CXCR4 inhibitor compound in the treatment and / or the prevention of severe ards

Abstract

The present invention relates to a CXCR4 inhibitor compound for use in the treatment of a severe acute respiratory distress syndrome (sARDS), wherein the sARDS is characterized by a dysregulation of NETosis which may be measured by the plasmatic levels of cDNA, citrullinated histone H3, neutrophil elastase, interleukin-8, myeloperoxidase-DNA complexes and / or calprotectin. The present invention also relates to a method of prognostic of severe ARDS, a pharmaceutical composition for use in the treatment of severe acute respiratory distress syndrome (ARDS), a kit of prognostic of severe ARDS in a subject and the use of NETosis biomarkers.

Description

[0001] CXCR4 INHIBITOR COMPOUND IN THE TREATMENT AND / OR THE PREVENTION OF SEVERE ARDS

[0002] FIELD OF INVENTION

[0003]

[0001] The present invention relates to the prevention and / or the treatment of severe acute respiratory distress syndrome characterized by a dysregulation of NETosis.

[0004] BACKGROUND OF INVENTION

[0005]

[0002] ARDS (acute respiratory distress syndrome) is a manifestation of acute lung injury (ALI), commonly resulting from sepsis, trauma, and severe pulmonary infections. ARDS is mainly associated with infection by viral agents such as influenza or SARS- Cov-2 but it can also be caused by infection by bacterial agents including common microorganisms such as Streptococcus pneumoniae, Haemophillus influenzae, Moraxella catarrhalis and Streptococcus pyogenes, or fungi.

[0006]

[0003] The evolution of ARDS is characterized by several phases: an exudative phase and a proliferative phase, which may be followed by a fibrotic phase in some patients. Although mechanical ventilation is the most important supportive therapy for patients with ARDS, it can paradoxically induce or aggravate lung injury. Thus, there is a need to develop new therapies for treating ARDS, and in particular the most severe cases of ARDS leading to very high mortality rates in patients.

[0007]

[0004] Currently, there is no reliable method to predict the evolution and outcome of ARDS in individual patients, underscoring a significant unmet medical need. The lack of predictive biomarkers hampers the ability to stratify patients accurately for treatment and clinical trial enrollment, which is critical for the development of targeted therapies. Recognizing the variability in patient responses and outcomes, the identification and validation of specific biomarkers for ARDS would enable the stratification of patients based on their risk profile and likely disease trajectory. This stratification would facilitate more personalized therapeutic approaches, optimizing clinical outcomes while minimizing exposure to potentially ineffective or harmful treatments. Consequently, there is a pressing need for the development and integration of predictive biomarkers into the clinical management of ARDS to enhance prognostication, guide therapeutic decisionmaking, and improve patient outcomes in this critically ill population.

[0008]

[0005] NETosis is a process by which immune cells, specifically polynuclear neutrophils release fibers composed of DNA and proteins to trap pathogens. By forming these Neutrophil Extracellular Traps (NETs), neutrophils are able to kill pathogens while limiting damage to host cells. It has been shown that not only bacteria but also fungi such as Candida albicans may induce NETosis. NETosis has also been observed in a context of a wider infection, such as sepsis (the NETs may be produced in lung capillaries and liver sinusoids for example).

[0009]

[0006] NETosis is known to be involved in numerous pathological conditions at different degree. In healthy subjects, NETs levels are low in blood. Plasma Blood levels of three NETs biomarkers were found to increase with COVID-19 clinical severity and high levels of NETs biomarkers were associated with lower survival rate among ARDS patients. The abundance of NETs is therefore a good indicator that a subject suffers from severe ARDS.

[0010]

[0007] On another hand, CXCR4 receptors are known to be involved in the treatment of ARDS.

[0011]

[0008] In this context, the inventors have found that CXCR4 inhibitor compounds, such as plerixafor, are particularly efficient in (i) reducing the likelihood that patients with ARDS will develop severe ARDS and (ii) are also more efficient in treating patients suffering with severe ARDS associated with heavy dysregulation of NETosis, compared with patients with mild or moderate ARDS. More generally, the inventors discovered that CXCR4 inhibitors are a particularly good treatment for patients with abundant NETs. SUMMARY

[0012]

[0009] The present invention relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment and / or the prevention of a severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the sARDS is characterized by a dysregulation of NETosis and wherein the subject has a PaO2 / FiO2 ratio inferior or equal to 200 mmHg, preferably inferior or equal to 100 mmHg.

[0013]

[0010] In some embodiments, the dysregulation of NETosis is characterized by the presence of at least one of the following NETosis biomarkers measured in a biological sample from the subject:

[0014] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0015] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0016] - citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,

[0017] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0018] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0019] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0020] - calprotectin levels equal to or greater than 5 pg / mL.

[0021] [Oi l] In some embodiments, the dysregulation of NETosis is characterized by the presence of at least one of the following NETosis biomarkers measured in a biological sample from the subject:

[0022] - the ratio of circulating DNA levels in the subject compared to circulating DNA levels in a healthy individual is equal to or greater than 1,4; - the ratio of citrullinated histone H3 levels in the subject compared to citrullinated histone H3 levels in a healthy individual is equal to or greater than 2,0;

[0023] - the ratio of citrullinated histone H3-DNA complexes levels in the subject compared to citrullinated histone H3-DNA complexes levels in a healthy individual is equal to or greater than 1,8;

[0024] - the ratio of neutrophil elastase levels in the subject compared to neutrophil elastase levels in a healthy individual is equal to or greater than 3,5;

[0025] - the ratio of interleukin-8 levels in the subject compared to interleukin-8 levels in a healthy individual is equal to or greater than 30;

[0026] - the ratio of myeloperoxidase-DNA complexes levels in the subject compared to myeloperoxidase-DNA complexes levels in a healthy individual is equal to or greater than 1,4; and / or

[0027] - the ratio of calprotectin levels in the subject compared to calprotectin levels in a healthy individual is equal to or greater than 10.

[0028]

[0012] In some embodiments, the compound is of formula (I), and formula (I) is:

[0029] ( I )

[0030] - Z and Y, which are identical or different, represent: o an alkyl group having from 1 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms, or o an aryl or heteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms, or o a cyclic or heterocyclic polyamine group having from 9 to 32 ring members and from 3 to 8 amine groups in the ring spaced by 2 or more carbon atoms from each other, optionally comprising an aryl or heteroaryl group having from 3 to 6 carbon atoms, a heteroatom, and optionally substituted by a halogen atom, a hydroxyl group, a heteroatom, an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, or o a D1D2N- group, wherein DI and D2, which may be identical or different, represent:

[0031] • a hydrogen atom, or

[0032] • an alkyl group having from 1 to 6 carbon atoms optionally substituted by at least one hydroxyl group, a halogen atom, a CF3 group, a CN group, an amine group, or an alkyl, O-alkyl or S-alkyl group having from 1 to 12 carbon atoms, or

[0033] • an aryl, an heteroaryl, a cycloalkyl, a heterocycloalkyl, an alkylaryl, an alkylheteroaryl or an alkylheteropolyaryl group having from 3 to 12 carbon atoms, optionally substituted by at least one hydroxyl group, halogen atom, CF3 group, CN group, amine group, or alkyl, O-alkyl or S-alkyl group having from 1 to 12 carbon atoms, or

[0034] • DI and D2 are linked together to from a N-containing aryl or heteroaryl group having from 3 to 12 carbon atoms and optionally substituted by at least one amine group optionally substituted by an alkylheteroaryl group having from 3 to 12 carbon atoms, and

[0035] - A represents: o an aryl or heteroaryl group having from 3 to 12 carbon atoms, or o an alkyl group having from 1 to 6 carbon atoms, or o -R4-Y’ -R5-, wherein R4 and R5 which are identical or different represent an alkyl group having from 1 to 6 carbon atoms and Y’ represents an aryl or heteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a halogen atom, a hydroxyl group, an amide group, an amine group, an alkoxy group, an ester group, a CF3 group, a CN group or an alkyl, O-alkyl or S-alkyl group having from 1 to 6 carbon atoms optionally substituted by a hydroxyl group, an amine group or an O-alkyl group having from 1 to 6 carbon atoms, and

[0036] - R and R’, which are identical or different, represent an alkyl group having from 1 to 6 carbon atoms or a R1NR2R3 group or a single bond, and

[0037] - R1 represents a single bond or an alkyl group having from 1 to 6 carbon atoms, and

[0038] - R2 and R3, which are identical or different, represent a hydrogen atom, an amine group, an alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, wherein said aryl or heteroaryl group having from 3 to 6 carbon atoms is optionally substituted by an alkyl group having from 1 to 6 carbon atoms.

[0039]

[0013] In some embodiments, the CXCR4 inhibitor compound is plerixafor, LY2510924, AMD3465, MSX-122, mavorixafor, motixafortide and a mixture thereof, preferably plerixafor.

[0040]

[0014] The present invention further relates to a method of prognostic of severe ARDS in a subject wherein the subject has a PaO2 / FiO2 ratio inferior or equal to 200 mmHg, preferably inferior or equal to 100 mmHg, comprising the following steps:

[0041] (a) measuring the levels of a NETosis biomarker in a biological sample from a subject, in particular from the subject’s plasma,

[0042] (b) comparing said NETosis biomarker with a subject of reference and / or threshold of NETosis,

[0043] (c) determining whether the subject is at risk of suffering from a severe ARDS depending on the levels of NETosis biomarker, and (d) optionally treating the subject with a CXCR4 inhibitor compound.

[0044]

[0015] In some embodiments, the CXCR4 inhibitor compound of step (d) is selected from the list consisting of plerixafor, LY2510924, AMD3465, MSX-122, mavorixafor, motixafortide and a mixture thereof, preferably plerixafor.

[0045]

[0016] In some embodiments, the NETosis biomarker measured in a biological sample from the subject is circulating DNA levels, citrullinated histone H3 levels, interleukin-8 levels, myeloperoxidase-DNA complexes levels, neutrophil elastase levels and / or calprotectin levels.

[0046]

[0017] In some embodiments, step (d) is the administration of a CXCR4 inhibitor compound to the patient with severe ARDS via one of the following routes: intravenous, intramuscular, subcutaneous, parenteral, oral, nasal, ocular, transmucosal, or transdermal.

[0047]

[0018] In some embodiments, step (d) corresponds to the continuous intravenous administration of a CXCR4 inhibitor compound to the patient with severe ARDS.

[0048]

[0019] The present invention further relates to a pharmaceutical composition for use in the treatment and / or in the prevention of severe acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the severe ARDS is characterized by a dysregulation of NETosis, wherein the subject has a PaO2 / FiO2 inferior to 200 mmHg, preferably inferior or equal to 100 mmHg ; and wherein said pharmaceutical composition comprises a CXCR4 inhibitor compound and a pharmaceutically acceptable excipient.

[0049]

[0020] In some embodiments, the severe ARDS results from a viral, a fungal or a bacterial infection.

[0050]

[0021] The present invention further relates to a kit for prognostic of a severe ARDS comprising the means for measuring in a biological sample from a subject at least one of NETosis biomarkers selected from the list consisting of circulating DNA levels, citrullinated histone H3 levels, neutrophil elastase levels, interleukin-8 levels, myeloperoxidase-DNA, complexes levels, and calprotectin levels.

[0022] The present invention further relates to the use of at least one NETosis biomarker selected from circulating DNA, citrullinated histone H3, citrullinated histone H3-DNA complexes, neutrophil elastase, interleukin- 8, myeloperoxidase-DNA complexes, and calprotectin as a biomarker for in vitro detection of severe ARDS in a biological sample, preferably the at least one biomarker is selected from circulating DNA, citrullinated histone H3 and neutrophil elastase.

[0051] DEFINITIONS

[0052]

[0023] In the present invention, the following terms have the following meanings:

[0053]

[0024] Within the meaning of the invention “a” or “an” refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, “an element” means one element or more than one element.

[0054]

[0025] Within the meaning of the invention “about” is used herein to mean approximately, roughly, around, or in the region of. When the term "about" is used in conjunction with a numerical range, it modifies that range by extending the boundaries above and below the numerical values set forth.

[0055]

[0026] “Alkyl” by itself or as part of another substituent refers to a linear, branched or cyclic saturated hydrocarbyl group. Typically, alkyl groups comprise from 1 to 12 carbon atoms, preferably from 1 to 6 carbon atoms, more preferably from 1 to 3 carbon atoms, furthermore preferably 1 to 2 carbon atoms. Alkyl groups may be linear or branched and may be substituted as indicated herein. Nonlimiting examples of alkyl groups include methyl, ethyl, zz-propyl, z-propyl, zz-butyl, zbutyl, .s-butyl and / -butyl, pentyl and its isomers (e.g., zz-pentyl, zso -pentyl), and hexyl and its isomers (e.g., zz-hexyl, / .so-hcxyl). Preferred alkyl groups include methyl, ethyl, / / propyl, z-propyl, / / -butyl, .s-butyl, and t- butyl. “Heteroalkyl”, in particular "heterocycloalkyl", refers to an alkyl, in particular a cycloalkyl, group comprising at least one heteroatom preferably selected from the group consisting of O, P, N, S and Si, which is more preferably N. “Alkylaryl” refers to an aryl group substituted by at least one alkyl group. “Alkoxy” refers to O-alkyl.

[0027] “Amine” refers to NH2 group. In some embodiments, the amine groups may be substituted, thus providing secondary or tertiary amine groups.

[0056]

[0028] “Aryl” refers to a cyclic, polyunsaturated, aromatic hydrocarbyl group comprising at least one aromatic ring. Aryl groups may have a single ring (z.e., phenyl) or multiple aromatic rings fused together (e.g., naphthyl) or linked covalently. Typically, aryl groups have from 5 to 12 carbon atoms, preferably from 6 to 10 carbon atoms. “Heteroaryl” refers to an aryl group comprising at least one heteroatom preferably selected from the group consisting of O, P, N, S and Si, which is more preferably N. “Arylalkyl” refers to an alkyl group substituted by at least one aryl group.

[0057]

[0029] “At least one” refers to one, two, three or more.

[0058]

[0030] “Halogen” refers to fluoro, chloro, bromo or iodo. Preferred halogen groups include fluoro and chloro.

[0059]

[0031] “Hydrate of a compound” refers to a molecular complex comprising the compound and one or more pharmaceutically acceptable solvent molecules, wherein the solvent is water.

[0060]

[0032] “Plerixafor®” or “plerixafor” refers to a bicyclam derivative which chemical name is 1 , l'-[ 1 ,4-phenylene-bis(methylene)] -bis- 1,4,8,11 -tetraazacyclotetradecane. Plerixafor is also designated under the name of AMD3100, and its CAS reference number is 110078-46-1.

[0061]

[0033] “Salt of a compound” refers to acid addition or base salts of said compound. Suitable acid addition salts are formed from acids which form non-toxic salts. Examples include the acetate, adipate, aspartate, benzoate, besylate, bicarbonate / carbonate, bisulphate / sulphate, borate, camsylate, citrate, cyclamate, edisylate, esylate, formate, fumarate, gluceptate, gluconate, glucuronate, hexafluorophosphate, hibenzate, hydrochloride / chloride, hydrobromide / bromide, hydroiodide / iodide, isethionate, lactate, malate, maleate, malonate, mesylate, methylsulphate, naphthylate, 2-napsylate, nicotinate, nitrate, orotate, oxalate, palmitate, pamoate, phosphate / hydrogen phosphate / dihydrogen phosphate, pyroglutamate, saccharate, stearate, succinate, tannate, tartrate, tosylate, trifluoroacetate and xinofoate salts. Suitable base salts are formed from bases which form non-toxic salts. Examples include the aluminium, arginine, benzathine, calcium, choline, diethylamine, 2(diethylamino)ethanol, diolamine, ethanolamine, glycine, 4(2hydroxyethyl)-morpholine, lysine, magnesium, meglumine, morpholine, olamine, potassium, sodium, tromethamine and zinc salts. In some embodiments, the salt is selected from the group consisting of hydrobromide, hydrochloride, dihydrobromide, dihydrochloride, zinc and copper salts.

[0062]

[0034] Within the meaning of the invention the “subject” is an animal, preferably a mammal, more preferably a human, receiving the composition of the invention. In the sense of the present invention, a subject may be a patient, i.e., a person receiving medical attention, undergoing or having underwent a medical treatment, or monitored for the development of a disease.

[0063]

[0035] “NETosis”: or neutrophil extracellular trap (NET) formation refers to a process where neutrophils release their chromatin contents along with antimicrobial peptides to trap and kill pathogens. While this mechanism is primarily a part of the immune response against pathogens, dysregulation of NETosis has been associated with various diseases. The process of NETosis includes the release of the granule components into the cytosol (containing proteins and protease including myeloperoxidase and calprotectin), the modification of histones leading to chromatin decondensation, the destruction of the nuclear envelope, as well as formation of pores in the plasma membrane. NETosis is therefore associated with an increase of the concentration of cell-free DNA, and with the increase of cytokines (including interleukin-8). Interleukin-8 is also known to be an activator of NETosis. The release of myeloperoxidase into the cytosol leads to an increase of myeloperoxidase-DNA complexes (MPO), and the modification of histones leads to a specific increase in the levels of citrullinated histone H3.

[0064]

[0036] “NETopathy”: is a disease associated with a dysregulation of NETosis, in particular an abnormal increase of NETosis. For example, the following diseases may be NETopathies: autoimmune diseases, thrombosis, cardiovascular diseases, cancer, chronic obstructive pulmonary disease (COPD), sepsis, inflammatory bowel disease (IBD), diabetes, autoinflammatory diseases.

[0037] ARDS refers to an acute respiratory distress syndrome. ARDS may be defined as a form of acute lung injury (ALI) and occurs as a result of a severe pulmonary injury that causes alveolar damage heterogeneously throughout the lung. ARDS is associated with at least one of the following symptoms: shortness of the breath, rapid breathing. ARDS is a rapidly progressive disorder that initially manifests as dyspnea, tachypnea, and hypoxemia, then quickly evolves into respiratory failure.

