Compositions comprising nangibotide and uses thereof

Abstract

The present invention relates to a pharmaceutical composition comprising nangibotide, and at least one amino acid selected from the group consisting of: arginine, lysine and histidine; wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide.

Description

COMPOSITIONS COMPRISING NANGIBOTIDE AND USES THEREOFFIELD OF INVENTION

[0001] The present invention relates to pharmaceutical compositions comprising nangibotide.

[0002] The present invention also relates to methods for producing such pharmaceutical compositions and to said pharmaceutical compositions for use in the treatment of a disease selected from the group consisting of: inflammatory diseases, cardiovascular diseases and infectious diseases.BACKGROUND OF INVENTION

[0003] Triggering receptor expressed on myeloid cells- 1 (TREM-1) is a transmembrane receptor expressed on innate immune cells and part of the family of TREM receptors. Activation of TREM- 1 triggers the release of pro-inflammatory cytokines and chemokines and contributes to the dysregulated immune response in sepsis (Siskind et al., 2022, Front. Immunol. 13:907387). TREM-like transcript 1 (TLT-1) also belongs to the TREM family and is specific to platelets and megakaryocytes, where it plays a role in hemostasis / thrombosis (Washington et al., 2004, Blood 104(4): 1042- 1047). A TLT-1- derived peptide (nangibotide) has been shown to exhibit anti-inflammatory properties by inhibiting TREM-1 signaling (Derive et al., 2012, J Immunol. 188:5585-5592). Administration of nangibotide in an adult minipig model of induced sepsis reduced cardiovascular failure and improved survival in animals after induction of fecal peritonitis (Derive et al., 2013, Shock 39(2): 176-182). Nangibotide has also been tested in a Phase 2a randomized controlled clinical trial in patients with septic shock (Francois et al., 2020, Intensive Care Med., 46(7): 1425-1437). Pharmaceutical compositions of nangibotide aretherefore needed that are safe and effective for administration to treat septic shock and other disorders.SUMMARY

[0004] The present invention relates to a pharmaceutical composition comprising: nangibotide, at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof, and at least one pharmaceutically acceptable buffer selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof, wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide.

[0005] The present invention also relates to a pharmaceutical composition comprising: nangibotide, at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof, and at least one pharmaceutically acceptable buffer selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof, wherein the amount of the at least one amino acid is of at least 15% by weight relative to the dry weight of the pharmaceutical composition.

[0006] For example, the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0007] The present invention also relates to a pharmaceutical composition comprising: nangibotide, at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof, and at least one pharmaceutically acceptable buffer selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof, wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide.

[0008] For example, the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0009] For example, the amount of the at least one pharmaceutically acceptable buffer is less than 10%, preferably it ranges from 3% to 7%, more preferably it ranges from 3% to 5%, even more preferably from 2% to 5%, better it ranges from 4% to 6%, still better it is about 5%, by weight relative to the weight of nangibotide.

[0010] The present invention also relates to a pharmaceutical composition comprising: nangibotide, and at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof; wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide; and wherein the pH of said pharmaceutical composition ranges from 4.5 to 8.5.

[0011] The present invention also relates to a pharmaceutical composition comprising: nangibotide, and at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof; wherein the amount of the at least one amino acid is of at least 15% by weight relative to the dry weight of the pharmaceutical composition; and wherein the pH of said pharmaceutical composition ranges from 4.5 to 8.5.

[0012] For example, the pharmaceutical composition has a pH ranging from 4.5 to 8.5, preferably from 4.5 to 7.4, more preferably from 5 to 7, even more preferably from 5 to 6, or better said pharmaceutical composition has a pH of about 5.5.

[0013] For example, the pharmaceutical composition further comprises at least one pharmaceutically acceptable buffer, preferably the at least one pharmaceutically acceptable buffer being selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof.

[0014] For example, the amount of the at least one pharmaceutically acceptable buffer is less than 10%, preferably it ranges from 3% to 7%, more preferably it ranges from 3% to 5%, even more preferably from 2% to 5%, better it ranges from 4% to 6%, still better it is about 5%, by weight relative to the weight of nangibotide.

[0015] For example, the pharmaceutical composition does not comprise a buffer.

[0016] For example, said pharmaceutical composition comprises less than 5.0% of dimeric nangibotide by weight relative to the total weight of all peptides in said pharmaceutical composition.

[0017] For example, said pharmaceutical composition comprises from 600 mg to 2500 mg of nangibotide, preferably from 800 mg to 2500 mg of nangibotide, more preferably from 1200 mg to 2000 mg of nangibotide.

[0018] For example, said pharmaceutical composition comprises at least 65%, preferably from about 65% to about 85%, more preferably from about 70% to about 80%, of nangibotide by weight relative to the dry weight of the pharmaceutical composition.

[0019] For example, the amount of the at least one amino acid ranges from 22% to 50%, preferably from 22% to 34%, more preferably from 22% to 30%, even more preferably from 23% to 29%, by weight relative to the weight of nangibotide.

[0020] For example, the pharmaceutical composition comprises (i) about 72.6% by weight of nangibotide, about 20% by weight of arginine, and about 3.9% by weight of citrate buffer (i.e., the sum of the weight of the citric acid and conjugate base) relative to the dry weight of the pharmaceutical composition or (ii) 71.3% by weight of nangibotide, about 19.7% by weight of arginine, and about 3.9% by weight of citrate buffer (i.e., the sum of the weight of the citric acid and conjugate base), relative to the total weight of the pharmaceutical composition.

[0021] For example, the pharmaceutical composition comprises about 1200 mg nangibotide, about 331 mg arginine, and about 65 mg sodium citrate.

[0022] For example, the pharmaceutical composition is a dry composition that when reconstituted with a liquid comprises about 120 mg / mL nangibotide, about 33.1 mg / mL arginine, and about 6.5 mg / ml sodium citrate.

[0023] For example, the pharmaceutical composition is a dry composition that when reconstituted with a liquid comprises:(i) about 120 mg / mL to about 200 mg / mL nangibotide(ii) at least one of amino acid selected from arginine, lysine, or histidine or a combination of thereof at a concentration of about 140 mM to about 300 mM; and(iii) a buffer selected from citrate buffer, a histidine buffer, a succinate buffer, an acetate buffer, a phosphate buffer, or tromethamine buffer from about 20 mM to about 60 mM;wherein the pH of the reconstituted pharmaceutical composition is about 4.5 to about 8.5.

[0024] For example, in the pharmaceutical composition:(i) the buffer is about 45-55 mM acetate;(ii) the buffer is about 20-30 mM citrate;(iii) the buffer is about 20-30 mM histidine;(iv) the buffer is about 45-55 mM phosphate;(v) the buffer is about 45-55 mM succinate; or(vi) the buffer is about 45-60 mM tromethamine.

[0025] For example, the pharmaceutical composition is a dry composition that when reconstituted with a liquid comprises about 4 mg / mL or less of dimeric nangibotide, about 1 mg / mL or less of Des-Gln-LR12, and about 0.5 mg / mL or less of Met(O)12-LR12.

[0026] For example, said pharmaceutical composition comprises less than or equal to 1.2% of Des-Gln-LR12, by weight relative to the total weight of all peptides in said pharmaceutical composition and / or less than or equal to 1.0% of Met(O)12-LR12 by weight relative to the total weight of all peptides in said pharmaceutical composition.

[0027] For example, said pharmaceutical composition is a powder.

[0028] The present invention also relates to a reconstituted pharmaceutical composition comprising the pharmaceutical composition as described herein and an aqueous medium, preferably said aqueous medium being selected from the group consisting of: water and a pharmaceutically acceptable aqueous solution, more preferably said aqueous medium being a saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.

[0029] The present invention also relates to a ready-to-use pharmaceutical composition comprising the pharmaceutical composition as described herein and an aqueous medium, preferably said aqueous medium being selected from the group consisting of: water and apharmaceutically acceptable aqueous solution, more preferably said aqueous medium being a saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.

[0030] For example, said ready-to-use pharmaceutical composition presents an osmolality of less than 500 mOsm / kg, preferably an osmolality ranging from 300 to 400 mOsm / kg, more preferably an osmolality ranging from 320 to 390 mOsm / kg, even more preferably an osmolality of about 380 mOsm / kg.

[0031] For example, said ready-to-use pharmaceutical composition comprises equal to or less than 10.0%, preferably equal to or less than 5.0%, of nangibotide dimers by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition.

[0032] The present invention also relates to the pharmaceutical composition as described herein or the reconstituted pharmaceutical composition as described herein or the ready-to- use pharmaceutical composition as described herein, for use as a medicament.

[0033] The present invention also relates to the pharmaceutical composition as described herein or the reconstituted pharmaceutical composition as described herein or the ready-to- use pharmaceutical composition as described herein, for use in the treatment of a disease in a subject in need thereof, wherein said disease is selected from the group consisting of: inflammatory diseases, preferably selected from the group consisting of: septic shock, sepsis, severe sepsis, systemic inflammatory response syndrome (SIRS), acute respiratory distress syndrome (ARDS), hemorrhagic shock, ischemiareperfusion and pancreatitis; cardiovascular diseases, preferably selected from the group consisting of: myocardial infarction, cerebral infarction, acute myocardial infarction, ischemia, coronary heart disease, acute coronary syndrome, stroke, aneurysm, stable angina pectoris, effort angina pectoris, cardiomyopathy, hypertensive heart disease, chronic heart failure, acute heart failure, cor pulmonale, cardiac dysrhythmias, inflammatory heart disease (such as endocarditis and myocarditis), peripheralarterial disease, SIRS-associated myocardial and / or vascular dysfunction, and atherosclerosis; and infectious diseases, preferably COVID- 19.

[0034] For example, said pharmaceutical composition or reconstituted pharmaceutical or ready-to-use pharmaceutical composition is for administration to said subject in need thereof via intravenous route of administration at a rate ranging from 0.1 to 3 mg / kg / hr (mg of nangibotide per kilogram of bodyweight of the subject per hour).

[0035] The present invention also relates to a process for preparing a pharmaceutical composition as described herein, comprising the steps of: a) mixing nangibotide with at least one amino acid selected from the group consisting of: arginine, lysine and histidine, wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide, thereby leading to a bulk solution; b) optionally adjusting the pH of the bulk solution obtained in step a) to a pH ranging from 5.5 to 6.5 or ranging from 5.0 to 7.0, preferably ranging from 5.5 to 6.0, preferably a pH of 5.5, preferably said pH adjustment being carried out using HC1; and c) optionally lyophilizing the bulk solution obtained in step a) or in step b) to obtain a pharmaceutical composition in a powder form.

[0036] The present invention also relates to a process for preparing a reconstituted pharmaceutical composition as described herein, comprising the steps of: a) mixing nangibotide with at least one amino acid selected from the group consisting of: arginine, lysine and histidine, wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide, thereby leading to a bulk solution;b) optionally adjusting the pH of the bulk solution obtained in step a) to a pH ranging from 5.5 to 6.5 or ranging from 5.0 to 7.0, preferably ranging from 5.5 to 6.0, preferably a pH of 5.5, preferably said pH adjustment being carried out using HC1; and c) optionally lyophilizing the bulk solution obtained in step a) or in step b) to obtain a pharmaceutical composition in a powder form; then- adding an aqueous medium to the pharmaceutical composition in a powder form obtained in step c), to obtain a reconstituted pharmaceutical composition, preferably the total volume of the reconstituted pharmaceutical composition is 10 mb, preferably in a 20 mb vial, preferably said aqueous medium is selected from the group consisting of: water and saline water solution, more preferably said aqueous medium is selected from the group consisting of: water and saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.

[0037] The present invention also relates to a process for preparing a ready-to-use pharmaceutical composition as described herein, comprising the steps of:- carrying out the process as described herein to obtain a reconstituted pharmaceutical composition; then- diluting the reconstituted pharmaceutical composition in an aqueous medium, preferably in a syringe or in an infusion bag, for example a syringe or in an infusion bag having a total volume of 50 mb, to obtain a ready-to-use pharmaceutical composition.

[0038] The present invention also relates to a purified nangibotide substance comprising equal to or less than 2.7% of dimeric nangibotide by weight relative to the total weight of all peptides of said purified nangibotide substance.

[0039] For example, the purified nangibotide substance comprises at least 85% of peptides by weight relative to the total weight of the purified nangibotide substance.

[0040] For example, said purified nangibotide substance comprises: equal to or less than about 1.2%, preferably equal to or less than about 1.0%, more preferably equal to or less than about 0.5%, of Des-Gln-LR12, by weight relative to the total weight of all peptides in said purified nangibotide substance; and / or equal to or less than about 1.0%, preferably equal to or less than about 0.75%, more preferably equal to or less than about 0.5%, of Met(O)12-LR12, by weight relative to the total weight of all peptides in said purified nangibotide substance.DEFINITIONS

[0041] In the present invention, the following terms have the following meanings:

[0042] The terms “a” and “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. Similarly, the expressions “at least one” and “one or more” are interchangeable.

[0043] “About” preceding a value is used to indicate that the value includes the inherent variation of error for the device or the method being employed to determine the value, or the variation that exists among the samples being measured. Unless otherwise stated or otherwise evident from the context, the term “about” means within 10% above or below the reported numerical value (except where such number would exceed 100% of a possible value or go below 0%). When used in conjunction with a range or series of values, the term “about” applies to the endpoints of the range or each of the values enumerated in the series, unless otherwise indicated. “About” appears before a value, the absolute value itself is also disclosed. For example, “about 10” is also a disclosure of the value 10. As used herein, the terms “about” and “approximately” may be used as equivalents.

[0044] “All peptides” or “peptides”, in a composition, refers to the total of all the different peptides contained in said composition. In particular, in the present application, the term “all peptides” refers to the total content of nangibotide, along with any derivatives of nangibotide (belonging to the peptide impurities) that may be present including dimeric nangibotide, Des- Gln-LR12, and Met(O)12-LR12 in a given composition.

[0045] “API” is an active pharmaceutical ingredient, i.e., nangibotide in the present application. An API is an agent that has a treatment effect; it may be a chemical or a biological ingredient.

[0046] “At least” means “greater than or equal to” and is thus different from “strictly greater than” which does not comprise “equal to”.

[0047] “Buffer” relates to a mixture of a weak acid and its conjugate base, or a weak base and its conjugate acid, allowing the pH of a pharmaceutical composition to resist significant changes in pH when a small amount of strong acid or base is added thereto, preferably to be maintained constant. “Buffering agent” refers to the specific chemical that is present in both an acidic form and a basic form in a buffer, allowing the pH of a pharmaceutical composition to resist significant changes in pH when a small amount of strong acid or base is added thereto, preferably to be maintained constant. A “binary buffer” refers to a buffer comprising only two entities, i.e., a conjugate acid-base pair, where the acid and its conjugate base are the only forms present in the system. Examples of binary buffers are succinate buffer (conjugate acid-base pair: succinic acid / succinate), lactic acid buffer (conjugate acid-base pair: lactic acid / lactate), acetate buffer (conjugate acid-base pair: acetic acid / acetate), histidine buffer (conjugate acid-base pair: histidine hydrochloride / histidine), MES buffer (conjugate acid-base pair: MES (2-(N-morpholino)ethanesulfonic acid) / conjugate base of MES), and tromethamine buffer (TRIS, tris(hydroxymethyl)aminomethane) (conjugate acidbase pair: tromethamine hydrochloride / conjugate base of tromethamine). A “ternary buffer” refers to a buffer comprising three entities due to multiple protonation states. Examples of ternary buffers are citrate buffer (conjugate acid-base pair: citric acid / citrate:three possible protonation states: HsCeHsO?, HzCeHsO? , HCeHsO?2) and phosphate buffer (conjugate acid-base uripair: phosphoric acid / phosphate ions: three possible protonation states: H3PO4, H2PO4 ., HPO42). An “acidic buffer” is a buffer whose pKa is less than 6; for example, the term “acidic buffer” may include acetate buffer (pKa —4.76), succinate buffer (pKa -4.21), lactic acid buffer (pKa -3.86), adipic acid buffer (pKa -4.41), glutamate buffer (pKa -4.25), citrate buffer (pKa -3.13, 4.76) and mixtures thereof. A “neutral buffer” is a buffer whose pKa ranges from 6 to 8; for example, the term “neutral buffer” may include MES buffer (pKa -6.15), histidine buffer (pKa -6.0), phosphate buffer (pKa -7.2) and mixtures thereof. An “alkaline buffer” is a buffer whose pKa is greater than 8; for example, the term “alkaline buffer” may include tromethamine buffer (pKa -8.1).

[0048] “Chronic disease” refers to a long-term, progressive illness, often associated with disability and the threat of serious complications. Chronic diseases evolve more or less rapidly for at least several months, in particular at least 3 months.

[0049] “Comprising” or “comprise” is to be construed in an open, inclusive sense, not limited to the features following this term.

[0050] “Consisting of’ or “consist” is to be construed in a closed, non-inclusive sense, limited to the features following this term.

[0051] “Consisting essentially of’ is to be construed as being limited to the features following this term with the exception that additional features that do not materially affect the basic and novel characteristics of the recited features, e.g., the basic and novel characteristics of the claims, may also be included.

[0052] “Des-Gln-LR12” is a peptide whose amino acid sequence is SEQ ID NO: 2(LEED AGEYGCM) and whose chemical formula is the following formula (III):Formula (III).

[0053] “Dose” refers to the amount of API administered at one time. When the API is administered by intravenous route of administration, one dose may be administered either by intravenous injection (=a single and rapid administration delivered quickly, often in seconds or minutes, for example via a syringe) or intravenous infusion (=a continuous administration over a period of time at a controlled rate, for example via a drip or an infusion pump). For example, the ready-to-use pharmaceutical composition as described herein may be administered to a subject from 8 hours to 1 week apart, preferably from 8 hours to 48 hours apart, preferably from 12 hours to 48 hours apart, preferably from 24 hours to 48 hours apart, more preferably from 8 hours to 24 hours apart, more preferably from 12 hours to 24 hours apart, even more preferably from 8 hours to 14 hours apart, better from 12 hours to 14 hours apart, still better about 12 hours.

[0054] “Drug substance” or “purified nangibotide substance” is a powder comprising or consisting of nangibotide. For example, the purified nangibotide substance may be obtained by the process described in part “Process for manufacturing purified nangibotide substance” of the present application. Preferably, the purified nangibotide substance comprises at least 80% w / w, preferably of at least 85% w / w, more preferably of at least 90% w / w, even more preferably at least 95% w / w, of all peptides, by weight relative to the total weight of the purified nangibotide substance. Preferably, the purified nangibotide substance comprises at least 80% w / w, preferably of at least 85% w / w, more preferably of at least 90% w / w, even more preferably at least 95% w / w, of nangibotide, by weight relative to the total weight of the purified nangibotide substance. Preferably, the purified nangibotide substance comprisesat least 80% w / w, preferably of at least 85% w / w, more preferably of at least 90% w / w, even more preferably at least 95% w / w, of nangibotide, by weight relative to the total weight of all peptides in said purified nangibotide substance. Preferably, the purified nangibotide substance comprises at least 85% w / w of all peptides by weight relative to the total weight of the purified nangibotide substance and at least 95% of nangibotide by weight relative to the total weight of all peptides in said purified nangibotide substance. Advantageously, the humidity content of the purified nangibotide substance is of equal or less than 10.0% w / w, preferably equal or less than 5.0 %w / w.

[0055] “Dry weight” refers to the weight excluding the water weight (in particular, excluding the humidity weight).

[0056] “Excipient” refers to any inactive ingredient, which is required for the formulation of an API in a suitable dosage form.

[0057] “From X to Y” refers to the range of values between X and Y, the limits X and Y being included in said range.

[0058] Numerical values are subject to measurement errors which place limits on their accuracy, as known by the skilled person. Thus, in the present application, the general convention in the scientific and technical literature is applied: the last decimal place of a numerical value indicates its degree of accuracy. Where no other error margins are given, the maximum margin is ascertained by applying the rounding-off convention to the last decimal place.

