EA042812B1 - SOLID FORMS AND COMBINED COMPOSITIONS CONTAINING A BETA-LACTAMASE INHIBITOR AND THEIR APPLICATIONS - Google Patents

Description

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The present patent application claims priority over U.S. Provisional Application No. 62/467,750, filed March 06, 2017; U.S. Provisional Application No. 62/467,752, filed March 6, 2017, U.S. Provisional Application No. 62/564,989, filed September 28, 2017, and U.S. Provisional Application No. 62/564,990, filed September 28, 2017 .; each of which is incorporated herein by reference in its entirety.

Application for federal research funding

The invention was made with the support of the Government of the United States of America under contract number R43AI096679 from the National Institutes of Health (NIH), under contract number R44AI096613c from the National Institutes of Health (NIH) and under contract number HHSN272201300019C from the National Institutes of Health (NIH). The government has certain rights to the present invention.

The field of technology to which the invention belongs

The invention relates to pharmaceutical compositions containing boron-containing compounds, polymorphic forms, preparations and their use as beta-lactamase enzyme inhibitors and as antibacterial agents in combination with a beta-lactam antibiotic.

Background of the Invention

Antibiotics are the most effective drugs for the treatment of bacterial infections. They are widely used in clinical practice due to their good antibacterial effect with limited side effects. Among them, a class of beta-lactam antibiotics (eg, penicillins, cephalosporins, monobactams, and carbapenems) is preferred due to their bactericidal effect and the fact that their target is absent in eukaryotic cells, resulting in low toxicity. To counteract the effectiveness of various beta-lactams, bacteria have evolved to produce variants of beta-lactam inactivating enzymes called beta-lactamases, as well as the ability to share this tool both vertically and horizontally between and within species. These beta-lactamases belong to the serine or metallo-beta-lactamases, respectively, based on the presence of the key serine or zinc at the active site of the enzyme. The rapid induction, selection, and spread of this bacterial resistance mechanism may severely limit the entire class of beta-lactam treatment options in the hospital setting and familiar social setting. There is a need for effective and safe therapeutic agents that can treat such resistant infections.

The essence of the invention

This document discloses a pharmaceutical composition containing:

(i) (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine- 8-carboxylic acid

a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof; and (ii) cefepime.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition is formulated as a uniform liquid suitable for injection.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition further comprises a pharmaceutically acceptable adjuvant.

According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is an amino acid or a monosaccharide derivative.

According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is L-arginine.

According to some embodiments of the pharmaceutical compositions, the pharmaceutically acceptable adjuvant is meglumine.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition further comprises an aqueous carrier.

In accordance with some embodiments of the pharmaceutical composition, the aqueous carrier is water for injection, 0.9% sodium chloride injection, 5% dextrose injection, 10% dextrose injection, sodium lactate injection, 5% dextrose injection, and 0 .9% sodium chloride, Lactated Ringer's Injection with 5% Dextrose, Sodium Chloride/Sodium Acetate/Sodium Gluconate/Potassium Chloride/Magnesium Chloride Injection, Sodium Chloride/Potassium Acetate/Magnesium Acetate Injection in 5% Dextrose, or any of these combinations.

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In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition has a pH of about 4 to about 9.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition has a pH of about 4 to about 6.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition is formulated as a powder for reconstitution.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical agent further comprises a pharmaceutically acceptable adjuvant.

According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is an amino acid or a monosaccharide derivative.

According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is L-arginine.

According to some embodiments of the pharmaceutical compositions, the pharmaceutically acceptable adjuvant is meglumine.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition is suitable for injection after reconstitution with an aqueous vehicle.

In accordance with some embodiments of the pharmaceutical composition, the aqueous carrier is water for injection, 0.9% sodium chloride injection, 5% dextrose injection, 10% dextrose injection, sodium lactate injection, 5% dextrose injection, and 0 .9% sodium chloride, Lactated Ringer's Injection with 5% Dextrose, Sodium Chloride/Sodium Acetate/Sodium Gluconate/Potassium Chloride/Magnesium Chloride Injection, Sodium Chloride/Potassium Acetate/Magnesium Acetate Injection in 5% Dextrose, or any of these combinations.

A pharmaceutical composition according to any one of 0-0, wherein the pharmaceutical composition is stable at about 25±2°C/RH 60%±RH 5% for at least 12 months.

According to some embodiments of the pharmaceutical composition (R)-3-(2(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1, 2]oxaborinine-8-carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate are crystalline.

According to some embodiments of the pharmaceutical composition, crystalline (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2Hbenzo[e][1, 2]oxaborinine-8-carboxylic acid is present in the form of its dihydrochloride or solvate.

According to some embodiments of the pharmaceutical composition, crystalline dihydrochloride (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4dihydro-2H-benzo[e][1 The ,2]oxaborinine-8-carboxylic acid is present in the form of a solvate monohydrate.

In accordance with some embodiments of the pharmaceutical composition, the crystalline form has an X-ray powder diffraction (XRPD) pattern substantially the same as that shown in FIG. 2.

In accordance with some embodiments of the pharmaceutical composition, the crystalline form has an X-ray powder diffraction (XRPD) pattern substantially the same as that shown in FIG. 3.

In accordance with some embodiments of the pharmaceutical composition, the crystalline form is characterized by an X-ray powder diffraction (XRPD) pattern containing characteristic peaks with a 2θ value of about 7.0 ± 0.1°, with a 2θ value of about 14.1 ± 0.1°, with a 2θ value about 20.2±0.1°, with a 2θ value of about 24.6±0.1°, and with a 2θ value of about 27.7±0.1°.

In accordance with some embodiments of the crystalline pharmaceutical composition, the X-ray powder diffraction (XRPD) pattern further comprises characteristic peaks with a 2θ value of about 10.5±0.1°, with a 2θ value of about 18.9±0.1°, with a 2θ value of about 23.7±0.1°, with a 2θ value of about 25.6±0.1° and with a 2θ value of about 29.6±0.1°.

According to some embodiments of the pharmaceutical composition, the pharmaceutical composition contains about 500 mg of (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H- benzo[e][1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof.

According to some embodiments of the pharmaceutical composition, the pharmaceutical composition contains about 750 mg of (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H- benzo[e][1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition contains about 2 g of cefepime.

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Also disclosed herein is a method for treating a bacterial infection in a subject in need thereof, comprising administering to the subject the pharmaceutical composition described herein.

In accordance with some embodiments of a method for treating a bacterial infection, the pharmaceutical composition is administered via intravenous (IV) infusion.

In accordance with some embodiments of a method for treating a bacterial infection, the infusion period of the pharmaceutical composition is about 2 hours.

This document also discloses the crystalline form of (R)-3-(2-(trans-4-(2aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2] oxaborinine-8-carboxylic acid

its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate.

This document also discloses the crystalline form of (R)-3-(2-(trans-4-(2aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2] oxaborinine-8-carboxylic acid

its dihydrochloride or solvate.

According to some embodiments of the crystal form, (R)-3(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1 The ,2]oxaborinine 8-carboxylic acid is present in the form of a monohydrate solvate.

In accordance with some embodiments of the crystalline form, the crystalline form has an X-ray powder diffraction (XRPD) pattern substantially the same as that shown in FIG. 2.

In accordance with some embodiments of the crystalline form, the crystalline form has an X-ray powder diffraction (XRPD) pattern substantially the same as that shown in FIG. 3.

In accordance with some embodiments of the crystalline form, the crystalline form is characterized by an X-ray powder diffraction (XRPD) pattern containing characteristic peaks with a 2Θ value of about 7.0 ± 0.1°, with a 2Θ value of about 14.1 ± 0.1°, with a 2Θ value about 20.2±0.1°, with a 2Θ value of about 24.6±0.1° and with a 2Θ value of about 27.7±0.1°.

In accordance with some embodiments of the crystal form, the X-ray powder diffraction (XRPD) pattern further comprises characteristic peaks with a 2Θ value of about 10.5±0.1°, with a 2Θ value of about 18.9±0.1°, with a 2Θ value of about 23, 7±0.1°, with a 2Θ value of about 25.6±0.1° and with a 2Θ value of about 29.6±0.1°.

According to some embodiments of the crystal form, (R)-3(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)hydroxy-3,4-dihydro-2H-benzo[e] dihydrochloride [1,2] oxaborinine-8carboxylic acid is anhydrous.

In accordance with some embodiments of the crystalline form, the crystalline form has an X-ray powder diffraction (XRPD) pattern substantially the same as that shown in FIG. 8.

In accordance with some embodiments of the crystalline form, the crystalline form is characterized by an X-ray powder diffraction (XRPD) pattern containing characteristic peaks with a 2Θ value of about 7.3±0.1°, with a 2Θ value of about 14.5±0.1°, with a 2Θ value about 18.0±0.1°, with a 2Θ value of about 19.7±0.1°, with a 2Θ value of about 24.0±0.1°, and with a 2Θ value of about 27.3±0.1°.

In accordance with some embodiments of the crystalline form, the X-ray powder diffraction (XRPD) pattern further comprises characteristic peaks with a 2Θ value of about 10.8±0.1°, with a 2Θ value of about 16.6±0.1°, with a 2Θ value of about 19, 6±0.1°, with a 2Θ value of about 23.3±0.1°, with a 2Θ value of about 24.3±0.1°, and with a 2Θ value of about 29.3±0.1°.

This document also discloses a pharmaceutical composition containing:

(i) the crystalline form disclosed herein, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof; and (ii) cefepime.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition is formulated as a uniform liquid suitable for injection.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition further comprises an aqueous carrier. In some embodiments of the pharmaceutical composition, the pharmaceutical composition has a pH of about to about 6. In some embodiments of the pharmaceutical composition, the pharmaceutical composition is formulated as a powder for reconstitution.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition is suitable for injection after reconstitution with an aqueous vehicle.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition further comprises a pharmaceutically acceptable adjuvant.

According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is an amino acid or a monosaccharide derivative.

According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is L-arginine.

According to some embodiments of the pharmaceutical compositions, the pharmaceutically acceptable adjuvant is meglumine.

Brief description of the drawings

New features of the present disclosure are set forth in characteristic detail in the claims of the appended claims. A better understanding of the features and advantages of the present disclosure will be achieved in view of the following detailed description, which sets forth illustrative embodiments that use the principles of the present disclosure, and the accompanying drawings, in which:

In FIG. 1A shows the solubility values of Compound 1 dihydrochloride in IPA/water mixtures at 5, 25, 40 and 50°C as a function of IPA/water content.

In FIG. 1B shows the solubility of compound 1 dihydrochloride in IPA/water mixtures as a function of temperature.

In FIG. 2 shows an X-ray powder diffraction pattern obtained for form 1 at room temperature.

In FIG. 3 shows an X-ray powder diffraction pattern modeled from single crystal data for form 1 obtained at 100K.

In FIG. 4 shows a TGMS thermogram (at a heating rate of 10°C/min) for form 1.

In FIG. 5 shows a DSC thermogram (at a heating rate of 10°C/min) for form 1.

In FIG. 6 shows the DVS isotherm for form 1.

In FIG. 7 shows stacked XRPD patterns for Compound 1 dihydrochloride at variable RH (Relative Humidity).

In FIG. 8 shows an X-ray powder diffraction pattern obtained for form 2 at room temperature.

In FIG. 9 shows the DSC thermogram for form 2.

In FIG. 10A shows the recorded weight changes as a function of the relative humidity level. Solid samples of form 2 were incubated at the indicated value of relative humidity at 25°C, at which time changes in mass were continuously recorded.

In FIG. 10B shows the recorded weight changes as a function of the relative humidity level. Conversion rates were calculated and plotted against the relative humidity value (from the linear portion of the curve in FIG. 10A). The line regression coefficient is 0.9995.

In FIG. 11 shows the water content of compound 1 dihydrochloride after incubation at various temperatures and RH conditions.

In FIG. 12 shows the X-ray powder diffraction pattern obtained for Form 3.

In FIG. 13 shows the X-ray powder diffraction pattern obtained for form 4.

In FIG. 14 shows the X-ray powder diffraction pattern obtained for form 5.

In FIG. 15 shows the X-ray powder diffraction pattern obtained for form 6.

Detailed disclosure of the invention

Disclosed herein are the crystalline forms of (R)-3-(2-(trans-4-(2aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine -8-carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate.

This document also discloses pharmaceutical compositions containing crystalline forms of (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2Hbenzo[e][ 1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof.

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This document also discloses pharmaceutical compositions containing solid forms of (R)-3(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e] [ 1,2]oxaborinine 8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof.

In the context of this document, (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine-8 -carboxylic acid is as shown in the structure below

According to some embodiments, (R)-3-(2-(trans-4-(2aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine -8-carboxylic acid is also referred to as compound 1. According to some embodiments, compound 1 exists in an equilibrium between a closed ring form (shown above) and an open acyclic form

According to some embodiments, Compound 1 binds to intramolecular dimers, trimers, and any combination thereof. In accordance with some embodiments, Compound 1 is present in the form of a pharmaceutically acceptable salt. In accordance with some embodiments, the pharmaceutically acceptable salt is the hydrochloride. In accordance with some embodiments, the pharmaceutically acceptable salt is a dihydrochloride. In accordance with some embodiments, compound 1 is present in the form of a pharmaceutically acceptable solvate. Solvates contain either stoichiometric or non-stoichiometric amounts of solvent and, in some embodiments, are formed during a crystallization process with pharmaceutically acceptable solvents such as water or organic solvents. In some embodiments, the solvent is not covalently bonded to compound 1. In some embodiments, the solvent is covalently bonded to compound 1. Hydrates are formed when the solvent is water or alcoholates are formed when the solvent is an alcohol. In general, the solvated forms are considered equivalent to the unsolvated forms for the purposes of the compounds and methods presented herein. According to some embodiments, the pharmaceutically acceptable solvate is a monohydrate solvate.

crystalline forms.

Disclosed herein are the crystalline forms of (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2 ]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof. In some embodiments, the crystalline form is Form 1. In some embodiments, the crystalline form is Form 2. In some embodiments, the crystalline form is Form 3. In some embodiments, the crystalline form is Form 4. In some embodiments, the crystalline form is Form 5. In some embodiments, the crystalline form is Form 6.

Form 1.

According to some embodiments, the dihydrochloride salt of Compound 1 is converted to a pure and stable crystalline form (Form 1) under controlled conditions. In some embodiments, form 1 is the monohydrate form of the dihydrochloride salt of compound 1. In some embodiments, the powder x-ray diffraction pattern (XRPD) of form 1 is essentially the same as shown in FIG. 2, with the corresponding tabulated peak data shown in Table 2. 1. In accordance with some embodiments, the X-ray powder diffraction pattern (XRPD) of Form 1 is substantially the same as shown in FIG. 3. Powder X-ray diffraction pattern (XRPD) shown in FIG. 2 is essentially the same as the X-ray powder diffraction (XRPD) pattern shown in FIG. 3.