[0065] ARDS may be defined according to the American-European Consensus Conference (AECC), i.e., ARDS is defined as a respiratory failure of acute onset with a PaOi / FiOi ratio < 200 mm Hg (regardless of the levels of positive end-expiratory pressure, PEEP), bilateral infiltrates on frontal chest radiograph, and a pulmonary capillary wedge pressure <18 mmHg (if measured) or no evidence of left atrial hypertension.

[0066] ARDS may be evaluated according to the Berlin definition for ARDS. According to this definition, patients are considered as having ARDS if they have: (1) acute respiratory failure not fully explained by cardiac failure or fluid overload, as judged by the treating physician; (2) bilateral opacities consistent with pulmonary edema on the chest radiograph or the computed tomography scan; (3) onset within 1 week after a known clinical insult or new / worsening respiratory symptoms. The Berlin definition categorizes ARDS according to the ratio of arterial oxygen partial pressure (PaOi) to fractional inspired oxygen (FiO2) (PaC FiCh): mild (PaCWFiOi: 201-300 mmHg), moderate (PaCE / FiCE: 101-200 mmHg), and severe (PaOi / FiOi < 100 mm Hg), with a PEEP> 5 cm H2O.

[0067] ARDS may be evaluated by the Murray score system. The Murray scoring system is based on 4 pulmonary variables: hypoxemia with PaCWFiOi ratio, positive end-expiratory pressure (PEEP) in cmH20, respiratory system compliance in mL / cmH20 and chest X- rays (CXR) quadrants infiltrated. Each of the 4 criteria is awarded a value from 0 to 4 according to the severity of the condition. The final score can be interpreted as follows: 0 points: no lung injury; 1 to 2.5 points: mild to moderate lung injury; >2.5 points: ARDS.

[0068]

[0038] “Marker”, in particular “biomarker” or “biological marker” refers to a variable that can be measured in a biological sample from a subject. A marker may reflect the presence, the risk, and / or the progression of a disease state, thus serving as a prognostic tool.

[0039] “c-DNA”: “Circulating DNA”, “cDNA”, “cfDNA” or “cell-free DNA” herein refers to DNA that exists outside a cell in a subject or the isolated form of such DNA, typically in a body fluid, such as blood. For example, the cfDNA molecule is a 15 single- or double- stranded DNA molecule, with a size ranging from 50 to 150 bp. The circulating DNA can arise from a tumor cell and therefore be called a tumor circulating DNA.

[0069]

[0040] “CXCR4”: CX-C chemokine receptor type 4, also named CD 184, also named fusin, is an alpha-chemokine receptor that plays a central role in cell migration but also in other essential processes such as the development of the immune system. CXCR4 is commonly expressed by leukemic cells of both myeloid and lymphoid lineage and the cytokine SDF-1 (also named CXCL12) is its ligand, which is released by the bone marrow.

[0070]

[0041] “CXCR4 inhibitor compound”: refers to a natural or synthetic compound inducing an inhibition or down-regulation of a biological activity associated with activation of CXCR4, including any of the downstream biological effects otherwise resulting from the binding of the CXCR4 to its natural ligand. A CXCR4 inhibitor compound may be a CXCR4 antagonist.

[0071]

[0042] “Therapeutically effective amount” or “therapeutically effective dose” refers to the amount or dose or concentration of a CXCR4 inhibitor compound as described herein that is aimed at, without causing significant negative or adverse side effects to the subject in need of treatment, preventing, reducing, alleviating or slowing down (lessening) one or more of the symptoms or manifestations of a NETopathy, i.e. a disease characterized by a dysregulation of NETosis, in particular of severe acute respiratory distress syndrome (sARDS) characterized by a dysregulation of NETosis.

[0072]

[0043] “Prognostic” or “prognosis” refers to the prediction of the likelihood or expected development of a disease, including whether the signs and symptoms will improve or worsen (and how quickly) or remain stable over time, and the likelihood of survival (including life expectancy). Herein the term is used to refer to the prediction of the evolution of ARDS in a subject, i.e. whether it will evolve from mild ARDS or moderate ARDS to severe ARDS.“Prophylactically treating” refers to reducing the incidence, severity, relapse, or occurrence of a disease. Herein, “prophylactically treating sARDS” refers to the reduction of the incidence of severe ARDS.

[0073]

[0044] “Treating” or “Treatment” refers to a therapeutic treatment and includes the eradication, removal, amelioration, modification, reduction, management, or control of a pathological condition, in particular of a ARDS and severe ARDS, “treating” or “treatment” also refers to a therapeutic treatment wherein the object is to prevent, reduce, alleviate, and / or slow down (lessen) one or more of the symptoms or manifestations of a disease.

[0074] DETAILED DESCRIPTION

[0075]

[0045] The present invention relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment of a disease in a subject in need thereof, wherein the disease is characterized by a dysregulation of NETosis, in particular by high levels of NETosis in comparison with a subject of reference.

[0076]

[0046] A disease characterized by a dysregulation of NETosis may be selected in the group consisting of: autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus (SLE), vasculitis; cardiovascular diseases such as thrombosis, deep vein thrombosis (DVT), pulmonary embolism, myocardial infection, and stroke; cancer; chronic obstructive pulmonary disease (COPD); acute respiratory distress syndrome; sepsis; inflammatory bowel disease (IBD), Crohn’s disease, ulcerative colitis; diabetes and its complications such as diabetic nephropathy and diabetic retinopathy; and autoinflammatory diseases such as familial Mediterranean fever and gout.

[0077]

[0047] This invention relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment of severe acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the severe ARDS is associated with a dysregulation of NETosis, in particular is characterized by the presence of an excessive amount of neutrophil extracellular traps (NETs).

[0078]

[0048] This invention also relates to a pharmaceutical composition for use in the treatment of severe acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the severe ARDS is associated with a dysregulation of NETosis, and wherein said pharmaceutical composition comprises a CXCR4 inhibitor compound and a pharmaceutically acceptable excipient.

[0079]

[0049] The dysregulation of NETosis may be characterized by the presence of an excessive amount of neutrophil extracellular traps (NETs).

[0080]

[0050] The dysregulation of NETosis may be characterized by an excessive level of any of the following NETosis biomarkers: circulating DNA (cDNA), citrullinated histone H3, neutrophil elastase, myeloperoxidase-DNA complexes, interleukin-8 and / or calprotectin in a biological sample from a subject.

[0081]

[0051] In some embodiments, the at least one NETosis biomarker in a biological sample from a subject is selected from the list consisting of circulating DNA (cDNA), citrullinated histone H3, neutrophil elastase, myeloperoxidase-DNA complexes, interleukin-8 and / or calprotectin, preferably the at least one NETosis biomarker in a biological sample from a subject is circulating DNA (cDNA).

[0082]

[0052] According to the invention, the CXCR4 inhibitor compound allows to treat a severe ARDS characterized by the dysregulation of NETosis. Severe ARDS may be an ARDS with a ratio of PaCE / FiCE inferior or equal to 100 mm Hg, with a Positive end- expiratory pressure (PEEP) superior or equal to 5 cm H2O, as defined in the Berlin Definition.

[0083]

[0053] The invention also relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment and / or the prevention of a severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein sARDS is characterized by a dysregulation of NETosis, wherein the subject has a PaOi / FiOi ratio inferior or equal to 200 mmHg, preferably has a PaOi / FiOi ratio inferior or equal to 100 mmHg.

[0084]

[0054] The invention also relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment and / or the prevention of a severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the subject is characterized by a dysregulation of NETosis and, wherein the subject has a PaOi / FiCh ratio inferior or equal to 200 mmHg, preferably has a PaOi / FiOi ratio inferior or equal to 100 mmHg.

[0085]

[0055] The invention also relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment and / or the prevention of a severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the subject has a PaOi / FiCh ratio inferior or equal to 200 mmHg, preferably has a PaOi / FiCh ratio inferior or equal to 100 mmHg.

[0086]

[0056] In some embodiments, the CXCR4 inhibitor compound or the pharmaceutically acceptable salt and / or hydrate thereof, is for use in the prevention of a severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the subject has a PaOi / FiCh ratio inferior or equal to 200 mmHg and, wherein the subject is characterized by the presence of at least one NETosis biomarker in a biological sample selected from:

[0087] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0088] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0089] - citrullinated histone H3-DNA complexes levels equal or greater than 150 ng / mL,

[0090] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0091] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0092] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0093] - calprotectin levels equal to or greater than 5 pg / mL.

[0057] In some embodiments, the CXCR4 inhibitor compound or the pharmaceutically acceptable salt and / or hydrate thereof, is for use in the prevention of a severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the subject has a PaOi / FiCh ratio inferior or equal to 200 mmHg and is at risk of developing a sARDS, wherein the subject is characterized by the presence of at least one NETosis biomarker in a biological sample selected from:

[0094] - the ratio of circulating DNA levels in the subject compared to circulating DNA levels in a healthy individual is equal to or greater than 1,4;

[0095] - the ratio of citrullinated histone H3 levels in the subject compared to citrullinated histone H3 levels in a healthy individual is equal to or greater than 2,0;

[0096] - the ratio of citrullinated histone H3-DNA complexes levels in the subject compared to citrullinated histone H3-DNA complexes levels in a healthy individual is equal to or greater than 1,8;

[0097] - the ratio of neutrophil elastase levels in the subject compared to neutrophil elastase levels in a healthy individual is equal to or greater than 3,5;

[0098] - the ratio of interleukin-8 levels in the subject compared to interleukin-8 levels in a healthy individual is equal to or greater than 30;

[0099] - the ratio of myeloperoxidase-DNA complexes levels in the subject compared to myeloperoxidase-DNA complexes levels in a healthy individual is equal to or greater than 1,4; and / or

[0100] - the ratio of calprotectin levels in the subject compared to calprotectin levels in a healthy individual is equal to or greater than 10.

[0101]

[0058] In some embodiments, the CXCR4 inhibitor compound or the pharmaceutically acceptable salt and / or hydrate thereof, is for use in the treatment of a severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein a subject has a PaOi / FiOi ratio inferior or equal to 100 mmHg, wherein the subject is characterized by the presence of at least one NETosis biomarker in a biological sample selected from:

[0102] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0103] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0104] - citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,

[0105] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0106] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0107] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0108] - calprotectin levels equal to or greater than 5 pg / mL.

[0109]

[0059] In some embodiments, the CXCR4 inhibitor compound or the pharmaceutically acceptable salt and / or hydrate thereof, is for use in the treatment of a severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein a subject has a PaOi / FiOi ratio inferior or equal to 100 mmHg, wherein the subject is characterized by the presence of at least one NETosis biomarker in a biological sample selected from:

[0110] - the ratio of circulating DNA levels in the subject compared to circulating DNA levels in a healthy individual is equal to or greater than 1,4;

[0111] - the ratio of citrullinated histone H3 levels in the subject compared to citrullinated histone H3 levels in a healthy individual is equal to or greater than 2,0;

[0112] - the ratio of citrullinated histone H3-DNA complexes levels in the subject compared to citrullinated histone H3-DNA complexes levels in a healthy individual is equal to or greater than 1,8;

[0113] - the ratio of neutrophil elastase levels in the subject compared to neutrophil elastase levels in a healthy individual is equal to or greater than 3,5;

[0114] - the ratio of interleukin-8 levels in the subject compared to interleukin-8 levels in a healthy individual is equal to or greater than 30; - the ratio of myeloperoxidase-DNA complexes levels in the subject compared to myeloperoxidase-DNA complexes levels in a healthy individual is equal to or greater than 1,4; and / or

[0115] - the ratio of calprotectin levels in the subject compared to calprotectin levels in a healthy individual is equal to or greater than 10.

[0116]

[0060] In some embodiments, the severe ARDS results from a viral, a fungal or a bacterial infection.

[0117]

[0061] In some embodiments, the severe ARDS is associated with a medical condition selected from the group consisting of barotrauma (volutrauma), pulmonary embolism (PE), ventilator- associated pneumonia (VAP), gastrointestinal bleeding (ulcer), dysmotility, aspiration, vascular injury, pneumothorax (by placing pulmonary artery catheter), tracheal injury / stenosis (result of intubation and / or irritation by endotracheal tube), blood clots, inhalational lung injury, lung contusion, chest trauma, near-drowning, trauma (e.g. fat embolism), cardiopulmonary bypass, burns, viral infection.

[0118]

[0062] Thus, the CXCR4 inhibitor compound for use in the present invention may be for treating severe ARDS, or for preventing the onset or the aggravation of a severe ARDS, wherein severe ARDS is caused by or related to infection by a virus, a bacterial or a fungus.

[0119]

[0063] The CXCR4 inhibitor compound for use in the present invention may be for prophylactic treatment of sARDS, or for reducing the likelihood of the worsening of ARDS into severe ARDS, or for slowing down the worsening of ARDS into severe ARDS, wherein sARDS is caused by or related to infection by a virus, a bacterial or a fungus.

[0120]

[0064] Examples of viruses that may cause severe ARDS include, but are not limited to, viruses of the coronaviridae family, viruses of the herpesviriadae family, respiratory syncytial virus and influenza viruses. Examples of viruses of the herpesviridae family that may cause ARDS include, without being limited to, herpes simplex virus (HSV) and cytomegalovirus (CMV). Examples of influenza viruses that may cause ARDS include, without being limited to, H5N1 (avian influenza A) and H1N1 (influenza A).

[0121]

[0065] The viral infection may be caused by a coronavirus.

[0122]

[0066] Thus, the CXCR4 inhibitor compound for use in the present invention may be for treating severe ARDS or for preventing the onset or the aggravation of severe ARDS, wherein said severe ARDS is caused by or related to infection by a virus, such as coronavirus.

[0123]

[0067] The CXCR4 inhibitor compound for use in the present invention may be for prophylactic treatment of sARDS, or for reducing the likelihood of the worsening of ARDS into severe ARDS, or for slowing down the worsening of ARDS into severe ARDS, wherein said sARDS is caused by or related to infection by a virus, such as coronavirus.

[0124]

[0068] The coronavirus may be a human coronavirus. The coronavirus may be an alpha coronavirus or a beta coronavirus.

[0125]

[0069] Examples of alpha coronaviruses include, without being limited to, human coronavirus 229E (HCoV-229E) and human coronavirus NL63 (HCoV-NL63) also sometimes known as HCoV-NH or New Haven human coronavirus. Examples of beta coronaviruses include, without being limited to, human coronavirus OC43 (HCoV- OC43), human coronavirus HKU1 (HCoV-HKUl), Middle East respiratory syndrome- related coronavirus (MERS-CoV) previously known as novel coronavirus 2012 or HCoV- EMC, severe acute respiratory syndrome coronavirus (SARS-CoV) also known as SARS- CoV-1 or SARS-classic, and severe acute respiratory syndrome coronavirus (SARS- CoV-2) also known as 2019-nCoV or novel coronavirus 2019.

[0126]

[0070] The coronavirus may be selected from the group comprising or consisting of HCoV-229E, HCoV-NL63, HCoV-OC43, HCoV-HKUl, MERS-CoV, SARS-CoV-1 and SARS-CoV-2. The viral infection may be caused by SARS-CoV-2 (causing the disease COVID-19).

[0071] Thus, the CXCR4 inhibitor compound for use in the present invention is for treating severe ARDS or for preventing the onset or aggravation of severe ARDS, wherein said ARDS is caused by or related to an infection by SARS-CoV-2.

[0127]

[0072] The CXCR4 inhibitor compound for use in the present invention may be for treating severe ARDS or for preventing the onset or aggravation of severe ARDS or for prophylactic treatment of sARDS, or for reducing the likelihood of the worsening of ARDS into severe ARDS, or for slowing down the worsening of ARDS into severe ARDS, wherein said sARDS is caused by or related to an infection by SARS-CoV-2.

[0128]

[0073] The subject may be suffering from COVID-19 and developed severe ARDS. The subject may be suffering from CO VID- 19 and be at risk of developing severe ARDS. The subject may have developed micro-thrombotic events (and optionally macro-thrombotic events). The subject may present at least one of the following symptoms: cough, shortness of breath or difficulty breathing. The subject may present at least one, preferably at least two, of the following symptoms: fever (i.e., any body temperature over 38°C), chills, repeated shaking with chills, muscle pain, headache, sore throat and new loss of taste or smell.

[0129]

[0074] The severe ARDS may be caused by an infection by bacterial agents selected from the list comprising: Streptococcus pneumoniae, Haemophillus influenzae, Moraxella catarrhalis and Streptococcus pyogenes, Staphyloccocus aureus, Legionella pneumophila, Clamydia pneumoniae, Mycoplasma pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumanii, Stenotrophompnas maltophilia.

[0130]

[0075] In some embodiments, the severe ARDS is associated with sepsis.

[0131]

[0076] The ARDS or severe ARDS may be caused by an infection by fungi selected from the list comprising: Candida spp, Aspergillus spp, Mucorales.