[0059] “Infectious disease” refers to a pathological condition or disorder resulting from an infection. Examples of infectious diseases include bacterial diseases (or bacterial infections), viral diseases (or viral infections), fungal diseases (or fungal infections), and parasitic diseases (or parasitic infections), which are infectious diseases caused by bacteria, viruses, fungi, and parasites, respectively.

[0060] “Met(O)12-LR12” is a peptide whose amino acid sequence is SEQ ID NO: 3 (LQEED AGEYGCM) in which the methionine at position 12 is oxidized into methionine sulfoxide also referred to as Met(O) or MetO, and whose chemical formula is the following formula (IV):Formula (IV).

[0061] “Nangibotide” or “nangibotide peptide” refers to the peptide having the INN Request No. 10637 (and CAS number 2014384-91-7). “Nangibotide” is also referred to as “LR12” or LR12 peptide . The ammo acid sequence of nangibotide is as set forth in SEQ ID NO: 1 (LQEED AGEYGCM), wherein the C-terminal methionine has been ami dated, and its chemical formula is the following formula (I):Formula (I).

[0062] “Nangibotide dimer” or “dimeric nangibotide” refers to a molecule composed of two nangibotide monomers covalently bound by a disulfide bond through the cysteine at position 11. Indeed, the sequence of nangibotide contains a cysteine, which leads to the formation of disulfide bridges between two molecules of nangibotide, creating nangibotidedimers. Nangibotide dimers were found to be inactive in a preclinical in vitro pharmacology model and thus they can negatively impact the nangibotide treatment efficacy. The chemical formula of a nangibotide dimer is the following formula (II):Formula (II).

[0063] “Osmolality” refers to an overall measure of the contribution of the various solutes present in a solution to the osmotic pressure of the solution. For example, osmolality may be determined by measurement of the depression of freezing point using an osmometer.

[0064] “pH” refers to the potential of hydrogen in a solution. According to the present invention, the pH of a powder, such as a lyophilized composition, may be measured after dissolving or suspending 1 gram of the powder in 10 mb of distilled water. According to the present invention, the pH is preferably measured at room temperature using a pH meter or a pH paper, more preferably the pH is measured at room temperature using a pH meter. A pH meter is an electronic device which determines the pH of a solution. Advantageously, a pH meter may be a potentiometric pH meter. Unless indicated otherwise, all pH values in the present application refer to pH values measured at room temperature, i.e., preferably at a temperature of20°C.

[0065] “Pharmaceutically acceptable” refers to an excipient or carrier or buffer that does not produce an adverse, allergic or other untoward reaction when administered to a mammal, such as a human. It includes for example any and all solvents, such as, for example, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents. In particular, for human administration, pharmaceutical compositions should meet sterility, pyrogenicity, general safety and purity standards as required by the regulatory offices such as the EMA (European Medicines Agency) or FDA (US Food and Drug Administration).

[0066] “Pharmaceutical composition” refers to the combination of at least one API and at least one pharmaceutically acceptable excipient.

[0067] “Powder” refers to a set of dry and more or less fine solid (micro)particles. Advantageously, the humidity content of any powder as described herein is of equal or less than 10.0% w / w. For example, the humidity content may be about 9.0% w / w or less, or about 8.0% w / w or less, or about 7.0% w / w or less, or about 6.0% w / w or less, or about 5.0% w / w or less, or 4.0% w / w or less, or about 3.0% w / w or less, or about 2.0% w / w or less, or about 1.0% w / w or less. “Humidity content” refers to the water content, for example in a powder.

[0068] “Purity” or “nangibotide purity” refers to percent of nangibotide in a composition by weight in comparison to the total weight of all peptides in the composition. The peptide purity may be determined by any method known by the man skilled in the art, including but not limited to, liquid chromatography (LC) and elemental nitrogen analysis. LC is employed to quantify the relative proportion of the peptide of interest to related impurities based on chromatographic peak areas, while elemental nitrogen analysis determines the peptide content (i.e., the content of all peptides) as a fraction of the total sample weight by measuring the nitrogen content and correlating it to the theoretical nitrogen content of the peptide. Peptide content may be determined by any means known in the art, for example by a method selected from the group consisting of: elemental analysis, quantitative amino acid analysis,and the Kjeldahl method. Preferably, the peptide content and thus the nangibotide purity of a composition may be determined by elemental nitrogen analysis.

[0069] “Qsp” refers to “quantity sufficient for” or “quantum satis”. The specification of quantum satis for an ingredient essentially means “add as much of this ingredient as is needed to achieve the desired result, but not more.”

[0070] “Ready- to-use pharmaceutical composition” refers to a medication that is prepared and can be administered to a patient directly without the need for further preparation or dilution. For example, a ready to use pharmaceutical composition comprising nangibotide may be prepared as follows: preparing a purified nangibotide substance (drug substance) > obtaining the purified nangibotide substance (drug substance) (powder) > preparing a pharmaceutical composition comprising the purified nangibotide substance (drug product) > obtaining the pharmaceutical composition comprising nangibotide (drug product) (powder) > preparing a reconstituted pharmaceutical composition > obtaining the reconstituted pharmaceutical composition (liquid) > further diluting the reconstituted pharmaceutical composition > obtaining the ready-to-use pharmaceutical composition (liquid) .

[0071] “Reconstituted pharmaceutical composition” refers to the pharmaceutical composition comprising nangibotide (drug product) that is reconstituted in a pharmaceutically acceptable aqueous solution.

[0072] “Room temperature” refers to a temperature ranging from 20°C to 25°C, preferably a temperature of 20°C.

[0073] “Rpm” or “revolutions per minute” refers to the number of turns in one minute.

[0074] “Sepsis” is defined as a life-threatening organ dysfunction caused by a dysregulated response of the human subject to infection (Singer et al., JAMA. 2016 Feb 23;315(8):801-10). Patients with sepsis may be clinically identified as patients suffering from a documented or suspected infection and from an organ dysfunction. Organ dysfunction in a human subject may be identified using an organ dysfunction score, notablyupon admission of the human subject in ICU or emergency unit. Examples of organ dysfunction scores include, without being limited to, the SOFA (Sequential Organ Failure Assessment) score, the qSOFA (quick SOFA) score, the MODS (Multiple Organ Dysfunction Score), the P-MODS (Pediatric Multiple Organ Dysfunction Score) and the LODS (Logistic Organ Dysfunction System). For example, organ dysfunction can be represented by an increase in the SOFA score of 2 points or more.

[0075] “Septic shock” is defined as a subset of sepsis, in which particularly profound circulatory, cellular, and metabolic abnormalities are associated with a greater risk of mortality than with sepsis alone (Singer et al., JAMA. 2016 Feb 23;315(8):801-l 0). Thus, sepsis encompasses septic shock. Patients with septic shock may be clinically identified as patients suffering from sepsis and having (i) persisting hypotension requiring vasopressors to maintain their mean arterial pressure >65 mm Hg despite adequate volume resuscitation and (ii) a serum lactate level >2 mmol / L (18mg / dL) (Sepsis-3 definition as described in Singer et al., JAMA. 2016 Feb 23;315(8):801 - 10).

[0076] “SIRS” (systemic inflammatory response syndrome) is characterized by systemic inflammation and widespread tissue injury. Clinically, SIRS may be defined as fulfilling at least two of the following four criteria: fever > 38.0°C or hypothermia < 36.0°C; tachycardia > 90 beats / minute; tachypnea > 20 breaths / minute; and leukocytosis > 12*109 / L or leucopenia < 4xlO9 / L (Bone et al., Chest. 1992 Jun;101(6):1644-55). SIRS may occur as a response to a nonspecific insult of either infectious or non-infectious origin. Examples of insults of non-infectious origin include, without being limited to, trauma, thermal injury, pancreatitis, autoimmune disorders and surgery. Examples of insults of infectious origin include bacterial infections (e.g., respiratory infections, abdominal infections and urinary tract infections (UTIs), fungal infections (e.g., respiratory infections) and viral infections (e.g., respiratory infections).

[0077] “Subject” refers to a warm-blooded animal, more preferably a mammal. The term “mammal” refers here to any mammal, including humans. A subject may be a “patient”,who / which is awaiting the receipt of, or is receiving medical care or was / is / will be the object of a medical procedure, or is monitored for the development of the targeted disease or condition. In one example, the subject may be a human.

[0078] “Therapeutically effective amount” or “effective amount” of an API or of a pharmaceutical composition refers to a nontoxic but sufficient amount of said API or pharmaceutical composition to provide the desired therapeutic effect.

[0079] “Total weight of all peptides” in a composition refers to the sum of the weights of all the peptides present in the composition. In the present application, the term “all peptides” refers to nangibotide, along with any derivatives of nangibotide that may be present including dimeric nangibotide, Des-Gln-LR12, and Met(O)12-LR12.

[0080] “Treatment” refers to a therapeutic (or curative) treatment, to a prophylactic (or preventative) treatment, or to both a therapeutic (or curative) treatment and a prophylactic (or preventative) treatment, wherein the object is to prevent, reduce, slow down (lessen), or cure one or more of the symptom(s) or manifestation(s) of the targeted pathologic condition or disorder, such as an inflammatory disease or disorder, in particular septic shock, sepsis or SIRS, a cardiovascular disease, disorder or condition, or an infectious disease. Those in need of treatment include those already suffering from an inflammatory disease or disorder, a cardiovascular disease, disorder or condition, or an infectious disease, as well as those prone to develop an inflammatory disease or disorder, a cardiovascular disease, disorder or condition, or an infectious disease, or those in whom an inflammatory disease or disorder, a cardiovascular disease, disorder or condition, or an infectious disease is to be prevented.

[0081] “Water for injection” refers to water that has been purified through a process comprising steps of prefiltration, softening, deionization and distillation, that is sterile and nonpyrogenic. These steps may be carried out by any method known by the man skilled in the art. For example, distillation may be carried out by vapor compression distillation, multiple effect distillation, or reverse osmosis + ultrafiltration. Water for injection is sterile and nonpyrogenic.

[0082] It should be understood that the expression “at least one of’ includes individually each of the recited objects after the expression and the various combinations of two or more of the recited objects unless otherwise understood from the context and use.

[0083] The use of the term “include,” “includes,” “including,” “have,” “has,” “having,” “contain,” “contains,” or “containing,” including grammatical equivalents thereof, should be understood generally as open-ended and non-limiting, for example, not excluding additional unrecited elements or steps, unless otherwise specifically stated or understood from the context.

[0084] The use of any and all examples, or exemplary language herein, for example, “such as” or “including,” is intended merely to illustrate better the present invention and does not pose a limitation on the scope of the invention unless claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the present invention.

[0085] Throughout the description, where compositions are described as having, including, or comprising specific components, or where processes and methods are described as having, including, or comprising specific steps, it is contemplated that, additionally, there are compositions of the present invention that consist essentially of, or consist of, the recited components, and that there are processes and methods according to the present invention that consist essentially of, or consist of, the recited processing steps.

[0086] In the application, where an element or component is said to be included in and / or selected from a list of recited elements or components, it should be understood that the element or component can be any one of the recited elements or components, or the element or component can be selected from a group consisting of two or more of the recited elements or components.

[0087] It should be understood that the order of steps or order for performing certain actions is immaterial so long as the present invention remains operable. Moreover, two or more steps or actions may be conducted simultaneously.

[0088] Further, it should be understood that elements and / or features of a composition or a method described herein can be combined in a variety of ways without departing from the spirit and scope of the present invention, whether explicit or implicit herein. For example, where reference is made to a particular compound, that compound can be used in various embodiments of compositions of the present invention and / or in methods of the present invention, unless otherwise understood from the context. In other words, within this application, embodiments have been described and depicted in a way that enables a clear and concise application to be written and drawn, but it is intended and will be appreciated that embodiments may be variously combined or separated without parting from the present teachings and invention(s). For example, it will be appreciated that all features described and depicted herein can be applicable to all aspects of the invention(s) described and depicted herein.DETAILED DESCRIPTION

[0089] Disclosed herein compositions that are designed to optimize efficacy of nangibotide. Applicant has observed that, under certain conditions, nangibotide can form dimers. These dimers are biologically inactive and their formation reduces the amount of active nangibotide monomer in a pharmaceutical composition. Accordingly, the presence of nangibotide dimers has the potential to reduce the therapeutic efficacy of a nangibotide therapeutic administration scheme. Applicant believes that nangibotide’s propensity for dimerization and concomitant negative effect on the treatment efficacy due to biological inactivity was heretofore unknown. Accordingly, disclosed herein are nangibotide compositions that minimize nangibotide dimer content, thereby maintaining therapeutic efficacy of the compositions.

[0090] In addition, in order to administer a sufficient therapeutic amount of nangibotide parenterally, e.g., by injection or intravenous administration, nangibotide must be solubilized in a liquid pharmaceutical vehicle such as saline water or water for injection. The nangibotide compositions disclosed herein are designed to take advantage of current understanding of the appropriate use of fluids in intensive care units (ICUs) and hospitals. For example, the compositions disclosed herein ensure a therapeutically effective dose of nangibotide by combining high concentrations of nangibotide in appropriately low volumes of liquid, thereby reducing the potential risks of volume overload and / or osmotic shock on organ failure and mortality, in particular in vulnerable subjects such as subjects suffering from septic shock or systemic inflammatory response syndrome. In order to comply with clinical prerequisites with regard to the fluid administration and osmotic pressure compliance, particularly for subjects vulnerable to volume overload and / or osmotic shock, the nangibotide pharmaceutical compositions disclosed herein can be prepared into a ready-to-use pharmaceutical composition wherein nangibotide presents an optimal solubility and wherein said formulation comprises the minimal possible amount of excipients and an optimal osmolality. For example, the nangibotide pharmaceutical compositions disclosed herein can be reconstituted and diluted into a ready-to-use pharmaceutical composition having a high concentration of nangibotide, allowing the administration to a patient of a volume of liquid parenteral formulation which is as low as possible, while also having a low nangibotide dimer content and an optimal osmolality.

[0091] The inventors of the present application have surprisingly discovered that a pharmaceutical composition comprising at least one amino acid, for example, such as arginine, lysine or histidine, allows said pharmaceutical composition to optimize nangibotide administration, with a high amount of nangibotide being efficiently solubilized and having a suitable stability, and the composition being compliant with regard to osmolality prerequisites. For example, the amount of the at least one amino acid may be present in the formulation in an amount of from 22% to 34% by weight relative to the weight of nangibotide in the pharmaceutical composition.Pharmaceutical composition comprising nangibotide (drug product)

[0092] The present invention relates to a pharmaceutical composition comprising nangibotide and at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixtures thereof. Such pharmaceutical composition may be referred to as “drug product”.

[0093] The present invention also relates to a pharmaceutical composition comprising nangibotide and at least one amino acid selected from the group consisting of: arginine, lysine histidine and mixtures thereof, wherein the amount of the at least one amino acid is of at least 10% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 10% to 30% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 10% to 15% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 10% to 30% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 20% to 30% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 20% to 25% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 25% to 30% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 22% to 34% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 27% to 29% by weight relative to the weight of nangibotide.

[0094] The present invention also relates to a pharmaceutical composition comprising nangibotide and at least one amino acid selected from the group consisting of: arginine, lysine histidine and mixtures thereof, wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 22% to 50% by weight relative to the weightof nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 22% to 40% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 22% to 35% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 22% to 34% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 22% to 32% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 22% to 30% by weight relative to the weight of nangibotide. For example, the amount of the at least one amino acid or mixture thereof ranges from 23% to 29% by weight relative to the weight of nangibotide. In some embodiments, the amount of the at least one amino acid in the pharmaceutical composition is of 22% to 34% or of 25% to 30% by weight relative to the weight of nangibotide.

[0095] In some embodiments, the amount of the at least one amino acid or mixture thereof is of at least 15% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the amount of the at least one amino acid or mixture thereof ranges from 15% to 40% by weight relative to the dry weight of the pharmaceutical composition. For example, the amount of the at least one amino acid or mixture thereof ranges from 15% to 30% by weight relative to the dry weight of the pharmaceutical composition. For example, the amount of the at least one amino acid or mixture thereof ranges from 15% to 25% by weight relative to the dry weight of the pharmaceutical composition. For example, the amount of the at least one amino acid or mixture thereof ranges from 15% to 20% by weight relative to the dry weight of the pharmaceutical composition.

[0096] In some embodiments, the amount of the at least one amino acid or mixture thereof is of at least 15% by weight relative to the total weight of the pharmaceutical, preferably the amount of the at least one amino acid or mixture thereof ranges from 15% to 40%, more preferably from 15% to 30%, even more preferably from 15% to 25%, better from 15% to 20% by weight relative to the total weight of the pharmaceutical composition.

[0097] The inventors have surprisingly discovered that such a ratio of the weight of the at least one amino acid selected from the group consisting of: arginine, lysine, histidine and mixtures thereof, to the weight of nangibotide, in a pharmaceutical composition, allows said pharmaceutical composition to have a high concentration of nangibotide which is efficiently solubilized and has a suitable stability, a low amount of nangibotide dimers, and to be formulated with minimal excipients while maintaining an optimal osmolality. In particular, such ratio allows the pharmaceutical composition to comprise equal to or less than 5.0% of nangibotide dimers by weight relative to the total weight of all peptides in said pharmaceutical composition comprising nangibotide (also referred to as drug product).

[0098] Thus, a pharmaceutical composition according to the invention allows nangibotide to be chemically stable. According to a particularly advantageous aspect, such chemical stability is maintained throughout the shelf-life of 36 months of the pharmaceutical composition according to the invention.

[0099] In some embodiments, the at least one amino acid is selected from the group consisting of: arginine, lysine and any mixture thereof.

[0100] In some embodiments, the at least one amino acid is selected from the group consisting of: arginine, histidine and any mixture thereof.

[0101] In some embodiments, the at least one amino acid is selected from the group consisting of: lysine, histidine and any mixture thereof.

[0102] In some embodiments, the at least one amino acid is arginine.

[0103] In some embodiments, the at least one amino acid is lysine.

[0104] In some embodiments, the at least one amino acid is histidine.

[0105] In some embodiments, the pH of said pharmaceutical composition ranges from 4.5 to 8.5. In some embodiments, the pH of said pharmaceutical composition ranges from 4.5 to 7.4, preferably from 5 to 7, more preferably from 5 to 6, even more preferably the pH of saidpharmaceutical composition is of about 5.5, 5.6 or 5.7. In some embodiments, the pH of said pharmaceutical compositions is between 5.5 to 6.5.

[0106] The pH of the pharmaceutical composition applies whether the pharmaceutical composition is a powder or a liquid (for example, a solution). The pH may be measured using standard techniques known to the person skilled in the art, for example as described in part “Definitions” of the present application.

[0107] In some embodiments, said pharmaceutical composition further comprises at least one pharmaceutically acceptable buffer.

[0108] In some embodiments, the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES (2-(N-morpholino)ethanesulfonic acid) buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof. For example, the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES (2-(N-morpholino)ethanesulfonic acid) buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof. For example, the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0109] In some embodiments, the at least one pharmaceutically acceptable buffer is citrate buffer. For example, the citrate buffer may be sodium citrate buffer. Citrate buffer may be prepared by mixing a citrate salt such as sodium citrate with citric acid, thereby leading to a buffer of citrate / citric acid.

[0110] In some embodiments, the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES (2-(N-morpholino)ethanesulfonic acid) buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof. For example, the at least one pharmaceutically acceptable buffer is selected from the group consisting of:succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES (2-(N- morpholino)ethanesulfonic acid) buffer, phosphate buffer, tromethamine buffer and any mixtures thereof. For example, the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof. Preferably, the pharmaceutical composition does not comprise citrate buffer.

[0111] In some embodiments, the at least one pharmaceutically acceptable buffer is at least one pharmaceutically acceptable binary buffer. Preferably, said pharmaceutically acceptable binary buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES (2-(N-morpholino)ethanesulfonic acid) buffer, tromethamine buffer and any mixtures thereof. Preferably, the pharmaceutical composition does not comprise citrate buffer.

[0112] In some embodiments, the at least one pharmaceutically acceptable buffer is an acidic buffer, preferably an acidic buffer selected from the group consisting of: acetate buffer, succinate buffer, lactic acid buffer, adipic acid buffer, glutamate buffer, citrate buffer and mixtures thereof.

