HK40056364B - Dry powder inhalation formulation and its use for the therapeutic treatment of lungs - Google Patents

Description

Dry powder inhalation formulations and their use in the treatment of lung diseases.

This invention relates to dry powder inhalation formulations and their use in the treatment of lung diseases.

Dry powder inhalation (DPI) therapy is now primarily used to deliver many treatments to the lungs, such as in the treatment of asthma, and much effort has been made to optimize dry powder inhalation formulations to prolong the retention of the active pharmaceutical ingredient (API) in the lungs.

Inhalation allows high doses of drugs to be delivered directly to the lungs without prior distribution in the body. On the one hand, this allows for the use of smaller amounts of the active pharmaceutical ingredient (API) and potentially waste in the body; on the other hand, direct in-situ administration allows for reduced toxicity when considering the toxicity of APIs.

Furthermore, due to the lungs' characteristic of a large absorption surface, inhalation is a promising route of administration for delivering APIs into the systemic circulation.

However, conventional immediate-release dry powder inhalation (DPI) formulations may still result in too short a residence time in the lungs due to multiple clearance mechanisms of exogenously inhaled particles (i.e., mucociliary and macrophage clearance) and rapid dissolution of the drug particles leading to rapid absorption into the systemic circulation. Therefore, this short residence time necessitates multiple doses, resulting in poor patient compliance. Immediate release of the API may also lead to poor local tolerance due to the rapid dissolution of the drug particles and the high peak concentration of API in the pulmonary fluid after lung deposition.

Specific excipients have been identified to prolong the sustained retention curve of the API in the lungs.

Therefore, further development of dry powder inhalation formulations is needed to increase the effectiveness of lung treatment.

In this context, for example, cisplatin-based controlled-release DPI formulations with high drug loading and high fine particle fraction are developed (Levet et al., Int J Pharm [International Journal of Pharmaceutics] 2016). These formulations exhibit controlled release and prolonged lung retention, resulting in low systemic distribution (Levet et al., Int J Pharm [International Journal of Pharmaceutics] 2017).

To this end, Levet et al. have investigated a specific formulation of an antitumor agent, namely cisplatin containing PEGylated excipients, to avoid premature clearance by the lung epithelial defense mechanism, and have also compared different carriers, such as polymer and lipid matrices, to allow for prolonged retention time in the lungs, thereby increasing inhalation efficiency.

Among other things, the lipid matrix described in the work of Levet et al. contains tristearin, which has been identified as a promising candidate for providing a lipid matrix.

Furthermore, even though lipid-derived excipients used to form the matrix are superior to polymers for toxicity reasons, only a few substances capable of forming lipid matrices are permitted and accepted for inhaled pharmaceutical compositions compared to other therapeutic routes (Pilcer et Amighi, Int J Pharm [International Journal of Pharmaceutics] 2010).

Unfortunately, although tristearin has been identified as a promising candidate, its market availability as a pharmaceutical compound is rather limited.

The present invention addresses at least a portion of these disadvantages by providing, for example, dry powder inhalation formulations for monotherapy or polytherapy, the formulations comprising at least one active pharmaceutical ingredient (API) and a lipid matrix, the lipid matrix comprising at least one triglyceride selected from the group consisting of monohydroxystearin, dihydroxystearin, trihydroxystearin, and mixtures thereof.

It has been surprisingly determined that at least one triglyceride (in pharmaceutical grade) selected from the group consisting of monohydroxystearin, dihydroxystearin, trihydroxystearin and mixtures thereof is capable of forming a lipid matrix with an API (active pharmaceutical ingredient) and exhibits an increased rate of pulmonary deposition in the lungs, which allows for pulmonary inhalation.

In fact, monohydroxystearin, dihydroxystearin, trihydroxystearin and mixtures thereof are compounds with physicochemical properties, such as a LogP value greater than 5, preferably between 14 and 25, more particularly between 18 and 25 and preferably about 20, as measured by the shake flask method and/or a melting point temperature greater than or equal to 40°C, preferably greater than or equal to 60°C, more preferably greater than or equal to 75°C.

This allows for the production of inhalation formulations in the form of dry powder, which are neither oil-based nor viscous pastes, making them suitable for use in dry powder inhalers.

Furthermore, according to the present invention, contrary to all expectations, it has been determined that by changing the ratio between the amount of API and the amount of at least one triglyceride selected from the group consisting of monohydroxystearin, dihydroxystearin, trihydroxystearin, and mixtures thereof, it is possible to increase the rate of pulmonary deposition of API relative to other conventional triglycerides such as tristearin, and to provide a regulated dissolution profile of API, which is not the case for conventional triglycerides.

Preferably, the weight ratio between the at least one API and the at least one triglyceride in the dry powder inhalation formulation according to the invention is: API/triglyceride is 0.1/99.9 to 99.9/0.1, preferably 10/90 to 88/12, preferably 15/85 to 85/15, preferably 25/75 to 75/25, more preferably 30/70 to 70/30, particularly 40/60 to 60/40, for example about 50/50.

Advantageously, in the formulations according to the invention, said at least one triglyceride is hydrogenated castor oil.

Castor oil has been described in a long list of common excipients (see US 2005/0042178) or grinding aids (WO2013/128283) in the literature.

In a preferred embodiment of the invention, the dry powder inhalation formulation comprises 0.1 wt% to 98 wt% of the at least one API relative to the total weight of the dry powder inhalation formulation. In a first specific embodiment of the invention, the active agent is a small chemical molecule with a solubility in alcohol of at least 0.1 w%/v, or at least 0.5 w%/v, more preferably at least 1 w%/v, and more particularly at least 5 w%/v. According to this first specific embodiment, the small chemical molecule is any active pharmaceutical ingredient (API) with an alcohol solubility of 0.1% w/v or higher, defined in this invention as an alcohol-soluble API, such as budesonide, paclitaxel, pemetrexed, itraconazole, voriconazole, clarithromycin, salbutamol, salbutamol sulfate, fluticasone, beclomethasone, mometasone, mometasone furoate, ciroxonide, formoterol, aformoterol, indacaterol, indacaterol maleate, olodaterol, olodaterol hydrochloride, salmeterol, ipratropium bromide, glycopyrronium bromide, tiotropium bromide, urodimethyl bromide, ibuprofen, vancomycin, vancomycin hydrochloride, tetrahydroripotassium, isoniazid, rifampin, pyrazinamide, docetaxel, vincristine, vincristine sulfate, etoposide, vinorelbine, gemcitabine, etc.

More specifically, relative to the weight of the dry powder inhalation formulation, the dry powder inhalation formulation according to the first specific embodiment contains 0.1 wt% to 95 wt%, preferably 1 wt% to 90 wt%, more preferably 1 wt% to 85 wt% of the at least one API.

In a preferred embodiment of the invention, the dry powder inhalation formulation comprises lipid matrix particles in which the alcohol-soluble API is dissolved or finely dispersed.

In a second embodiment of the invention, the active agent is a small chemical molecule with a solubility in alcohol of less than 0.5 w%/v. In this second embodiment, the small chemical molecule is any API with a solubility in alcohol of less than 0.5% w/v, particularly less than 0.1% w/v, and even more particularly less than 0.01% w/v, defined as an alcohol-insoluble API, such as cisplatin, carboplatin, oxaliplatin, pemetrexed disodium, azacytidine, beclomethasone dipropionate, tobramycin, adecyl bromide, etc.

More specifically, relative to the weight of the dry powder inhalation formulation, the dry powder inhalation formulation according to the second embodiment contains 0.1 wt% to 95 wt%, preferably 1 wt% to 90 wt%, more preferably 1 wt% to 85 wt% of the at least one API.