[0132]

[0077] The CXCR4 inhibitor compound as described herein may be a compound of formula (I), wherein formula (I) is: - Z and Y, which are identical or different, represent: o an alkyl group having from 1 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms, or o an aryl or heteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms, or o a cyclic or heterocyclic polyamine group having from 9 to 32 ring members and from 3 to 8 amine groups in the ring spaced by 2 or more carbon atoms from each other, optionally comprising an aryl or heteroaryl group having from 3 to 6 carbon atoms, a heteroatom, and optionally substituted by a halogen atom, a hydroxyl group, a heteroatom, an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, or o a D 1D2N- group, wherein D 1 and D2, which may be identical or different, represent:

[0133] • a hydrogen atom, or

[0134] • an alkyl group having from 1 to 6 carbon atoms optionally substituted by at least one hydroxyl group, a halogen atom, a CF3 group, a CN group, an amine group, or an alkyl, O-alkyl or S-alkyl group having from 1 to 12 carbon atoms, or

[0135] • an aryl, an heteroaryl, a cycloalkyl, a heterocycloalkyl, an alkylaryl, an alkylheteroaryl or an alkylheteropolyaryl group having from 3 to 12 carbon atoms, optionally substituted by at least one hydroxyl group, halogen atom, CF3 group, CN group, amine group, or alkyl, O-alkyl or S-alkyl group having from 1 to 12 carbon atoms, or

[0136] • DI and D2 are linked together to from a N-containing aryl or heteroaryl group having from 3 to 12 carbon atoms and optionally substituted by at least one amine group optionally substituted by an alkylheteroaryl group having from 3 to 12 carbon atoms, and

[0137] - A represents: o an aryl or heteroaryl group having from 3 to 12 carbon atoms, or o an alkyl group having from 1 to 6 carbon atoms, or o -R4-Y’-R5-, wherein R4 and R5 which are identical or different represent an alkyl group having from 1 to 6 carbon atoms and Y’ represents an aryl or heteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a halogen atom, a hydroxyl group, an amide group, an amine group, an alkoxy group, an ester group, a CF3 group, a CN group or an alkyl, O-alkyl or S-alkyl group having from 1 to 6 carbon atoms optionally substituted by a hydroxyl group, an amine group or an O-alkyl group having from 1 to 6 carbon atoms, and

[0138] - R and R’, which are identical or different, represent an alkyl group having from 1 to 6 carbon atoms or a R1NR2R3 group or a single bond, and

[0139] - R1 represents a single bond or an alkyl group having from 1 to 6 carbon atoms, and

[0140] - R2 and R3, which are identical or different, represent a hydrogen atom, an amine group, an alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, wherein said aryl or heteroaryl group having from 3 to 6 carbon atoms is optionally substituted by an alkyl group having from 1 to 6 carbon atoms. wherein:

[0141] Z and Y, which are identical or different, represent: o an alkyl group having from 1 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms, or o an aryl or heteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms, or o a cyclic or heterocyclic polyamine group having from 9 to 32 ring members and from 3 to 8 amine groups in the ring spaced by 2 or more carbon atoms from each other, optionally comprising an aryl or heteroaryl group having from 3 to 6 carbon atoms, a heteroatom, and optionally substituted by a halogen atom, a hydroxyl group, a heteroatom, an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, and

[0142] A represents an aryl or heteroaryl group having from 3 to 12 carbon atoms, and

[0143] R and R’ , which are identical or different, represent an alkyl group having from 1 to 6 carbon atoms or a R1NR2R3 group, and

[0144] Ri represents a single bond or an alkyl group having from 1 to 6 carbon atoms, and

[0145] R2 and R3, which are identical or different, represent a hydrogen atom, an amine group, an alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, wherein said aryl or heteroaryl group having from 3 to 6 carbon atoms is optionally substituted by an alkyl group having from 1 to 6 carbon atoms.

[0146]

[0079] The CXCR4 inhibitor compound may be of formula (I), wherein:

[0147] Z and Y are identical and represent cyclic polyamine moieties having from 9 to 20 ring members and from 3 to 6 amine groups in the ring spaced by 2 or more carbon atoms from each other, and

[0148] A represents an aryl or heteroaryl group having from 3 to 8 carbon atoms, and R and R’, which are identical or different, represent a methylene linked to a nitrogen atom of Z or Y, the nitrogen atoms being otherwise unsubstituted.

[0149]

[0080] Z and / or Y may be cyclic polyamine containing 9-32 ring members of which 3- 8 are nitrogen atoms, said nitrogen atoms being separated from each other by at least 2 carbon atoms. Said cyclic polyamine may contain 10-20 ring members, preferably 14 or 16 ring members, more preferably 14 ring members. Said cyclic poly amine may contain 3-6 nitrogen atoms, preferably 4 or 5 nitrogen atoms, more preferably 5 nitrogen atoms. Said nitrogen atoms may be separated from each other by at least 2 carbon atoms, in particular by 2 or 3 carbon atoms. Said cyclic poly amine may contain additional heteroatoms besides nitrogen, in particular oxygen or sulfur, and / or may be fused to an additional ring system, in particular to a pyridine.

[0150]

[0081] In particular, A may be selected from the group consisting of phenylene, pyridine, thiophene, pyrazine and imidazole.

[0082] The compound of formula (I) may be selected from the group consisting of the compounds represented by the following structures:

[0151] compound 11

[0152]

[0083] The compound of formula (I) may be l,l'-[l,4-phenylene-bis(methylene)]-bis- 1,4,8, 11-tetraazacyclotetradecane, also named plerixafor, AMD3100 or AMD-3100 (CAS No.: 110078-46-1).

[0084] The compound of formula (I) may be represented by the following formula (II): wherein:

[0153] DI and D2, which may be identical or different, represent: o an alkyl group having from 1 to 6 carbon atoms optionally substituted by at least one hydroxyl group, a halogen atom, a CF3 group, a CN group, an amine group, or an alkyl, O-alkyl or S-alkyl group having from 1 to 12 carbon atoms, or o an aryl, an heteroaryl, a cycloalkyl, a heterocycloalkyl, an alkylaryl, an alkylheteroaryl or an alkylheteropoly aryl group having from 3 to 12 carbon atoms, optionally substituted by at least one hydroxyl group, halogen atom, CF3 group, CN group, amine group, or alkyl, O-alkyl or S- alkyl group having from 1 to 12 carbon atoms, or o DI and D2 are linked together to from a N-containing aryl or heteroaryl group having from 3 to 12 carbon atoms and optionally substituted by at least one amine group optionally substituted by an alkylheteroaryl group having from 3 to 12 carbon atoms, and

[0154] X represents: o an alkyl group having from 1 to 6 carbon atoms, or o -R4-Y’-R5-, wherein R4 and R5 which are identical or different represent an alkyl group having from 1 to 6 carbon atoms and Y’ represents an aryl or heteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a halogen atom, a hydroxyl group, an amide group, an amine group, an alkoxy group, an ester group, a CF3 group, a CN group or an alkyl, O-alkyl or S-alkyl group having from 1 to 6 carbon atoms optionally substituted by a hydroxyl group, an amine group or an O-alkyl group having from 1 to 6 carbon atoms.

[0155]

[0085] Y’ may be selected from the group consisting of phenylene, pyridine and thiophene.

[0156]

[0086] The compound of formula (I) may be represented by the above-mentioned formula (II), wherein:

[0157] DI and D2, which may be identical or different, represent an aryl or heteroaryl group having from 3 to 12 carbon atoms, optionally substituted by a hydroxyl group or an alkyl group having from 1 to 6 carbon atoms, and

[0158] X represents an alkyl group having from 1 to 6 carbon atoms.

[0159]

[0087] DI may be a monocyclic or bicyclic fused ring system containing at least one nitrogen atom, for example a 5, 6, 7, 8 -tetrahydroquinoline.

[0088] D2 may be a monocyclic or fused bicyclic unsubstituted or substituted ring system containing at least one heteroatom selected from N, O and S. D2 may be selected from the group consisting of substituted or unsubstituted dihydroquinoline, tetrahydroquinoline, pyranopyridine, dihydropyranopyridine, thiapyranopyridine, dihydrothiapyranopyridine, dihydronaphthyridine, tetrahydronaphthyridine, imidazole, oxazole, thiazole, benzimidazole, benzothiazole, and benzoxazole.

[0160]

[0089] In some embodiments, the compound of formula (I) is represented by the following formula (III): wherein:

[0161] El represents an alkyl group having from 1 to 12 carbon atoms, or a heteroaryl group having from 3 to 12 carbon atoms, and

[0162] E2 represents a heteroalkyl group having from 1 to 12 carbon atoms, substituted by an amine group, and

[0163] E3 represents a heteroalkyl group having from 1 to 12 carbon atoms.

[0090] In some embodiments, the compound of formula (I) is selected from the group consisting of the compounds represented by the following structures: compound 14 compound 15 compound 18

[0091] The compound of formula (I) may be N'-(lH-benzimidazol-2- ylmethyl)-N'- [(8S)-5,6,7,8-tetrahydroquinolin-8-yl]butane-l,4-diamine, also named AMD070 or mavorixafor.

[0164]

[0092] The CXCR4 inhibitor compound as described herein may be selected from the group consisting of an indole-based compound, a N-substituted indole compound, a bicyclam compound, a cyclam mimetic compound, a paraxylylenediamine-based compound, a guanidine-based compound, a tetrahydroquinoline-based compound, and a 1 ,4-phenylenebis(methylene) compound.

[0165]

[0093] The CXCR4 inhibitor compound as described herein may be selected from the group consisting of plerixafor (AMD3100), burixafor (TG-0054), LY2510924, AMD3329, AMD3465, mavorixafor (AMD070), MSX-122, CTCE-9908, WZ811, and B KT- 140.

[0166]

[0094] The CXCR4 inhibitor compound as described herein may be selected from the group consisting of plerixafor (AMD3100), LY2510924, AMD3465, MSX-122, mavorixafor (AMD070), and mixtures thereof.

[0167]

[0095] The CXCR4 inhibitor compound as described herein may be 2-[4-[6-amino-2- [ [4- [ [3 - (cyclohexylamino)propylamino] methyl] cyclohexyl] methylamino] pyrimidin-4- yl]piperazin-l-yl]ethylphosphonic acid, also named TG-0054 or burixafor (CAS No.: 1191448-17-5).

[0168]

[0096] The CXCR4 inhibitor compound as described herein may be: acetic acid;(5S)-N- [(2S)-l-amino-l-oxo-6-(propan-2-ylamino)hexan-2-yl]-2-benzyl-l l-[3- (carbamoylamino)propyl]-5-[(4-hydroxyphenyl)methyl]-14-(naphthalen-2-ylmethyl)- 3,6,9,12,15,18,23-heptaoxo-8-[4-(propan-2-ylamino)butyl]-l,4,7,10,13,16,19- heptazacyclotricosane-20-carboxamide, also named LY2510924 (CAS No.: 1088715-84-7) .

[0169]

[0097] The CXCR4 inhibitor compound as described herein may be 7-[[4-(4,7, 10,17- tetrazabicyclo [11.3.1 ]heptadeca- 1(17), 13, 15 -trien-7 -ylmethyl)phenyl] methyl] - 4,7,10,17-tetrazabicyclo[11.3.1]heptadeca-l(17),13,15-triene, also named AMD3329 (CAS No.: 170861-87-7).

[0170]

[0098] The CXCR4 inhibitor compound as described herein may be N-[[4-(l,4,8,l 1- Tetraazacyclotetradec- 1 -ylmethyl)phenyl]methyl] -2-pyridinemethanamine hexahydrobromid, also named AMD3465 (CAS No.: 185991-07-5).

[0171]

[0099] The CXCR4 inhibitor compound as described herein may be N-{4-[(2- pyrimidinylamino)methyl]benzyl}-2-pyrimidinamine, also named MSX-122 (CAS No.: 897657-95-3).

[0172]

[0100] The CXCR4 inhibitor compound as described herein may be CTCE-9908 (CAS No.: 1030384-98-5).

[0173]

[0101] The CXCR4 inhibitor compound as described herein may be Nl,N4-Di-2- pyridinyl-l,4-benzenedimethanamine, also named WZ811 (CAS No.: 55778-02-4).

[0174]

[0102] The CXCR4 inhibitor compound as described herein may be N2-(4- fluorobenzoyl)-L-arginyl-L-arginyl-3-(2-naphthalenyl)-L-alanyl-L-cysteinyl-L-tyrosyl- N5-(aminocarbonyl)-L-ornithyl-L-lysyl-D-lysyl-L-prolyl-L-tyrosyl-L-arginyl-N5- (aminocarbonyl)-L-omithyl-L-cysteinyl-L-argininamide, cyclic (4^ l 3)-disulfidc, also named BKT-140, also named motixafortide, also named BL-8040 (CAS No.: 664334-36- 5).

[0175]

[0103] The CXCR4 inhibitor compound as described herein may be a nucleic acid. Exemplary CXCR4 nucleic acid inhibitors include, but are not limited to, antisense oligonucleotides, siRNAs, shRNAs, microRNAs, aptamers, ribozymes and decoy oligonucleotides .

[0176]

[0104] The CXCR4 inhibitor compound as described herein may be a CXCR4 antisense oligonucleotide.

[0177]

[0105] The CXCR4 inhibitor compound as described herein may bind to a ligand binding site on the CXCR4 receptor and thus interfere with the binding of the ligand to the CXCR4 receptor.

[0106] The CXCR4 inhibitor compound as described herein may bind to a non-ligand binding site on the CXCR4 receptor and thus interfere with the binding of the ligand to the CXCR4 receptor.

[0178]

[0107] The CXCR4 inhibitor compound as described herein may bind to a CXCR4 receptor ligand and thus interfere with the binding of the ligand to the CXCR4 receptor.

[0179]

[0108] The CXCR4 inhibitor compound as described herein, may inhibit the expression of a polynucleotide (e.g., mRNA) expressing CXCR4.

[0180]

[0109] The CXCR4 inhibitor compound as described herein may be an antibody or an antibody fragment.

[0181]

[0110] The CXCR4 inhibitor as described herein may be a small molecule, for example, AMD3100, ALX40-4C, T22, T140, Met-SDF-1 beta, T134, or AMD-3465.

[0182]

[0111] The CXCR4 inhibitor as described herein may be ALX40-4C (CAS No.: 143413-49-4).

[0183]

[0112] The CXCR4 inhibitor as described herein may be [Tyr5,12, Lys7]-polyphemusin II, also named T22 (CAS No.: 142960-16-5).

[0184]

[0113] The CXCR4 inhibitor as described herein may be (4S)-6-chloro-4-(2- cyclopropylethynyl)-4-(trifluoromethyl)-lH-3,l-benzoxazin-2-one, also named Met- SDF-1 beta (CAS No.: 154598-52-4).

[0185]

[0114] The CXCR4 inhibitor as described herein may be Arg-Arg-Trp-Cys-Tyr-Arg- Lys-D-Lys-Pro-Tyr-Arg-Cit-Cys-Arg, also named T134 (CAS No.: 205586-56-7).

[0186]

[0115] The CXCR4 inhibitor as described herein may be derivatives of T140, including but not limited to: TN14003, TC14012 (CAS No.: 368874-34-4), and TE14011.

[0187]

[0116] The CXCR4 inhibitor as described herein may be TCI 4012 (CAS No.: 368874-34-4).

[0117] The CXCR4 inhibitor as described herein may be 4-fluorobenzoyl-Arg-Arg- Nal-Cys-Tyr-Cit-Lys-d-Lys-Pro-Tyr-Arg-Cit-Cys-Arg-NH2, also named 4F- benzoyl-TN 14003.

[0188]

[0118] The CXCR4 inhibitor compound as described herein may be an antagonist to CXCR4.

[0189]

[0119] The CXCR4 inhibitor compound may decrease the function as compared to a reference function induced by CXCR4 activation, such as, for example, a function inferior or equal to 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10%, or less of the reference function.

[0190]

[0120] The inhibition of CXCR4 may be measured after contacting a cell with a compound tested for its impact on function, and the reference function correspond to a function induced by CXCR4 activation measured in a cell not contacted with said compound.

[0191]

[0121] In some embodiments, the CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, is used in the treatment of a severe acute respiratory distress syndrome (sARDS), wherein the sARDS is characterized by a dysregulation of NETosis, especially by a dysregulation of at least one NETosis biomarker.

[0192]

[0122] In some embodiments, the at least one NETosis biomarker is selected from the list consisting of circulating DNA (cDNA), citrullinated histone H3, neutrophil elastase, myeloperoxidase-DNA complexes, interleukin-8 and / or calprotectin, preferably the at least one NETosis biomarker is circulating DNA (cDNA).

[0193]

[0123] In some embodiments, the at least one NETosis biomarker measured in a biological sample from a subject is 1.1 time, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2 times times, 3 times, 4 times, 5 times or more higher in a subject in comparison with the same at least one NETosis biomarker measured in a biological sample from a subject of reference, preferably may be two times or more higher in a subject in comparison with the same at least one NETosis biomarker measured in a biological sample from a subject of reference.

[0124] Said increase may be measured by any methods suitable for measuring the NETosis biomarkers in a subject, such as the one disclosed therein: measure of the levels of circulating DNA (cDNA), measure of the levels of citrullinated histone H3, measure of the levels of neutrophil elastase, measure of the levels of myeloperoxidase-DNA complexes, measure of the levels of interleukin- 8, and / or measure of the levels of calprotectin.

[0194]

[0125] In some embodiments, the at least one NETosis biomarker may be measured in a biological sample from a subject, such as in a tissue sample from a subject, in blood sample from a subject, such as in plasma sample from a subject and / or in serum sample from a subject.

[0195]

[0126] Severe ARDS may be associated with at least one of the following biomarkers of NETosis:

[0196] - levels of circulating DNA (cDNA), such as levels of plasmatic circulating DNA (cDNA)

[0197] - levels of citrullinated histone H3, such as blood levels or plasma levels,

[0198] - levels of neutrophil elastase, such as blood levels or plasma levels,

[0199] - levels of myeloperoxidase-DNA complexes, such as blood levels or plasma levels,

[0200] - levels of interleukin- 8, such as blood levels or plasma levels,

[0201] - levels of calprotectin, such as blood levels or plasma levels.

[0202]

[0127] The severe ARDS may be characterized by a dysregulation of NETosis, such as of at least one NETosis biomarker, characterized by an excessive amount of neutrophil extracellular traps (NETs) that can be measured by the levels of circulating DNA (cDNA) in comparison with the amount of neutrophil extracellular traps (NETs) in a subject of reference.

