Nanoformulation of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate

Abstract

The present invention relates to a stable nanosuspension of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate (Vericiguat, the compound of formula (I)), a method for preparing the stable nanosuspension, nanoparticles comprising the compound of formula (I), and solid pharmaceutical compositions made from the nanosuspension.

Description

[Technical Field]

[0001] The present invention relates to a stable nanosuspension of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate (Vericiguat, the compound of formula (I)), a method for preparing the stable nanosuspension, nanoparticles comprising the compound of formula (I), and solid pharmaceutical compositions made from the nanosuspension. [Background technology]

[0002] The number of poorly water-soluble drug candidates emerging from drug discovery has increased dramatically over the past few decades. Their formulation into effective dosage forms presents a variety of challenges.

[0003] Methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate of formula (I), known from WO 2011 / 147809 [ka] (Vericiguat), is one of these active pharmaceutical ingredients (API) with limited dissolution behavior that affects bioavailability. WO 2011 / 147809 mentions solid and liquid formulations, including suspensions, for oral administration. WO 2011 / 147809 does not address the topic of the limited dissolution behavior of the compound of formula (I), nor any means for increasing its dissolution. Furthermore, WO 2011 / 147809 does not disclose crystalline forms of the compound of formula (I).

[0004] WO 2013 / 076168 relates, inter alia, to a process for producing the compound of formula (I) and intermediates used in this process. WO 2013 / 076168 further relates to the compound of formula (I) in the crystalline form of modification I and to the crystalline compound of formula (I) in the form of a didimethyl sulfoxide solvate of the compound of formula (I). The didimethyl sulfoxide solvate of the compound of formula (I) is used as an intermediate in a process for obtaining the compound of formula (I) in the crystalline form of modification I with high purity. WO 2013 / 076168 further mentions the stability of the compound of formula (I) in the crystalline form of modification I during storage in micronization, meaning that transformation and recrystallization do not occur. WO 2013 / 076168 does not address the topic of the limited solubility behavior of the compound of formula (I), nor any means for increasing its solubility.

[0005] The publication by Follmann et al. (2017) relates to the discovery of the compound of formula (I) for the treatment of heart failure. Follmann et al. describe process steps for producing the compound of formula (I) in crystalline form of modification I, including a final step as described in Example 13 of WO 2013 / 076168, which uses a didimethyl sulfoxide solvate of the compound of formula (I) as an intermediate. This process yields a highly purified compound of formula (I) in crystalline form of modification I in a dry form.

[0006] WO 2020 / 126983, published after the first filing date of this application, relates to an active compound product of the compound of formula (I) in the crystalline form of modification I, which has improved properties, for example, with respect to the isolation of the active compound product, the discharge of the active compound product after isolation and drying, and the transportability, sieving, and pulverizability of the active compound product. These improved properties make it possible to carry out the production process on a technical scale. Improved micronization is said to be measurable, for example, by easier feeding of the active compound product into a jet mill. WO 2020 / 126983 further mentions that micronization is achieved, for example, by grinding in a jet mill. Jet mills perform dry grinding. Jet mills are suitable for grinding particles down to the micrometer range. The grinding action in a jet mill is created by high-speed collisions between particles driven by multiple jets of air or steam. Dry grinding is not suitable for producing nanoparticles. WO 2020 / 126983 does not address the topic of the limited solubility behavior of the compound of formula (I), nor any measures to increase its solubility.

[0007] WO 2020 / 014504 relates to the use of sGC stimulators, including vericiguat, for the treatment of mitochondrial disorders. WO 2020 / 014504 also generally relates to formulations of sGC stimulators, specifically mentioning stabilizers and nanoparticles. However, WO 2020 / 014504 does not provide any specific teachings on stable nanoformulations of vericiguat or methods for preparing them, except for the general statement that the formulations can be prepared using conventional dissolution and mixing procedures.

[0008] Chinese Patent No. 108721296 relates to the use of vericiguat for treating altitude sickness. In Example 1, Chinese Patent No. 108721296 describes a specific composition of solid vericiguat, but does not disclose crystalline forms or nanoformulations of vericiguat.

[0009] The anticipated clinical dose of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate (Vericiguat, compound of Formula (I)) ranges from 2.5 to 15 mg once daily, depending on the target indication. A standard immediate-release tablet formulation has been developed. As the dose increases, a favorable food effect is observed, manifested as decreased bioavailability with standard immediate-release formulations such as tablets and increased bioavailability with food intake.

[0010] Food can affect the pharmacokinetics of drug products through several mechanisms, including delayed gastric emptying, stimulation of bile flow, changes in gastrointestinal (GI) pH, alterations in luminal metabolism, or interactions between the drug and the food itself. The drug absorption process can be influenced by many factors, including calorie content (low-calorie vs. high-calorie meals), nutritional composition (protein-, carbohydrate-, or high-fat meals), quantity, temperature of the meal itself, and fluid intake. Food also increases blood flow to the liver (splanchnic blood flow). Therefore, the resulting changes in first-pass extraction can cause differences in bioavailability between the fed and fasted states.

[0011] The preparation of drug nanoparticles or nanocrystals is one way to formulate drugs with limited dissolution behavior, because reducing the size of drug crystals increases their specific surface area, which can improve the dissolution rate and, therefore, their bioavailability. Furthermore, ultrafine particles tend to exhibit higher saturation solubilities, which also enhance the dissolution rate.

[0012] Within the meaning of the present invention, a sufficient dissolution rate is defined as the capacity in which sufficient drug particles in the envisaged dose range can be dissolved within the transit time at the absorption site. In contrast, a drug has a limited dissolution rate if the dissolution rate is too slow for all drug particles to dissolve during the time it takes to pass through the absorption site (Butler, Dressman, 2010).

[0013] There are several approaches to producing drug nanoparticles, such as wet bead milling (also called nanomilling in the context of nanosuspension preparation), homogenization, liquid antisolvent precipitation, melt emulsification, precipitation using supercritical fluids, evaporative precipitation, and microemulsions.

[0014] The particle size during milling generally depends on (i) process equipment parameters, including specific energy input and stress intensity; (ii) the mechanical and physicochemical properties of the drug particles; and (iii) the physical stability of the milled suspension, i.e., the mitigation of aggregation and / or Ostwald ripening in the presence of various stabilizers. "Stress intensity (SI)" is defined according to Kwade et al. (1996) as follows:

number

[0015] "Ostwald ripening" describes the phenomenon in which smaller particles in solution dissolve and deposit on larger particles, minimizing the surface area ratio and achieving a more thermodynamically stable state, promoting the formation of larger particles. The preparation of drug nanosuspensions with desired particle size and sufficient storage stability requires a wet bead milling (or nanomilling) process, as defined below and described in Examples 1-3. Selecting the optimal stabilizer blend is a laborious and resource-intensive task, but it is a critical one with potentially serious consequences. Poorly formulated nanosuspensions of drugs can suffer from aggregation, Ostwald ripening, rapid particle settling, and cake formation during milling and storage, which can lead to various problems in downstream processing of the respective suspensions and poor product performance of the final dose, such as unexpectedly slow dissolution rates.

[0016] According to Kwade et al. (1996), for a fixed specific energy input there is an optimum stress intensity for which the finest product is achieved. As the specific energy input, and therefore the fineness of the product, increases, the optimum stress intensity decreases. Since specific energy is proportional to the product of stress intensity and stress frequency, comminution results can also be correlated to stress frequency and stress intensity. As stress intensity increases, the stress frequency required for a given product fineness decreases.

[0017] Potential particle size increase or particle growth during milling and storage can result in loss of the high surface area associated with drug nanoparticles, which reduces the significant benefits intended from the nanomilling process.

[0018] Stabilization of nanoparticles in solution is always necessary to utilize their specific properties. Aggregation, growth, or fusion must be avoided over time. Due to their high specific surface area, nanoparticles are always in a "high-energy state" and therefore inherently unstable. Therefore, special measures must be taken to stabilize nanoparticle suspensions.

[0019] Stabilization of drug nanosuspensions produced by wet bead milling can be achieved with a variety of excipients, depending on the underlying stabilization mechanism, which is unpredictable. Prior art data suggests that wet bead milling has been used effectively to prepare nanosuspensions of numerous poorly water-soluble drugs, and that various polymers and / or surfactants can be used to ensure adequate physical stability of the nanosuspension. Interestingly, only a few nanosuspensions have achieved final drug particle sizes below 100 nm. Thus, there is a significant gap in the pharmaceutical nanotechnology literature regarding the preparation of drug nanoparticles by wet bead milling.

[0020] As outlined in a review by Li et al. (2016), the preparation of drug nanoparticles by wet bead milling (nanomiruing) is a well-known and established technique for oral and other forms of application. According to Li et al. (2016), wet bead milling followed by various drying processes has become an established and proven formulation approach for enhancing the bioavailability of drugs, particularly poorly water-soluble drugs. Although the physical stability of wet-milled suspensions (nanosuspensions) has received significant attention, a fundamental understanding of the process remains lacking. The selection of surfactants and their optimal concentrations is crucial for physical instability, a tedious and resource-intensive task. A first-principles predictive method for selecting the appropriate stabilizer or stabilizers for a given drug remains lacking. It can be concluded that no correlation exists between physicochemical drug properties (molecular weight, melting point, logP, solubility, and density) and the formation of stable nanosuspensions. Insufficient concentrations of stabilizers, such as polymers or surfactants, may not prevent aggregation of drug nanoparticles, but excessive amounts (especially when the surfactant concentration exceeds the critical micelle concentration (CMC)) are thought to accelerate Ostwald ripening. Changes in the solid state during milling can affect bioavailability and further manufacturing. Also, cross-linking from one mill to another can be important.

[0021] (2008) point out the impact and importance of identifying the appropriate stabilizer or stabilizers and of process parameters such as the effect of the number of homogenization cycles on particle size and the order of mixing of components on the physical properties of the nanosuspension. George & Gosh (2013) analyzed the mechanism of stabilization as a function of drug properties and concluded that optimizing nanosuspensions using a media milling approach is a complex process as it involves many factors that affect the properties of the nanosuspension product.

[0022] According to Desai (2012), the main challenges in designing nanosuspensions for oral delivery are maintaining the colloidal stability and particle size of the nanosuspension during storage and in the gastrointestinal tract after oral administration, as well as converting the nanosuspension into a palatable and patient-friendly oral formulation.

[0023] Publications by Choi & Han (2018) and Jermain et al. (2018) provide recent updates on nanocrystal technology for poorly water-soluble drugs. As summarized by Jermain et al. (2018), nanoparticles are much more unstable than microparticles due to the extra Gibbs free energy contribution associated with particle size reduction, primarily due to increased surface energy. Addressing this extra contribution is important for the formulation of pharmaceutical nanoparticles because pharmaceutical nanoparticles tend to aggregate to minimize their total energy (Van Eerdenbrugh et al., 2008). As outlined by Jermain et al. (2018), thermodynamic stabilization can be used as a method to stabilize drug nanoparticles using surfactants or block copolymers for particle stability. For maximum effectiveness, the two methods are often combined (Lee et al., 2008). Careful selection of the amount of stabilizer is just as important as the type of stabilizer. For example, one obstacle to stabilization is Ostwald ripening, as mentioned above. Too little stabilizer will result in aggregation of nanoparticles, while too much stabilizer will promote Ostwald ripening (Merisko-Liversidge et al., 2003).