[0113] In some embodiments, the at least one pharmaceutically acceptable buffer is a neutral buffer, preferably a neutral buffer selected from the group consisting of: MES buffer, histidine buffer, phosphate buffer and mixtures thereof.

[0114] In some embodiments, the at least one pharmaceutically acceptable buffer is an alkaline buffer, preferably an alkaline buffer which is tromethamine buffer.

[0115] In some embodiments, the at least one pharmaceutically acceptable buffer is a histidine buffer.

[0116] In some embodiments, the at least one pharmaceutically acceptable buffer is succinate buffer.

[0117] In some embodiments, the at least one pharmaceutically acceptable buffer is an acetate buffer.

[0118] In some embodiments, the at least one pharmaceutically acceptable buffer is a tromethamine buffer.

[0119] In some embodiments, the at least one pharmaceutically acceptable buffer is a phosphate buffer.

[0120] In some embodiments, the at least one amino acid is arginine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof.

[0121] In some embodiments, the at least one amino acid is arginine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof. In some embodiments, the at least one amino acid is arginine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer and any mixtures thereof. Preferably, the pharmaceutical composition does not comprise citrate buffer.

[0122] In some embodiments, the at least one amino acid is arginine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0123] In some embodiments, the at least one amino acid is arginine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer,acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0124] In some embodiments, the at least one amino acid is arginine and the at least one pharmaceutically acceptable buffer is at least one pharmaceutically acceptable binary buffer. Preferably, said pharmaceutically acceptable binary buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, tromethamine buffer and any mixtures thereof.

[0125] In some embodiments, the at least one amino acid is arginine and the at least one pharmaceutically acceptable buffer is an acidic buffer, preferably an acidic buffer selected from the group consisting of: acetate buffer, succinate buffer, lactic acid buffer, adipic acid buffer, glutamate buffer, citrate buffer and mixtures thereof.

[0126] In some embodiments, the at least one amino acid is arginine and the at least one pharmaceutically acceptable buffer is a neutral buffer, preferably a neutral buffer selected from the group consisting of: MES buffer, histidine buffer, phosphate buffer and mixtures thereof.

[0127] In some embodiments, the at least one amino acid is arginine and the at least one pharmaceutically acceptable buffer is an alkaline buffer, preferably an alkaline buffer which is tromethamine buffer.

[0128] In some embodiments, the at least one amino acid is lysine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof.

[0129] In some embodiments, the at least one amino acid is lysine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof. In someembodiments, the at least one amino acid is lysine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer and any mixtures thereof. Preferably, the pharmaceutical composition does not comprise citrate buffer.

[0130] In some embodiments, the at least one amino acid is lysine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0131] In some embodiments, the at least one amino acid is lysine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0132] In some embodiments, the at least one amino acid is lysine and the at least one pharmaceutically acceptable buffer is at least one pharmaceutically acceptable binary buffer. Preferably, said pharmaceutically acceptable binary buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, tromethamine buffer and any mixtures thereof.

[0133] In some embodiments, the at least one amino acid is lysine and the at least one pharmaceutically acceptable buffer is an acidic buffer, preferably an acidic buffer selected from the group consisting of: acetate buffer, succinate buffer, lactic acid buffer, adipic acid buffer, glutamate buffer, citrate buffer and mixtures thereof.

[0134] In some embodiments, the at least one amino acid is lysine and the at least one pharmaceutically acceptable buffer is a neutral buffer, preferably a neutral buffer selected from the group consisting of: MES buffer, histidine buffer, phosphate buffer and mixtures thereof.

[0135] In some embodiments, the at least one amino acid is lysine and the at least one pharmaceutically acceptable buffer is an alkaline buffer, preferably an alkaline buffer which is tromethamine buffer.

[0136] In some embodiments, the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof.

[0137] In some embodiments, the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof. In some embodiments, the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer and any mixtures thereof. Preferably, the pharmaceutical composition does not comprise citrate buffer.

[0138] In some embodiments, the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0139] In some embodiments, the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0140] In some embodiments, the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is at least one pharmaceutically acceptable binary buffer.Preferably, said pharmaceutically acceptable binary buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, tromethamine buffer and any mixtures thereof.

[0141] In some embodiments, the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is an acidic buffer, preferably an acidic buffer selected from the group consisting of: acetate buffer, succinate buffer, lactic acid buffer, adipic acid buffer, glutamate buffer, citrate buffer and mixtures thereof.

[0142] In some embodiments, the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is a neutral buffer, preferably a neutral buffer selected from the group consisting of: MES buffer, histidine buffer, phosphate buffer and mixtures thereof.

[0143] In some embodiments, the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is an alkaline buffer, preferably an alkaline buffer which is tromethamine buffer.

[0144] In some embodiments, the at least one amino acid is a mixture of at least two amino acids selected from the group consisting of: arginine, lysine, and histidine, and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof.

[0145] In some embodiments, the at least one amino acid is a mixture of at least two amino acids selected from the group consisting of: arginine, lysine, and histidine, and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof. In some embodiments, the at least one amino acid is a mixture of at least two amino acids selectedfrom the group consisting of: arginine, lysine, and histidine, and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer and any mixtures thereof. Preferably, the pharmaceutical composition does not comprise citrate buffer.

[0146] In some embodiments, the at least one amino acid is a mixture of at least two amino acids selected from the group consisting of: arginine, lysine, and histidine, and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0147] In some embodiments, the at least one amino acid is a mixture of at least two amino acids selected from the group consisting of: arginine, lysine, and histidine, and the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

[0148] In some embodiments, the at least one amino acid is a mixture of at least two amino acids selected from the group consisting of: arginine, lysine, and histidine, and the at least one pharmaceutically acceptable buffer is at least one pharmaceutically acceptable binary buffer. Preferably, said pharmaceutically acceptable binary buffer is selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, tromethamine buffer and any mixtures thereof.

[0149] In some embodiments, the at least one amino acid is a mixture of at least two amino acids selected from the group consisting of: arginine, lysine, and histidine, and the at least one pharmaceutically acceptable buffer is an acidic buffer, preferably an acidic buffer selected from the group consisting of: acetate buffer, succinate buffer, lactic acid buffer, adipic acid buffer, glutamate buffer, citrate buffer and mixtures thereof.

[0150] In some embodiments, the at least one amino acid is a mixture of at least two amino acids selected from the group consisting of: arginine, lysine, and histidine, and the at least one pharmaceutically acceptable buffer is a neutral buffer, preferably a neutral buffer selected from the group consisting of: MES buffer, histidine buffer, phosphate buffer and mixtures thereof.

[0151] In some embodiments, the at least one amino acid is a mixture of at least two amino acids selected from the group consisting of: arginine, lysine, and histidine, and the at least one pharmaceutically acceptable buffer is an alkaline buffer, preferably an alkaline buffer which is tromethamine buffer.

[0152] In some embodiments, the pharmaceutical composition further comprises at least one pharmaceutically acceptable buffer as described above, and: the amount of the at least one amino acid ranges from 22.0% to 34.0%, preferably from 22.0% to 25.0%, preferably from 25.0% to 30.0%, more preferably from 26.0% to 28.0%, even more preferably is about 27.6%, by weight relative to the weight of nangibotide, and / or the amount of the at least one pharmaceutically acceptable buffer ranges from 3.0% to 7.0%, preferably from 3.0% to 5.5%, preferably from 4.0% to 6.0%, more preferably from 5.0% to 6.0%, even more preferably is about 5.4%, by weight relative to the weight of nangibotide.

[0153] In some embodiments, the pharmaceutical composition comprises, by weight relative to the dry weight of the pharmaceutical composition: at least 65% nangibotide, about 15% to 25% arginine, less than 10% of the at least one pharmaceutically acceptable buffer as described herein, for example less than 10% of citrate buffer.

[0154] In some embodiments, the pharmaceutical composition comprises, by weight relative to the total weight of the pharmaceutical composition:at least 65% nangibotide, about 15% to 25% arginine, less than 10% of the at least one pharmaceutically acceptable buffer as described herein, for example less than 10% of citrate buffer.

[0155] For example, the pharmaceutical composition may comprise about 65% to about 85% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 65% to about 80% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 65% to about 70% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 70% to about 80% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 70% to about 75% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 68% to about 76% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 75% to about 85% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 15% to about 20% arginine (w / w). For example, the pharmaceutical composition may comprise about 20% to about 25% arginine (w / w). For example, the pharmaceutical composition may comprise about 18% to about 23% arginine (w / w). For example, the pharmaceutical composition may comprise about 2% to about 10% citrate buffer (w / w) (e.g., citric acid plus its conjugate base). For example, the pharmaceutical composition may comprise about 2% to about 8% citrate buffer (w / w). For example, the pharmaceutical composition may comprise about or about 3% to about 7% citrate buffer (w / w). For example, the pharmaceutical composition may comprise about 2% to about 6% citrate buffer (w / w). For example, the pharmaceutical composition may comprise about 3% to about 6% citrate buffer (w / w). For example, the pharmaceutical composition may comprise 72.6% nangibotide (w / w), 20% arginine (w / w), and 3.9% citrate buffer (w / w). For example, the pharmaceutical composition is a dry powder, e.g., a lyophilizate. The dry powder may have a residual water content of, for example, about 1% to about 4% (w / w), or 1% to about 3% (w / w).

[0156] In some embodiments, the pharmaceutical composition comprises equal to or less than 5.0% by weight (w / w) of the total pharmaceutical composition, e.g., dimeric nangibotide, Des-Gln-LR12, Met(O)12-LR12 or other impurities. For example, the pharmaceutical composition comprises 5.0% or less of dimeric nangibotide (w / w), e.g., less than 3.0%, or e.g., less than 2.5%. For example, the pharmaceutical composition comprises 2.0% or less of Des-Gln-LR12, Met(O)12-LR12 and other impurities (w / w).

[0157] In some embodiments, the pharmaceutical composition comprises, by weight relative to the dry weight of the pharmaceutical composition: at least 65%, preferably about 72.6%, of nangibotide, from 15% to 25%, preferably about 20.0%, of arginine, less than 10%, preferably about 3.9%, of citrate buffer (i.e., the sum of the weights of citric acid and citrate), equal to or less than 5.0%, preferably about 2.4% or less, of nangibotide dimer, equal to or less than 1.2%, preferably about 0.6% or less, of Des-Gln-LR12, equal to or less than 1.0%, preferably about 0.3% or less, of Met(O)12-LR12, equal to or less than 0.5%, preferably about 0.1% or less, of other impurities.

[0158] In some embodiments, the pharmaceutical composition comprises, by weight relative to the dry weight of the pharmaceutical composition: about 72.6% of nangibotide, about 20.0% of arginine, about 2.4% of dimeric nangibotide, about 3.9% of citrate buffer (i.e., the sum of the weights of citric acid and citrate), about 0.6% of Des-Gln-LR12,- about 0.3% of Met(O)12-LR12, about 0.1% of other impurities.For example, the pharmaceutical composition may be in the form of a dry powder, e.g., a lyophilizate.

[0159] In some embodiments, the pharmaceutical composition comprises, by weight relative to the total weight of the pharmaceutical composition: at least 65%, preferably about 71.3%, of nangibotide, from 15% to 25%, preferably about 19.7%, of arginine, less than 10%, preferably about 3.9%, of citrate buffer (i.e., the sum of the weights of citric acid and citrate), equal or less than 10.0%, preferably about 1.8%, of water, equal to or less than 5.0%, preferably about 2.4%, of nangibotide dimer, equal to or less than 1.2%, preferably about 0.6%, of Des-Gln-LR12, equal to or less than 1.0%, preferably about 0.3%, of Met(O)12-LR12, equal to or less than 0.5%, preferably about 0.1%, of other impurities.For example, the pharmaceutical composition may be in the form of a dry powder, e.g., a lyophilizate.

[0160] In some embodiments, the pharmaceutical composition comprises, by weight relative to the total weight of the pharmaceutical composition: about 65%-75%, preferably about 70-75%, of nangibotide, about 15% to 25%, preferably about 18-22%, of arginine, about 2%-5% of citrate buffer (i.e., the sum of the weights of citric acid and citrate), equal or less than 10.0%, preferably about 1.8%, of water, equal to or less than 5.0%, preferably about 2.4% or less, of nangibotide dimer, equal to or less than 1.2%, preferably about 0.6% or less, of Des-Gln-LR12, equal to or less than 1.0%, preferably about 0.3% or less, of Met(O)12-LR12, equal to or less than 0.5%, preferably about 0.1% or less, of other impurities.For example, the pharmaceutical composition may be in the form of a dry powder, e.g., a lyophilizate.

[0161] For example, the pharmaceutical composition may comprise:1200 mg of nangibotide;331 mg of arginine;65 mg of sodium citrate;30 mg of water;40 mg of nangibotide dimers;10 mg of Des-Gln-LR12;5 mg of Met(O)12-LR12; and2 mg of other impurities.For example, the pharmaceutical composition may be in the form of a dry powder, e.g., a lyophilizate. For example, the dry powder may be contained in a sterile vial, e.g., a 10 mL or a 20 mL vial, preferably a 20 mL vial. When the pharmaceutical composition is reconstituted with about 9.5 mL of liquid, e.g., 0.9% NaCl, the pharmaceutical composition comprises 120 mg / mL nangibotide, 33.1 mg / mL (190 mM) arginine, and 6.5 mg / mL sodium citrate. The pH of the reconstituted pharmaceutical composition may be from 5 to 6, preferably 5.5.

[0162] For example, the pharmaceutical composition may comprise:1100- 1300 mg of nangibotide;280-360 mg of arginine; and50-70 mg of sodium citrate.Optionally, the composition may further comprise no more than 40 mg of nangibotide dimers; no more than 10 mg of Des-Gln-LR12; no more than 5 mg of Met(O)12-LR12; and no more than 2 mg of other impurities.For example, the pharmaceutical composition may be in the form of a dry powder, e.g., a lyophilizate. For example, the dry powder may be contained in a sterile vial, e.g., a 10 mL or a 20 mL vial, preferably a 20 mL vial.

[0163] For example, the pharmaceutical composition may be a dry powder, that when reconstituted, comprises: 120 mg / mL-200 mg / mL nangibotide, 150 - 300 mM arginine, and 6-10 mg / mL sodium citrate. The pH may be in the range of 5 to 6, such as 5.5, 5.6, or 5.7, for example.

[0164] In some embodiments, the pharmaceutical composition comprises, by weight relative to the dry weight of the pharmaceutical composition: at least 65% nangibotide, about 15% to about 25% of an amino acid selected from arginine, lysine, or histidine or combinations thereof, about 1% to about 10% of the at least one pharmaceutically acceptable buffer as described herein, for example a pharmaceutically acceptable buffer selected from acetate, citrate, histidine, succinate, phosphate or tromethamine, or combinations thereof.

[0165] In some embodiments, the pharmaceutical composition comprises, by weight relative to the total weight of the pharmaceutical composition: at least 65% nangibotide, about 15% to about 25% of an amino acid selected from arginine, lysine, or histidine or combinations thereof, about 1% to about 10% of the at least one pharmaceutically acceptable buffer as described herein, for example a pharmaceutically acceptable buffer selected from acetate, citrate, histidine, succinate, phosphate or tromethamine, or combinations thereof.

[0166] For example, the pharmaceutical composition may comprise about 65% to about 85% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 65% to about 80% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 65% to about 70% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 70% to about 80% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 70% to about 75% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 68% to about 76% nangibotide (w / w). For example, the pharmaceutical composition may comprise about 75% to about 85% nangibotide (w / w).

[0167] For example, the pharmaceutical composition may comprise about 15% to about 20% of the amino acid(s) (w / w). For example, the pharmaceutical composition may comprise about 20% to about 25% of the amino acid(s) (w / w). For example, the pharmaceutical composition may comprise about 18% to about 23% of the amino acid(s) (w / w).

[0168] For example, the pharmaceutical composition may comprise about 2% to about 10% of the pharmaceutically acceptable buffer(s) (w / w) (e.g., the acid plus its conjugate base). For example, the pharmaceutical composition may comprise about 2% to about 8% of the pharmaceutically acceptable buffer(s) (w / w). For example, the pharmaceutical composition may comprise about or about 3% to about 7% of the pharmaceutically acceptable buffer(s) (w / w). For example, the pharmaceutical composition may comprise about 2% to about 6% of the pharmaceutically acceptable buffer(s) (w / w). For example, the pharmaceutical composition may comprise about 3% to about 6% of the pharmaceutically acceptable buffer(s) (w / w). In some embodiments, the pharmaceutically acceptable buffer is acetate. In some embodiments, the pharmaceutically acceptable buffer is citrate. In other embodiments, the pharmaceutically acceptable buffer is histidine. In yet other embodiments, the pharmaceutically acceptable buffer is succinate. In still other embodiments, the pharmaceutically acceptable buffer is phosphate. In still further embodiments, the pharmaceutically acceptable buffer is tromethamine.

[0169] For example, the pharmaceutical composition may comprise about 72.6% nangibotide (w / w), about 20% arginine (w / w), and about 3.9% citrate buffer (w / w).

[0170] For example, the pharmaceutical composition may comprise about 71% nangibotide (w / w), about 19.7% arginine (w / w), and about 3.9% citrate buffer (w / w).

[0171] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% arginine (w / w), and about 2-5% citrate buffer (w / w).

[0172] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% arginine (w / w), and about 2-10%, e.g., 2- 5% histidine buffer (w / w).

[0173] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% arginine (w / w), and about 2-10%, e.g., 2- 5% succinate buffer (w / w).

[0174] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% arginine (w / w), and about 2-10%, e.g., 2- 5% acetate buffer (w / w).

[0175] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% arginine (w / w), and about 2-10%, e.g., 2- 5% phosphate buffer (w / w).

[0176] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% arginine (w / w), and about 2-10%, e.g., 2- 5% tromethamine buffer (w / w).

[0177] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% lysine (w / w), and about 2-10%, e.g., 2-5% citrate buffer (w / w).

[0178] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% lysine (w / w), and about 2-10%, e.g., 2-5% histidine buffer (w / w).

[0179] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% lysine (w / w), and about 2-10%, e.g., 2-5% succinate buffer (w / w).

[0180] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% lysine (w / w), and about 2-10%, acetate buffer (w / w).

[0181] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% lysine (w / w), and about 2-10%, e.g., 2-5% phosphate buffer (w / w).

[0182] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% lysine (w / w), and about 2-10%, e.g., 2-5% tromethamine buffer (w / w).

[0183] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% histidine (w / w), and about 2-10%, e.g., 2- 5% citrate (w / w).

[0184] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% histidine (w / w), and about 2-10%, e.g., 2- 5% histidine buffer (w / w).

[0185] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% histidine (w / w), and about 2-10%, e.g., 2- 5% succinate buffer (w / w).

[0186] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% histidine (w / w), and about 2-10%, e.g., 2- 5% acetate buffer (w / w).

[0187] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% histidine (w / w), and about 2-10%, e.g., 2- 5% phosphate buffer (w / w).

[0188] For example, the pharmaceutical composition may comprise about 65-85%, e.g., about 70-80% nangibotide (w / w), about 15-25% histidine (w / w), and about 2-10%, e.g., 2- 5% tromethamine buffer (w / w).

[0189] For example, the pharmaceutical composition is a dry powder, e.g., a lyophilizate. The dry powder, e.g., the lyophilizate, may have a residual water content of, for example, about 1% to about 4% (w / w), or 1% to about 3% (w / w).

[0190] In some embodiments, the pharmaceutical composition comprises equal to or less than 5.0% of any derivatives of nangibotide by weight (w / w) of the total pharmaceutical composition, e.g., dimeric nangibotide, Des-Gln-LR12, Met(O)12-LR12 or other impurities. For example, the pharmaceutical composition comprises 5.0% or less of dimeric nangibotide (w / w), e.g., less than 3.0%, or e.g., less than 2.5%. For example, the pharmaceutical composition comprises 2.0% or less of Des-Gln-LR12, Met(O)12-LR12 and other impurities (w / w).

