Field of the invention The present invention relates to a process for decreasing the particle size of the bromide salt of [(3R)-1-(3-phenoxypropyl)quinuclidin-1-ium-3-yl] 2-hydroxy-2,2-bis(2- thienyl)acetate of Formula I

and increase the level of crystallinity of Formula I by a subsequent conditioning method. The present invention has application in the manufacture of chemical compounds, such as active pharmaceutical ingredients and for use in pharmaceutical formulations, such as inhalation formulations.

The present invention is also relates to the production of active pharmaceutical ingredient particle size and distribution thereof that are to form a dry powder formulation to be administered to the lung by using e.g. a drug powder inhaler (DPI) device. In particular the present invention provides the characteristics and the preferred manufacturing processing of particles with improved crystallinity.

Background of the invention

[(3R)-1 -(3-Phenoxypropyl)quinuclidin-1 -ium-3-yl] 2-hydroxy-2,2-bis(2-thienyl)acetate bromide compound which is also known as aclidinium bromide {CAS no. 320345-99- 1) is a long-acting, inhaled anticholinergic agent with a strong affinity and selectivity for all muscarinic receptor subtypes (M1-M5) and kinetic selectivity for the 3 receptor over the M2 receptor. The claimed and approved indication is the maintenance bronchodilator treatment to relieve symptoms in adult patients with chronic obstructive pulmonary disease (COPD). The preparation of aclidinium bromide and a purification thereof applying solid phase extraction is described in patent application WO2001004118. Further process for the preparation of aclidinium bromide applying a reaction between 2,2-dithien-2-ylacetic acid 1-azabicycio[2.2.2]oct-3(R)yl ester and 3-phenoxypropylbromide in ketones and cyclic ethers is disclosed in patent application WO2008009397.

A pharmaceutical composition for inhalation comprising 200 μg of aclidinium is disclosed in patent application WO2009112273 including an average particle diameter of the active pharmaceutical compound within 2-5 pm and a d(10) of 90-160 prn, a d(50) of 170-270 pm and d(90) of 290-400 pm particle size distribution of the carrier particles.

Another pharmaceutical composition for inhalation comprising 400 pg of aclidinium is disclosed in patent application WO2009112274. The patent application describes an average particle diameter of the active pharmaceutical compound within 2-5 pm and a d(10) of 90-160 pm, a d(50) of 170-270 pm and d(90) of 290-400 pm particle size distribution of the carrier particles.

A further pharmaceutical composition intended for inhalation comprising aclidinium in the form of a dry powder in admixture with lactose powder is disclosed in patent application WO2013175013. The present formulation provides a delivered dose of aclidinium equivalent to about 322 mg aclidinium free base and/or a fine particle dose equivalent to about 140 mg aclidinium bromide.

The particle size characteristics necessary for use by inhalation are ensured by micronization which may have a damaging effect on the physical properties of the micronized particles. Particles may undergo morphological alteration during the milling process which leads an undesirable surface morphological transformation resulting in the formation of amorphous phases that is unsuitable for pharmaceutical formulation designed for inhalation. Furthermore, micronization may generate considerable heat leading to an inappropriate micronized material in case of low melting active pharmaceutical ingredients.

A process for the decrease of the amorphous content of a given solid material and increase the crystallinity thereof is provided in patent application WO2011048412 applying ultrasound for a solid compound which is less than 100% crystalline. Therefore the objective of the present invention to provide a novel process for the preparation of a crystalline aclidinium bromide with a particle size less than 10 pm which is suitable for industrial scale application and easily reproducible.

Summary of the invention

The present invention relates to a process for the preparation of crystalline aclidinium bromide having a particle size less than about 10 pm measured by laser diffraction method comprising a micronization of the crystalline aclidinium bromide to obtain a particle size less than about 10 pm and a subsequent storing and conditioning step of the micronized aclidinium bromide to increase the crystallinity of the micronized aclidinium bromide.

An advantage of the new process consists in good physical and chemical stability of aclidinium bromide which makes it suitable for preparation of a dosage form intended for inhalation.

The novel process offers a robust, scalable and reproducible preparation method of aclidinium bromide having a particle size less than about 10 pm and supplies physically and chemically stable particles for the formulation of a dry powder used for inhalation.

Detailed description of the invention

The aim of the present invention is to provide a process for the preparation of aclidinium bromide with particle size decreased and a subsequent conditioning step for the improvement of the crystallinity thereof.