[0203]

[0128] This invention relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment of severe ARDS in a subject in need thereof, wherein the subject suffers from severe ARDS associated with at least one of the following NETosis biomarkers measured in a biological sample from a subject, in particular from the subject’s blood: - circulating DNA levels equal to or greater than 200 ng / mL, preferably equal to or greater than 300 ng / mL, more preferably equal to or greater than 400 ng / mL;

[0204] - citrullinated histone H3 levels equal to or greater than 200 ng / mL, preferably equal to or greater than 300 ng / mL, more preferably equal to or greater than 400 ng / mL, or citrullinated histone H3 levels equal to or greater than 0.4 (measured by an optical density (OD) - absorbance measured at 450 nm), preferably equal to or greater than 0.6, more preferably equal to or greater than 0.8;

[0205] - neutrophil elastase levels equal to or greater than 200 ng / mL, preferably equal to or greater than 300 ng / mL, more preferably equal to or greater than 400 ng / mL;

[0206] - interleukin-8 levels equal to or greater than 100 pg / mL, preferably equal to or greater than 200 pg / mL, more preferably equal to or greater than 300 pg / mL;

[0207] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, preferably equal to or greater than 300 ng / mL, more preferably equal to or greater than 400 ng / mL; and / or

[0208] - calprotectin levels equal to or greater than 5 pg / mL, preferably equal to or greater than 10 pg / mL, more preferably equal to or greater than 15 pg / mL.

[0209]

[0129] The level of the at least one NETosis biomarker can be measured by any method known by the skilled person in the art, such as ELISA test, optical density (such as absorbance measured at 450 nm), or any techniques allowing the measure of DNA or RNA molecule (such as PCR or RT-PCR).

[0210]

[0130] In some embodiments, the at least one NETosis biomarker is circulating DNA levels, such as blood levels, equal to or greater than 200 ng / mL, preferably equal to or greater than 300 ng / mL, more preferably equal to or greater than 400 ng / mL.

[0211]

[0131] In some embodiments, the NETosis biomarkers are circulating DNA levels equal to or greater than 200 ng / mL in combination with at least one further NETosis biomarker measured in a biological sample from a subject, selected from one of the following NETosis biomarkers:

[0212] - citrullinated histone H3 levels equal to or greater than 200 ng / mL or citrullinated histone H3 levels equal to or greater than 0.4 (measured by an optical density (OD - absorbance measured at 450 nm); - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0213] - interleukin-8 levels equal to or greater than 100 pg / mL;

[0214] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL; and / or

[0215] - calprotectin levels equal to or greater than 5 pg / mL.

[0216]

[0132] The levels of circulating DNA in a subject associated with severe ARDS may be any concentration equal to or greater than 200 ng / mL, 210 ng / mL, 220 ng / mL, 230 ng / mL, 240 ng / mL, 250 ng / mL, 260 ng / mL, 270 ng / mL, 280 ng / mL, 290 ng / mL, 300 ng / mL, 310 ng / mL, 320 ng / mL, 330 ng / mL, 340 ng / mL, 350 ng / mL, 360 ng / mL, 370 ng / mL, 380 ng / mL, 390 ng / mL, 400 ng / mL, 410 ng / mL, 420 ng / mL, 430 ng / mL, 440 ng / mL, 450 ng / mL, 460 ng / mL, 470 ng / mL, 480 ng / mL, 490 ng / mL, 500 ng / mL, 510 ng / mL, 520 ng / mL, 530 ng / mL, 540 ng / mL, 550 ng / mL, preferably equal to or greater than 280 ng / mL.

[0217]

[0133] The at least one NETosis biomarker may be the ratio of circulating DNA (in ng / mL) of the subject (cDNAsubject) compared with the concentration of circulating DNA measured in a healthy subject (cDNAheaithy). ). It can be blood concentration, such as plasmatic concentration.

[0218]

[0134] In some embodiments, sARDS is characterized by the subject having a cDNAsubject / cDNAheaithy ratio equal to or greater than 1,1 ; 1,2 ; 1,3 ; 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. In a preferred embodiment, sARDS is characterized by the subject having a cDNAsubject / cDNAheaithy ratio equal to or greater than 1,4.

[0219]

[0135] In some embodiments, the subject in need of treatment and / or prevention is characterized by a cDNAsubject / cDNAheaithy ratio equal to or greater than 1,1 ; 1,2 ; 1,3 ; 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. In a preferred embodiment, the subject in need of treatment and / or prevention is characterized by a cDNAsubject / cDNAheaithy ratio equal to or greater than 1,4.

[0220]

[0136] In some embodiments, the dysregulation of NETosis is characterized by the subject having a cDNAsubject / cDNAheaithy ratio equal to or greater than 1,1 ; 1,2 ; 1,3 ; 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ;

[0221] 3.2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. In a preferred embodiment, the dysregulation of NETosis is characterized by the subject having a cDNAsubject / cDNAheaithy ratio equal to or greater than 1,4.

[0222]

[0137] The levels of citrullinated histone H3 associated with severe ARDS may be any DO equal to or greater than 0.4 (absorbance measured at 450 nm) or concentration equal or greater than 200 ng / mL.

[0223]

[0138] In some embodiments, the at least one NETosis biomarker is citrullinated histone H3 (H3Cit) levels, such as blood levels, equal to or greater than 0.4 (absorbance measured at 450 nm) or concentration equal or greater than 200 ng / mL.

[0224]

[0139] Citrullinated histone may be detected in the blood in its complexed form with DNA: citrullinated histone H3-DNA. In some embodiments, the at least one NETosis biomarker is citrullinated histone H3-DNA levels equal to or greater than 150 ng / mL.

[0225]

[0140] The at least one NETosis biomarker may be the ratio of the citrullinated histone H3 levels (H3Cit), measured by the absorbance at 450 nm or ELISA kit (in ng / mL), in the biological sample of the subject (H3Citsubject), compared with the citrullinated histone H3 levels measured in a healthy subject (H3Citheaithy). It can be blood level, such as plasmatic level.

[0226]

[0141] In some embodiments, sARDS is characterized by the subject having a H3Cit subject / H3Cit healthy ratio equal to or greater than 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ;

[0227] 2.3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. In a preferred embodiment, sARDS is characterized by the subject having a H3Cit subject / H3Cit healthy ratio equal to or greater than 2 ; or having a H3Cit-DNAsubject / H3Cit- DNAheaithy ratio equal to or greater than 1,8.

[0228]

[0142] In some embodiments, the subject in need of treatment and / or prevention is characterized by a H3Cit subject / H3Cit healthy ratio equal to or greater than 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. In a preferred embodiment, the subject in need of treatment and / or prevention is characterized by a H3Citsubject / H3Citheaithy ratio equal to or greater than 2 ; or having a H3Cit-DNAsubject / H3Cit-DNAheaithy ratio equal to or greater than 1,8.

[0229]

[0143] In some embodiments, the dysregulation of NETosis is characterized by the subject having a H3Cit subject / H3Cit healthy ratio equal to or greater than 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. . In a preferred embodiment, the dysregulation of NETosis is characterized by the subject having a H3Citsubject / H3Citheaithy ratio equal to or greater than 2 ; or having a H3Cit-DNAsubject / H3Cit-DNAheaithy ratio equal to or greater than 1 ,8.

[0230]

[0144] The levels of neutrophil elastase associated with severe ARDS may be any concentration equal to or greater than 200 ng / mL.

[0231]

[0145] In some embodiments, the at least one NETosis biomarker is neutrophil elastase levels equal to or greater than 200 ng / mL, preferably equal to or greater than 250 ng / mL, more preferably equal to or greater than 300 ng / mL. It can be blood levels, such as plasmatic levels.

[0232]

[0146] The at least one NETosis biomarker may be the ratio of the neutrophil elastase levels (NE) in ng / mL, measured in the biological sample of the subject (NEsubject), compared with the levels of neutrophil elastase measured in a healthy subject (NEheaithy). The biological sample may be a blood sample such as a plasmatic sample.

[0233]

[0147] In some embodiments, sARDS is characterized by the subject having a NEsubject / NEheaithy ratio equal to or greater than 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5. In a preferred embodiment, sARDS is characterized by the subject having a NEsubject / NEheaithy ratio equal to or greater than 3,5.

[0234]

[0148] In some embodiments, the subject in need of treatment and / or prevention is characterized by a NEsubject / NEheaithy ratio equal to or greater than 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5. In a preferred embodiment, the subject in need of treatment and / or prevention is characterized by a NEsubject / NEheaithy ratio equal to or greater than 3,5.

[0149] In some embodiments, the dysregulation of NETosis is characterized by the subject having a NEsubject / NEheaithy ratio equal to or greater than 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5. In a preferred embodiment, the dysregulation of NETosis is characterized by the subject having a NEsubject / NEheaithy ratio equal to or greater than 3,5.

[0235]

[0150] The levels of interleukin-8 associated with severe ARDS may be any concentration equal to or greater than 100 pg / mL.

[0236]

[0151] In some embodiments, the at least one NETosis biomarker is interleukin-8 levels equal to or greater than 100 pg / mL, preferably equal to or greater than 150 pg / mL, more preferably equal to or greater than 200 pg / mL.

[0237]

[0152] The at least one NETosis biomarker may be the ratio of the levels of interleukin- 8 (in pg / mL), measured in the biological sample of the subject (IL-8subject), compared with levels of interleukin-8 measured in a healthy subject (IL-8heaithy). It can be blood levels, such as plasmatic levels.

[0238]

[0153] In some embodiments, sARDS is characterized by the subject having a IL-8subject / IL-8heaithy ratio equal to or greater than 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70. In a preferred embodiment, sARDS is characterized by the subject having a IL-8Subject / IL-8heaithy ratio equal to or greater than 30.

[0239]

[0154] In some embodiments, the subject in need of treatment and / or prevention is characterized by a IL-8Subject / IL-8heaithy ratio equal to or greater than 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70. In a preferred embodiment, the subject in need of treatment and / or prevention is characterized by a IL-8Subject / IL-8heaithy ratio equal to or greater than 30.

[0240]

[0155] In some embodiments, the dysregulation of NETosis is characterized by the subject having a IL- 8 subject / IL- 8heaithy ratio equal to or greater than 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70. In a preferred embodiment, the dysregulation of NETosis is characterized by the subject having a IL-8subject / IL-8heaithy ratio equal to or greater than 30.

[0241]

[0156] The levels of myeloperoxidase-DNA complexes associated with severe ARDS may be any concentration equal to or greater than 200 ng / mL.

[0242]

[0157] In some embodiments, the at least one NETosis biomarker is myeloperoxidase- DNA complexes levels equal to or greater than 200 ng / mL, preferably equal to or greater than 250 ng / mL, more preferably equal to or greater than 300 ng / mL.

[0243]

[0158] The at least one NETosis biomarker may be the ratio of the levels of myeloperoxidase-DNA complexes (in ng / mL) measured in the biological sample of the subject (MPOsubject), compared with the levels of myeloperoxidase-DNA complexes measured in a healthy subject (MPOheaithy). It can be blood levels, such as plasmatic levels.

[0244]

[0159] In some embodiments, sARDS is characterized by the subject having a MPOsubject / MPOheaithy ratio equal to or greater than 1,1 ; 1,2 ; 1,3 ; 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ;

[0245] 3,8 ; 3,9 ; or 4,0. In a preferred embodiment, sARDS is characterized by the subject having a MPOsubject / MPOheaithy ratio equal to or greater than 1,4.

[0246]

[0160] In some embodiments, the subject in need of treatment and / or prevention is characterized by a MPOsubject / MPOheaithy ratio equal to or greater than 1,1 ; 1,2 ; 1,3 ; 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ;

[0247] 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. In a preferred embodiment, the subject in need of treatment and / or prevention is characterized by a MPOsubject / MPOheaithy ratio equal to or greater than 1,4.

[0248]

[0161] In some embodiments, the dysregulation of NETosis is characterized by the subject having a MPOsubject / MPOheaithy ratio equal to or greater than 1,1 ; 1,2 ; 1,3 ; 1,4 ;

[0249] 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. In a preferred embodiment, the dysregulation of NETosis is characterized by the subject having a MPOsubject / MPOheaithy ratio equal to or greater than 1,4.

[0250]

[0162] The levels of calprotectin associated with severe ARDS may be any concentration equal to or greater than 5 pg / mL.

[0251]

[0163] In some embodiments, the at least one NETosis biomarker is calprotectin levels equal to or greater than 5 pg / mL, preferably greater than 7 pg / mL, more preferably greater than lOpg / mL.

[0252]

[0164] The at least one NETosis biomarker may be the ratio of the levels of calprotectin levels (CP) in pg / mL, measured in the biological sample of the subject (CPsubject), compared with the levels of calprotectin measured in a healthy subject (CPheaithy). It can be blood level, such as plasmatic level.

[0253]

[0165] In some embodiments, sARDS is characterized by the subject having CPsubject / CPheaithy ratio equal to or greater than 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 65, 70, 75. In a preferred embodiment, sARDS is characterized by the subject having CPsubject / CPheaithy ratio equal to or greater than 10.

[0254]

[0166] In some embodiments, the subject in need of treatment and / or prevention is characterized by a CPsubject / CPheaithy ratio equal to or greater than 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 65, 70, 75. In a preferred embodiment, the subject in need of treatment and / or prevention is characterized by a CPsubject / CPheaithy ratio equal to or greater than 10.

[0255]

[0167] In some embodiments, the dysregulation of NETosis is characterized by the subject having a CPsubject / CPheaithy ratio equal to or greater than 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 65, 70, 75. In a preferred embodiment, the dysregulation of NETosis is characterized by the subject having a CPsubject / CPheaithy ratio equal to or greater than 10.

[0256]

[0168] The CXCR4 inhibitor compound may be administered intravenously, intramuscularly, subcutaneously, parenterally, orally, nasally, ocularly, transmucosally, or transdermally.

[0257]

[0169] The CXCR4 inhibitor compound may be administered once, twice, three times a day.

[0258]

[0170] The CXCR4 inhibitor compound may be plerixafor administered by continuous intravenous infusion.

[0259]

[0171] The CXCR4 inhibitor compound may be plerixafor administered by continuous subcutaneous administration.

[0260]

[0172] Preferably, the CXCR4 inhibitor compound is administered by continuous intravenous or subcutaneous administration.

[0261]

[0173] The invention also relates to a method of prognostic of severe ARDS in a subject characterized by a dysregulation of NETosis in a subject, comprising the following steps: a) measuring the levels of at least one NETosis biomarker in a biological sample from a subject, in particular from the subject’s blood, from the subject’s plasma,

[0262] (b) comparing said NETosis biomarker with a subject of reference and / or threshold of NETosis,

[0263] (c) determining whether the subject is at risk of suffering from a severe ARDS depending on the levels of the NETosis biomarker in the subject, and

[0264] (d) optionally treating the subject with a CXCR4 inhibitor compound.

[0265]

[0174] The invention also relates to a method of prognostic of severe ARDS wherein the subject having sARDS has a PaCE / FiCE ratio inferior or equal to 200 mmHg, preferably inferior or equal to 100 mmHg, comprising the following steps: a) measuring the levels of at least one NETosis biomarker in a biological sample from a subject, in particular from the subject’s blood, from the subject’s plasma, (b) comparing said NETosis biomarker with a subject of reference and / or threshold of NETosis,

[0266] (c) determining whether the subject is at risk of suffering from a severe ARDS depending on the levels of the NETosis biomarker in the subject, and

[0267] (d) optionally treating the subject with a CXCR4 inhibitor compound.

[0268]

[0175] The NETosis biomarker measured in step (a) may be selected in the group comprising: plasmatic cDNA levels, citrullinated histone H3 levels, neutrophil elastase levels, interleukin- 8 levels, myeloperoxidase-DNA complexes levels, and / or calprotectin levels.

[0269]

[0176] The NETosis biomarker measured in the method of prognostic as described herein may be cDNA levels.

[0270]

[0177] In some embodiments, the step (b) of the prognostic method herein may be comparing the concentration of circulating DNA (in ng / mL) of the subject (cDNAsubject) with the concentration of circulating DNA measured in a healthy subject (cDNAheaithy). It can be blood concentration, such as plasmatic concentration.

[0271]

[0178] The method of prognostic of severe ARDS may be comparing the concentration of circulating DNA (in ng / mL) of the subject (cDNAsubject) with concentration of circulating DNA measured in a healthy subject (cDNAheaithy) in step (b); and determining in step (c) that the subject suffers from a severe ARDS when: the ratio cDNAsubject / cDNAheaithy is equal to or greater than 1,1 ; 1,2 ; 1,3 ; 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. In a preferred embodiment, step (c) is determining that the subject suffers from a severe ARDS when the ratio cDNAsubject / cDNAheaithy is equal to or greater than 1,4.

[0272]

[0179] For example, a healthy subject has an average plasmatic concentration of circulating DNA of 140 ng / mL.

[0273]

[0180] Step (c) of the method of prognostic of severe ARDS herein may be determining in step (c) that the subject suffers from a severe ARDS when: the ratio cDNAsubject / cDNAheaithy is equal to or greater than 1,1 ; 1,2 ; 1,3 ; 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0.

[0274]

[0181] Methods of quantifying cell-free DNA are known from the person skilled in the art. For example, plasmatic cDNA levels may be measured by PCR.

[0275]

[0182] The NETosis biomarker measured in the method of prognostic as described herein may be citrullinated histone H3 (H3Cit) levels.

[0276]

[0183] In some embodiments citrullinated histone may be detected in the blood in its form complexed with DNA: citrullinated histone H3-DNA. In some embodiments, the NETosis biomarker measured in the method of prognostic as described herein may be citrullinated histone H3-DNA levels.

[0277]

[0184] The method of prognostic of severe ARDS may be comparing citrullinated histone H3 levels (H3Cit), measured by the absorbance at 450 nm or ELISA kit (in ng / mL), in the biological sample of the subject (H3Citsubject) with levels of citrullinated histone H3 measured in a healthy subject (H3Citheaithy) in step (b); and determining in step (c) that the subject suffers from a severe ARDS when: the ratio H3Citsubject / H3Citheaithy is equal to or greater than 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0 . It can be blood level, such as plasmatic level.

[0278]

[0185] For example, a healthy subject has an average plasmatic concentration of H3Cit of 100 ng / mL or an optical density of 0.15 (absorbance at 450 nm).