[0024] Several commercial products based on nanoformulations produced via wet bead milling are known (table revised after the book "Nanomedicine in Health and Disease" (Hunter, 2011):

[0025] [Table 1]

[0026] As outlined above, common technical issues regarding nanoformulations that must be resolved for each individual compound include: a) Selection of stabilizers b) Mechanical processing: influence of process parameters (choice of mill, specific energy input and stress intensity, duration of grinding, tip speed, etc.), material of grinding beads, size of grinding beads, volumetric filling of grinding beads, and physicochemical properties of the drug. c) Further processing: risk of loss of beneficial effect due to nanosize upon drying of the nanosuspension, for example due to aggregation or Ostwald ripening, or further processing prior to storage of the final dosage form, and resuspension at the absorption site. [Prior art documents] [Patent documents]

[0027] [Patent Document 1] International Publication No. 2011 / 147809 [Patent Document 2] International Publication No. 2013 / 076168 [Patent Document 3] International Publication No. 2020 / 126983 [Patent Document 4] International Publication No. 2020 / 014504 [Patent Document 5] Chinese Patent No. 108721296 [Patent Document 6] U.S. Patent No. 5,145,684 [Patent Document 7] U.S. Patent No. 8,258,132 [Patent Document 8] U.S. Patent No. 6,375,986 [Patent Document 9] U.S. Patent No. 7,276,249 [Patent Document 10] U.S. Patent No. 7,320,802 [Patent Document 11] U.S. Patent No. 6,592,903 [Patent Document 12] U.S. Patent No. 9,101,540 Summary of the Invention [Problem to be solved by the invention]

[0028] In view of the prior art, one object of the present invention is to solve the problem that due to the limited dissolution behavior of vericiguat, its bioavailability in preclinical and clinical studies may decrease at higher doses. [Means for solving the problem]

[0029] This object of the present invention is solved by providing a stable nanosuspension of the compound of formula (I) according to the present invention, a process for preparing the stable nanosuspension, nanoparticles comprising the compound of formula (I), and pharmaceutical compositions in solid form made from the nanosuspension, which are expected to generally increase bioavailability, which is limited by a low dissolution rate, and to avoid the food effect.

[0030] According to one embodiment, the present invention provides a stable nanosuspension of the compound of formula (I) (vericiguat), for which no stable nanoformulation has been previously described.

[0031] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a dispersing agent, the maximum concentration of the stabilizer(s) being the solubility limit of the stabilizer(s) in the suspension, said nanoparticles having an average particle size, expressed as d50, of less than or equal to 500 nm.

[0032] Within the meaning of the present invention, the compound of formula (I) is methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate. [ka]

[0033] Within the meaning of the present invention, a "compound of formula (I) in crystalline form of modification (I)" is understood to mean, for example, a compound of formula (I) having defined peak maxima in 2θ angles at 5.9, 6.9 and 22.7 or at 5.9, 6.9, 16.2, 16.5, 24.1, 22.7 and 24.7; or at 1707, 1633 and 1475 cm -1 or 1707, 1633, 1566, 1475, 1255 and 1223 cm -1 or with the aid of a melting point of 257 °C.

[0034] Within the meaning of the present invention, "drug nanoparticles" or "nanoparticles of the compound of formula (I) in the crystalline form of modification I" are defined as particles of an active pharmaceutical ingredient (API) or particles of the compound of formula (I) in the crystalline form of modification I, respectively, having a particle size, expressed as d50, of 500 nm or less, or 400 nm or less, or 300 nm or less, or 250 nm or less, or 200 nm or less, or 150 nm or less, or 100 nm or less.

[0035] Within the meaning of the present invention, particle size distribution is expressed as d value. d value can be considered as the diameter of a sphere that divides the sample mass into a specified percentage when the particles are arranged in ascending order of mass. For example, d10 is the diameter of particles whose diameter is less than this value, which makes up 10% of the sample mass. d50 is the diameter of particles whose mass is smaller than this value and whose mass is larger than this value, which makes up 50% of the sample mass. d90 represents the diameter of particles whose mass is smaller than this value and whose mass is larger than this value, which makes up 90% of the sample mass. The size of drug nanoparticles can be measured, for example, by dynamic light scattering (DLS) using Zetasizer Nano-ZS (Malvern Panalytical) or static light scattering (SLS) using Mastersizer 3000 (Malvern Panalytical).

[0036] A "stable nanosuspension" within the meaning of the present invention is defined as a nanosuspension comprising nanoparticles of the compound of formula (I) in crystalline form of modification I and one or more stabilizers in a dispersing agent, said nanoparticles having an average particle size, expressed as d50, of 500 nm or less, or 300 nm or less, or 250 nm or less, or 200 nm or less, or 150 nm or less, or 100 nm or less, and wherein the average particle size, expressed as d50, remains 500 nm or less, or 300 nm or less, or 250 nm or less, or 200 nm or less, or 150 nm or less, or 100 nm or less upon storage for at least one week at a temperature of at least 40°C.

[0037] Within the meaning of the present invention, a "stable nanosuspension" is further defined as a nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a ratio of 8:1 to 1:1 w / w, said nanoparticles having an average particle size, expressed as d50, of 500 nm or less, said particle size of the nanoparticles, measured either by DLS or SLS, remaining 500 nm or less when stored at 40° C. for one week or more. Long-term measurements of particle growth are shown in Example 2b and in FIG. 2.

[0038] Within the meaning of the present invention, "stabilizers" are defined as substances and combinations thereof that improve grinding properties, cause surface wetting and surface stabilization, and provide physical stability of the nanosuspension.

[0039] The subject of the present disclosure is a stable nanosuspension, wherein the one or more stabilizers are selected from the group consisting of polyvinylpyrrolidone (PVP), vinylpyrrolidone-vinyl acetate copolymer, ethylene oxide-propylene oxide block copolymer, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), d-alpha tocopheryl polyethylene glycol 1000 succinate (vitamin E-TPGS), polysorbate, hydroxypropyl cellulose (HPC), polyoxyl-35 castor oil, polyoxyl-15 hydroxystearate, Na-desoxycholate, and combinations thereof.

[0040] The subject of the present disclosure is a stable nanosuspension, wherein the one or more stabilizers are selected from the group consisting of polyvinylpyrrolidone (PVP), vinylpyrrolidone-vinyl acetate copolymer (e.g., PVP VA 64), ethylene oxide-propylene oxide block copolymer (e.g., poloxamer 188), sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), d-alpha tocopheryl polyethylene glycol 1000 succinate (vitamin E-TPGS), polysorbate (Tween®), hydroxypropyl cellulose (HPC), polyoxyl-35 castor oil (Cremophor EL®), polyoxyl 15 hydroxystearate (Solutol HS 15®), Na-desoxycholate, and combinations thereof.

[0041] The subject of the present disclosure is a stable nanosuspension, wherein the one or more stabilizers are selected from the group consisting of sodium dodecyl sulfate, polyvinylpyrrolidone K10-K50, hydroxypropyl methylcellulose, vitamin E TPGS, polysorbate 20-80, and combinations thereof.

[0042] One embodiment of the present invention is a stable nanosuspension according to the present invention, wherein the one or more stabilizers are selected from the group consisting of polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), vinylpyrrolidone-vinyl acetate copolymer, ethylene oxide-propylene oxide block copolymer, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), polysorbate, hydroxypropyl cellulose (HPC), polyoxyl-35 castor oil, polyoxyl 15 hydroxystearate, Na-desoxycholate, and combinations thereof.

[0043] One embodiment of the present invention is a stable nanosuspension according to the present invention, wherein the one or more stabilizers are selected from the group consisting of sodium dodecyl sulfate, polyvinylpyrrolidone K10-K50 in combination with sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose, polysorbate 20-80, and combinations thereof.

[0044] According to one embodiment of the present invention, the polyvinylpyrrolidone (PVP) is selected from PVP K10 to K50. According to a further embodiment of the present invention, the polyvinylpyrrolidone (PVP) is selected from PVP K12 to K30. According to a further embodiment of the present invention, the polyvinylpyrrolidone (PVP) is selected from PVP K12, PVP K17, and PVP K30. According to one embodiment of the present invention, the vinylpyrrolidone-vinyl acetate copolymer is PVP VA64. According to one embodiment of the present invention, the ethylene oxide-propylene oxide block copolymer is poloxamer 188. According to one embodiment of the present invention, the polysorbat is selected from polysorbat 20 to 80. According to one embodiment of the present invention, the polysorbat is polysorbat 80.

[0045] According to one embodiment of the present invention, the one or more stabilizers contained in the nanoparticles according to the present invention are a combination of SDS and PVP K12. According to one embodiment of the present invention, the one or more stabilizers contained in the nanoparticles according to the present invention are a combination of SDS and PVP K17. According to one embodiment of the present invention, the one or more stabilizers contained in the nanoparticles according to the present invention are a combination of SDS and Poloxamer 188.

[0046] One embodiment of the present invention is a stable nanosuspension according to the present invention, wherein the one or more stabilizers is sodium dodecyl sulfate. One embodiment of the present invention is a stable nanosuspension according to the present invention, wherein the one or more stabilizers is polysorbate 20-80. One embodiment of the present invention is a stable nanosuspension according to the present invention, wherein the one or more stabilizers is polysorbate 80.

[0047] Within the meaning of the present invention, the maximum concentration of one or more stabilizers is the solubility limit of the stabilizer(s) in the suspension. In the case of pharmaceutical compositions prepared using the nanosuspension of the present invention, the maximum concentration of one or more stabilizers is further limited by the acceptable daily intake (ADI), if an ADI is defined for each stabilizer. According to the European Food Safety Authority, the ADI is an estimate of the amount of a substance in food or drinking water that can be ingested daily over a lifetime without significant health risks. It is usually expressed as milligrams of substance per kilogram of body weight and applies to chemicals such as food additives, pesticide residues, and veterinary drugs.

[0048] Within the meaning of the present invention, a "dispersant" is defined as a polar liquid in which the compound of formula (I) is insoluble. Examples include, but are not limited to, water; alcohols such as primary, secondary, and tertiary alcohols, including ethanol, propanol, isopropanol, butanol, isobutanol, and tert-butanol; and polyhydric alcohols, including glycerol. According to one embodiment of the present invention, water is used as the dispersant.

[0049] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a dispersing agent, the maximum concentration of the stabilizer(s) being the solubility limit of the stabilizer(s) in the suspension, said nanoparticles having an average particle size, expressed as d50, of less than or equal to 400 nm.

[0050] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a dispersing agent, the maximum concentration of the stabilizer(s) being the solubility limit of the stabilizer(s) in the suspension, said nanoparticles having an average particle size, expressed as d50, of less than or equal to 300 nm.

[0051] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a dispersing agent, the maximum concentration of the stabilizer(s) being the solubility limit of the stabilizer(s) in the suspension, said nanoparticles having an average particle size, expressed as d50, of less than or equal to 250 nm.

[0052] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a dispersing agent, the maximum concentration of the stabilizer(s) being the solubility limit of the stabilizer(s) in the suspension, said nanoparticles having an average particle size, expressed as d50, of less than or equal to 200 nm.