Therefore, relative to the total weight of the dry powder inhalation formulation, the dry powder inhalation formulation contains 0.1 wt% to 95 wt%, preferably 1 wt% to 90 wt%, more preferably 1 wt% to 85 wt%, such as 1 wt% to 60 wt% or 1 wt% to 50 wt% of the alcohol-insoluble API.

In a preferred embodiment of the invention, the dry powder inhalation formulation comprises lipid matrix particles in which the alcohol-insoluble API is dispersed.

In a third embodiment of the invention, the active agent is a macromolecule. In this third embodiment of the invention, the dry powder inhalation formulation contains 0.1 wt% to 95 wt%, preferably 1 wt% to 90 wt%, more preferably 1 wt% to 85 wt% of the at least one API, relative to the weight of the dry powder inhalation formulation.

Therefore, relative to the total weight of the dry powder inhalation formulation, the dry powder inhalation formulation contains 0.1 wt% to 95 wt%, preferably 1 wt% to 90 wt%, more preferably 1 wt% to 85 wt%, for example 1 wt% to 60 wt% or 1 wt% to 50 wt% of the macromolecules.

In another preferred embodiment, the formulation according to the invention further comprises a lung-retention prolonging excipient, such as a polyethylene glycol-modified excipient or a polysaccharide, such as chitosan or dextran.

Preferably, according to the invention, the extended lung retention excipient is a polyethylene glycol-modified excipient and is present in an amount of 0.1 wt% to 20 wt%, preferably 0.2 wt% to 10 wt%, more preferably 0.5 wt% to 5 wt% relative to the total weight of the dry powder inhalation formulation.

More specifically, according to the present invention, the polyethylene glycol excipient is derived from vitamin E or phospholipids, such as tocopheryl polyethylene glycol succinate (TPGS) or distearate ethanolamine phosphate polyethylene glycol 2000 (DSPE-mPEG-2000).

In another preferred embodiment, the formulation according to the invention further comprises one or more excipients, such as excipients that improve the physicochemical and/or aerodynamic properties of inhaled dry powder.

The term "one or more excipients" refers to compounds selected from: sugar alcohols; polyols (e.g., sorbitol, mannitol, and xylitol); crystalline sugars, including monosaccharides (e.g., glucose, arabinose) and disaccharides (e.g., lactose, maltose, sucrose, dextrose, trehalose, maltitol); inorganic salts (e.g., sodium chloride and calcium carbonate); organic salts (e.g., sodium lactate, potassium phosphate or sodium phosphate, sodium citrate, urea); polysaccharides (e.g., dextran, chitosan, starch, cellulose, hyaluronic acid, and their derivatives); oligosaccharides (e.g., cyclodextrin and dextrin); titanium dioxide; silicon dioxide; magnesium stearate; lecithin; amino acids (e.g., leucine, isocyanate, iodine ... Leucine, histidine, threonine, lysine, valine, methionine, phenylalanine; derivatives of amino acids (such as acesulfame potassium, aspartame); lauric acid or its derivatives (e.g., esters and salts); palmitic acid or its derivatives (e.g., esters and salts); stearic acid or its derivatives (e.g., esters and salts); erucic acid or its derivatives (e.g., esters and salts); benzyl acid or its derivatives (e.g., esters and salts); sodium stearoyl fumarate; sodium stearoyl lactylate; phosphatidylcholine; phosphatidylglycerol; natural and synthetic lung surfactants; lauric acid and its salts (e.g., sodium lauryl sulfate, magnesium lauryl sulfate); triglycerides; glycolipids; phospholipids; cholesterol; talc.

In some preferred embodiments, the excipient is mannitol, dextrose, or lactose.

In some preferred embodiments, the excipient is phospholipid or cholesterol.

In some preferred embodiments, the excipients are mannitol, dextrose, hyaluronic acid, lactose, phospholipids, or cholesterol.

In some preferred embodiments, the excipients are mannitol, dextran, phospholipids, or cholesterol.

In some embodiments, the at least one excipient is a carrier.

In a preferred embodiment of the formulation of the present invention, the formulation is in the form of fine particles having a geometric particle size distribution (PSD) d ₅₀ of less than or equal to 30 μm, preferably less than 15 μm, preferably less than or equal to 10 μm, and preferably less than or equal to 5 μm.

In another preferred embodiment of the formulation of the present invention, the formulation is in the form of fine particles having a geometric particle size distribution (PSD) d ₉₀ of less than or equal to 60 μm, preferably less than or equal to 30 μm, more preferably less than or equal to 15 μm, more preferably less than or equal to 10 μm and more preferably less than or equal to 7 μm.

In another preferred embodiment of the formulation of the present invention, the formulation is in the form of particles having a volume average diameter D[4,3] of less than or equal to 40 μm, preferably less than or equal to 20 μm, more preferably less than or equal to 15 μm, preferably less than or equal to 10 μm, and preferably less than or equal to 6 μm.

Preferably, according to the invention, the improved excipient and/or the excipient are present in an amount of 0.1 wt% to 80 wt%, preferably less than 70 wt%, more preferably less than 60 wt%, and particularly less than 50 wt%, for example, less than 50 wt%, relative to the total weight of the dry powder inhalation formulation.

The term “fine particulate dose” or “FPD” generally refers to the mass of particles with an aerodynamic diameter of less than 5 μm relative to the mass of the nominal dose (i.e., the mass of the dose loaded in the inhalation device).

Fine particle dose or fine particle fraction represents the fraction of a drug formulation that can be deeply inhaled and is theoretically used to assess pharmacological activity (Dunbar et al., Kona 16:7-45, 1998).

In an advantageous embodiment of the invention, the formulation is in the form of fine particles having a median mass aerodynamic diameter (MMAD) of less than or equal to 6 μm, preferably less than or equal to 5 μm, and more preferably less than or equal to 4 μm.

MMAD refers to the diameter of particles deposited in an impactor, where 50% (w/w) of the particles have a smaller diameter and 50% (w/w) have a larger diameter.

The term "aerodynamic diameter" or "dae" for a particle can be defined as the diameter of a sphere with a unit density (i.e., density of 1) that has the same settling velocity in still air as the particle under consideration. "Dae" provides a useful measurement of inhalable particles and takes into account factors affecting their aerodynamic properties. "Dae" can be used to compare particles of different physical sizes, taking into account their density, shape, and geometry.

The methods used to measure “dae” are those described in the European or United States Pharmacopeia, which use an impactor or impact device, such as a glass impactor, a multistage liquid impactor (MsLI), an Anderson cascade impactor, or a next-generation impactor (NGI). These allow for the measurement of the aerodynamic properties of DPI formulations (including MMAD, geometric standard deviation, lung deposition pattern, fine particle dose, and fine particle fraction) under simulated respiratory conditions.

The total dose of particles with an aerodynamic diameter of less than 5 μm can be calculated from the collection efficiency curve by interpolation and is considered as fine particle dose (FPD) or fine particle fraction (FPF), expressed as a percentage of the nominal API dose (i.e. the dose contained in the DPI device).

Preferably, the dry powder inhalation formulation according to the invention is packaged, for example, in a blister pack or capsule, for use in a dry powder inhaler or in a sealed and/or disposable dry powder inhaler.

In one specific embodiment, the active agent is a small chemical molecule possessing bronchodilatory activity, glucocorticoid activity, anti-inflammatory activity, and anti-infective activity (e.g., antibiotic, anti-tuberculosis, antifungal, antiviral), etc.