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Table 1

2ΘΗ d value [A] Intensity [%] 2ΘΗ d[A] value Intensity [%] 7.0 12.54 19 28.6 3.12 25 8.6 10.5 11.2 12.7 13.1 13.6 14.1 14.9 16.1 16.9 17.2 17.6 18.9 19.3 20.2 20.6 21.2 21.7 22.1 22.6 22.9 23.7 24.6 25.0 25.6 25.9 26.1 10.29 8.39 7.93 6.96 6.73 6.50 6.28 5.93 5.51 5.24 5.15 5.02 4.69 4.60 4.39 4.31 4.19 4.09 4.03 3.93 3.88 3.75 3.61 3.56 3.48 3.44 3.41 25 88 12 42 12 17 16 thirty 68 18 AND 14 52 100 42 46 15 17 23 20 45 88 27 34 42 34 34 28.8 29.6 30.1 30.3 31.2 31.4 31.5 31.9 32.7 32.8 33.0 33.4 33.6 33.8 34.2 34.4 34.7 35.0 35.7 36.1 36.4 36.6 36.8 37.2 37.5 37.7 38.0 3.09 3.02 2.97 2.94 2.87 2.85 2.84 2.80 2.73 2.73 2.71 2.68 2.66 2.65 2.62 2.60 2.58 2.56 2.51 2.48 2.47 2.45 2.44 2.41 2.40 2.39 2.37 13 21 14 17 12 14 13 18 16 15 27 15 12 18 13 12 14 13 16 9 10 18 8 8 23 7 7 26.5 3.37 20 38.2 2.36 7 26.7 3.33 25 38.6 2.33 16 26.9 3.32 19 39.1 2.30 7 27.7 3.22 16 39.6 2.28 19 28.1 3.17 26 39.9 2.26 12

In accordance with some embodiments, Form 1 shows an X-ray powder diffraction pattern that includes at least three characteristic peaks selected from peaks with a 2Θ value of about 7.0±0.1°, with a 2Θ value of about 10.5±0, 1°, with a 2Θ value of about 14.1 ±0.1°, with a 2Θ value of about 18.9±0.1°, with a 2Θ value of about 20.2±0.1°, with a 2Θ value of about 23.7± 0.1°, with a 2Θ value of about 24.6±0.1°, with a 2Θ value of about 25.6±0.1°, with a 2Θ value of about 27.7±0.1° and with a 2Θ value of about 29, 6±0.1°. In accordance with some embodiments, Form 1 shows an X-ray powder diffraction (XRPD) pattern that contains characteristic peaks with a 2Θ value of about 7.0±0.1°, with a 2Θ value of about 14.1±0.1°, with a 2Θ value of about 20.2±0.1°, with a 2Θ value of about 24.6±0.1° and with a 2Θ value of about 27.7±0.1°. In accordance with some embodiments, Form 1 shows an X-ray powder diffraction (XRPD) pattern that further contains characteristic peaks with a 2Θ value of about 10.5±0.1°, with a 2Θ value of about 18.9±0.1°, with a 2Θ value about 23.7±0.1°, with a 2Θ value of about 25.6±0.1° and with a 2Θ value of about 29.6±0.1°. In accordance with some embodiments, Form 1 shows an X-ray powder diffraction (XRPD) pattern that contains characteristic peaks with a 2Θ value of about 10.5±0.1°, with a 2Θ value of about 12.7±0.1°, with a 2Θ value of about 16.1±0.1°, with a 2Θ value of about 18.9±0.1°, with a 2Θ value of about 19.3±0.1°, with a 2Θ value of about 20.2±0.1°, with a value 2Θ is about 20.6±0.1°, with a 2Θ value of about 22.9±0.1°, with a 2Θ value of about 23.7±0.1°, and with a 2Θ value of about 25.6±0.1°. In accordance with some embodiments, Form 1 shows an X-ray powder diffraction (XRPD) pattern that contains characteristic peaks with a 2Θ value of about 10.5±0.1°, with a 2Θ value of about 12.7±0.1°, with a 2Θ value of about 14.9±0.1°, with a 2Θ value of about 16.1±0.1°, with a 2Θ value of about 18.9±0.1°, with a 2Θ value of about 19.3±0.1°, with a value 2Θ about 20.2±0.1°, with 2Θ value about 20.6±0.1°, with 2Θ value about 22.9±0.1°, with 2Θ value about 23.7±0.1°, with a 2Θ value of about 25.0±0.1°, with a 2Θ value of about 25.6±0.1°, with a 2Θ value of about 25.9±0.1° and with a 2Θ value of about 26.1±0.1 °. In accordance with some embodiments, Form 1 shows an X-ray powder diffraction (XRPD) pattern that contains characteristic peaks with a 2Θ value of about 8.6±0.1°, with a 2Θ value of about 10.5±0.1°, with a 2Θ value of about 12.7±0.1°, with a 2Θ value of about 14.9±0.1°, with a 2Θ value of about 16.1±0.1°, with a 2Θ value of about 18.9±0.1°, with a value 2Θ about 19.3±0.1°, with a value of 2Θ about

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20.2±0.1°, with a 2Θ value of about 20.6±0.1°, with a 2Θ value of about 22.1 ±0.1°, with a 2Θ value of about

22.6±0.1°, with a 2Θ value of about 22.9±0.1°, with a 2Θ value of about 23.7±0.1°, with a 2Θ value of about

24.6±0.1°, with a 2Θ value of about 25.0±0.1°, with a 2Θ value of about 25.6±0.1°, with a 2Θ value of about

25.9±0.1°, with a value of 20 about 26.1±0.1°, with a value of 20 about 26.5±0.1°, with a value of 2Θ about

26.7±0.1°, with a 2Θ value of about 28.1 ±0.1°, with a 2Θ value of about 28.6±0.1°, with a 2Θ value of about

29.6±0.1°, with a 2Θ value of about 33.0±0.1° and with a 2Θ value of about 37.5±0.1°.

Form 2.

In some embodiments, the dihydrochloride salt of Compound 1 can be converted to a pure and stable crystalline form (Form 2) under controlled conditions. In some embodiments, Form 2 is the anhydrous form of the dihydrochloride salt of Compound 1. The powder X-ray diffraction (XRPD) pattern of Form 2 is essentially the same as shown in FIG. 8, with the corresponding tabulated peak data presented in Table 1. 2.

table 2

2Θ [°] d value [A] Intensity [%] 2Θ [°] d[A] value Intensity [%] 7.3 12.17 84 28.2 3.16 17 8.7 10.11 23 28.6 3.12 22 10.8 8.21 81 28.9 3.09 28 11.2 7.86 33 29.3 3.04 31 12.9 6.87 47 30.2 2.96 17 13.5 6.56 20 30.4 2.94 24 13.8 6.42 20 30.6 2.92 16 14.5 6.09 32 31.2 2.86 32 15.2 5.84 23 31.7 2.82 17 15.9 5.56 45 31.8 2.81 18 16.3 5.42 63 32.2 2.78 31 16.6 5.34 100 32.3 2.77 32 17.2 5.15 22 32.6 2.75 20 17.5 5.06 18 33.3 2.69 28 18.0 4.91 26 33.5 2.67 20 19.2 4.62 56 34.1 2.63 25 19.6 4.54 64 34.5 2.60 19 19.7 4.50 79 34.8 2.57 22 20.5 4.32 44 35.1 2.55 13 21.1 4.20 95 35.3 2.54 14 21.4 4.15 26 35.6 2.52 14 21.6 4,I 18 35.8 2.51 16 22.1 4.01 18 36.5 2.46 21 22.8 3.89 thirty 36.9 2.43 16 23.1 3.84 29 37.3 2.41 14 23.3 3.82 22 37.9 2.37 14 24.0 3.71 75 38.2 2.35 14 24.3 3.66 62 38.5 2.34 AND 25.1 3.54 22 38.8 2.32 AND 25.3 3.51 19 39.2 2.30 12 25.9 3.43 31 39.3 2.29 15 26.3 3.38 32 39.7 2.27 17 26.6 3.35 thirty 40.2 2.24 10 26.8 3.32 26 40.6 2.22 AND 27.0 3.30 24 40.8 2.21 10 27.3 3.26 36 41.3 2.18 AND 27.8 3.21 22

In accordance with some embodiments, Form 2 shows an X-ray powder diffraction pattern that includes at least three characteristic peaks selected from peaks with a 2Θ value of about 7.3±0.1°, with a 2Θ value of about 10.8±0, 1°, with a 2Θ value of about 14.5±0.1°, with a 2Θ value of about 16.6±0.1°, with a 2Θ value of about 18.0±0.1°, with a 2Θ value of about 19.6± 0.1°, with a 2Θ value of about 19.7±0.1°, with a 2Θ value of about 23.3±0.1°, with a 2Θ value of about 24.0±0.1°, with a 2Θ value of about 24, 3±0.1°, with a 2Θ value of about 27.3±0.1° and with a 2Θ value of about

29.3±0.1°. In accordance with some embodiments, Form 2 exhibits a powder

- 7 042812 X-ray diffraction pattern (XRPD), which contains characteristic peaks with a 2Θ value of about 7.3±0.1°, with a 2Θ value of about 14.5±0.1°, with a 2Θ value of about 18.0±0.1 °, with a 2Θ value of about 19.7±0.1°, with a 2Θ value of about 24.0±0.1° and with a 2Θ value of about 27.3±0.1°. In accordance with some embodiments, Form 2 shows an X-ray powder diffraction (XRPD) pattern that further contains characteristic peaks with a 2Θ value of about 10.8±0.1°, with a 2Θ value of about 16.6±0.1°, with a 2Θ value about 19.6±0.1°, with a 2θ value of about 23.3±0.1°, with a 2θ value of about 24.3±0.1°, and with a 2θ value of about 29.3±0.1°. In accordance with some embodiments, Form 2 shows an X-ray powder diffraction (XRPD) pattern that contains characteristic peaks with a 2Θ value of about 7.3±0.1°, with a 2Θ value of about 10.8±0.1°, with a 2Θ value of about 12.9±0.1°, with a 2Θ value of about 15.9±0.1°, with a 2Θ value of about 16.3±0.1°, with a 2Θ value of about 16.6±0.1°, with a value 2Θ about 19.2±0.1°, with 2Θ value about 19.7±0.1°, with 2Θ value about 20.5±0.1°, with 2Θ value about 21.1±0.1°, with a 2Θ value of about 24.0±0.1° and with a 2Θ value of about 24.3±0.1°. In accordance with some embodiments, Form 2 shows an X-ray powder diffraction (XRPD) pattern that contains characteristic peaks with a 2Θ value of about 7.3±0.1°, with a 2Θ value of about 10.8±0.1°, with a 2Θ value of about 11.2±0.1°, with a 2Θ value of about 12.9±0.1°, with a 2Θ value of about 14.5±0.1°, with a 2Θ value of about 15.9±0.1°, with a value 2Θ about 16.3±0.1°, with 2Θ value about 16.6±0.1°, with 2Θ value about 19.2±0.1°, with 2Θ value about 19.7±0.1°, with a 2Θ value of about 20.5±0.1°, with a 2Θ value of about 21.1±0.1°, with a 2Θ value of about 22.8±0.1°, with a 2Θ value of about 24.0±0.1 °, with a 2Θ value of about 24.3±0.1°, with a 2Θ value of about 25.9±0.1°, with a 2Θ value of about 26.3±0.1°, with a 2Θ value of about 26.6±0 ,1°, with a 2Θ value of about 29.3±0.1° and with a 2Θ value of about 32.3±0.1°.

Form 3.

According to some embodiments, the dihydrochloride salt of Compound 1 can be converted to a novel form (Form 3) under controlled conditions. The X-ray powder diffraction pattern (XRPD) of Form 3 is essentially the same as shown in FIG. 12, with the corresponding tabulated peak data shown in Table 12. 3.

Table 3

2θ[°] d value [A] Intensity [%1 8.68 10.18 100 15.92 5.56 82.5 18.91 4.69 81.8 21.58 4,P 65.7 22.86 3.89 83.7

Form 4.

According to some embodiments, the dihydrochloride salt of Compound 1 can be converted to a new form (Form 4) under controlled conditions. The X-ray powder diffraction (XRPD) pattern 4 is essentially the same as shown in FIG. 13, with the corresponding tabulated peak data shown in Table 13. 4.

Table 4

2Θ [°] d value [A] Intensity [%] 8.34 10.58 72.7 8.75 10.11 71.0 8.82 10.01 72.0 9.22 9.58 80.9 18.94 4.68 62.2 20.38 4.35 72.0 22.90 3.88 100

Form 5.

According to some embodiments, the dihydrochloride salt of Compound 1 can be converted to a novel form (Form 5) under controlled conditions. The X-ray powder diffraction pattern (XRPD) of Form 5 is essentially the same as shown in FIG. 14, with the corresponding tabulated peak data shown in Table 14. 5.

- 8 042812

Table 5

2Θ [°] d value [A] Intensity [%] 7.90 11.18 47.3 10.65 8.29 62.9 15.86 5.57 91.2 18.82 4.71 100

Form 6.

In accordance with some embodiments, the dihydrochloride salt of Compound 1 can be converted to a crystalline form (Form 6) under controlled conditions. The X-ray powder diffraction pattern (XRPD) of Form 6 is essentially the same as shown in FIG. 15, with the corresponding tabulated peak data presented in Table 1. 6.

_________________Table 6

2Θ [°] d value [A] Intensity [%] 8.74 10.11 100 8.81 10.02 72.2 13.38 6.61 48.8 16.30 5.43 49.1 20.06 4.42 77.6 22.86 3.89 87.7 28.30 3.15 44.7

Embodiments

Embodiment P1. Crystalline form of (R)-3-(2-(trans-4-(2aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborinine-8-carboxylic acids

its pharmaceutically acceptable salt and/or solvate.

Embodiment P2. Crystalline form of (R)-3-(2-(trans-4-(2aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborinine-8-carboxylic acids

its dihydrochloride and/or solvate.

Embodiment P3. A crystalline form according to embodiment P2, wherein (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro2H-benzo[e][1 The ,2]oxaborinine-8-carboxylic acid is present in the form of a solvate monohydrate.

Embodiment P4. A crystalline form according to embodiment P3, wherein the crystalline form has at least one of the following properties:

(a) an X-ray powder diffraction (XRPD) pattern essentially the same as shown in FIG. 2; or (b) an X-ray powder diffraction pattern (XRPD) substantially the same as shown in FIG. 3; or (c) an X-ray powder diffraction (XRPD) pattern containing characteristic peaks with a 2Θ value of about 7.0±0.1°, with a 2Θ value of about 14.1±0.1°, with a 2Θ value of about 20.2±0, 1°, with a 2Θ value of about 24.6±0.1° and with a 2Θ value of about 27.7±0.1°.

Embodiment P5. The crystalline form according to embodiment P4, wherein the powder X-ray diffraction pattern (XRPD) additionally contains characteristic peaks with a 2Θ value of about 10.5±0.1°, with a 2Θ value of about 18.9±0.1°, with a 2Θ value of about 23 .7±0.1°, with a 2Θ value of about 25.6±0.1° and with a 2Θ value of about 29.6±0.1°.

Embodiment P6. A crystalline form according to any one of embodiments P3-P5, wherein the crystalline form is Form 1.

Embodiment P7. A crystalline form according to embodiment P2, wherein (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro2H-benzo[e][1 ,2]oxaborinine-8-carboxylic acid is anhydrous.

Embodiment P8. The crystalline form according to embodiment P7, wherein the crystalline form has at least one of the following properties:

- 9 042812 (a) X-ray powder diffraction pattern (XRPD) essentially the same as shown in FIG. 8 or (b) an X-ray powder diffraction (XRPD) pattern containing characteristic peaks with a 2θ value of about 7.3±0.1°, with a 2θ value of about 14.5±0.1°, with a 2θ value of about 18.0±0 .1°, with a 2θ value of about 19.7±0.1°, with a 2θ value of about 24.0±0.1°, and with a 2θ value of about 27.3±0.1°.

Embodiment P9. The crystalline form according to embodiment P8, wherein the X-ray powder diffraction pattern (XRPD) further comprises characteristic peaks with a 2θ value of about 10.8±0.1°, with a 2θ value of about 16.6±0.1°, with a 2θ value of about 19 .6±0.1°, with a 2θ value of about 23.3±0.1°, with a 2θ value of about 24.3±0.1°, and with a 2θ value of about 29.3±0.1°.

Embodiment P10. A crystalline form according to any one of embodiments P7-P9, wherein the crystalline form is Form 2.

Pharmaceutical compositions

This document discloses a pharmaceutical composition comprising: (i) (R)-3-(2-(trans-4-(2aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e] [1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof; and (ii) cefepime.

According to some embodiments, (R)-3-(2-(trans-4-(2aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine -8-carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate are amorphous. According to some embodiments, (R)-3-(2-(trans4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine -8 carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate are crystalline. This document also discloses a pharmaceutical composition containing: (i) crystalline (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H- benzo[e][1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof; and (ii) cefepime.

According to some embodiments of the pharmaceutical composition, (R)-3-(2-(mrans4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2 ]oxaborinin-8-carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate is present in the form of a crystalline form disclosed in this document. According to some embodiments, the crystalline form of (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1 ,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof is Form 1. In some embodiments, crystalline form (R)-3-(2(trans-4-(2 -aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborinine-8carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate is Form 2 In accordance with some embodiments, the pharmaceutical compositions described herein are provided in unit dosage form. In the context of this document, a unit dosage form is a composition containing an amount of Compound 1 that is suitable for administration to an animal subject, preferably a mammal, in a single dose in accordance with good medical practice. The preparation in a unit or unit dosage form, however, does not imply that the dosage form is administered once a day or once during the period of therapy. It is contemplated that such dosage forms are administered one, two, three, or more times per day, and may be administered as an infusion over a period of time (e.g., from about 30 minutes to about 2-6 hours) or may be administered as a continuous infusion, and they can be given more than once during the period of implementation of therapy, although a single administration is not specifically excluded. In some embodiments, Compound 1 and cefepime are formulated in separate containers. In some embodiments, Compound 1 and cefepime are formulated in the same container.