[0053] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a dispersing agent, the maximum concentration of the stabilizer(s) being the solubility limit of the stabilizer(s) in the suspension, said nanoparticles having an average particle size, expressed as d50, of less than or equal to 150 nm.

[0054] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a dispersing agent, the maximum concentration of the stabilizer(s) being the solubility limit of the stabilizer(s) in the suspension, said nanoparticles having an average particle size, expressed as d50, of less than or equal to 100 nm.

[0055] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of the compound of formula (I) in the crystalline form of modification I and one or more stabilizers in a dispersing agent, the maximum concentration of the one or more stabilizers being the solubility limit of the one or more stabilizers in the suspension, said nanoparticles having an average particle size, expressed as d50, of less than or equal to 500 nm, and a ratio of compound of formula (I) to one or more stabilizers of between 16:1 and 1:2 w / w.

[0056] When a combination of two stabilizers is used, and these two stabilizers are a surfactant and a polymer (e.g., SDS+polyvinylpyrrolidone or SDS+ethylene oxide-propylene oxide block copolymer), the ratio of Compound (I):polymer is 16:1 to 1:2 w / w or 8:1 to 2:1 w / w, and the concentration of the surfactant is 0.1 to 0.2% w / v.

[0057] One embodiment of the present invention is a stable nanosuspension of the present invention, wherein the mean particle size, expressed as d50, remains equal to or less than 300 nm when stored at a temperature of at least 40° C. for at least one week.

[0058] One embodiment of the present invention is a stable nanosuspension according to the invention, wherein one or more stabilizers are used in a ratio of compound of formula (I):stabilizer(s) of 8:1 to 2:1 w / w.

[0059] According to one embodiment of the present invention, the mean particle size, expressed as d50 and measured either by DLS or SLS, of a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a ratio of 8:1 to 1:1 w / w remains equal to or less than 300 nm upon storage at 40°C for at least one week.

[0060] According to one embodiment of the present invention, the mean particle size, expressed as d50 and measured either by DLS or SLS, of a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a ratio of 8:1 to 1:1 w / w remains equal to or less than 300 nm upon storage at 40°C for at least 2 weeks.

[0061] According to one embodiment of the present invention, the mean particle size, expressed as d50 and measured either by DLS or SLS, of a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a ratio of 8:1 to 1:1 w / w remains equal to or less than 300 nm upon storage at 40°C for at least 4 weeks.

[0062] According to one embodiment of the present invention, the mean particle size, expressed as d50 and measured either by DLS or SLS, of a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a ratio of 8:1 to 1:1 w / w remains equal to or less than 300 nm upon storage at 40°C for at least 8 weeks.

[0063] According to one embodiment of the present invention, the mean particle size, expressed as d50 and measured either by DLS or SLS, of a stable nanosuspension comprising nanoparticles of compound of formula (I) in crystalline form of modification I and one or more stabilizers in a ratio of 8:1 to 1:1 w / w remains equal to or less than 300 nm upon storage at 40°C for at least 13 weeks.

[0064] One embodiment of the present invention is methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate of formula (I) in the crystalline state of modification I [ka] and one or more stabilizers in a dispersing agent, wherein an X-ray diffractogram of the compound is characterized by exhibiting peak maxima at 2θ angles of 5.9, 6.9, and 22.7, and the one or more stabilizers are selected from the group consisting of polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), vinylpyrrolidone-vinyl acetate copolymer, ethylene oxide-propylene oxide block copolymer, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), polysorbate, hydroxypropyl cellulose, and the like. the maximum concentration of the one or more stabilizers is the solubility limit of the one or more stabilizers in the suspension; the dispersing agent is selected from the group consisting of water, primary, secondary and tertiary alcohols and polyhydric alcohols; and the nanoparticles have an average particle size, expressed as d50, of 500 nm or less and an average particle size, expressed as d50, that remains 500 nm or less when stored at a temperature of at least 40°C for at least one week.

[0065] One embodiment of the present invention is a stable nanosuspension according to the present invention, wherein the one or more stabilizers are selected from the group consisting of sodium dodecyl sulfate, polyvinylpyrrolidone K10-K50 in combination with sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose, polysorbate 20-80, and combinations thereof.

[0066] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate of formula (I) in the crystalline form of modification I and one or more stabilizers in a dispersing agent, wherein the X-ray diffractogram of said compound is characterized in that it exhibits peak maxima at 2θ angles of 5.9, 6.9 and 22.7, and the one or more stabilizers are selected from the group consisting of sodium dodecyl sulfate, polyisoprene, sodium dodecyl sulfate (SDS), and methyl 4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate. the maximum concentration of the one or more stabilizers is the solubility limit of the one or more stabilizers in the suspension; the dispersing agent is selected from the group consisting of water, primary, secondary and tertiary alcohols and polyhydric alcohols; and the nanoparticles have an average particle size, expressed as d50, of 500 nm or less and an average particle size, expressed as d50, that remains 500 nm or less when stored at a temperature of at least 40°C for at least one week.

[0067] One embodiment of the present invention is a stable nanosuspension according to the present invention, in which the ratio of the compound of formula (I) to one or more stabilizers is 16:1 to 1:2 w / w. One embodiment of the present invention is a stable nanosuspension according to the present invention, in which the ratio of the compound of formula (I) to one or more stabilizers is 8:1 to 2:1 w / w. When a combination of two stabilizers is used, and these two stabilizers are a surfactant and a polymer (e.g., SDS + polyvinylpyrrolidone or SDS + ethylene oxide-propylene oxide block copolymer), the ratio of compound (I) to polymer is 16:1 to 1:2 w / w or 8:1 to 2:1 w / w, and the concentration of the surfactant is 0.1 to 0.2% w / v.

[0068] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate of formula (I) in the crystalline form of modification I and one or more stabilizers in a dispersing agent, characterized in that the X-ray diffractogram of said compound exhibits peak maxima at 2θ angles of 5.9, 6.9 and 22.7, and the one or more stabilizers are selected from the group consisting of polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), vinylpyrrolidone-vinyl acetate copolymer, ethylene oxide-propylene oxide block copolymer, sodium dodecyl sulfate (SDS), hydroxybenzoates, ... the maximum concentration of the one or more stabilizers is the solubility limit of the one or more stabilizers in the suspension; the dispersing agent is selected from the group consisting of water, primary, secondary, and tertiary alcohols, and polyhydric alcohols; the nanoparticles have an average particle size, expressed as d50, of 500 nm or less, an average particle size, expressed as d50, that remains 500 nm or less upon storage at a temperature of at least 40°C for at least one week; and the ratio of the compound of formula (I) to the one or more stabilizers is 16:1 to 1:2 w / w.

[0069] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate of formula (I) in the crystalline form of modification I and one or more stabilizers in a dispersing agent, wherein the X-ray diffractogram of said compound is characterized in that it exhibits peak maxima at 2θ angles of 5.9, 6.9 and 22.7, and the one or more stabilizers are polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS) or ethylene oxide in combination with SDS. the maximum concentration of the one or more stabilizers is the solubility limit of the one or more stabilizers in the suspension; the dispersing agent is selected from the group consisting of water, primary, secondary and tertiary alcohols and polyhydric alcohols; the nanoparticles have an average particle size expressed as d50 of 500 nm or less, an average particle size expressed as d50 that remains 500 nm or less upon storage for at least one week at a temperature of at least 40°C; the ratio of compound of formula (I) to polyvinylpyrrolidone or ethylene oxide-propylene oxide block copolymer is 16:1 to 1:2 w / w or 8:1 to 2:1 w / w, respectively; and the concentration of SDS is 0.1 to 0.2% w / v.

[0070] One embodiment of the present invention is a stable nanosuspension according to the invention, wherein the nanoparticles have an average particle size, expressed as d50, of 500 nm or less, or 300 nm or less, or 250 nm or less, or 200 nm or less, or 150 nm or less, or 100 nm or less.

[0071] One embodiment of the present invention is a stable nanosuspension according to the invention, wherein the nanoparticles have a mean particle size, expressed as d50, of less than or equal to 300 nm.

[0072] One embodiment of the present invention is a stable nanosuspension according to the invention, wherein the mean particle size, expressed as d50, remains equal to or less than 300 nm when stored at a temperature of at least 40° C. for at least one week.

[0073] One embodiment of the present invention is a stable nanosuspension comprising nanoparticles of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate of formula (I) in the crystalline form of modification I and one or more stabilizers in a dispersing agent, wherein the X-ray diffractogram of said compound is characterized in that it exhibits peak maxima at 2θ angles of 5.9, 6.9 and 22.7, and the one or more stabilizers are selected from the group consisting of polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), vinylpyrrolidone-vinyl acetate copolymer, ethylene oxide-propylene oxide block copolymer, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), polysorbate, hydroxypropyl methylcellulose (HPMC), ... the maximum concentration of the one or more stabilizers is the solubility limit of the one or more stabilizers in the suspension; the dispersing agent is selected from the group consisting of water, primary, secondary and tertiary alcohols and polyhydric alcohols; and the nanoparticles have an average particle size, expressed as d50, of 500 nm or less, and an average particle size, expressed as d50, that remains 500 nm or less, or 300 nm or less, or 250 nm or less, or 200 nm or less, or 150 nm or less, or 100 nm or less upon storage at a temperature of at least 40°C for at least 1 week, or at least 2 weeks, or at least 4 weeks, or at least 8 weeks, or at least 13 weeks.

[0074] To obtain stable nanosuspensions according to the present invention, it was necessary to identify specific stabilizers and combinations of stabilizers (Figure 1). This work is highly dependent on the specific compounds. Figure 2 illustrates the long-term stability of the nanosuspensions of the present invention, which is a prerequisite for their direct use and processing into further pharmaceutical forms such as granules and tablets. Contrary to the prior art teaching that stabilizer concentrations above the CMC promote Ostwald ripening, it was surprisingly found that stable nanosuspensions can be obtained at concentrations of at least the stabilizer SDS four times above the CMC (Figure 3, API:SDS = 2:1). As shown in Figure 4, the particle size of micronized vericiguat after nanomilling is significantly smaller than that of vericiguat that is not micronized.

[0075] In order to obtain a stable nanosuspension, the wet bead milling parameters had to be carefully selected in addition to the choice and concentration of the stabilizer(s).

[0076] One embodiment of the present invention is a process for preparing a stable nanosuspension according to the present invention, comprising: a. suspending the compound of formula (I) in crystalline form of modification I in a dispersing agent and one or more stabilizers according to the invention, the maximum concentration of the stabilizer(s) being the solubility limit of the stabilizer(s) in the suspension; b. Specific energy input of 10,000 kJ / kg or more and 0.004 10 -3 Nm~1·10 -3 wet bead milling the suspension produced in step a at a stress intensity of 1000 Nm; It is a process including:

[0077] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, wherein the dispersant is selected from the group consisting of water, primary, secondary, and tertiary alcohols, and polyhydric alcohols.

[0078] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, wherein the one or more stabilizers are selected from the group consisting of polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), vinylpyrrolidone-vinyl acetate copolymer, ethylene oxide-propylene oxide block copolymer, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), polysorbate, hydroxypropyl cellulose (HPC), polyoxyl-35 castor oil, polyoxyl 15 hydroxystearate, Na-desoxycholate, and combinations thereof.

