number of doses of a therapeutic agent, decreasing the number of doses of a therapeutic agent, maintaining the same number of doses of a therapeutic agent, substituting at least one other treatment or therapeutic agent, or combining a treatment or therapeutic agent with at least another treatment or additional therapeutic agent.

[0048] Additionally, health care providers may, for example, authorize or deny prescription of a treatment, authorize or deny insurance coverage for a treatment, authorize or deny reimbursement for a treatment, determine eligibility for a treatment, and the like.

[0049] In some embodiments, a clinical laboratory may, for example, collect or obtain samples, process samples, submit samples, receive samples, move samples, analyze or measure samples, quantify samples, provide results obtained after analyzing / measuring / quantifying samples, receive results obtained after analyzing / measuring / quantifying samples, compare / score results obtained after analyzing / measuring / quantifying one or more samples, provide a comparison / score from one or more samples, obtain a comparison / score from one or more samples, or perform other related actions.

[0050] II. Treatment of ARDS with AP In certain aspects, the present disclosure relates to methods for maintaining or improving lung function in a subject population determined to respond particularly well to treatment with AP, particularly patients with ARDS.

[0051] A recent analysis of data from the STOP-AKI study, extensively described by Pickkers et al. (Pickkers P, Mehta RL, Murray PT, et al. Effect of Human Recombinant Alkaline Phosphatase on 7-Day Creatinine Clearance in Patients With Sepsis-Associated Acute Kidney Injury: A Randomized Clinical Trial. JAMA. 2018;320(19):1998-2009), confirmed a statistically significant correlation between thresholds corresponding to ARDS severity and improvement in lung function (evidenced by improved P / F ratios versus placebo). Stratifying patients in the STOP-AKI clinical trial according to markers of lung function, particularly the P / F ratio, surprisingly identified specific effects of AP administration for specific subgroups. The parameter or parameters (e.g., a set of thresholds such as a P / F ratio threshold) defining each of these subgroups can be used, for example, to individualize AP therapy to specific subgroups, patients selected for treatment, make decisions about AP treatment (e.g., modify dosing or dosing schedule), or assess the likelihood of a positive outcome.

[0052] Thus, in some embodiments, the methods disclosed herein relate to administering AP (e.g., RecAP) to a subject determined to have either moderate or severe ARDS, including administering AP such as RecAP to the subject. Methods for classifying a subject's ARDS severity as moderate or severe are well known to those skilled in the art. As described above, the 2012 Berlin definition classifies a P / F ratio of >100 mmHg and ≦200 mmHg as moderate ARDS, and a P / F ratio of ≦100 mmHg (≦13.33 kPa) as severe ARDS. Therefore, a person with moderate or severe ARDS has a P / F ratio ≦200 mmHg. The Berlin definition further requires a minimum positive end-expiratory pressure (PEEP) of 5 cmH2O to account for the P / F ratio. Thus, in a preferred embodiment, the present invention provides alkaline phosphatase (AP) for use in a method of treating acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the subject has moderate or severe ARDS with a P / F ratio of ≦200 mmHg when the P / F ratio is measured using a minimum positive end-expiratory pressure (PEEP) of 5 cmH2O. Administration of AP is particularly effective when administered at a dose of 500 U / kg or greater to subjects with moderate to severe ARDS. AP is particularly effective when administered at a dose of 1,000 U / kg or greater.

[0053] In some embodiments, the AP (e.g., RecAP) is administered at a dose of about 500 U / kg or more, about 600 U / kg or more, about 700 U / kg or more, about 800 U / kg or more, about 900 U / kg or more, about 1000 U / kg or more, about 1100 U / kg or more, about 1200 U / kg or more, about 1300 U / kg or more, about 1400 U / kg or more, about 1500 U / kg or more, about 1600 U / kg or more, about 1700 U / kg or more, about 1800 U / kg or more, about 1900 U / kg or more, or about 2000 U / kg or more per dose. In some embodiments, the AP (e.g., RecAP) is administered at a dose of more than 2000 U / kg per dose. In some embodiments, the AP (e.g., RecAP) is administered at a dose of less than 500 U / kg per dose.

[0054] In some embodiments, AP (e.g., RecAP) is administered at a dose of about 500 U / kg to about 1500 U / kg, about 600 U / kg to about 1400 U / kg, about 700 U / kg to about 1300 U / kg, about 800 U / kg to about 1200 U / kg, or about 900 U / kg to about 1100 U / kg. In some specific embodiments, AP is administered at a dose of about 1000 U / kg.

[0055] In some embodiments, the AP is a human AP. In some embodiments, the AP is a recombinant AP. In some embodiments, the AP is a chimeric AP. In a particular embodiment, the chimeric AP is RecAP (SEQ ID NO: 1). In some embodiments, an AP disclosed herein has about 70% or more, about 75% or more, about 80% or more, about 85% or more, about 90% or more, about 95% or more, about 96% or more, about 97% or more, about 98% or more, or about 99% or more sequence identity to the amino acid sequence of SEQ ID NO: 1. In some embodiments, the AP is a functional fragment (i.e., a fragment of an AP, e.g., an AP that maintains about 10% or more, about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, 70% or more, about 80% or more, or about 90% or more of the AP activity of the corresponding full-length AP). In some embodiments, the AP is a variant or derivative of an AP disclosed herein. Other APs that may be used as disclosed herein are discussed in detail below.

[0056] In some embodiments, the AP is RecAP (e.g., clinical grade RecAP used in the present disclosure) and is administered at a dose of about 0.1 mg / kg or more, about 0.2 mg / kg or more, about 0.3 mg / kg or more, about 0.4 mg / kg or more, about 0.5 mg / kg or more, about 0.6 mg / kg or more, about 0.7 mg / kg or more, about 0.8 mg / kg or more, about 0.9 mg / kg or more, about 1 mg / kg or more, about 1.1 mg / kg or more, about 1.3 mg / kg or more, about 1.4 mg / kg or more, about 1.5 mg / kg or more, about 1.6 mg / kg or more, about 1.7 mg / kg or more, about 1.8 mg / kg or more, about 1.9 mg / kg or more, about 2 mg / kg or more, about 2.1 mg / kg or more, about 2.2 mg / kg or more, about 2.3 mg / kg or more, or about 2.4 mg / kg or more per dose. In some embodiments, the AP is administered at a dose of greater than 2.4 mg / kg per dose. In some embodiments, the AP is RecAP (e.g., clinical-grade RecAP used in the present disclosure) and is administered at a dose of about 100 U / kg or more, about 200 U / kg or more, about 300 U / kg or more, about 400 U / kg or more, about 500 U / kg or more, about 600 U / kg or more, about 700 U / kg or more, about 800 U / kg or more, about 900 U / kg or more, about 1000 U / kg or more, about 1100 U / kg or more, about 1200 U / kg or more, about 1300 U / kg or more, about 1400 U / kg or more, about 1500 U / kg or more, about 1600 U / kg or more, about 1700 U / kg or more, about 1800 U / kg or more, about 1900 U / kg or more, or about 2000 U / kg or more. AP is RecAP and is administered at a dose of less than 100 U / kg. AP is RecAP and is administered at a dose of more than 2000 U / kg.

[0057] In some embodiments, the AP is RecAP (e.g., clinical-grade RecAP used in the present disclosure) and is administered at a dose of about 0.8 mg / kg to about 2.4 mg / kg, about 0.9 mg / kg to about 2.3 mg / kg, about 1 mg / kg to about 2.2 mg / kg, about 1.1 mg / kg to about 2.1 mg / kg, about 1.2 mg / kg to about 2 mg / kg, about 1.3 mg / kg to about 1.9 mg / kg, about 1.4 mg / kg to about 1.8 mg / kg, or about 1.5 mg / kg to about 1.7 mg / kg. In some specific embodiments, the AP is administered at a dose of about 1.6 mg / kg.

[0058] In some embodiments, the AP is RecAP (e.g., clinical-grade RecAP used in the present disclosure) and has a specific activity of about 100 U / mg or more, about 200 U / mg or more, about 300 U / mg or more, about 400 U / mg or more, about 500 U / mg or more, about 600 U / mg or more, about 700 U / mg or more, about 800 U / mg or more, about 900 U / mg or more, about 1000 U / mg or more, about 1100 U / mg or more, about 1200 U / mg or more, about 1300 U / mg or more, about 1400 U / mg or more, about 1500 U / mg or more, about 1600 U / mg or more, about 1700 U / mg or more, about 1800 U / mg or more, about 1900 U / mg or more, or about 2000 U / mg or more.

[0059] In some embodiments, the AP is RecAP (e.g., clinical-grade RecAP used in the present disclosure) and has a specific activity of about 1000 U / mg. In some embodiments, the AP is RecAP and has a specific activity of about 600 U / mg to about 700 U / mg, about 500 U / mg to about 800 U / mg, about 400 U / mg to about 900 U / mg, about 300 U / mg to about 1000 U / mg, about 200 U / mg to about 1100 U / mg, or 100 U / mg to about 1200 U / mg. In some embodiments, the AP is RecAP and has a specific activity of less than 100 U / mg. In some embodiments, the AP is RecAP and has a specific activity of greater than 1200 U / mg.

[0060] In some embodiments, only one dose of AP (e.g., RecAP) is administered per treatment (e.g., once daily for 1-7 days). In other embodiments, two or more doses of AP are administered. In some embodiments, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 doses of AP are administered (e.g., at least twice daily for 1-7 days).

[0061] In some embodiments, the AP dose is administered daily, hi other embodiments, the AP dose is administered every 2, 3, 4, 5, 6, or 7 days.

[0062] In some embodiments, a single dose is administered daily, hi some embodiments, two, three, or more doses are administered daily.

[0063] In some embodiments, the treatment with AP is for less than about 4 days, hi some embodiments, the treatment with AP is for less than 3 days, less than 2 days, or less than 1 day.