In some embodiments, administration of compound 1, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof is via any of the accepted modes of administration for agents that serve similar functions, including, but not limited to, orally, subcutaneously, intravenously, intranasally, topically, transdermally, intraperitoneally, intramuscularly, intrapulmonaryly, vaginally, rectally, or into the eye. In accordance with some embodiments, the administration is an oral administration. In accordance with some embodiments, the administration is intravenous. In accordance with some variants of implementation of the introduction is an intramuscular injection.

Standard pharmaceutical formulation techniques are used, such as those disclosed in Remington's The Science and Practice of Pharmacy, 21st Ed., Lippincott Williams & Wilkins (2005), incorporated by reference for such disclosures.

In accordance with some embodiments, the pharmaceutical composition contains a pharmaceutically acceptable adjuvant. The term pharmaceutically acceptable adjuvant as used herein means one or more compatible solid or encapsulating substances that are suitable for administration to a mammal. The term compatible in the context of this document means that the components of the composition are able to mix with this compound and with each other in such a way that there is no interaction that would significantly reduce the pharmaceutical effectiveness of the composition in situations of normal use. In some embodiments, the pharmaceutically acceptable adjuvant is of sufficiently high purity and low enough toxicity to make it suitable for administration, preferably to an animal, preferably a mammal, being treated.

Some examples of substances that can serve as pharmaceutically acceptable adjuvants include amino acids such as alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine , proline, serine, threonine, tryptophan, tyrosine and valine. In some embodiments, the amino acid is arginine. According to some embodiments, the amino acid is L-arginine;

monosaccharides such as glucose (dextrose), arabinose, mannitol, fructose (levulose) and galactose;

derivatives of monosaccharides, such as a monosaccharide in which one or more -OH groups have been replaced. In accordance with some embodiments of the monosaccharide derivative, one or more -OH groups on the monosaccharide have been replaced with one or more -NH2 or -NH-CH3 groups. In some embodiments, the monosaccharide derivative is meglumine. Other examples of the monosaccharide derivative include amino alcohols;

cellulose and its derivatives such as sodium carboxymethylcellulose, ethylcellulose and methylcellulose;

solid lubricants such as talc, stearic acid and magnesium stearate;

polyols such as propylene glycol, glycerin, sorbitol, mannitol and polyethylene glycol;

emulsifiers such as polysorbates;

wetting agents such as sodium lauryl sulfate;

diluents such as calcium carbonate, sodium carbonate, mannitol and lactose;

binders such as starches (corn starch and potato starch), gelatin and sucrose;

disintegrants such as starch, alginic acid and croscarmellose;

glidants such as silica;

coloring agents, such as dyes for the chemical and food industries (FD&C);

sweeteners and flavors such as aspartame, saccharin, menthol, peppermint and fruit flavors;

preservatives such as benzalkonium chloride, PHMB, chlorobutanol, thimerosal, phenylmercuric acetate, phenylmercuric nitrate, parabens, and sodium benzoate;

tonicity regulators such as sodium chloride, potassium chloride, mannitol and glycerin;

antioxidants such as sodium bisulfite, a compound of acetone and sodium bisulfite, sodium formaldehyde, sulfoxylate, thiourea and EDTA;

a pH adjuster such as NaOH, sodium carbonate, sodium acetate, HCl and citric acid;

cryoprotectants such as sodium or potassium phosphates, citric acid, tartaric acid, gelatin and carbohydrates such as dextrose, mannitol and dextran.

In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical composition is formulated as a uniform liquid suitable for injection. In accordance with some embodiments, the pharmaceutical composition is formulated for intravenous/intramuscular administration. In accordance with some embodiments of the pharmaceutical composition, the composition further comprises a pharmaceutically acceptable adjuvant. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is an amino acid, a monosaccharide, or a monosaccharide derivative. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is an amino acid or a monosaccharide derivative. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is an amino acid. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is arginine. In accordance with some embodiments of the pharmaceutical composition, the pharmaceutical

- 11 042812 a highly acceptable adjuvant is L-arginine. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is a monosaccharide or a monosaccharide derivative. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is a monosaccharide derivative. According to some embodiments of the pharmaceutical compositions, the pharmaceutically acceptable adjuvant is meglumine.

liquid composition.

In accordance with some embodiments, the pharmaceutical composition for intravenous/intramuscular administration is formulated as a homogeneous liquid suitable for injection. In accordance with some embodiments, the pharmaceutical composition further comprises an aqueous carrier. In some embodiments, the aqueous vehicle is water for injection, 0.9% sodium chloride injection, 5% dextrose injection, 10% dextrose injection, sodium lactate injection, 5% dextrose injection, and 0.9 % sodium chloride, lactated Ringer's injection with 5% dextrose, sodium chloride/sodium acetate/sodium gluconate/potassium chloride/magnesium chloride injection, sodium chloride/potassium acetate/magnesium acetate injection in 5% dextrose, or any combination thereof. In accordance with some embodiments, the aqueous carrier is water for injection. According to some embodiments, the aqueous vehicle is 0.9% sodium chloride injectable solution.

pH of the liquid composition.

In accordance with some embodiments, the pH of the homogeneous liquid is from about 4 to about 9. In accordance with some embodiments, the pH of the homogeneous liquid is about 4, about 4.4, about 5, about 5.5, about 6, about 6.5 , about 7, about 7.5, about 8, about 8.5, or about 9. In some embodiments, the pH of the homogeneous liquid is from about 4 to about 6.

Recovery powder.

In accordance with some embodiments, the pharmaceutical composition for intravenous/intramuscular administration is formulated as a powder for reconstitution. In accordance with some embodiments, the pharmaceutical composition is suitable for injection after reconstitution with an aqueous vehicle.

In some embodiments, the aqueous vehicle is water for injection, 0.9% sodium chloride injection, 5% dextrose injection, 10% dextrose injection, sodium lactate injection, 5% dextrose injection, and 0.9 % sodium chloride, lactated Ringer's injection with 5% dextrose, sodium chloride/sodium acetate/sodium gluconate/potassium chloride/magnesium chloride injection, sodium chloride/potassium acetate/magnesium acetate injection in 5% dextrose, or any combination thereof.

The stability of the recovery powder.

The pharmaceutical compositions described herein are stable under various storage conditions, including refrigerated, ambient, and accelerated degradation conditions. In some embodiments, the chilled state is about 2°C, about 2.1°C, about 2.2°C, about 2.3°C, about 2.4°C, about 2.5°C , about 2.6°C, about 2.7°C, about 2.8°C, about 2.9°C, about 3°C, about 3.1°C, about 3.2°C, about 3 .3°C, about 3.4°C, about 3.5°C, about 3.6°C, about 3.7°C, about 3.8°C, about 3.9°C, about 4° C, about 4.1°C, about 4.2°C, about 4.3°C, about 4.4°C, about 4.5°C, about 4.6°C, about 4.7°C , near

4.8°C, about 4.9°C, about 5°C, about 5.1°C, about 5.2°C, about 5.3°C, about 5.4°C, about 5.5 °С, about

5.6°C, about 5.7°C, about 5.8°C, about 5.9°C, about 6°C, about 6.1°C, about 6.2°C, about 6.3 °С, about

6.4°C, about 6.5°C, about 6.6°C, about 6.7°C, about 6.8°C, about 6.9°C, about 7°C, about 7.1 °С, about

7.2°C, about 7.3°C, about 7.4°C, about 7.5°C, about 7.6°C, about 7.7°C, about 7.8°C, about 7 .9°C or about 8°C. Accelerated degradation conditions include temperatures and/or relative humidity (RH) that are about equal to or above ambient levels (eg, 25 ^° C.±2 ^° C./RH 60%±RH 5%). In some cases, accelerated degradation conditions are about 30°C, about 35°C, about 40°C, about 45°C, about 50°C, about 55°C, or about 60°C. In other cases, accelerated degradation conditions are relative humidity above 55%, about 65%, about 70%, about 75%, or about 80%. In additional cases, accelerated degradation conditions are about 40° C. or 60° C. at ambient humidity. In other additional cases, accelerated degradation conditions are about 40±2°C/RH 75%±RH 5%. In other additional cases, accelerated degradation conditions are about 30±2°C/RH 65%±RH 5%. In accordance with some variants of implementation of the pharmaceutical composition is stable at about 5±3°C for at least 12 months. According to

- 12 042812 In some embodiments, the pharmaceutical composition is stable at about 25±2°C/RH 60%±RH 5% for at least 12 months. In accordance with some embodiments, the pharmaceutical composition is stable at about 30±2°C/RH 65%±RH 5% for at least 6 months. In accordance with some embodiments, the pharmaceutical composition is stable at about 40±2°C/RH 75%±RH 5% for at least 6 months.

Embodiments

Embodiment P'1. Pharmaceutical composition containing:

(i) (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine- 8-carboxylic acid

a pharmaceutically acceptable salt and/or solvate thereof; and (ii) cefepime.

Embodiment R'2. The pharmaceutical composition according to embodiment P'1, wherein the pharmaceutical composition is formulated as a uniform liquid suitable for injection.

Embodiment P'3. A pharmaceutical composition according to embodiment P'1 or P'2 further comprising a pharmaceutically acceptable adjuvant.

Embodiment P'4. The pharmaceutical composition according to embodiment P'3, wherein the pharmaceutically acceptable adjuvant is selected from amino acids.

Embodiment R'5. A pharmaceutical composition according to embodiment P'4, wherein the amino acid is L-arginine.

Embodiment R'6. The pharmaceutical composition according to any of the embodiments P'1-P'5 further comprising an aqueous carrier.

Embodiment P'7. The pharmaceutical composition according to embodiment P'6 wherein the aqueous carrier is selected from water for injection, 0.9% sodium chloride injection, 5% dextrose injection, 10% dextrose injection, sodium lactate injection, c5 % dextrose and 0.9% sodium chloride, Ringer's injection solution with lactate and 5% dextrose, sodium chloride/sodium acetate/sodium gluconate/potassium chloride/magnesium chloride injection, sodium chloride/potassium acetate/magnesium acetate injection 5% dextrose; and any combinations thereof.

Embodiment P'8. A pharmaceutical composition according to any one of the P'2-P'7 embodiments having a pH of about 4 to about 9.

Embodiment R'9. A pharmaceutical composition according to any of the embodiments of P'2-P'8 having a pH of about 4 to about 6.

Embodiment R'10. The pharmaceutical composition according to embodiment P'1, wherein the pharmaceutical composition is formulated as a powder for reconstitution.

Embodiment R'11. A pharmaceutical composition according to embodiment P'10 further comprising a pharmaceutically acceptable adjuvant.

Embodiment P'12. The pharmaceutical composition according to embodiment P'11, wherein the pharmaceutically acceptable adjuvant is selected from amino acids.

Embodiment P'13. A pharmaceutical composition according to embodiment P'12, wherein the amino acid is L-arginine.

Embodiment P'14. The pharmaceutical composition according to any of the embodiments P'10-P'13, wherein the pharmaceutical composition is suitable for injection after reconstitution with an aqueous vehicle.

Embodiment R'15. A pharmaceutical composition according to embodiment P'14, wherein the aqueous carrier is selected from water for injection, 0.9% sodium chloride injection, 5% dextrose injection, 10% dextrose injection, sodium lactate injection, c5 % dextrose and 0.9% sodium chloride, Ringer's injection solution with lactate and 5% dextrose, sodium chloride/sodium acetate/sodium gluconate/potassium chloride/magnesium chloride injection, sodium chloride/potassium acetate/magnesium acetate injection 5% dextrose; and any combinations thereof.

Embodiment P'16. The pharmaceutical composition according to any of the embodiments P'10-P'15, wherein the pharmaceutical composition is stable at about 5±3°C for at least 12 months.

Embodiment R'17. A pharmaceutical composition according to any of the embodiments of P'10-P'15, wherein the pharmaceutical composition is stable at about 25±2°C/RH 60%±RH 5% for at least 12 months.

- 13 042812

Embodiment R'18. A pharmaceutical composition according to any of the embodiments P'10-P'15, wherein the pharmaceutical composition is stable at about 30±2°C/RH

65%±RH 5% for at least 6 months.

Embodiment R'19. A pharmaceutical composition according to any of the embodiments of P'10-P'15, wherein the pharmaceutical composition is stable at about 40±2°C/RH 75%±RH 5% for at least 6 months.

Embodiment P'20. A method of administering to a subject in need thereof a pharmaceutical composition comprising:

(i) ^)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine-8 -carboxylic acid, its pharmaceutically acceptable salt and/or solvate; and (ii) cefepime.

Embodiment R'21. The method of Embodiment P'20 wherein (R)-3-(2(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][ 1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt and/or solvate thereof, and cefepime are formulated in separate containers.

Embodiment R'22. A method according to an embodiment of P'20 or P'21 wherein (R)3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo [e][ 1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt and/or solvate thereof are constituted in the first container.

Embodiment R'23. The method according to any of the embodiments P'20-P'22, wherein (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro- The 2H-benzo[e][1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt and/or solvate thereof is present in the form of a first reconstitution powder.

Embodiment R'24. The method according to any of the embodiments P'20-P'23, wherein the cefepime is formulated in a second container.

Embodiment P'25. The method according to any of the embodiments of P'20-P'24, wherein the cefepime is present in the form of a second reconstitution powder.

Embodiment R'26. The method according to embodiment P'23 or P'25, wherein the method comprises:

(a) mixing the first reconstitution powder and the second reconstitution powder to form a third reconstitution powder;

(b) reconstituting the third reconstitution powder from step (a) with an aqueous carrier to obtain a homogeneous liquid; and (c) administering the homogeneous liquid from step (b) to a subject in need thereof.

Embodiment R'27. A method according to an embodiment P'23 or P'25, wherein the method comprises:

(a) recovering the first powder for recovery with a first aqueous vehicle to obtain a first homogeneous liquid; and (b) recovering the second powder for recovery with a second aqueous vehicle to obtain a second homogeneous liquid; and (c) mixing the first homogeneous liquid from step (a) and the second homogeneous liquid from step (b) to obtain a third homogeneous liquid; and (d) administering the third homogeneous liquid from step (c) to a subject in need thereof.

Embodiment P'28: The method according to embodiment P'26 or P'27, wherein each of the aqueous vehicles is independently selected from water for injection, 0.9% sodium chloride for injection, 5% dextrose for injection, 10% dextrose injection, sodium lactate injection, 5% dextrose and 0.9% sodium chloride injection, Ringer's solution with lactate and 5% dextrose injection, sodium chloride/sodium acetate/sodium gluconate/potassium chloride/magnesium chloride injection, chloride sodium/potassium acetate/magnesium acetate in 5% dextrose for injection and any combination thereof.

Embodiment R'29. The method of Embodiment P'20 wherein (R)-3-(2(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][ 1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt and/or solvate thereof, and cefepime are formulated in one container.

Embodiment P'30. The method of Embodiment P'29 wherein (R)-3-(2(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][ 1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt and/or solvate thereof, and cefepime are present in the form of a powder for reconstitution.

Embodiment R'31. The method of embodiment P'30, wherein the method comprises:

(a) reconstituting the powder to be reconstituted with an aqueous vehicle to form a homogeneous liquid, and

- 14 042812 (b) administering the homogeneous liquid from step (a) to a subject in need thereof.

Embodiment R'32. The method according to embodiment P'31, wherein the aqueous vehicle is selected from water for injection, 0.9% sodium chloride injection, 5% dextrose injection, 10% dextrose injection, sodium lactate injection, 5% dextrose and 0.9% sodium chloride, Ringer's injection with lactate and 5% dextrose, sodium chloride/sodium acetate/sodium gluconate/potassium chloride/magnesium chloride injection, sodium chloride/potassium acetate/magnesium acetate injection in 5% dextrose and any combination of them.

Embodiment R'33. The method according to any one of embodiments P'20-P'32, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable adjuvant.

Embodiment R'34. The method of embodiment P'33, wherein the pharmaceutically acceptable adjuvant is selected from amino acids.

Embodiment R'35. The method of embodiment P'34 wherein the pharmaceutically acceptable adjuvant is L-arginine.

Embodiment R'36. A method for treating a bacterial infection in a subject in need thereof, comprising administering to the subject a pharmaceutical composition according to any one of embodiments P'1-P'19.

Embodiment R'37. A method for treating a bacterial infection in a subject in need thereof, comprising administering to the subject a pharmaceutical composition in accordance with the method of any one of embodiments P'20-P'35.

Embodiment R'38. The method according to any of the embodiments P'20-P'37, wherein the pharmaceutical composition contains about 500 mg of (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy- 3,4-dihydro-2H-benzo[e][1,2]oxaborinine-8-carboxylic acid.