A conventional micronization process of any drug compounds may involve the injection of a relatively coarse powder into a system which applies multiple highspeed collisions. A typical non-micronized powder used as source of the micronization process contains particle with size substantially greater than 30 pm, more precisely greater than 20 pm. The objective of a micronization process is to reduce the primary particle size to a size which is small enough to be delivered to the respiratory system and airways. It is known that an appropriate size may be 0.1 pm to 10 pm and preferably 0.5 pm to 6 pm.

The multiple high-speed collisions occurred in micronization provide the milling energy required to the breakage of the particles down to the size demanded. It is also known that such milling energy may also induce the generation of non-crystalline material, especially on the surface of the particles. Such non-crystalline material may be an amorphous material.

The particle size and distribution thereof can be measured by laser diffraction techniques that involves a light from a laser source passes through a dispersion of particles suspended in a transparent gas or liquid. The particles scatter the light: large particles scatter the light at small angles; small particles scatter the light at large angles relative to the laser beam. The scattered light is collected by a series of photodetectors located at various angles. The scattering intensity is analyzed and scattering pattern created applying a light scattering theory which assumes the particles to be spherical. Particle size and distribution thereof is reported as volume equivalent sphere diameter.

Various techniques may be used for the determination of the crystalline content of a material. X-Ray Powder Diffraction (XRPD) may be a useful technique for the identification of a crystalline material. Crystalline particles have distinct, characteristic pattern (fingerprint) for a given solid form. Amorphous materials do not show a diffraction pattern having no periodic array with long-range order and provide a broad hump or noise (halo). Differential Scanning Calorimetry (DSC) may also prove a clear melting point and the corresponding heat of fusion which is related to the level of crystallinity in a given sample. Differential Scanning Calorimetry curve of a crystalline sample reveals a sharp melting phenomenon comparing to an inconsistent behaviour of an amorphous phase. Raman spectroscopy may give an indication of a crystalline compound as Raman spectra of an amorphous compound can be easily distinguished. Raman spectroscopy also allows distinguishing between crystalline solid phases.

For the purposes of the present patent application Differential Scanning Calorimetry is the preferred method to determine the amorphous content of a given sample. Differential Scanning Calorimetry analysis can be measured by various commercially available apparatus including a Perkin Elmer instrument's Pyris 1 , referred as DSC for measurement of a DSC curve and determine crystalline content of a given sample in the present application.

Typically an appropriate amount of material is weighted into the sample pan of the 5 DSC instrument and subjected to a heating ramp of 10°C/min up to 240°C. The endotherm of the melting point phenomenon and the integral of the heat flow provide qualitative and quantitative information in respect to the crystallinity. Known compositions of amorphous and crystalline phases of a given compound allows direct comparison of the two samples and clearly shows quantitative information on the L0 content of the crystalline compound.

It has been found from detailed studies of aclidinium bromide that the presence of the non-crystalline or amorphous aclidinium bromide material may be dependent on the increase of the specific surface area obtained by the micronization process and consequently on the energy applied for micronization. Additionally, the micronized L5 product may be conditioned to decrease the content of the amorphous material and improve the crystallinity of the micronized product.

In one aspect, the present invention provides a process for the preparation of crystalline aclidinium bromide having a particle size less than about 10 μηη measured by laser diffraction method comprising a micronization of the crystalline aclidinium 20 bromide to obtain a particle size less than about 10 pm and a subsequent storing and conditioning step of the micronized aclidinium bromide to increase the crystallinity of the micronized aclidinium bromide.

Preferably, the particles of the micronized aclidinium bromide have a volume diameter equal or less than 10 μηι, more preferably at least the 90 wt% of the particles of the

25 micronized aclidinium bromide have a diameter equal or less than 10 pm as determined by measuring the characteristic equivalent sphere diameter, known as volume diameter, by laser diffraction technique described above; preferably Malvern or equivalent apparatus. The parameters taken into account are the volume diameters in microns of 10%, 50% and 90% of the particles expressed as d(10), d(50) and

30 d(90), respectively.

Preferably, not more than 10 wt% of the particles of the micronized aclidinium bromide have a volume diameter d(10) less than 0.5 pm. Preferably, not more than 50 wt% of the particles of the micronized aclidinium bromide have a volume diameter d(50) less than 2 pm. Preferably, at least 90 wt% of the particles of the micronized aclidinium bromide have a volume diameter d(90) less than 10 μιτι. Preferably, 100 wt% of the micronized aclidinium bromide have a volume diameter equal or less than 10 pm.