[0064] In certain embodiments, the AP is administered as a daily dose of about 1000 U / kg administered for three consecutive days. In some specific embodiments, when the AP is RecAP, the AP is administered as a daily dose of 1.6 mg / kg administered for three consecutive days. In some embodiments, the dose of each AP, e.g., RecAP (e.g., clinical-grade RecAP used in the present disclosure), is about 0.10 mg / kg to about 3 mg / kg, about 0.20 mg / kg to about 2.9 mg / kg, about 0.3 mg / kg to about 2.8 mg / kg, about 0.4 mg / kg to about 2.7 mg / kg, about 0.5 mg / kg to about 2.6 mg / kg, about 0.6 mg / kg to about 2. 0.5mg / kg, about 0.7mg / kg to about 2.4mg / kg, about 0.8mg / kg to about 2.3mg / kg, about 0.9mg / kg to about 2.2mg / kg, about 1mg to about 2.1mg / kg, about 1.1mg / kg to about 2mg / kg, about 1.2mg / kg to about 1.9mg / kg, about 1.3mg / kg to about 1.8mg / kg, or about 1.4mg / kg to about 1.7mg / kg. In some embodiments, the dose of each AP, e.g., RecAP, is about 0.1 mg or more AP / kg, about 0.2 mg or more AP / kg, about 0.3 mg or more AP / kg, about 0.4 mg or more AP / kg, about 0.5 mg or more AP / kg, about 0.6 mg or more AP / kg, about 0.7 mg or more AP / kg, about 0.8 mg or more AP / kg, about 0.9 mg or more AP / kg, about 1 mg or more AP / kg, about 1.1 mg or more AP / kg, about 1.2 mg or more AP / kg, about 1.3 mg or more AP / kg, about 1.4 mg or more AP / kg, about 1.5 mg or more AP / kg. P / kg, about 1.6 mg or more AP / kg, about 1.7 mg or more AP / kg, about 1.8 mg or more AP / kg, about 1.9 mg or more AP / kg, about 2 mg or more AP / kg, about 2.1 mg or more AP / kg, about 2.2 mg or more AP / kg, about 2.3 mg or more AP / kg, about 2.4 mg or more AP / kg, about 2.5 mg or more AP / kg, about 2.6 mg or more AP / kg, about 2.7 mg or more AP / kg, about 2.8 mg or more AP / kg, about 2.9 mg or more AP / kg, or about 3 mg or more AP / kg.

[0065] The AP can be administered via different routes, for example, intravenously, intrarectally, intrabronchially, or orally. In some specific embodiments, the AP is administered intravenously, for example, via intravenous infusion. In some embodiments, the AP is administered intravenously by continuous infusion.

[0066] Although short-term maintenance of lung function can have immediate life-saving results, it is preferable that the effects of AP on lung function be long-term.

[0067] The P / F ratio can be determined by methods known in the art. Arterial pO2 measured by arterial blood gas (ABG) is the preferred method for calculating the P / F ratio. However, if pO2 is unknown because ABGs are not available, pO2 can be approximated using SpO2 measured by pulse oximetry, as shown in the table below. It should be noted that when SpO2 is between 98% and 100%, estimation of pO2 from SpO2 becomes unreliable. [Table 1] Example: Assume a patient on 40% oxygen has a pulse oximetry SpO2 of 95%. Referring to the table above, an SpO2 of 95% corresponds to a pO2 of 80 mmHg. P / F ratio = 80 ÷ 0.40 = 200. The patient may be stabilized on 40% oxygen but still have acute respiratory failure. If the patient is discontinued on room air, the pO2 will be only 40 mmHg (well below the 60 mmHg cutoff for acute respiratory failure on room air).

[0068] Supplemental oxygen can be administered by either a mask or a nasal cannula ("NC"). Venturi masks (bench masks) deliver a controlled flow of oxygen at a specific fixed concentration (FIO2) of 24%, 28%, 31%, 35%, 40%, or 50%. Non-rebreather ("NRB") masks are designed to deliver approximately 100% oxygen. Providing supplemental oxygen at 40% or more means the physician is treating acute respiratory failure, since only a patient with acute respiratory failure would require that amount of oxygen. Nasal cannulas deliver oxygen at an adjustable flow rate in liters of oxygen per minute (L / min or "LPM"). The actual FIO2 (percent oxygen) delivered by a nasal cannula is somewhat more variable and less reliable than when using a mask, but it can be estimated as shown in the table below. The FIO2 obtained from the nasal cannula flow rate can be used to calculate the P / F ratio. [Table 2] Example: A patient has a pO2 of 85 mmHg on ABG receiving 5 L / min of oxygen. 5 L / min is 40% oxygen (an FIO2 of 0.40), so the P / F ratio = 85 ÷ 0.40 = 212.5.

[0069] In some aspects, administration of an AP disclosed herein to a subject can improve a number of respiratory function-related parameters in the subject, such as P / F ratio, arterial oxygen saturation (pO2), pulse oxygen saturation (SpO2), oxygenation index, and / or respiratory index.

[0070] In some embodiments, administration of AP (e.g., RecAP) to a subject results in an increase in the P / F ratio compared to the P / F ratio of an untreated subject. In some embodiments, the increase in the P / F ratio is 5 mmHg or more, 10 mmHg or more, 20 mmHg or more, 30 mmHg or more, 40 mmHg or more, 50 mmHg or more, 60 mmHg or more, 70 mmHg or more, 80 mmHg or more, 90 mmHg or more, 100 mmHg or more, 110 mmHg or more, 120 mmHg or more, 130 mmHg or more, 140 mmHg or more, 150 mmHg or more, 160 mmHg or more, 170 mmHg or more, 180 mmHg or more, 190 mmHg or more, or 200 mmHg or more relative to the P / F ratio of an untreated subject.

[0071] In some embodiments, administering AP (e.g., RecAP) to a subject increases the subject's P / F ratio relative to a baseline P / F ratio. In some embodiments, the increase in the subject's P / F ratio is 5 mmHg or more, 10 mmHg or more, 20 mmHg or more, 30 mmHg or more, 40 mmHg or more, 50 mmHg or more, 60 mmHg or more, 70 mmHg or more, 80 mmHg or more, 90 mmHg or more, 100 mmHg or more, 110 mmHg or more, 120 mmHg or more, 130 mmHg or more, 140 mmHg or more, 150 mmHg or more, 160 mmHg or more, 170 mmHg or more, 180 mmHg or more, 190 mmHg or more, or 200 mmHg or more relative to the baseline P / F ratio. In some embodiments, administration of AP (e.g., RecAP) to a patient with ARDS reduces the severity of ARDS from "severe" to "moderate," "moderate" to "mild," "severe" to "mild," or "moderate" or "severe" to "no ARDS" (i.e., P / F >300 mmHg).

[0072] In this context, the baseline P / F ratio is defined as the P / F ratio before or around the start of treatment. The baseline P / F ratio is preferably determined on day -2, -1, 0, or 1, i.e., preferably within 48 hours before the start of AP treatment and within 24 hours after the start of AP treatment. More preferably, the baseline P / F ratio is determined within 24 hours before the start of AP treatment and within 12 hours after the start of AP treatment, more preferably between 12 hours before the start of AP treatment and 6 hours after the start of AP treatment, more preferably between 12 hours before the start of AP treatment and the start of AP treatment, and most preferably between 6 hours before the start of AP treatment and the start of AP treatment. AP treatment is preferably continued until the P / F ratio is at least 200 or more, more preferably 300 or more, and most preferably until the subject is considered to no longer have ARDS.

[0073] In some embodiments, an increase in pulmonary function, e.g., an increase in the P / F ratio, is observed on day 1, day 2, day 3, day 4, day 5, day 6, day 7, day 8, day 9, day 10, day 11, day 12, day 13, day 14, day 15, day 16, day 17, day 18, day 19, day 20, day 21, day 22, day 23, day 24, day 25, day 26, day 27, or day 28 after AP administration.

[0074] In some embodiments of the present disclosure, the subject's ARDS is accompanied by or caused by sepsis or viral infection. In some embodiments, the AP (e.g., RecAP) is administered to the subject only if the subject's ARDS is accompanied by or caused by sepsis or viral infection, and sepsis or viral infection is determined within 96 hours before the decision to start treatment. In other embodiments, the AP (e.g., RecAP) is administered to the subject only if the subject's ARDS is accompanied by or caused by sepsis or viral infection, and sepsis or viral infection is determined within 72 hours of detecting ARDS.

[0075] In some embodiments of the present disclosure, the subject's ARDS is accompanied by or caused by sepsis or a viral infection, and treatment of the subject with AP (e.g., RecAP) is initiated within 24 hours after the diagnosis of sepsis or a viral infection. The presence of sepsis can be detected by using criteria developed to determine sepsis, such as SIRS or SOFA. Since its first definition in 1992, the definition of "sepsis" has been reevaluated several times, focusing on the then-prevailing view that the host's systemic inflammatory response syndrome (SIRS) to infection is the cause of sepsis (Bone RC, Balk RA, Cerra FB, et al. American College of Chest Physicians / Society of Critical Care Medicine Consensus Conference: definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis. Crit Care Med. 1992;20(6):864-874). In 2001, the list of diagnostic criteria was expanded, but no alternative criteria were provided (Levy MM, Fink MP, Marshall JC, et al. International Sepsis Definitions Conference. 2001 SCCM / ESICM / ACCP / ATS / SIS International Sepsis Definitions Conference. Intensive Care Med. 2003;29(4):530-538). In 2016, the now widely used "SEPSIS-3" criteria were recommended, and the SIRS criteria were discontinued.Instead, the Sequential [Sepsis-related] Organ Failure Assessment (SOFA) is used to diagnose sepsis (Singer M, Deutschman CS, Seymour CW, et al. The Third International Consensus Definitions for Sepsis and Septic Shock (Sepsis-3). JAMA. 2016;315(8):801-810. doi:10.1001 / jama.2016.0287). The STOP-AKI study, described in the Examples, was conducted from 2014 to 2017 and utilized both SIRS and SOFA scores to identify and / or stratify patients with sepsis. For the definition and determination of "sepsis" used herein, it is preferable to use the SEPSIS-3 criteria based on the SOFA score and described in detail by Singer et al. (Singer M, Deutschman CS, Seymour CW, et al. The Third International Consensus Definitions for Sepsis and Septic Shock (Sepsis-3). JAMA. 2016;315(8):801-810. doi:10.1001 / jama.2016.0287).

[0076] In some aspects of the present disclosure, AP is administered to subjects at risk of sepsis to prevent a decline in lung function.