Embodiment R'39. The method according to any of the embodiments P'20-P'37, wherein the pharmaceutical composition contains about 750 mg of (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy- 3,4-dihydro-2H-benzo[e][1,2]oxaborinine-8-carboxylic acid.

Embodiment R'40. The method according to any of the embodiments of P'20-P'39, wherein the pharmaceutical composition contains about 2 g of cefepime.

Embodiment R'41. The method according to any of the embodiments P'20-P'10, wherein the pharmaceutical composition is administered via intravenous (IV) infusion.

Embodiment P'42. The method of embodiment P'41 wherein the infusion period of the pharmaceutical composition is about 2 hours.

Route of administration for intravenous/intramuscular administration

According to some embodiments, the pharmaceutical composition is administered via intravenous/intramuscular injection. In accordance with some embodiments, the pharmaceutical composition is administered by infusion to a patient over a period of time. According to various embodiments, the infusion time (infusion period) ranges from 5 minutes to continuous infusion, 10 minutes to 8 hours, 30 minutes to 4 hours, and 1 to 3 hours. In one embodiment, the pharmaceutical composition administered by infusion over a period of 30 minutes. In one embodiment, the pharmaceutical composition is administered by infusion over a period of 1 hour. In one embodiment, the pharmaceutical composition is administered by infusion over a period of 1.5 hours. In one embodiment, the pharmaceutical composition is administered by infusion over a period of 2 h. In accordance with one embodiment, the pharmaceutical composition is administered by infusion over a period of 3 hours. In accordance with one embodiment, the pharmaceutical composition is administered by infusion over a period of 4 hours. In accordance with some embodiments, the infusion is repeated with the desired interval between doses, which includes, for example, 6 hours, 8 hours, 12 hours or 24 hours.

In accordance with some embodiments, the pharmaceutical composition is administered over a period of one day. In accordance with some embodiments, the pharmaceutical composition is administered over a period of two days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of three days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of four days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of five days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of six days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of seven days. In accordance with some embodiments, the pharmaceutical composition is administered over a period

- 15 042812 which is eight days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of nine days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of ten days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of 11 days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of 12 days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of 13 days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of 14 days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of 28 days. In accordance with some embodiments, the pharmaceutical composition is administered over a period of 29 days or longer. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of one day. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of two days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of three days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of four days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of five days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of six days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of seven days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of eight days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of nine days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of ten days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of 11 days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of 12 days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of 13 days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of 14 days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of 28 days. In some embodiments, the infusion is repeated at the desired dosing interval, which includes, for example, 6 hours, 8 hours, 12 hours, or 24 hours, over a period of 29 days or longer. According to certain embodiments, only one infusion is given on the last day of the treatment period. In some embodiments, the infusion is repeated every 8 hours for a period of one day. In some embodiments, the infusion is repeated every 8 hours over a period of two days. In some embodiments, the infusion is repeated every 8 hours for a period of three days. In some embodiments, the infusion is repeated every 8 hours over a period of four days. In some embodiments, the infusion is repeated every 8 hours over a period of five days. In some embodiments, the infusion is repeated every 8 hours for a period of six days. In some embodiments, the infusion is repeated every 8 hours for a period of seven days. In some embodiments, the infusion is repeated every 8 hours for a period of eight days. In some embodiments, the infusion is repeated every 8 hours for a period of nine days. In some embodiments, the infusion is repeated every 8 hours for a period of ten days. According to some embodiments,

- 16 042812 fusion is repeated every 8 hours for a period of 11 days. In some embodiments, the infusion is repeated every 8 hours over a period of 12 days. In some embodiments, the infusion is repeated every 8 hours for a period of 13 days. In some embodiments, the infusion is repeated every 8 hours over a period of 14 days. In some embodiments, the infusion is repeated every 8 hours for a period of 28 days. In some embodiments, the infusion is repeated every 8 hours for a period of 29 days or longer. According to some embodiments, only one infusion is performed on the last day of the administration period. In some embodiments, the infusion is repeated every 8 hours over a period of 10 days, with three doses on days 1-9 and one dose on day 10.

In some embodiments, the infusion is repeated every 12 hours for a period of one day. In some embodiments, the infusion is repeated every 12 hours over a period of two days. In some embodiments, the infusion is repeated every 12 hours over a period of three days. In some embodiments, the infusion is repeated every 12 hours over a period of four days. In some embodiments, the infusion is repeated every 12 hours over a period of five days. In some embodiments, the infusion is repeated every 12 hours for a period of six days. In some embodiments, the infusion is repeated every 12 hours for a period of seven days. In some embodiments, the infusion is repeated every 12 hours for a period of eight days. In some embodiments, the infusion is repeated every 12 hours for a period of nine days. In some embodiments, the infusion is repeated every 12 hours for a period of ten days. In some embodiments, the infusion is repeated every 12 hours over a period of 11 days. In some embodiments, the infusion is repeated every 12 hours over a period of 12 days. In some embodiments, the infusion is repeated every 12 hours for a period of 13 days. In some embodiments, the infusion is repeated every 12 hours over a period of 14 days. In some embodiments, the infusion is repeated every 12 hours over a period of 28 days. In some embodiments, the infusion is repeated every 12 hours for a period of 29 days or longer. According to some embodiments, only one infusion is performed on the last day of the administration period.

In some embodiments, the infusion is repeated every 24 hours for a period of one day. In some embodiments, the infusion is repeated every 24 hours for a period of two days. In some embodiments, the infusion is repeated every 24 hours for a period of three days. In some embodiments, the infusion is repeated every 24 hours for a period of four days. In some embodiments, the infusion is repeated every 24 hours for a period of five days. In some embodiments, the infusion is repeated every 24 hours for a period of six days. In some embodiments, the infusion is repeated every 24 hours for a period of seven days. In some embodiments, the infusion is repeated every 24 hours for a period of eight days. In some embodiments, the infusion is repeated every 24 hours for a period of nine days. In some embodiments, the infusion is repeated every 24 hours for a period of ten days. In some embodiments, the infusion is repeated every 24 hours for a period of 11 days. In some embodiments, the infusion is repeated every 24 hours for a period of 12 days. In some embodiments, the infusion is repeated every 24 hours for a period of 13 days. In some embodiments, the infusion is repeated every 24 hours for a period of 14 days. In some embodiments, the infusion is repeated every 24 hours for a period of 28 days. In some embodiments, the infusion is repeated every 24 hours for a period of 29 days or longer.

In some embodiments, the infusion is repeated every 48 hours for a period of two days. In some embodiments, the infusion is repeated every 48 hours over a period of four days. In some embodiments, the infusion is repeated every 48 hours for a period of six days. In some embodiments, the infusion is repeated every 48 hours

- 17 042812 within a period of eight days. In some embodiments, the infusion is repeated every 48 hours for a period of ten days. In some embodiments, the infusion is repeated every 48 hours for a period of 12 days. In some embodiments, the infusion is repeated every 48 hours for a period of 14 days. In some embodiments, the infusion is repeated every 48 hours for a period of 28 days or longer.

In accordance with some embodiments, the pharmaceutical composition contains from 100 to 5000 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains from 250 to 2000 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, 1000, 1100, 1200, 1300, 1400, 1500 , 1600, 1700, 1800, 1900 or 2000 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 250 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 500 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 750 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 1000 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 1250 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 1500 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 1750 mg of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 2000 mg of cefepime.

In some embodiments, the daily dose of cefepime is 250 mg to 12 g. In some embodiments, the daily dose of cefepime is 250 mg, 500 mg, 750 mg, 1000 mg, 1250 mg, 1500 mg, 1750 mg, 2000 mg, 2250 mg, 2500 mg, 2750 mg, 3000 mg, 3.5 g, 4 g, 4.5 g, 5 g, 5.5 g, 6 g, 6.5 g, 7 g, 7.5 g, 8 g, 8.5 g, 9 g, 9.5 g, or 10 g. In some embodiments, the daily dose of cefepime is 250 mg. In some embodiments, the daily dose of cefepime is 500 mg. In some embodiments, the daily dose of cefepime is 1 g. In some embodiments, the daily dose of cefepime is 2 g. In some embodiments, the daily dose of cefepime is 4 g. In some embodiments, the daily dose of cefepime is 6 d. In some embodiments, the daily dose of cefepime is 8 g. In some embodiments, the daily dose of cefepime is 10 g. In some embodiments, the daily dose of cefepime is 12 g.

In some embodiments, the pharmaceutical composition contains 100 to 2000 mg of Compound 1. In some embodiments, the pharmaceutical composition contains 125 to 1000 mg of Compound 1. In some embodiments, the pharmaceutical composition contains 125, 187.5, 200 , 250, 300, 350, 375, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, or 1000 mg of compound 1. In some embodiments, the pharmaceutical composition contains 100 mg of compound 1. According to some embodiments, the pharmaceutical composition contains 125 mg of Compound 1. According to some embodiments, the pharmaceutical composition contains 150 mg of Compound 1. According to some embodiments, the pharmaceutical composition contains 187.5 mg of Compound 1. According to some embodiments, the pharmaceutical composition contains 187.5 mg of Compound 1. in some embodiments, the pharmaceutical composition contains 200 mg of Compound 1. In some embodiments, the pharmaceutical composition contains 250 mg of Compound 1. In some embodiments, the pharmaceutical composition contains 300 mg of Compound 1. In some embodiments, the pharmaceutical composition contains 350 mg of Compound 1 1. In some embodiments, the pharmaceutical composition contains 375 mg of Compound 1. In some embodiments, the pharmaceutical composition contains 400 mg of Compound 1. In some embodiments, the pharmaceutical composition contains 450 mg of Compound 1. In some embodiments, the pharmaceutical composition contains 450 mg of Compound 1. In some embodiments, the pharmaceutical composition contains 500 mg of compound 1. In some embodiments, the pharmaceutical composition contains 550 mg of compound 1. In some embodiments, the pharmaceutical composition contains 600 mg of compound 1. In some embodiments, the pharmaceutical composition contains 650 mg of compound 1. In some embodiments, the pharmaceutical composition contains 750 mg of compound 1. In some embodiments, the pharmaceutical composition contains 800 mg of compound 1. In some embodiments, the pharmaceutical composition contains 850 mg of compound 1. In some embodiments, the pharmaceutical composition contains 900 mg of compound 1. In some embodiments, the pharmaceutical composition contains 950 mg of compound 1. B

- 18 042812 according to some embodiments, the pharmaceutical composition contains 1000 mg of compound 1.

In accordance with some embodiments, the pharmaceutical composition contains from 100 to 2000 mg of compound 1 and from 0.5 to 5 g of cefepime. In accordance with certain embodiments, the pharmaceutical composition contains from 125 to 1000 mg of compound 1 and from 0.5 to 2.5 g of cefepime. In accordance with some embodiments, the pharmaceutical composition contains 125, 150, 187.5, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, or 1000 mg of Compound 1 and 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950 or 1000 mg of Compound 1. According to certain embodiments, the pharmaceutical composition contains 250 mg compounds 1 and 2 g of cefepime. According to some embodiments, the pharmaceutical composition contains 500 mg of compound 1 and 2 g of cefepime. According to certain embodiments, the pharmaceutical composition contains 750 mg of compound 1 and 2 g of cefepime. According to certain embodiments, the pharmaceutical composition contains 125 mg of compound 1 and 1 g of cefepime. According to some embodiments, the pharmaceutical composition contains 250 mg of compound 1 and 1 g of cefepime. According to certain embodiments, the pharmaceutical composition contains 375 mg of compound 1 and 1 g of cefepime. According to certain embodiments, the pharmaceutical composition contains 62.5 mg of compound 1 and 2 g of cefepime. According to some embodiments, the pharmaceutical composition contains 125 mg of compound 1 and 2 g of cefepime. According to certain embodiments, the pharmaceutical composition contains 187.5 mg of compound 1 and 2 g of cefepime.

In some embodiments, the daily dose of Compound 1 is about 200 mg to 5 g. In some embodiments, the daily dose of Compound 1 is about 200, 250 mg, 500 mg, 750 mg, 1000 mg, 1250 mg, 1500 mg. , 1750 mg, 2000 mg, 2250 mg, 2500 mg, 2750 mg, 3000 mg, 3.5 g, 4 g, 4.5 g, or 5 g. In some embodiments, the daily dose of Compound 1 is 250 mg. In some embodiments, the daily dose of Compound 1 is 500 mg. In some embodiments, the daily dose of Compound 1 is 750 mg. In some embodiments, the daily dose of Compound 1 is 1000 mg. In some embodiments, the daily dose of Compound 1 is 1250 mg. In some embodiments, the daily dose of Compound 1 is 1500 mg. In some embodiments, the daily dose of Compound 1 is 1750 mg. In some embodiments, the daily dose of Compound 1 is 2000 mg. In some embodiments, the daily dose of Compound 1 is 2250 mg. In some embodiments, the daily dose of Compound 1 is 2500 mg. In some embodiments, the daily dose of Compound 1 is 2750 mg. In some embodiments, the daily dose of Compound 1 is 3000 mg. In some embodiments, the daily dose of Compound 1 is 3.5 g. In some embodiments, the daily dose of Compound 1 is 4 g. In some embodiments, the daily dose of Compound 1 is 4.5 g. embodiments, the daily dose of Compound 1 is 5 g.

In accordance with some embodiments, the pharmaceutical composition is stored in a sterile container. In some embodiments, Compound 1, a pharmaceutically acceptable salt, MES or pharmaceutically acceptable salt and MES thereof are co-administered with an antibiotic. In some embodiments, the antibiotic is a beta-lactam antibiotic. In some embodiments, the beta-lactam antibiotic is cefepime.

In some embodiments, compound 1, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof, and cefepime are provided in separate containers. In some embodiments, compound 1, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof, and cefepime are provided in a single container. In accordance with some embodiments, the container is an intravenous solution bag. In accordance with some embodiments, the container is a glass bottle. In accordance with some embodiments, the container is an amber glass bottle.

This document discloses a method of administering to a subject in need thereof a pharmaceutical composition comprising:

(i) (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine- 8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof; and (ii) cefepime.

In accordance with some embodiments of a method for administering a pharmaceutical

- 19 042812 compositions (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2Hbenzo[e][1,2]oxaborinine-8 α-carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate, and cefepime are formulated in separate containers. According to some embodiments of the method of administering the pharmaceutical composition (R)-3(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][ 1,2]oxaborinine 8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof are present in the form of a crystalline form as disclosed herein. According to some embodiments of the method of administering the pharmaceutical composition, the crystalline form is Form 1. According to some embodiments of the method of administering the pharmaceutical composition, (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido) -2hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborinine-8-carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate are in the first container. According to some embodiments of the method of administering the pharmaceutical composition (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e] The [1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt, a solvate or a pharmaceutically acceptable salt and solvate thereof are present in the form of a first reconstitution powder. According to some embodiments of the method of administering the pharmaceutical composition, cefepime is formulated in a second container. In accordance with some embodiments of the method of administering the pharmaceutical composition, cefepime is present in the form of a second reconstitution powder.

According to some embodiments of a method for administering a pharmaceutical composition, the method includes:

(a) mixing the first reconstitution powder and the second reconstitution powder to form a third reconstitution powder;

(b) reconstituting the third reconstitution powder from step (a) with an aqueous carrier to obtain a homogeneous liquid; and (c) administering the homogeneous liquid from step (b) to a subject in need thereof.

According to some embodiments of a method for administering a pharmaceutical composition, the method includes:

(a) recovering the first powder for recovery with a first aqueous vehicle to obtain a first homogeneous liquid; and (b) recovering the second powder for recovery with a second aqueous vehicle to obtain a second homogeneous liquid; and (c) mixing the first homogeneous liquid from step (a) and the second homogeneous liquid from step (b) to obtain a third homogeneous liquid; and (d) administering the third homogeneous liquid from step (c) to a subject in need thereof.

In accordance with some embodiments of the method of administering the pharmaceutical composition, each of the aqueous carrier is independently water for injection, 0.9% sodium chloride injection, 5% dextrose injection, 10% dextrose injection, sodium lactate injection, c 5% Dextrose and 0.9% Sodium Chloride, Ringer's Injection with Lactate and 5% Dextrose, Sodium Chloride/Sodium Acetate/Sodium Gluconate/Potassium Chloride/Magnesium Chloride Injection, Sodium Chloride/Potassium Acetate/Magnesium Acetate Injection at 5 % dextrose or any combination thereof. In accordance with some embodiments of the pharmaceutical composition, the composition further comprises a pharmaceutically acceptable adjuvant. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is an amino acid or a monosaccharide derivative. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is L-arginine. According to some embodiments of the pharmaceutical compositions, the pharmaceutically acceptable adjuvant is meglumine.