[0191] In some embodiments, the pharmaceutical composition, when reconstituted, comprises: nangibotide at a concentration of about 100 mg / mL to about 250 mg / mL, e.g., about 100 mg / mL to about 200 mg / mL, e.g., about 120 mg / ml to about 200 mg / mL, e.g., about 120 mg / mL, or e.g., 120 mg / mL to about 150 mg / mL, or about 200 mg / mL, and an amino acid selected from arginine, lysine, or histidine or a combination of thereof at a concentration of about 140 mM to about 300 mM, e.g., about 150 mM to about 250 mM, e.g., about 150 mM to about 300 mM, e.g., about 180 mM to about 200 mM, or e.g., about 300 mM; and a buffer.

[0192] The buffer may be selected from a citrate buffer, a histidine buffer, a succinate buffer, an acetate buffer, a phosphate buffer, and a tromethamine buffer. The buffer may be selected from a histidine buffer, a succinate buffer, an acetate buffer, a phosphate buffer, anda tromethamine buffer. In some instances, the buffer is not citrate. For example, the concentration of the buffer is from about 20 mM to about 60 mM, e.g., about 20 to about 30 mM or about 45 to about 60 mM. For example, the buffer is 20-30 mM citrate. For example, the buffer is about 20-30 mM histidine. For example, the buffer is about 45-55 mM acetate. For example, the buffer is about 45-55 mM succinate. For example, the buffer is about 45- 55 mM phosphate. For example, the buffer is about 45-60 mM tromethamine. In some embodiments, the buffer is not citrate.

[0193] The pH of the reconstituted pharmaceutical composition may range from 4.5 to 8.5. For example, the pH when using a citrate buffer, a succinate buffer, a histidine buffer or an acetate buffer may be in the range of 5.0 to 6.0 or 5.5 to 6.5, e.g., 5.3, e.g., 5.4, e.g., 5.5., e.g., 5.6, e.g., 5.7. In some embodiments, the buffer is not citrate. For example, when a phosphate buffer is used, the pH is about 6.0 to about 7.5, e.g., about 6.0, e.g., about 6.5, e.g., about 7.O., e.g., about 7.5. For example, when a tromethamine buffer is used, the pH is about 7.0 to about 8.5, e.g., about 7.0, e.g., about 7.5, e.g., about 8.0.

[0194] For example, the pharmaceutical composition is reconstituted from a lyophilizate in a vial, e.g., a 10 mb or a 20 mL vial, preferably a 20 mL vial, to form a reconstituted pharmaceutical composition in a liquid form. The reconstituted pharmaceutical composition may then be further diluted in order to be administered to the patient according to an appropriate dosing regimen.

[0195] In some embodiments, where the amino acid is histidine and the buffer is histidine, the total concentration of histidine in the formulation may range from 22% to 50%, preferably from 22% to 34%, more preferably from 22% to 30%, even more preferably from 23% to 29%, by weight relative to the weight of nangibotide.

[0196] Alternatively, in some embodiments, said pharmaceutical composition does not comprise a buffer.

[0197] Thus, the pharmaceutical composition as described herein comprises a high amount of nangibotide and stability characteristics for clinical administration while maintaining a minimal amount of excipients.

[0198] In some embodiments, the pharmaceutical composition comprises from 600 mg to 2500 mg of nangibotide, preferably from 800 mg to 2500 mg of nangibotide, more preferably from 1200 mg to 2000 mg of nangibotide. Otherwise expressed, in some embodiments, the pharmaceutical composition comprises from 600 mg to 2500 mg of nangibotide per unit dosage, preferably from 800 mg to 2500 mg of nangibotide per unit dosage, more preferably from 1200 mg to 2000 mg of nangibotide per unit dosage. Examples of unit dosage include vials. Thus, the pharmaceutical composition comprises a high amount of nangibotide, with a view to achieving the highest treatment efficacy.

[0199] In some embodiments, the pharmaceutical composition comprises about 600 mg, about 650 mg, about 700 mg, about 750 mg, about 800 mg, about 850 mg, about 900 mg, about 950 mg, about 1000 mg, about 1050 mg, about 1100 mg, about 1150 mg, about 1200 mg, about 1250 mg, about 1300 mg, about 1350 mg, about 1400 mg, about 1450 mg, about 1500 mg, about 1550 mg, about 1600 mg, about 1650 mg, about 1700 mg, about 1750 mg, about 1800 mg, about 1850 mg, about 1900 mg, about 1950 mg, about 2000 mg, about 2050 mg, about 2100 mg, about 2150 mg, about 2200 mg, about 2250 mg, about 2300 mg, about 2350 mg, about 2400 mg, about 2450 mg, or about 2500 mg, of nangibotide. Otherwise expressed, in some embodiments, the pharmaceutical composition comprises about 600 mg, about 650 mg, about 700 mg, about 750 mg, about 800 mg, about 850 mg, about 900 mg, about 950 mg, about 1000 mg, about 1050 mg, about 1100 mg, about 1150 mg, about 1200 mg, about 1250 mg, about 1300 mg, about 1350 mg, about 1400 mg, about 1450 mg, about 1500 mg, about 1550 mg, about 1600 mg, about 1650 mg, about 1700 mg, about 1750 mg, about 1800 mg, about 1850 mg, about 1900 mg, about 1950 mg, about 2000 mg, about 2050 mg, about 2100 mg, about 2150 mg, about 2200 mg, about 2250 mg, about 2300 mg, about 2350 mg, about 2400 mg, about 2450 mg, or about 2500 mg, of nangibotide per unit dosage.Examples of unit dosage include vials, such as a 10 mL or a 20 mL vial, preferably a 20 mL vial.

[0200] In some embodiments, the pharmaceutical composition comprises at least about 65% of nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises at least about 60% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises at least about 70% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises at least about 75% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises at least about 80% of nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises at least about 85% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises at least about 90% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises about 60% to about 85% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises about 65% to about 85% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises about 60% to about 70% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises about 65% to about 70% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises about 70% to about 80% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises about 70% to about 85% nangibotide by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises about 75% to about 90% nangibotide by weight relative to the dry weight of thepharmaceutical composition. Thus, the pharmaceutical composition comprises a high concentration of nangibotide and therefore a very low quantity of excipients, with a view to limiting the volume of pharmaceutical composition to be administered to the patient while achieving the highest treatment efficacy.

[0201] In some embodiments, the pharmaceutical composition comprises at least about 65%, preferably at least about 70%, more preferably at least about 75%, even more preferably at least about of 80%, better at least 85%, still better at least 90%, of nangibotide by weight relative to the total weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises at least about 90% of nangibotide by weight relative to the total weight of the pharmaceutical composition.

[0202] In some embodiments, the pharmaceutical composition comprises at least about 90% of nangibotide by weight relative to the total weight of all peptides in the pharmaceutical composition and equal to or less than about 5.0% of nangibotide dimer by weight relative to the total weight of all peptides in the pharmaceutical composition.

[0203] In some embodiments, the pharmaceutical composition comprises at least about 90% of nangibotide by weight relative to the total weight of all peptides in the pharmaceutical composition, equal to or less than about 5.0% of nangibotide dimer by weight relative to the total weight of all peptides in the pharmaceutical composition and equal to or less than about 5.0% of other peptidic impurities (i.e., other than nangibotide dimer) by weight relative to the total weight of all peptides in the pharmaceutical composition.

[0204] In some embodiments, the peptidic impurities other than nangibotide dimer may include Des-Gln-LR12 and Met(O)12-LR12.

[0205] In some embodiments, the pharmaceutical composition comprises at least about 90% of nangibotide by weight relative to the total weight of all peptides in the pharmaceutical composition, equal to or less than about 5.0% of nangibotide dimer by weight relative to the total weight of all peptides in the pharmaceutical composition, equal to or less than about5.0% of other peptidic impurities (i.e., other than nangibotide dimer) by weight relative to the total weight of all peptides in the pharmaceutical composition including equal to or less than about 1.0% of Met(O)12-LR12 by weight relative to the total weight of all peptides in the pharmaceutical composition and equal to or less than about 1.2% of Des-Gln-LR12 by weight relative to the total weight of all peptides in the pharmaceutical composition.

[0206] In some embodiments, the pharmaceutical composition comprises less than or equal to about 2.5%, less than or equal to about 2%, less than or equal to about 1.5%, less than or equal to about 1.2%, less than or equal to about 1%, less than or equal to about 0.5% or less than or equal to about 0.3%, of Des-Gln-LR12, by weight relative to the total weight of all peptides in the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises less than or equal to about 1.2%, preferably less than or equal to about 1.0%, more preferably less than or equal to about 0.5%, of Des-Gln-LR12, by weight relative to the total weight of all peptides in the pharmaceutical composition.

[0207] In some embodiments, the pharmaceutical composition comprises less than or equal to about 2.5%, less than or equal to about 2%, less than or equal to about 1.5%, less than or equal to about 1.2%, less than or equal to about 1%, less than or equal to about 0.75%, less than or equal to about 0.5%, or less than or equal to about 0.3%, of Met(O)12-LR12, by weight relative to the total weight of all peptides in the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises less than or equal to about 1.0%, preferably less or equal to about 0.75%, more preferably less than or equal to about 0.5%, of Met(O)12-LR12, by weight relative to the total weight of all peptides in the pharmaceutical composition.

[0208] Advantageously, the solubility of nangibotide in the pharmaceutical composition is at least about 250 mg / mL, at least about 225 mg / mL, at least about 200 mg / mL, at least about 175 mg / mL, or at least about 150 mg / mL, when reconstituted in aqueous solution. For example, the solubility of nangibotide in the pharmaceutical composition is at least about 120 mg / mL, when reconstituted in an aqueous solution. Advantageously, the solubility ofnangibotide in the pharmaceutical composition is of at least about 175 mg / mL when reconstituted in aqueous solution.

[0209] For example, the concentration of nangibotide in the pharmaceutical composition is about 100 mg / ml to about 250 mg / mL when reconstituted in an aqueous solution. For example, the concentration of nangibotide in the pharmaceutical composition is about 100 mg / ml to about 150 mg / mL when reconstituted in an aqueous solution. For example, the concentration of nangibotide in the pharmaceutical composition is about 150 to about 200 mg / mL when reconstituted in an aqueous solution. For example, the concentration of nangibotide is about 100 mg / mL to about 200 mg / mL when reconstituted in an aqueous solution. For example, the concentration of nangibotide is about 110 mg / mL to about 130 mg / mL when reconstituted in an aqueous solution. For example, the concentration of nangibotide is about 120 mg / mL when reconstituted in an aqueous solution. The pharmaceutical composition may be reconstituted from a lyophilizate, for example.

[0210] Nangibotide may be incorporated into the pharmaceutical composition also referred to as “drug product”, via a “purified nangibotide substance”. In some embodiments, said purified nangibotide substance is as disclosed in the part “Purified nangibotide substance (drug substance)” of the present application.

[0211] In some embodiments, the amount of the at least one amino acid ranges from 20% to 30% by weight relative to the dry weight of the purified nangibotide substance.

[0212] In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount ranging from 70.0% to 80.0%, preferably from 75.2% to 79.0%, more preferably about 77.3%, by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 60% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 65% by weight relative to the dry weight of the pharmaceutical composition. In someembodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 70% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 75% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 80% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 85% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 90% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 60% to about 85% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 65% to about 85% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 60% to about 70% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 65% to about 70% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 70% to about 80% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 70% to about 85% by weight relative to the dry weight of the pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount of about 75% to about 90% by weight relative to the dry weight of the pharmaceutical composition.

[0213] In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount ranging from 70.0% to 80.0%, preferably from 75.2% to 79.0%, more preferably about 77.3%, by weight relative to the total weight of the pharmaceutical composition.

[0214] In some embodiments, the pharmaceutical composition further comprises at least one pharmaceutically acceptable buffer as described herein and: the amount of the at least one amino acid ranges from 20.0% to 30.0%, preferably from 22% to 30%, preferably from 25% to 30%, preferably from 20% to 25%, preferably from 22.0% to 28.0%, more preferably from 22.3% to 27.6%, even more preferably is about 24.5%, by weight relative to the dry weight of the purified nangibotide substance, and / or the amount of the at least one pharmaceutically acceptable buffer ranges from 3.0% to 7.0%, preferably from 4.0% to 6.0%, preferably from 3% to 5%, more preferably from 4.5% to 5.5%, even more preferably is about 4.8%, by weight relative to the dry weight of the purified nangibotide substance.

[0215] In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount ranging from 70.0% to 80.0%, preferably from 75.2% to 79.0%, more preferably about 77.3% by weight relative to the dry weight of the pharmaceutical composition, and said pharmaceutical composition further comprises at least one pharmaceutically acceptable buffer, and: the amount of the at least one amino acid ranges from 20.0% to 30.0%, preferably from 22.0% to 28.0%, more preferably from 22.3% to 27.6%, even more preferably is about 24.5%, by weight relative to the dry weight of the purified nangibotide substance, and / or the amount of the at least one pharmaceutically acceptable buffer ranges from 3.0% to 7.0%, preferably from 4.0% to 6.0%, more preferably from 4.5% to 5.5%, even more preferably is about 4.8%, by weight relative to the dry weight of the purified nangibotide substance.

[0216] In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount ranging from 70.0% to 80.0%, preferably from 73.0% to 77.0%, more preferably about 75.2% by weight relative to the dry weight of the pharmaceutical composition, and said pharmaceutical composition further comprises at least one pharmaceutically acceptable buffer, and: the amount of the at least one amino acid ranges from 25.0% to 30.0%, preferably from 26.0% to 28.0%, more preferably from 27.0% to 28.0%, even more preferably is about 27.6%, by weight relative to the weight of nangibotide, and / or the amount of the at least one pharmaceutically acceptable buffer ranges from 3.0% to 7.0%, preferably from 4.0% to 6.0%, more preferably from 5.0% to 6.0%, even more preferably is about 5.4%, by weight relative to the weight of nangibotide.

[0217] In some embodiments, the pharmaceutical composition comprises the purified nangibotide substance in an amount ranging from 70.0% to 80.0%, preferably from 73.0% to 77.0%, more preferably about 75.2% by weight relative to the total weight of the pharmaceutical composition, and said pharmaceutical composition further comprises at least one pharmaceutically acceptable buffer, and: the amount of the at least one amino acid ranges from 25.0% to 30.0%, preferably from 26.0% to 28.0%, more preferably from 27.0% to 28.0%, even more preferably is about 27.6%, by weight relative to the weight of nangibotide, and / or the amount of the at least one pharmaceutically acceptable buffer ranges from 3.0% to 7.0%, preferably from 4.0% to 6.0%, more preferably from 5.0% to 6.0%, even more preferably is about 5.4%, by weight relative to the weight of nangibotide.

[0218] In certain embodiments, where the at least one amino acid is histidine and the at least one pharmaceutically acceptable buffer is a histidine buffer, the weight of histidine in the pharmaceutical composition may range from 22% to 50%, preferably from 22% to 34%, more preferably from 22% to 30%, even more preferably from 23% to 29%, by weight relative to the weight of nangibotide.

[0219] In some embodiments, the pharmaceutical composition is a powder. Preferably, the humidity content of the pharmaceutical composition which is in the form of a powder is equal to or less than 10.0% w / w, or equal to or less than 5.0% w / w, preferably equal to or less than 3.0% w / w. Preferably, the pharmaceutical composition is a lyophilized powder. Preferably, the pharmaceutical composition is a white to off-white powder; or a white or off-white powder.

[0220] In some embodiments, the pharmaceutical composition comprises equal to or less than 3.0% of water by weight relative to the total weight of said pharmaceutical composition.

[0221] The present invention also relates to a vial containing a pharmaceutical composition according to the invention. In some embodiments, a vial contains from about 600 mg to about 2500 mg of nangibotide. For example, the vial may be a 10 mb or a 20 mb vial, preferably a 20 mb vial.Reconstituted pharmaceutical composition

[0222] The present invention also relates to a reconstituted pharmaceutical composition comprising: the pharmaceutical composition, also referred to as “drug product”, which is described in part “Pharmaceutical composition comprising nangibotide (drug product)” of the present application; and an aqueous medium, preferably said aqueous medium being selected from the group consisting of: water (e.g., water for injection such as water for injection USP) and a pharmaceutically acceptable aqueous solution, more preferably said aqueous medium being a saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.

[0223] The reconstituted pharmaceutical composition is obtained by the reconstitution of the pharmaceutical composition, also referred to as “drug product”, which is described in part “Pharmaceutical composition comprising nangibotide (drug product)” of the presentapplication, in water or in a pharmaceutically acceptable aqueous solution. For example, the volume of the reconstituted pharmaceutical composition ranges from 9 mL to 11 mL, preferably it is about 10 mL. For example, the reconstitution of the pharmaceutical composition, in order to obtain the reconstituted pharmaceutical composition, is carried out in a vial, for example a 10 mL vial (i.e., a vial having a total volume of 10 mL) or a 20 mL vial (i.e., a vial having a total volume of 20 mL), preferably a 20 mL vial.

[0224] In some embodiments, all the features described above for the pharmaceutical composition referred to as “drug product” (part “Pharmaceutical composition comprising nangibotide (drug product)” of the present application) apply mutatis mutandis for the reconstituted pharmaceutical composition. In particular, all the features described above regarding the weight of nangibotide in the pharmaceutical composition, the amount of the at least one amino acid by weight relative to the weight of nangibotide, the amount of the at least one amino acid by weight relative to the weight and dry weight of the pharmaceutical composition, the pH, the presence or absence of at least one pharmaceutically acceptable buffer, the type of buffer, the amount of the at least one pharmaceutically acceptable buffer by weight relative to the weight of nangibotide, the amount of the at least one pharmaceutically acceptable buffer by weight relative to the weight and dry weight of the pharmaceutical composition, the weight of Des-Gln-LR12 and Met(O)12-LR12 relative to the total weight of all peptides in said pharmaceutical composition, apply mutatis mutandis for the reconstituted pharmaceutical composition.

[0225] In some embodiments, the reconstituted pharmaceutical composition has a pH ranging from 4.5 to 8.5, preferably from 4.5 to 7.4, more preferably from 5 to 7, even more preferably from 5 to 6, better of about 5.5, 5.6 or 5.7. For example, in certain embodiments, the pH of said reconstituted pharmaceutical composition ranges from 5.5 to 6.5.

[0226] Advantageously, the chemical stability of nangibotide is maintained in the reconstituted pharmaceutical composition according to the invention.

[0227] In particular, in some embodiments, the reconstituted pharmaceutical composition comprises equal to or less than 10.0% of nangibotide dimers by weight relative to the total weight of all peptides in said reconstituted pharmaceutical composition. A nangibotide dimer content of equal to or less than 10.0% by weight relative to the total weight of all peptides in said reconstituted pharmaceutical composition allows a sufficient therapeutic efficacy of the ready-to-use pharmaceutical composition obtained by the dilution of the reconstituted pharmaceutical composition. In particular, in some embodiments, the reconstituted pharmaceutical composition comprises equal to or less than 5.0% of nangibotide dimers by weight relative to the total weight of all peptides in said reconstituted pharmaceutical composition.

[0228] In some embodiments, the reconstituted pharmaceutical composition comprises: the purified nangibotide substance in an amount ranging from 70.0% to 80.0%, preferably from 75.2% to 79.0%, more preferably about 77.3%, by weight relative to the dry weight of the reconstituted pharmaceutical composition; at least one pharmaceutically acceptable buffer, in an amount ranging from 2.0% to 6.0%, preferably from 3.0% to 5.0%, more preferably about 3.7%, by weight relative to the dry weight of the reconstituted pharmaceutical composition, and at least one amino acid selected from the group consisting of: arginine, lysine and histidine, preferably arginine or lysine, more preferably arginine, in an amount ranging from 15.0% to 35.0%, preferably from 18.0% to 22.0%, more preferably about 19.0%, by weight relative to the dry weight of the reconstituted pharmaceutical composition.

[0229] In some embodiments, the reconstituted pharmaceutical composition comprises, by weight relative to the dry weight of the reconstituted pharmaceutical composition: at least 65%, preferably about 72.6%, of nangibotide, from 15% to 25%, preferably about 20.0%, of arginine, less than 10%, preferably about 3.9%, of citrate buffer (i.e., the sum of the weights of citric acid and citrate),equal to or less than 5.0%, preferably about 2.4%, of nangibotide dimer, equal to or less than 1.2%, preferably about 0.6%, of Des-Gln-LR12, equal to or less than 1.0%, preferably about 0.3%, of Met(O)12-LR12, equal to or less than 0.5%, preferably about 0.1%, of other impurities.