In another specific embodiment, the small chemical molecule is any active pharmaceutical ingredient that is absorbed by the lungs for systemic or local treatment, such as budesonide, salbutamol, fluticasone, beclomethasone, mometasone, cisasonide, formoterol, salbutamol, afortol, indacaterol, olodaterol, salmeterol, ipratropium, adine, glycopyrronium, tiotropium bromide, fumedoxomil, mometasone, cisasonide, formoterol, afortol, ibuprofen, tobramycin, vancomycin, tetrahydrolipresteen, clarithromycin, isoniazid, rifampin, pyrazinamide, itraconazole, voriconazole, aztreonam, ethambutol, and streptomycin. Kanamycin, Amikacin, Colistin, Polymyxin E Sodium Mesylate, Chloramphenicol, Ciprofloxacin, Rifapentine, Doxycycline, Cycloserine E, Ethionamide, Gatifloxacin, Levofloxacin, Moxifloxacin, Ofloxacin, Fosfomycin, Para-aminosalicylic acid, Denofossodium Tetrasodium, Lancovirate, Ribavirin, Zanamivir, Laminavir, Rupmitine, Pentamidine, Amphotericin B, Posaconazole, Isaconazole, Capofungin, Micafungin, Anifungin, Iloprost, Levothyroxine, their salts, solvates, hydrates, polymorphs and esters, combinations thereof, analogs and derivatives thereof.

In another embodiment, the active agent is a macromolecule, such as a peptide, protein, antibody, antibody fragment, nanobody, or nucleic acid.

Preferably, according to the present invention, the macromolecule is insulin, proinsulin, synthetic insulin, semi-synthetic insulin, bevacizumab, pembrolizumab, atezolizumab, nivolumab, ipilimumab, Toll-like receptor agonist, auxin-releasing peptide, IgG monoclonal antibody, small interfering RNA (siRNA), alfa streptase, cyclosporine A, α-1 antitrypsin, interleukin antagonist, interferon-α, interferon-β, interferon-γ, interferon-ω, interleukin-2, anti-IgE mAb, catalase, calcitonin, parathyroid hormone, human growth hormone, insulin-like growth factor-I, heparin, rhG-CSF, GM-CSF, Epo-Fc, FSH-Fc, sFc-γRIIb, mRNA.

In another preferred embodiment, the active agent is an antitumor agent, such as for lung cancer and lung tumors.

Lung cancer is the leading cause of cancer death and morbidity worldwide. In most cases, lung cancer is diagnosed at an advanced stage. Therefore, patients often already have metastases in the lungs or other organs, a condition known as extrapulmonary metastasis. Treatment methods are primarily used in combination, including surgery, radiation therapy, chemotherapy, targeted therapy, and immunotherapy.

Chemotherapy is used in up to 60% of lung cancer patients, primarily at advanced stages of the disease. Chemotherapy is currently administered via intravenous injection or infusion, or orally, a systemic route, also known as systemic chemotherapy. Due to (i) the widespread distribution of chemotherapeutic agents throughout the body and (ii) the lack of selectivity for cancer cells, chemotherapy causes severe systemic toxicity. Therefore, oncologists are in dire need of new, more effective, and better-tolerated treatment methods.

Chemotherapy is actually about finding the maximum amount of antitumor agents to be administered by minimizing the irreversible toxic side effects and adverse side effects caused by chemotherapy.

Therefore, very complex treatment protocols are anticipated, which combine very different treatments and behaviors (such as surgery) with a first cycle of radiation therapy, followed by complex cycles of injection or intravenous infusion of antitumor drugs, which may be the same or different for each cycle and reduced or adjusted based on the side effects identified in the patient's body.

Each antitumor agent or molecule has dose-limiting toxicities (DLTs), such as nephrotoxicity and neurotoxicity, which require the introduction of rest periods into the treatment regimen to allow the patient's body to recover from adverse side effects when they are irreversible.

In addition, antitumor agents have a half-life, commonly referred to as their half-life. Half-life is the time required for the concentration or amount of a drug in the body to decrease by half. The half-life of antitumor agents typically ranges from 12 to 36 hours, but it can be short because it represents the duration of human exposure to the beneficial effects of the antitumor agent.

However, due to dose-limiting toxicity (DLT) of antitumor agents, the amount of dose administered over several administration periods remains limited and, as previously mentioned, should be separated from each other by rest periods or withdrawal periods.

The combination of half-life and dose-limiting toxicity, along with the fact that antitumor agents are administered orally, by injection, or by intravenous infusion, results in low concentrations of antitumor agents that effectively reach the site of solid tumors, limited effects on the tumor itself, and high systemic toxicity to the patient.

Another limitation of antitumor agents is the cumulative dose, which is the total dose resulting from repeated exposure of the same part of the body or the whole body to the antitumor agent.

For cisplatin administered intravenously, the cumulative dose over 4 to 6 cycles is 300 mg/ ^m² .

These considerations have prompted the researchers of this invention to develop more targeted therapies and on-site injections/infusions.

Inhalation therapy comes in different types, and inhalers in the form of nebulizers can be distinguished by different ways of delivering APIs to the respiratory tract. Inhalers can also be of different types, with dry powder inhalers (DPIs) being one of them.

Compared to nebulizers, dry powder inhalers (DPIs) are well-suited for chemotherapy. They allow for the administration of high doses of APIs, but also allow for the administration of poorly water-soluble compounds (i.e., most chemotherapy agents in cancer). Furthermore, DPIs limit aerosol pollution due to (i) they are initiated and driven solely by the patient's inhalation flow and (ii) negligible exhaled drug doses. Finally, DPIs can be designed as single-use devices.

Therefore, there is a need to develop pulmonary routes for antitumor agents and adapt them to current inhalation devices and formulations used in clinical trials to deliver effective anticancer therapies to lungs carrying tumors.

Prolonging the retention curve of antitumor agents in the lungs and ensuring that absorption is not too rapid has many advantages. However, achieving a prolonged release curve remains challenging because the lungs have a high absorption surface area, leading to rapid systemic absorption of the drug.

As previously mentioned, cisplatin-based controlled-release DPI formulations have been developed to have high drug loading and high particle size fraction, resulting in controlled release and prolonged pulmonary retention, leading to low systemic distribution (Levet et al., Int J Pharm [International Journal of Pharmaceutics] 2017). According to this literature, in a mouse model of lung cancer, this method resulted in in vivo tumor response comparable to half-dose intravenous regimens (1.0 mg/kg and 0.5 mg/kg, respectively). The intravenous regimen yielded good results for extrapulmonary metastases, while inhaled cisplatin appeared to be more active against lung tumors.

DPI formulations composed of paclitaxel-based nanocarriers have also been developed, exhibiting increased residence time in the lungs, limited systemic distribution, and specific targeting of lung cancer cells via folate receptor (FR) targeting (Rosière et al., Int JPharm [International Journal of Pharmaceutics] 2016; Rosière et al., Mol Pharm [Molecular Pharmaceutics], 2018). FR, especially FR-α, is overexpressed on the surface of cancer cells in many lung tumors (i.e., over 70% of adenocarcinomas) and is a promising membrane receptor for targeting lung cancer. In a mouse model of lung cancer, significantly longer survival was observed with inhaled therapy targeting FR in combination with intravenous administration (in the commercial form of paclitaxel) compared to intravenous administration alone.

To recover, despite encouraging preclinical results in terms of pharmacokinetic distribution and safety from the use of DPI chemotherapy, their efficacy remains quite limited.

While there are many advantages to achieving a prolonged release curve for antitumor agents, this challenge is exacerbated in the case of inhaled chemotherapy because the lungs have a significantly large absorption surface, causing the API to be rapidly absorbed and delivered to the systemic circulation.