This document also discloses a method of administering to a subject in need thereof a pharmaceutical composition comprising:

(i) (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborynine- 8-carboxylic acid, a pharmaceutically acceptable salt or solvate thereof; and (ii) cefepime.

According to some embodiments of the method of administering the pharmaceutical composition (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2Hbenzo[e][1 ,2]oxaborinine-8-carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate, and cefepime are in one container. According to some embodiments of the method of administering the pharmaceutical composition (R)-3-(2(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e][ 1,2]oxaborinin-8

- 20 042812 carboxylic acid, its pharmaceutically acceptable salt, solvate or pharmaceutically acceptable and solvate are present in the form of a crystalline form, which is disclosed in this document. According to some embodiments of the method of administering the pharmaceutical composition, the crystalline form is Form 1. According to some embodiments of the method of administering the pharmaceutical composition, (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido) -2hydroxy-3,4-dihydro-2H-benzo[e][1,2]oxaborinine-8-carboxylic acid, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt thereof, and solvate and cefepime are present in the form of a powder for reconstitution.

According to some embodiments of a method for administering a pharmaceutical composition, the method includes:

(a) reconstituting the reconstituting powder with an aqueous vehicle to form a homogeneous liquid; and (b) administering the homogeneous liquid from step (a) to a subject in need thereof.

In accordance with some embodiments of the method of administering the pharmaceutical composition, the aqueous carrier is water for injection, 0.9% sodium chloride injection, 5% dextrose injection, 10% dextrose injection, sodium lactate injection, 5% dextrose injection. and 0.9% sodium chloride, Lactated Ringer's Injection 5% Dextrose, Sodium Chloride/Sodium Acetate/Sodium Gluconate/Potassium Chloride/Magnesium Chloride Injection, Sodium Chloride/Potassium Acetate/Magnesium Acetate Injection in 5% Dextrose or any combination of them. In accordance with some embodiments of the pharmaceutical composition, the composition further comprises a pharmaceutically acceptable adjuvant. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is an amino acid or a monosaccharide derivative. According to some embodiments of the pharmaceutical composition, the pharmaceutically acceptable adjuvant is L-arginine. According to some embodiments of the pharmaceutical compositions, the pharmaceutically acceptable adjuvant is meglumine.

Methods of treatment

The present disclosure also relates to methods for inhibiting the growth of bacteria, for example, by reducing the resistance of bacteria to a β-lactam antibiotic, such methods include contacting a bacterial cell culture or a bacterial-infected cell culture, tissue, or organism with Compound 1, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate. In some embodiments, the bacteria to be inhibited upon administration of Compound 1, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof are bacteria that are resistant to beta-lactam antibiotics. The term resistant is well understood by those skilled in the art (see, e.g., Payne et al., Antimicrobial Agents and Chemotherapy 38 767-772 (1994), Hanaki et al., Antimicrobial Agents and Chemotherapy 30 1120-1126 (1995) ).

These methods are useful for inhibiting bacterial growth in a number of situations. In certain embodiments, compound 1, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof is administered to an in vitro experimental cell culture to prevent the growth of beta-lactam resistant bacteria. In certain other embodiments, compound 1, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof is administered to a mammal, including a human, to prevent the growth of beta-lactam resistant bacteria in vivo. The method according to this embodiment comprises administering a therapeutically effective amount of a beta-lactamase inhibitor for a therapeutically effective period of time to a mammal, including a human. Preferably, the beta-lactamase inhibitor is administered in the form of a pharmaceutical composition as described above. In some embodiments, the antibiotic is co-administered with a beta-lactamase inhibitor. In some embodiments, the antibiotic is a beta-lactam antibiotic. In some embodiments, the beta-lactam antibiotic is cefepime.

In another aspect, provided herein are methods of treating a bacterial infection, the method comprising administering to a subject a pharmaceutical composition comprising Compound 1, a pharmaceutically acceptable salt, solvate or pharmaceutically acceptable salt and solvate thereof, and a pharmaceutically acceptable adjuvant as described above. In some embodiments, the bacterial infection is an upper or lower respiratory tract infection, a urinary tract infection, an intra-abdominal infection, or a skin infection. In some embodiments, the infection being treated or prevented contains a bacterium that includes

- 21 042812

Elizabethkingia meningoseptica, Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas acidovorans, Pseudomonas alcaligenes, Pseudomonas putida, Stenotrophomonas maltophilia, Burkholderia cepacia, Aeromonas hydrophilia, Escherichia coli, Citrobacter freundii, Salmonella typhimurium, Salmonella typhi, Salmonella paratyphi, Salmonella enteritidis, Shigella dysenleriae, Shigella flexneri , Shigella sonnei, Enlerobacler cloacae, Enlerobacter aerogenes, Klebsiella pneumoniae, Klebsiella oxytoca, Serratia marcescens, Francisella tularensis, Morganella morganii, Proteus mirabilis, Proteus vulgaris, Providencia alcalifaciens, Providenciarettgeri, Providencia stuartii, Acinetobacter baumannii, Acinetobacter calcoaceticus, Acinetobacter haemolyticus, Yersinia enterocolitica , Yersinia pestis, Yersinia pseudotuberculosis, Yersinia intermedia, Bordetella pertussis, Bordetella parapertussis, Bordetella bronchiseptica, Haemophilus influenzae, Haemophilus parainfluenzae, Haemophilus haemolyticus, Haemophilus parahaemolyticus, Haemophilus ducreyi, Pasteurella multocida, Pasteurella haemolytica, Branhamella catarrhalis, Helicobacter pylori, Campylobacter fetus, Campylobacter jejuni, Campylobacter coli, Borrelia burgdorferi, Vibrio cholerae, Vibrio parahaemolyticus, Legionella pneumophila, Listeria monocytogenes, Neisseria gonorrhoeae, Neisseria meningitidis, Kingella, Moraxella, Gardnerella vaginalis, Bacteroides fragilis, Bacteroides distasonis, Bacteroides 3452A homology groups, Bacteroides gavaluoides Bacteroides thetaiotaomicron, Bacteroides uniformis, Bacteroides eggerthii, Bacteroides splanchnicus, Clostridium difficile, Mycobacterium tuberculosis, Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium leprae, Corynebacterium diphtheriae, Corynebacterium ulcerans, Streptococcus pneumoniae, Streptococcus agalactiae, Streptococcus pyogenes, Enterococcus faecalis, Enterococcus faecium, Staphylococcus aureus , Staphylococcus epidermidis, Staphylococcus saprophyticus, Staphylococcus intermedins, Staphylococcus hyicus subsp. hyicus, Staphylococcus haemolyticus, Staphylococcus hominis, or Staphylococcus saccharolyticus.

In some embodiments, the infection being treated or prevented contains a bacterium that includes Elizabethkingia meningoseptica, Pseudomonas aeruginosa, Pseudomonas fluorescens, Stenotrophomonas maltophilia, Escherichia coli, Citrobacter freundii, Salmonella typhimurium, Salmonella typhi, Salmonella paratyphi, Salmonella enteritidis, Shigella dysenteriae, Shigella flexneri, Shigella sonnei, Enterobacter cloacae, Enterobacter aerogenes, Klebsiella pneumoniae, Klebsiella oxytoca, Serratia marcescens, Acinetobacter calcoaceticus, Acinetobacter haemolyticus, Yersinia enterocolitica, Yersinia pestis, Yersinia pseudotuberculosis, Yersinia intermedia, Haemophilus influenzae, Haemophilus parainfluenzae, Haemophilus haemolyticus, Haemophilus parahaemolyticus, Helicobacter pylori, Campylobacter fetus, Campylobacter jejuni, Campylobacter coli, Vibrio cholerae, Vibrio parahaemolyticus, Legionella pneumophila, Listeria monocytogenes, Neisseria gonorrhoeae, Neisseria meningitidis, Moraxella, Bacteroides fragilis, Bacteroides vulgatus, Bacteroides ovalus, Bacteroides thetaiotaomicron, Bacteroides uniformis, Bacteroides eggerthii or Bacteroides splanchnicus.

In one aspect, this document provides a method of treating bacterial infections by administering Compound 1, a pharmaceutically acceptable salt, MES or pharmaceutically acceptable salt and MES thereof, in combination with a beta-lactam antibiotic. In some embodiments, the beta-lactam antibiotic is cefepime. In certain embodiments, the bacterial infection is a complicated intra-abdominal infection, a complicated urinary tract infection (including pyelonephritis), pneumonia, an uncomplicated urinary tract infection, or an uncomplicated infection of the skin and skin structures. In accordance with certain embodiments, the bacterial infection is a complicated intra-abdominal infection or a complicated urinary tract infection (including pyelonephritis). In accordance with certain embodiments, the bacterial infection is a complicated intra-abdominal infection. In certain embodiments, the bacterial infection is a complicated urinary tract infection (including pyelonephritis).

In another aspect, this document provides a method of treating febrile neutropenic patients by administering Compound 1, a pharmaceutically acceptable salt, MES or pharmaceutically acceptable salt and MES in combination with a beta-lactam antibiotic. In some embodiments, the beta-lactam antibiotic is cefepime.

Some definitions.

Unless otherwise indicated, all technical and scientific terms used in this document have the same meanings that are commonly understood by a person skilled in the art. While any methods and materials similar or equivalent to those described herein may be used in the practice or testing of the embodiments described herein, certain methods, devices, and materials will now be described. In the context of this document and the appended claims, the singular forms include the corresponding plural forms, unless the context clearly dictates otherwise. Thus, for example, a reference to an adjuvant is a reference to one or more adjuvants and their equivalents known to those skilled in the art, and so on.

- 22 042812

The term about is used to indicate that a value includes the standard error for the device or method used to determine the value.

The use of the term or in claims generally means and/or, unless it is explicitly stated that it refers only to alternatives, or the alternatives are mutually exclusive, although the present disclosure justifies a definition that refers only to alternatives and to and/or. The terms contain, have, and include are open linking verbs. Any forms or tenses of one or more of these verbs, such as contains, containing, has, having, includes, and includes are also open. For example, any method that provides for, has or includes one or more steps is not limited to only that one or more steps, but also extends to other steps not included in the list.

Optional or optional may mean that the structure, phenomenon, or circumstance described below may or may not occur, and that the description includes cases where events occur and cases where they do not occur.

In the context of this document, the term therapeutic means an agent used to treat, control, alleviate, prevent or improve an undesirable condition or disease in a patient.

Administration, when used in conjunction with a therapeutic agent, means administering the therapeutic agent systemically or locally, such as directly into the target tissue or application to the target tissue, or administering the therapeutic agent to a patient, wherein the therapeutic agent has a beneficial effect on the target tissue. The administration of the pharmaceutical composition may be by injection, topical administration and oral administration, or by other means alone or in combination with other known techniques.

The term animal as used herein includes, without limitation, humans and non-human vertebrates such as wild, domestic and farm animals. In the context of this document, the terms patient, subject, and individual are intended to include living organisms that may experience certain conditions as described herein. Examples include humans, monkeys, cows, sheep, goats, dogs, cats, mice, rats, and their transgenic varieties. In accordance with a preferred embodiment, the patient is a primate. In accordance with certain embodiments, the primate or subject is a human. In certain cases, the person is an adult. In certain cases, a person is a child. In additional cases, the person is 12 years of age or younger. In certain cases, the person is elderly. In other cases, the person is 60 years of age or older. Other examples of subjects include experimental animals such as mice, rats, dogs, cats, goats, sheep, pigs and cows.

By pharmaceutically acceptable is meant that the carrier, diluent or adjuvant must be compatible with the other ingredients of the composition and not harmful to its recipient.

The term "pharmaceutical composition" means a composition containing at least one active ingredient, such as Compound 1, the composition being studied for a specific effective result in a mammal (eg, without limitation, a human). Those skilled in the art will understand and appreciate the appropriate techniques for determining whether an active ingredient has the desired effective result, based on the needs of the skilled person.

A therapeutically effective amount or effective amount as used herein refers to the amount of an active compound or pharmaceutical agent that elicits in a tissue, system, animal, individual, or human a biological or therapeutic response that is being sought by the researcher, veterinarian, physician, or other clinician, and which includes one or more of the following: (1) disease prevention; for example, preventing a disease, condition, or disorder in an individual who may be predisposed to the disease, condition, or disorder but is not yet experiencing or exhibiting the pathology or symptoms of the disease, (2) suppressing the disease; for example, suppression of a disease, condition, or disorder in an individual who is experiencing or exhibiting the pathology or symptoms of the disease, condition, or disorder (ie, delaying further progression of the pathology and/or symptom), and (3) attenuating the disease; for example, amelioration of a disease, condition, or disorder in an individual who is experiencing or exhibiting the pathology or symptoms of the disease, condition, or disorder (ie, reversal of development of the pathology and/or symptoms). The terms treat, receive treatment, treatment or treatment as used herein refer to both therapeutic treatment in accordance with some embodiments and prophylactic or preventive measures in accordance with other embodiments, the purpose being to prevent or slow down (alleviate ) an undesirable physiological condition, disorder or disease or in obtaining favorable or desired clinical results. In the context described herein, beneficial or desirable clinical results include, without limitation, relief of symptoms; reduction in the severity of the condition, disorder or disease; stabilization (ie, no worsening) of the condition, disorder, or disease; delaying the onset or retardation of the condition, disorder, or disease; amelioration of a condition, disorder, or disease state and remission (either partial or complete), either detectable or undetected, or aggravation or amelioration of a condition, disorder, or disease. Treatment includes providing a clinically meaningful response without excessive levels of side effects. Treatment also includes extending life compared to life expectancy in the absence of treatment. The preventive benefits of treatment include preventing the condition, delaying the development of the condition, stabilizing the condition, or reducing the likelihood of the condition occurring. As used herein, treating, receiving treatment, treatment, or providing treatment includes prophylaxis, in accordance with some embodiments. The term essentially the same in the context of this document refers to an X-ray powder diffraction pattern or differential scanning calorimetry scan that is not identical to that depicted herein, but which falls within experimental error when considered by one skilled in the art.

Examples

Methods of analysis.

Powder x-ray diffraction.

XRPD diffraction patterns were obtained using a high throughput XRPD system (HT-XRPD). The plates were mounted on a Bruker GADDS diffractometer equipped with a Hi-Star area detector. The XRPD platform was calibrated using silver behenate for large interatomic distances d and corundum for small interatomic distances d.

Data collection was carried out at room temperature using monochromatic CuKa radiation in the 2θ range from 1.5 to 41.5°, which is the most expressive part of the XRPD diffractogram. A diffraction pattern for each well was obtained in two ranges of 2θ values (1.5°<2θ 21.5° for the first frame and 19.5°<2θ 41.5° for the second) with an exposure time of 90 s for each frame. No background subtraction or curve smoothing was applied to the XRPD patterns. The carrier material used in the XRPD analysis was X-ray transparent and contributed only a minor amount to the background.

High Resolution X-Ray Powder Diffraction (HR-XRPD).

Powder data were obtained on a D8 Advance diffractometer using CuKa1 radiation (1.54016 A) with a germanium monochromator at room temperature. Data was acquired in single 2θ mode (detector scan) at 2θ values from 4° to 45° in 0.016° 2θ steps, 3426.5 s per step on a LynxEye solid state detector. The sample was measured in a glass capillary 8 mm long with an outer diameter of 0.5 mm.

Variable Temperature Powder X-Ray Diffraction (VT-XRPD).

For experiments at variable temperature, an ANSYCO HT camera was used, installed in the system of a D8 Advance diffractometer (Bruker), designed with the Bragg-Brentano geometry and equipped with a LynxEye solid-state detector. The radiation used for data collection was CuKa1 (λ= 1.54056 A), which was monochromatic with a germanium crystal. The material was placed on a fixed sample holder that was mounted inside the chamber. VT-XRPD: The temperature change rate was 10°C/min. Diffraction patterns were obtained in the range of values 2θ 9-24.5 with a step 2θ 0.01569 and a measurement time of 8 s per step. The period of data collection for the temperature value was 2x60 min.

Analysis by differential scanning calorimetry (DSC).

Melting properties were obtained from DSC thermograms recorded using a DSC822e heat flow instrument (Mettler-Toledo GmbH, Switzerland). DSC822e was calibrated for temperature and enthalpy using a small piece of indium (melting point 156.6°C; AHf = 28.45 J-g' ¹ ). Samples were sealed in standard 40 μl aluminum crucibles, pierced and heated in DSC from 25 to 300°C at a heating rate of 10°C min' ¹ . Dry N2 gas with a flow rate of 50 ml min' ¹ was used to purge the DSC equipment during the measurement.