[0079] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, wherein the one or more stabilizers are selected from the group consisting of sodium dodecyl sulfate, polyvinylpyrrolidone K10-K50 in combination with sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose, polysorbate 20-80, and combinations thereof.

[0080] One embodiment of the present invention is a process for preparing a stable nanosuspension according to the present invention, comprising: a. suspending the compound of formula (I) in crystalline form of modification I in a dispersing agent selected from the group consisting of water, primary, secondary, and tertiary alcohols, and polyhydric alcohols, and one or more stabilizers selected from the group consisting of polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), vinylpyrrolidone-vinyl acetate copolymers, ethylene oxide-propylene oxide block copolymers, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), polysorbates, hydroxypropyl cellulose (HPC), polyoxyl-35 castor oil, polyoxyl 15 hydroxystearate, Na-desoxycholate, and combinations thereof, wherein the maximum concentration of the one or more stabilizers is the solubility limit of the one or more stabilizers in the suspension; b. Specific energy input of 10,000 kJ / kg or more and 0.004 10-3 Nm~1·10 -3 wet bead milling the suspension produced in step a at a stress intensity of 1000 Nm; It is a process including:

[0081] Within the meaning of the present invention, "wet bead milling" is used synonymously with "wet ball milling" and also with "wet milling". The capacity and fineness can be adjusted by adjusting the diameter of the balls / beads.

[0082] To obtain nanoformulations according to the present invention, milling parameters including milling bead size, milling bead material, tip speed, milling duration, and volume filling by milling beads were optimized.

[0083] Within the meaning of the present invention, the term "milling" is used synonymously with the term "grinding".

[0084] To obtain nanoformulations according to the invention, a sufficient specific energy input and stress intensity are required. The specific energy input and / or stress intensity is sufficient if the desired particle size is reached with sufficient stabilization. If the particle size is too high, either the specific energy input and / or stress intensity and / or stabilization is not sufficient.

[0085] Within the meaning of the present invention, specific energy input is defined as the net energy input related to the product mass.

[0086] Within the meaning of the present invention, stress intensity is defined according to Kwade et al. (1996). According to Kwade, for a fixed specific energy input, there exists an optimum stress intensity for which the finest product is achieved. As the specific energy input increases, and thus the fineness of the product, increases, the optimum stress intensity decreases. For a constant specific energy input, the stress intensity determines the fineness of the product.

[0087] Within the meaning of the present invention, "wet bead milling" or "nanomiruing" is defined as the preparation of drug nanoparticles by wet media milling (nanomiruing). As known to those skilled in the art, milling beads are used in "wet bead milling" or "nanomiruing". The basic principle behind nanomilling is to increase the surface area-to-volume ratio of the API by reducing the particle size, measured as d90, to less than 800 nm, typically in the range of several hundred to several hundred nm, measured as d50. This conversion of drug particles into nanocrystals allows for greater interaction with water, increasing the dissolution rate. Generally speaking, smaller particles dissolve more quickly.

[0088] According to one embodiment of the present invention, the wet bead milling of the suspension is carried out in the presence of grinding beads having a size of 0.05 to 0.8 mm. According to one embodiment of the present invention, the grinding beads are made of a material selected from the group consisting of ceramic, glass, polymer, and steel. Examples of ceramics within the meaning of the present invention include, for example, sintered or fused yttrium-stabilized zirconium oxide, cerium-stabilized zirconium oxide, and zirconium silicate. Examples of glasses include zirconia-doped glasses. Examples of polymers include polystyrene. According to one embodiment of the present invention, the grinding beads are made of yttrium-stabilized zirconium oxide. According to one embodiment of the present invention, the volumetric filling of the grinding beads is 40 to 85% v / v. According to one embodiment of the present invention, the volumetric filling of the grinding beads is 50 to 85% v / v. According to one embodiment of the present invention, the volumetric filling of the grinding beads is 60 to 85% v / v. According to one embodiment of the present invention, the volumetric filling of the grinding beads is 60 to 80% v / v.

[0089] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, wherein step b. is carried out in the presence of grinding beads having a size of 0.05 to 0.8 mm.

[0090] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, wherein step b. is carried out in the presence of grinding beads made from a material selected from the group selected from ceramic, glass, polymer and steel.

[0091] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, wherein the volume filling with beads in step b. is 50-85% v / v.

[0092] Specific energy input of 10,000 kJ / kg or more and 0.004·10 -3 Nm~1·10 -3 The stress intensity in Nm is related to, for example, the tip speed and the duration of the crushing.

[0093] According to one embodiment of the present invention, the specific energy input is 10,000 kJ / kg or more, and the stress intensity is 0.004·10 -3 Nm~1·10 -3 In the case of a specific energy input of 10,000 kJ / kg or more and a stress intensity of 0.004 10 -3 Nm~1·10 -3 In the case of a specific energy input of 10,000 kJ / kg or more and a stress intensity of 0.004 10 -3 Nm~1·10 -3 In the case of a specific energy input of 10,000 kJ / kg or more and a stress intensity of 0.004 10 -3 Nm~1·10 -3 If the cutting speed is 8 to 15 m / s, the grinding time is 30 minutes to 2 hours.

[0094] According to one embodiment of the present invention, the grinding beads are made of yttrium-stabilized zirconium oxide, the grinding beads have a size of 0.05-0.6 mm, the volume filling by the grinding beads is 60-80%, the tip speed is 12-15 m / s, and the grinding time is 60 minutes.

[0095] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, wherein step b. is carried out in the presence of grinding beads having a size of 0.05-0.8 mm made from a material selected from the group consisting of ceramic, glass, polymer, and steel, and the volume filling by the beads in step b. is 50-85% v / v.

[0096] One embodiment of the present invention includes an additional step prior to wet bead nano-milling, which is carried out to break down coarse particles and improve the results of the subsequent nano-milling step. Coarse particles can be broken down, for example, by micronization or by wet bead milling using beads with a size of 1-2 mm.

[0097] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, wherein in step a., the compound of formula (I) in crystalline form of modification I is micronized in a first step before suspending it in a dispersing agent and one or more stabilizers.

[0098] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, wherein in step a, the compound of formula (I) in crystalline form of modification I is milled in a first wet bead milling step in the presence of milling beads having a size of 1-2 mm before suspending it in a dispersing agent and one or more stabilizers.

[0099] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, comprising: a. micronizing in a first step, prior to suspending the compound of formula (I) in crystalline form of modification I in a dispersing agent selected from the group consisting of water, primary, secondary, and tertiary alcohols, and polyhydric alcohols, and one or more stabilizers selected from the group consisting of polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), vinylpyrrolidone-vinyl acetate copolymers, ethylene oxide-propylene oxide block copolymers, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), polysorbates, hydroxypropyl cellulose (HPC), polyoxyl-35 castor oil, polyoxyl 15 hydroxystearate, Na-desoxycholate, and combinations thereof, wherein the maximum concentration of the stabilizer(s) is the solubility limit of the stabilizer(s) in the suspension; b. Wet bead milling the suspension produced in step a with a specific energy input of 10,000 kJ / kg or more and a stress intensity of 0.004·10-3 Nm to 1·10-3 Nm, wherein the milling beads used have a size of 0.05 to 0.8 mm. It is a process including:

[0100] One embodiment of the present invention is a process for preparing a stable nanosuspension according to any embodiment of the present invention, comprising: a. grinding the compound of formula (I) in crystalline form of modification I in a first wet bead milling step in the presence of milling beads having a size of 1 to 2 mm, before suspending it in a dispersing agent selected from the group consisting of water, primary, secondary, and tertiary alcohols, and polyhydric alcohols, and one or more stabilizers selected from the group consisting of polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), vinylpyrrolidone-vinyl acetate copolymers, ethylene oxide-propylene oxide block copolymers, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), polysorbates, hydroxypropyl cellulose (HPC), polyoxyl-35 castor oil, polyoxyl-15 hydroxystearate, Na-desoxycholate, and combinations thereof, wherein the maximum concentration of the stabilizer(s) is the solubility limit of the stabilizer(s) in the suspension; b. wet bead milling the suspension produced in step a with a specific energy input of 10,000 kJ / kg or more and a stress intensity of 0.004 10-3 Nm to 1 10-3 Nm, wherein the milling beads used have a size of 0.05 to 0.8 mm; It is a process including:

[0101] One embodiment of the present invention is a process for preparing a stable nanosuspension according to the present invention, comprising: a. suspending the compound of formula (I) in crystalline form of modification I in a dispersing agent and one or more stabilizers according to the invention, the maximum concentration of the stabilizer(s) being the solubility limit of the stabilizer(s) in the suspension; b. Specific energy input of 15,000 kJ / kg or more, or 20,000 kJ / kg or more, or 25,000 kJ / kg or more and 0.004 10 -3 Nm~1·10 -3 wet bead milling the suspension produced in step a at a stress intensity of 1000 Nm; It is a process including:

[0102] One embodiment of the present invention is a process for preparing a stable nanosuspension according to the present invention, comprising: a. suspending the compound of formula (I) in crystalline form of modification I in a dispersing agent selected from the group consisting of water, primary, secondary, and tertiary alcohols, and polyhydric alcohols, and one or more stabilizers selected from the group consisting of polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), vinylpyrrolidone-vinyl acetate copolymers, ethylene oxide-propylene oxide block copolymers, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), polysorbates, hydroxypropyl cellulose (HPC), polyoxyl-35 castor oil, polyoxyl 15 hydroxystearate, Na-desoxycholate, and combinations thereof, wherein the maximum concentration of the one or more stabilizers is the solubility limit of the one or more stabilizers in the suspension; b. wet bead milling the suspension produced in step a with a specific energy input of 10,000 kJ / kg or more, or 15,000 kJ / kg or more, or 20,000 kJ / kg or more, or 25,000 kJ / kg or more and a stress intensity of 0.004 10-3 Nm to 1 10-3 Nm, wherein the milling beads used have a size of 0.05 to 0.8 mm; It is a process including:

[0103] It is known that reducing the particle size of an API can increase the dissolution rate, thus leading to higher bioavailability and potentially affecting the degree of food effect. However, selecting a process for reducing particle size, selecting parameters for process equipment, i.e., preparing stable nanosuspensions and stable nanoparticles, and selecting the optimal stabilizer for drug nanosuspensions are difficult and not obvious in light of the prior art.

[0104] The upper limit of the specific energy input is related to the degree of amorphization of the compound of formula (I) caused by the milling process. Complete amorphization should be avoided because in that case the crystalline structure of the compound of formula (I) is totally lost and recrystallization occurs in an uncontrolled manner. As shown in Figure 5a-c, surprisingly, amorphization of compound (I) in the crystalline modification I does not occur during the wet bead nanomilling process.

[0105] The choice of mill is crucial for producing nanoparticles. Nanomilling must be carried out as wet bead milling, for example in a planetary mill or stirred media mill. In contrast, jet mills (another type of mill used for micronization) perform dry milling. Jet mills are suitable for grinding particles down to the micrometer range. The grinding action in jet mills is created by high-velocity collisions between particles driven by multiple jets of air or steam. Dry milling is not suitable for producing nanoparticles.