Furthermore, because the lungs have strong defense mechanisms, the elimination of APIs in inhaled therapy is quite rapid due to the lungs' highly efficient clearance mechanisms against foreign deposited particles.

Sebti et al. (Sebti et al., Eur J Pharm Biopharm, 2006a; Sebti et al., Eur J Pharm Biopharm, 2006b) developed solid lipid macroparticles with budesonide and a lipid matrix composed of cholesterol, but no delay in budesonide release was observed.

To this end, Depreter et Amighi (Depreter and Amighi, Eur J Pharm Biopharm [European Journal of Pharmaceutics and Biopharmaceutics] 2010) developed insulin microparticles coated with a lipid matrix composed of cholesterol, for which only a slight delay in insulin release was observed.

To this end, as previously mentioned, Levet et al. have worked to extend the retention time in the lungs, thereby improving the efficiency of inhaled chemotherapy and have identified a lipid matrix composed of tristearin and TPGS as a promising candidate.

Furthermore, even though lipid-derived excipients used to form the matrix are superior to polymers, only a few substances capable of forming lipid matrices are permitted and accepted for use in inhaled pharmaceutical compositions for toxicity reasons, compared to other therapeutic routes (Pilcer and Amighi, Int J Pharm [International Journal of Pharmaceutics] 2010).

Unfortunately, while tristearate-based matrices are considered promising, their market availability as pharmaceutical compounds is rather limited, whereas the availability of at least one triglyceride selected from the group consisting of monohydroxystearin, dihydroxystearin, trihydroxystearin, and mixtures thereof is much more extensive.

Furthermore, the tristearin-based matrix is not promising for controlling the release profile of voriconazole, and the preparation method (i.e., spray drying) and excipients are similar to those described by Levet et al. for cisplatin.

Furthermore, according to the present invention, it has been determined that at least one triglyceride selected from the restriction group according to the present invention, in addition to obtaining a sustained release profile and prolonging the retention of chemotherapy in the lungs, also allows for a significant increase in lung deposition rate relative to the closest triglyceride, tristearate.

Preferably, the antitumor agent is cisplatin, carboplatin, oxaliplatin, docetaxel, paclitaxel, pemetrexed, etoposide, or vinorelbine.

Other embodiments of the dry powder inhalation formulation according to the invention are mentioned in the appended claims.

This invention relates to a method for preparing a dry powder inhalation formulation according to the present invention, comprising the following steps:

a) With or without a solvent, the one or more APIs are mixed with a predetermined amount of at least one triglyceride, said triglyceride being selected from the group comprising monohydroxystearin, dihydroxystearin, trihydroxystearin, and mixtures thereof.

b) The aforementioned mixture is used to produce inhalable particles by bottom-up or top-down methods, such as a micronization step, spray drying of a suspension or solution, spray condensation of an active drug or API in a solution of the at least one triglyceride, or extrusion, followed by jet milling of a physical mixture of the API and the at least one triglyceride.

For example, the present invention relates to a method for preparing a dry powder inhalation formulation according to the present invention, comprising the steps of: suspending or dissolving powder of one or more APIs in a predetermined amount of at least one triglyceride to form a particulate suspension or solution of one or more APIs (e.g., by melt and/or by extrusion), said triglyceride being selected from the group comprising monohydroxystearin, dihydroxystearin, trihydroxystearin and mixtures thereof, and then reducing the particle size of the solution or suspension of one or more APIs obtained after cooling at high speed and/or high pressure (e.g., by spray-coagulation) or extrusion to obtain said dry powder inhalation formulation (e.g., by jet milling).

In another example, the manufacturing method includes the following steps:

a) Mix at least one API with at least one triglyceride to form a homogeneous mixture.

b) Extruding the homogeneous mixture at a suitable temperature using a (twin-screw) extruder to obtain a homogeneous lipid matrix containing the API.

c) Cut the extrudate to obtain coarse particles/cylinders.

d) Optionally, the triglycerides are converted into a stable polymorph by storing the coarse particles/cylinders under appropriate storage conditions, and

e) Grind the particles using a suitable grinder to obtain inhalation microparticles (DPIs) containing the API and at least one of the triglycerides.

In another instance, the present invention relates to a method for preparing a dry powder inhalation formulation according to the present invention, comprising the following steps:

a) Suspending or dissolving powders of one or more APIs in a solvent to form a particulate suspension or solution of one or more APIs.

b) Optionally, the particle size of one or more APIs is reduced under cooling during high-speed and/or high-pressure homogenization to form a suspension of one or more APIs particles with reduced size.

c) Mixing a predetermined amount of at least one triglyceride selected from the group consisting of monohydroxystearin, dihydroxystearin, trihydroxystearin, and mixtures thereof with the microcrystalline suspension or solution in a solvent to obtain a mixture of the at least one API and the at least one triglyceride.

d) Spray drying of the mixture of the at least one API and the at least one triglyceride to obtain the dry powder inhalation formulation.

In a preferred embodiment of the method according to the invention, the high speed for reducing size is 10,000-30,000 rpm, preferably 15,000-26,000 rpm, and the application time is 8-15 minutes, preferably 9-12 minutes.

Preferably, according to the invention, the high pressure used for the homogenization step is gradually increased from a first pressure of 2000-10000 psi, preferably 4000-6000 psi, through a predetermined number of pre-grinding cycles of 8-12, preferably 9-11, to a second pressure of 8000-12000 psi, preferably 9000-11000 psi, through a predetermined number of pre-grinding cycles of 8-12, preferably 9-11, to a third pressure of 18000-24000 psi, preferably 19000-22000 psi, through a predetermined number of pre-grinding cycles of 18-22, preferably 19-21.

In a particularly preferred embodiment of the invention, the microcrystals of the microcrystalline suspension have a geometric particle size distribution (PSD) d ₅₀ of less than or equal to 30 μm, preferably less than 15 μm, preferably less than or equal to 10 μm, and preferably less than or equal to 5 μm.

In another particularly preferred embodiment of the invention, the microcrystals of the microcrystalline suspension have a geometric particle size distribution (PSD) d ₉₀ of less than or equal to 60 μm, preferably less than or equal to 30 μm, more preferably less than or equal to 15 μm, more preferably less than or equal to 10 μm, and more preferably less than or equal to 7 μm.

In another particularly preferred embodiment of the invention, the microcrystals of the microcrystalline suspension have a volume average diameter D[4,3] of less than or equal to 40 μm, preferably less than or equal to 20 μm, more preferably less than or equal to 15 μm, preferably less than or equal to 10 μm, and preferably less than or equal to 6 μm.

In an advantageous preferred embodiment according to the invention, a polyethylene glycol-based excipient or other excipient is further added.

Other embodiments of the method according to the invention are mentioned in the appended claims.

This invention also relates to the use of dry powder inhalation formulations in the treatment of lung diseases.

Preferably, it is anticipated that the use of the invention can be used to treat local lung diseases such as asthma, COPD, lung infections (e.g., cystic fibrosis, aspergillosis, tuberculosis, etc.) or systemic diseases such as diabetes, pain, etc.

In a variant of the invention, the dry powder inhaled chemotherapy formulation is used in multiple therapies for the treatment of lung cancer such as any lung tumor, such as lung metastases, such as osteosarcoma metastases, small cell lung cancer, or non-small cell lung cancer.

Advantageously, the multiple therapy includes a primary therapy selected from the group consisting of: intravenous or infusion chemotherapy, immunotherapy, tumor ablation surgery, ablation surgery for the partial or complete removal of organs carrying tumors, curative surgery, radiotherapy and combinations thereof, and one or more inhaled chemotherapy as an additional therapy.