[0279]

[0186] Step (b) of the method of prognostic herein may be comparing citrullinated histone H3 levels (H3Cit), measured by the absorbance at 450 nm or using an ELISA kit (ng / mL), in the biological sample of the subject (H3Cit subject) with levels of citrullinated histone H3 measured in a healthy subject (H3Cit healthy). It can be blood level, such as plasmatic level.

[0280]

[0187] Step (c) of the method of prognostic herein may be determining that the subject suffers from a severe ARDS when: the ratio H3Cit subject / H3Cit healthy is equal to or greater than 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0.

[0281]

[0188] In a preferred embodiment, step (c) is determining that the subject suffers from a severe ARDS when the ratio H3Citsubject / H3Citheaithy is equal to or greater than 2 ; or when the ratio H3Cit-DNAsubject / H3Cit-DNAheaithy is equal to or greater than 1,8.

[0282]

[0189] Methods of quantifying citrullinated histone H3 levels are known from the person skilled in the art. For example, plasmatic citrullinated histone H3 may be quantified with a modified ELISA, as described by Thalin et al., using a Cell Death detection kit without streptavidin-precoated wells. The optical densities (ODs) may be measured at a wavelength of 450nm with a reference correction wavelength at 620 nm, by using a microplate photometer.

[0283]

[0190] The NETosis biomarker measured in the method as described herein may be neutrophil elastase levels.

[0284]

[0191] The method of prognostic of severe ARDS may be comparing neutrophil elastase levels (NE) in ng / mL, in the biological sample of the subject (NEsubject) with levels of neutrophil elastase measured in a healthy subject (NEheaithy) in step (b); and determining in step (c) that the subject suffers from a severe ARDS when: the ratio NEsubject / NEheaithy is equal to or greater than 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5. It can be blood level, such as plasmatic level.

[0285]

[0192] For example, a healthy subject has an average plasmatic concentration of neutrophil elastase of 110 ng / mL.

[0286]

[0193] Step (b) of the method of prognostic herein may be comparing neutrophil elastase levels (NE) in ng / mL, in the biological sample of the subject (NEsubject) with levels of neutrophil elastase levels measured in a healthy subject (NEheaithy).

[0287]

[0194] Step (c) of the method of prognostic herein may be determining that the subject suffers from a severe ARDS when: the ratio NEsubject / NEheaithy is equal to or greater than 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5.

[0288]

[0195] In a preferred embodiment, step (c) is determining that the subject suffers from a severe ARDS when the ratio NEsubject / NEheaithy is equal to or greater than 3,5.

[0289]

[0196] Methods of quantifying neutrophil elastase levels are known from the person skilled in the art. For example, plasmatic neutrophil elastase may be quantified by ELISA Test.

[0290]

[0197] The NETosis biomarker measured in the method as described herein may be interleukin-8 levels.

[0291]

[0198] The method of prognostic of severe ARDS may be comparing the levels of interleukin-8 (in pg / mL) in the biological sample of the subject (IL-8subject) with levels of interleukin-8 measured in a healthy subject (IL-8heaithy) in step (b); and determining in step (c) that the subject suffers from a severe ARDS when: the ratio IL-8Subject / IL-8heaithy is equal to or greater than 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70. It can be blood level, such as plasmatic level.

[0292]

[0199] For example, a healthy subject has an average plasmatic concentration of interleukin- 8 of 3 pg / mL.

[0293]

[0200] Step (b) of the method of prognostic of severe ARDS herein may be comparing the levels of interleukin-8 (in pg / mL) in the biological sample of the subject (IL-8subject) with levels of interleukin measured in a healthy subject (IL-8heaithy).

[0294]

[0201] Step (c) of the method of prognostic of severe ARDS herein may be determining that the subject suffers from a severe ARDS when: the ratio IL-8Subject / IL-8heaithy is equal to or greater than 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70.

[0202] In a preferred embodiment, step (c) is determining that the subject suffers from a sever ARDS when the ratio IL-8subject / IL-8heaithy is equal to or greater than 30.

[0295]

[0203] Methods of quantifying IL-8 levels are known from the person skilled in the art. For example, plasmatic IL-8 levels may be measured by ELISA test.

[0296]

[0204] Myeloperoxidase-DNA complexes (MPO) levels is a known reliable marker of NETosis, that directly measures a process specific to NETosis.

[0297]

[0205] The method of prognostic of severe ARDS may be comparing the levels of myeloperoxidase-DNA complexes (in ng / mL) in the biological sample of the subject (MPOsubject) with levels of myeloperoxidase-DNA complexes measured in a healthy subject (MPOheaithy) in step (b); and determining in step (c) that the subject suffers from a severe ARDS when: the ratio MPOsubject / MPOheaithy is equal to or greater than 1,1 ; 1,2 ;

[0298] 1.3 ; 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ; 2,4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0. It can be blood level, such as plasmatic level.

[0299]

[0206] For example, a healthy subject has an average plasmatic concentration of MPO of 150 ng / mL.

[0300]

[0207] Step (b) of the method of prognostic of severe ARDS herein may be comparing the levels of myeloperoxidase-DNA complexes (in ng / mL) in the biological sample of the subject (MPOsubject) with levels of myeloperoxidase-DNA complexes measured in a healthy subject (MPOheaithy).

[0301]

[0208] Step (c) of the method of prognostic of severe ARDS herein may be determining that the subject suffers from a severe ARDS when: the ratio MPOsubject / MPOheaithy is equal to or greater than 1,1 ; 1,2 ; 1,3 ; 1,4 ; 1,5 ; 1,6 ; 1,7; 1,8 ; 1,9 ; 2,0 ; 2,1 ; 2,2 ; 2,3 ;

[0302] 2.4 ; 2,5 ; 2,6 ; 2,7 ; 2,8 ; 2,9 ; 3,0 ; 3,1 ; 3,2 ; 3,3 ; 3,4 ; 3,5 ; 3,6 ; 3,7 ; 3,8 ; 3,9 ; or 4,0.

[0303]

[0209] In a preferred embodiment, step (c) is determining that the subject suffers from a sever ARDS when the ratio MPOsubject / MPOheaithy is equal to or greater than 1,4.

[0210] Methods of quantifying myeloperoxidase-DNA complexes levels are known from the person skilled in the art. For example, plasmatic myeloperoxidase-DNA complexes may be quantified by enzyme-linked immunosorbent assay (ELISA) using a modified approach of the Cell Death Detection ELISA kit and the capture of antimyeloperoxidase antibody. A calibration range made from a stock solution of NETs may be used to measure myeloperoxidase-DNA complexes, with the results then been expressed as standard NETs.

[0304]

[0211] The method of prognostic of severe ARDS may be comparing calprotectin levels (CP) in pg / mL, in the biological sample of the subject (CP subject) with levels of calprotectin measured in a healthy subject (CPheaithy) in step (b); and determining in step (c) that the subject suffers from a severe ARDS when: the ratio CP subject / CPheaithy is equal to or greater than 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 65, 70, 75. It can be blood level, such as plasmatic level.

[0305]

[0212] For example, a healthy subject has an average plasmatic concentration of calprotectin between 0.1 pg / mL and 1 pg / mL.

[0306]

[0213] Step (b) of the method of prognostic of severe ARDS herein may be comparing calprotectin levels (CP) in pg / mL, in the biological sample of the subject (CPsubject) with levels of calprotectin measured in a healthy subject (CPheaithy).

[0307]

[0214] Step (c) of the method of prognostic of severe ARDS herein may be determining in step (c) that the subject suffers from a severe ARDS when the ratio CPsubject / CPheaithy is equal to or greater than 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 65, 70, 75.

[0308]

[0215] In a preferred embodiment, step (c) is determining that the subject suffers from a severe ARDS when the ratio CPsubject / CPheaithy is equal to or greater than 10.

[0216] Methods of quantifying calprotectin levels are known from the person skilled in the art. For example, plasmatic calprotectin levels may be quantified by ELISA test.

[0309]

[0217] In some embodiments, the step (c) of determining whether the subject is at risk of suffering from a severe ARDS depends on the levels of NETosis biomarker which is determined in view of the measured carried out at step (a) and on the comparison of step (b).

[0310]

[0218] In some embodiments, the step (c) of determining whether the subject is at risk of suffering from a severe ARDS depends on the levels of at least one of the following NETosis biomarker:

[0311] - circulating DNA levels equal to or greater than 200 ng / mL preferably equal to or greater than 300 ng / mL, more preferably equal to or greater than 400 ng / mL,

[0312] - citrullinated histone H3 levels equal to or greater than 200 ng / mL or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0313] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0314] - interleukin- 8 levels equal to or greater than 100 pg / mL,

[0315] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0316] - calprotectin plasma levels equal to or greater than 5 pg / mL.

[0317]

[0219] In some embodiments, the severity of ARDS is determined in step (c) as follows: the greater the increase of the measured value of said NETosis biomarker in the subject as compared to the value of said biomarker in the subject of reference in step (b), the higher is the severity of ARDS.

[0318]

[0220] In some embodiments, at step c) of the method, the subject is determined at low risk or at high risk of suffering from a severe ARDS depending on the levels of the NETosis biomarker in the subject.

[0319]

[0221] In some embodiments, the step (c) of determining whether the subject is at risk of suffering from a severe ARDS allows to stratify patient, i.e. determining what stage of ARDS he / she suffers from, especially whether he / she suffers from a severe ARDS.

[0222] In some embodiments, at step c) of the method, the subject is determined at low risk of suffering from a severe ARDS because the levels of the NETosis biomarker in the subject is / are:

[0320] - circulating DNA levels lower than 200 ng / mL,

[0321] - citrullinated histone H3 levels lower than 200 ng / mL or measured by an optical density (OD) lower than 0.4,

[0322] - neutrophil elastase levels lower than 200 ng / mL,

[0323] - interleukin-8 levels lower than 100 pg / mL,

[0324] - myeloperoxidase-DNA complexes levels lower than 200 ng / mL, and / or

[0325] - calprotectin plasma levels lower than 5 pg / mL.

[0326]

[0223] In some embodiments, at step c) of the method, the subject is determined at high risk of suffering from a severe ARDS because the levels of the NETosis biomarker in the subject is / are:

[0327] - circulating DNA levels equal to or greater than 200 ng / mL,

[0328] - citrullinated histone H3 levels equal to or greater than 200 ng / mL or measured by an optical density (OD) lower than 0.4,

[0329] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0330] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0331] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0332] - calprotectin plasma levels equal to or greater than 5 pg / mL.

[0333]

[0224] In some embodiments, when the subject is determined at low risk of suffering from a severe ARDS at step c), the subject is not treated with a CXCR4 inhibitor compound at step d).

[0334]

[0225] In some embodiments, when the subject is determined at high risk of suffering from a severe ARDS at step c), the subject is treated with a CXCR4 inhibitor compound at step d). The CXCR4 inhibitor compound is defined in the present disclosure. Especially, the CXCR4 inhibitor compound can be plerixafor.

[0335]

[0226] In the context of the method, the subject may be affected with ARDS, in particular with severe ARDS. The subject may be at risk of developing an ARDS. Examples of risk factors for developing ARDS include, but are not limited to, diabetes (such as, for example, type 2 diabetes mellitus (T2DM)), high blood pressure, obesity, pneumonia, aspiration of gastric contents, inhalation injury, pulmonary contusion, pulmonary vasculitis, and drowning.

[0336]

[0227] The CXR4 inhibitor compound used at step (d) for treating the subject is defined in the present disclosure herein above. The CXCR4 inhibitor compound can for example be plerixafor. All embodiments defined herein above regarding the CXR4 inhibitor compound, and / or the method of administration are applicable in the context of the prognostic method.

[0337]

[0228] The invention also relates to a method for treating severe ARDS in a subject, comprising the following steps:

[0338] (a) measuring the level, in a biological sample from a subject, of a NETosis biomarker,

[0339] (b) comparing said NETosis biomarker levels with the levels in a subject of reference or a threshold of NETosis,

[0340] (c) determining whether the subject suffers from a severe ARDS depending on the levels of NETosis biomarker, and

[0341] (d) treating the subject with a CXCR4 inhibitor compound.

[0342]

[0229] Step (a) of the method for treating severe ARDS may be measuring in a biological sample from a subject, in particular from the subject’s plasma, at least one of the following biomarkers of dysregulation of NETosis: circulating DNA levels equal to or greater than 200 ng / mL preferably equal to or greater than 300 ng / mL, more preferably equal to or greater than 400 ng / mL,

[0343] - citrullinated histone H3 levels equal to or greater than 200 ng / mL or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0344] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0345] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0346] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0347] - calprotectin plasma levels equal to or greater than 5 pg / mL.

[0230] The invention also relates to a method for prophylactically treating severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, comprising administering a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof to the subject, wherein the subject has a PaCL / FiCh ratio inferior or equal to 200 mmHg and / or wherein the subject is characterized by the presence of at least one NETosis biomarker in a biological sample selected from:

[0348] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0349] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0350] - citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,

[0351] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0352] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0353] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0354] - calprotectin levels equal to or greater than 5 pg / mL.

[0355]

[0231] The invention also relates to a method for treating severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, comprising administering a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof to the subject, wherein the subject has a PaCL / FiCh ratio inferior or equal to 100 mmHg and / or wherein the subject is characterized by the presence of at least one NETosis biomarker in a biological sample selected from:

[0356] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0357] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0358] - citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,

[0359] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0360] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0361] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or - calprotectin levels equal to or greater than 5 pg / mL.

[0362]

[0232] As disclosed herein, the subject in need thereof is a human patient susceptible to severe acute respiratory distress syndrome.

[0363]

[0233] As disclosed herein, the subject in need thereof is a human patient having a PaOi / FiOi ratio inferior or equal to 200 mmHg, preferably having a PaCWFiOi ratio inferior or equal to 100 mmHg.

[0364]

[0234] As disclosed herein, the subject in need thereof is a human patient characterized by the presence of at least one NETosis biomarker in a biological sample selected from:

[0365] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0366] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0367] - citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,

[0368] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0369] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0370] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0371] - calprotectin levels equal to or greater than 5 pg / mL.

[0372]

[0235] As disclosed herein, the subject in need thereof is a human patient characterized by the presence of at least one NETosis biomarker in a biological sample selected from:

[0373] - the ratio of circulating DNA levels in the subject compared to circulating DNA levels in a healthy individual is equal to or greater than 1,4;

[0374] - the ratio of citrullinated histone H3 levels in the subject compared to citrullinated histone H3 levels in a healthy individual is equal to or greater than 2 ;

[0375] - the ratio of citrullinated histone H3-DNA complexes levels in the subject compared to citrullinated histone H3-DNA complexes levels in a healthy individual is equal to or greater than 1,8; - the ratio of neutrophil elastase levels in the subject compared to neutrophil elastase levels in a healthy individual is equal to or greater than 3,5;

[0376] - the ratio of interleukin-8 levels in the subject compared to interleukin-8 levels in a healthy individual is equal to or greater than 30;

[0377] - the ratio of myeloperoxidase-DNA complexes levels in the subject compared to myeloperoxidase-DNA complexes levels in a healthy individual is equal to or greater than 1,4; and / or

[0378] - the ratio of calprotectin levels in the subject compared to calprotectin levels in a healthy individual is equal to or greater than 10.

[0379]

[0236] The invention also relates to the use of a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof in the manufacture of a medicament for treating severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the subject in need thereof has a PaCL / FiCh ratio inferior or equal to 100 mmHg and / or wherein the subject has at least one NETosis biomarker in a biological sample selected from:

[0380] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0381] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0382] - citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,

[0383] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0384] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0385] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0386] - calprotectin levels equal to or greater than 5 pg / mL.

[0387]

[0237] The invention also relates to the use of a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof in the manufacture of a medicament for prophylactically treating severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the subject in need thereof has a PaCL / FiCh ratio inferior or equal to 200 mmHg and / or wherein the subject has at least one NETosis biomarker in a biological sample selected from:

[0388] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0389] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0390] - citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,

[0391] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0392] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0393] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0394] - calprotectin levels equal to or greater than 5 pg / mL.

[0395]

[0238] The invention also relates to the use of a composition comprising a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, and a pharmaceutically acceptable excipient in the manufacture of a medicament for treating severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the subject in need thereof has a PaCL / FiCL ratio inferior or equal to 100 mmHg and / or wherein the subject has at least one NETosis biomarker in a biological sample selected from:

[0396] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0397] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0398] - citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,

[0399] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0400] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0401] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0402] - calprotectin levels equal to or greater than 5 pg / mL.

[0239] The invention also relates to the use of composition comprising a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof and a pharmaceutically acceptable excipient in the manufacture of a medicament for prophylactically treating severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the subject in need thereof has a PaC FiCh ratio inferior or equal to 200 mmHg and / or wherein the subject has at least one NETosis biomarker in a biological sample selected from:

[0403] - circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),

[0404] - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0405] - citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,

[0406] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0407] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0408] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0409] - calprotectin levels equal to or greater than 5 pg / mL.

[0410]

[0240] The invention also relates to a kit for prognostic of severe ARDS comprising the means for measuring in a biological sample from a subject, at least one of NETosis biomarkers selected from the list consisting of circulating DNA levels, citrullinated histone H3 levels, neutrophil elastase levels, interleukin-8 levels, myeloperoxidase-DNA, complexes levels, and calprotectin levels.

[0411]

[0241] The means of measuring in a biological sample from a subject, in particular from the subject’s plasma, may allow for the quantification of said at least one NETosis biomarker, wherein the value of the measured NETosis biomarker is indicative of the progression and severity of ARDS in the subject.