[0106] Subject of the present disclosure are particles comprising the compound of formula (I) in crystalline form of modification I and one or more stabilizers, in a ratio of compound of formula (I):one or more stabilizers of between 16:1 and 1:2 w / w, said particles having a mean particle size, expressed as d50, in dry form of less than or equal to 500 nm.

[0107] One embodiment of the present invention is dry nanoparticles comprising the compound of formula (I) in crystalline form of modification I and one or more stabilizers, in a ratio of compound of formula (I):one or more stabilizers of between 16:1 and 1:2 w / w, and having a mean particle size, expressed as d50, of less than or equal to 500 nm.

[0108] One embodiment of the present invention is dry nanoparticles comprising the compound of formula (I) in crystalline form of modification I and a combination of two stabilizers, PVP and SDS, in a ratio of compound of formula (I) to PVP of 16:1 to 1:2 w / w and a concentration of SDS of 0.1 to 0.2% w / v, the nanoparticles having a mean particle size, expressed as d50, of less than or equal to 500 nm.

[0109] One embodiment of the present invention is dry nanoparticles comprising the compound of formula (I) in crystalline form of modification I and a combination of two stabilizers, an ethylene oxide-propylene oxide block copolymer and SDS, in a ratio of compound of formula (I) to ethylene oxide-propylene oxide block copolymer of 16:1 to 1:2 w / w and a concentration of SDS of 0.1 to 0.2% w / v, the nanoparticles having an average particle size, expressed as d50, of less than or equal to 500 nm.

[0110] Within the meaning of the present invention, the terms "dry" or "dry form", as used for example in the terms "dry nanoformulation" or "(nano)particles in dry form" or "(nano)particles in dry form having an average particle size expressed as d50 of 500 nm or less", are defined as a nanosuspension according to the invention that has been transferred to a solid state by a drying step. Dried nanoformulations or nanoparticles still contain one or more stabilizers but no longer contain a dispersant. The removal of the dispersant is achieved by carrying out a drying step.

[0111] The subject of the present disclosure is a pharmaceutical composition in solid form prepared using the nanosuspension of the present invention in dry form, which is prepared by a pharmaceutical process including compression and coating, optionally including granulation.

[0112] The subject of the present disclosure is a pharmaceutical composition in solid form prepared using the nanosuspension of the present invention in dry form, which is prepared by pharmaceutical processes including granulation, compression and coating.

[0113] According to one embodiment of the present invention, drying of the nanosuspension according to the invention can be carried out by spray drying or by concomitant granulation by fluidized bed granulation.

[0114] The production of solid dosage forms can be carried out by wet granulation processes (high shear granulation or fluidized bed granulation). High shear granulation is a compacting process for wet granulation. A binder liquid is fed to powder particles in a closed container using mixing tools and a chopper. Dense granules are formed through the resulting liquid-solid bridges. Fluidized bed granulation is also a wet granulation process that involves adding a binder liquid to primary particles to form agglomerated granules. The particles are fluidized from below while the binder liquid is sprayed from above.

[0115] According to one embodiment of the present invention, top spray granulation is used in fluid bed granulation.

[0116] Tableting is preferably carried out using the granules initially produced, after which the solid dosage form may be coated.

[0117] In wet granulation, the active compound product is suspended in a granulation liquid. The granulation liquid used contains a solvent, a hydrophilic binder, and a wetting agent. The hydrophilic binder is dispersed in the granulation fluid or, preferably, dissolved therein. Solvents that can be used in the granulation liquid include organic solvents such as ethanol or acetone, or water or a mixture thereof. Water is preferred as the solvent. The hydrophilic binder used is a pharmaceutically acceptable hydrophilic additive, preferably one that dissolves in the solvent of the granulation fluid. Preferred examples of hydrophilic polymers used herein include hydroxypropylmethylcellulose (HPMC), sodium carboxymethylcellulose, methylcellulose, hydroxypropylcellulose (HPC), low-substituted hydroxypropylcellulose (L-HPC), hydroxypropylcellulose LF, polyvinylpyrrolidone, polyvinyl alcohol, vinylpyrrolidone-vinyl acetate copolymer (e.g., Kollidon® VA64, BASF), gelatin, guar gum, partially hydrolyzed starch, alginate, or xanthan. It is particularly preferred to use hydroxypropylmethylcellulose (HPMC) as the hydrophilic binder. The hydrophilic binder is present in a concentration of 1% to 12%, preferably 1% to 6% (based on the total weight of the pharmaceutical dosage form). The wetting agents used are pharmaceutically acceptable compounds such as sodium lauryl sulfate, polysorbate, polyethylene glycol (15)-hydroxystearate, or polyethylene glycol hexadecyl ether.

[0118] The wet granulation premix contains additional pharmaceutically acceptable additives such as fillers, binders, and disintegrants (disintegrants). Fillers and binders are, for example, cellulose powder, microcrystalline cellulose, silicified microcrystalline cellulose, hydroxypropyl cellulose, lactose monohydrate, mannitol, maltitol, sorbitol, and xylitol, preferably microcrystalline cellulose or mannitol, or a mixture of microcrystalline cellulose and mannitol / lactose monohydrate. Disintegrants (disintegrants) are, for example, carboxymethylcellulose, croscarmellose (cross-linked carboxymethylcellulose), crospovidone (cross-linked polyvinylpyrrolidone), low-substituted hydroxypropyl cellulose (L-HPC), sodium carboxymethyl starch, sodium potato starch glycolate, partially hydrolyzed starch, wheat starch, corn starch, rice starch, and potato starch.

[0119] The resulting granules are then converted into a solid dosage form. Pharmaceutically acceptable additives that can be added include lubricants, glidants, flow regulators, and disintegrants (disintegrants). Lubricants, glidants, and flow regulators include, for example, fumaric acid, stearic acid, sodium stearyl fumarate, magnesium stearate, high molecular weight fatty alcohols, starch (wheat, rice, corn, or potato starch), talc, highly dispersible (colloidal) silicon dioxide, and glycerol distearate. Disintegrants (disintegrants) include, for example, carboxymethylcellulose, croscarmellose (cross-linked carboxymethylcellulose), crospovidone (cross-linked polyvinylpyrrolidone), low-substituted hydroxypropylcellulose (L-HPC), sodium carboxymethyl starch, partially hydrolyzed starch, wheat starch, corn starch, rice starch, and potato starch.

[0120] The solid dosage forms are optionally coated in a further step under conventional conditions well known to those skilled in the art by the addition of coating and film-forming agents such as hydroxypropylcellulose, hydroxypropylmethylcellulose (e.g., hydroxypropylmethylcellulose 5 cP or 15 cP), polyvinylpyrrolidone, vinylpyrrolidone-vinyl acetate copolymer (e.g., Kollidon® VA64, BASF), shellac, glyceryl triacetate, triethyl citrate, talc as anti-blocking agent and / or colorant / pigment, e.g., titanium dioxide, iron oxide, indigotin or suitable color coatings.

[0121] One embodiment of the present invention includes a solid dosage form containing the active compound product of the compound of Formula (I) in the form of nanoparticles, and further containing microcrystalline cellulose, lactose monohydrate, hydroxypropyl methylcellulose 3 cP and / or 5 cP, sodium lauryl sulfate or polysorbate 20, croscarmellose sodium, magnesium stearate, talc, iron oxide, and titanium dioxide. The nanoparticles still contain one or more stabilizers, but the dispersant has been removed by drying.

[0122] Further embodiments of the present invention include solid dosage forms containing the active compound product of a compound of Formula (I) produced by the process for forming nanoparticles according to the present invention, wherein the solid dosage form contains 1.25 to 20 mg of the active compound product of a compound of Formula (I) per solid dosage form. Further embodiments include solid dosage forms containing 1.25 mg, 2.5 mg, 5.0 mg, 7.5 mg, 10 mg, 12.5 mg, 15 mg, 17.5 mg, or 20 mg of the active compound product of a compound of Formula (I) produced by the process according to the present invention per solid dosage form.

[0123] One embodiment of the present invention is dry nanoparticles comprising the compound of formula (I) methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate in crystalline form of modification I and one or more stabilizers, wherein the X-ray diffractogram of said compound is characterized by exhibiting peak maxima at 2θ angles of 5.9, 6.9 and 22.7, the ratio of compound of formula (I) to one or more stabilizers is between 16:1 and 1:2 w / w, and the dry nanoparticles have an average particle size, expressed as d50, of less than or equal to 500 nm.

[0124] One embodiment of the present invention is a pharmaceutical composition in solid form comprising dry nanoparticles of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate of formula (I) in crystalline form of modification I and one or more stabilizers having an average particle size, expressed as d50, of less than or equal to 500 nm, wherein the X-ray diffractogram of said compound is characterized in that it exhibits peak maxima at 2θ angles of 5.9, 6.9 and 22.7, prepared using nanoparticles according to the invention.

[0125] One embodiment of the present invention is a pharmaceutical composition according to the present invention, wherein the solid form is selected from the group consisting of granules and tablets.

[0126] One embodiment of the present invention is a dry nanoparticle or pharmaceutical composition according to the present invention that does not contain a dispersing agent.

[0127] Aside from solubility in water / microorganism-removing media, stabilizers in nanoformulations must satisfy additional important properties. They must ensure short-term stabilization of the nanosuspension against (re)agglomeration. This property is unavoidable in nanoformulation processes and is a fundamental requirement for producing nanoparticles through milling processes. Furthermore, stabilizers must ensure long-term stabilization of the nanosuspension. Once a nanosuspension is generated, it must remain stable for a distinct period of time until drying can occur. During this time, reagglomeration, crystal growth (Ostwald ripening), and sedimentation must be inhibited by the appropriate selection of stabilizers. Furthermore, it must be ensured that nano-sized particles remain present after reconstitution of the dried material. During spray drying, the nanosuspension undergoes an almost instantaneous transition to solid particles. This phase change implies possible changes to the active compound nanoparticles, including melting, aggregation, or dissolution of the polymer. Identifying appropriate spray drying parameters and suitable excipients must ensure that the active compound particles remain as discrete nanoparticles after dissolution of the dried material.

[0128] An additional obstacle and challenge is the further processing of nanosuspensions containing methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate (Vericiguat, the compound of formula (I)) into granules and tablets as the final dosage form. Potential particle size increase or particle growth can also occur during subsequent processing, such as granulation and compression, resulting in a loss of the high surface area associated with the drug nanoparticles, which reduces the significant benefits intended from the nanomilling process.

[0129] As shown in Figures 6-11, the stable nanosuspensions according to the present invention and the dried pharmaceutical forms made from them resulted in increased drug release. This was observed for the nanosuspensions as well as the granules and tablets made from them. Increased drug release was observed not only in 0.01N HCl but also in bioequivalent media. (FeSSIF) Confocal laser scanning microscopy (CLSM) photographs of granules containing only the micronized compound of Formula (I) (Figure 12a) clearly showed light spots representing the fluorescent compound of Formula (I). In contrast, as shown in Figure 12b, CLSM photographs of the micronized and nanomilled compound of Formula (I) clearly show a much smoother surface without light spots. These photographs clearly show the preserved nanoparticles after micronization and nanomilling.