Thermogravimetric analysis combined with mass spectroscopy (TGMS).

Weight loss due to loss of solvent or water from the crystals was determined by TGA/SDTA (TGA: Thermogravimetric Analysis; SDTA: Simultaneous Differential Thermal Analysis). Monitoring the mass of the sample during heating in a TGA/SDTA851e instrument (MettlerToledo GmbH, Switzerland) resulted in a mass versus temperature curve. TGA/SDTA851e was temperature calibrated using indium and aluminum. Samples were weighed into 100 μl aluminum crucibles and sealed. The sealing lids were pierced and the crucibles were heated

- 24 042812 in TGA from 25 to 300 or 400°C (depending on the experiment) with a heating rate of 10°C min ^-1 . Dry N2 gas was used for purge.

Gases released from TGA samples were analyzed using an Omnistar mass spectrometer.

GSD 301 T2 (Pfeiffer Vacuum GmbH, Germany). The latter is a quadrupole mass spectrometer that analyzes masses in the range of 0-200 atomic mass units.

Dynamic Vapor Sorption (DVS) analysis.

Differences in hygroscopicity (moisture absorption) of various forms of solid material provided a measurable indicator of their relative stability with increasing relative humidity. Moisture sorption isotherms for small samples were obtained using the DVS-1 system from Surface Measurement Systems (London, UK); this instrument is suitable for use with as little as a few milligrams of sample with an accuracy of 0.1 µg. The following humidity profile was used: Relative humidity was cycled from 45 to 95% (sorption), back to zero 0% (desorption) and up to 45% at a constant temperature of 25°C. The change steps consisted of 10% RH (45 to 75% Rh), 5% RH (0 to 45% RH), and 20% (75 to 95% RH). Mass equilibrium per step was established using a minimum hold time of 1 hour. At the end of the DVS experiments, recovered solid material was measured by HTXRPD.

Example 1 Preparation of the dihydrochloride salt of compound 1

(2)

To a solution of tert-butyl-3 -((R)-2-(2-(( 1 r,4R)-4-((tert-butoxycarbonyl)(2-((tert-butoxycarbonyl)amino)ethyl)amino)cyclohexyl )acetamido)-2-((3aS,4S,6S,7aR)-3а,5,5-trimethylhexahydro-4,6-methanobenzo[b][1,3,2]dioxaborol-2-yl)ethyl)-2 -methoxybenzoate (structure 2, 110.9 g, 136.7 mmol) in 1,4-dioxane (137 ml) hydrochloric acid (540 ml, 3 M in water) was added. The resulting mixture was heated to reflux and stirred at this temperature for 100 minutes. The resulting mixture was cooled to room temperature over 30 minutes, then extracted with methyl tert-butyl ether (3x200 ml). The aqueous phase was concentrated in vacuo (at 57-29 mmHg and external bath temperature 20-72°C). The residue was diluted with water to a total weight of 130 g. Isopropanol (90 ml) was added to this solution over about 10 minutes. This solution was seeded with crystals of compound 1-(HCl) ₂ (100 mg). To this mixture was added isopropanol (100 ml) over approximately 5 minutes. The resulting slurry was stirred for approximately 15 h, then filtered under vacuum, the solid was washed with isopropanol (2x200 ml) and then with methyl tert-butyl ether (400 ml). The solid was further dried under high vacuum to give the title compound (structure 1) (43.9 g) as a white crystalline solid, a mixture of Forms 1 and 2.

Example 2. Crystal forms of the dihydrochloride of compound 1.

Example 2A. Solubility of compound 1 dihydrochloride in IPA/water mixtures In order to design a recrystallization process for compound 1 dihydrochloride salt from water as solvent and IPA as antisolvent, the solubility of API in these combinations was determined. Suspensions in various water/IPA mixtures were prepared and incubated for 24 h at 5, 25, 40 and 50°C. At the end of the incubation time, the solids were separated from the mother liquors by centrifugation and filtration, and the concentration of Compound 1 in solution was determined by HPLC analysis. The remaining solids were analyzed using HT-XRPD. The results of determining the solubility depending on the content of the antisolvent and temperature are presented in Table. 6 and in FIG. 1A and 1B.

Table 6

Solubility values of compound 1 dihydrochloride in mg/ml in IPA/water mixtures at four temperatures, 5, 25, 40 and 50°C

Solvent mixture Temperature 5°C 25°C 40°C 50°C IPA/H2O (40:60) 270.7 429.4 566.7 713.8 IPA/H2O (50:50) 217.3 335.9 453.9 528.4 IPA/H2O (60:40) 128.9 205.4 424.2 439.5 IPA/H2O (70:30) 78.6 100.9 189.2 235.0 IPA/H2O (80:20) 28.0 41.7 81.0 102.5 IPA/H2O (90:10) 4.2 4.2 5.6 7.2

Example 2B. Preparation of crystalline dihydrochloride compound 1.

To the dihydrochloride of compound 1 from Example 1 (675 g, Sample A) was added 1.85 L of isopropanol/water (40/60 v/v). The mixture was stirred at 19-26° C. until a clear solution was obtained. The solution was filtered through a membrane filter (5 μm). Isopropanol (550 ml) was added to the filtrate followed by seed crystals (6.75 g). To this mixture was added isopropanol (8.7 L) over 1 hour. The resulting mixture was stirred for 63 hours then filtered under vacuum. The liquid from the wet cake was removed in vacuo for 1 h 20 min. The solid was washed with isopropanol (700 ml) then methyl tert-butyl ether (700 ml). The solid was dried in ambient air for 2 hours 45 minutes. 612 g of compound 1 dihydrochloride monohydrate were isolated (sample B). Chloride according to the results of ion chromatography 14.8% wt./wt. Water content according to the results of the analysis by the method of Karl Fischer = 3.8% wt./wt.

Example 2C. Impurity profile in crystalline dihydrochloride monohydrate compound 1.

In table. 7 shows the impurity profile in Compound 1 dihydrochloride (sample A) and Compound 1 crystalline dihydrochloride (sample B) as measured by the HPLC method described in Table 1. 8.

Table 7

SAMPLE Comm. 1 RRT_0.86 RRT_l,06 RRT_1.15 RRT_1,21 RRT_1.26 Sample A 97.5 0.4 0.7 0.18 0.7 0.55 Sample B 99.7 0.16 below LOQ ο,ι below LOQ below LOQ

Table 8

Column details Zorbax SB-C18 Column length 15 cm Column inner diameter 4.6mm Particle size in column 5 µm Mobile phase A 0.05% v/v TFA in water Mobile phase B 0.05% v/v TFA in acetonitrile Gradient Time %A %IN 0 98 2 15 2 98 20 2 98 20.01 98 2 25 98 2 Thinner UHP Water Type 1 Flow 1.0 ml/min. Column temperature 40°C Wavelength 210 nm Injection volume 5 µl Sample concentration 0.3 mg/ml Total run time 25 min. Typical hold time compound 1: 4.9 min. Calculations Relative retention time for peak X (RRT X): Retention time for peak X/Retention time for compound 1 Purity (% area): (Peak area for compound 1)/(Σ peak area)* 100% Impurities (% area): Impurity peak area/(U peak area)* 100%, where The area of the peaks is the arithmetic sum of the areas of all the peaks

Abbreviations: RRT = relative retention time; TFA = trifluoroacetic acid

2D example. Characterization using XRPD.

Compound 1 crystalline dihydrochloride (from Example 2B) was analyzed by HRXRPD (FIG. 2). From the single crystal data available for the monohydrate (FIG. 3), it was concluded that the crystalline form was the dihydrochloride salt monohydrate of Compound 1. This was named Form 1. in Table 1. Table 1 shows the positions of the peaks and their intensities for form 1.

Example 2E. TG/MS and DSC analysis.

Analysis of compound 1 dihydrochloride monohydrate by TG/MS (FIG. 4) showed that 3.4% weight loss was recorded between 25 and 80°C. This mass is due to the loss of 1 molecule of water per molecule of compound 1 dihydrochloride. This result confirms the XRPD analysis that the material consisted of the monohydrate form. The compound decomposed at temperatures above 280°C.

- 26 042812

In FIG. 5 shows the DSC curve for form 1 in the case of a programmed temperature increase from 0 to 400°C. The DSC curve shows an endothermic phenomenon with an onset temperature of approximately 50°C. This phenomenon corresponds to the onset of dehydration of the monohydrate. This dehydration process is followed by decomposition of the compound at approximately 290° C., as signaled by a major endothermic event. From this experiment, it is concluded that Form 1 is stable up to elevated temperatures, up to about 50°C.

Example 2F. Analysis of dynamic vapor sorption.

Compound 1 dihydrochloride monohydrate was analyzed by dynamic vapor sorption (FIG. 6). The material was analyzed to obtain an isothermal desorption/sorption profile starting at 45-95-0-45% relative humidity levels (5% increments at low relative humidity (RH)) at 25°C. The temperature was kept constant at approximately 25°C during the period of the experiment. The curve marked with diamonds shows the change in mass of the Form 1 sample when exposed to an increasing level of relative humidity (RH from 45 to 95%). It can be seen that the mass uptake is relatively small and that at high humidity the dihydrochloride of Compound 1 remained as Form 1. This was confirmed by the mass stability and the VH-XRPD experiment performed (see FIG. 7).

The curve marked with squares shows the change in weight of the Form 1 sample when exposed to a series of relative humidity values from RH 95% to RH 0%. It can be seen that the mass of the sample does not change significantly from approximately RH 55% to RH 15%. Upon subsequent exposure to RH values of 10, 5 and 0%, the sample loses approximately 4% of its mass, which corresponds to the loss of one molecule of water per dihydrochloride salt of compound 1. The VH-XRPD experiment confirmed that the loss of water was accompanied by conversion to an anhydrous form (Form 2) (see Fig. 7). The curve marked with triangles shows the change in mass of form 2 caused by dehydration of crystalline form 1 when exposed to increasing relative humidity (from RH 0% to RH 50%). There is a slight increase in weight up to RH 25% followed by a sharp increase in weight when Form 2 is exposed to RH 30%. The VH-XRPD experiment showed that Form 2 converted to Form 1 at %RH values in the RH range of 25-35% (see FIG. 7). These observations demonstrate that Form 2 undergoes significant changes in moisture content and physical form at relative humidity levels that are typical of those found in the pharmaceutical industry.

Example 2G. XRPD analysis at variable humidity.

Variable Humidity XRPD Analysis (VH-XRPD) was performed to confirm the hypothesis of solid form change at relative humidity levels between 40 and 0%. The sample was exposed to a humidity profile representing 45-75-2015-10-30-35-45% relative humidity levels at 30°C. Two diffractograms were recorded at each stage. The first diffraction pattern was recorded when the target humidity was reached, and the second after the sample had equilibrated for 1 hour at each humidity level. When a phase transition occurred, and if the conversion rate was higher than the scan time, the diffractogram showed peaks for the original form at the beginning (lower 2θ angles) and peaks for the converted form at the end of the diffraction pattern (higher 2θ angles). The most intense peaks for form 1 (dihydrochloride monohydrate) are in the range of 2θ values from 9.5 to 24.5°.

Stacked diffraction patterns recorded at various humidity levels are shown in FIG. 7. The measured RH level and the type of RH change step that occurs (sorption or desorption) are on the right side. The material shows a clear change in the diffractogram, indicating a phase transition induced by moisture sorption. Upon desorption, the material was converted at a relative humidity of 20 to 15% into Form 2, an anhydrous form. During sorption form 2 was converted back to form 1 at a relative humidity of 30 to 35%. DVS analysis showed significant moisture sorption in the RH range from 40 to 95%. The VH-XRPD results did not provide evidence of additional shape changes at these values.

Example 2H. Characteristics of form 2.

As a result of X-ray powder diffraction experiments at variable humidity, carried out on form 1, a second crystalline form of the dihydrochloride of compound 1 was discovered. This form was called form 2. To further characterize form 2, some material was obtained by drying form 1 in vacuum at 40 ° C for 18 hours. FIG. 8 shows the XRPD diffraction pattern for Form 2. Analysis of a solid sample of Form 2 by TGA demonstrated the anhydrous nature of this solid. The results of the thermogravimetric analysis and the thermogram obtained by DSC analysis showed no thermal phenomena prior to decomposition at 280° C. and no loss of weight on drying (FIG. 9). In table. Figure 2 shows the positions of the peaks and their intensities for form 2.

VH-XRPD and DVS analyzes performed on starting materials indicated that there was a dynamic and reversible relationship between the anhydrous form 2 and the monohydrate form 1.

- 27 042812 The rate of conversion of Form 2 to Form 1 as a function of relative humidity was studied by means of an isothermal liquid sorption experiment at fixed values of relative humidity. Briefly, a Form 2 solid sample was obtained by drying Form 1. This sample was placed in a DVS machine and incubated for 1 hour at the indicated humidity level while recording the change in weight. The results are shown in FIG. 10A and 10V. The conversion rate depended on the level of relative humidity. The highest speed was achieved at 45% RH, while the lowest speed was achieved at 30% RH. The right section shows a linear relationship between the moisture level applied and the conversion rate.

Summary.

Compound 1 dihydrochloride was physically characterized using X-ray powder diffraction, thermal analysis, and dynamic vapor sorption techniques. Loss on drying studies by TG/MS analysis demonstrated that the material consisted of the monohydrate crystalline solid form of the dihydrochloride compound 1. This particular form was termed Form 1. The material exhibited degradation at temperatures above 280°C. Characterization using DVS analysis showed the nature of dehydration/hydration at values of relative humidity from 0 to 40% at 25°C with a clear hysteresis. The presence of hysteresis indicated a phase transition from a monohydrate to an anhydrous crystalline form. This phase transition can be confirmed by an x-ray powder diffraction study under variable conditions which showed the conversion of form 1 (monohydrate) to form 2 (anhydrous).

Physical characterization of compound 1 dihydrochloride confirmed the existence of two crystalline forms. The main form is the monohydrate, called form 1, and the other form is the anhydrous form, form 2. Dynamic vapor sorption studies have shown that under ambient conditions, the dihydrochloride monohydrate form of compound 1 is the most stable form. In this study, the anhydrous form was observed only at low levels of relative humidity or drying at elevated temperature and/or under vacuum. The interconversion between the two solid forms is fast and reversible.

Example 3 Solubility of compound 1 dihydrochloride monohydrate in water as a function of pH.

A known amount of compound 1 was weighed directly into 5 ml vials and 1 ml of each phosphate buffer was added; the resulting solutions were maintained at a controlled temperature (25°C) and stirred with a magnetic stirrer for up to 24 hours. Aliquots of the resulting suspensions or solutions were centrifuged (14000 rpm, 10 min) and taken at two predetermined time points (4 24 h), appropriately diluted and analyzed by HPLC-UV (HPLC with detection in UV) using the parameters of the analysis method shown in table. 9.

Table 9

Description Meaning Column type ZorbaxSB-C18 Column length [cm] 15 Inner diameter [cm] 0.46 Particle size [µm] 5 mobile phase A: 0.05% v/v TFA in water; B: 0.05% v/v TFA in acetonitrile Stage 1: time-reserve. A-reserve B T=0 min. 98%A - 2%B Stage 2: time-reserve A-reserve B T=15 min. 2%A / 98%B up to 20.0 min. Stage 3: time-reserve A-reserve B T=20.01 min. 98%A / 2%B up to 25.0 min. Flow rate [ml/min] 1 Column temperature [°C] 40 Detector type UV Wavelength (nm) 210 Injection volume (µl) 5

Compound 1 dihydrochloride monohydrate solubility data and pH values are given in Table 1. 10.

- 28 042812

Table 10

Medium (phosphate buffer) pH (4 hours) pH (24 hours) Conc, (mg/ml) 4 hours Conc, (mg/ml) 24 hours Visual observation (4 hours) Visual observation (24 hours) pH 4 3.9 3.9 >143 132.8 Medium (phosphate buffer) pH (4 hours) pH (24 hours) Conc, (mg/ml) 4 hours Conc, (mg/ml) 24 hours Visual observation (4 hours) Visual observation (24 hours) pH 5 4.9 4.9 >135 107.0 Clear, clear solution The solution was clear, but a gel-like precipitate adhered to the vial. pH 6 6.2 6.2 >135 101.6 pH 7 7.1 7.0 >133 87.8 The solution was yellow and a sticky precipitate could be observed. pH 8 8.1 8.1 >127 68.3 pH 9 8.7 8.9 >109 61.4

Example 4 Physical Stability of Compound 1 Dihydrochloride

Conducted a study of the physical stability of the dihydrochloride compound 1 (a mixture of form 1 and form 2). A solid sample of compound 1 dihydrochloride was incubated under four relative humidity and temperature conditions: 5°C/RH ambient, 25°C/RH 60%, 30°C/RH 65%, and 40°C/RH 75%. At regular intervals, the compound was analyzed by HR-XRPD and TGMS. After three days of exposure, the original crystalline form of compound 1 dihydrochloride showed complete conversion to form 1 (monohydrate) under all four test sets of conditions. No significant change in solid form was observed when incubated for an extended period of time (at least up to 1 month).