[0077] In some aspects, the subject is initiated into AP treatment within 48 hours after the detection of ARDS. In a preferred embodiment, the subject is initiated into AP treatment within 48 hours, preferably within 24 hours, more preferably within 12 hours, and most preferably within 6 hours after the detection of moderate or severe ARDS.

[0078] In some aspects of the methods disclosed herein, administration of at least one dose of AP results in a reduction in the duration of or cessation of mechanical ventilation in a subject undergoing mechanical ventilation.

[0079] In some embodiments of the methods disclosed herein, administration of AP results in an increase in lung function or prevents lung function from declining below a critical threshold. Thus, in some embodiments, administration of AP can prevent lung function from declining below a critical threshold. Thus, in some embodiments, to determine the risk that the subject's lung function will decline below a certain threshold level, an index of lung function (e.g., P / F ratio) is determined before administering AP to maintain lung function.

[0080] In some embodiments, the methods disclosed herein involve detecting a change in a marker of lung function (e.g., the P / F ratio), alone or in combination with detecting a change in the levels of one, two, three, or more biomarkers.

[0081] In some embodiments, the methods disclosed herein include predicting an increased clinical response to AP (e.g., RecAP) therapy based on a detected pulmonary function parameter (e.g., baseline P / F ratio). In some embodiments, the disclosed methods include assessing whether the pulmonary function parameter (e.g., baseline P / F ratio) is within a certain range or whether it is above or below a certain threshold (e.g., a P / F ratio threshold for ARDS severity). Thus, for example, if the pulmonary function parameter (e.g., baseline P / F ratio), alone or in combination with other biomarkers, indicates that a patient would benefit from AP therapy, therapy can be initiated, maintained, or modified (e.g., increasing or decreasing the dosage, or increasing or decreasing the frequency of administration).

[0082] Conversely, for example, if pulmonary function parameters (e.g., baseline P / F ratio), alone or in combination with other biomarkers, indicate that a patient will not benefit from AP therapy, the therapy can be discontinued, interrupted, or modified (e.g., by increasing or decreasing the dosage, or by increasing or decreasing the frequency of administration).

[0083] In other words, specific levels of lung function parameters (e.g., baseline P / F ratio), alone or in combination with other molecular or clinical biomarkers, correlate with the clinical effect of AP therapy and help predict clinical outcomes in specific populations of patients with sepsis and / or ARDS.

[0084] In some jurisdictions, the present invention provides alkaline phosphatase (AP) for use in a method of treating acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the subject has a P / F ratio of ≦200 mmHg. Further provided is use of alkaline phosphatase (AP) for the manufacture of a medicament for treating acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the subject has a P / F ratio of ≦200 mmHg.

[0085] The present invention also provides alkaline phosphatase (AP) for use in a method of treating acute respiratory distress syndrome (ARDS) in a subject in need thereof, said method comprising administering to said subject an effective amount of alkaline phosphatase (AP); (i) the subject had moderate or severe ARDS prior to treatment with AP, and (ii) The AP is administered in at least one dose of 300 U / kg to 2,000 U / kg.

[0086] In moderate or severe ARDS, according to the 2012 Berlin definition, the P / F ratio is ≤200 mmHg when measured using a minimum positive end-expiratory pressure (PEEP) of 5 cmH2O.

[0087] In a preferred embodiment, the subject has moderate ARDS with a P / F ratio of >100 mmHg and <200 mmHg before treatment with AP. In another preferred embodiment, the subject has severe ARDS with a P / F ratio of <100 mmHg before treatment with AP.

[0088] In some preferred embodiments, there is provided an AP for use according to the invention or a use according to the invention, wherein said AP is a human AP.

[0089] In some preferred embodiments, the AP is a recombinant AP, preferably a chimeric AP, more preferably an AP having 80% or more, 85% or more, 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 100% sequence identity to the amino acid sequence of RecAP (SEQ ID NO: 1). In some preferred embodiments, the AP is a recombinant AP having 80% or more, 85% or more, 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 100% sequence identity to the amino acid sequence of RecAP (SEQ ID NO: 1), with the proviso that the amino acid at position 279 is leucine (L), the amino acid at position 328 is valine (V), and the amino acid at position 478 is leucine (L).

[0090] In a preferred embodiment, there is provided an AP for use according to the invention or for use according to the invention, wherein administration of said AP results in an increase in pulmonary function in a subject, said increase in pulmonary function preferably comprising an increase in the P / F ratio in said subject compared to the P / F ratio in the subject in the absence of treatment.

[0091] In some preferred embodiments, an AP for use according to the invention or a use according to the invention is provided, wherein the ARDS is associated with or caused by sepsis, preferably the sepsis is detected within 96 hours prior to administration of the AP.

[0092] In a more preferred embodiment, there is provided an AP for use according to the invention or for use according to the invention, wherein the ARDS is associated with or caused by sepsis and the sepsis is detected within 72 hours before the detection of the ARDS. Preferably, the AP treatment is initiated within 24 hours after the sepsis and / or ARDS is detected.

[0093] In other preferred embodiments, there is provided an AP for use according to the present invention or for use according to the present invention, wherein the ARDS is associated with or caused by a viral infection. The viral infection preferably comprises a coronavirus infection, more preferably an infection by a severe acute respiratory syndrome (SARS)-associated coronavirus. In some preferred embodiments, the coronavirus is SARS-CoV-2.

[0094] In a preferred embodiment, an AP for use according to the present invention or a use according to the present invention is provided, wherein the AP is administered once a day. In a preferred embodiment, the AP is administered intravenously. In a preferred embodiment, the AP is administered in three doses a day.

[0095] In some preferred embodiments, there is provided an AP for use according to the present invention or a use according to the present invention, wherein the AP is RecAP, and the dose of RecAP is 0.6 mg / kg (or 375 U / kg) to 3.2 mg / kg (or 2,000 U / kg), preferably 0.8 mg / kg (or 500 U / kg) to 2.0 mg / kg (or 1250 U / kg), more preferably about 1.6 mg / kg (or 1000 U / kg).

[0096] In some preferred embodiments, AP for use according to the present invention or uses according to the present invention are provided, wherein the AP is administered in at least one dose of 500 U / kg to 2,000 U / kg. Preferably, the AP dose is 500 U / kg or 1,000 U / kg of RecAP. In some preferred embodiments, the AP dose is 0.8 mg / kg or 1.6 mg / kg of RecAP.

[0097] In a preferred embodiment, an AP for use according to the present invention or a use according to the present invention is provided, wherein administration of at least one dose of the AP results in a maintenance or increase in the P / F ratio in a subject. Preferably, administration of at least one dose of the AP results in a shortening or cessation of mechanical ventilation in a subject undergoing mechanical ventilation.

[0098] The term "biomarker" as used herein refers to a factor that is a characteristic indicator of a biological process, biological event, and / or pathological condition, e.g., a predictor of clinical response to treatment with AP (e.g., RecAP). As used herein, the term "biomarker" encompasses both clinical markers and molecular biomarkers (biological markers). Thus, in the context of the present disclosure, the term "biomarker" encompasses, for example, "biological biomarkers" or "molecular biomarkers." In some embodiments, biological or molecular biomarkers used to assess pulmonary function include markers of inflammation, such as C-reactive protein (CRP), fibrinogen, interleukin-6 (IL-6), or interleukin-8 (IL-8). More specific biomarkers associated with the diagnosis and outcome of ARDS are receptor for advanced glycation end products (RAGE), angiopoietin-2 (Ang-2), surfactant protein D (SP-D), interleukin-8, Fas and Fas ligand, procollagen peptide (PCP) I and III, octane, acetaldehyde, and 3-methylheptane. Generally, they are cell-specific in response to epithelial or endothelial damage or are involved in inflammatory or infectious responses (Garcia-Laorden MI, Lorente JA, Flores C, Slutsky AS, Villar J. Biomarkers for the acute respiratory distress syndrome: how to make the diagnosis more precise. Ann Transl Med. 2017;5 (14):283).

[0099] As disclosed above, the term "biomarker" also encompasses "clinical biomarkers," also referred to as "clinical condition markers," that can predict response to biological therapy, e.g., gender, age, concomitant medications, smoking status, body mass index (BMI), etc.

[0100] As described above, a cutoff approach based on baseline P / F values ​​is applied to classify ARDS as "severe ARDS" or "moderate ARDS." For example, differences in P / F levels observed in subjects with "severe ARDS," "moderate-to-severe ARDS," or "mild ARDS" can be used to predict clinical outcomes when patients are treated with AP (e.g., RecAP). Thus, if a subject's P / F ratio is below a certain threshold (e.g., 200 mmHg), the subject is a candidate for a specific AP therapy (e.g., therapy with a specific AP regimen including the administration of one or more doses of RecAP).

[0101] In some embodiments, simply determining that the P / F ratio is below a predetermined threshold level is sufficient to identify a subject as a candidate for a particular AP therapy (e.g., treatment with RecAP therapy). Thus, in some embodiments of the methods disclosed herein, the P / F ratio can be used alone. However, in other embodiments, the P / F ratio can be combined with other measures of pulmonary function (e.g., using spirometry) and / or molecular or clinical biomarkers, such as, for example, a marker selected from the group consisting of C-reactive protein (CRP), fibrinogen, interleukin-6 (IL-6) or interleukin-8 (IL-8), receptor for advanced glycation end products (RAGE), angiopoietin-2 (Ang-2), surfactant protein D (SP-D), interleukin-8, Fas and Fas ligand, procollagen peptide (PCP) I and III, octane, acetaldehyde, and 3-methylheptane.

[0102] These findings can be applied, for example, to methods for determining new treatments (e.g., by selecting patients as candidates for a particular AP therapy), treating, preventing, enhancing, or maintaining lung function (e.g., to treat ARDS), monitoring the effectiveness of AP therapy, or adjusting formulations, dosing regimens, or routes of administration.

[0103] The methods disclosed herein include prescribing, initiating, and / or modifying prophylaxis and / or treatment based on the severity of the subject's ARDS, which is typically classified based on the subject's P / F ratio alone or in combination with one or more additional biomarkers, as described above.