[0130] Finally, it must be ensured that the nanoparticles are retained in the solid formulations made from the nanosuspensions and that they are still present at the absorption site upon resuspension. As shown in Tables 7 and 8, nanosized particles are still present after reconstitution of the dry material obtained by drying the nanosuspensions, depending on the nature and amount of excipients / stabilizers used.

[0131] The fact that it was possible to transfer the nanosuspension to a solid state and at the same time maintain the excellent bioavailability of the dry nanosuspension was a surprising advantage over the prior art.

[0132] One embodiment of the present invention is a stable nanosuspension comprising the compound of formula (I) in crystalline form of modification I according to the invention and one or more stabilizers in a dispersing agent for use in the treatment and / or prevention of heart failure, worsening chronic heart failure, worsening chronic heart failure with reduced ejection fraction, worsening chronic heart failure with preserved ejection fraction, angina pectoris, hypertension, pulmonary hypertension, ischemia, vascular disorders, cognitive disorders, renal failure, thromboembolic disorders, fibrotic disorders and arteriosclerosis.

[0133] One embodiment of the present invention is dry nanoparticles comprising the compound of formula (I) in crystalline form of modification I according to the invention and one or more stabilizers for use in the treatment and / or prevention of heart failure, worsening chronic heart failure, worsening chronic heart failure with reduced ejection fraction, worsening chronic heart failure with preserved ejection fraction, angina pectoris, hypertension, pulmonary hypertension, ischemia, vascular disorders, cognitive disorders, renal failure, thromboembolic disorders, fibrotic disorders and arteriosclerosis.

[0134] One embodiment of the present invention is a pharmaceutical composition in solid form comprising dry nanoparticles of compound of formula (I) in crystalline form of modification I according to the invention and one or more stabilizers in a dispersing agent, for use in the treatment and / or prevention of heart failure, worsening chronic heart failure, worsening chronic heart failure with reduced ejection fraction, worsening chronic heart failure with preserved ejection fraction, angina pectoris, hypertension, pulmonary hypertension, ischemia, vascular disorders, cognitive disorders, renal failure, thromboembolic disorders, fibrotic disorders and arteriosclerosis.

[0135] One embodiment of the present invention is a method for the treatment and / or prevention of heart failure, worsening of chronic heart failure, worsening of chronic heart failure with reduced ejection fraction, worsening of chronic heart failure with preserved ejection fraction, angina pectoris, hypertension, pulmonary hypertension, ischemia, vascular disorders, cognitive disorders, renal failure, thromboembolic disorders, fibrotic disorders and arteriosclerosis in humans and animals by administering an effective amount of a pharmaceutical composition in solid form prepared with a nanosuspension comprising the compound of formula (I) in crystalline form of modification I according to the invention and one or more stabilizers in a dispersing agent.

[0136] The improved bioavailability of this nanosuspension was demonstrated in rat studies compared with a Tylose suspension containing only micronized material of the compound of Formula (I). The nanoscale effect on bioavailability improvement was maintained even after drying of the suspension in a fluidized-bed granulator, a highly sensitive manufacturing step prone to API particle aggregation or Ostwald ripening and therefore risking loss of the nanoparticle benefits. Relative bioavailability studies were conducted in rats using granules and minitablets. As shown in Example 5, Table 3, Example 6, Tables 5-7, and Figures 6-11, formulations containing nanosized processing starting materials resulted in higher in vitro and in vivo bioavailability than formulations containing micronized starting materials.

[0137] Due to the dissolution-limited behavior of the compound of formula (I) (Vericiguat), its bioavailability in preclinical and clinical studies decreased at higher doses. Nanoformulations can generally increase dissolution-rate-limited bioavailability and avoid the food effect.

[0138] The data presented are surprising beyond the state of the art, as the development of stable nanosuspensions using SDS as a stabilizer would not have been expected. Furthermore, it was surprising that the choice of stabilizer was a critical process parameter for producing nanoparticles by wet bead milling. The advantages of using SDS instead of vitamin E TPGS as a stabilizer, as demonstrated by the smaller particle size and lack of amorphization after milling (both of which result in smaller nanoparticles, important parameters for increasing bioavailability), were also surprising beyond the state of the art.

[0139] The ratio of Compound (I):SDS used in the experiment was 2:1. Since the content of Compound (I) was 2%, the content of SDS was 1%. The critical micelle concentration (CMC) of SDS is 0.23% or 8.2 mM. Therefore, the concentration of SDS used in the experiment was more than four times its CMC. Therefore, the concentration of surfactants, such as SDS, used to achieve stable nanoformulations according to the present invention clearly exceeds the critical micelle concentration (CMC). As outlined in the prior art, surfactant concentrations clearly exceeding the CMC are thought to promote Ostwald ripening. Therefore, it was surprising that Ostwald ripening was not observed in the present invention.

[0140] The nanoparticles were physically stabilized using sodium dodecyl sulfate (SDS) in the API / SDS concentration range of 8:1 to 2:1 w / w. Surprisingly, the vitamin E TPGS-stabilized nanoparticles known from the prior art showed poorer results with respect to higher particle size and poorer bioavailability compared to similarly prepared nanoparticles using SDS as a stabilizer, as shown in Figure 1 and Example 5, Table 3, Example 6, Tables 4-6, and Figure 10.

[0141] In a first step, an extensive screening of stabilizers revealed that not all excipients described in the literature were suitable for the preparation of nanosuspensions of this compound: human serum albumin, egg lecithin, hydroxypropylmethylcellulose acetate succinate (HPMC-AS) or sodium oleate resulted in undesirable products with oversized particles or aggregation phenomena.

[0142] A beneficial effect of the stable nanosuspensions and nanoparticles according to the present invention is an increased bioavailability of the compound of formula (I). [Brief explanation of the drawings]

[0143] [Figure 1a]Figure 1 shows the results of a screening experiment of nanosuspensions made with a compound of formula (I) after wet bead milling in a planetary mill with different stabilizers (Example 1). [Figure 1b]

[0023] Figure 1 shows the results of a screening experiment of nanosuspensions made with a compound of formula (I) synthesized according to WO2020126983, published after the first filing date of this application, after wet bead milling in a planetary mill with different stabilizers (Example 1). [Figure 2] The long-term stability of different nanoparticles obtained in the screening of Example 1 at elevated temperatures for up to 13 weeks is shown. The milling conditions were as follows: Compound (I): Stabilizer 1 ratio = 5:1; volume filling with milling beads = 50%; milling bead size = 0.3-0.4 mm; milling time = 90 min; mill: planetary mill, storage at 40 °C. The particle size is given as Dn(50) and polydispersity index (PDI). [Figure 3] Figure 1 shows particle size distribution and Compound (I):stabilizer ratio for different concentrations of micronized and nano-milled Compound (I) milled in a planetary mill. API = Compound (I) (Example 3, scale-up). [Figure 4] Figure 1 shows the particle size distribution after nanomilling of Compound (I) (Vericiguat) (20% API suspended relative to total mass). Data are given as volume fraction Q3 (volume percentage, vol%, Q3 indicates that the particles are measured in three dimensions, and X% indicates the percentage of particles smaller than the indicated size). Vericiguat material in both curves was micronized and not sonicated. Left curve: nanomilled in a Netzsch mill with 3.3% SDS for 90 minutes. Right curve: no nanomilling. [Figure 5a] 1 shows the XRPD measured for Suspension 1 of Example 4. The lower figure shows the XRPD of Compound (I) in crystalline modification I as a reference, and the upper figure shows the XRPD of Compound (I) after wet bead milling under the conditions shown for Suspension 1 in Table 2. The comparison shows that no amorphization occurs during the wet bead milling procedure. [Figure 5b]1 shows the XRPD measured for Suspension 2 of Example 4. The lower figure shows the XRPD of Compound (I) in crystalline modification I as a reference, and the upper figure shows the XRPD of Compound (I) after wet bead milling under the conditions shown for Suspension 2 in Table 2. The comparison shows that no amorphization occurs during the wet bead milling procedure. [Figure 5c] 1 shows the XRPD measured for Suspension 3 of Example 4. The lower figure shows the XRPD of Compound (I) in crystalline modification I as a reference, and the upper figure shows the XRPD of Compound (I) after wet bead milling under the conditions shown for Suspension 3 in Table 2. The comparison shows that no amorphization occurs during the wet bead milling procedure. [Figure 6] 1 shows the amount of Compound (I) (Vericiguat) as granules released into a flow-through cell using three different media. [Figure 7] Figures 6 and 7 show the cumulative amount of Compound (I) (vericiguat) released as granules measured in a flow-through cell using three different media. Figures 6 and 7 clearly demonstrate the increased drug release using nanomilled Compound (I) (vericiguat) compared to micronized Compound (I) (vericiguat), the only difference between the batches investigated. This advantage of nanomilled Compound (I) (vericiguat) is independent of the media tested. [Figure 8]

[0023] Figure 1 shows drug dissolution rate data for tablets incorporating micronized or nanomilled Compound (I) (Vericiguat) drug substance at 10 mg dosage strengths. Drug dissolution rates were measured in 0.01 N HCl (mean of n = 6). [Figure 9]

[0023] Figure 1 shows drug dissolution rate data for tablets incorporating micronized or nanomilled Compound (I) (Vericiguat) drug substance at 15 mg dosage strengths. Drug dissolution rates were measured in 0.01 N HCl (mean of n = 6). [Figure 10]An overview of all in vivo results obtained from different formulations containing Compound (I) (solution, suspension, granules, minitablets): low dose: 0.3 mg / kg, medium dose: 1 mg / kg (solution only), high dose: 2.1 mg / kg for granules, 3 mg / kg for solution, suspension (using two different particle sizes: a tylose suspension with microcrystalline material and a suspension with nanosized Compound (I)) and minitablets (made from either micronized or micronized + nanomilled Compound (I)). [Figure 11] Figure 1 shows dissolution data for tablet cores (uncoated) containing 15 mg of Compound (I) in micronized or micronized + nanomilled form. The dissolution profiles were measured in a biorelevant medium (FeSSIF) where no-sink conditions exist. "No-sink conditions" means that less than three times the dose contained in the tablet dissolves in the release medium. This results in a higher differentiation. Looking at the profiles, the formulation with nanomilled Compound (I) is clearly superior to the formulation with micronized Compound (I), with the particle size of Compound (I) being the only difference between the two tablet cores. [Figure 12] Figures 12a and 12b show confocal laser scanning microscopy (CLSM) images of granules containing micronized (Figure 12a, #WU-000704-01) and nanomilled (Figure 12b, #WU-000704-02) compounds of Formula (I), respectively. Fluorescence was stimulated at 405 nm, and emission was measured at 420-500 nm with a gain of 550 using an HC PL APO CS2 63x / 1.40 OIL objective. Figure 12a clearly shows the light spots representing the fluorescent compound of Formula (I) that has only been micronized. Figure 12b clearly shows a much smoother surface without the light spots representing the fluorescent compound of Formula (I). DETAILED DESCRIPTION OF THE INVENTION

[0144] Example

[0145] [Table 2]

[0146] Wet bead grinding equipment Method A: Planetary Mill Most of the screening experiments in Example 1 were carried out in a Pulverisette 5 planetary ball mill (PBM). For sample preparation, the finely divided compound of formula (I) was pre-dispersed in an aqueous polymer surfactant solution and transferred to a milling chamber (PET 23 ml) filled with milling beads. Water contained in the solution was used as the dispersant. Silibeads® zirconium oxide (yttrium-stabilized) from Sigmund Lindner (Germany) was used as the milling media (0.4-0.6 mm) with a bulk density of 3.9 kg / L. The milling bead filling level was 60% of the milling chamber volume. Care was taken to avoid leaving air in the milling chamber, as this would adversely affect the milling process. The milling time was 90 minutes at 400 revolutions per minute. Four milling chambers could be fed into the PBM for each milling run. Therefore, after separation of the milling media, approximately 40-45 g of suspension could be produced per milling run.