[0412]

[0242] The kit of prognostic may comprise an Enzyme-Linked ImmunoSorbent Assay

[0413] (ELISA) for the quantitative detection of at least one of said NETosis biomarker and: a. a set of one or more specific capture antibodies, each antibody being unique to at least one of said NETosis biomarkers associated with the severity and progression of ARDS, b. a detection antibody that binds specifically to at least one of said NETosis biomarkers, wherein the detection antibody may be labelled with an enzyme to produce a measurable signal upon the addition of an appropriate substrate, c. a standardized biomarker solution of known concentration to generate a calibration curve allowing for the quantification of the at least one of said NETosis biomarkers in a biological sample from a subject, d. a reagent for the enzyme- substrate reaction that yields a quantifiable product correlating with the at least one of said NETosis biomarker’s concentration, e. a series of buffers and solutions for the preparation of samples, washing, and reaction processes inherent to ELISA protocols, f. a control sample to ensure the accuracy and precision of the assay, and / or g. detailed instructions for sample preparation, reagent handling, and interpretation of results to accurately prognosticate severe ARDS.

[0414]

[0243] The invention also relates to the identification of early-stage biomarker that allows efficient prognostic of severe ARDS. These biomarkers have been selected as allowing the identification of severe ARDS in a patient at early stage especially when compared with classical marker of severe ARDS as disclosed in the Berlin Definition.

[0415]

[0244] Thus, the invention also relates on the use of at least one NETosis biomarker selected from circulating DNA, citrullinated histone H3, neutrophil elastase, interleukin- 8, myeloperoxidase-DNA complexes, and calprotectin as a biomarker for in vitro detection of severe ARDS in a biological sample. As comprised herein, citrullinated histone H3 may be measured in its complexed form with DNA: citrullinated histone H3- DNA.

[0416]

[0245] In some embodiments, the at least one biomarker is selected from circulating DNA, citrullinated histone H3 and neutrophil elastase.

[0246] In some embodiments, the invention comprises the measure of the level of the two biomarkers circulating DNA and citrullinated histone H3.

[0417]

[0247] In some embodiments, the invention comprises the measure of the level of the three biomarkers circulating DNA, citrullinated histone H3 and neutrophil elastase.

[0418]

[0248] In some embodiments, the invention comprises the measure of the level of the four biomarkers circulating DNA, citrullinated histone H3, neutrophil elastase and interleukin- 8.

[0419]

[0249] In some embodiments, the invention comprises the measure of the level of the five biomarkers circulating DNA, citrullinated histone H3, neutrophil elastase, interleukin-8 and myeloperoxidase-DNA complexes.

[0420]

[0250] In some embodiments, the invention comprises the measure of the level of the six biomarkers circulating DNA, citrullinated histone H3, neutrophil elastase, interleukin- 8, myeloperoxidase-DNA complexes and calprotectin.

[0421]

[0251] In some embodiments, the invention comprises the measure of the level of the seven biomarkers circulating DNA, citrullinated histone H3, citrullinated histone H3- DNA, neutrophil elastase, interleukin- 8, myeloperoxidase-DNA complexes and calprotectin.

[0422]

[0252] This invention relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment of a severe acute respiratory distress syndrome (sARDS), wherein the sARDS is characterized by a dysregulation of NETosis.

[0423]

[0253] The present invention further relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment of severe ARDS in a subject in need thereof, wherein the severe ARDS is associated with at least one of the following NETosis biomarkers measured in a biological sample from a subject:

[0424] - circulating DNA levels equal to or greater than 200 ng / mL, - citrullinated histone H3 levels equal to or greater than 200 ng / mL,

[0425] - neutrophil elastase levels equal to or greater than 200 ng / mL,

[0426] - interleukin-8 levels equal to or greater than 100 pg / mL,

[0427] - myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or

[0428] - calprotectin levels equal to or greater than 5 pg / mL.

[0429]

[0254] In some embodiments, the CXCR4 inhibitor compound is selected from the list consisting of plerixafor, LY2510924, AMD3465, MSX-122, Mavorixafor and a mixture thereof.

[0430]

[0255] In some embodiments, the compound is of formula (I), and formula (I) is:

[0431] - Z and Y, which are identical or different, represent: o an alkyl group having from 1 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms, or o an aryl or heteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O- alkyl group having from 1 to 6 carbon atoms, or o a cyclic or heterocyclic polyamine group having from 9 to 32 ring members and from 3 to 8 amine groups in the ring spaced by 2 or more carbon atoms from each other, optionally comprising an aryl or heteroaryl group having from 3 to 6 carbon atoms, a heteroatom, and optionally substituted by a halogen atom, a hydroxyl group, a heteroatom, an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, or o a D1D2N- group, wherein DI and D2, which may be identical or different, represent: • a hydrogen atom, or

[0432] • an alkyl group having from 1 to 6 carbon atoms optionally substituted by at least one hydroxyl group, a halogen atom, a CF3 group, a CN group, an amine group, or an alkyl, O-alkyl or S-alkyl group having from 1 to 12 carbon atoms, or

[0433] • an aryl, an heteroaryl, a cycloalkyl, a heterocycloalkyl, an alkylaryl, an alkylheteroaryl or an alkylheteropoly aryl group having from 3 to 12 carbon atoms, optionally substituted by at least one hydroxyl group, halogen atom, CF3 group, CN group, amine group, or alkyl, O-alkyl or S-alkyl group having from 1 to 12 carbon atoms, or

[0434] • DI and D2 are linked together to from a N-containing aryl or heteroaryl group having from 3 to 12 carbon atoms and optionally substituted by at least one amine group optionally substituted by an alkylheteroaryl group having from 3 to 12 carbon atoms, and

[0435] - A represents: o an aryl or heteroaryl group having from 3 to 12 carbon atoms, or o an alkyl group having from 1 to 6 carbon atoms, or o -R4-Y’-R5-, wherein R4 and R5 which are identical or different represent an alkyl group having from 1 to 6 carbon atoms and Y’ represents an aryl or heteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a halogen atom, a hydroxyl group, an amide group, an amine group, an alkoxy group, an ester group, a CF3 group, a CN group or an alkyl, O-alkyl or S-alkyl group having from 1 to 6 carbon atoms optionally substituted by a hydroxyl group, an amine group or an O-alkyl group having from 1 to 6 carbon atoms, and

[0436] - R and R’, which are identical or different, represent an alkyl group having from 1 to 6 carbon atoms or a R1NR2R3 group or a single bond, and

[0437] - R1 represents a single bond or an alkyl group having from 1 to 6 carbon atoms, and

[0438] - R2 and R3, which are identical or different, represent a hydrogen atom, an amine group, an alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, wherein said aryl or heteroaryl group having from 3 to 6 carbon atoms is optionally substituted by an alkyl group having from 1 to 6 carbon atoms.

[0256] In some embodiments, the CXCR4 inhibitor compound is plerixafor, LY2510924, AMD3465, MSX-122, mavorixafor and a mixture thereof.

[0439]

[0257] The present invention also relates to a method of prognostic of severe ARDS characterized by a dysregulation of NETosis in a subject, comprising the following steps:

[0440] (a) measuring the levels of a NETosis biomarker in a biological sample from a subject, in particular from the subject’s plasma,

[0441] (b) comparing said NETosis biomarker with a subject of reference and / or threshold of NETosis,

[0442] (c) determining whether the subject is at risk of suffering from a severe ARDS depending on the levels of NETosis biomarker, and

[0443] (d) optionally treating the subject with a CXCR4 inhibitor compound.

[0444]

[0258] In some embodiments, the CXCR4 inhibitor compound is selected from the list consisting of plerixafor, LY2510924, AMD3465, MSX-122, mavorixafor and a mixture thereof.

[0445]

[0259] In some embodiments, the NETosis biomarker measured in a biological sample from the subject is circulating DNA levels, citrullinated histone H3 levels, interleukin-8 levels, myeloperoxidase-DNA complexes levels, neutrophil elastase levels and / or calprotectin levels.

[0446]

[0260] In some embodiments, step (d) is the administration of a CXCR4 inhibitor compound to the patient with severe ARDS via one of the following routes: intravenous, intramuscular, subcutaneous, parenteral, oral, nasal, ocular, transmucosal, or transdermal.

[0447]

[0261] In some embodiments, step (d) corresponds to the continuous intravenous administration of a CXCR4 inhibitor compound to the patient with severe ARDS.

[0448]

[0262] The present invention further relates to a pharmaceutical composition for use in the treatment of severe acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the severe ARDS is characterized by a dysregulation of NETosis, and wherein said pharmaceutical composition comprises a CXCR4 inhibitor compound and a pharmaceutically acceptable excipient.

[0263] In some embodiments, severe ARDS results from a viral, a fungal or a bacterial infection.

[0449]

[0264] The present invention also relates to a kit for prognostic of a severe ARDS comprising the means for measuring in a biological sample from a subject at least one of NETosis biomarkers selected from the list consisting of circulating DNA levels, citrullinated histone H3 levels, neutrophil elastase levels, interleukin-8 levels, myeloperoxidase-DNA complexes levels, and calprotectin levels.

[0450]

[0265] The present invention further relates to the use of at least one NETosis biomarker selected from circulating DNA, citrullinated histone H3, neutrophil elastase, interleukin- 8, myeloperoxidase-DNA complexes, and calprotectin as a biomarker for in vitro detection of severe ARDS in a biological sample, preferably the at least one biomarker is selected from circulating DNA, citrullinated histone H3 and neutrophil elastase.

[0451]

[0266] The present invention also relates to a CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment of a disease in a subject in need thereof, wherein the disease is characterized by a dysregulation of NETosis, in particular by high levels of NETosis in comparison with a subject of reference.

[0452] BRIEF DESCRIPTION OF THE DRAWINGS

[0453]

[0267] Figures 1A-C: are graphs showing the association between NETosis levels and the severity of ARDS, modeled in a murine model of sepsis induced by intraperitoneal (IP) administration. Fig. 1A shows the percentage survival of mice over 7 days; n=12 per group at the start of the study; *p<0.05, compared to IM group. Fig. IB shows the quantification of plasmatic DNA levels (pg / mL) of non-induced mice (IM) or induced mice (2M and 3M) on days 1 and 3. Mean ± SEM; n=4 per group; *p<0.05, compared to IM group and #p<0.05, compared to 2M group. Fig. 1C shows the quantification of plasmatic CXCL1 / KC levels (pg / mL) of non-induced mice (IM) or induced mice (2M and 3M) on days 1 and 3. Mean ± SEM; n=4 per group; *p<0.05 and ****p<0.0001, compared to IM group and ##p<0.01, compared to 2M group.

[0454]

[0268] Figure 2: is graph showing the effect of plerixafor on severe ARDS modeled in mice by IP administration of LPS at 10 mg / kg. This graph shows the percentage survival of mice over 7 days; n=5 per group at the start of the study (n=4 for IM group).

[0455]

[0269] Figures 3A-D: are graphs showing the association between NETosis process and the severity of ARDS, modeled in a murine model of sepsis induced by intratracheal (IT) administration of LPS. Fig. 3A shows the percentage survival of mice over 7 days; n=7 per group at the start of the study (n=5 for IM group); **p<0.01, compared to IM group and ##p<0.01, compared to 2M group. Fig. 3B shows oxygen saturation in mice, as a percentage of day 0. Mean ± SEM; n=10 per group; **p<0.01 and ***p<0.001, compared to IM group. Fig. 3C shows the quantification of Neutrophil Elastase levels (pg / mL / mg of tissues) in lung homogenates of non-induced mice (IM) or induced mice (2M and 3M). Mean ± SEM; n=7 per group (n=5 for IM group); ****p<0.0001, compared to 1 M group and ####p<0.0001, compared to 2M group. Fig. 3D shows the quantification of Citrullinated Histone H3 levels (ng / mL / mg of tissues) in lung homogenates of noninduced mice (IM) or induced mice (2M and 3M). Mean ± SEM; n=7 per group (n=5 for IM group); ***p<0.001, compared to IM group and ###p<0.001, compared to 2M group.

[0456]

[0270] Figures 4A-B: are graphs showing the effect of plerixafor on severe ARDS modeled in mice by IT administration of LPS at 10 mg / kg. Fig. 4A shows the percentage survival of mice over 7 days; n=10 per group at the start of the study; ***p<0.001, compared to IM group and ###p<0.001, compared to 2M group. Fig. 4B shows oxygen saturation in mice on day 3, as a percentage of day 0. Mean ± SEM; n=10 per group; ***p<0.001, compared to IM group and #p<0.05, compared to 2M group.

[0457]

[0271] Figures 5A-F: are graphs showing the association between NETosis levels and the severity of ARDS, in a murine model of sepsis induced by cecal ligation and puncture. Fig. 5A shows the percentage survival of mice over 7 days; n=13-15 per group at the start of the study; ***p<0.001, compared to IM group and #p<0.05, compared to 2M group. Fig. 5B shows the quantification of Citrullinated Histone H3 levels (ng / mL / mg of tissues) in lung homogenates of non-induced mice (IM) or induced mice (2M and 3M). Mean ± SEM; n=5; *p<0.05, compared to IM group. Fig. 5C shows the quantification of plasmatic Myeloperoxidase levels (pg / mL) of non-induced mice (IM) or induced mice (2M and 3M). Mean ± SEM; n=5; **p<0.01, ***p<0.001 compared to IM group and #p<0.05, compared to 2M group. Fig. 5D shows the quantification of plasmatic CXCL1- KC levels (pg / mL) of non-induced mice (IM) or induced mice (2M and 3M). Mean ± SEM; n=5; *p<0.05, compared to IM group. Fig. 5E shows the quantification of plasmatic Neutrophil Elastase (pg / mL) levels of non-induced mice (IM) or induced mice (2M and 3M). Mean ± SEM; n=5; **p<0.01 and ***p<0.001, compared to IM group. Fig. 5F shows the quantification of plasmatic DNA levels (pg / mL) of non-induced mice (IM) or induced mice (2M and 3M). Mean ± SEM; n=5; **p<0.01, compared to IM group and #p<0.05, compared to 2M group.

[0458]

[0272] Figures 6A-B are graphs showing the effect of plerixafor on ARDS severities, modeled in a murine model of sepsis induced by cecal ligation and puncture. Fig. 6A shows the percentage survival of mice over 7 days in the model of severe ARDS; n=8 per group at the start of the study; *p<0.05 and ***p<0.001, compared to IM group and #p<0.05, compared to 2M group. Fig. 6B shows the quantification of plasmatic DNA levels (pg / mL) in the moderate and severe models of ARDS, n=8 per group. Mean ± SEM; ***p<0.001, compared to IM group and #p<0.05, compared to 4M group.

[0459]

[0273] Figures 7A-B are graphs showing the effect of different CXCR4 antagonists on severe ARDS, in a murine model of sepsis induced by cecal ligation and puncture. Fig. 7A shows the quantification of plasmatic Myeloperoxidase levels (pg / mL) of mice. Mean ± SEM; n=8; **p<0.01, compared to IM group and #p<0.05, compared to 2M group. Fig. 7B shows the quantification of plasmatic DNA levels (pg / mL) of mice. Mean ± SEM; n=8; ***p<0.001, compared to IM group and #p<0.05, compared to 2M group. EXAMPLES

[0460]

[0274] The present invention is further illustrated by the following examples.

[0461] EXAMPLE 1: ARDS severity induced by intraperitoneal route is associated to NETosis

[0275] Three groups of C57B1 / 6 mice were allocated for this study (see Table 1). The model of ARDS was induced by intraperitoneal (IP) administration of Lipopolysaccharide (LPS) using two doses to model varying levels of severity. On day 0, 24 mice were induced by IP administration of PBS containing 5 or 10 mg / kg of LPS (groups 2M and 3M, respectively). 12 mice received only PBS in the same way as groups 2M and 3M and served as non-induced control group (sham, group IM).

[0462]

[0276] Table 1: Study design of example 1.

[0463]

[0277] During the study, mortality and morbidity observations, clinical score and body weight measurements were performed on 4 mice from each group. Mice reaching a body weight loss superior to 20% of their initial body weight and / or reaching humane endpoints were euthanized and the time of death was precisely recorded. This enabled to establish a survival curve which was represented in the form of the Kaplan Meier estimator. As shown in Figure 1A, 75% of mice induced with the dose of 10 mg / kg LPS were euthanized at day 3 (p<0.05) while only 25% of mice were euthanized at day 2 for mice induced with the dose of 5 mg / kg LPS. All mice from the non-induced control group survived up to study termination (day 7).

[0464]

[0278] Additionally, 4 mice for each group were euthanized on days 1 and 3, plasma was collected for NETosis analysis. Concentrations of plasmatic CXCL1 / KC (equivalent to the human IL- 8 cytokine) and DNA were quantified. As shown in Figure IB, on day 1, plasmatic DNA levels were only detected at low concentrations in non-induced mice (0.274 ± 0.003 pg / mL) and those treated with the dose of 5 mg / kg LPS (0.359 ± 0.016 pg / mL). However, on day 1, LPS at 10 mg / kg significantly increased the production of plasmatic DNA compared to sham control group (0.746 ± 0.146 pg / mL; p<0.05). In the same way, as shown in Figure 1C, CXCL1 / KC levels were detected at low concentration in plasma of non-induced mice (79.3 ± 11.2 pg / mL). LPS at 5 or 10 mg / kg significantly modulate the production of plasmatic CXCL1 / KC in a dose-dependent manner (1 342.7 ± 185.0 pg / mL; p<0.05 for 5 mg / kg LPS and 3 332.3 ± 440.5 pg / mL; p<0.0001 for 10 mg / kg LPS) compared to sham control. It is important to note that on day 3, mice in the 3M group recovered plasma DNA (0.334 ± 0.033 pg / mL) and mice recovered plasma CXCL1-KC in the 2M (92.3 ± 20.0 pg / mL) and 3M group (81.5 ± 19.0 pg / mL) concentrations similar to those in the control group (103.3 ± 16.2 pg / mL).

[0465]

[0279] In conclusion, as shown in Figure 1, ARDS severity, assessed by survival (Figure 1A), is well correlated with an increase in NETosis, assessed by plasmatic DNA (Figure IB) and plasmatic CXCL1 / KC (Figure 1C) quantification. In addition, NETosis has been shown to be an early process, quantifiable on day 1.