[0104] The present disclosure provides methods for determining whether to treat a patient having ARDS with a treatment regimen comprising administration of AP, the methods comprising: (a) determining a P / F ratio (or another pulmonary function parameter), and optionally measuring or directing a clinical laboratory to measure levels of additional biomarkers, such as C-reactive protein (CRP), fibrinogen, interleukin 6 (IL-6) or interleukin 8 (IL-8), receptor for advanced glycation end products (RAGE), angiopoietin-2 (Ang-2), surfactant protein D (SP-D), interleukin-8, Fas and Fas ligand, procollagen peptides (PCP) I and III, octane, acetaldehyde, and / or 3-methylheptane, in a sample obtained from the patient; and (b) determining whether the patient has a higher or lower P / F ratio (or another pulmonary function parameter) compared to a predetermined threshold level and / or compared to a P / F ratio or another pulmonary function parameter in a control, and determining whether the patient has a certain P / F ratio (or another pulmonary function parameter) compared to a predetermined threshold level and / or compared to a P / F ratio or another pulmonary function parameter in a control. Optionally, treating the patient or instructing a healthcare provider to treat the patient with a therapeutic regimen comprising administration of AP (e.g., RecAP), discontinuing treatment, not initiating treatment, refusing treatment, or instructing a healthcare provider to discontinue, not initiate, or refuse treatment, if the sample is determined to have a higher or lower level of an additional biomarker (e.g., C-reactive protein (CRP), fibrinogen, interleukin 6 (IL-6) or interleukin 8 (IL-8), receptor for advanced glycation end products (RAGE), angiopoietin-2 (Ang-2), surfactant protein D (SP-D), interleukin-8, Fas and Fas ligand, procollagen peptides (PCP) I and III, octane, acetaldehyde, and / or 3-methylheptane, etc.) compared to a predetermined threshold level for each biomarker or compared to the level of each biomarker in one or more controls.

[0105] In one aspect, the disclosure provides a method for determining whether to treat a patient having ARDS with a treatment regimen comprising administration of AP, the method comprising (a) determining the P / F ratio (or another pulmonary function parameter), and optionally, measuring in a sample obtained from the patient, C-reactive protein (CRP), fibrinogen, interleukin 6 (IL-6) or interleukin 8 (IL-8), receptor for advanced glycation end products (RAGE), angiopoietin-2 (Ang-2), surfactant protein D (SP-D), interleukin-8 (IL-8), Fas and Fas ligand, procollagen peptide (PCP) I and III, octane, acetaldehyde, and / or 3 and (b) measuring or directing a clinical laboratory to measure the level of an additional biomarker, such as methylheptane, and (b) treating the patient with or directing a healthcare provider to treat, a treatment regimen comprising administration of AP if the patient is determined to have a P / F ratio (or another pulmonary function parameter) that is higher or decreased compared to a predetermined threshold level and / or compared to the P / F ratio or another pulmonary function parameter in a control, and optionally, a higher or lower level of at least one of any additional biomarkers in the sample compared to a predetermined biomarker threshold level or compared to the biomarker level in one or more controls.

[0106] In certain aspects, the disclosure provides methods for determining whether to treat a patient having ARDS with a treatment regimen comprising administration of AP, the method comprising: (a) determining the P / F ratio (or another pulmonary function parameter), and optionally measuring levels of additional biomarkers, such as C-reactive protein (CRP), fibrinogen, interleukin-6 (IL-6) or interleukin-8 (IL-8), receptor for advanced glycation end products (RAGE), angiopoietin-2 (Ang-2), surfactant protein D (SP-D), interleukin-8, Fas and Fas ligand, procollagen peptide (PCP) I and III, octane, acetaldehyde, and / or 3-methylheptane, in a sample obtained from the patient. and (b) discontinuing treatment with a treatment regimen comprising administering AP (e.g., RecAP) to the patient, not initiating treatment, refusing treatment, or instructing a healthcare provider to discontinue, not initiate, or refuse treatment, if the patient is determined to have a higher or increased P / F ratio (or another pulmonary function parameter) compared to a predetermined threshold level and / or compared to the P / F ratio or another pulmonary function parameter in a control, and optionally, a higher or lower level of at least one of any additional biomarkers in the sample compared to a predetermined biomarker threshold level or compared to the biomarker level in one or more controls.

[0107] Also provided is a method for selecting a patient diagnosed with ARDS as a candidate for treatment with AP, the method comprising (a) determining the P / F ratio (or another pulmonary function parameter), and optionally, detecting in a sample obtained from the patient, C-reactive protein (CRP), fibrinogen, interleukin 6 (IL-6) or interleukin 8 (IL-8), receptor for advanced glycation end products (RAGE), angiopoietin-2 (Ang-2), surfactant protein D (SP-D), interleukin-8 (inteleukin-8), Fas and Fas ligand, procollagen peptide (PCP) I and III, octane, acetaldehyde, and / or 3- measuring or directing a clinical laboratory to measure the level of an additional biomarker, such as methylheptane; and (b) treating the patient or directing a healthcare provider to treat with AP if the patient is determined to have a P / F ratio (or another pulmonary function parameter) that is higher or decreased compared to a predetermined threshold level and / or compared to the P / F ratio or another pulmonary function parameter in a control, and optionally, a higher or lower level of at least one of any additional biomarkers in the sample compared to a predetermined threshold level or compared to the biomarker level in one or more controls.

[0108] Also provided is a method for selecting a patient diagnosed with ARDS as a candidate for treatment with AP, the method comprising (a) determining the P / F ratio (or another pulmonary function parameter) and, optionally, measuring the levels of additional biomarkers, such as C-reactive protein (CRP), fibrinogen, interleukin 6 (IL-6) or interleukin 8 (IL-8), receptor for advanced glycation end products (RAGE), angiopoietin-2 (Ang-2), surfactant protein D (SP-D), interleukin-8, Fas and Fas ligand, procollagen peptide (PCP) I and III, octane, acetaldehyde, and / or 3-methylheptane, in a sample obtained from the patient. and (b) interrupting treatment of the patient with AP (e.g., RecAP), not initiating treatment, refusing treatment, or instructing a healthcare provider to interrupt, not initiate, or refuse treatment if the patient is determined to have a higher or increased P / F ratio (or another pulmonary function parameter) compared to a predetermined threshold level and / or compared to the P / F ratio or another pulmonary function parameter in a control, and optionally, a lower or decreased level of at least one of any additional biomarkers in the sample compared to a predetermined threshold level or compared to the biomarker level in one or more controls.

[0109] In some embodiments, the disclosed methods may involve prescribing and / or performing one or more additional assays. For example, a P / F ratio (or another pulmonary function parameter) determining assay may be repeated to rule out a false negative result, and / or one or more additional P / F ratio (or another pulmonary function parameter) determining assays may be performed to monitor the subject's condition. On the other hand, it may be desirable to repeat a P / F ratio (or another pulmonary function parameter) determining assay to rule out a false positive result.

[0110] In some embodiments, the presence of a P / F ratio (or another pulmonary function parameter) above or below a predetermined threshold level in patients with ARDS can be used in combination with one or more clinical or molecular biomarkers specific to sepsis or specific to a particular infection that leads to sepsis in patients with ARDS that is accompanied by or caused by sepsis.

[0111] Those skilled in the art will understand that in accordance with the methods disclosed herein, including but not limited to therapeutic, diagnostic, and monitoring methods, the P / F ratio (or another pulmonary function parameter) can be used as a positive selector, i.e., certain actions (e.g., treating the patient) are taken if the P / F ratio (or another pulmonary function parameter) in the patient is below or above a predetermined P / F ratio (or another pulmonary function parameter) threshold level, or if the P / F ratio (or another pulmonary function parameter) is increased or decreased relative to the P / F ratio (or another pulmonary function parameter) in one or more controls.

[0112] Those skilled in the art will understand that in accordance with the methods disclosed herein, including but not limited to the treatment, diagnosis, and monitoring methods, the P / F ratio (or another pulmonary function parameter) can be used as a negative selector, i.e., no particular action (e.g., treating the patient) is taken if the P / F ratio (or another pulmonary function parameter) in the patient is below or above a predetermined P / F ratio (or another pulmonary function parameter) threshold level, or if the P / F ratio (or another pulmonary function parameter) is increased or decreased relative to the P / F ratio (or another pulmonary function parameter) in one or more controls.

[0113] In certain aspects, the present disclosure includes methods for facilitating a decision by a healthcare provider, healthcare benefit provider, or clinical laboratory as to whether a patient will benefit from treatment with an AP.

[0114] In certain embodiments, the methods disclosed herein include making a diagnosis, which may be a differential diagnosis, based at least in part on the patient's P / F ratio (or another pulmonary function parameter). In some embodiments, the methods disclosed herein include notifying the subject of the results of a P / F ratio (or another pulmonary function parameter) determining assay and / or the results of a diagnosis based at least in part on the P / F ratio (or another pulmonary function parameter). The patient can be notified verbally, in writing, and / or electronically. The diagnosis may also be recorded in the patient's medical record.

[0115] The term "medical record" or "patient medical record" refers to a description of a patient's examination and / or treatment, and typically includes one or more of the patient's medical history and condition, a physician's physical examination, the results of diagnostic tests and procedures, and the patient's medications and therapeutic treatments. A medical record is typically prepared by one or more physicians and / or physician assistants and is a written, transcribed, or otherwise recorded record and / or history of various illnesses or injuries requiring medical care, and / or vaccinations, and / or allergies, and / or treatments, and / or prognosis, and / or health information often relating to parents, siblings, and / or occupation. The record may be reviewed by a physician in diagnosing a condition.

[0116] The medical record may be on paper and / or maintained on a computer-readable medium. The medical record may be maintained by a laboratory, a clinic, a hospital, a healthcare maintenance organization, an insurance company, and / or a personal medical record website. In some embodiments, the diagnosis based at least in part on the determined P / F ratio is recorded on or in a medical alert article, such as a card, a wearable article, and / or a radio frequency identification (RFID) tag. As used herein, the term "wearable article" refers to any article that can be worn on a subject's body, including, but not limited to, a tag, a bracelet, a necklace, an armband, or a headband.

[0117] As used herein, the term "diagnosis" refers to detecting a disease or determining the stage or extent of a disease. Typically, a diagnosis of a disease is made based on the evaluation of one or more factors and / or symptoms indicative of the disease. That is, a diagnosis can be made based on the presence, absence, or amount of a factor that indicates the presence or absence of a disease or disorder. Each factor or symptom considered to be indicative of the diagnosis of a particular disease need not be exclusively related to that particular disease; for example, there may be a differential diagnosis that can be inferred from the diagnostic factor or symptom. Similarly, a factor or symptom indicative of a particular disease may be present in an individual who does not have the particular disease.