[0147] Method B: Stirred Media Mill To produce larger quantities of nanosuspensions (eg, for animal studies), either a picoliq (Hosokawa Alpine) or a Netzsch Labstar stirred media mill was used.

[0148] Drug nanosuspensions were prepared by wet milling the micronized compound of formula (I) in a Hosokawa Picoliq-type stirred media mill with a milling chamber volume of 19 ml. Milling was performed using a stirrer. The rotation speed was 10,000 rpm. The milling media was identical to that of PBM, including the use of water as a dispersant. The milling media fill level was 80% of the milling chamber volume. The mill was operated in batch mode. 13.6 g of suspension was milled. The entire system was cooled by a cryostat so that a maximum temperature of 29°C was measured at the milling chamber outlet during milling.

[0149] Alternatively, drug nanosuspensions were prepared by wet milling the micronized compound of formula (I) in a Netzsch LabStar-type stirred media mill with a milling chamber volume of 120 ml. Milling was performed using a disk agitator. The rotation speed was 2770 rpm. The milling media was identical to that of the PBM. The milling media fill level was 80% of the milling chamber volume. The mill was operated in circular mode. The feed vessel held a total of 1 L, and 600 g of suspension was milled, which corresponds to approximately 500 ml. The entire system was cooled by a cryostat so that a maximum temperature of 23°C was measured at the milling chamber outlet.

[0150] The specific energy demand for producing nanoparticles by wet bead milling in a stirred media mill was a minimum of 10,000 kJ / kg. The stress intensity (Kwade et al., 1996) was 0.004 10 -3 Nm~1·10 -3 Nm.

[0151] Example 1 Nanosuspension screening experiments Crystalline form of modification I of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate (Vericiguat, Compound I) was milled to nanoscale particle size (d50 after milling < 200 nm) by a wet bead milling process according to Examples 1-3.

[0152] Figure 1 shows the results of a screening experiment after applying a wet bead milling process in a planetary mill. The compound of formula (I) in crystalline form in modification I was prepared according to Example 13, Method E of WO 2013 / 076168. Prior to nanomilling, the compound of formula (I) in crystalline form in modification I was micronized. For the screening experiments, many different formulations were tested. In these experiments, the duration of milling was 60 minutes. The milling bead material used was ZrO2 with a milling bead size of 0.5 mm (0.4-0.6 mm).

[0153] In a first step, extensive screening of stabilizers revealed that not all excipients described in the literature were suitable for producing nanosuspensions of this compound. The best results were found to be achieved with sodium dodecyl sulfate (SDS) and vitamin E TPGS as single stabilizers, as well as combinations of two stabilizers, such as surfactants and polymers (e.g., SDS + polyvinylpyrrolidone K17 (PVP K17)). Eudragit® EPO, egg lecithin, polyvinyl acetate (PVA), sodium oleate, and human serum albumin (HSA) were not suitable for nanomilling for various reasons. Human serum albumin, egg lecithin, hydroxypropylmethylcellulose acetate succinate (HPMC-AS), or sodium oleate resulted in undesirable products with oversized particles or aggregation.

[0154] The ratio of Compound (I) to one or more stabilizers was 16:1 to 1:2 w / w or 8:1 to 2:1 w / w. When a combination of two stabilizers, such as a surfactant and a polymer (e.g., SDS + polyvinylpyrrolidone or SDS + ethylene oxide-propylene oxide block copolymer), was used, the ratio of Compound (I) to polymer was 16:1 to 1:2 w / w or 8:1 to 2:1 w / w, and the surfactant concentration was 0.1 to 0.2% w / v.

[0155] In the meantime, a new API quality of the compound of formula (I) in the crystalline form of modification I, synthesized and micronized according to WO 2020126983, published after the first filing date of the present application, was introduced. Therefore, the screening was repeated with this new API quality, and the results could be reproduced (Figure 1b).

[0156] SDS was the preferred stabilizer for further investigation, as it was also used in the further granulation process, however PVP VA64, d-alpha tocopheryl polyethylene glycol 1000 succinate (vitamin E TPGS), Cremophor® EL, PVP K30, and hydroxypropyl cellulose (HPC) were also used for further investigation in the Picoliq mill.

[0157] Example 2 Stability testing of nanosuspensions a) Particle size immediately after grinding and over a period of at least 7 days The purpose of these experiments was to evaluate the stability of the nanosuspension over a period of at least 7 days. Furthermore, we assessed whether milling would induce particle disintegration. For nanomilling, the following parameters were applied: micronized compound (I):stabilizer ratio 2:1, compound (I) content in the suspension 2%, milling bead material yttrium-stabilized ZrO2, milling bead size 0.4–0.6 mm, milling duration 60 min, Picoliq mill.

[0158] [Table 3]

[0159] The results show that the dispersions are largely stable, but show some small differences in particle size. However, the HPC formulation shows a significant increase in d95 after 7 days at ambient temperature.

[0160] b) Long-term measurement of particle growth over 13 weeks The stability of the different nanoparticles obtained in the screening of Example 1 was measured at elevated temperatures for up to 13 weeks. The milling conditions were as follows: Compound (I):stabilizer ratio = 8:1; volumetric filling with milling beads: 60%; milling bead material: ZrO2, milling bead size: 0.4-0.6 mm; milling time: 60 min; mill: planetary mill; storage at 40 °C and 75% RH. The particle size distribution of the nanosuspension was measured by dynamic light scattering (DLS). DLS was performed using a Zetasizer Nano-ZS (Malvern Analytical) at an appropriate concentration (e.g., 0.2 mg / mL, diluted with demineralized water). The Dn(50) and polydispersity index (PDI) were obtained as responses. The results are shown in Figure 2.

[0161] Example 3 Scale-up of nanosuspension preparation The next step in the development of nanosuspensions was scale-up. To achieve this, at least two problems had to be solved: In a first step, the concentration of the compound of formula (I) (Vericiguat) in the nanosuspension had to be increased from 2% to 20%. This was necessary mainly to reduce the drying operation and process time. A 2% suspension is too dilute and not suitable for fluidized bed granulation. These experiments were carried out in a planetary mill.

[0162] Figure 3 shows the results of these experiments in a planetary mill. "LD particle size" indicates particle size measurements by static light scattering (SLS). During the experiments, it became clear that increasing the SDS concentration above 5% was not possible due to gelation. Therefore, for higher concentrations of the compound of formula (I) in the nanosuspension, the compound of formula (I):stabilizer ratio had to be changed in favor of a higher API share. Particle size measurements indicate that higher concentrations of the compound of formula (I) in the nanosuspension, up to 20%, are possible without a significant increase in particle size. Conversely, the d90 values ​​decreased slightly with increasing concentrations of the compound of formula (I) and the compound (I):stabilizer ratio.

[0163] Experiments have shown that increasing the concentration of Compound (I) is possible, however, consideration should be given to starting with a lower concentration of stabilizer (see Example 3 above).

[0164] In the second step, the process had to be transferred to a larger mill so that larger quantities of product could be produced. For this purpose, the process was transferred to a 120 ml Netzsch Labstar mill. 60 g of nanosuspension was produced for further processing in fluidized bed granulation (FBG). This experiment was carried out using the Netzsch mill in batch mode. The results of this experiment are shown in Figure 4.

[0165] The median particle size was 120 nm. The suspension was further processed by fluid bed granulation.

[0166] Example 4 Compound (I) in the crystalline form of modification I after wet bead milling in a Picoliq mill To analyze whether amorphization of compound (I), used in the crystalline form of micronized modification I in the wet bead milling process to produce nanosuspensions, occurs during wet bead milling, three nanosuspensions were produced by wet bead milling in a Picoliq mill under various conditions, as shown in Table 2.

[0167] [Table 4]

[0168] XRPD analyses of these three nanosuspensions are shown in Figures 5a-c. XRPD analysis revealed that the crystalline morphology of modification I was preserved during wet bead milling, even under high mechanical stresses such as long milling times and high rotation speeds, as exemplified for suspension 3.

[0169] Example 5 Further processing of the nanosuspension into granules and tablets The preblend used was improved using wet granulation technology (fluid bed granulation): the active compound is suspended in a granulation liquid and sprayed onto the preblend to ensure a uniform distribution of the active compound in the resulting granules.

[0170] Process steps leading up to final tablet formulation To investigate the effect of particle size on the bioavailability of methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate (Vericiguat, Compound (I)), several formulations were prepared using suspensions containing drug substance that was first micronized and then nanomilled, and suspensions containing drug substance that was only micronized. The compositions of these suspensions are outlined in the following paragraphs. In the context of the present invention, micronization is carried out by milling, for example, in a spiral jet mill or a fluidized-bed opposed jet mill.

[0171] First step—Preparation of a suspension containing micronized drug substance: To incorporate the micronized drug substance into a suspension, the binder hydroxypropyl methylcellulose (HPMC 5cP) and the wetting agent sodium dodecyl sulfate (SDS) were dissolved in water. After a clear solution was obtained, micronized methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate (Vericiguat, Compound (I)) was suspended in the binder solution and uniformly distributed (compare Table 3, "Suspension").

[0172] Second step - Preparation of granules using the suspension containing the micronized drug substance: To produce the granules, the resulting suspension was sprayed onto a premix of the filler cellulose microcrystals (Avicel PH 101) and lactose monohydrate and a portion of the disintegrant croscarmellose sodium (AcDiSol) using fluidized bed granulation techniques. The granules were dried and sieved (mesh size 0.8 mm) (compare Table 3, "Granules").

[0173] Third step - Preparation of tablets based on granules containing micronized drug substance: The resulting granules were post-blended in two steps with the disintegrant croscarmellose sodium and the lubricant magnesium stearate in part 2. A portion of this ready-to-press blend was compressed into tablets (9 mm diameter circular format) or mini-tablets (1.2 mm diameter) (Table 3, compare "Tablets").

[0174] This three-step procedure was used to manufacture two different batches of tablets with different dosages (Table 3, compare batches 2 and 4).

[0175] A comparable approach was implemented for the use of micronized + nanomilled drug substance.

[0176] First step - Preparation of suspension containing micronized nanomilled drug substance: To incorporate micronized nanomilled Compound (I) physically stabilized with sodium dodecyl sulfate (SDS) into granules, an aqueous suspension of micronized nanomilled Compound (I) and sodium dodecyl sulfate (SDS) was taken and mixed with an aqueous solution of hydroxypropyl methylcellulose (HPMC 5cP) in the relative amounts shown in Table 3 below. Of the amount of water used, 30% was used to prepare the nanoparticles and 70% was used to prepare the aqueous solution of binder (HPMC 5cP) (Table 3, compare "Suspension").