The process for micronization of aclidinium bromide comprises the following steps: a/ feeding the solid aclidinium bromide into the jet mill; b/ applying pressure using nitrogen;

c/ collecting the micronized aclidinium bromide;

d/ storing and conditioning the micronized aclidinium bromide.

In another aspect, the present invention provides a process for preparing aclidinium bromide having a particle size less than 10 pm measured by laser diffraction technique applying a micronization energy less than 8000 kJ/kg.

Preferably the micronization energy applied for the micronization of aclidinium bromide between 2000 kJ/kg and 4000 kJ/kg, more preferably between 2000 kJ/kg and 3000 kJ/kg, most preferably about 2300 kJ/kg.

The corresponding solid feed rate of aclidinium bromide into the jet mill is preferably between 0.055-0.110 kg/h. The pressure applied for micronization is preferably between 0.2-0.6 MPa (2-6 bars) at the solid material injection line and between 0.2- 0.6 MPa (2-6 bars) at the grinding line using nitrogen.

More preferably, the solid feed rate of aclidinium bromide into the jet mill is between 0.086-0.110 kg/h. The pressure applied for micronization is between 0.2-0.4 MPa (2-4 bars) at the solid material injection line and between 0.2-0.4 MPa (2-4 bars) at the grinding line using nitrogen.

Most preferably, the solid feed rate of aclidinium bromide into the jet mill is between 0.110 kg/h. The pressure applied for micronization is 0.2 MPa (2 bars) at the solid material injection line and 0.2 MPa (2 bars) at the grinding line using nitrogen.

Preferably, the micronization is performed with a fluid energy mill. One of those skilled in the art appreciates that a fluid energy mill or micronizer is the most frequently used type of mill in the pharmaceutical industry due to the advantages such easy cleaning and very simple mechanism as well as it provides small particles with a relatively narrow size distribution. The fluid energy mill used for the micronization of aclidinium bromide is more preferably a spiral jet mill or a loop jet mill, most preferably a spiral jet mill.

5 The micronization step preferably decreases the particle size of aclidinium bromide to a volume diameter equal or less than 10 pm, more preferably at least the 90 wt% of the particles of the micronized aclidinium bromide have a diameter equal or less than 10 pm as determined by measuring the characteristic equivalent sphere diameter, known as volume diameter, by laser diffraction technique described above; preferably

L0 Malvern or equivalent apparatus. The parameters taken into account are the volume diameters in microns of 10%, 50% and 90% of the particles expressed as d(10), d(50) and d(90), respectively.

Preferably, not more than 10 wt% of the particles of the micronized aclidinium bromide have a volume diameter d(10) less than 0.5 pm. Preferably, not more than L5 50 wt% of the particles of the micronized aclidinium bromide have a volume diameter d(50) less than 2 pm.

Preferably, at least 90 wt% of the particles of the micronized aclidinium bromide have a volume diameter d(90) less than 10 pm.

More preferably, at least 90 wt% of the particles of the micronized aclidinium bromide 10 have a volume diameter d{90) less than 8 pm.

Most preferably, at least 90 wt% of the particles of the micronized aclidinium bromide have a volume diameter d(90) less than 6 pm.

Preferably, 100 wt% of the micronized aclidinium bromide have a volume diameter equal or less than 0 pm.

25 More preferably, 100 wt% of the micronized aclidinium bromide have a volume diameter equal or less than 8 pm.

Most preferably, 100 wt% of the micronized aclidinium bromide have a volume diameter equal or less than 6 pm.

Preferably, the micronized aclidinium bromide may be in the form of crystals in the 30 presence of non-crystalline or amorphous aclidinium bromide. After micronization the micronized aclidinium bromide may contain amorphous phase preferably in content of less than 10% determined by DSC, more preferably in content of less than 8% determined by DSC, most preferably in content of less than 5% determined by DSC.