The present invention also relates to corresponding treatment methods.

Detailed Implementation

Other features and advantages of the invention will become apparent from the following non-limiting description and with reference to examples and drawings.

In the accompanying figures, Figure 1 shows the FPF values (%) of the comparative example (composition F5 in Levet et al.) and examples 1 and 2 (mean ± SD, n = 2-3).

Figure 2 shows the release profiles of cisplatin from the inhalable portion of the DPI formulations prepared according to Examples 1 and 2, compared to a comparative formulation consisting only of cisplatin microparticles.

Figure 3 shows the FPF values (mean ± SD, n = 3 and 1, respectively) of Example 10 compared to a comparative formulation consisting only of pemetrexed microparticles, indicating that Example 10 is superior to the conventional DPI formulation in terms of lung deposition.

Figure 4 shows the release curves (mean ± SD, n = 3) of pemetrexed from the inhalable portion of the DPI formulation prepared according to Example 10, compared to a comparative formulation consisting solely of pemetrexed microparticles.

Figure 5 shows the FPF values (mean ± SD) of Example 13, demonstrating the high lung deposition rate of the insulin-based DPI composition disclosed in this invention.

Figure 6 shows the release curves of insulin from the inhaled portion of the DPI formulation prepared according to Example 13 (up to 240 minutes, mean ± SD, n = 2) compared to the comparative formulation described by Depreter et al. (up to 180 minutes) (B).

Figure 7 shows the release curves (mean ± SD, n = 3) of cisplatin from the inhalable portion of the DPI formulation prepared according to Example 16, compared to a comparative formulation composed of cisplatin microparticles.

Figure 8 shows the FPF values (mean ± SD, n = 3) of Examples 19 and 20 compared to Comparative Examples BUD-TS4 and BUD-TS5. (***) p < 0.001, t-test, indicating that the budesonide composition disclosed in this invention is superior to other triglyceride-based budesonide DPI formulations in terms of lung deposition.

Figure 9 shows the release profiles (n=1) of budesonide from the inhalable portion of micronized budesonide comparative examples and DPI formulations prepared according to Examples 19, 20 and 21, demonstrating the controlled release profiles of budesonide from the compositions of the present invention and the possibility of adjusting the release profile by adjusting the drug/lipid ratio.

Example.

Example 1. Preparation of cisplatin dry powder formulation for inhalation n°1

In short, coarse cisplatin microcrystals obtained from bulk powder (Shanghai Jinhe Bio-technology Co., Ltd., Shanghai, PRC) were first suspended in 50 mL of isopropanol to achieve a concentration of 5% w/v. The microcrystals were then reduced in size by high-speed (10 min, 24000 rpm) homogenization (X620 motor and T10 dispersion shaft, Ingenieurbüro CAT M. Zipperer GmbH, Staufen, Germany) and high-pressure homogenization (EmulsiFlex-C5 high-pressure homogenizer, Avestin Inc., Ottawa, Canada) at 5000 psi, then 10000 psi, for 10 pre-grinding cycles each, followed by 20 grinding cycles at 20000 psi. A heat exchanger was connected to the homogenization valve and maintained at -15°C using an F32-MA cooling circulator (ulabo GmbH, Seeelbach, Germany). At the end of the process, equal portions of the sample were removed from the suspension to measure the particle size distribution (PSD) of cisplatin crystals by laser diffraction (see below).

Then, hydrogenated castor oil (BASF, Ludwigshafen, Germany) and TPGS (Sigma-Aldrich, St. Louis, USA) dissolved in heated isopropanol were added to the microcrystalline suspension to obtain a final concentration of 1.32% w/v cisplatin and 0.68% w/v hydrogenated castor oil/TPGS (99:1 w/w) mixture, which was then spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain the human DPI formulation.

The operating parameters used during spray drying are as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air flow rate of 800 L/min, and drying air flow rate of 35 ^m³ /h. The unit is equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

The geometrical PSD of bulk cisplatin (cisplatin microparticles from the size reduction process and cisplatin dry powder formulations) was measured as suspended and individualized particles. Measurements were performed using a Mastersizer 3000 laser diffractometer (Malvern Instruments Ltd., Worcestershire, UK) connected to a Hydro MV dispenser (Malvern Instruments Ltd.) equipped with a 40W ultrasonic probe. Bulk cisplatin and aliquots from the size reduction process were measured in cisplatin-saturated isopropanol. Cisplatin dry powder formulations were pre-dispersed, vortexed, and measured in cisplatin-saturated 0.1% w/v poloxamer 407 (BASF, Ludwigshafen, Germany) 0.9% NaCl aqueous solution. PSD is expressed as the volume median diameter d (0.5) (where 50% of the particles are below the represented diameter), the volume average diameter D [4,3], and the percentage of fine particles (% of particles below 5 μm) determined by the cumulative undersize curve. PSD results are within the range anticipated according to the present invention.

Example 2. Preparation of cisplatin dry powder formulation for inhalation n°2

In short, coarse cisplatin microcrystals obtained from bulk powder (Shanghai Jinhe Bio-technology Co., Ltd., Shanghai, PRC) were first suspended in 50 mL of isopropanol to achieve a concentration of 5% w/v. The microcrystals were then reduced in size by high-speed (10 min, 24000 rpm) homogenization (X620 motor and T10 dispersion shaft, Ingenieurbüro CAT M. Zipperer GmbH, Staufen, Germany) and high-pressure homogenization (EmulsiFlex-C5 high-pressure homogenizer, Avestin Inc., Ottawa, Canada) at 5000 psi, then 10000 psi, for 10 pre-grinding cycles each, followed by 20 grinding cycles at 20000 psi. A heat exchanger was connected to the homogenization valve and maintained at -15°C using an F32-MA cooling circulator (ulabo GmbH, Seeelbach, Germany). At the end of the process, equal portions of the sample were removed from the suspension to measure the particle size distribution (PSD) of cisplatin crystals by laser diffraction (see below).

Then, hydrogenated castor oil (BASF, Ludwigshafen, Germany) and TPGS (Sigma-Aldrich, St. Louis, USA) dissolved in heated isopropanol were added to the microcrystalline suspension to obtain a final concentration of 1.0% w/v cisplatin and 1.0% w/v hydrogenated castor oil/TPGS (99:1 w/w) mixture, which was then spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavier, Switzerland) to obtain the human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air 800 L/min, and drying air flow rate 35 ^m³ /h. The unit was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

The geometric PSD of bulk cisplatin (cisplatin microparticles from the size reduction process and cisplatin dry powder formulations) was measured as suspended and individualized particles. As described in Example 1, this was performed using a Mastersizer 3000 laser diffractometer (Malvern Instruments Ltd., Worcestershire, UK) connected to a Hydro MV dispenser (Malvern Instruments Ltd.) equipped with a 40W ultrasonic probe.

Example 3. - Deposition rate analysis of cisplatin dry powder formulations prepared according to Examples 1 and 2.

The fine particulate fraction values of the formulations according to the comparative example (composition F5 in Levet et al.) and according to Examples 1 and 2 have been analyzed.

The fine particle fraction (FPF) – which is the percentage of cisplatin-based particles with an aerodynamic diameter (dae) less than 5 μm relative to the recovered dose – and the aerodynamic PSD, characterized by the mass median aerodynamic diameter (MMAD), were determined using the MsLI (Copley Scientific, Nottingham, UK) as described in European Pharmacopoeia 8.0. (2014). 20 mg of each DPI formulation, pre-sieved through a 355 mm stainless steel sieve, was weighed into No. 3 HPMC capsules (Quali-V-I, Qualicaps, Madrid, Spain) (Comparative Example and according to Example 2) and deposited in the MsLI using an RS.01 dry powder inhaler (RPCPlastiape, Osnago, Italy), which was attached to the inhalation port with its adapter (n=3).