The water content of each sample was determined by TGMS analysis and is shown in FIG. 11. After 3 days of exposure, the water content increased from 3.1% to values from 3.8 to 4.1%, corresponding to 1 or 1.1 water molecules per molecule of dihydrochloride compound 1. The water content is consistent with the monohydrate nature of dihydrochloride form 1 Compound 1. TGMS assay results recorded at 2 weeks and 1 month incubation are also closer to 3.8% water (1 water molecule per API molecule).

These data show that Form 1 is physically stable and non-hygroscopic under the test conditions. No change in the crystalline phase was observed for samples incubated for 4 weeks. Moreover, the absence of significant changes in water content was recorded when exposed to various conditions and at various points in time.

Example 5 Screening for compound 1 dihydrochloride polymorphs.

Obtaining an amorphous powder of dihydrochloride compound 1.

Approximately 20 mg of compound 1 dihydrochloride was weighed into nine separate 1.8 ml glass vials and nine selected solvent systems were added to these vials in stages until a clear solution was obtained. In several solvent systems (2-propanol water, 1,4-dicosan/water, tetrahydrofuran/water, ethanol/water, acetone/water, and acetonitrile/water; all at 90/10 v/v), the API did not dissolve; therefore, these experiments were not taken into account. Solutions (prepared from methanol/water, 90/10 v/v; dimethyl sulfoxide/water, 1/1 v/v, and water) were frozen in liquid nitrogen followed by drying using a Crys Alpha 2 lyophilizer -4 LO for 24 h. Additional drying at 50° C. and 5 mbar pressure for 4 h was applied. The resulting solids were analyzed by HT-XRPD. An amorphous solid was obtained from one solvent, water. These conditions were scaled up to give 1600 mg of an amorphous solid for polymorph screening.

Polymorph screening experiments.

The comprehensive polymorph screening plan is characterized by the use of various crystallization techniques in combination with a range of solvents and solvent mixtures. The versatility of screening will depend on the solubility profile of the API. Certain crystallization modes, such as suspension equilibration, require intermediate solubility, while anti-solvent crystallization relies on solvents with good solubility and solvents with near zero solubility. Polymorph screening consisted of the following crystallization modes:

solvent equilibration at RT and 50°C in 25 solvent systems;

evaporative crystallization;

crystallization on cooling with hot filtration;

- 29 042812 crystallization with rapid cooling (crash-crystallization) using the addition of an anti-solvent;

crystallization during thermal cycling;

ultrasonic treatment;

diffusion from the vapor phase to solids;

diffusion from the vapor phase into the liquid.

Solids from various crystallization experiments were analyzed by high throughput XRPD (HT-XRPD). If appropriate, the mother liquors were allowed to evaporate completely and the remaining solids were analyzed by HT-XRPD. Thereafter, all solids were subjected to artificial accelerated aging conditions for 48 hours at 40° C. and 75% RH (AAC).

Four new diffraction patterns were identified from the polymorph screening in addition to the two known dihydrochloride forms of Compound 1, Form 1 and Form 2. Form 1 was the most common solid form found in this polymorph screening. The new diffractograms were named forms 3, 4, 5, and 6. All of these forms were detected at low frequency and had a weakly crystalline structure. All of them were physically unstable, since the conversion to form 1 was observed under short-term exposure to stressful conditions (2 days at 40°C and RH 75%). The HT-XRPD patterns of the new forms 3-6 are shown in FIG. 12-15 respectively. In table. 11 is a brief description of the experimental conditions under which these forms were obtained.

Table 11

Summary of Compound 1 Dihydrochloride Solid Forms Detected in Polymorph Screening

Form Physical stability after exposure to AAS Crystallization conditions Solvents for crystallization 1 stable - - 2 (with traces of form 1) turned into shape 1 equilibration with solvent at 50°C 1,2-dimethoxyethane, DMF, DMA, DMSO/DCM (5/95), DMSO/MTBE (5/95), DMA/toluene (5/95), DMSO/anisole (75/25), NMP/THF ( 50/50), THF/water (80/20) thermal cycling NMP/THF (50/50), DMSO/Anisole (25/75) antisolvent water (S), THF (AS) 3 turned into shape 1 solvent equilibration at RT (wet solid) iso-amyl alcohol 4 turned into shape 1 solvent equilibration at RT (wet solid) TFE crystallization at cooling TFE hydrate screening water/IP A (2.6/97.4) 5 turned into shape 1 Solvent equilibration at RT (dried solid) iso-amyl alcohol equilibration with solvent at 50°C iso-amyl alcohol hot filtration chloroform/isopropyl acetate (50/50)

- 30 042812

6 turned into shape 1 equilibration with solvent at 50°C 1,2-Dimethoxyethane, DMSO/DCM (5/95), DMF/anisole (5/95) crystallization at cooling ethanol/para-xylene (50/50) evaporative crystallization 1-propanol screening of hydrates at 35°C and 50°C water/IP A (0.7/99.3, 2.6/97.4)

The physical stability of the forms was determined when exposed to 40°C and RH 75% for 2 days (AAS). (S) means solvent, (AS) means anti-solvent. All ratios are about./about.

All of the above forms, with the exception of Form 3, were further analyzed by DSC, TGMS and HPLC. A brief description of the analytical characteristic is presented below.

Form 3 was obtained as a pure crystalline phase in one experiment. No further analysis could be performed on this form due to limited physical stability. This form was found in a solid obtained from isoamyl alcohol after drying at ambient conditions. Drying in vacuo converted to form 1. Also, upon short term stress, form 3 converted to form 1. Form 4 was found in a small number of crystallization experiments performed in TFE, 1,4dioxane and IPA/water. Thermal analysis confirmed that Form 4 is a mixed solvated/hydrated form. Based on the various solvent/water content values determined for several samples of Form 4, the present inventors have assumed that Form 4 is a class of isostructural mixed hydrate/solvates. A mixed hydrated/solvated with isoamyl alcohol form was identified from solvent equilibration experiments carried out in isoamyl alcohol. This new solid form was called Form 5, which appeared to be physically stable when dried under vacuum; however, conversion to form 1 was observed upon exposure to AAS.

Form 6 was predominantly detected in hydrate screening experiments performed in IPA/water mixtures, but was also identified in a small number of solvent equilibration experiments performed in various solvents such as 1,2 dimethoxyethane, DMSO/DCM and DMF/anisole. Form 6 was mainly found in vacuum dried solids; however, conversion of Form 1 was observed after AAS. Analytical characterization indicated that Form 6 is a mixed hydrated/solvated form. In view of the various solvents that may be included in the structure of form 6, the authors of the present invention assume that this form combines a class of isostructural mixed hydrate/solvates containing molecules of various organic solvents.

Hydrate screening.

Solvent equilibration experiments were carried out in several water/organic solvent mixtures to study the formation of other hydrated forms. Suspensions of amorphous dihydrochloride of compound 1 were prepared in 15 water/solvent mixtures with various water activities. The suspensions were stirred for 7 days at 5, 20, 35 and 50°C.

At the end of the equilibration time, the remaining solids were analyzed by HT-XRPD in the dry state after drying at ambient conditions and after drying in vacuum. Mother fluids were analyzed by Karl Fischer titration to determine the water content. Three experiments showed no solids after exposure to equilibration conditions. Solutions from these experiments were evaporated in vacuo (200 mbar) and the resulting dried solids were analyzed by HT-XRPD. All solids were then exposed to 40° C. and 75% RH for two days.

Under most crystallization conditions, the monohydrate form 1 crystallized.

Form 4, Form 6 and mixtures thereof with Form 1 were identified from IPA/water mixtures at low water contents (<5%). At water activity values above 0.5, the monohydrate form 1 was identified in all cases. Based on the performance in the analysis, Form 4 and 6 are mixed hydrated/solvated forms, which, upon short-term exposure to stressful conditions, were converted to Form 1.

Solvent equilibration experiments performed in ethyl acetate/water showed differences in solid form between ambient and vacuum dried solids. Form 1 was identified in most ambient dried solids. Drying in vacuum identified mixtures of Forms 1 and 2. Form 6 was identified in a mixture with Form 1 at water activity values below 0.3 at temperatures above 35°C. The presence of form 2 was observed using elevated temperatures and high water content, as well as at temperatures below 20°C and low water content.

A similar trend was observed in the case of experiments carried out in acetonitrile e/water. All solids dried at ambient conditions were Form 1. When solids were dried in vacuo, partial conversion to Form 2 was observed in some cases. This observation was noted in the case of solvent equilibration experiments carried out at temperatures above 35°C. Experiments carried out at temperatures below 25°C did not show the transformation of any transformation of the solid form upon drying. A physical stability study carried out for 2 days at 40°C/RH 75% led to the identification of form 1. All new powder diffraction patterns identified in this study showed conversion to the original form 1.

Example 6 Stability of the Combined Formulation.

The stability of aqueous solutions of compound 1 dihydrochloride in combination with meropenem at pH 5.0 was studied in comparison with aqueous solutions of compound 1 dihydrochloride in combination with cefepime at pH 5.0 after 24 hours. 12

Table 12

% present at pH 5.0 at 25°C Past tense 24 hours 24 hours 20 hours Solution composition compound 1 meropenem cefepime compound 1 100 - - meropenem - 89 - cefepime - - 99 compound 1 + meropenem 57 20 - % present at pH 5.0 at 25°C Past tense 24 hours 24 hours 20 hours Solution composition compound 1 meropenem cefepime compound 1 + cefepime 99.9 - 99.1

Example 7 Efficiency of the Combined Formulation.

The results presented in examples 7A-7D demonstrate that the combination of cefepime/compound 1 is more effective than the combination of meropenem/compound 1 against a number of organisms.

Example 7A. Elizabethkingia meningoseptica.

Elizabethkingia meningoseptica is a gram-negative bacillus that expresses 2 chromosomal metallo-beta-lactamases (both carbapenemases) and class A serine cephalosporinase CME (AAC, 2012, 1686-1692). The results of the study MIC (minimum inhibitory concentration) (N=10 strains) are presented in table. 13.

Table 13

Connections Range MICso (mcg/ml) Ml Sei (mcg/ml) %S@8 cefepime 16-32 16 32 0 Cefepime/compound 1@4 2-8 4 4 100 Meropenem 16-128 64 128 0 Meropenem/compound 1@4 4-16 8 16 70

Example 7B. Stenotrophomonas maltophilia.

Stenotrophomonas maltophilia is a Gram-negative bacillus. It is an opportunistic pathogen with MDR. It has been associated with infections with high morbidity and mortality in severely immunocompromised and debilitated individuals. The results of the MIC study are presented in table. 14.

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Table 14

Connections Range MICso (mcg/ml) MIC90 (mcg/ml) %S cefepime 1 - 128 32 64 20 Cefepime/compound 1@4 0.5 - 16 4 16 80 Meropenem 64->128 128 >128 0 Meropenem/compound 1@4 16->128 64 >128 0

Example 7C. Pseudomonas aeruginosa.

507 isolates are insensitive to meropenem (MIC values for meropenem >8 µg/mL). The results of the MIC study are presented in table. 15.

Table 15

Connections Range MICso (mcg/ml) MIC90 (mcg/ml) %S@8 cefepime 1 - 128 32 256 30.8 Cefepime/compound 1@4 1->64 8 >64 60 Meropenem 16->128 32 >128 0 Meropenem/compound 1@4 16->128 16 64 23.9

Example 7D. Pseudomonas aeruginosa.

554 isolates are insensitive to cefepime (MIC values for cefepime >8 µg/mL). The results of the MIC study are presented in table. 16.

Table 16

Connections Range MICso (mcg/ml) MIC90 (mcg/ml) %S@8 cefepime 16->512 32 256 0 Cefepime/compound 1@4 <0.06 - >64 8 >64 55.8 Meropenem <0.12->128 16 >128 36.6 Meropenem/compound 1@4 <0.06 - >64 8 64 52

Example 8 Randomized Drug Interaction Study to Evaluate the Safety and Pharmacokinetic Characteristics of Compound 1 in Healthy Adult Volunteers.

Example 8A. Phase 1. Review of the clinical trial.

Example 8 describes four Phase 1 clinical studies designed to evaluate the safety, tolerability, and PK of various dose levels of Compound 1 when administered as single or multiple doses, either alone or in combination with cefepime, containing the following components (and set out in Table 17 ):

1) single incremental (SAD) and multiple incremental (MAD) doses of Compound 1 in healthy volunteers; And

2) single dose drug interactions of compound 1, cefepime and metronidazole alone and compound 1 in combination with cefepime with and without metronidazole (part 1) and repeated doses of compound 1 in combination with cefepime (part 2) 10 days later, in healthy volunteers .

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Table 17

Purpose(s) of the study Study plan Dosing regimens Number of subjects Processing time Safety, Tolerability, and PK in Healthy Adult Volunteers Two-part, randomized, double-blind, placebo-controlled Part 1: Single incremental doses of 62.5, 125, 250, 500, 1000 and 1500 mg Compound 1 or placebo. Includes simulation of QT concentration-effect relationship. Part 2: Multiple incremental doses of 250, 500, and 750 mg compound 1 or placebo as a 2-hour q8h IV infusion Part 1: 6 investigational medicinal products and 2 placebo at each dose level Part 2: 9 investigational medicinal products and 3 placebo at each dose level single dose 10 days DDI; Safety, Tolerability, and PK in Healthy Adult Volunteers Two-part, randomized, three-way cross-over drug-drug interaction study Part 1: Single doses of 750 mg compound 1 alone, 2 g cefepime alone, 500 mg metronidazole alone, or compound 1 in combination with cefepime with or without metronidazole metronidazole Part 1: 18 subjects Single dose in each of 5 periods dosing Part 2: cefepime 2 g with compound 1,500 mg or 750 mg as 2 hourly IV q8h infusions for 10 days Part 2: 24 subjects (10 subjects/dose level) and 4 placebo subjects 10 days

Abbreviations: ~ = approximately;

BAT = best available therapy;

DDI = drug interaction;

h = hour;

IV = intravenous;

PK = pharmacokinetic parameters;

q8h = three times a day with an interval of 8 hours;

TBD = to be determined.

Assessing the achievement of target pharmacokinetic parameters and pharmacokinetic/pharmacodynamic parameters for compound 1 in humans.

Results from in vitro and in vivo preclinical PK/PD studies, human PK data from the phase 1 study, and human plasma binding data were used to determine the human therapeutic doses required to achieve target PK/PD characteristics. In vitro studies have shown that the binding of compound 1 to human serum proteins is 37%, leaving an unbound proportion of 63%. In table. 18 summarizes the calculated values for the total AUC _0-24 , the corresponding proportion of unbound compound (fAUC _0-24 ), and the estimated ratio/AUC _0-24 /MIC, based on the established CLSI inflection point for cefepime of 8 mg/L when administered at a dose of 2 g three times a day.

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Table 18

Dose level (q8 hr) Total AUCo-24 (h*mg/l) /AUCo-24 (h*mg/l) Ratio /AUCo-24/MIC (for М1С=8 mg/l) 250 123 77 9.7 500 248 156 19.5

Abbreviations: AUCo _-24 = area under the plasma concentration versus time curve from time zero to h;

fAUC ₀ - ₂₄ = area under the unbound drug plasma concentration versus time curve from time zero to 24 hours;

MIC = minimum inhibitory concentration; q8 hr = every 8 hours

The fAUC _0-24 /MIC ratios (based on a MIC value of 8 mg/L) for all doses >250 mg every 8 hours exceeded the PK/PD targets required to achieve arrest of bacterial growth for the isolates studied to date. Similarly, fAUC _0-24 /MIC values for doses of approximately 375 mg every 8 hours (interpolated from the data shown) exceeded the fAUC _0-24 /MIC ratios required to achieve 1 log bacterial kill. Based on these data, a dose of 500 mg every 8 hours provides approximately three times the drug exposure required to achieve the target PK/PD to stop bacterial growth.

Brief description of the protocol.