[0230] In some embodiments, the reconstituted pharmaceutical composition comprises, by weight relative to the dry weight of the reconstituted pharmaceutical composition: about 72.6% of nangibotide, about 20.0% of arginine, about 2.4% of dimeric nangibotide, about 3.9% of citrate buffer (i.e., the sum of the weights of citric acid and citrate), about 0.6% of Des-Gln-LR12,- about 0.3% of Met(O)12-LR12, about 0.1% of other impurities.

[0231] For example, the reconstituted pharmaceutical composition may comprise: about 120 mg / mL of nangibotide; about 33.1 mg / mL of arginine; about 6.5 mg / mL of sodium citrate; about 4 mg / mL of nangibotide dimers; about 1 mg / mL of Des-Gln-LR12; about 0.5 mg / mL of Met(O)12-LR12; and about 0.2 mg / mL of other impurities.

[0232] The present invention also relates to a vial containing a reconstituted pharmaceutical composition according to the invention. In some embodiments, a vial comprises from about 600 mg to about 2500 mg of nangibotide and an aqueous medium qsp 10 mb.Ready-to-use pharmaceutical composition

[0233] The present invention also relates to a ready-to-use pharmaceutical composition comprising: the pharmaceutical composition, also referred to as “drug product”, which is described in part “Pharmaceutical composition comprising nangibotide (drug product)” of the present application; and an aqueous medium, preferably said aqueous medium being selected from the group consisting of: water (e.g., water for injection USP) and a pharmaceutically acceptable aqueous solution, more preferably said aqueous medium being a saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.

[0234] The ready-to-use pharmaceutical composition is obtained by the dilution of the reconstituted pharmaceutical composition, in water or in a pharmaceutically acceptable aqueous solution. For example, the volume of the ready-to-use pharmaceutical composition ranges from 40 mb to 60 mb, preferably it is about 50 mb. For example, the dilution of the reconstituted pharmaceutical composition, in order to obtain the ready-to-use pharmaceutical composition, is carried out in a syringe or in an infusion bag, for example in a 50 mb syringe or in a 50 mb infusion bag (i.e., a syringe or in an infusion bag having a total volume of 50 mb).

[0235] In some embodiments, all the features described above for the pharmaceutical composition referred to as “drug product” (part “Pharmaceutical composition comprising nangibotide (drug product)” of the present application) apply mutatis mutandis for the ready- to-use pharmaceutical composition. In particular, all the features described above regarding the weight of nangibotide in the pharmaceutical composition, the amount of the at least one amino acid by weight relative to the weight of nangibotide, the amount of the at least one amino acid by weight relative to the dry weight of the pharmaceutical composition, the pH, the presence or absence of at least one pharmaceutically acceptable buffer, the type of buffer,the amount of the at least one pharmaceutically acceptable buffer by weight relative to the weight of nangibotide, the amount of the at least one pharmaceutically acceptable buffer by weight relative to the dry weight of the pharmaceutical composition, the weight of Des-Gln- LR12 and Met(O)12-LR12 relative to the total weight of all peptides in said pharmaceutical composition, apply mutatis mutandis for the ready-to-use pharmaceutical composition.

[0236] In some embodiments, the ready-to-use pharmaceutical composition has a pH ranging from 4.5 to 8.5, preferably from 4.5 to 7.4, more preferably from 5 to 7, even more preferably from 5 to 6, better of about 5.5, 5.6 or 5.7. For example, in certain embodiments, the pH of said ready-to-use pharmaceutical composition ranges from 5.5 to 6.5.

[0237] Advantageously, the chemical stability of nangibotide is maintained in the ready-to- use pharmaceutical composition according to the invention. More advantageously, the chemical stability of nangibotide is maintained during the intravenous administration to a patient of the ready-to-use pharmaceutical composition.

[0238] In particular, in some embodiments, the ready-to-use pharmaceutical composition comprises equal to or less than 10.0% of nangibotide dimers by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition. A nangibotide dimer content of equal to or less than 10.0% by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition allows a sufficient therapeutic efficacy of the ready-to-use pharmaceutical composition. In particular, in some embodiments, the ready-to- use pharmaceutical composition comprises equal to or less than 5.0% of nangibotide dimers by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition.

[0239] In some embodiments, the ready-to-use pharmaceutical composition comprises: the purified nangibotide substance in an amount ranging from 70.0% to 80.0%, preferably from 75.2% to 79.0%, more preferably about 77.3%, by weight relative to the dry weight of the ready-to-use pharmaceutical composition;at least one pharmaceutically acceptable buffer, in an amount ranging from 2.0% to 6.0%, preferably from 3.0% to 5.0%, more preferably about 3.7%, by weight relative to the dry weight of the ready-to-use pharmaceutical composition, and at least one amino acid selected from the group consisting of: arginine, lysine and histidine, preferably arginine or lysine, more preferably arginine, in an amount ranging from 15.0% to 35.0%, preferably from 18.0% to 22.0%, more preferably about 19.0%, by weight relative to the dry weight of the ready-to-use pharmaceutical composition.

[0240] In some embodiments, the ready-to-use pharmaceutical composition comprises, by weight relative to the dry weight of the ready-to-use pharmaceutical composition: at least 65%, preferably about 72.6%, of nangibotide, from 15% to 25%, preferably about 20.0%, of arginine, less than 10%, preferably about 3.9%, of citrate buffer (i.e., the sum of the weights of citric acid and citrate), equal to or less than 5.0%, preferably about 2.4%, of nangibotide dimer, equal to or less than 1.2%, preferably about 0.6%, of Des-Gln-LR12, equal to or less than 1.0%, preferably about 0.3%, of Met(O)12-LR12, equal to or less than 0.5%, preferably about 0.1%, of other impurities.

[0241] In some embodiments, the ready-to-use pharmaceutical composition comprises, by weight relative to the dry weight of the ready-to-use pharmaceutical composition: about 72.6% of nangibotide, about 20.0% of arginine, about 2.4% of dimeric nangibotide, about 3.9% of citrate buffer (i.e., the sum of the weights of citric acid and citrate), about 0.6% of Des-Gln-LR12,- about 0.3% of Met(O)12-LR12, about 0.1% of other impurities.

[0242] In some embodiments, the ready-to-use pharmaceutical composition presents an osmolality of equal to or less than 500 mOsm / kg, preferably an osmolality ranging from 300 to 500 mOsm / kg, more preferably an osmolality ranging from 300 to 400 mOsm / kg, even more preferably an osmolality ranging from 320 to 390 mOsm / kg, better an osmolality of about 380 mOsm / kg. Thus, the ready-to-use pharmaceutical composition has an optimal osmolality which complies with the clinical prerequisites to avoid volume overload and / or osmotic shock in a patient.

[0243] In the ready-to-use pharmaceutical composition, the ratio of the weight amount of the at least one amino acid selected from the group consisting of: arginine, lysine and histidine, preferably of arginine or lysine, more preferably of arginine, to the weight amount of nangibotide, allows an increase of the solubility of nangibotide in the ready-to-use pharmaceutical composition to at least 120 mg / mL in the aqueous medium and maintains a low content of nangibotide dimers of equal to or less than 10.0% of nangibotide dimers by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition.

[0244] More precisely, the content of nangibotide dimers in the ready-to-use pharmaceutical composition may be of equal to or less than 5.0% of nangibotide dimers by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition at the time of the manufacture of said ready-to-use pharmaceutical composition (for example via the process described in part “Process for manufacturing a ready-to-use pharmaceutical composition” of the present application). Then, the nangibotide dimers content may increase but at a slower rate than in compositions that do not allow to limit nangibotide dimers formation. Thus, about 12 hours but also about 24 hours or even about 48 hours, after the preparation of said ready-to-use pharmaceutical composition (for example via the process described in part “Process for manufacturing a ready-to-use pharmaceutical composition” of the present application), there may be equal to or less than 10.0% of nangibotide dimers by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition. Indeed, whatever the timing after the preparation of the ready-to-usepharmaceutical composition, the amount of nangibotide dimers in the ready-to-use pharmaceutical composition to be administered to the patients should never exceed 10% by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition. Therefore, the content of nangibotide dimers in the ready-to-use pharmaceutical composition may be of equal to or less than 5.0% of nangibotide dimers by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition, at the time of the preparation of said ready-to-use pharmaceutical composition, and of equal to or less than 10.0% of nangibotide dimers by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition, about 12 hours but also about 24 hours or even about 48 hours, after the preparation of said ready-to-use pharmaceutical composition.

[0245] In some embodiments, the ready-to-use pharmaceutical composition comprises at least one amino acid selected from the group consisting of: arginine, lysine and histidine, preferably arginine or lysine, more preferably arginine, at a concentration ranging from about 100 mM to about 250 mM.

[0246] The present invention also relates to a syringe containing a ready-to-use pharmaceutical composition according to the invention. In some embodiments, a syringe contains from about 600 to about 2500 mg of nangibotide, preferably from 800 mg to 2500 mg of nangibotide, more preferably from 1200 mg to 2000 mg of nangibotide. For example, in the syringe, nangibotide may be diluted in about 40 mb to about 60 mb of aqueous solution, preferably in about 50 mb of aqueous solution.

[0247] The present invention also relates to an infusion bag containing a ready-to-use pharmaceutical composition according to the invention. In some embodiments, an infusion bag contains from about 600 to about 2500 mg of nangibotide, preferably from 800 mg to 2500 mg of nangibotide, more preferably from 1200 mg to 2000 mg of nangibotide.Therapeutic uses

[0248] The present invention further relates to the pharmaceutical composition, the reconstituted pharmaceutical composition or the ready-to-use pharmaceutical composition, as described in the present application, for use as a medicament. The present invention also relates to a method of treating a disease by administering to a subject in need thereof an effective amount of the pharmaceutical composition, the reconstituted pharmaceutical composition or the ready-to-use pharmaceutical composition as described in the present application. The present invention also relates to the use of the pharmaceutical composition, the reconstituted pharmaceutical composition or the ready-to-use pharmaceutical composition as described in the present application for the manufacture of a medicament for the treatment of a disease in a subject in need thereof. The present invention also relates to the use of the pharmaceutical composition, the reconstituted pharmaceutical composition or the ready-to-use pharmaceutical composition as described in the present application for the treatment of a disease in a subject in need thereof.

[0249] The present invention further relates to the pharmaceutical composition, the reconstituted pharmaceutical composition or the ready-to-use pharmaceutical composition, as described in the present application, for use in the treatment of a disease selected from the group consisting of: inflammatory diseases, cardiovascular diseases, and infectious diseases, in a subject in need thereof. The present invention also relates to a method of treating a disease selected from the group consisting of: inflammatory diseases, cardiovascular diseases, and infectious diseases, by administering to a subject in need thereof an effective amount of the pharmaceutical composition, the reconstituted pharmaceutical composition or the ready-to-use pharmaceutical composition as described in the present application. The present invention also relates to the use of the pharmaceutical composition, the reconstituted pharmaceutical composition or the ready-to-use pharmaceutical composition as described in the present application for the manufacture of a medicament for the treatment of a disease selected from the group consisting of: inflammatory diseases, cardiovascular diseases, and infectious diseases, in a subject in need thereof. The present invention also relates to the useof the pharmaceutical composition, the reconstituted pharmaceutical composition or the ready-to-use pharmaceutical composition as described in the present application for the treatment of a disease selected from the group consisting of: inflammatory diseases, cardiovascular diseases, and infectious diseases, in a subject in need thereof.

[0250] In some embodiments, the disease to be treated is an inflammatory disease. Preferably, the inflammatory disease may be selected from the group consisting of: septic shock, sepsis, severe sepsis, systemic inflammatory response syndrome (SIRS), sepsis- associated organ dysfunction, acute respiratory distress syndrome (ARDS), severe acute respiratory syndrome (SARS), acute kidney injury (AKI), ischemia-reperfusion, pancreatitis, pneumonia, endotoxemia, and hemorrhagic shock. More preferably, the inflammatory disease may be selected from the group consisting of: septic shock, sepsis, severe sepsis, SIRS, ARDS, ischemia-reperfusion, pancreatitis and hemorrhagic shock. Even more preferably, the inflammatory disease is an acute inflammatory disease and may be selected from the group consisting of: septic shock, sepsis, SIRS, ARDS, AKI, pancreatitis and hemorrhagic shock. In some embodiments, the inflammatory disease is septic shock, sepsis or SRIS. In some embodiments, the inflammatory disease is septic shock. In some embodiments, the inflammatory disease is sepsis.

[0251] In some embodiments, the inflammatory disease, for example the septic shock or sepsis, may be induced by an infection. In particular, the infection may be a bacterial, fungal, parasitic, or viral infection. Examples of such infections include, without being limited to: respiratory infections, abdominal infections, and urinary tract infections (UTIs).

[0252] In some embodiments, symptoms of septic shock or sepsis ameliorated by the administration of the ready-to-use pharmaceutical composition may include sepsis-induced cardiovascular dysfunction, organ failure, inflammation, fever, and hypothermia.

[0253] In some embodiments, the disease to be treated is a cardiovascular disease. Preferably, the cardiovascular disease may be selected from the group consisting of: myocardial infarction, acute myocardial infarction, cerebral infarction, ischemia, coronaryheart disease, acute coronary syndrome, stroke, aneurysm, stable angina pectoris, effort angina pectoris, cardiomyopathy, hypertensive heart disease, chronic heart failure, acute heart failure, cor pulmonale, cardiac dysrhythmias, inflammatory heart diseases (such as endocarditis and myocarditis), vasculitis, peripheral arterial disease, SIRS-associated myocardial and / or vascular dysfunction, and atherosclerosis. More preferably, the cardiovascular disease may be selected from the group consisting of: myocardial infarction, cerebral infarction, acute myocardial infarction, ischemia, coronary heart disease, acute coronary syndrome, stroke, aneurysm, stable angina pectoris, effort angina pectoris, cardiomyopathy, hypertensive heart disease, chronic heart failure, acute heart failure, cor pulmonale, cardiac dysrhythmias, inflammatory heart disease, peripheral arterial disease, SIRS-associated myocardial and / or vascular dysfunction, and atherosclerosis.

[0254] In some embodiments, the disease to be treated is an infectious disease. Preferably, the infectious disease may be selected from the group consisting of: bacterial diseases (or bacterial infections), viral diseases (or viral infections), fungal diseases (or fungal infections), and parasitic diseases (or parasitic infections). More preferably, the infectious disease may be selected from the group consisting of: Escherichia coli infections and SARSCoV-2 infections (also referred to as coronavirus disease 2019 or COVID-19). Even more preferably, the infectious disease is COVID-19.

[0255] The present invention further relates to the pharmaceutical composition, the reconstituted pharmaceutical composition or the ready-to-use pharmaceutical composition, as described in the present application, for use in the treatment of a disease selected from the group consisting of: stroke, sickle cell diseases, vaso-occlusive crisis in sickle cell diseases patients, trauma.

[0256] In some embodiments, the disease to be treated is selected from the group consisting of:an inflammatory disease, preferably selected from the group consisting of: septic shock, sepsis, severe sepsis, SIRS, ARDS, hemorrhagic shock, ischemia-reperfusion and pancreatitis; a cardiovascular disease, preferably selected from the group consisting of: myocardial infarction, cerebral infarction, acute myocardial infarction, ischemia, coronary heart disease, acute coronary syndrome, stroke, aneurysm, stable angina pectoris, effort angina pectoris, cardiomyopathy, hypertensive heart disease, chronic heart failure, acute heart failure, cor pulmonale, cardiac dysrhythmias, inflammatory heart disease (such as endocarditis and myocarditis), peripheral arterial disease, SIRS-associated myocardial and / or vascular dysfunction, and atherosclerosis; and an infectious disease, preferably COVID- 19.

[0257] In some embodiments, the disease to be treated is selected from the group consisting of: an inflammatory disease, preferably selected from the group consisting of: septic shock, sepsis, severe sepsis, SIRS, ARDS, hemorrhagic shock, ischemia-reperfusion and pancreatitis; a cardiovascular disease, preferably selected from the group consisting of: myocardial infarction, cerebral infarction, acute myocardial infarction, ischemia, coronary heart disease, acute coronary syndrome, stroke, aneurysm, stable angina pectoris, effort angina pectoris, cardiomyopathy, hypertensive heart disease, chronic heart failure, acute heart failure, cor pulmonale, cardiac dysrhythmias, inflammatory heart disease (such as endocarditis and myocarditis), peripheral arterial disease, SIRS-associated myocardial and / or vascular dysfunction, and atherosclerosis; an infectious disease, preferably COVID- 19; and sickle cell diseases, vaso-occlusive crisis in sickle cell diseases patients, trauma.

[0258] In some embodiments, the administration of the pharmaceutical composition, of the reconstituted pharmaceutical composition, preferably of the ready-to-use pharmaceutical composition, to the subject is carried out by infusion or injection, preferably by intravenousinfusion or intravenous injection. In some embodiments, the administration of the ready-to- use pharmaceutical composition to the subject is carried out by continuous infusion, preferably by continuous intravenous infusion.

[0259] In some embodiments, the administration of the ready-to-use pharmaceutical composition to the subject is carried out at a dose (or rate) ranging from about 0.1 mg / kg / h to about 3 mg / kg / h, preferably from about 0.3 mg / kg / h to about 3 mg / kg / h (mg of nangibotide per kilogram of bodyweight of the subject per hour).

[0260] In some embodiments, the administration of the ready-to-use pharmaceutical composition to the subject is carried out at a dose (or rate) ranging from about 0.1 mg / kg / h to about 3 mg / kg / h, preferably from about 0.1 mg / kg / h to about 1 mg / kg / h, more preferably from about 0.3 mg / kg / h to about 1 mg / kg / h (mg of nangibotide per kilogram of bodyweight of the subject per hour).

[0261] Indeed, the high nangibotide concentration in the drug product as described herein and thus the high nangibotide concentration in the ready-to-use pharmaceutical composition as described herein allows a high rate of administration of nangibotide, for example ranging from about 0.1 mg / kg / h to about 3 mg / kg / h (mg of nangibotide per kilogram of body weight of the subject per hour). This high rate of administration improves patient comfort by limiting administration time and reduces the risk of volume overload.

[0262] In some embodiments, the administration of the ready-to-use pharmaceutical composition to the subject is carried out at a dose (or rate) of about 0.1 mg / kg / h, 0.2 mg / kg / h, 0.3 mg / kg / h, 0.4 mg / kg / h, 0.5 mg / kg / h, 0.6 mg / kg / h, 0.7 mg / kg / h, 0.8 mg / kg / h, 0.9 mg / kg / h or 1 mg / kg / h (mg of nangibotide per kilogram of bodyweight of the subject per hour). In some embodiments, the dose is 1 mg / kg / h of nangibotide.

[0263] In some embodiments, the administration of the ready-to-use pharmaceutical composition to the subject, preferably by continuous infusion, is carried out for at least 24 hours, at least 48 hours, at least 72 hours, at least 96 hours, or at least 120 hours. In someembodiments, the administration of the ready-to-use pharmaceutical composition to the subject, preferably by continuous infusion, is carried out for up to 7, 6, 5, 4, or 3 days. In some embodiments, the administration of the ready-to-use pharmaceutical composition to the subject, preferably by continuous infusion, is carried out for up to 5 days.

[0264] In some embodiments, the subject is selected from the group consisting of: a mouse, a rat, a guinea pig, a rabbit, a dog, a cat, a pig, a human, and a non-human primate. Preferably, the subject is a human. More preferably, the subject is an adult human. Alternatively, the subject is an adolescent human. Alternatively, the subject is a pediatric human. For example, a pediatric human is a human ranging from a newborn with a mean body weight of 3.45kg to an adolescent (e.g., under the age of 20, or under the age of 18) with a mean body weight of 54.3kg.