A deposition rate of 100 ± 5 L/min was obtained by connecting two HCP5 air pumps (Copley Scientific, Nottingham, UK) in series with a TPK critical flow controller (Copley Scientific, Nottingham, UK) and measuring it using a DFM3 flow meter (Copley Scientific, Nottingham, UK).

At this flow rate, the cutoff diameters between the stages of MsLI are 10.0, 5.3, 2.4, 1.3, and 0.4 mm. The micropore collector (MOC) filter (i.e., stage 5) comprises a Fluoropore 9cm PTFE membrane with a pore size of 0.45 mm bonded to a high-density polyethylene carrier (Merck Millipore, Darmstadt, Germany). As required by European Pharmacopoeia 8.0. (2014), a critical flow controller is used to ensure a deposition time of 2.4 s and a critical flow rate of <0.5 for the P3/P2 ratio at 100 L/min.

Following impact, the first four stages of MsLI were subjected to a first rinse using 20 mL of pre-filled 0.5% w/v poloxamer 407 in ultrapure water/isopropanol (60:40 v/v) as the diluent, a second rinse using 25 mL of DMF, and a third rinse using the diluent, adjusted to 100.0 mL and sonicated for 30 min. Drug deposition in the capsule, device, inhalation port, and MOC filter was determined after dissolution with 100.0 mL of the diluent and sonication for 30 min. Impact mass in each stage was determined by quantifying cisplatin content using empirically validated electrothermal atomic absorption spectrometry (ETAAS) as described by Levet et al. (Levet, Int J Pharm [International Journal of Pharmaceutics] 2016).

The results were then plotted in Copley Inhaler Trial Data Analysis Software 1 (Copley Scientific, Nottingham, UK) to obtain an FPD below 5 μm. This was done by interpolating the recovery mass with the cutoff diameter of the corresponding stage. FPF is expressed as a percentage of the nominal dose.

Figure 1 shows the FPF values (%) (mean ± SD, n = 2-3) of the comparative example (composition F5 in Levet et al.) and examples 1 and 2, indicating that the cisplatin composition disclosed in this invention is superior to other triglyceride-based cisplatin DPI formulations in terms of lung deposition.

Example 4. Analysis of the dissolution rate of cisplatin from cisplatin dry powder formulations prepared according to Examples 1 and 2.

Dissolution properties of DPI formulations were established using the method described by Levet et al. (Levet et al., Int J Pharm [International Journal of Pharmaceutics] 2016). This method, derived from the paddle disc method of USP39, utilizes a modified V-type dissolution device for transdermal patches. Cisplatin release profiles were determined from the entire inhalable fraction (dae ≤ 5 μm) of the DPI formulations selected using a rapid screening impactor (FSI, Copley Scientific, Nottingham, UK). Appropriate masses of each DPI formulation, equivalent to a 3 mg cisplatin deposition dose, were weighed into HPMC capsules No. 3 (Quali-V-I Qualicaps, Madrid, Spain). The deposits were then performed using an RS.01 DPI device (RPC Plastiape) onto a hydrophobic PTFE membrane filter (Merck Millipore, Darmstadt, Germany) with a 0.45 mm pore size, where the FSI (2.4 s, 100 L/min) was equipped with a corresponding pre-separator insert. The Fluoropore filter with the deposited powder facing upwards was then covered with a 0.4 mm hydrophilic polycarbonate filter (Merck-Milipore, Germany) and secured to the petri dish-PTFE disc assembly (Copley, Nottingham, UK) using the provided clips and PTFE screen. The disc assembly was then immersed in the dissolution vessel of an AT7 dissolution apparatus (Sotax AG, Aesch, Switzerland) containing 400 mL of modified simulated lung fluid (mSLF) (Son and McConville, 2009) – a medium simulating a combination of lung electrolytes and surfactants.

Dissolution tests were conducted at 37±0.2℃ and pH 7.35±0.05, depending on the tank conditions. The paddle rotation speed (with a center-to-center distance of 25±2 mm between the paddle and the disc assembly) was set at 50±4 rpm. A 2.0 mL sample volume was filtered through a 0.22 mm cellulose acetate injection filter (VWR, Leuven, Belgium) at predetermined intervals between 2 minutes and 24 hours, and replaced with 2.0 mL of free, preheated mSLF.

At the end of the dissolution assay, the disc assembly was opened into the dissolution vessel and sonicated for 30 minutes to establish a 100% cisplatin dissolution value.

Figure 2 shows the release profiles of cisplatin from the inhalable portion of the DPI formulations prepared according to Examples 1 and 2, compared to a comparative formulation consisting only of cisplatin microparticles.

Example 5. Preparation of Insulin Dry Powder Formulation n°1

Insulin was first suspended in isopropanol (2% w/v) and the powder was dispersed by sonication in a Branson 2510 bath at 40 kHz for 10 minutes. Particle size was then reduced using an EmulsiFlex-C5 high-pressure homogenizer (Avestin Inc., Ottawa, Canada). The insulin suspension was first subjected to pre-grinding low-pressure homogenization cycles to further reduce particle size (10 cycles at 7000 PSI, 10 cycles at 12,000 PSI). Finally, HPH was applied at 24,000 PSI for 30 cycles. These cycles were performed by directly recirculating the treated suspension into the sample bath (closed loop). Because HPH causes a temperature increase in the sample (30°C after 20 cycles at 24,000 PSI), all operations were performed using a heat exchanger placed before the homogenization valve, maintaining the sample temperature at 5 ± 1°C.

At the end of the process, equal portions of the sample were removed from the suspension to measure the particle size distribution (PSD) of insulin crystals by laser diffraction (see below).

Then, hydrogenated castor oil (BASF, Ludwigshafen, Germany) and TPGS (Sigma-Aldrich, St. Louis, USA) dissolved in heated isopropanol were added to the microcrystalline suspension to obtain a final concentration of 1.0% w/v insulin and a 1.0% w/v hydrogenated castor oil/TPGS (99:1 w/w) mixture, which was then spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavier, Switzerland) to obtain the human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air 800 L/min, and drying air flow rate 35 ^m³ /h. The unit was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

The geometric PSD of bulk insulin (insulin microparticles and dry powder formulations from the size reduction process) was measured as suspended and individualized particles, and is within the scope of this invention. As described in Example 1, this was performed using a Mastersizer 3000 laser diffractometer (Malvern Instruments Ltd., Worcestershire, UK) connected to a Hydro MV dispenser (Malvern Instruments Ltd.) equipped with a 40W ultrasonic probe.

Example 6. Preparation of Insulin Dry Powder Formulation n°2

Insulin was first suspended in isopropanol (2% w/v) and the powder was dispersed by sonication in a Branson 2510 bath at 40 kHz for 10 minutes. Particle size was then reduced using an EmulsiFlex-C5 high-pressure homogenizer (Avestin Inc., Ottawa, Canada). The insulin suspension was first subjected to pre-grinding low-pressure homogenization cycles to further reduce particle size (10 cycles at 7000 PSI, 10 cycles at 12,000 PSI). Finally, HPH was applied at 24,000 PSI for 30 cycles. These cycles were performed by directly recirculating the treated suspension into the sample bath (closed loop). Because HPH causes a temperature increase in the sample (30°C after 20 cycles at 24,000 PSI), all operations were performed using a heat exchanger placed before the homogenization valve, maintaining the sample temperature at 5 ± 1°C.