Population: 42 subjects in total, divided between two parts of the study as follows:

part 1: crossover with single doses of 5 periods: 18 healthy male subjects and female subjects;

part 2: 10-day randomized repeat dose: 24 healthy male and female subjects.

Inclusion Criteria.

No exceptions will be allowed for inclusion/exclusion criteria. A subject must meet the following inclusion criteria in order to be eligible for enrollment in the study.

1. Willingness to participate in the study, willingness to give written informed consent, and compliance with study limitations.

2. Sex: male or female with a negative pregnancy test (human chorionic β-gonadotropin [β-hCG]) at Screening and Day -1; women may or may not be of reproductive potential.

3. Age: 18-55 years old inclusive, at screening.

4. Weight: >50 kg.

5. Body Mass Index (BMI): >18.5 and <30.0 kg/m ² .

6. Normal blood pressure, defined as systolic values >90 and <140 mmHg. Art. and a diastolic value less than 90 mm Hg. Art. (screening and Day -1). Values out of range for inclusion may be retested if there is clinical evidence that the value is out of range.

7. Urine dipstick protein results are negative or show trace amounts at screening and on Day -1.

8. All values for hematological and clinical biochemical analyzes of blood and urine are either in the normal range or are defined as acceptable exceptions in Table. 3 at screening and on Day -1. Values outside these ranges may be retested if there is clinical evidence that the value is out of range. Screening laboratories are identified in Table 1. 9 and 10.

9. Ability and willingness to abstain from alcohol 48 hours (2 days) prior to visiting the clinical research site prior to the follow-up visit.

10. Men who are not surgically sterilized and women of childbearing potential must agree to use highly effective contraceptive methods during the study and for 90 days after the last dose of study drug. A woman is considered of reproductive potential if she is not post-menopausal (>1 year without menses) or is surgically sterilized by bilateral oophorectomy, hysterectomy, bilateral tubal ligation, or successful Essure® placement with documented confirmatory testing at least 3 months after the procedure.

Highly effective contraception is defined as a contraceptive method that has a failure rate of <1% when used systematically and correctly. These methods are:

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intrauterine device (IUD) or intrauterine system (IUS);

double barrier methods of contraception (eg, male condom with a diaphragm, male condom with a cervical cap);

monogamous relationship with a partner with a vasectomy;

complete abstinence from alcohol in accordance with the lifestyle of the subject.

11. All over-the-counter [OTC] medicines, dietary supplements, and herbal medicines (eg, St. John's wort extract) must be discontinued at least 14 days prior to visiting the clinical research site. An exception is made for acetaminophen, which is allowed at doses <3 g per day before visiting the clinical research site.

12. Suitable veins for catheterization/multiple venipunctures as assessed by the investigator at screening.

exclusion criteria.

A subject that satisfies any of the following exclusion criteria will not be eligible for inclusion in the study.

1. Sponsor or Principal Investigator employee.

2. A woman who is pregnant, breastfeeding, or plans to try to become pregnant during this study or within 90 days of dosing study drug.

3. A man with a female partner who is pregnant or breastfeeding during this study, or plans to try to become pregnant during this study or within 90 days of dosing study drug.

4. Use of any investigational medicinal product or device within 30 days prior to screening (90 days for an injectable biological agent).

5. The presence of congenital or acquired immunodeficiency syndrome.

6. The presence of ongoing cardiovascular, respiratory, hepatic, renal, gastrointestinal, endocrine, autoimmune, hematological, neoplastic or neurological disorders.

7. Any medical condition that, in the opinion of the investigator, places subjects at unacceptable risk.

8. Clinically significant ECG abnormality defined as mean QTcF >450 ms, short QTcF (<300 ms), or ECG evidence of atrial fibrillation, atrial flutter, complete right or left bundle branch block, Wolff syndrome Parkinson-White or pacemaker (screening or Day -1).

9. Current or history of second- and third-degree heart block and/or clinically significant prolongation of the PR interval, as determined by the investigator (screening or Day -1).

10. A history of drug allergy with a severity that requires emergency medical attention, such as treatment with epinephrine in the emergency department.

11. Any hypersensitivity reaction after administration of a cephalosporin, penicillin, or other beta-lactam antibacterial agent in history.

12. Hypersensitivity to metronidazole or other nitroimidazole derivatives in history.

13. Requires regular use of chronic drug or prescription drug or vaccination within 14 days prior to Day -1. Hormonal contraceptives are allowed.

14. Use of >3 g acetaminophen for 3 or more days in the 14 days prior to screening.

15. Physical activity, sunbathing and contact sports within 48 hours (2 days) before visiting the clinical research site and during the study period.

16. Previous donation of more than 450 mL of blood within 60 days prior to receiving a dose at a clinical trial site or planned donation before 30 days have elapsed since the final dose of study drug.

17. Plasma or platelet donation within 7 days of receiving a dose and throughout the study.

18. Current potential drug or alcohol abuse, as defined in the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition, revised text criteria.

19. Recent history of alcohol consumption exceeding 2 standard drinks per day on average (1 standard drink = 10 g of alcohol).

20. Positive test result for alcohol or drugs at screening or on Day -1.

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21. Positive screening test result for hepatitis B surface antigen (HBsAg), antibodies to hepatitis C virus (HCV), or antibodies to human immunodeficiency virus (HIV) 1 and 2.

22. Temperature >38.5°C or acute illness at entry (Day -1).

23. Prior participation in Compound 1 study.

Investigational drugs.

Identification information for the study product(s) and placebo.

Compound 1 was provided in glass vials containing 500 mg of lyophilized solid. Compound 1 was reconstituted in situ with sterile water for injection.

The following products were obtained locally from a commercial pharmacy or wholesaler:

cefepime for IV infusion, metronidazole, 500 mg, immediate-release tablets, USP sodium chloride injection 0.9%, water for injection.

Goals.

Part 1.

Home.

Assess the pharmacokinetic parameters (PK) of compound 1 and cefepime after single doses alone and in combination (Part 1A).

Additional.

Assess the PK of metronidazole PK after single doses of metronidazole alone and in combination with Compound 1 and cefepime (Part 1B).

Assess the PK of Compound 1 and the PK of cefepime after single doses of co-administered Compound 1 and cefepime co-administered with metronidazole (Part 1B).

To evaluate the safety and tolerability of a single dose of compound 1 in combination with cefepime, with and without metronidazole, compared with compound 1 alone, cefepime and metronidazole in healthy adult volunteers.

Part 2.

Home.

To evaluate the safety and tolerability of a repeat dose (10 days) at two dose levels of compound 1 in combination with cefepime in healthy adult volunteers.

Additional.

To evaluate the residual plasma concentrations of compound 1 and cefepime with repeated doses (10 days) at two dose levels of compound 1 co-administered with cefepime in healthy adult volunteers.

Research.

Assess bactericidal activity in serum and urine at peak and residual concentrations.

Research plan.

It is a phase 1, randomized, single-center, drug-drug interaction study in healthy male and female volunteers. The study consisted of two independent parts. Each part included separate groups of subjects. Eligible subjects were allowed to participate in either Part 1 or Part 2, but not both.

Part 1.

Part 1 included 18 subjects in a randomized evaluation with a single dose for five treatments according to the plan with an overlap of consecutive 3 periods (Part 1A) and an overlap of 2 periods (Part 1B). Subjects who did not meet any of the barrier rules in Part 1A were allowed to re-randomize in Part 1B.

Part 2.

Part 2 included 24 subjects in a randomized, double-blind evaluation of three treatments according to a parallel group design.

Rationale.

Given the available preclinical data and data from single and multiple dose administration in humans, further development of Compound 1 in combination with a β-lactam antibiotic was warranted. This study provided an assessment of the safety and pharmacokinetic parameters of Compound 1 in combination with cefepime with and without metronidazole according to a crossover design with single doses administered over multiple periods in Part 1 and the safety, tolerability and pharmacokinetic parameters (PK) of multiple doses of Compound 1 administered in combinations with cefepime in part 2.

Compound 1 was administered as an intravenous (IV) infusion over 2 hours.

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Rationale for the selection of subjects.

Because of the design of the study, the low risk of clinically significant toxicity at expected levels of exposure, and the absence of any potential therapeutic utility in this study population, healthy subjects were selected. Moreover, the use of healthy subjects as opposed to patients allowed for a clearer interpretation of study results as there were no confounding factors resulting from changes in disease state and/or concomitant medications.

Example 8B. Part 1 study with multiple doses.

Part 1A.

All treatments in Part 1A were administered as a single 2 hour IV infusion and included the treatments in Table 1A. 19. All subjects received all three treatments. Dosing occurred on Day 1, Day 4 and Day 7. The sequence of administration was randomized. Descriptions of treatments in part 1A are presented in table. 19.

Table 19

Treatment Processing Description A 750 mg Compound 1 IN 2 g cefepime WITH 750 mg compound 1 + 2 g cefepime

Part 1B.

Part 1B included two treatments that were performed according to the treatment sequence defined in the randomization code. Dosing occurred on Day 10 and Day 13. Treatments are described in Table 1. 20.

Table 20

Treatment Processing Description D metronidazole 500 mg + compound 1 750 mg + cefepime 2 g E metronidazole 500 mg + cefepime 2 g

Research notes.

In each processing period, certain evaluation procedures were repeated.

Subjects were released after completion of all evaluation procedures on Day 15.

If the subject withdrew from the study before Day 15, all ET assessment procedures were completed at the time of termination. A scheduled score received within 1 day ET could be used instead of the ET score.

Additional general or symptom-targeted assessment procedures could be performed at other time points at the discretion of the principal investigator.

Screening included measurement of height, weight and calculation of BMI. Day -1 includes weight measurement and BMI calculation.

Clinical laboratory evaluation included serological analysis (HBsAg, anti-HCV antibodies and anti-HIV 1 and 2 antibodies) at screening.

Samples for PK studies were collected before each treatment and at 0.5, 1, 2 (end of infusion), 2.25, 2.5, 3, 4, 6, 8, 12, 16, 24, 36 and 48 hours after start of infusion.

Adverse events (AE) were recorded from visit (Day -1) to the follow-up visit. Any AEs continuing at follow-up were followed up until resolution or stabilization.

Subjects received one dose of the assigned randomized treatment on Days 1, 4, 7, 10, and 13. All subjects received all treatments in a predetermined order according to the randomization scheme. Compound 1 and cefepime were administered IV. Metronidazole was administered orally.

Included FSH (follicle stimulating hormone) screening in post-menopausal women.

Example 8C. Part 2 multi-dose study.

In Part 2, subjects were randomized in a 5:5:2 ratio to one of the three treatments in Table 1. 21.

Table 21

Treatment Processing Description 2A 500 mg compound 1 + 2 g cefepime 2B 750 mg compound 1 + 2 g cefepime 2C placebo (corresponding to compound 1 + cefepime)

Treatments were administered every 8 hours (q8h) as a 2-hour IV infusion over 10 hours. Subjects received a total of 28 doses, with one dose administered on Day 10.

Subjects were released after review of laboratory and ECG results on Day 10 and completion of all evaluation procedures on Day 11.

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If the subject withdrew from the study before Day 11, all ET assessments were completed at the time of discontinuation in the case of subjects who did not complete all assessments after the final dose of study drug.

Subjects who discontinued participation were required to complete a follow-up visit 7 days (±1 day) after the last dose of study drug.

Additional general or symptom-targeted assessment procedures could be performed at other time points at the discretion of the principal investigator.

Growth measurement and BMI calculation only at screening. Measurement of body weight at screening and on Day -1 (introduction).

Day 1 and 10 ECGs were performed after the first infusion of the day.

Vital signs include supine systolic and diastolic blood pressure, pulse, body temperature, and respiratory rate. On Days 1-9, vital signs were recorded before and 2 hours after the start of the first infusion of the day. On Day 10, vital signs were recorded before and after dosing (at any time).

Study drug was administered as a 2-hour intravenous infusion every 8 hours (a total of 28 doses with the last dose on the morning of Day 10).

Samples were collected before each dose on Days 1 and 2 and before the morning dose on Days 3, 5 and 10.

Blood samples were collected on Day 1 before dosing and on Day 10 at the end of the infusion and 8 hours after the start of the infusion. Urine samples were collected on Day 1 pre-dose (one sample 12 h pre-dose) and on Day 10 as pooled whole urine samples collected at intervals of 0-4 and 4-8 h after the start of the infusion.

AEs were recorded from visit (Day -1) to the subsequent follow-up visit. Any AEs continuing at follow-up were followed up until resolution or stabilization.

Included FSH (follicle stimulating hormone) screening in post-menopausal women.

Results.

There was no significant difference in the pharmacokinetic parameters AUC( ₀ . _inf ) and C _max for compound 1 or cefepime when the two drugs were co-administered compared to either drug alone.

There was no significant pharmacokinetic interaction between metronidazole and the cefepime/compound 1 combination compared to administration of metronidazole alone or the cefepime/compound 1 combination without metronidazole. Both single and repeated doses of study drug were well tolerated. There were no clinically significant abnormalities in vital signs, ECG, or laboratory findings. All adverse events were mild.

Example 9. Treatment of a bacterial infection using complex therapy with cefepime and compound 1.

Subjects with multi-anatomical infections due to resistant pathogens, including lower respiratory infections, complicated intra-abdominal infections (cIAIs), and complicated urinary tract infections (cUTIs), such as pyelonephritis, are treated with the cefepime/compound 1 combination as follows in table. 22.

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Claims (5)

Table 22 Dosing regimens Considered population Duration treatment Compound 1, 500 mg, in combination with cefepime, 2 g, IV q8h patients with cUTI 5-14 days Compound 1, 500 mg, in combination with cefepime, 2 g, IV q8h patients with cIAI 5-14 days Compound 1, 500 mg, in combination with cefepime, 2 g, IV q8h patients infected with resistant pathogens in multiple areas 5-14 days Compound 1, 750 mg, in combination with cefepime, 2 g, IV q8h patients with cUTI 5-14 days Compound 1, 750 mg, in combination with cefepime, 2 g, IV q8h patients with cIAI 5-14 days Compound 1, 750 mg, in combination with cefepime, 2 g, IV q8h patients infected with resistant pathogens in multiple areas 5-14 days Abbreviations: ~ = approximately; cIAI = complicated intra-abdominal infection; cUTI = complicated urinary tract infection; h = hour; IV = intravenous; q8h = three times a day with an interval of 8 hours. Processing in table. 22 is administered every 8 hours (q8h) as a 2-hour IV infusion over 10 hours. Subjects will receive a total of 28 doses, with one dose administered on Day 10. CLAIM 1. Anhydrous crystalline form of dihydrochloride (R)-3-(2-(trans-4-(2-aminoethylamino)cyclohexyl)acetamido)-2-hydroxy-3,4-dihydro-2H-benzo[e] [1,2 ]oxaborinine-8-carboxylic acid wherein the crystalline form is characterized by an X-ray powder diffraction (XRPD) pattern containing at least three characteristic peaks selected from peaks with a 2Θ value of 7.3±0.1°, with a 2Θ value of 10.8±0.1°, with a value of 2Θ 14.5±0.1°, with 2Θ value 16.6±0.1°, with 2Θ value 18.0±0.1°, with 2Θ value 19.6±0.1°, with 2Θ value 19, 7±0.1°, with 2Θ value 23.3±0.1°, with 2Θ value 24.0±0.1°, with 2Θ value 24.3±0.1°, with 2Θ value 27.3± 0.1°and with a 2Θ value of 29.3±0.1°. 2. The anhydrous crystalline form according to claim 1, wherein the crystalline form is characterized by an X-ray powder diffraction pattern (XRPD) containing characteristic peaks with a 2Θ value of 7.3±0.1°, with a 2Θ value of 14.5±0.1°, with with a 2Θ value of 18.0±0.1°, with a 2Θ value of 19.7±0.1°, with a 2Θ value of 24.0±0.1°, and with a 2Θ value of 27.3±0.1°. 3. Anhydrous crystalline form according to claim 2, wherein the powder X-ray diffraction pattern (XRPD) additionally contains characteristic peaks with a 2Θ value of 10.8±0.1°, with a 2Θ value of 16.6±0.1°, with a 2Θ value of 19 .6±0.1°, with a 2Θ value of 23.3±0.1°, with a 2Θ value of 24.3±0.1°, and with a 2Θ value of 29.3±0.1°. 4. Pharmaceutical composition containing: (i) a crystalline form according to any one of paragraphs. 1-3 and (ii) cefepime. 5. Pharmaceutical composition according to claim 4, wherein the pharmaceutical composition is formulated as a homogeneous liquid suitable for injection. -