[0118] The term "diagnosis" also encompasses determining the therapeutic efficacy of a drug therapy (e.g., AP therapy) or predicting the pattern of response to a drug therapy. Diagnostic methods may be used independently or in combination with other diagnostic and / or staging methods known in the medical arts for a particular disease.

[0119] The term "differential diagnosis" as used herein refers to determining which of two or more diseases with similar symptoms is likely to be the cause of the target symptom based on the analysis of clinical data.This term is also used to refer to determining whether a patient is susceptible to AP treatment according to whether the determined P / F ratio (or other pulmonary function parameter) of the patient is above or below a predetermined threshold level, or whether it is elevated or reduced compared with the level of one or more controls.

[0120] The term "prognosis" as used herein refers to the prediction of the likely course and outcome of a clinical condition or disease (e.g., sepsis or ARDS). Prognosis may be determined by evaluating disease factors or symptoms that are typically indicative of a favorable or unfavorable disease course or outcome. As used herein, the phrase "determining a prognosis" refers to a process by which a person skilled in the art can predict the course or outcome of a patient's condition. The term "prognosis" does not refer to the ability to predict the course or outcome of a condition with 100% accuracy. Instead, those skilled in the art will understand that the term "prognosis" refers to an increased probability that a particular course or outcome may occur. That is, a patient who exhibits a particular symptom has a higher likelihood of experiencing the course or outcome compared to an individual who does not exhibit that symptom.

[0121] As used herein, the terms "good prognosis" and "positive prognosis," or "poor prognosis" and "negative prognosis," are relative terms for predicting the likely course and / or likely outcome of a condition or disease (e.g., sepsis or ARDS). A good prognosis and a positive prognosis predict a better outcome of a condition than a poor prognosis and a negative prognosis. In a general sense, a "good prognosis" is a relatively better outcome than many other possible prognoses that may be associated with a particular condition, whereas a poor prognosis predicts a relatively worse outcome than many other possible prognoses that may be associated with a particular condition. Typical examples of good prognosis and positive prognosis include increased lung function, maintained lung function, and an increased P / F ratio (or another lung function parameter).

[0122] The present disclosure includes a method of treating ARDS in a subject, or an AP for use in a method of treating ARDS in a subject, based on a change in the expression of the P / F ratio (or another pulmonary function parameter). The present disclosure provides an AP for use in a method of treating a patient with ARDS, or a method of treating ARDS in a patient, wherein the method comprises administering an AP to the patient when the patient's P / F ratio (or another pulmonary function parameter) is determined to be low or decreased compared to a predetermined P / F ratio (or another pulmonary function parameter) threshold level or compared to the P / F ratio (or another pulmonary function parameter) in one or more controls.

[0123] The present disclosure provides a method of treating a patient with ARDS, or an AP for use in a method of treating ARDS in a patient, the method comprising: (a) measuring pO2 and FiO2 (or deriving pO2 and FiO2 from another measurable pulmonary function parameter) to determine a P / F ratio (or another pulmonary function parameter); and (b) administering an AP to the patient if the patient's P / F ratio (or another pulmonary function parameter) is low or decreased compared to a predetermined P / F ratio (or another pulmonary function parameter) threshold level or compared to the P / F ratio (or another pulmonary function parameter) in one or more controls.

[0124] Also provided is a method of treating a patient with ARDS, or an AP for use in a method of treating ARDS in a patient, wherein the method includes (a) determining the P / F ratio (or another pulmonary function parameter) of the patient, and (b) interrupting or not initiating administration of an AP (e.g., RecAP) to the patient if the patient's P / F ratio (or another pulmonary function parameter) is high or increased compared to a predetermined P / F ratio (or another pulmonary function parameter) threshold level or compared to one or more control P / F ratio (or another pulmonary function parameter) levels.

[0125] The present disclosure also provides an AP for use in a method of treating a patient with ARDS or a method of treating ARDS in a patient, the method including (a) measuring the patient's pO2 and FiO2 (or deriving pO2 and FiO2 from another measurable pulmonary function parameter) to determine the patient's P / F ratio (or another pulmonary function parameter), and (b) determining whether the P / F ratio (or another pulmonary function parameter) is high or increased, or low or decreased, compared to a predetermined P / F ratio (or another pulmonary function parameter) threshold level. In some embodiments, the method further includes administering or advising a healthcare provider to administer an AP (e.g., RecAP) to the patient when the patient's P / F ratio (or another pulmonary function parameter) is determined to be low or decreased compared to a predetermined P / F ratio (or another pulmonary function parameter) threshold level or compared to one or more control P / F ratio (or other pulmonary function parameter) levels, or discontinuing or refusing to administer an AP when the patient's P / F ratio (or another pulmonary function parameter) is determined to be high or increased compared to a predetermined P / F ratio (or another pulmonary function parameter) threshold level or compared to one or more control P / F ratio (or other pulmonary function parameter) levels.

[0126] Also provided is an AP for use in a method of treating a patient with or for treating ARDS in a patient, the method comprising: (a) measuring the patient's pO2 and FiO2 (or deriving pO2 and FiO2 from another measurable pulmonary function parameter) to determine the patient's P / F ratio (or another pulmonary function parameter); and (b) administering the AP to the patient if the patient's P / F ratio (or another pulmonary function parameter) is determined to be low or decreased compared to a predetermined P / F ratio (or other pulmonary function parameter) threshold level or compared to one or more control P / F ratio (or other pulmonary function parameter) levels, or interrupting, not initiating, or refusing to administer the AP to the patient if the patient's P / F ratio (or another pulmonary function parameter) is determined to be high or increased compared to the predetermined P / F ratio (or other pulmonary function parameter) threshold level or compared to one or more control P / F ratio (or other pulmonary function parameter) levels.

[0127] The present disclosure also provides a method for measuring the efficacy or pharmacodynamics of AP in a patient diagnosed with ARDS, comprising: (a) making a first determination of the patient's P / F ratio (or another pulmonary function parameter); (b) administering AP (e.g., RecAP); and (c) making a second determination of the patient's P / F ratio (or another pulmonary function parameter), wherein an increase in the patient's P / F ratio (or another pulmonary function parameter) in the second determination compared to the patient's P / F ratio (or another pulmonary function parameter) in the first determination indicates that the patient is responding to treatment with AP (e.g., RecAP).

[0128] The present disclosure also provides a method for measuring the efficacy or pharmacodynamics of AP in a patient diagnosed with ARDS, comprising: (a) performing a first determination of the patient's P / F ratio (or another pulmonary function parameter); (b) administering AP (e.g., RecAP); and (c) performing a second determination of the patient's P / F ratio (or another pulmonary function parameter), wherein a decrease in the patient's P / F ratio (or another pulmonary function parameter) in the second determination compared to the patient's P / F ratio (or another pulmonary function parameter) in the first determination indicates that the patient is responding to treatment with AP (e.g., RecAP).

[0129] In some embodiments, the second assessment is performed 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 15 days, 16 days, 17 days, 18 days, 19 days, 20 days, 21 days, 22 days, 23 days, 24 days, 25 days, 26 days, 27 days, or 28 days, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 13 weeks, 14 weeks, 15 weeks, or 16 weeks after administration of the AP (e.g., RecAP), or at any time point therebetween.

[0130] In certain embodiments, in all of the treatment methods disclosed herein, a "loading" dose of AP is administered to achieve a desired level of lung function in the patient. If the AP loading dose does not significantly affect the patient's lung function, a decision can be made to discontinue treatment (e.g., to switch to an alternative therapy).

[0131] If the loading dose results in an increase in the patient's lung function, a decision can 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 healthcare providers to administer treatment, and that final treatment decisions will be based on the healthcare provider's sound judgment.

[0132] The formulation, dosing schedule, and route of administration of the AP (e.g., RecAP) can be adjusted to provide an effective amount for an optimal therapeutic response according to the methods disclosed herein. With regard to administration of the AP, the AP may be administered via any suitable means, composition, and route known in the art. With regard to the dosing schedule, a single bolus can be administered, several divided doses can be administered, or the dose can be proportionally reduced or increased, as indicated by the exigencies of the therapeutic situation.

[0133] III. Alkaline phosphatase (AP) Alkaline phosphatase (AP; EC 3.1.3.1 in the IUBMB enzyme nomenclature) is an enzyme that catalyzes the reaction of phosphatase monoesters and HO to alcohol and phosphate. Other names for AP include alkaline phosphomonoesterase, phosphomonoesterase, glycerophosphatase, alkaline phosphohydrolase, alkaline phenylphosphatase, and orthophosphate monoester phosphohydrolase (alkaline optimal). The systematic name for AP is phosphate monoester phosphohydrolase (alkaline optimal).

[0134] APs are broad-specificity enzymes that also catalyze transphosphorylation. At least four distinct but related APs are known in humans and other mammals: 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 form is located on chromosome 1.

[0135] The term "AP of the present disclosure" refers to isolated alkaline phosphatase, including its splice variants, isoforms, and polymorphs. Recombinant and chimeric APs are also included. In certain embodiments, the AP is RecAP. The amino acid sequence of RecAP is shown in Figure 2. In some embodiments, an AP as used herein has about 70% or more, about 75% or more, about 80% or more, about 85% or more, about 90% or more, about 95% or more, about 96% or more, about 97% or more, about 98% or more, about 99% or more, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 1. In some embodiments, the AP is a functional fragment (i.e., a fragment of the AP, e.g., an AP that maintains about 10% or more, about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, 70% or more, about 80% or more, or about 90% or more of the AP activity of the corresponding full-length AP). In some embodiments, the AP is a variant or derivative of an AP disclosed herein.

[0136] AP for use according to the present disclosure can be a commercially available AP enzyme or any composition comprising an AP enzyme, and any means by which a functional AP enzyme can be produced in the context of the present invention, such as a DNA or RNA nucleic acid encoding an AP protein.

[0137] The nucleic acid encoding the AP may be embedded in a suitable vector, such as a plasmid, a phagemid, a phage, a (retro)virus, a transposon, a gene therapy vector, or any other vector capable of inducing or conferring the production of the AP. Also, natural or recombinant microorganisms, such as bacteria, fungi, protozoa, and yeasts, may be applied as a source of AP in the context of the present disclosure.