[0177] Second step - Preparation of granules using the suspension of micronized + nanomilled drug substance: The resulting suspension was sprayed onto a premix of fillers cellulose microcrystals and lactose monohydrate and a portion of the disintegrant croscarmellose sodium (AcDiSol) using fluidized bed granulation technology. The granules (preblend) were dried and sieved (mesh size 0.8 mm) (Table 3, compare "Granules"). The granules contain nanoparticles in dry form. The dispersant (here water) was removed in a drying step.

[0178] Third step - Preparation of tablets based on granules containing micronized + nanomilled drug substance: The resulting granules were post-blended in two steps with the second part of the disintegrant croscarmellose sodium (AcDiSol) and the lubricant magnesium stearate. A portion of this ready-to-press blend was compressed into tablets (circular format, 9 mm diameter). For administration of the tablets in in vivo studies in rats (Example 6b), mini-tablets were compressed (1.2 mm diameter) (compare Table 3, "Tablets"). The tablets contain nanoparticles in dry form that no longer contain a dispersant (here water).

[0179] Using this three-step procedure, two different batches of tablets using micronized and nanomilled Compound (I) were manufactured, differing only in dosage strength (Table 3, compare batches 1 and 3).

[0180] [Table 5]

[0181] The manufactured granules (15 mg dose strength) of batches 3 and 4 were subjected to dissolution testing in three different media (phosphate buffer pH 6.8, FaSSIF, and FeSSIF) using a mini flow-through cell dissolution apparatus (USP Convention 2011, Revision Bulletin, Official Feb 1, 2012, General Chapter <711> The cells were characterized by flow-through experiments (n=2±SD (1 mg API per cell, 2 ml / min flow rate) through a dissolution medium). The results are summarized in Figures 6 and 7.

[0182] Figures 6 and 7 clearly demonstrate the increased drug release of granules made with micronized + nanomilled Compound (I) (Vericiguat) compared to granules made with only micronized Compound (I) (Vericiguat) (composition according to Table 3), the only difference between the batches investigated. This advantage of micronized + nanomilled Compound (I) (Vericiguat) is independent of the vehicle tested.

[0183] Additionally, all four manufactured batches of tablets (compare Table 3) were characterized by drug dissolution behavior using a USP II paddle apparatus (USP Convention 2011, Revision Bulletin, Official Feb 1, 2012, 711 Dissolution) with 900 mL of 0.01 N HCl at 75 rpm. A comparison is shown in Figure 8 for the 10 mg dose and Figure 9 for the 15 mg dose.

[0184] The percentage of drug dissolved was higher in the two tablet batches containing micronized + nanomilled Compound (I) (Vericiguat) compared to both batches containing micronized drug substance, regardless of the dose strength investigated.

[0185] Additionally, 15 mg tablets containing micronized + nanomilled or micronized only Compound (I) (Vericiguat) with compositions according to Table 4 were characterized by drug dissolution behavior using a USP II paddle apparatus (USP Convention 2011, Revision Bulletin, Official Feb 1, 2012, 711 Dissolution) with a 500 mL FeSSIF at 75 rpm. The dissolution data are shown in Figure 11. The data clearly demonstrate the increased drug release using micronized + nanomilled Compound (I) (Vericiguat) compared to micronized Compound (I) (Vericiguat), the only difference between the batches investigated.

[0186] [Table 6]

[0187] Example 6 In vivo experiments a) Nanosuspension All in vivo studies were performed in catheterized male Wistar rats (n=3) in accordance with the German Animal Protection Act. Three doses were applied in different formulations (suspension, granules, minitablets): 0.3, 1.0, and 3.0 mg / kg (granules at the highest dose of 2.1 mg / kg), covering the entire therapeutic range. The applied volume of the solution and suspension was 5 mL / kg. Plasma samples were collected over 48 hours post-dose and analyzed by LC-MS / MS. Pharmacokinetic parameters were estimated from the plasma concentration-time profiles using standard PK software. Relative bioavailability was calculated as the AUC ratio obtained from the different test arms, comparing two different formulations applied via the same administration route at normalized doses.

[0188] For comparison, the test compound (Compound (I)) was orally administered as a solution (100% PEG or PEG / EtOH / water (40 / 10 / 50, V / V / V)) containing Compound (I), which is less likely to precipitate in vivo. Instead of disintegrated standard tablets, a suspension containing 0.5% Tylose solution in which the compound was suspended as a micronized substance was administered. The following two nanosuspensions were tested at different dose strengths: a) Nanosuspension stabilized by SDS a. 0.6 mg / mL micronized + nanomilled Compound (I) + 0.3 mg / mL SDS solution b. 1 mg / mL micronized + nanomilled Compound (I) + 0.5 mg / mL SDS solution c. 6 mg / mL micronized + nanomilled Compound (I) + 3 mg / mL SDS solution b) Vitamin E TPGS stabilized nanosuspension a. 0.6 mg / mL micronized + nanomilled Compound (I) + 0.3 mg / mL Vitamin E TPGS solution b. 1 mg / mL micronized + nanomilled Compound (I) + 0.5 mg / mL Vitamin E TPGS solution c. 6 mg / mL micronized + nanomilled Compound (I) + 3 mg / mL Vitamin E TPGS solution

[0189] Following animal testing, the particle size of the applied nanosuspension was checked to ensure that no significant changes in particle size had occurred. The bioavailability of the nanosuspension is shown in Table 5.

[0190] [Table 7]

[0191] b) Granules and tablets Granules based on microsuspensions or nanosuspensions (produced in micronized or micronized + nanomilled form by fluidized bed granulation with Compound (I)) were administered in minicapsules (doses: 0.3 and 2.1 mg / kg due to limited filling volume, see Table 3 for composition). Minitablets (diameter = 1.2 mm) made from the granules were also administered in minicapsules. A dose of 0.3 mg / kg corresponded to the ratio of the components of one tablet of 15 mg granules, so that a dose of 3 mg / kg corresponded to 10 tablets.

[0192] The above granules based on nanosuspensions of the compound of formula (I) were compared in vivo with granules based on microsuspensions of the compound of formula (I) and with the compound of formula (I) in PEG solution. Dose proportionality was observed. The results are shown in Tables 5, 6, and Figure 10.

[0193] [Table 8]

[0194] [Table 9]

[0195] At the lower dose (0.3 mg / kg), the micro- and nano-formulations were comparable. AUC and C maxAt higher doses (2.1 or 3 mg / kg), the nanoformulation was superior, as indicated by an increase in . The superiority of the formulation containing micronized+nanomimized Compound (I) over the formulation containing only micronized Compound (I) was demonstrated for all formulation types, i.e., suspension, granules, and tablets.

[0196] Example 7 Screening experiments on stability and reconstitution after drying of nanosuspensions To analyze whether nanosized particles are still present after reconstitution of the dry material obtained by drying the nanosuspension, the following screening experiment was carried out: Different active ingredient / stabilizer combinations were nanomilled in a planetary ball mill (PBM) of type Pulverisette 5 as described in Method A, except that 5 ml vials were used and the milling time was 60 min at 400 rpm.

[0197] The particle size distribution was measured immediately after grinding. For the determination of the stability of the suspension, the nanosuspension was stored at 40°C for 7 days at room temperature without stress (without stirring).

[0198] To measure the stability of nanoparticles after drying and reconstitution, nanosuspensions were milled in a drying chamber at 40 °C for 14–18 h and then dried immediately. Water was added to the dried film, which was then mixed with a magnetic stirrer for up to 5 min. The particle size distribution was measured.

[0199] These tests were carried out using various combinations of HPC or PVP K30 and SDS as a stabilizer and lactose as a matrix former in the drying process, and the results are shown in Tables 7 and 8.

[0200] [Table 10]

[0201] [Table 11]

[0202] [References] TIFF0007780442000016.tif216168

Claims

1. Methyl {4,6-diamino-2-[5-fluoro-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-5-yl}carbamate of formula (I) in the crystalline form of modification I 【Chemistry 1】 and one or more stabilizers in a dispersant, the X-ray diffractogram of the compound is characterized by peak maxima at 2θ angles of 5.9, 6.9, and 22.7; the one or more stabilizers are polyvinylpyrrolidone (PVP) in combination with sodium dodecyl sulfate (SDS), or are selected from the group consisting of vinylpyrrolidone-vinyl acetate copolymer, ethylene oxide-propylene oxide block copolymer, sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose (HPMC), polysorbate, hydroxypropyl cellulose (HPC), polyoxyl-35 castor oil, polyoxyl-15 hydroxystearate, Na-desoxycholate, and combinations thereof; the maximum concentration of the one or more stabilizers is the solubility limit of the one or more stabilizers in the suspension; the dispersant is selected from the group consisting of water, primary, secondary and tertiary alcohols, and polyhydric alcohols; the nanoparticles have an average particle size, expressed as d50, of less than or equal to 500 nm; and A stable nanosuspension having a mean particle size, expressed as d50, that remains below 500 nm when stored at a temperature of at least 40°C for at least one week.

2. 2. The stable nanosuspension of claim 1, wherein the one or more stabilizers are selected from the group consisting of sodium dodecyl sulfate, polyvinylpyrrolidone K10-K50 in combination with sodium dodecyl sulfate (SDS), hydroxypropyl methylcellulose, polysorbate 20-80, and combinations thereof.

3. 3. The stable nanosuspension according to claim 1 or 2, wherein the ratio of the compound of formula (I) to one or more stabilizers is from 16:1 to 1:2 w / w.

4. 4. The stable nanosuspension of claim 1, wherein the nanoparticles have an average particle size, expressed as d50, of less than or equal to 300 nm.

5. 5. The stable nanosuspension of claim 1, wherein the mean particle size, expressed as d50, remains equal to or less than 300 nm when stored for at least one week at a temperature of at least 40°C.

6. 6. A process for preparing a stable nanosuspension according to any one of claims 1 to 5, comprising: a) suspending the compound of formula (I) in crystalline form of modification I in water, a dispersant selected from the group consisting of primary, secondary and tertiary alcohols and polyhydric alcohols, and one or more stabilizers according to claim 1 or 2, wherein the maximum concentration of the one or more stabilizers is the solubility limit of the one or more stabilizers in the suspension; b. Specific energy input of 10,000 kJ / kg or more and 0.004 10 －3 Nm to 1.10 －3 wet bead milling the suspension produced in step a at a stress intensity of 0.05-0.8 mm; A process involving:

7. 7. The process according to claim 6, wherein in step a) the compound of formula (I) in crystalline form of modification I is micronized in a first step before suspending it in a dispersing agent and one or more stabilizers.

8. 7. The process according to claim 6, wherein in step a) the compound of formula (I) in crystalline form of modification I is milled in a first wet bead milling step in the presence of milling beads having a size of 1-2 mm before suspending it in a dispersing agent and one or more stabilizers.

9. 9. The process of any one of claims 6 to 8, wherein the grinding beads are made from a material selected from the group selected from ceramic, glass, polymer, and steel.

10. 10. The process of any one of claims 6 to 9, wherein the volume filling with grinding beads is 50 to 85% v / v.

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