At the end of the process, equal portions of the sample were removed from the suspension to measure the particle size distribution (PSD) of insulin crystals by laser diffraction (see below).

Then, hydrogenated castor oil (BASF, Ludwigshafen, Germany) and TPGS (Sigma-Aldrich, St. Louis, USA) dissolved in heated isopropanol were added to the microcrystalline suspension to obtain a final concentration of 1.5% w/v insulin and a 0.5% w/v hydrogenated castor oil/TPGS (99:1 w/w) mixture, which was then spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavier, Switzerland) to obtain the human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air 800 L/min, and drying air flow rate 35 ^m³ /h. The unit was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

The geometric PSD of bulk insulin (insulin microparticles and dry powder formulations from the size reduction process) was measured as suspended and individualized particles, and is within the scope of this invention. As described in Example 1, this was performed using a Mastersizer 3000 laser diffractometer (Malvern Instruments Ltd., Worcestershire, UK) connected to a Hydro MV dispenser (Malvern Instruments Ltd.) equipped with a 40W ultrasonic probe.

Example 7. Preparation of Budesonide dry powder formulation for inhalation n°1

First, budesonide (1% w/v) was dissolved in isopropanol under magnetic stirring. Then, hydrogenated castor oil (BASF, Ludwigshafen, Germany) and TPGS (Sigma-Aldrich, St. Louis, USA) dissolved in heated isopropanol were added to the microcrystalline suspension to obtain a final concentration of 1.0% w/v budesonide and a 1.0% w/v hydrogenated castor oil/TPGS (99:1 w/v) mixture, which was then spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain the human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air 800 L/min, and drying air flow rate 35 ^m³ /h. The unit is equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

The geometrical PSD of the bulk budesonide dry powder formulation was measured as suspended and individualized particles, and within the scope of this invention. As described in Example 1, the measurement was performed using a Mastersizer 3000 laser diffractometer (Malvern Instruments Ltd., Worcestershire, UK) connected to a Hydro MV dispenser (Malvern Instruments Ltd.) equipped with a 40W ultrasonic probe.

Example 8. Preparation of Budesonide dry powder formulation for inhalation n°2

First, budesonide (1.5% w/v) was dissolved in isopropanol under magnetic stirring. Then, hydrogenated castor oil (BASF, Ludwigshafen, Germany) and TPGS (Sigma-Aldrich, St. Louis, USA), dissolved in heated isopropanol, were added to the microcrystalline suspension to obtain a final concentration of 1.5% w/v budesonide and a 0.5% w/v hydrogenated castor oil/TPGS (99:1 w/w) mixture, which was then spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain the human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air 800 L/min, and drying air flow rate 35 ^m³ /h. The unit is equipped with a B-296 dehumidifier (BüchiLabortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

The geometrical PSD of the bulk budesonide dry powder formulation was measured as suspended and individualized particles, and within the scope of this invention. As described in Example 1, the measurement was performed using a Mastersizer 3000 laser diffractometer (Malvern Instruments Ltd., Worcestershire, UK) connected to a Hydro MV dispenser (Malvern Instruments Ltd.) equipped with a 40W ultrasonic probe.

Example 9. Preparation of budesonide dry powder formulation n°3.

10 g of micronized budesonide, 9.9 g of hydrogenated castor oil, and 0.1 g of TPGS were mixed until homogeneous in a mixer (Willy A. Bachofen AG, Muttenz, Switzerland). The homogeneous blend was then extruded at an appropriate temperature using a twin-screw extruder (Process-11, Thermo Fischer Scientific, Massachusetts, USA) to obtain a homogeneous lipid matrix. The extrudate was then cut to obtain coarse particles, which were placed in an incubator under appropriate storage conditions to convert the lipid matrix into a stable polymorph. Finally, the particles were ground using a jet mill (at appropriate particle feed rate, injection pressure, and grinding pressure) to obtain inhaled microparticles (DPIs) for human use.

Example 10. Preparation of pemetrexed inhalation powder formulation n°1

In short, crude pemetrexed disodium (heptahydrate form) microcrystals obtained from bulk powder (Carbosynth Limited, Berkshire, UK) were first suspended in 50 mL of isopropanol to achieve a concentration of 1% w/v in the presence of 0.05% w/v TPGS (Sigma-Aldrich, St. Louis, USA). The size was reduced by high-speed (10 min, 24000 rpm) homogenization (X620 motor and T10 dispersion shaft, Ingenieurbüro CAT M. Zipperer GmbH, Staufen, Germany) and high-pressure homogenization (EmulsiFlex-C3 high-pressure homogenizer, Avestin Inc., Ottawa, Canada) at 25000 psi for 20 grinding cycles. A heat exchanger was connected to the homogenization valve and maintained at +5°C using an F32-MA cooling circulator (Julabo GmbH, Seeelbach, Germany). At the end of the process, equal portions of the sample were removed from the suspension to measure the particle size distribution (PSD) of the pemetrexed crystals by laser diffraction (see below).

Then, 1% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany) was dissolved in a heated (50°C) microcrystalline suspension and spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavier, Switzerland) to obtain the human-use DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air flow rate of 800 L/min, and drying air flow rate of 35 ^m³ /h. The unit was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

The geometrical PSD of bulk pemetrexed (pemetrexed microparticles from the size reduction process and pemetrexed dry powder formulation) was measured as suspended and individualized particles, and within the scope of this invention. As described in Example 1, the measurement was performed using a Mastersizer 3000 laser diffractometer (Malvern Instruments Ltd., Worcestershire, UK) connected to a Hydro MV dispenser (Malvern Instruments Ltd.) equipped with a 40W ultrasonic probe.

Example 11. Deposition rate analysis of the pemetrexed dry powder formulation prepared according to Example 10.

The in vitro lung deposition pattern and FPF value of the preparation based on Example 10 were analyzed.

The FPF (Frequency Productivity) – which is the percentage of recovered dose associated with pemetrexed-based particles with an aerodynamic diameter (dae) less than 5 μm – and the aerodynamic PSD (Percentage of Fibre Depression) were determined using an NGI (Copley Scientific, Nottingham, UK) as described in European Pharmacopoeia 8.0. (2014) and an NGI characterized by MMAD. Each DPI formulation (Example 10 and a comparative formulation consisting only of pemetrexed microparticles) was weighed in 3 HPMC capsules (Quali-V-I, Qualicaps, Madrid, Spain) with a pre-sieved mass of 20 mg through a 355 mm stainless steel sieve and deposited in the NGI using an RS.01 dry powder inhaler (RPC Plasciape, Osnago, Italy), which was mounted on the inhaler via its adapter (n=3).

A deposition rate of 100 ± 5 L/min was obtained by connecting two HCP5 air pumps (Copley Scientific, Nottingham, UK) in series with a TPK critical flow controller (Copley Scientific, Nottingham, UK) and measuring it using a DFM3 flow meter (Copley Scientific, Nottingham, UK).

At this flow rate, the cutoff diameters between the stages of NGI are 6.12, 3.42, 2.18, 1.31, 0.72, 0.40, and 0.24 μm. As required by European Pharmacopoeia 8.0. (2014), a critical flow controller is used to ensure a deposition time of 2.4 s and a critical flow rate of <0.5 for the P3/P2 ratio at 100 L/min.