[0138] AP-containing compositions for use according to the present disclosure can include eukaryotic AP (e.g., mammalian AP), which can be tissue-nonspecific AP, such as liver / bone or kidney type, or tissue-specific AP types, such as placental type AP, intestinal type AP, and placental-like AP. The latter, also known as germ cell AP, are localized in the testis, thymus, and certain germ cell tumors and are closely related to both the placental and intestinal types of AP.

[0139] In some embodiments, the mammalian AP is a human AP or a bovine AP. Non-limiting 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, the AP comprises a polymorphism. In some embodiments, the AP is a placental AP, a placental-like AP, an intestinal AP, a liver / bone / kidney AP, or a combination thereof. In some embodiments, the AP is a recombinant AP.

[0140] From the viewpoint of three-dimensional structure, AP is roughly composed of two domains: a crown domain and an active site domain. The active site domain can be divided into separate parts such as catalytic residues and three metal ion sites (Zn1, Zn2, Mg3). From the viewpoint of primary structure, the crown domain is adjacent to the amino acids that form the active site domain. The amino acid sequence of AP and the relative positions of the catalytic domain and crown domain are known to those skilled in the art.

[0141] In some embodiments of the present disclosure, 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 different APs, and at least one of the different phosphatases is a human phosphatase. In some embodiments, the AP is, for example, ECAP (Escherichia coli AP) or one of the seven known BIAPs (bovine intestinal APs).

[0142] In some embodiments, the AP is an isolated or recombinant AP comprising a crown domain and a catalytic domain, wherein the crown domain and catalytic domain are obtained from a different AP, and the different AP is a human AP. This is particularly useful when the engineered phosphatase is subsequently used in human therapy. APs for use in the disclosed methods can also be engineered (e.g., genetically engineered) APs of human origin that are non-immunogenic or weakly immunogenic.

[0143] The modified APs disclosed herein can be used, for example, in "in vitro" or "ex vivo" diagnostics or therapeutics. Such modified phosphatases can include, for example, human AP and E. coli AP, or can be composed of bovine AP and E. coli AP.

[0144] In some embodiments of the present disclosure, the AP is an isolated or recombinant AP comprising a crown domain and a catalytic domain, wherein the crown domain and catalytic domain are obtained from different APs, and the crown domain is the crown domain of placental-type AP (ALPP) and the catalytic domain is the catalytic domain of intestinal-type AP (ALPI). In some embodiments, at least one of the different APs is a human phosphatase. In other embodiments, both of the different APs are human phosphatases.

[0145] Domain-swap mutants based on human AP and suitable for the methods disclosed herein are listed in Table 1.

[0146] [Table 3]

[0147] In some embodiments, the AP is a combination of the catalytic domain of ECAP or any of the human forms (ALPI, ALPP, GCAP, or TNAP) with the crown domain of BIAP. Additionally, combinations of the catalytic domain of BIAP with any of the human forms of the crown domain can also be generated.

[0148] In some embodiments, the modified AP is an AP that, under natural conditions, binds to the cell membrane via a glycosylphosphatidylinositol (GPI) anchor but has been modified so that it no longer binds to the cell membrane. All isozymes are functionally active in the cell membrane, and the GPI-anchor-deficient forms do not naturally exist at detectable levels. While serum AP activity has been demonstrated, it is generally accepted that the enzyme still exists in isolated membrane fractions or membrane vesicles. AP activity in milk is also present in fractions containing membrane vesicles. The GPI anchor is stored intracellularly as a precursor molecule, where it binds to the binding site via transamidase. The GPI anchor skeleton is the same as in mammals, but cell-type-dependent modifications are known.

[0149] In some embodiments, when treating human subjects, the AP is human. This is preferred due to the fact that AP types obtained from other species may be immunogenic in human subjects, and treatment may induce immunological reactions and pathological side effects. In some subjects, fatal side effects (i.e., anaphylactic shock) may occur, and therefore, the risk of immunological side effects is preferably minimized by using human AP types.

[0150] Because isolation of human-derived AP is impractical, human recombinant forms of AP proteins can be routinely produced in various recombinant expression platforms. However, expression and purification of GPI-modified and membrane-anchored proteins are extremely challenging. GPI proteins are difficult to separate from the membrane and are difficult to isolate and purify. Thus, in some embodiments, the recombinant AP contains a modification in the GPI signal sequence that results in a secreted AP, i.e., the AP is not bound to the cell membrane.

[0151] Although there is no general sequence involved in GPI anchor binding, there are some specific consensus characteristics: (i) a hydrophobic stretch of amino acids at the C-terminus (at least 11 amino acids, preferably 11 or more amino acids); (ii) upstream of the hydrophobic region, a spacer of hydrophilic amino acids (5–12 amino acids); (iii) GPI binds to the small amino acids glycine, aspartic acid, asparagine, alanine, serine, or cysteine; and (iv) The two subsequent amino acids downstream of the GPI attachment site must be small amino acids, most often selected from glycine, aspartic acid, asparagine, alanine, serine, or cysteine.

[0152] In some embodiments, the recombinant AP comprises a modification in the GPI signal sequence that results in a biologically active secreted AP, i.e., that exhibits activity against biologically relevant substrates. In some embodiments, the secreted AP is a human AP. In some embodiments, the secreted human AP is human liver / kidney / bone-type phosphatase, human intestinal-type AP, or human placental-like alkaline phosphatase.

[0153] Based on the above consensus characteristics, one skilled in the art can introduce modifications, for example, by inserting one or more amino acids that disrupt part of the consensus and result in an AP that cannot bind a GPI anchor. Thus, in some embodiments, the recombinant AP contains a modification of the GPI signal sequence that results in a secreted AP, and this modification includes the mutation or deletion of at least one amino acid in the sequence encompassing the consensus GPI signal sequence.

[0154] In some embodiments, the AP is an AP described in U.S. Patent No. 8,557,545. In some embodiments, the AP is a chimeric AP or chimeric AP-like protein, e.g., as described in U.S. Patent Application Publication Nos. 2017 / 0009216 and 2014 / 0193388. Preferably, the AP is a recombinant alkaline phosphatase comprising the catalytic domain of ALPI (intestinal alkaline phosphatase) and the crown domain of ALPP (placental alkaline phosphatase). More preferably, the AP is RecAP (corresponding to SEQ ID NO: 1 and described in U.S. Patent Application Publication No. 2017 / 0009216).

[0155] In some embodiments, an AP of the present disclosure comprises (i) a sequence having about 70% or more, about 75% or more, about 80% or more, about 85% or more, about 90% or more, about 95% or more, about 96% or more, about 97% or more, about 98% or more, or 99% or more sequence identity to the crown domain of human ALPP, and (ii) a sequence having about 70% or more, about 75% or more, about 80% or more, about 85% or more, about 90% or more, about 95% or more, about 96% or more, about 97% or more, about 98% or more, or 99% or more sequence identity to the catalytic domain of human ALPI.

[0156] In some embodiments, the sequence having sequence identity to the crown domain of ALPP is located in the protein of the present invention at approximately the same position as the crown domain of ALPP in the naturally occurring ALPP protein.

[0157] The percentage of identity of an amino acid or nucleic acid sequence, i.e., 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 the residues in a reference sequence, after aligning the two sequences and introducing gaps, if necessary, to achieve the maximum percent identity. In a preferred embodiment, the calculation of the percentage sequence identity is performed without introducing gaps. Methods and computer programs for alignment are well known in the art, such as "Align 2" or the BLAST service of the National Center for Biotechnology Information (NCBI).

[0158] In some embodiments, the present invention provides alkaline phosphatase (AP) for use in a method of treating acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the method comprises administering an effective amount of alkaline phosphatase (AP) to the subject, wherein the subject has moderate or severe ARDS prior to treatment with the AP, and wherein administration of the AP results in an increase in respiratory function. In a preferred embodiment, the subject has severe ARDS prior to treatment with the AP. In another preferred embodiment, the subject has moderate ARDS prior to treatment with the AP.

[0159] In some preferred embodiments, AP is administered in at least one dose of 500 U / kg to 2,000 U / kg. Preferably, the ARDS is associated with or caused by sepsis, or the ARDS is associated with or caused by a viral infection. If the ARDS is associated with or caused by sepsis, it is preferred that sepsis be detected within 96 hours prior to administration of AP, more preferably within 72 hours prior to detection of ARDS. It is preferred that treatment be initiated within 24 hours after detection of sepsis.

[0160] In some preferred embodiments, the AP is a human AP and / or the AP is a recombinant AP. When the AP is recombinant, it is preferably a chimeric AP. Preferably, the chimeric AP has 80% or more, 85% or more, 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 100% sequence identity to the amino acid sequence of RecAP (SEQ ID NO: 1).

[0161] In some preferred embodiments, there is provided an AP for use according to the present invention, wherein the increase in respiratory function comprises an increase in the P / F ratio compared to the P / F ratio in the absence of treatment with the AP.

[0162] In some preferred embodiments, AP is provided for use according to the present invention, wherein treatment is initiated within 24 hours of detection of ARDS.

[0163] In some preferred embodiments, AP is administered once daily and / or intravenously. Preferably, AP is administered in three doses per day. Preferably, each AP dose is about 0.8 mg / kg or about 1.6 mg / kg of RecAP, and / or each AP dose is about 500 U / kg or about 1000 U / kg of RecAP. "About" in this context means ±20%, preferably ±10%, and most preferably ±5% of the indicated dose. For an average person (e.g., a person weighing approximately 60-80 kg), each dose is about 30,000 U to about 160,000 U, preferably about 30,000 U to about 80,000 U, more preferably about 60,000 U to about 80,000 U. In "mg" units, the dose is about 48 mg to about 256 mg, preferably 48 mg to about 128 mg, and more preferably 96 mg to 128 mg. Furthermore, there is provided an AP for use according to the present invention, wherein the dose is about 30,000 U to about 160,000 U, preferably about 30,000 U to about 80,000 U, and more preferably about 60,000 U to about 80,000 U. There is also provided an AP for use according to the present invention, wherein the dose is about 48 mg to about 256 mg, preferably 48 mg to about 128 mg, and more preferably 96 to 128 mg.