The inventive process of the present application comprises a subsequent storing and conditioning step to increase the crystallinity of the micronized aclidinium bromide,

5 The conditioning step includes storage at various temperatures and under various relative humidity conditions for an appropriate time period. The storage at defined condition improves the crystallinity of the micronized aclidinium bromide by the decrease of the amorphous content due to the recrystallization thereof. Preferably, the storage condition to be applied is a temperature range between 25 to 80°C and a

D relative humidity of 0% to 75% for at least 4 days in open vessel. More preferably the storage condition to be applied is a temperature of 25°C, 50°C and 80°C and relative humidity of 0% and 75% for a time period of 4 days in open vessel.

The conditioning step described above preferably decreases the amorphous content of the micronized aclidinium bromide below 1%, more preferably below 0.5%, most 5 preferably to 0%.

The aclidinium bromide of the present invention is substantially in a pure form. "Substantially in a pure form" means at least 95% chemical purity. The chemical purity of aclidinium bromide used in the present invention is preferably at least 99%, more preferably 99.5%, most preferably 99.9%. The chemical purity may be determined o according to the method known to the person skilled such as High-Performance Liquid Chromatography (HPLC).

Examples

Instrumentation:

5 Micronizer: Hosokawa Alpine 50 AS spiral jet-mill

Solid feeder: ThreeTec ZD 5 FB-C-1M-50 flat-tray feeder Example 1

Preparation of aclidinium bromide having a particle size less than 6 urn

Certain amount of non-micronized, crystalline aclidinium bromide with an average particle size distribution of about 20-30 m was placed into the flat-tray solid feeder. 5 The crystalline material was micronized applying a 0.110 kg/h of solid feed rate and 2 bars of pressure on the solid injection line and the grinding line, respectively.

Example 2

Preparation of aclidinium bromide having a particle size less than 6 urn

Certain amount of non-micronized, crystalline aclidinium bromide with an average o particle size distribution of about 20-30 pm was placed into the flat-tray solid feeder.

The crystalline material was micronized applying a 0.110 kg/h of solid feed rate and 4 bars of pressure on the solid injection line and the grinding line, respectively.

Example 3

Preparation of aclidinium bromide having a particle size less than 6 urn

5 Certain amount of non-micronized, crystalline aclidinium bromide with an average particle size distribution of about 20-30 pm was placed into the fiat-tray solid feeder. The crystalline material was micronized applying a 0.110 kg/h of solid feed rate and 6 bars of pressure on the solid injection line and the grinding line, respectively.

Example 4

o Preparation of aclidinium bromide having a particle size less than 6 urn

Certain amount of non-micronized, crystalline aclidinium bromide with an average particle size distribution of about 20-30 pm was placed into the flat-tray solid feeder. The crystalline material was micronized applying a 0.086 kg/h of solid feed rate and 2 bars of pressure on the solid injection line and the grinding line, respectively.

5 Example 5

Preparation of aclidinium bromide having a particle size less than 6 urn

Certain amount of non-micronized, crystalline aclidinium bromide with an average particle size distribution of about 20-30 pm was placed into the flat-tray solid feeder. The crystalline material was micronized applying a 0.086 kg/h of solid feed rate and 4 bars of pressure on the solid injection line and the grinding line, respectively.

Example 6

Preparation of aclidinium bromide having a particle size less than βμιπ

5 Certain amount of non-micronized, crystalline aclidinium bromide with an average particle size distribution of about 20-30 Mm was placed into the flat-tray solid feeder. The crystalline material was micronized applying a 0.086 kg/h of solid feed rate and 6 bars of pressure on the solid injection line and the grinding line, respectively.

Example 7

o Preparation of aclidinium bromide having a particle size less than 6 pm

Certain amount of non-micronized, crystalline aclidinium bromide with an average particle size distribution of about 20-30 pm was placed into the flat-tray solid feeder. The crystalline material was micronized applying a 0.055 kg/h of solid feed rate and 2 bars of pressure on the solid injection line and the grinding line, respectively.

L5 Example 8

Preparation of aclidinium bromide having a particle size less than 6 um

Certain amount of non-micronized, crystalline aclidinium bromide with an average particle size distribution of about 20-30 pm was placed into the flat-tray solid feeder. The crystalline material was micronized applying a 0.055 kg/h of solid feed rate and !0 4 bars of pressure on the solid injection line and the grinding line, respectively.

Example 9

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 7.5% of amorphous phase of aclidinium bromide prepared according to Example 4 was kept at 25°C for 4 days in is closed vessel.