[0265] In some embodiments, the subject suffers or is suffering from a shock or hypotensive shock or distributive shock, which refers to a diminished or insufficient perfusion that impairs organ function and is generally associated with decreased arterial blood pressure. Accordingly, in some embodiments, the subject received or is receiving fluid therapy, i.e., a therapy aiming at restoring and / or maintaining the volume and composition of the body fluids to normal, in particular with respect to the water-electrolyte balance. Fluid therapy thus aims at correcting and / or preventing volume and / or electrolyte deficit. In some embodiments, the subject is vulnerable to hypertonic solutions and / or to volume overload.

[0266] In some embodiments, a subject suffering from an inflammatory disease or disorder, in particular septic shock, sepsis, or SIRS is successfully “treated”, if, after being administered a ready-to-use pharmaceutical composition as described herein, the subject shows at least one of the following: o a reversal of a hypotensive shock, wherein a hypotensive shock reversal is defined as the absence of any vasopressor therapy during at least 24 hours (i.e., not requiring to restart a vasopressor therapy in the 24 hours following the end of a vasopressor therapy); o a decrease of a severity score used to assess the severity of the disease and / or the prognosis of the subject upon admission in ICU or emergency unit, such as theAPACHE II (Acute Physiology And Chronic Health Evaluation II) score, the APACHE (Acute Physiology And Chronic Health Evaluation III) III score, the APACHE IV (Acute Physiology And Chronic Health Evaluation IV) score, the SAPS II (Simplified Acute Physiology Score II) score or the SAPS III (Simplified Acute Physiology Score III) score; o a decrease of an organ dysfunction score used to assess the presence of organ dysfunction in the subject upon admission in ICU or emergency unit, such as the SOFA score, the qSOFA score, the MODS, the P-MODS or the LODS; o a decrease of the SOFA score and / or of the qSOFA score, preferably with reference to the SOFA score and / or the qSOFA score assessed upon admission in ICU or emergency unit or before the start of the administration of a ready-to-use pharmaceutical composition as described herein; o a decrease in the requirement for cardiovascular support, for example a decrease in the use of vasopressor therapy; o a decrease in the requirement for respiratory support, for example a decrease in the use of invasive mechanical ventilation (IMV); o a decrease in the requirement for renal support, for example a decrease in the use of continuous or discontinuous renal replacement therapy also referred to as RRT (e.g., dialysis); o an increase in the quality of life, in particular in the post-sepsis or post-shock quality of life, that may be assessed for example through an evaluation of survival and quality- adjusted life years (QALYs), estimated from the EQ5D. For example, health-related quality of life (HRQoL) scores from the EQ 5D 5L may be calculated and converted to utility scores.The above parameters for assessing successful treatment and improvement in the disease are readily measurable by routine procedures familiar to a physician.Purified nangibotide substance (drug substance)

[0267] The present invention also relates to a purified nangibotide substance comprising nangibotide, wherein said purified nangibotide substance comprises: equal to or less than 2.7% of nangibotide dimers by weight relative to the total weight of all peptides in said purified nangibotide substance; and / or at least about 85%, preferably at least about 86%, of peptides, by weight relative to the total weight of said purified nangibotide substance.

[0268] Such purified nangibotide substance may be referred to as “drug substance”.

[0269] The peptides present in said purified nangibotide substance comprise or consist of nangibotide. The peptides present in said purified nangibotide peptide substance may further comprise at least one peptide selected from the group consisting of: nangibotide dimer, Des- Gln-LR12, Met(O)12-LR12 and any mixtures thereof.

[0270] In some embodiments, the purified nangibotide substance comprises at least 80% w / w, preferably of at least 85% w / w, more preferably of at least 90% w / w, even more preferably at least 95% w / w, of nangibotide, by weight relative to the total weight of the purified nangibotide substance.

[0271] In some embodiments, the purified nangibotide substance comprises at least 80% w / w, preferably of at least 85% w / w, more preferably of at least 90% w / w, even more preferably at least 95% w / w, of nangibotide, by weight relative to the total weight of all peptides in said pharmaceutical composition

[0272] In some embodiments, the purified nangibotide substance comprises equal to or less than about 5.0%, equal to or less than about 4.5%, equal to or less than about 4.0%, equal to or less than about 3.5%, equal to or less than about 3.0%, equal to or less than about 2.7%, or equal to or less than 2.5% of nangibotide dimers by weight relative to the total weight of all peptides in said purified nangibotide substance. In a preferred embodiment, the purifiednangibotide substance comprises equal to or less than 2.7% of nangibotide dimers by weight relative to the total weight of all peptides in said purified nangibotide substance.

[0273] In some embodiments, the purified nangibotide substance further comprises arginine. Preferably, the purified nangibotide substance further comprises arginine in an amount ranging from 2% to 5% by weight relative to the total weight of said purified nangibotide substance.

[0274] In some embodiments, the purified nangibotide substance is a composition comprising:- at least about 85%, preferably at least about 86%, of all peptides, by weight relative to the total weight of said purified nangibotide substance; and / or- equal to or less than about 5.0%, equal to or less than about 4.5%, equal to or less than about 4.0%, equal to or less than about 3.5%, equal to or less than about 3.0%, equal to or less than about 2.7%, or equal to or less than 2.5% of nangibotide dimers by weight relative to the total weight of all peptides in said purified nangibotide substance; and- optionally, from 2% to 5% of arginine by weight relative to the total weight of said purified nangibotide substance.

[0275] In some embodiments, the purified nangibotide substance comprises equal to or less than about 2.5%, equal to or less than about 2.0%, equal to or less than about 1.5%, equal to or less than about 1.2%, equal to or less than about 1.0%, equal to or less than about 0.5% or equal to or less than about 0.3%, of Des-Gln-LR12, by weight relative to the total weight of all peptides in said purified nangibotide substance.

[0276] In some embodiments, the purified nangibotide substance comprises equal to or less than about 2.5%, equal to or less than about 2.0%, equal to or less than about 1.5%, equal to or less than about 1.2%, equal to or less than about 1.0%, equal to or less than about 0.75%, equal to or less than about 0.5% or equal to or less than about 0.3%, of Met(O)12-LR12, by weight relative to the total weight of all peptides in said purified nangibotide substance.

[0277] In some embodiments, the purified nangibotide substance comprises: equal to or less than about 1.2%, preferably equal to or less than about 1.0%, more preferably equal to or less than about 0.5%, of Des-Gln-LR12, by weight relative to the total weight of all peptides in said purified nangibotide substance; and / or equal to or less than about 1.0%, preferably equal to or less than about 0.5%, of Met(O)12-LR12, by weight relative to the total weight of all peptides in said purified nangibotide substance.

[0278] In some embodiments, the purified nangibotide substance comprises at least about 85%, preferably at least about 86%, of all peptides, by weight relative to the total weight of said purified nangibotide substance, wherein said peptides comprise: at least about 95% of nangibotide by weight relative to the total weight of all peptides in said purified nangibotide substance; and equal to or less than about 2.7% of nangibotide dimers by weight relative to the total weight of all peptides in said purified nangibotide substance; and equal to or less than about 1.2% of Des-Gln-LR12 by weight relative to the total weight of all peptides in said purified nangibotide substance; and / or equal to or less than about 1.0% of Met(O)12-LR12 by weight relative to the total weight of all peptides in said purified nangibotide substance.

[0279] In some embodiments, the purified nangibotide substance comprises: equal to or less than about 5.0%, equal to or less than about 4.5%, equal to or less than about 4.0%, equal to or less than about 3.5%, equal to or less than about 3.0%, equal to or less than about 2.5%, equal to or less than about 2.0%, equal to or less than about 1.5%, equal to or less than about 1.0%, or equal to or less than about 0.75%, of acetate, by weight relative to the total weight of said purified nangibotide substance; and / or equal to or less than about 750 ppm, equal to or less than about 700 ppm, equal to or less than about 650 ppm, equal to or less than about 600 ppm, equal to or less than about 550 ppm, equal to or less than about 500 ppm, equal to or less than about 450ppm, or equal to or less than about 410 ppm, of acetonitrile, by weight relative to the total weight of said purified nangibotide substance; and / or equal to or less than about 7000 ppm, equal to or less than about 6500 ppm, equal to or less than about 6000 ppm, equal to or less than about 5500 ppm, equal to or less than about 5000, or equal to or less than about 4500 ppm, of methyl-t-butyl ether, by weight relative to the total weight of said purified nangibotide substance; and / or equal to or less than about 7000 ppm, equal to or less than about 6500 ppm, equal to or less than about 6000 ppm, equal to or less than about 5500 ppm, equal to or less than about 5000, or equal to or less than about 4500 ppm, of n-heptane, by weight relative to the total weight of said purified nangibotide substance

[0280] In some embodiments, the purified nangibotide substance comprises: equal to or less than about 1.0% w / w of acetate; and / or equal to or less than about 500 ppm of acetonitrile, preferably equal to or less than about 410 ppm of acetonitrile; and / or equal to or less than about 5000 ppm of methyl-t-butyl ether; and / or equal to or less than about 5000 ppm of n-heptane, all the percentages being expressed by weight relative to the total weight of said purified nangibotide substance.

[0281] In some embodiments, the purified nangibotide substance comprises: equal to or less than about 1.0% of acetate by weight relative to the total weight of said purified nangibotide substance; equal to or less than about 500 ppm of acetonitrile by weight relative to the total weight of said purified nangibotide substance; equal to or less than about 5000 ppm of methyl-t-butyl ether by weight relative to the total weight of said purified nangibotide substance; equal to or less than about 5000 ppm of n-heptane by weight relative to the total weight of said purified nangibotide substance; andat least about 85%, preferably at least about 86%, of all peptides, by weight relative to the total weight of said purified nangibotide substance, wherein said peptides comprise: o at least about 95% of nangibotide by weight relative to the total weight of all peptides in said purified nangibotide substance; o equal to or less than 2.7% of nangibotide dimers by weight relative to the total weight of all peptides in said purified nangibotide substance; o equal to or less than about 1.2% of Des-Gln-LR12 by weight relative to the total weight of all peptides in said purified nangibotide substance; o equal to or less than about 1.0% of Met(O)12-LR12 by weight relative to the total weight of all peptides in said purified nangibotide substance.

[0282] In some embodiments, the purified nangibotide substance is a powder, preferably a lyophilized powder. Preferably, the purified nangibotide substance is a powder having a humidity of equal to or less than 10.0% w / w.

[0283] In some embodiments, the purified nangibotide substance comprises equal to or less than 10.0% of water by weight relative to the total weight of said purified nangibotide substance.

[0284] The purified nangibotide substance as described herein allows the obtention of a drug product comprising a high concentration of nangibotide, which allows the obtention of a ready-to-use pharmaceutical composition having a reduced volume to be administered to the patient.

[0285] The present invention also relates to a vial containing a purified nangibotide substance according to the invention. In some embodiments, a vial contains from about 600 mg to about 2500 mg of nangibotide. For example, the vial may be a 10 mb or a 20 mb vial, preferably a 20 mb vial.Process for manufacturing purified nangibotide substance

[0286] The present invention also relates to a process for manufacturing a purified nangibotide substance, preferably a purified nangibotide substance according as described in part “Purified nangibotide substance (drug substance)” of the present application.

[0287] In some embodiments, the process for preparing a purified nangibotide substance comprises: i. synthesizing the nangibotide on solid phase, leading to a crude nangibotide composition; ii. providing a first peptide solution comprising the crude nangibotide composition; iii. purifying the first peptide solution by ion exchange column chromatography (IEC), leading to a first pre -purified substance of nangibotide; iv. further purifying the pre -purified substance of nangibotide by reversed-phase high performance liquid chromatography (HPLC), thereby leading to HPLC pooled fractions comprising a second pre -purified substance of nangibotide; v. preparing a solution containing at least one solubility enhancing agent, preferably arginine, between 5 mM and 25 mM to be added to the pooled fractions during step vi.; vi. concentrating the pooled fractions using membrane diafiltration or ultrafiltration, typically on a 150 to 300 Dalton membrane, to a concentrated solution of at least 60 g / L of nangibotide, preferably at least 120 g / L of nangibotide; and vii. filtering and lyophilizing the concentrated solution obtained in step vi) to obtain the purified nangibotide substance.

[0288] In some embodiments, concentrating the pooled fractions comprises 2 steps of concentration.

[0289] In some embodiments, the HPLC pooled fractions comprising the second prepurified substance of nangibotide are cooled to a temperature of 15°C or less and / or are bubbled with nitrogen.

[0290] According to some embodiments, the purified nangibotide substance obtained in step vii. comprises at least one solubility enhancing agent, preferably arginine in an amount ranging from 2-5% w / w, by weight relative to the weight of the purified nangibotide substance.

[0291] Such a process for manufacturing a purified nangibotide substance allows to further reduce the formation of nangibotide dimers while maintaining nangibotide solubility.

[0292] In some embodiments of the process, the concentrated nangibotide solution is filtered and lyophilized to obtain a purified nangibotide substance having a peptide purity of at least 95%, 96%, or 97%. In some embodiments of the process provided herein, the concentrated nangibotide solution is filtered and lyophilized to obtain a purified nangibotide substance having a peptide purity of at least 95%.

[0293] In some embodiments of the process disclosed herein, nangibotide dimer formation is reduced by using buffers at pH between 5 and 6 for IEC, RPC and diafiltration, and adjusting the pH between 5 and 6 of the pooled fractions from RPC; cooling the solutions below 15°C; and / or bubbling the solutions with nitrogen.

[0294] The improved manufacturing process provided herein controls timing between steps to allow an acceptable and reproducible control of the nangibotide dimer content in the final drug product. To optimize the practicability of administration in a hospital setting, the drug product is highly concentrated, with a suitable stability.Process for manufacturing a pharmaceutical composition comprising nangibotide (drug product)

[0295] The present invention also relates to a process for manufacturing a pharmaceutical composition comprising nangibotide, preferably a pharmaceutical composition according as described in part “Pharmaceutical composition comprising nangibotide (drug product)” of the present application.

[0296] The process for manufacturing a pharmaceutical composition comprising nangibotide, may comprise the steps of: a) mixing a purified nangibotide substance with at least one amino acid selected from the group consisting of: arginine, lysine and histidine, preferably arginine or lysine, more preferably arginine, and optionally with at least one pharmaceutically acceptable buffer, preferably at least one pharmaceutically acceptable buffer as described herein, preferably wherein: the amount of the at least one amino acid ranges from 20.0% to 30.0%, preferably from 22.0% to 28.0%, more preferably from 22.3% to 27.6%, even more preferably is about 24.5%, by weight relative to the dry weight of the purified nangibotide substance, and / or the amount of the optional at least one pharmaceutically acceptable buffer ranges from 3.0% to 7.0%, preferably about 3% to about 5%, preferably from 4.0% to 6.0%, more preferably from 4.5% to 5.5%, even more preferably is about 4.8%, by weight relative to the dry weight of the purified nangibotide substance; thereby leading to a bulk solution; then b) optionally adjusting the pH of the bulk solution obtained in step a) to a pH of about 4.5 to about 8.5, preferably about 5.5 to about 6.5, preferably about 5.5, preferably said pH adjustment being carried out using HC1; then c) optionally lyophilizing the bulk solution obtained in step a) or in step b) to obtain a pharmaceutical composition in a powder form.

[0297] For example, the at least one amino acid is selected from lysine, arginine, histidine, and combinations thereof. For example, the pharmaceutically acceptable buffer may be selected from a citrate buffer, a histidine buffer, a succinate buffer, an acetate buffer, a phosphate buffer, or a tromethamine buffer. For example, the pharmaceutically acceptable buffer may be selected from a histidine buffer, a succinate buffer, an acetate buffer, a phosphate buffer, or a tromethamine buffer. For example, the pharmaceutically acceptablebuffer may be selected from a histidine buffer, a succinate buffer, an acetate buffer, and a phosphate buffer.

[0298] In some embodiments, the purified nangibotide substance supplied in step a) is obtained by carrying out the steps i) to vii) of the process for preparing a purified nangibotide substance as described in part “Process for manufacturing purified nangibotide substance” of the present application.

[0299] The process for manufacturing a pharmaceutical composition comprising nangibotide, may comprise the steps of: a) mixing nangibotide with at least one amino acid selected from the group consisting of: arginine, lysine and histidine, wherein the amount of the at least one amino acid ranges is of at least 22% by weight relative to the weight of nangibotide, for example from 22% to 34 % by weight relative to the weight of nangibotide, and optionally adding at least one pharmaceutically acceptable buffer, preferably at least one pharmaceutically acceptable buffer as described herein, thereby leading to a bulk solution; b) optionally adjusting the pH of the bulk solution obtained in step a) to a pH ranging from about 4.5 to about 8.5, preferably from about 5.5 to about 6.5, more preferably of about 5.5, preferably said pH adjustment being carried out using HC1; and c) optionally lyophilizing the bulk solution obtained in step a) or in step b) to obtain a pharmaceutical composition in a powder form.

[0300] In some embodiments, the nangibotide used in step a) is supplied via a purified nangibotide substance, preferably said purified nangibotide substance is obtained by carrying out the steps i) to vii) of the process for preparing a purified nangibotide substance as described in part “Process for manufacturing purified nangibotide substance” of the present application.

[0301] In some embodiments, the bulk solution obtained in step a) or in step b) is introduced in vial(s) before optional step c) of lyophilization. This may allow obtaining the pharmaceutical composition in a powder form directly present in the container (i.e., the vial) for storage and distribution. For example, the vial may be a 10 mb or a 20 mb vial, preferably a 20 mb vial.

[0302] In some embodiments, the bulk solution obtained in step a) or in step b) is colorless.

[0303] In some embodiments, the pharmaceutical composition comprising nangibotide manufactured by the process as described herein is a pharmaceutical composition according as described in part “Pharmaceutical composition comprising nangibotide (drug product)” of the present application. In this case, all the features described above for the pharmaceutical composition referred to as “drug product” (part “Pharmaceutical composition comprising nangibotide (drug product)” of the present application) apply mutatis mutandis for the pharmaceutical composition manufactured by the process for manufacturing a pharmaceutical composition comprising nangibotide. In particular, all the features described above regarding the weight of nangibotide in the pharmaceutical composition, the amount of the at least one amino acid by weight relative to the weight of nangibotide, the pH, the presence or absence of at least one pharmaceutically acceptable buffer, the nature and amount of the at least one pharmaceutically acceptable buffer by weight relative to the weight of nangibotide, the weight of Des-Gln-LR12 and Met(O)12-LR12 relative to the total weight of all peptides in said pharmaceutical composition, apply mutatis mutandis for the pharmaceutical composition comprising nangibotide manufactured by the process for manufacturing a pharmaceutical composition comprising nangibotide.

[0304] This process for manufacturing a pharmaceutical composition comprising nangibotide allows a high concentration of nangibotide to be present in the obtained pharmaceutical composition and a low amount of nangibotide dimers to be formed in said pharmaceutical composition. Thus, the pharmaceutical composition comprising nangibotideobtained by the process as described herein is highly concentrated in nangibotide and has a suitable nangibotide stability.Process for preparing a reconstituted pharmaceutical composition

[0305] The present invention also relates to a process for preparing a reconstituted pharmaceutical composition, preferably a reconstituted pharmaceutical composition as described in part “Reconstituted pharmaceutical composition” of the present application.

[0306] The process for preparing a reconstituted pharmaceutical composition may comprise the following steps:- carrying out the process for manufacturing a pharmaceutical composition as described in part “Process for manufacturing a pharmaceutical composition comprising nangibotide (drug product)” of the present application; then- adding an aqueous medium to the pharmaceutical composition in a powder form obtained in step c) of the process for manufacturing a pharmaceutical composition, to obtain a reconstituted pharmaceutical composition.