[0164] In some preferred embodiments, there is provided an AP for use according to the present invention, wherein administration of at least one dose of the AP results in a reduction in the duration of or cessation of mechanical ventilation in a subject undergoing mechanical ventilation.

[0165] Preferably, administration of at least one dose of AP results in a maintenance or increase in the P / F ratio in the subject. More specifically, the AP for use according to the present invention results in a reduction in the risk of mortality in the subject. Treatment of one or more subjects who are part of a group of subjects suffering from moderate or severe acute respiratory distress syndrome results in a reduction in the mortality rate of said group of subjects.

[0166] Although the application may describe features as part of the same embodiment or as part of separate embodiments, the scope of the present invention also includes embodiments including any combination of all or some of the features described herein.

[0167] The present invention is further described in the following examples, which do not limit the scope of the invention but merely serve to clarify it. [Example]

[0168] Example 1 Human recombinant alkaline phosphatase for acute respiratory distress syndrome Acute respiratory distress syndrome (ARDS) adversely affects long-term pulmonary outcomes and survival rates. Administration of the detoxification enzyme alkaline phosphatase improved pulmonary inflammation parameters in preclinical studies. We investigated the efficacy and safety of human recombinant alkaline phosphatase (RecAP) in patients with ARDS (Figure 2).

[0169] In Part 1 of the adaptive Phase 2a / 2b STOP-AKI trial, patients were randomized to receive 0.4 mg / kg, 0.8 mg / kg, or 1.6 mg / kg RecAP or placebo once daily for 3 days to establish the optimal dose.

[0170] In Part 2, this dose was compared with placebo. The primary endpoint was the time-adjusted area under the curve (AUC) of endogenous creatinine clearance from days 1 to 7. 1-7 The primary secondary endpoint was renal replacement therapy (RRT). Pulmonary function was also analyzed. Overall, 301 patients were enrolled. No differences in (serious) adverse events were observed between RecAP and placebo. RecAP appeared to be safe and well tolerated.

[0171] method STOP-AKI Study Design and Participants The STOP-AKI trial was an international, randomized, double-blind, placebo-controlled, four-arm, parallel-group, dose-ranging, phase 2a / 2b trial conducted in critically ill adult patients with sepsis-associated acute kidney injury. The protocol and primary results of the STOP-AKI study were previously published (Pickkers P, Mehta RL, Murray PT, et al. Effect of Human Recombinant Alkaline Phosphatase on 7-Day Creatinine Clearance in Patients With Sepsis-Associated Acute Kidney Injury: A Randomized Clinical Trial. JAMA. 2018;320(19):1998-2009).

[0172] ICU patients aged 18 years or older with a diagnosis of sepsis (Levy et al. Crit Care Med 2003;31:1250-6) and a primary diagnosis of AKI (Mehta et al. Crit Care 2007;11:R31) were eligible for study participation. Inclusion and exclusion criteria and main results are described in Pickkers et al. (JAMA 2018).

[0173] Measuring Results The main objectives of this study were to determine the optimal therapeutic dose of RecAP and to evaluate its effects on renal function. Effects on other organs, such as the lungs, were also investigated.

[0174] We also investigated the safety, tolerability, pharmacokinetics (first 120 patients), immunogenicity, systemic and urinary biomarkers, and the effect of RecAP on quality of life and other non-renal parameters in patients with SA-AKI. Pulmonary function analysis is described in detail below. Pulmonary function assessed by arterial oxygen tension (PaO2) / inspiratory oxygen tension (FiO2) (P / F ratio), Carrico index, positive end-expiratory pressure (PEEP), and tidal volume in mechanically ventilated patients. Ventilator-free days: total number of days alive and independent of mechanical ventilation from randomization through day 28. A ventilator-free day was defined as a day on which the patient was not receiving mechanical ventilation (invasive or non-invasive mechanical ventilation). - Number of days on a ventilator: For patients who were on a ventilator at the start of this period, the number of days from the start of the first administration of the study drug to the time they were weaned from the ventilator (total number from Day 1 to Day 28).

[0175] statistical analysis For patients who provided informed consent and were randomly assigned to a treatment group, data were analyzed according to the intention-to-treat principle. Details regarding sample size calculations are provided below.

[0176] Sample size calculation The planned sample size was n1 = 30 patients per treatment group in Part 1, with an additional n2 = 85 patients recruited to the optimal RecAP dose and placebo treatment groups in Part 2 (total sample size of n = 290 patients). Custom-programmed simulations were performed using SAS software V.9.2 to determine the power and type I error rate of the selected sample size and design under various dose-response scenarios. Each scenario assumed a standard deviation for the primary endpoint of 49 mL / min, a placebo response of 60 mL / min, and a RecAP response of 60 mL / min (no treatment effect) to 79 mL / min (strong treatment effect).

[0177] 50,000 simulations were performed, demonstrating a one-sided type I error rate of 2.4% (thus adequately controlled at a one-sided 2.5% significance level). Power was defined as the probability of rejecting the null hypothesis (no difference between treatment groups) when one or more RecAP treatment groups had a treatment effect defined as a response of 69.5 mL / min. This was investigated across seven scenarios, with 10,000 simulations performed for each. The selected design achieved 79%–86% power for scenarios with a strong treatment effect for the medium and high RecAP dose groups. Responses for the low-dose groups varied, with a power of 66%–67% for scenarios in which only the high-dose group showed a strong treatment effect. Sample size determination was based on the number of patients required for an intention-to-treat (ITT) analysis; therefore, patients who were randomly assigned but discontinued before study completion were not replaced.

[0178] The per-protocol analysis compared the intervention and placebo groups for patients who received study medication according to the study protocol and had ≤2 missing ECC values ​​from days 1 to 7, as detailed in the statistical analysis plan.

[0179] For descriptive statistics, continuous variables are presented as mean with standard deviation or median with interquartile range, depending on the distribution. Normally distributed variables were compared using Student's t-test. Non-normally distributed variables were compared using the Mann-Whitney U test. Categorical (and binary) variables are presented as numbers with percentages and analyzed using the chi-square test. Survival analysis with Kaplan-Meier curves was used for graphical presentation. Cox proportional hazards regression analysis was used to estimate hazard ratios for survival on study days 1–28 with RecAP versus placebo, as well as hazard ratios for RRT, shock, and ventilator-free days.

[0180] Analysis of the primary efficacy endpoint was performed by analysis of variance. All analyses performed on secondary endpoints were exploratory only, so no multiplicity adjustment was required. Post-hoc multivariate analyses were performed to establish the robustness of the RecAP effect. All statistical tests (SAS software version 9.4, SAS Institute Inc., Cary, North Carolina, USA) were performed on the intention-to-treat population and two-sided analyses were performed using a 5% significance level.

[0181] result participants Of 326 patients who passed the initial screening at 53 sites in 11 countries in the European Union and North America, 301 patients were enrolled (Figure 4). Patients received RecAP at 0.4 mg / kg (n = 30), 0.8 mg / kg (n = 32), or 1.6 mg / kg (n = 29 in Part 1 and n = 82 in Part 2), or placebo (n = 30 in Part 1 and n = 86 in Part 2). Randomization resulted in balanced demographic and patient characteristics, with the exception of minor baseline differences in renal function between groups (Figure 5).

[0182] safety Treatment-emergent serious adverse events were reported in 43% of patients receiving RecAP 1.6 mg / kg and 50% of patients receiving placebo (see Figure 11). The incidence and nature of adverse events (AEs) were not dose-dependent with RecAP. Antidrug antibody titers were slightly above the limit of detection in nine patients treated with RecAP.

[0183] Consider In this multinational, double-blind, randomized controlled trial in patients with SA-AKI, we observed that the addition of RecAP to standard treatment significantly impacted survival and was associated with improved and sustained recovery of renal function and long-term clinical outcomes, including the composite MAKE endpoint. Because this was a dose-finding, proof-of-principle phase 2 trial, we did not assess renal dysfunction (e.g., short-term ECC) and more patient-centered long-term clinical outcomes (e.g., MAKE). 60-90日目 The study was designed to include endpoints relevant to both treatment and survival.

[0184] In conclusion, RecAP therapy was considered safe and well tolerated. In sepsis patients with AKI, RecAP treatment improved long-term renal function and MAKE 60-90日目 , and a significant improvement in survival rate was observed.

[0185] Example 2 Determination of RecAP enzyme activity and protein concentration Activity Assay: Determination of recAP enzyme activity was based on the conversion (hydrolysis) of 4-nitrophenol phosphate to 4-nitrophenol (yellow color). The change in optical density at 405 nm per unit time is a measure of alkaline phosphatase activity. The assay buffer consisted of 0.25 M glycine buffer (pH 9.6) containing 2 mM MgCl, 0.1 mM ZnCl, and 8.5 mM 4-nitrophenol phosphate at 25 °C.

[0186] The definition of a unit (U) of recAP, expressed in U / mL, is the amount of enzyme that causes the hydrolysis of 1 μmol of 4-nitrophenol phosphate per minute at pH 9.6 and 25°C.

[0187] Protein concentration: Determination of total protein concentration in RecAP drug substance and formulations was performed by UV / Vis analysis. RecAP solutions were analyzed at 280 nm and the absorbance was a measure of protein content (mg / mL) using the following formula: Concentration (mg / mL) = [A / (a × b)] × DF (A = A280, b = path length, a = mass extinction coefficient in 1.01 mL mg-1 cm-1, DF is the dilution factor). ***

[0188] Example 3 Searching for P / F ratio In preparation for studies on its possible beneficial effects in the treatment of COVID-19, it is interesting to investigate the effect of recAP (recombinant alkaline phosphatase) on relevant biomarkers or endpoints, as observed in the STOP-AKI study (described in Example 1 and Pickkers et al. (JAMA 2018)). One relevant endpoint is the P / F ratio, which is the ratio of arterial oxygen tension to inspired oxygen tension. Therefore, the aim of the current investigation is to investigate the possible impact of recAP on the P / F ratio, as observed in the STOP-AKI study.