[0466] EXAMPLE 2: Plerixafor is effective in severe ARDS model induced by intraperitoneal route

[0467]

[0280] Three groups of C57B1 / 6 mice were allocated for this study (see Table 2). PBS containing 10 mg / kg LPS (groups 2M and 3M) were intraperitoneally administered to 10 mice on day 0 to induce severe ARDS model. 4 mice received only PBS in the same way as groups 2M and 3M and served as non-induced control group (sham, group IM). 4 hours after LPS induction and then twice a day until termination, mice were subcutaneously (SC) treated with water as vehicle (groups IM and 2M) or plerixafor at 20 mg / kg / day (group 3M).

[0281] Table 2: Study design of example 2.

[0468]

[0282] During the study, mortality and morbidity observations, clinical score and body weight measurements were performed. Mice reaching a body weight loss superior to 20% of their initial body weight and / or reaching humane endpoints were euthanized and the time of death was precisely recorded. This enabled to establish a survival curve which was represented in the form of the Kaplan Meier estimator. As shown in Figure 2, plerixafor significantly enhances the survival of mice administered with 10 mg / kg of LPS by completely abolished mortality compared to the 2M group (which induced 50% of mortality). All mice from the non-induced control group (IM) survived up to study termination (day 7).

[0283] In conclusion, as shown in Figure 2, plerixafor is effective in a severe ARDS model (10 mg / kg LPS), as assessed by survival. In fact, plerixafor enhances survival with severe dose of LPS (10 mg / kg) administered intraperitoneally. As shown in Example 1, ARDS severity is linked to high level of NETosis. We can therefore suggest, based on plerixafor’ s mechanism of action, that its efficacy on severe ARDS is related to their high level of NETosis.

[0469] EXAMPLE 3: ARDS severity induced by intratracheal route is associated to NETosis

[0470]

[0284] Three groups of C57B1 / 6 mice were allocated for this study (see Table 3). The model of ARDS was induced by intratracheal (IT) administration of Lipopolysaccharide (LPS) using two doses to model varying levels of severity. On day 0, 14-20 mice were induced by IT administration of PBS containing 5 or 10 mg / kg of LPS (groups 2M and 3M, respectively). 5-10 mice received only PBS in the same way as groups 2M and 3M and served as non-induced control group (sham, group IM).

[0471]

[0285] Table 3: Study design of example 3.

[0472]

[0286] During the study, mortality and morbidity observations, clinical score and body weight measurements were performed. Mice reaching a body weight loss superior to 20% of their initial body weight and / or reaching humane endpoints were euthanized and the time of death was precisely recorded. This enabled to establish a survival curve which was represented in the form of the Kaplan Meier estimator. As shown in Figure 3A, mice induced with the dose of 10 mg / kg LPS were all euthanized at day 5 (p<0.01) while only one mouse was euthanized at day 4 for the mice induced with the dose of 5 mg / kg LPS. All mice from the non-induced control group survived up to study termination (day 7).

[0473]

[0287] Blood oxygen saturation was monitored on day 3, using the "Mouse OxPlus" system. As shown in Figure 3B, LPS administration induced a significant decrease in oxygen saturation in groups 2M (90.6 ± 1.7%; p<0.01) and 3M (87.2 ± 2.8%; p<0.001) compared to sham control (99.5 ± 1.2%).

[0474]

[0288] Additionally, when mice were euthanized for ethical reason or at termination at day 7, lungs were collected for NETosis analysis. Concentrations of Citrullinated Histone H3 (H3Cit) and Neutrophil Elastase (NE) were evaluated in lung homogenates by ELISA. As shown in Figure 3C, NE levels were detected at low concentration in non-induced mice (23 874.5 ± 6 146.4 pg / mL / mg of tissues). LPS at 5 mg / kg didn’t significantly modulate the production of NE (33 361.5 ± 8 610.5 pg / mL / mg of tissues) compared to sham control while 10 mg / kg LPS significantly increased its production in lungs (130 251.4 ± 17 252.2 pg / mL / mg of tissues; p<0.0001). In the same way, as shown in Figure 3D, H3Cit levels were only detected at low concentrations in non-induced mice (0.028 ± 0.0005 ng / mL / mg of tissues) and those treated with the dose of 5 mg / kg LPS (0.040 ± 0.0046 ng / mL / mg of tissues). However, LPS at 10 mg / kg significantly increased the production of H3Cit in lung homogenates compared to sham control group (0.202 ± 0.0415 ng / mL / mg of tissues; p<0.001).

[0475]

[0289] In conclusion, as shown in Figure 3, the ARDS severity, as assessed by survival (Figure 3A) and oxygen saturation (Figure 3B), is well correlated with an increase in NETosis, assessed by NE (Figure 3C) and H3Cit (Figure 3D) quantification in lung homogenates.

[0476] EXAMPLE 4: Plerixafor is effective in severe ARDS model induced by intratracheal route

[0477]

[0290] Three groups of C57B1 / 6 mice were allocated for this study (see Table 4). 50 pL of PBS containing 10 mg / kg LPS (groups 2M and 3M) were intratracheally administered to 20 mice on day 0 to induce severe ARDS model. 10 mice received only PBS in the same way as groups 2M and 3M and served as non-induced control group (sham, group IM). 4 hours after LPS induction and then twice a day until termination, mice were subcutaneously (SC) treated with water as vehicle (groups IM and 2M) or plerixafor at 20 mg / kg / day (group 3M).

[0478]

[0291] Table 4: Study design of example 4.

[0479]

[0292] During the study, mortality and morbidity observations, clinical score and body weight measurements were performed. Mice reaching a body weight loss superior to 20% of their initial body weight and / or reaching humane endpoints were euthanized and the time of death was precisely recorded. This enabled to establish a survival curve which was represented in the form of the Kaplan Meier estimator. As shown in Figure 4A, plerixafor significantly enhances the survival of mice administered with 10 mg / kg of LPS compared to the 2M group. All mice from the non-induced control group (IM) survived up to study termination (day 7).

[0293] Blood oxygen saturation was monitored on day 3 using the "Mouse OxPlus" system. As shown in Figure 4B, plerixafor significantly enhances oxygen saturation (p<0.05) in mice administered 10 mg / kg of LPS (94.9 ± 1.7%) compared to the 2M group (87.2 ± 2.8%). Mice from the non-induced group (sham group) have an oxygen saturation of 99.5 + 1.2%.

[0480]

[0294] In conclusion, as shown in Figure 4, the plerixafor is effective in a severe ARDS model (10 mg / kg LPS), as assessed by survival (Figure 4A) and oxygen saturation (Figure 4B). In fact, plerixafor enhances survival and oxygen saturation with severe dose of LPS (10 mg / kg) administered intratracheally. As shown in Example 3, ARDS severity is linked to high level of NETosis. We can therefore suggest, based on plerixafor’ s mechanism of action, that its efficacy on severe ARDS is related to their high level of NETosis.

[0481] EXAMPLE 5: Clinical trial CHOPIN (Cxcr4Hi neutrOPhils in InflueNza), study of the role of CXCR4-expressing neutrophils in lung injury during severe influenza infection.

[0482]

[0295] CHOPIN is a prospective cohort study, etiological, pathophysiological, and multicentric, conducted by the University hospital of Bordeaux (CHU de Bordeaux), the University hospital of Tours (CHU de Tours) and the University hospital of Lille (CHU de Lille).

[0483]

[0296] The aim of this study is to compare the percentage of blood CXCR4-expressing neutrophils between influenza survivors (i.e. patients who have survived at day 28 after been admitted to the intensive care unit (ICU)) and non-survivors (i.e. patients who are dead at day 28). Percentages of CXCR4-expressing neutrophils in bronchoalveolar lavage (BAL) or tracheal aspirates (TA) are also compared. To this aim, blood and BAL / TA sampling neutrophils isolation, immuno staining and flow cytometry acquisition are performed within the 24 hours and at day 3 after admission to ICU. Mortality rate is measured at day 28 and day 90.

[0484] Design of the study

[0485]

[0297] Experiments are conducted separately in 3 groups of patients: Group of interest: patients admitted to ICU for Acute respiratory Distress

[0486] Syndrome (ARDS) caused by severe influenza infection (n=30)

[0487] Two control groups:

[0488] (i) Control group 1 : ARDS caused by lung infection by bacteria (n=30)

[0489] (ii) Control group 2: ARDS caused by inflammation of extra-pulmonary origin (n=15).

[0490]

[0298] Blood sampling and BAL and TA are performed within the 24 hours and at day 3 after admission to ICU.

[0491]

[0299] The blood samples represent a total of 30mL at each time, the bronchoalveolar lavage (BAL) a volume of 50mL at each time, and the tracheal aspirates (TA) a volume of approximately 5mL. These samples are centrifuged and the supernatant is kept for global analysis at the end of the study without there being any biological collection beyond the duration of the study.

[0492]

[0300] The immune cells from the biological fluid samples (blood or BAL / TA sample) are identified by CD45 labeling, followed by neutrophils by CDl lb+, CD15+, CD14" labeling. CXCR4 labeling is used to measure the proportion of neutrophils expressing this molecule, as well as the mean fluorescence intensity.

[0493]

[0301] The samples are run on a flow cytometer within 3 days of labeling to maintain good fluorescence quality (Fortessa X20 cytometer). The proportion of neutrophils expressing CXCR4 and mean fluorescence intensity of CXCR4 on neutrophils are analyzed using a flow cytometry software.

[0494] Statistical analysis:

[0495]

[0302] Continuous variables are expressed as median and interquartile range and compared between survivors and non-survivors by the Wilcoxon-Mann- Whitney test assuming a non-normal distribution. All comparisons are performed with a two-sided type I error risk of 5% (Prism software n°9.0). Table 5: Participant follow-up summary table

[0496]

[0303] Clinical examination taking a history of illness, treatments, occupational and smoking exposure, postal code of place of residence, allergies, weight, height, vital parameters, cardiopulmonary inspection and auscultation, complete neurological examination, palpation, percussion and abdominal palpation, palpation of lymph nodes, inspection and palpation of skin, inspection and palpation of bone framework.

[0497]

[0304] Biological assessment2: biological tests are carried out as part of the treatment.

[0498] They usually include, within 24 hours of admission, a blood count, a blood ionogram, a liver function test, an assessment of renal function, a coagulation test, arterial blood gases to calculate the PaC FiCh ratio with lactate determination, and microbiological samples (blood cultures, tracheal aspiration, legionella and pneumococcal antigenuria).

[0499]

[0305] Paraclinical tests3: whether or not to perform a chest X-ray or CT scan is left to the discretion of the practitioner in charge of the patient.

[0500]

[0306] Mortality at D28 and D90 is determined either by knowledge of death before discharge from intensive care, or by knowledge of the patient's hospitalization status in a department other than intensive care, or by telephone call from the patient's attending physician. Calls from the patient or his or her family is only be made as a last resort.

[0501] Objectives

[0502] Main

[0503]

[0307] The main objective of the study is to compare the rate of CXCR4-expressing polynuclear neutrophils measured in the blood of patients within 24 hours of their admission to the ICU in the patients who have survived at day 28 after been admitted to the ICU with said rate in the patients who are dead at day 28.

[0504] Secondary objectives

[0505]

[0308] The secondary objectives are as follows: to compare the rate of CXCR4-expressing polynuclear neutrophils measured in the pulmonary compartment of patients within 24 hours of their admission to the ICU, in the patients who have survived at day 28 after been admitted in ICU with said rate in the patients who are dead at day 28. to compare the rate of CXCR4-expressing polynuclear neutrophils measured in the blood of patients on day 3 after their admission to the ICU, in the patients who have survived at day 28 after been admitted to the ICU with said rate in the patients who are dead at day 28. to compare the rate of CXCR4-expressing polynuclear neutrophils measured in the pulmonary compartment of patients on day 3 after their admission to the ICU, in the patients who have survived at day 28 after been admitted to the ICU with said rate in the patients who are dead at day 28. to compare the mean fluorescence intensity of CXCR4 in polynuclear neutrophils measured by flow cytometry in blood samples and pulmonary compartment samples taken from patients within 24 hours of admission to the ICU and on day 3 after their admission to the ICU, in the patients who have survived at day 28 after been admitted to the ICU with said fluorescence intensity in the patients who are dead at day 28. to compare the clusterin and C5 complement levels measured in the plasma of patients within 24h and on day 3 after admission to the ICU, in the patients who have survived at day 28 after been admitted to the ICU with said rate in the patients who are dead at day 28.

[0506]

[0309] The same comparisons are made between the patients who have survived at day 90 after been admitted to the ICU with said rate in the patients who are dead at day 90, and according to their ratio PaCL / FiCh (indicating the initial severity of the lung damage).

[0507] Additional sample analysis

[0508]

[0310] The supernatants from blood, BAL samples and TA samples taken within 24 hours and on the 3rd day after admission to intensive care are kept at -80°C until the end of the study for further analysis.

[0509]

[0311] Using these samples, assays are conducted at the end of the study for measuring levels of:

[0510] - CXCL-12, CXCL-17, a panel of pro-inflammatory interleukins and chemokines, a panel of anti-inflammatory interleukins and chemokines, Neutrophil Extracellular Traps: Myeloperoxydase-DNA (MPO-DNA) complex, circulating DNA, citrullinated histone H3, neutrophil elastase and calprotectin markers of complement and clusterin Parameters measured

[0511]

[0312] The main parameters measured are: percentage of CXCR4-expressing polynuclear neutrophils compared to the total number of polynuclear neutrophils measured by flow cytometry in the blood of patients within 24 hours after admission to the ICU. death rate at day 28 after admission in the ICU.

[0512]

[0313] These parameters are measured separately in the groups of patients with severe influenza infection, infection by bacteria and inflammation of extra-pulmonary origin.

[0513]

[0314] The secondary parameters measured are: percentage of CXCR4-expressing polynuclear neutrophils compared to the total number of polynuclear neutrophils measured by flow cytometry in the blood of patients on day 3 after admission to the ICU. percentage of CXCR4-expressing polynuclear neutrophils compared to the total number of polynuclear neutrophils measured by flow cytometry in the pulmonary compartment of patients within 24h and on day 3 after admission to the ICU. mean fluorescence intensity of CXCR4 in polynuclear neutrophils measured by flow cytometry in the blood of patients within 24h and on day 3 after admission to the ICU. mean fluorescence intensity of CXCR4 in polynuclear neutrophils measured by flow cytometry in the pulmonary compartment of patients within 24h and on day 3 after admission to the ICU.

[0514] - clusterin and C5 complement levels measured in the plasma of patients within 24h and on day 3 after admission to the ICU. death rate at day 90 after admission to the ICU. PaOi / FiOi ratio after optimization of mechanical ventilation, within 24 hours after admission to the ICU. During the analysis, the patients are sorted into two groups: PaOi / FiOi ratio inferior or equal to 150 ; and PaCWFiOi ratio strictly superior to 150, since this ratio corresponds to the severity threshold for treatment intensification (ventral decubitus and use of curares). As this ratio is sensitive to mechanical ventilation settings, and in particular to the setting of positive end- expiratory pressure, the value retained for analysis is the value after ventilatory optimization in the 24 hours following admission to intensive care.

[0515]

[0315] The additional parameters measured are:

[0516] CXCL-12 and CXCL-17 measured in the plasma of patients within 24h and on day 3 after admission to the ICU.

[0517] - pro-inflammatory interleukins and chemokines levels measured in the plasma of patients within 24h and on day 3 after admission to the ICU.

[0518] - anti-inflammatory interleukins and chemokines levels measured in the plasma of patients within 24h and on day 3 after admission to the ICU.

[0519] Neutrophils Extracellular Traps markers (MPO-DNA complex, circulating DNA, citrullinated histone H3, neutrophil elastase and calprotectin) measured in the plasma of patients within 24h and on day 3 after admission to the ICU.

[0520] Inclusion criteria

[0521]

[0316] Inclusion criteria (common to the three groups of patients) are the following:

[0522] Person 18-year-old or older

[0523] Person suffering from an acute respiratory distress syndrome defined according to the Berlin criteria by:

[0524] 1) Acute respiratory failure evolving for a week or less

[0525] 2) Bilateral opacities on chest imaging 3) No evidence of predominant hydrostatic oedema

[0526] 4) Hypoxemia with PaC FiCh ratio < 300 mmHg with positive expiratory pressure set at 5cm H2O or more.

[0527] Person having been in need of mechanical ventilation after oro-tracheal intubation for less than 24h.

[0528] Person affiliated to or benefiting from a social security scheme.

[0529] Signed informed consent from a family member or trusted support person, or failing that, from a close relative, for inclusion after the patient has been admitted to the ICU.

[0530]

[0317] As the microbiological documentation (bacterial culture or viral PCR) is not always available within the first 48 hours of intubation, it is not possible to include the exclusion criteria detailed below as inclusion criteria.

[0531] Non-inclusion criteria

[0532]

[0318] Non-inclusion criteria are the following:

[0533] Person with neutropenia (< 500 / mm3)

[0534] Person with a known qualitative anomaly of polynuclear neutrophils (complete or partial myeloperoxidase deficiency, chronic septic granulomatosis, neutrophil immune deficiency syndrome, Shwachman-Diamond syndrome, Chediak- Higashi syndrome).

[0535] Person participating in another interventional study testing an immunomodulator or antiviral treatment in this indication.

[0536] Person with an infection by Human Immunodeficiency Virus (HIV) at the acquired immunodeficiency syndrome (AIDS) stage.

[0537] Person with a contra-indication to bronchoalveolar lavage for patients:

[0538] 1) Severe bronchospasm 2) Uncontrolled shock

[0539] 3) Intracranial hypertension

[0540] 4) Severe refractory hypoxemia ( PaC FiCh < 60 mmHg)

[0541] Person under specific legal protection or deprived of their liberty: pregnant or breast-feeding women, person under guardianship or legal protection.

[0542] Exclusion criteria

[0543]

[0319] As the microbiological documentation (bacterial culture or viral PCR) is not always available within the first 48 hours of intubation, a patient may be admitted until the results of the test are available, and be excluded a posteriori.