The amorphous content determined after the 4-day storage was decreased to 3.0%. Example 10

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 7.5% of amorphous phase of aclidinium bromide prepared according to Example 4 was kept at 25°C under 75% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%. Example 11

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 7.5% of amorphous phase of aclidinium bromide prepared according to Example 4 was kept at 50°C under 0% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%. Example 12

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 7.5% of amorphous phase of aclidinium bromide prepared according to Example 4 was kept at 50 ^C C under 75% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%. Example 13

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 7.5% of amorphous phase of aclidinium bromide prepared according to Example 4 was kept at 80°C under 0% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%. Example 14

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 7.5% of amorphous phase of aclidinium bromide prepared according to Example 4 was kept at 80°C under 75% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%. Example 15

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 4.9% of amorphous phase of aclidinium bromide prepared according to Example 2 was kept at 25°C for 4 days in closed vessel.

The amorphous content determined after the 4-day storage was decreased to 1.4%. Example 16

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 4.9% of amorphous phase of aclidinium bromide prepared according to Example 2 was kept at 25°C under 75% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%. Example 17

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 4.9% of amorphous phase of aclidinium bromide prepared according to Example 2 was kept at 50°C under 0% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%. Example 18

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 4.9% of amorphous phase of aclidinium bromide prepared according to Example 2 was kept at 50°C under 75% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%. Example 19

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 4.9% of amorphous phase of aclidinium bromide prepared according to Example 2 was kept at 80°C under 0% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%. Example 20

Conditioning of the micronized aclidinium bromide

The micronized aclidinium bromide containing 4.9% of amorphous phase of aclidinium bromide prepared according to Example 2 was kept at 80°C under 75% relative humidity for 4 days in open vessel.

The amorphous content determined after the 4-day storage was 0%.

Analysis - XRPD (X-Ray Powder Diffractometry)

Diffractograms were obtained with laboratory XPERT PRO PD PANalytical diffractometer, used radiation CuKa {λ = 1.542A).

Generator settings:

- excitation voltage 45 kV

- anodic current 40 mA.

Scan description:

- scan type - gonio

- measurement range 2 - 40° 2Θ

- step size 0.01° 2Θ - step time: 0.5 s.

Samples were measured as received on Si plate (zero background holder).

Incident beam optics: programmable divergence slits (irradiated length 10 mm), 10 mm mask, 1/4° anti-scatter fixed slit, 0.02 rad Soller slits.

Diffracted beam optics: X'Celerator detector, scanning mode, active length 2.122°, 0.02 rad Soller slits, anti-scatter slit 5.0 mm, Ni filter.

Analysis - DSC (Differential Scanning Calorimetry)

DSC curves were obtained in DSC Pyris 1 (Perkin Elmer). About 5 mg of sample was weight into aluminum cup and crimped (not hermetically) immediately after weighting. Sample was measured by the following temperature program: start at 30°C, hold for 1 minute and heat with temperature ramp 10°C/min to 240°C.

The DSC method was used for evaluation of amorphous content. Pure amorphous material recrystallized (exotherm) at temperatures with T ₀ NSET- 112°C - 120°C and melt (endotherm) at temperature with T ₀ NSET= 220°C - 225°C. Calibration was performed with amorphous sample batch and crystalline sample batch. Evaluation was done based on exothermic peak area integration.

Analysis - PSD (Laser granulometry of particle size distribution) Particle size distribution was determined by Malvern Mastersizer equipped with wet dispersion unit Hydro 2000S (Malvern).

About 50 mg of sample was dispersed in silicon oil (polydimethylsiloxane oil, Tegiloxan 3, Evonik Industries).

Operating conditions:

API Particle Rl (estimated) = 1 ,543

Dispersant Rl (estimated) = 1 ,43

Internal ultrasound conditions: pre-measurement 40% for 5 min, 1 min delay Stirring speed: 1500 rpm Measurement time: 10s

Background time: 10s

Number of measurement per stage: 5

Analysis - SSA (Specific Surface Area)

Specific surface area was measured by BET based on nitrogen adsorption with NOVA2000e (Quantachrom).

Approximately 0.5 g - 1.0 g of the sample was conditioned at 60°C for 2 hours at vacuum.

Operating conditions:

Adsorbate gas: nitrogen

Thermal delay: 600s

P0 mode: calculate

BET measurement: 12 points adsorption in interval 0.04 - 0.3 P/P0, Equilibrium: 0.05 mm Hg

Equilibrium time: 100s

Equilibrium timeout: 300s