[0307] The process for preparing a reconstituted pharmaceutical composition may comprise the following steps: a) mixing a purified nangibotide substance with at least one amino acid selected from the group consisting of: arginine, lysine and histidine, preferably arginine or lysine, more preferably arginine, and optionally with at least one pharmaceutically acceptable buffer, preferably at least one pharmaceutically acceptable buffer as described herein, preferably wherein: the amount of the at least one amino acid ranges from 20.0% to 30.0%, preferably from 22.0% to 28.0%, more preferably from 22.3% to 27.6%, even more preferably is about 24.5%, by weight relative to the dry weight of the purified nangibotide substance, and / or the amount of the optional at least one pharmaceutically acceptable buffer ranges from 3.0% to 7.0%, preferably about 3% to about 5%, preferably from4.0% to 6.0%, more preferably from 4.5% to 5.5%, even more preferably is about 4.8%, by weight relative to the dry weight of the purified nangibotide substance; thereby leading to a bulk solution; then b) optionally adjusting the pH of the bulk solution obtained in step a) to a pH of about 4.5 to about 8.5, preferably about 5.5 to about 6.5, preferably about 5.5, preferably said pH adjustment being carried out using HC1; then c) optionally lyophilizing the bulk solution obtained in step a) or in step b) to obtain a pharmaceutical composition in a powder form; then- adding an aqueous medium to the pharmaceutical composition in a powder form obtained in step c) of the process for manufacturing a pharmaceutical composition, to obtain a reconstituted pharmaceutical composition.

[0308] In some embodiments, the step of adding an aqueous medium to the pharmaceutical composition is carried out extemporaneously, preferably less than 2 hours before starting treatment.

[0309] In some embodiments, said aqueous medium is selected from the group consisting of: water and a pharmaceutically acceptable aqueous solution. Preferably, said aqueous medium is selected from the group consisting of: water (e.g., water for injection (WFI) USP) and saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.

[0310] In some embodiments, the bulk solution obtained in step a) or in step b) of the process for manufacturing a pharmaceutical composition is introduced in vial(s) before step c) of lyophilization. In this case, the step of adding an aqueous medium to the pharmaceutical composition during the process for preparing a ready-to-use pharmaceutical composition is carried out directly in the vial. For example, said vial may be a vial of 10 mL or 20 mb, preferably of 20 mL.Process for preparing a ready-to-use pharmaceutical composition

[0311] The present invention also relates to a process for preparing a ready-to-use pharmaceutical composition, preferably a ready-to-use pharmaceutical composition as described in part “Ready-to-use pharmaceutical composition” of the present application.

[0312] The process for preparing a ready-to-use pharmaceutical composition may comprise the following steps:- carrying out the process for manufacturing a reconstituted pharmaceutical composition as described in part “Process for preparing a reconstituted pharmaceutical composition” of the present application; then- diluting the reconstituted pharmaceutical composition in an aqueous medium, preferably in a syringe or in an infusion bag, for example a syringe or in an infusion bag having a total volume of 50 mL.

[0313] The process for preparing a ready-to-use pharmaceutical composition may comprise the following steps: a) mixing a purified nangibotide substance with at least one amino acid selected from the group consisting of: arginine, lysine and histidine, preferably arginine or lysine, more preferably arginine, and optionally with at least one pharmaceutically acceptable buffer, preferably at least one pharmaceutically acceptable buffer as described herein, preferably wherein: the amount of the at least one amino acid ranges from 20.0% to 30.0%, preferably from 22.0% to 28.0%, more preferably from 22.3% to 27.6%, even more preferably is about 24.5%, by weight relative to the dry weight of the purified nangibotide substance, and / or the amount of the optional at least one pharmaceutically acceptable buffer ranges from 3.0% to 7.0%, preferably about 3% to about 5%, preferably from 4.0% to 6.0%, more preferably from 4.5% to 5.5%, even more preferably isabout 4.8%, by weight relative to the dry weight of the purified nangibotide substance; thereby leading to a bulk solution; then b) optionally adjusting the pH of the bulk solution obtained in step a) to a pH of about 4.5 to about 8.5, preferably about 5.5 to about 6.5, preferably about 5.5, preferably said pH adjustment being carried out using HC1; then c) optionally lyophilizing the bulk solution obtained in step a) or in step b) to obtain a pharmaceutical composition in a powder form; then- adding an aqueous medium to the pharmaceutical composition in a powder form obtained in step c), to obtain a reconstituted pharmaceutical composition; then- diluting the reconstituted pharmaceutical composition in an aqueous medium, preferably in a syringe or in an infusion bag, for example a syringe or in an infusion bag having a total volume of 50 mL, to obtain a ready-to-use pharmaceutical composition.

[0314] In some embodiments, the step of diluting the reconstituted pharmaceutical composition in an aqueous medium is carried out extemporaneously, preferably less than 2 hours before starting treatment.

[0315] In some embodiments, said aqueous medium is selected from the group consisting of: water and a pharmaceutically acceptable aqueous solution. Preferably, said aqueous medium is selected from the group consisting of: water (e.g., water for injection (WFI) USP) and saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.

[0316] For the administration to the subject, a syringe (50 mL) and an automatic syringe pump may be used to dispense by continuous intravenous infusion a dose ranging from 0.1 to 3 mg / kg / h, preferably a dose of 1 mg / kg / h (mg of nangibotide per kilogram of body weight of the subject per hour).

[0317] These steps allow a reduction of the total amount of liquid to be administered per day to the subject.BRIEF DESCRIPTION OF THE DRAWINGS

[0318] Figure 1 is a graph showing the results from flow cytometric quantification (DCFDA reduction) of neutrophils ROS production after 2 hours stimulation with LPS, LPS + l Oug / mL nangibotide, LPS + 50ug / mL nangibotide, LPS + l Oug / mL nangibotide -dimer (dimer), LPS + 50pg / mL nangibotide -dimer (dimer). *p<0.005; **p< 0.01 (T test).EXAMPLES

[0319] The present disclosure is further illustrated by the following examples. The following examples are included for illustrative purposes only and are not intended to limit the scope of the disclosure. It will be understood by those of skill in the art that numerous and various modifications can be made to yield essentially similar results without departing from the spirit of the present disclosureExample 1: Absence of biological activity of the nangibotide dimerMaterials and Methods

[0320] Neutrophils produce reactive oxygen species (ROS) in presence of LPS (lipopolysaccharides) following activation of the TREM- 1 pathway and are thus used in the present example in order to assess TREM-1 activation and the biological activity of the nangibotide monomer and dimer on the TREM-1 pathway. ROS production induced by TREM-1 activation and its modulation by nangibotide can be quantified by flow cytometry. Any decrease in ROS production by nangibotide (LPS+LR12 conditions) translates into a decrease in mean fluorescence intensity (MFI) as compared to the ROS production observed following the TREM-1 activation in presence of LPS (LPS condition).

[0321] Primary human neutrophils were isolated from the peripheral blood of healthy donors by immunomagnetic negative cell sorting with EasySep™ Human Monocyte / Neutrophil Isolation Kits (StemCell, Canada) following the manufacturer’sinstructions. Purity was assessed by flow cytometry. Cells were suspended in RPMI 1640 medium containing GlutaMAX and supplemented with 10% FCS, 25 mM HEPES, 100 U / ml penicillin and streptomycin before stimulation. Human primary neutrophils were incubated in resting conditions (also referred to as control conditions or Ctrl 2), or with 100ng / mL LPS from E. coli serotype 0127:B8 (Sigma-Aldrich), with or without nangibotide or nangibotide(dimer (dimer) at the indicated concentrations.

[0322] ROS production was quantified by using a fluorogenic substrate (DCFDA or 2',7'-dichlorofluorescein diacetate), a chemically reduced form of fluorescein used as an indicator of the presence of ROS in cells. Upon cleavage of the acetate groups by intracellular esterases and oxidation, the nonfluorescent DCFDA is converted to the highly fluorescent 2', 7'-dichloro fluorescein (DCF). 2.5x105isolated human neutrophils were incubated 2 hours at 37°C - 5% CO2 with 5uM of DCFDA, with or without lOOng / mL LPS, and with or without nangibotide or nangibotide-dimer (dimer) at the indicated concentrations. Data were acquired using flow cytometry. Results are expressed as mean fluorescence intensity (MFI).Results

[0323] As shown on Figure 1, LPS induced a significant increase of ROS production by human primary neutrophils after 2 hours stimulation. Nangibotide monomer was able to decrease in a dose-dependent manner the LPS-induced release of ROS. Surprisingly, nangibotide dimer did not display any TREM-1 inhibiting property. Indeed, nangibotide- dimer was not able to decrease the production of ROS by LPS-activated neutrophils.

[0324] These results show that, despite the fact that the nangibotide-dimer is composed of 2 nangibotide peptides covalently bound by a disulfide bond through the cysteine at position 11, it is not able to induce any pharmacological inhibition of TREM-1.Example 2: Preparation of the purified nangibotide substance (drug substance)

[0325] A stepwise solid-phase synthesis of nangibotide peptide (15mol) was performed using Fmoc / tBu chemistry in a 270L SPPS reactor. Sequential acylation reactions are carriedout at 20°C ±3°C under stirring at 30 ± 20 rpm using appropriate Na-Fmoc-protected amino acids, which are carboxyl-activated using Oxyma, DITU and DIC in DMF. Capping step by acylation with acetic anhydride is repeated after each amino acid coupling. Upon completion of the sequential acylation reactions, the peptide-resin is washed with IP A, filtered and dried in vacuo.

[0326] The resin-bound peptide is added into the cleavage reactor with a rate such that the product temperature does not exceed 25°C. Cleavage and side-chain deprotection of the peptidyl-resin are carried out by treatment with a prepared solution of TFA / DTT / TIS / H2O for 3-4h at 20°C ±5°C. After cooling down the cleavage solution to 0°C ±5°C, MBTE and n- hexane precipitation is carried out. The precipitate is filtered, washed with MBTE, and dried under vacuum.

[0327] The dried crude nangibotide peptide is re-solubilized in aqueous ammonia, its pH is adjusted to 6.5-7.0, and let under stirring for 3h at 50-55°C, before removing the spent resin by filtration. Purification of the dissolved crude solution is carried through ion-exchange chromatography (IEC) with load of <30 g / L Column Volume (CV) on a 450 mm diameter column (2.5cm / min), followed by Reversed-phase chromatography (RPC) with load of 15-22 g / L CV on a 450 mm diameter column (4.7cm / min) for which fractions are eluted with NH4O Ac / Acetonitrile (MeCN), collected in water and kept under cold storage (2-8°C). The pooled fractions are concentrated (in 2 steps) and purified by ultrafiltration / diafiltration using 300 Dalton membrane. 15mM Arginine is added after the 1st concentration (approx. 65 g / L). The solution is then further processed to a concentrated solution of 170g / L. Nitrogen inerting of the bulk solutions (cooled below 15°C) is performed to maintain a low-level oxygen content.

[0328] The concentrated nangibotide solution is immediately filtered and lyophilized to obtain a purified nangibotide substance containing i- 85% peptide content by weight of the total purified nangibotide substance weight and ii- 4.7% arginine by weight of the totalpurified nangibotide substance weight. The amount of the nangibotide dimer was 0.7% by weight relative to the total weight of the peptide content.Example 3: Preparation of the pharmaceutical composition (drug product)

[0329] The pH during the formulation process of the purified nangibotide substance “drug substance” according to Example 2 is maintained between 5 and 6. The timing between the solubilization of the purified nangibotide substance “drug substance” (comprising the active pharmaceutical ingredient, API), the addition of excipients (pH adjustment, arginine), and the filling of vials and the temperature adjustment in the freeze-dryer is set below 24 hours. Nitrogen inerting of the bulk solutions is performed by bubbling nitrogen to the solutions in order to get an oxygen content below 5 ppm. At the time of administration, the pharmaceutical composition (drug product powder in a 20mL vial) is reconstituted with 9.5 mb of NaCl 0.9% w / w to obtain the reconstituted pharmaceutical composition and then further diluted in a 50 mb syringe with NaCl 0.9% w / w to obtain the ready-to-use pharmaceutical composition. The concentration of obtained nangibotide solution is 24 mg / mL with an osmolality of 380 mOsm / kg.

[0330] The quantitative composition of the drug product is presented in Table 1.* containing 1200mg API (nangibotide)Table 1: Composition of the nangibotide drug product

[0331] Nangibotide-dimer content measured by analytical reversed phase HPLC-UV and expressed as a percentage by weight relative to the weight of the nangibotide peptide for every process step are presented in Table 2 below.Table 2: Content of nangibotide dimer, Des-Gln-LR12 and Met(O)12-LR12 % of the purified nangibotide substance, of the drug product and of the reconstituted drug product; the % being relative to the peptide content of the purified nangibotide substance, drug product and the reconstituted drug product respectively.Comparative Examples 4-9.: Preparation of comparative pharmaceutical compositionsComparative Example 4a (NaCl):

[0332] The purified nangibotide substance (of example 2) is formulated using the following excipients: 50 mM sodium citrate pH 5.5; 50 mM NaCl. The concentration of bulk solution (before freeze drying) is 10 mg / mL (due to solubility of nangibotide limit).Comparative Example 4b (NaCl, pH 6.5):

[0333] The purified nangibotide substance (of example 2) is formulated using the following excipients: 50 mM sodium phosphate pH 6.5; 50mM NaCl. The concentration of bulk solution (before freeze drying) is 20 mg / mL (due to solubility of nangibotide limit).Comparative Example 5 (mannitol):

[0334] The purified nangibotide substance (of example 2) is formulated using the following excipients: 50 mM sodium citrate pH 5.5; 200 mM mannitol. The concentration of bulk solution (before freeze drying) is 10 mg / mL (due to solubility of nangibotide limit).Comparative Example 6 (betaine):

[0335] The purified nangibotide substance (of example 2) is formulated using the following excipients: 50 mM sodium citrate pH 5.5; 150 mM betaine. The concentration of bulk solution (before freeze drying) is 10 mg / mL (due to solubility of nangibotide limit).Comparative Example 7a (lysine):

[0336] The purified nangibotide substance (of example 2) is formulated using the following excipients: 5 OmM sodium citrate pH 5.5; 50 mM lysine; 50 mM NaCl. The concentration of bulk solution (before freeze drying) is 40 mg / mL (due to solubility of nangibotide limit).Comparative Example 7b (arginine):

[0337] The purified nangibotide substance (of example 2) is formulated using the following excipients: 50mM sodium citrate pH 5.5; 50 mM arginine. The concentration of bulk solution (before freeze drying) is 40 mg / mL (due to solubility of nangibotide limit).Comparative Example 8 (sodium citrate; arginine; pH 5.5):

[0338] The purified nangibotide substance (of example 2) is formulated using the following excipients: 25mM sodium citrate pH 5.5; 225 mM arginine; 65 mM HC1. The concentration of bulk solution (before freeze drying) is 150 mg / mL; osmolality 512 mOsm / kg.Series of Comparative Examples 9 (90-420 mM Arginine; pH 5.5):

[0339] Formulations using incremental concentrations of the purified nangibotide substance (of example 2) and incremental concentrations of the following excipients: Arginine 90 to 420 mM, using only HC1 for pH adjustment to 5.5.

[0340] The results regarding the nangibotide solubility, the nangibotide stability and osmotic pressure of the obtained pharmaceutical compositions is presented in Table 3 below.Table 3: Solubility, stability and osmolality of the compared pharmaceutical compositions

[0341] Among all different formulations tested, citrate or phosphate buffers alone only allowed for up to 20 mg / mL nangibotide solubility (comparative examples 4a and 4b). The use of excipients such as mannitol or betaine did not lead to increased solubility of nangibotide (examples 5 and 6). Both arginine and lysine at 50mM were able to increase the solubility of nangibotide up to 40mg / mL, (comparative examples 7a and 7b). Increasing concentrations of arginine from 90 to 420 mM (example 9) were associated with higher solubility of nangibotide, but were also associated with significant higher dimer increase during formulation, despite the fact that pH was maintained as per in the pharmaceutical composition of the invention.

[0342] The combination of arginine and citrate buffer, according to the invention, allowed thus not only to enhance the nangibotide solubility but also to obtain the lowest dimer content products while using the minimum possible amount of excipients and keeping the osmolality of the obtained pharmaceutical composition below 500 mOsm / kg.Comparative Example 11: Comparative process regarding the preparation of purified nangibotide substance

[0343] The dried crude nangibotide peptide (of example 2) is re-solubilized in aqueous ammonia, pH adjusted to 6.5-7.0, and under stirring for 3h at 50-55°C, before removing the spent resin by filtration. Purification of the dissolved crude solution is carried through ionexchange chromatography (IEC) with load of <30 g / L CV on a 450mm diameter column (2.5cm / min), followed by Reversed-phase chromatography (RPC) at room temperature - 25°C) with load of 15-22 g / L CV on a 450mm diameter column (4.7cm / min) for which fractions are eluted with NH4OAc / MeCN, collected in water, without Nitrogen inerting. The pooled fractions are concentrated and purified by ultrafiltration / diafiltration using 300 Dalton membrane to a concentrated solution of 15g / L. The concentrated nangibotide solution is filtered and lyophilized to obtain a purified substance of nangibotide peptide.

[0344] Nangibotide-dimer contents for every process step are presented in Table 4.Table 4: nangibotide-dimer content

[0345] The results of the comparative example 11 show that the process steps according to the invention, in particular the N2 inerting and / or the reversed phase, ultrafiltration / diafiltration steps at chilled temperature of less than 15°C presented in example2 may further inhibit the nangibotide-dimer formation in the purified substance of nangibotide peptide.Examples 12-18: Preparation of pharmaceutical compositions according to the invention and of comparative pharmaceutical compositions

[0346] Material and methods

[0347] Pharmaceutical compositions were prepared using incremental concentrations of nangibotide peptide in the purified nangibotide substance (of example 2); incremental amounts of amino acids (arginine or lysine) from 15% to 26% by weight relative to the dry weight of the purified nangibotide substance; and incremental amounts of sodium citrate buffer from 5% to 7% by weight relative to the dry weight of the purified nangibotide substance. The obtained pharmaceutical compositions were lyophilized powders (= drug product in powder form) containing from 1200 mg to 2000 mg of nangibotide (per vial).

[0348] In order to prepare these pharmaceutical compositions in powder form, the buffer solution was first prepared according to table 5. The purified nangibotide substance and atleast one amino acid selected from arginine and lysine were added into the prepared buffer solution. The pH during the formulation process was adjusted to get a bulk solution (before the freeze-drying step) around 5.5. To further limit dimer formation during the manufacturing process, nitrogen inerting of the bulk solutions was performed in order to get an oxygen content below 5 ppm.

[0349] The prepared bulk solution was visually checked and filtered using 0.2 pm disposable filters. Then, 20 mb vials were filled with the filtered bulk solutions. Filling volumes varied from lOmL to 15mL depending on the nangibotide concentration in each bulk solution (from 80 mg / mL to 200 mg / mL). The vials containing the filtered bulk solutions were then lyophilized to get pharmaceutical compositions in the form of powders (= drug product in storage form).

[0350] Exemplary pharmaceutical compositions according to the invention are: nangibotide 120mg / mL & Arginine (155mM), nangibotide 120mg / mL & Lysine (190mM), nangibotide 120mg / mL & Arginine (190mM), nangibotide 200mg / mL & Arginine (300mM), nangibotide 200mg / mL & Lysine (300mM).

[0351] The following pharmaceutical compositions were compared to the above formulations as shown in Table 5: nangibotide 80mg / mL & Arginine (80mM), and nangibotide 100mg / mL & Arginine (120mM).

[0352] The material and methods are summarized in the following Table 5:*134.9mg of purified nangibotide substance comprises 120mg of nangibotideTable 5: Preparation of the compared pharmaceutical compositions

[0353] Results

[0354] The results are presented in the following Table 6.Table 6: Results obtained with the compared pharmaceutical compositions

[0355] The results show that there are precipitates for the two comparative examples having an amount of the amino acid of less than 20% by weight relative to the dry weight of the purified nangibotide substance. Those precipitates, observed in the bulk solution before the filtration, are not suitable for further processing. Thus, it is not possible to obtain a pharmaceutical composition in a powder form (=drug product in a storage form) when the amount of the amino acid is of less than 20% by weight relative to the dry weight of the purified nangibotide substance.

[0356] The results show that, upon reconstitution with 9.5 mL of NaCl 0.9% w / w, the pharmaceutical composition (drug product powder in a 20mL vial) comprises (by weight relative to the peptide content) less than 5.0% w / w of nangibotide dimer by weight relative to the peptide content, less than 1.2% of Des-Gln-LR12 by weight relative to the peptide content and less than 1.0% of Met(O)12-LR12 by weight relative to the peptide content. The osmolality is confirmed to be less than the acceptable limit of 500 mOsm / kg.