[0544]

[0320] A person who does not satisfy any of the following criteria, is excluded from the clinical trial:

[0545] Criteria for the group of ARDS caused by severe influenza infection : a person with an infection caused by influenza virus proven by a positive polymerase chain reaction test.

[0546] Criteria for the group of ARDS caused by lung infection by bacteria: a person with proven bacterial infection (over 105units forming colony per mL for tracheal aspirates and over 104units forming colony per mL for BAL).

[0547] Criteria for the group of ARDS caused by a systemic inflammation of extra- pulmonary origin: o a person with acute pancreatitis according to the 2013 international recommendations (typical pain, lipaemia above 3 times normal and / or imaging abnormality); or o a person with an acute bacterial peritonitis based on clinical diagnosis and bacteria isolated on peritoneal swabs (as recommended by the French Society for Anesthesia and Intensive Care - SFAR, RFE 2018). EXAMPLE 6: ARDS severity induced by cecal ligation and puncture is associated to NETosis

[0548]

[0321] Three groups of C57B1 / 6 mice were allocated for this study (see Table 6). The model of ARDS was induced cecal ligation and puncture (CLP) with two levels of severity. On day 0, 16 mice were induced by CLP with a 50% cecum ligation, to mimic moderate ARDS ; and 16 mice were induced by CLP with a 75% cecum ligation, to mimic severe ARDS (groups 2M and 3M, respectively). 16 mice received the surgery in the same way as groups 2M and 3M without CLP and served as non-induced control group (sham, group IM). Table 6: Study design of example 6.

[0549]

[0322] During the study, mortality and morbidity observations, clinical score and body weight measurements were performed for 7 days for 16 mice per group. Mice reaching a body weight loss superior to 20% of their initial body weight and / or reaching humane endpoints were euthanized and the day of death was recorded. This enabled to establish a survival curve which was represented in the form of the Kaplan Meier estimator. As shown in Figure 5A, mice induced with 75% cecal ligation had a mortality rate of 66.7% on day 7 (p <0.01) while mice induced with 50% cecal ligation had a mortality rate of 21.5%. All mice from the non-induced control group survived up to study termination (day 7).

[0550]

[0323] Additionally, for 5 mice per group lungs and blood were collected for NETosis analysis after 30 hours. Concentration of Citrullinated Histone H3 (H3cit) were evaluated in lung homogenates and concentrations of Neutrophil Elastase (NE), Myeloperoxidase (MPO) and circulating DNA ( cDNA) were evaluated in plasma by ELISA or by fluorometric method.

[0551]

[0324] As shown in Figure 5B, H3cit levels were only detected at low concentrations in non-induced mice (3.24 ± 0.16 ng / mL / mg of tissues) and those with 50% cecal ligation (3.93 ± 0.23 ng / mL / mg of tissues). However, 75% cecal ligation significantly increased the production of H3cit in lung homogenates compared to the sham control group (4.34 ± 0.47 ng / mL / mg of tissues; p<0.05).

[0552]

[0325] As shown in Figure 5C, the MPO levels were detected at low concentration in non-induced mice (709.6 ± 23.8 pg / mL). 50% cecal ligation significantly induced the production of MPO (1000.8 ± 61.9 pg / mL, p<0.01) compared to the sham control, as well as 75% cecum ligation, which further increased its production in plasma (1366.93 ± 50.92 pg / mL; p<0.001).

[0553]

[0326] As shown in Figure 5D, the CXCL1 / KC levels were detected at low concentration in non-induced mice (88.6 ± 31.4 pg / mL). 50% cecal ligation non- significantly increased the production of CXCL1 / KC (577.2 ± 29.1 pg / mL, p<0.01) compared to the sham control, while 75% cecum ligation significantly increased its production in plasma (685.3 ± 294.3 pg / mL; p<0.05).

[0554]

[0327] As shown in Figure 5E, the NE levels were detected at low concentration in noninduced mice (29190 ± 1480 pg / mL). 50% cecal ligation significantly increased the production of NE (115920 ± 24679 pg / mL, p<0.01) compared to the sham control, while 75% cecum ligation further significantly increased its production in plasma (168010 ± 10272 pg / mL; p<0.001). As shown in Figure 5F, cDNA levels were detected at low concentration in non-induced mice (0.221 ± 0.007 pg / mL). 50% cecal ligation didn’t significantly modulate the production of cDNA (0.470 ± 0.145 pg / mL) compared to the sham control, while 75% cecum ligation significantly increased its production in plasma (1.187 ± 0.243 pg / mL; p<0.001).

[0555]

[0328] In conclusion, as shown in Figure 5, the levels of NETosis biomarkers measured on day 1 in models of moderate ARDS and severe ARDS: H3Cit (Figure 5B) quantification in lung homogenates and MPO (Figure 5C), CXCL1-KC (Figure 5D), NE (Figure 5E), cDNA (Figure 5F) quantification in plasma, are good predictors of mortality (Figure 5A).

[0556] EXAMPLE 7: Plerixafor is effective in severe ARDS induced by cecal ligation and puncture

[0329] Five groups of C57B1 / 6 mice were allocated for this study (see Table 7). CLP surgery was conducted (groups 2M to 5M) on 10 mice on day 0 to induce severe or moderate ARDS model. 10 mice received surgery without CLP in the same way as groups 2M to 5M and served as non-induced control group (sham, group IM). One hour after CLP surgery mice were subcutaneously (SC) treated with water as vehicle (groups IM, 2M and 4M) or plerixafor at 10 mg / kg (group 3M and 5M).

[0557] Table 7: Study design of example 7.

[0558]

[0330] During the study, mortality and morbidity observations, clinical score and body weight measurements were performed for 7 days for 8-10 mice per group. Mice reaching a body weight loss superior to 20% of their initial body weight and / or reaching humane endpoints were euthanized and the day of death was recorded. This enabled to establish a survival curve which was represented in the form of the Kaplan Meier estimator. As shown in Figure 6A, plerixafor significantly enhanced the survival of mice induced with 75% cecal ligation, with 50% survival, compared to the control group, where mice induced with 75% cecal ligation not treated with plerixafor had only 10% survival (p<0.05). All mice from the non-induced control group (IM) survived up to study termination (day 7).

[0559]

[0331] Additionally, for 8 mice per group, blood was collected for NETosis analysis after 24 hours. Concentration of circulating DNA (cDNA) was evaluated in plasma by fluorometric method. As shown in Figure 6b, cDNA level was detected at low concentration in non-induced mice (0.230 ± 0.006 pg / mL). 50% cecal ligation significantly increased the production of cDNA (0.448 ± 0.042 pg / mL; p<0.001) compared to sham control, while 75% cecal ligation led to a further significant increase in its plasma concentration (0.817 ± 0.175 pg / mL; p<0.001). Treatment with plerixafor did not affect the cDNA production in the 50% cecal ligation group (moderate ARDS; 0.563 ± 0.098 pg / mL), whereas it significantly reduced DNA levels in the 75% cecal ligation group (severe ARDS; 0.451 ± 0.055 pg / mL; p<0.05).

[0560]

[0332] In conclusion, as shown in Figure 6, the plerixafor is effective in a model of severe ARDS (75% cecal ligation), assessed by survival (Figure 6A). Moreover, while plerixafor had no effect on NETosis plasmatic markers in a model of moderate ARDS, plerixafor significantly reduced the concentrations of NETosis markers in plasma, such as cDNA, in a model of severe ARDS (Figure 6B). EXAMPLE 8: CXCR4 antagonists are effective in severe ARDS induced by cecal ligation and puncture

[0561]

[0333] Four groups of C57B1 / 6 mice were allocated for this study (see Table 8). CLP surgery was conducted (groups 2M to 4M) on 24 mice on day 0 to induce severe ARDS model. 8 mice received surgery without CLP in the same way as groups 2M to 4M and served as non-induced control group (sham, group IM). One hour after CLP surgery mice were subcutaneously (SC) treated with water as vehicle (groups IM and 2M), plerixafor at 10 mg / kg (group 3M) or motixafortide (another CXCR4 antagonist) at 12 mg / kg (group 4M).

[0562] Table 8: Study design of example 8.

[0563]

[0334] Blood samples were collected for NETosis analysis 24 hours later. Concentrations of Myeloperoxidase (MPO) and circulating DNA (cDNA) were evaluated in plasma by ELISA or by fluorometric method. As shown in Figure 7A, the MPO level was detected at low concentration in non-induced mice (2995 ± 204 pg / mL). 75% cecal ligation significantly increased the production of MPO (4325 ± 230 pg / mL, p<0.01) compared to the sham control. Plerixafor and motixafortide treatments decreased MPO production in plasma compared to the 75% cecal ligation with vehicle (3691 ± 241 pg / mL; p<0.05 and 3522 ± 210 pg / mL; p=0.06, respectively). In the same way, as shown in Figure 7B, H3cit level in lung homogenates was significantly increased by the 75% cecal ligation (0.817 ± 0.175 ng / mL / mg of tissues) compared to the sham control (0.230 ± 0.006 ng / mL / mg of tissues, p<0.001). Plerixafor and motixafortide treatments decreased the production of H3cit in lung homogenates compared to the sham control group (0.451 ± 0.055 ng / mL / mg of tissues; p<0.05 and 0.465 + 0.045 ng / mL / mg of tissues; p=0.18, respectively).

[0564]

[0335] In conclusion, as shown in Figure 7, plerixafor and motixafortide are effective in a model of severe ARDS (75% cecal ligation), as assessed by NETosis markers such as MPO (Figure 7A) and cDNA (Figure 7B). These findings suggest a class effect of CXCR4 antagonists on severe ARDS, in animals that will develop or suffer from severe ARDS that may be identified by their elevated level of NETosis.

[0565] RECTIFIED SHEET

Claims

CLAIMS1. A CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof, for use in the treatment and / or the prevention of a severe acute respiratory distress syndrome (sARDS) in a subject in need thereof, wherein the sARDS is characterized by a dysregulation of NETosis and wherein the subject has a PaOi / FiCh ratio inferior or equal to 200 mmHg, preferably inferior or equal to 100 mmHg.

2. The CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof for use according to claim 1, wherein the dysregulation of NETosis is characterized by the presence of at least one of the following NETosis biomarkers measured in a biological sample from the subject:- circulating DNA levels equal to or greater than 200 ng / mL, or measured by an optical density (OD) equal to or greater than 0.4 (absorbance measured at 450 nm),- citrullinated histone H3 levels equal to or greater than 200 ng / mL,- citrullinated histone H3-DNA complexes levels equal to or greater than 150 ng / mL,- neutrophil elastase levels equal to or greater than 200 ng / mL,- interleukin-8 levels equal to or greater than 100 pg / mL,- myeloperoxidase-DNA complexes levels equal to or greater than 200 ng / mL, and / or- calprotectin levels equal to or greater than 5 pg / mL.

3. The CXCR4 inhibitor compound or a pharmaceutically acceptable salt and / or hydrate thereof for use according to claim 1, wherein the dysregulation of NETosis is characterized by the presence of at least one of the following NETosis biomarkers measured in a biological sample from the subject:- the ratio of circulating DNA levels in the subject compared to circulating DNA levels in a healthy individual is equal to or greater than 1,4;- the ratio of citrullinated histone H3 levels in the subject compared to citrullinated histone H3 levels in a healthy individual is equal to or greater than 2,0;- the ratio of citrullinated histone H3-DNA complexes levels in the subject compared to citrullinated histone H3-DNA complexes levels in a healthy individual is equal to or greater than 1,8;- the ratio of neutrophil elastase levels in the subject compared to neutrophil elastase levels in a healthy individual is equal to or greater than 3,5;- the ratio of interleukin-8 levels in the subject compared to interleukin-8 levels in a healthy individual is equal to or greater than 30;- the ratio of myeloperoxidase-DNA complexes levels in the subject compared to myeloperoxidase-DNA complexes levels in a healthy individual is equal to or greater than 1,4; and / or- the ratio of calprotectin levels in the subject compared to calprotectin levels in a healthy individual is equal to or greater than 10.

4. The CXCR4 inhibitor compound for use according to any one of claims 1 to 3, wherein the compound is of formula (I), and formula (I) is:- Z and Y, which are identical or different, represent: o an alkyl group having from 1 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms, oro an aryl or heteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a hydroxyl group, a halogen atom, a CN group, a CF3 group, a SO group, a NO2 group, an amine group, a difluor or an alkyl, S-alkyl or O- alkyl group having from 1 to 6 carbon atoms, or o a cyclic or heterocyclic polyamine group having from 9 to 32 ring members and from 3 to 8 amine groups in the ring spaced by 2 or more carbon atoms from each other, optionally comprising an aryl or heteroaryl group having from 3 to 6 carbon atoms, a heteroatom, and optionally substituted by a halogen atom, a hydroxyl group, a heteroatom, an alkyl, S-alkyl or O-alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, or o a D1D2N- group, wherein DI and D2, which may be identical or different, represent:• a hydrogen atom, or• an alkyl group having from 1 to 6 carbon atoms optionally substituted by at least one hydroxyl group, a halogen atom, a CF3 group, a CN group, an amine group, or an alkyl, O-alkyl or S-alkyl group having from 1 to 12 carbon atoms, or• an aryl, an heteroaryl, a cycloalkyl, a heterocycloalkyl, an alkylaryl, an alkylheteroaryl or an alkylheteropoly aryl group having from 3 to 12 carbon atoms, optionally substituted by at least one hydroxyl group, halogen atom, CF3 group, CN group, amine group, or alkyl, O-alkyl or S-alkyl group having from 1 to 12 carbon atoms, or• DI and D2 are linked together to from a N-containing aryl or heteroaryl group having from 3 to 12 carbon atoms and optionally substituted by at least one amine group optionally substituted by an alkylheteroaryl group having from 3 to 12 carbon atoms, and- A represents: o an aryl or heteroaryl group having from 3 to 12 carbon atoms, or o an alkyl group having from 1 to 6 carbon atoms, or o -R4-Y’-R5-, wherein R4 and R5 which are identical or different represent an alkyl group having from 1 to 6 carbon atoms and Y’ represents an aryl orheteroaryl group having from 3 to 6 carbon atoms, optionally substituted by a halogen atom, a hydroxyl group, an amide group, an amine group, an alkoxy group, an ester group, a CF3 group, a CN group or an alkyl, O-alkyl or S- alkyl group having from 1 to 6 carbon atoms optionally substituted by a hydroxyl group, an amine group or an O-alkyl group having from 1 to 6 carbon atoms, and- R and R’, which are identical or different, represent an alkyl group having from 1 to 6 carbon atoms or a R1NR2R3 group or a single bond, and- R1 represents a single bond or an alkyl group having from 1 to 6 carbon atoms, and- R2 and R3, which are identical or different, represent a hydrogen atom, an amine group, an alkyl group having from 1 to 6 carbon atoms or an aryl or heteroaryl group having from 3 to 6 carbon atoms, wherein said aryl or heteroaryl group having from 3 to 6 carbon atoms is optionally substituted by an alkyl group having from 1 to 6 carbon atoms.

5. The CXCR4 inhibitor compound for use according to any one of claims 1 to 3, wherein the CXCR4 inhibitor compound is plerixafor, LY2510924, AMD3465, MSX-122, mavorixafor, motixafortide and a mixture thereof, preferably plerixafor.

6. A method of prognostic of severe ARDS in a subject wherein the subject has a PaOi / FiOi ratio inferior or equal to 200 mmHg, preferably inferior or equal to 100 mmHg, comprising the following steps:(a) measuring the levels of a NETosis biomarker in a biological sample from a subject, in particular from the subject’s plasma,(b) comparing said NETosis biomarker with a subject of reference and / or threshold of NETosis,(c) determining whether the subject is at risk of suffering from a severe ARDS depending on the levels of NETosis biomarker, and(d) optionally treating the subject with a CXCR4 inhibitor compound.

7. The method according to claim 6, wherein the CXCR4 inhibitor compound is selected from the list consisting of plerixafor, LY2510924, AMD3465, MSX-122, mavorixafor, motixafortide and a mixture thereof, preferably plerixafor.

8. The method of any one of claims 6 or 7, wherein the NETosis biomarker measured in a biological sample from the subject is circulating DNA levels, citrullinated histone H3 levels, interleukin-8 levels, myeloperoxidase-DNA complexes levels, neutrophil elastase levels and / or calprotectin levels.

9. The method of any one of claims 6 to 8, wherein step (d) is the administration of a CXCR4 inhibitor compound to the patient with severe ARDS via one of the following routes: intravenous, intramuscular, subcutaneous, parenteral, oral, nasal, ocular, transmucosal, or transdermal.

10. The method of any one of claims 6 to 9, wherein step (d) corresponds to the continuous intravenous administration of a CXCR4 inhibitor compound to the patient with severe ARDS.

11. A pharmaceutical composition for use in the treatment and / or in the prevention of severe acute respiratory distress syndrome (ARDS) in a subject in need thereof, wherein the severe ARDS is characterized by a dysregulation of NETosis, wherein the subject has a PaCE / FiCE inferior to 200 mmHg, preferably inferior or equal to 100 mmHg; and wherein said pharmaceutical composition comprises a CXCR4 inhibitor compound and a pharmaceutically acceptable excipient.

12. The CXCR4 inhibitor compound for use, the method of prognostic, or a pharmaceutical composition for use according to any one of claims 1 to 11, wherein severe ARDS results from a viral, a fungal or a bacterial infection.

13. A kit for prognostic of a severe ARDS comprising the means for measuring in a biological sample from a subject at least one of NETosis biomarkers selected from the list consisting of circulating DNA levels, citrullinated histone H3 levels, neutrophil elastase levels, interleukin-8 levels, myeloperoxidase-DNA, complexes levels, and calprotectin levels.

14. Use of at least one NETosis biomarker selected from circulating DNA, citrullinated histone H3, citrullinated histone H3-DNA complexes, neutrophil elastase, interleukin- 8, myeloperoxidase-DNA complexes, and calprotectin as a biomarker for in vitro detection of severe ARDS in a biological sample, preferably the at least one biomarker is selected from circulating DNA, citrullinated histone H3 and neutrophil elastase.