[0189] The relevant tables of the Analysis Data Model (ADaM) dataset provided by AM-Pharma, specifically the Adverse Drug Reaction (ADRe) dataset, were loaded into the statistical analysis program R (r-project.org; version 3.4.4 (2018-03-15)). This included patients randomly assigned to treatment groups receiving either placebo or 1.6 mg / kg recAP, for which P / F ratios were determined. For all patients with P / F ratio measurements of 300 or less, PEEP was ≥ 5 cmH2O, as defined by the 2012 Berlin definition. The baseline P / F ratio values ​​determined in the screening samples and the sample collected on Day 1 were averaged to increase the stability of the values ​​and reduce the occurrence of missing values, and this was labeled as MeanBase. Patients with missing MeanBase values ​​were excluded from the analysis. Absolute and normalized values ​​are plotted as a function of time after the first dose. Graphical exploration was repeated after subsetting into tertiles of MeanBase values. Mean values ​​were plotted with bootstrapped confidence intervals. Trends observed in these plots were checked at relevant time points using linear regression and non-parametric tests.

[0190] The changes in the P / F ratio, which is the ratio of arterial oxygen partial pressure to inhaled oxygen partial pressure over time after the start of treatment, are shown in Figures 12 to 14. When the complete dataset is considered, the absolute P / F ratio does not change in a specific way over time, and there is no clear difference between placebo and treatment. See Figure 12. The number of patients decreases over the course of the study due to death or discontinuation of the study for other reasons, and the width of the bootstrap confidence interval increases accordingly. However, when the P / F ratio of each patient is determined as the "fold change" relative to the baseline value, patients administered recAP show an increase over time compared to patients treated with placebo (Figure 13).

[0191] As seen in Figure 14, this difference between patients receiving recAP treatment and those receiving placebo treatment is largely due to patients showing a P / F ratio of less than 200 mmHg at baseline (i.e., moderate to severe ARDS patients), while only minor transient differences are seen in the mild ARDS group (200 mmHg < P / F ratio < 300 mmHg) and the group of patients who did not show ARDS at baseline (P / F ratio > 300 mmHg). In conclusion, patients showing moderate to severe ARDS (P / F ratio < 200 mmHg) at baseline benefit the most from recAP treatment.

[0192] As can be seen in Figure 15, in a group of patients with moderate to severe ARDS (P / F ratio < 200 mmHg) at baseline, treatment with alkaline phosphatase resulted in a significant reduction in mortality compared to placebo-treated patients with a P / F ratio < 200 mmHg. Thus, in one embodiment, an AP for use according to the present invention is provided, wherein administration of at least one dose of the AP results in a reduction in the risk of death in the subject. In a preferred embodiment, this reduction is a reduction in the risk of all-cause mortality. Preferably, the risk of death in the subject is reduced compared to if the subject were not treated with the AP. Also provided is an AP for use according to the present invention, wherein administration of at least one dose of the AP results in a reduction in mortality in the subject group. In a preferred embodiment, this reduction is a reduction in all-cause mortality. Preferably, mortality is reduced in a group of subjects treated with the AP compared to a group of subjects with severe to moderate ARDS not treated with the AP.

[0193] Mortality, mortality risk, all-cause mortality, and all-cause mortality risk have their usual meanings in the context of the present invention. Mortality or all-cause mortality in the context of this specification refers to the mortality rate from all causes of death in a population in a particular time period. Mortality risk or all-cause mortality risk in this context refers to the risk of death from all causes for an individual in a particular time period.

[0194] It is understood that the Detailed Description section, and not the Summary and Abstract sections, is intended to be used to interpret the claims. The Summary and Abstract sections may describe one or more example embodiments of the invention, but not all, as contemplated by the inventors, and therefore are not intended to limit the scope of the invention and the appended claims in any way.

[0195] The present invention has been described above with the aid of functional components that illustrate the implementation of specific functions and relationships thereof. The boundaries of these functional components have been arbitrarily defined herein for the convenience of description. Alternative boundaries can be defined so long as the specific functions and relationships thereof are appropriately performed.

[0196] The foregoing description of specific embodiments sufficiently reveals the general nature of the present invention so that those skilled in the art can, by applying knowledge within the art, readily modify and / or adapt the specific embodiments, etc., for various uses without undue experimentation and without departing from the general concept of the present invention. Therefore, such adaptations and modifications are intended to be within the meaning and range of equivalents of the disclosed embodiments, based on the teaching and guidance presented herein. It should be understood that the phrases or terminology herein are intended to be descriptive rather than limiting, as the terms or phrases herein would be interpreted by one of ordinary skill in the art in light of the teaching and guidance.

[0197] The breadth and scope of the present invention should not be limited by any of the above-described exemplary embodiments, but should be defined only in accordance with the following claims and their equivalents.

[0198] All publications, patents, patent applications, and / or other documents cited in this application are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication, patent, patent application, and / or other document was individually indicated to be incorporated by reference for all purposes. Aspects of the present invention include the following. <1> 1. An alkaline phosphatase (AP) for use in a method of treating acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the subject has moderate or severe ARDS. <2> The subject has a 5cmH 2 Moderate or severe ARDS with a P / F ratio of ≤200 mmHg when measured using a minimum positive end-expiratory pressure (PEEP) of O <1> AP for use as described in. <3> The subject has severe ARDS with a P / F ratio of ≦100 mmHg before treatment with the AP. <2> AP for use as described in. <4> The subject has moderate ARDS with a P / F ratio of >100 mmHg and <200 mmHg before treatment with the AP. <2> AP for use as described in. <5> The AP is administered in at least one dose of 500 U / kg to 2,000 U / kg. <1> ~ <4> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <6> The ARDS is associated with or caused by sepsis. <1> ~ <5> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <7> The ARDS is associated with or caused by a viral infection. <1> ~ <6> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <8> wherein the AP is a human AP; <1> ~ <7> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <9> The AP is a recombinant AP, preferably the recombinant AP is chimeric. <1> ~ <8> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <10> The AP has 80% or more, 85% or more, 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 100% sequence identity to the amino acid sequence of RecAP (SEQ ID NO: 1). <1> ~ <9> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <11> administration of the AP results in an increase in respiratory function, preferably wherein the increase in respiratory function comprises an increase in the subject's P / F ratio compared to the subject's P / F ratio in the absence of treatment with the AP; <1> ~ <10> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <12> The AP treatment is initiated within 48 hours after detection of ARDS. <1> ~ <11> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <13> administration of at least one dose of the AP results in a reduction in the duration of or cessation of mechanical ventilation in a subject receiving mechanical ventilation; <1> ~ <12> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <14> AP is administered once daily, <1> ~ <13> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <15> AP is administered intravenously, <1> ~ <14> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <16> AP is administered in three doses per day, <1> ~ <15> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <17> The dose of AP is 0.8 mg / kg or 1.6 mg / kg of RecAP; <1> ~ <16> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <18> The dose of AP is 500 U / kg or 1000 U / kg of RecAP; <1> ~ <17> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <19> administration of at least one dose of the AP results in a maintenance or increase in the P / F ratio in the subject; <1> ~ <18> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <20> administration of at least one dose of the AP results in a reduced risk of death in the subject; <1> ~ <19> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5. <21> administration of at least one dose of said AP results in a reduction in mortality in a group of subjects including said subject; <1> ~ <19> 1. The method of claim 1, wherein the AP is for use as described in any one of claims 1 to 5.

Claims

1. 1. A medicament for treating acute respiratory distress syndrome (ARDS) in a subject, said medicament comprising alkaline phosphatase (AP), said subject having moderate or severe ARDS.

2. The subject has a blood pressure of 5 cmH ２ 2. The method of claim 1, wherein the patient has moderate or severe ARDS with a P / F ratio of ≦200 mmHg as measured using a minimum positive end-expiratory pressure (PEEP) of O.

3. 3. The method of claim 2, wherein the subject has severe ARDS with a P / F ratio of ≦100 mmHg prior to treatment with the AP.

4. 3. The method of claim 2, wherein the subject has moderate ARDS with a P / F ratio >100 mmHg and ≦200 mmHg before treatment with the AP.

5. The method of any one of claims 1 to 4, wherein the AP is administered in at least one dose of 500 U / kg to 2,000 U / kg.

6. The method according to any one of claims 1 to 5, wherein the ARDS is associated with or caused by sepsis.

7. The method according to any one of claims 1 to 6, wherein the ARDS is associated with or caused by a viral infection.

8. The agent according to any one of claims 1 to 7, wherein the AP is a human AP.

9. The agent according to any one of claims 1 to 8, wherein said AP is a recombinant AP, preferably said recombinant AP is chimeric.

10. The agent according to any one of claims 1 to 9, wherein the AP has 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, 99% or more, or 100% sequence identity to the amino acid sequence of RecAP (SEQ ID NO: 1).

11. 11. The method of claim 1, wherein administration of the AP results in an increase in respiratory function, preferably wherein the increase in respiratory function comprises an increase in the P / F ratio of the subject compared to the P / F ratio of the subject in the absence of treatment with the AP.

12. The method according to any one of claims 1 to 11, wherein the treatment with AP is initiated within 48 hours after the detection of ARDS.

13. The method of any one of claims 1 to 12, wherein administration of at least one dose of the AP results in a reduction in the duration of or cessation of mechanical ventilation therapy in a subject receiving mechanical ventilation therapy.

14. The method according to any one of claims 1 to 13, wherein the AP is administered once a day.

15. The method according to any one of claims 1 to 14, wherein the AP is administered intravenously.

16. The method according to any one of claims 1 to 15, wherein the AP is administered in three doses per day.

17. The method of any one of claims 1 to 16, wherein the dose of AP is 0.8 mg / kg or 1.6 mg / kg of RecAP (SEQ ID NO: 1).

18. The method of any one of claims 1 to 17, wherein the dose of AP is 500 U / kg or 1000 U / kg of RecAP (SEQ ID NO: 1).

19. The method of any one of claims 1 to 18, wherein administration of at least one dose of the AP results in a maintenance or increase in the P / F ratio in the subject.

20. The method of any one of claims 1 to 19, wherein administration of at least one dose of the AP results in a reduction in the risk of death in the subject.

21. The method of any one of claims 1 to 19, wherein administration of at least one dose of said AP results in a reduction in mortality in a group of subjects including said subject.

22. 1. Use of alkaline phosphatase (AP) in the manufacture of a medicament for treating acute respiratory distress syndrome (ARDS) in a subject, wherein the subject has moderate or severe ARDS.

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