[0357] Therefore, a pharmaceutical composition according to the invention in a powder form, having an amount of arginine, lysine or histidine of at least 22% by weight relative to the dry weight of the purified nangibotide substance, allows the obtention of a pharmaceutical composition to have less than 5.0% w / w of nangibotide dimer by weight relative to the peptide content, high nangibotide amounts of 1200 mg and 2000 mg and an osmolality of less than the acceptable limit of 500 mOsm / kg. All these technical advantages contribute to allow the pharmaceutical composition in its ready-to-use form to, at least: be suitable for administration with a high rate of administration, improving the patient comfort and limiting the risk of volume overload; decreasing the ratio between the weight content of excipients to the weight content of nangibotide, limiting the daily exposure to excipients, i.e., the total amount of the excipients that would be taken or used in a day basis (in mg / kg of excipients per kilo body weight per hour); be suitable for administration to pediatric population with body weight below 30kg; be tolerated by the patient body (osmolality of less than the acceptable limit of 500 mOsm / kg); andhave a sufficient therapeutic effect of the nangibotide thanks to high purity and stability of nangibotide.

[0358] On the contrary, pharmaceutical compositions in a powder form (=drug product in a storage form) were not obtained when the amount of the amino acid was of less than 20% by weight relative to the dry weight of the purified nangibotide substance.Example 19: Preparation of pharmaceutical compositions (drug product) according to the invention

[0359] Material and methods

[0360] Pharmaceutical compositions as described herein were prepared using 120mg / mL of nangibotide (batch No 30-22-0289-01, PolyPeptide Torrance) with different buffer solutions (Citrate, Histidine, Succinate, Acetate, Phosphate, and Tromethamine) at different pH and three solubilizing agents (Arginine, Lysine, Histidine).

[0361] First, different buffer solutions containing at least one solubilizing agent were prepared according to Table 7.

[0362] Second, nangibotide peptide was added slowly and gradually to approximatively 20 mL of buffer solution under continuous stirring and refrigerated conditions.

[0363] Third, pH of the bulk was adjusted to pH 5.7, 7.0 or 8.0 using HCl 4N and / or NaOH 10%-20% before adding buffer solution for final volume make up to 30 mL. Resulting bulk solution was visually checked before aliquoting into 1 mL cryotubes and remaining solution into 50 mL conical centrifuge tubes, which were stored at -80°C.

[0364] For some specific pharmaceutical compositions, 15 mL of bulk solution was stored for 8 h at 4°C and then stored at -80°C to assess the manufacturing process holding time stability (SPT), i.e., the stability of a pharmaceutical composition comprising nangibotide (drug product) as described herein. After -80°C storage up to a maximal overnight duration, 3 mL of bulk solution was diluted with 12 mL NaCl 0.9% to yield a 24 mg / mL ready-to-use pharmaceutical composition as described herein, which was leftfor 24 h at room temperature and then stored at -80°C to assess the ready-to-use stability (IUT), i.e., the stability of a ready-to-use pharmaceutical composition as described herein. For each test, 2x 1 mL were aliquoted into 1 mL cryotubes, which were stored at -80°C.

[0365] Analytical measurements were carried out on all samples: after thawing cryotubes at room temperature, or in a stability chamber at 25°C for SPT and IUT samples, samples were diluted in SEC-HPLC mobile Phase (i.e., potassium hydrogen phosphate, pH 6.7) and transferred to the refrigerated autosampler of the HPLC equipment for SEC-HPLC-UV analysis (TSK GEL G2000SWXL, 5 pm, 7.8mm x 300mm column; 1 mL / min; 20 min run; 20 uL injection) within the next 2-12 hours to determine the percentage area of nangibotide dimer. A correction factor was applied to align the results with UPLC-PDA / UV, the reference method for quantifying dimers during drug product release testing. The correction factor, determined as 0.82 or 0.83, was calculated based on the UPLC-PDA / UV dimer result at release for the reference standard NGA02 (3.8%) used during SEC-HPLC and the corresponding SEC-HPLC results (4.64% or 4.57%).

[0366] 50 mL conical centrifuge tubes were stored at 4°C for thawing until further pH and osmolality measurements. pH was determined at room temperature using a pH meter. Osmolality was determined using an osmometer.

[0367] Table 7: Preparation of buffer solutions comprising a solubilizing agent

[0368] Results

[0369] The results are presented in the following tables:

[0370] Table 8: Analytical results of 120mg / mL nangibotide in different buffers in the presence of 190 mM Arginine* based on correction factor of 0.82 calculated from UPLC-PDA / UV % dimer result at release (i.e., 3.8%) and SEC-HPLC result (i.e., 4.64%o) of the reference standard NGA02

[0371] Table 9: Analytical results of 120mg / mL nangibotide in different buffers in the presence of 190 mM Lysine* based on correction factor of 0.82 calculated from UPLC-PDA / UV % dimer result at release (i.e., 3.8%) and SEC-HPLC result (i.e., 4.64%o) of the reference standard NGA02

[0372] Table 10: Analytical results of 120mg / mL nangibotide in different buffers in the presence of 190 mM Histidine* based on correction factor of 0.83 calculated from UPLC-PDA / UV %> dimer result at release (i.e., 3.8%) and SEC-HPLC result (i.e., 4.57%o) of the reference standard NGA02

[0373] Table 11: Analytical results of 120mg / mL nangibotide in the presence of 190 mM Arginine assessed for process holding time stability (SPT) after 8 h at 4°C* based on correction factor of 0.83 calculated from UPLC-PDA / UV %> dimer result at release (i.e., 3.8%) and SEC-HPLC result (i.e., 4.57%o) of the reference standard NGA02

[0374] Table 12: Analytical results of24 mg / mL nangibotide in the presence of 190 mMArginine assessed for ready-to-use stability (IUT) after 24 h at room temperature* based on correction factor of 0.83 calculated from UPLC-PDA / UV % dimer result at release (i.e., 3.8%) and SEC-HPLC result (i.e., 4.57%o) of the reference standard NGA02

[0375] Table 13: Analytical results 120 mg / mL nangibotide in different buffers in the presence of 190 mM Histidine after 24 h at room temperature* based on correction factor of 0.64 calculated from UPLC-PDA / UV %> dimer result at release (ie, 1.7° / o) and SEC-HPLC result (ie, 2.67°%) of the reference standard NGA01

[0376] Nangibotide dissolved within 21-36 min under most conditions and within 66 min at the maximum. No major precipitation or gelation occurred in any of the bulk products.

[0377] As can be seen in Tables 8 to 12, most pH measurements were within ±0.5 unit from the theoretical value.

[0378] Percentage of dimers UPLC-PDA / UV extrapolated using a correction factor from SEC-HPLC results remained within specifications under all conditions (i.e., < 5.0% by weight relative to the total weight of all peptides in the pharmaceutical composition (drug product)).

[0379] The assessment of process holding time stability (SPT) after 8 h at 4°C in citrate buffer, pH 5.7 and phosphate buffer, pH 7.0 containing Arg showed that the percentage of dimers remained within specifications under all conditions (i.e., < 5.0% by weight relative to the total weight of all peptides in the pharmaceutical composition (drug product)).

[0380] The assessment of ready-to-use stability (IUT) after 24 hours at room temperature (20°C) in citrate buffer, pH 5.7 and phosphate buffer, pH 7.0 containing Arg showed that the percentage of dimers remained within specifications under all conditions (i.e., < 10.0% by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition).

[0381] Therefore, these examples demonstrate that nangibotide is able to dissolve in the pharmaceutical compositions (drug product) as described herein. In addition, the pharmaceutical compositions (drug product) as described herein allows the formation of a low content of nangibotide dimers, in particular of equal to or less than 5.0% by weight relative to the total weight of all peptides in the pharmaceutical composition. In addition, the ready-to-use pharmaceutical compositions as described herein allows the formation of a low content of nangibotide dimers, in particular of equal to or less than 10.0% by weight relative to the total weight of all peptides in the pharmaceutical composition after 24 hours at room temperature.

Claims

1. CLAIMS1. A pharmaceutical composition comprising:- nangibotide, at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof, and at least one pharmaceutically acceptable buffer selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof, wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide.

2. A pharmaceutical composition comprising:- nangibotide, at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof, and at least one pharmaceutically acceptable buffer selected from the group consisting of: succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof, wherein the amount of the at least one amino acid is of at least 15% by weight relative to the dry weight of the pharmaceutical composition.

3. The pharmaceutical composition according to claim 1 or 2, wherein the at least one pharmaceutically acceptable buffer is selected from the group consisting of: succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

4. A pharmaceutical composition comprising:- nangibotide, at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof, andat least one pharmaceutically acceptable buffer selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof, wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide.

5. The pharmaceutical composition according to claim 4, wherein the at least one pharmaceutically acceptable buffer is selected from the group consisting of: citrate buffer, succinate buffer, acetate buffer, histidine buffer, phosphate buffer, tromethamine buffer, and any mixtures thereof.

6. The pharmaceutical composition according to any one of claims 1 to 5, wherein the amount of the at least one pharmaceutically acceptable buffer is less than 10%, preferably it ranges from 3% to 7%, more preferably it ranges from 3% to 5%, even more preferably from 2% to 5%, better it ranges from 4% to 6%, still better it is about 5%, by weight relative to the weight of nangibotide.

7. A pharmaceutical composition comprising: nangibotide, and at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof; wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide; and wherein the pH of said pharmaceutical composition ranges from 4.5 to 8.5.

8. A pharmaceutical composition comprising: nangibotide, and at least one amino acid selected from the group consisting of: arginine, lysine, histidine and any mixture thereof; wherein the amount of the at least one amino acid is of at least 15% by weight relative to the dry weight of the pharmaceutical composition; and wherein the pH of said pharmaceutical composition ranges from 4.5 to 8.5.

9. The pharmaceutical composition according to claim 8, wherein said pharmaceutical composition has a pH ranging from 4.5 to 8.5, preferably from 4.5 to 7.4, more preferably from 5 to 7, even more preferably from 5 to 6, or better said pharmaceutical composition has a pH of about 5.5.

10. The pharmaceutical composition according to any one of claims 7 to 9, wherein said pharmaceutical composition further comprises at least one pharmaceutically acceptable buffer, preferably the at least one pharmaceutically acceptable buffer being selected from the group consisting of: citrate buffer, succinate buffer, lactic acid buffer, acetate buffer, histidine buffer, MES buffer, phosphate buffer, tromethamine buffer, adipic acid buffer, glutamate buffer, and any mixtures thereof.

11. The pharmaceutical composition according to claim 10, wherein the amount of the at least one pharmaceutically acceptable buffer is less than 10%, preferably it ranges from 3% to 7%, more preferably it ranges from 3% to 5%, even more preferably from 2% to 5%, better it ranges from 4% to 6%, still better it is about 5%, by weight relative to the weight of nangibotide.

12. The pharmaceutical composition according to any one of claims 7 to 9, wherein said pharmaceutical composition does not comprise a buffer.

13. The pharmaceutical composition according to any one of claims 1 to 12, wherein said pharmaceutical composition comprises less than 5.0% of dimeric nangibotide by weight relative to the total weight of all peptides in said pharmaceutical composition.

14. The pharmaceutical composition according to any one of claims 1 to 13, wherein said pharmaceutical composition comprises from 600 mg to 2500 mg of nangibotide, preferably from 800 mg to 2500 mg of nangibotide, more preferably from 1200 mg to 2000 mg of nangibotide.

15. The pharmaceutical composition according to any one of claims 1 to 14, wherein said pharmaceutical composition comprises at least 65%, preferably from about65% to about 85%, more preferably from about 70% to about 80%, of nangibotide by weight relative to the dry weight of the pharmaceutical composition.

16. The pharmaceutical composition according to any one of claims 1 to 15, wherein the amount of the at least one amino acid ranges from 22% to 50%, preferably from 22% to 34%, more preferably from 22% to 30%, even more preferably from 23% to 29%, by weight relative to the weight of nangibotide.

17. The pharmaceutical composition according to any one of claims 1 to 16, wherein the pharmaceutical composition comprises (i) about 72.6% by weight of nangibotide, about 20% by weight of arginine, and about 3.9% by weight of citrate buffer (i.e., the sum of the weight of the citric acid and conjugate base) relative to the dry weight of the pharmaceutical composition or (ii) 71.3% by weight of nangibotide, about 19.7% by weight of arginine, and about 3.9% by weight of citrate buffer (i.e., the sum of the weight of the citric acid and conjugate base), relative to the total weight of the pharmaceutical composition.

18. The pharmaceutical composition according to any one of claims 1 to 17, wherein the pharmaceutical composition comprises about 1200 mg nangibotide, about 331 mg arginine, and about 65 mg sodium citrate.

19. The pharmaceutical composition according to any one of claims 1 to 18, wherein the pharmaceutical composition is a dry composition that when reconstituted with a liquid comprises about 120 mg / mL nangibotide, about 33.1 mg / mL arginine, and about 6.5 mg / ml sodium citrate.

20. The pharmaceutical composition according to any one of claims 1 to 19, wherein the pharmaceutical composition is a dry composition that when reconstituted with a liquid comprises:(i) about 120 mg / mL to about 200 mg / mL nangibotide(ii) at least one of amino acid selected from arginine, lysine, or histidine or a combination of thereof at a concentration of about 140 mM to about 300 mM; and(iii) a buffer selected from citrate buffer, a histidine buffer, a succinate buffer, an acetate buffer, a phosphate buffer, or tromethamine buffer from about 20 mM to about 60 mM; wherein the pH of the reconstituted pharmaceutical composition is about 4.5 to about 8.5.

21. The pharmaceutical composition according to claim 20, wherein(i) the buffer is about 45-55 mM acetate;(ii) the buffer is about 20-30 mM citrate;(iii) the buffer is about 20-30 mM histidine;(iv) the buffer is about 45-55 mM phosphate;(v) the buffer is about 45-55 mM succinate; or(vi) the buffer is about 45-60 mM tromethamine.

22. The pharmaceutical composition according to any one of claims 1 to 21, wherein the pharmaceutical composition is a dry composition that when reconstituted with a liquid comprises about 4 mg / mL or less of dimeric nangibotide, about 1 mg / mL or less of Des-Gln-LR12, and about 0.5 mg / mL or less of Met(O)12-LR12.

23. The pharmaceutical composition according to any one of claims 1 to 22, wherein said pharmaceutical composition comprises less than or equal to 1.2% of Des-Gln- LR12, by weight relative to the total weight of all peptides in said pharmaceutical composition and / or less than or equal to 1.0% of Met(O) 12-LR12 by weight relative to the total weight of all peptides in said pharmaceutical composition.

24. The pharmaceutical composition according to any one of claims 1 to 23, wherein said pharmaceutical composition is a powder.

25. A reconstituted pharmaceutical composition comprising the pharmaceutical composition according to any one of claims 1 to 24 and an aqueous medium, preferably said aqueous medium being selected from the group consisting of: water and a pharmaceutically acceptable aqueous solution, more preferably said aqueous medium being a saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.I l l26. A ready-to-use pharmaceutical composition comprising the pharmaceutical composition according to any one of claims 1 to 24 and an aqueous medium, preferably said aqueous medium being selected from the group consisting of: water and a pharmaceutically acceptable aqueous solution, more preferably said aqueous medium being a saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.

27. The ready-to-use pharmaceutical composition according to claim 26, wherein said ready-to-use pharmaceutical composition presents an osmolality of less than 500 mOsm / kg, preferably an osmolality ranging from 300 to 400 mOsm / kg, more preferably an osmolality ranging from 320 to 390 mOsm / kg, even more preferably an osmolality of about 380 mOsm / kg.

28. The ready-to-use pharmaceutical composition according to claim 26 or 27, wherein said ready-to-use pharmaceutical composition comprises equal to or less than 10.0%, preferably equal to or less than 5.0%, of nangibotide dimers by weight relative to the total weight of all peptides in said ready-to-use pharmaceutical composition.

29. The pharmaceutical composition according to any one of claims 1 to 24 or the reconstituted pharmaceutical composition according to claim 25 or the ready-to-use pharmaceutical composition according to any one of claims 26 to 28, for use as a medicament.

30. The pharmaceutical composition according to any one of claims 1 to 24 or the reconstituted pharmaceutical composition according to claim 25 or the ready-to-use pharmaceutical composition according to any one of claims 26 to 28, for use in the treatment of a disease in a subject in need thereof, wherein said disease is selected from the group consisting of: inflammatory diseases, preferably selected from the group consisting of: septic shock, sepsis, severe sepsis, systemic inflammatory response syndrome (SIRS), acute respiratory distress syndrome (ARDS), hemorrhagic shock, ischemiareperfusion and pancreatitis;cardiovascular diseases, preferably selected from the group consisting of: myocardial infarction, cerebral infarction, acute myocardial infarction, ischemia, coronary heart disease, acute coronary syndrome, stroke, aneurysm, stable angina pectoris, effort angina pectoris, cardiomyopathy, hypertensive heart disease, chronic heart failure, acute heart failure, cor pulmonale, cardiac dysrhythmias, inflammatory heart disease (such as endocarditis and myocarditis), peripheral arterial disease, SIRS-associated myocardial and / or vascular dysfunction, and atherosclerosis; and infectious diseases, preferably COVID- 19.

31. The pharmaceutical composition or reconstituted pharmaceutical composition or ready-to-use pharmaceutical composition, for use according to claim 30, wherein said pharmaceutical composition or reconstituted pharmaceutical or ready-to-use pharmaceutical composition is for administration to said subject in need thereof via intravenous route of administration at a rate ranging from 0.1 to 3 mg / kg / hr (mg of nangibotide per kilogram of bodyweight of the subject per hour).

32. A process for preparing a pharmaceutical composition according to any one of claims 1 to 24, comprising the steps of: a) mixing nangibotide with at least one amino acid selected from the group consisting of: arginine, lysine and histidine, wherein the amount of the at least one amino acid is of at least 22% by weight relative to the weight of nangibotide, thereby leading to a bulk solution; b) optionally adjusting the pH of the bulk solution obtained in step a) to a pH ranging from 5.5 to 6.5 or ranging from 5.0 to 7.0, preferably ranging from 5.5 to 6.0, preferably a pH of 5.5, preferably said pH adjustment being carried out using HC1; and c) optionally lyophilizing the bulk solution obtained in step a) or in step b) to obtain a pharmaceutical composition in a powder form.

33. A process for preparing a reconstituted pharmaceutical composition according to claim 25, comprising the steps of:- carrying out the process according to claim 32; then- adding an aqueous medium to the pharmaceutical composition in a powder form obtained in step c) of the process according to claim 32, to obtain an aqueous reconstituted pharmaceutical composition, preferably qsp 10 mL in a 20 mb vial, preferably said aqueous medium is selected from the group consisting of: water and saline water solution, more preferably said aqueous medium is selected from the group consisting of: water and saline water solution having 0.9% sodium chloride by weight relative to the weight of the saline water solution.

34. A process for preparing a ready-to-use pharmaceutical composition according to any one of claims 26 to 28, comprising the steps of:- carrying out the process according to claim 33 to obtain a reconstituted pharmaceutical composition; then- diluting the reconstituted pharmaceutical composition in an aqueous medium, preferably in a syringe or in an infusion bag, for example a syringe or in an infusion bag having a total volume of 50 mL.

35. A purified nangibotide substance comprising equal to or less than 2.7% of dimeric nangibotide by weight relative to the total weight of all peptides of said purified nangibotide substance.

36. The purified nangibotide substance according to claim 35, wherein at least 85% of peptides by weight relative to the total weight of the purified nangibotide substance.

37. The purified nangibotide substance according to claim 35 or 36, wherein said purified nangibotide substance comprises: equal to or less than about 1.2%, preferably equal to or less than about 1.0%, more preferably equal to or less than about 0.5%, of Des-Gln-LR12, by weight relative to the total weight of all peptides in said purified nangibotide substance; and / orequal to or less than about 1.0%, preferably equal to or less than about 0.75%, more preferably equal to or less than about 0.5%, of Met(O)12-LR12, by weight relative to the total weight of all peptides in said purified nangibotide substance.

Images