Following impaction, pemetrexed material deposited in the capsule, device, inlet, pre-separator, seven stages, and NGI MOC was collected using ultrapure water/DMF (30:70 v/v) as the diluent phase and sonicated for 30 min. Impaction mass in each stage was determined by validated HPLC quantification of pemetrexed content. The chromatographic system (HP 1200 series, Agilent Technologies, Diegem, Belgium) was equipped with a quaternary pump, autosampler, and diode array detector. Separation was performed on a reversed-phase Hypersil Gold C18 column (5 mm, 250 mm x 4.6 mm) (Thermo Fisher Scientific, Waltham, USA). The mobile phase consisted of ultrapure water/acetonitrile (86:14) acidified with 0.4% formic acid, delivered at a flow rate of 1 mL/min. Quantification was performed at 256 nm. The injection volume was 20 μL, the temperature was set to 30 °C, and the analysis run time was 15 minutes.

The results were then plotted in Copley Inhaler Trial Data Analysis Software 1 (Copley Scientific, Nottingham, UK) to obtain an FPD below 5 μm. This was done by interpolating the recovery mass with the cutoff diameter of the corresponding stage. FPF is expressed as a percentage of the nominal dose.

Figure 3 shows the FPF values (mean ± SD, n = 3 and 1, respectively) of Example 10 compared to a comparative formulation consisting only of pemetrexed microparticles, indicating that Example 10 is superior to the conventional DPI formulation in terms of lung deposition.

Example 12. Analysis of the dissolution rate of pemetrexed from a pemetrexed dry powder formulation prepared according to Example 10.

The dissolution properties of DPI formulations were established by applying a modified method described by Pilcer et al. (Pilcer et al., J Pharm Sci [Pharmaceutical Science Journal] 2013). A dissolution system specifically developed for DPI release profile studies (Copley Scientific, Nottingham, UK) was used, with the method adapted from the "paddle disc method" (Eur. Ph. 7). To study the release profiles of particles deposited in the lungs, the pemetrexed formulation was first fractionated using NGI. A cup for stage 3 was selected, equipped with a removable disc insert to collect particles. Of particular interest was the cutoff diameter of 2.18–3.42 μm for stage 3 at a selected inhalation rate (100 L/min, 2.4 s), allowing for the selection of particles targeting the lungs. Capsules containing an appropriate amount of the formulation according to Example 10 were weighed to collect approximately 6 mg of pemetrexed in stage 3. The disc insert was then covered with a polycarbonate membrane (0.4 μm pore size) (Merck Millipore) and placed in a paddle dissolution apparatus (Erweka DT6; ERWEKA GmbH, Heusenstamm, Hesse, Germany) filled with 400 mL of mSLF (Son and McConville, 2009) – a medium simulating a combination of lung electrolytes and surfactants.

Dissolution tests were conducted at 37±0.2℃ and pH 7.35±0.05, depending on the tank conditions. The paddle rotation speed (with a center-to-center distance of 25±2 mm between the paddle and the disc assembly) was set at 50±4 rpm. A 2.0 mL sample volume was filtered through a 0.22 mm cellulose acetate injection filter (VWR, Leuven, Belgium) at predetermined intervals between 2 minutes and 24 hours, and replaced with 2.0 mL of free, preheated mSLF.

At the end of the dissolution assay, the disc assembly was opened into the dissolution vessel and sonicated for 30 minutes to establish a 100% pemetrexed dissolution value.

Figure 4 shows the release curves (mean ± SD, n = 3) of pemetrexed from the inhalable portion of the DPI formulation prepared according to Example 10, compared to a comparative formulation consisting only of pemetrexed microparticles.

The two dissolution profiles were compared using a similarity factor f2 (Shah et al., Pharm Res, 1998). The profiles were significantly different (f2 < 50). Furthermore, the cumulative release values of Example 10 at all time points were significantly lower than those of pemetrexed microcrystals (p < 0.05, t-test), for example, at 1 hour, they were 53 ± 9% and 97.9 ± 0.9%, respectively (p < 0.01), indicating a controlled release profile of pemetrexed from the compositions of the present invention.

Example 13. Preparation of Insulin Dry Powder Formulation n°3

First, insulin (Sigma-Aldrich) was suspended in isopropanol (1% w/v) using magnetic stirring, and powder dispersion was ensured by sonication in a Branson 2510 bath at 40 kHz for 10 minutes. Particle size was then reduced using an EmulsiFlex-C3 high-pressure homogenizer (Aves-tin Inc., Ottawa, Canada) at 22,000 PSI for 30 cycles. These cycles were performed by directly recirculating the treated suspension into the sample cell (closed loop). Due to the temperature rise caused by HPH, all operations were performed using a heat exchanger placed before the homogenizer valve, maintaining the sample temperature at 5 ± 1 °C.

At the end of the process, equal portions of the sample were removed from the suspension to measure the particle size distribution (PSD) of insulin crystals by laser diffraction (see below).

Then, 1% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany) was dissolved in a heated (50°C) microcrystalline suspension and spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavier, Switzerland) to obtain the human-use DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air flow rate of 800 L/min, and drying air flow rate of 35 ^m³ /h. The unit was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

The geometric PSD of bulk insulin (insulin microparticles and dry powder formulations from the size reduction process) was measured as suspended and individualized particles, and is within the scope of this invention. As described in Example 1, this was performed using a Mastersizer 3000 laser diffractometer (Malvern Instruments Ltd., Worcestershire, UK) connected to a Hydro MV dispenser (Malvern Instruments Ltd.) equipped with a 40W ultrasonic probe.

Example 14. - Deposition rate analysis of insulin dry powder formulation prepared according to Example 13.

The FPF values of the preparation based on Example 13 have been analyzed.

Faster FPF (FPF) – the percentage of insulin-based particles recovered from a dose of less than 5 μm – was determined using a Faster Screening Impactor (FSI) (Copley Scientific, Nottingham, UK). The FSI employs a two-stage separation process, where the first largest non-inhalable clumps are captured in a liquid trap, followed by a fine-cut impaction stage of 5 microns (corresponding to FPF). 10 mg of DPI formulation (according to Example 13) pre-sieved through a 355 mm stainless steel sieve was weighed into HPMC capsules (Quali-V-I, Qualicaps, Madrid, Spain) and deposited in the FSI using RS.01 dry powder inhalers (RPCPlastiape, Osnago, Italy), which were attached to the inhaler via their adapters (n=3).

Two HCP5 air pumps (Copley Scientific, Nottingham, UK) connected in series to a TPK critical flow controller (Copley Scientific, Nottingham, UK) were used to achieve a deposition rate of 100 ± 5 L/min, as measured by a DFM3 flow meter (Copley Scientific, Nottingham, UK). The critical flow controller was used to ensure a deposition time of 2.4 s at 100 L/min.

Following impaction, insulin material deposited in the capsule, device, inhalation port, pre-separator, and on a Fluoropore 9cm PTFE membrane with a pore size of 0.45 mm bonded to a high-density polyethylene carrier (Merck Millipore, Darmstadt, Germany) was collected using 0.01 M HCl as a diluent and sonicated for 30 min. Impaction mass at each stage was determined by quantifying insulin content using the HPLC method described in European Pharmacopoeia 9.2. (2017). FPF is expressed as a percentage of the nominal dose.

Figure 5 shows the FPF values (mean ± SD) of Example 13, demonstrating the high lung deposition rate of the insulin-based DPI composition disclosed in this invention.

Example 15. - Analysis of the dissolution rate of insulin from an insulin dry powder formulation prepared according to Example 13.

The dissolution properties of the DPI formulation according to Example 13 were established by applying a modified method described in Depreter et al. (Depreter et al., Eur J Pharm Biopharm [European Journal of Pharmaceutics and Biopharmaceutics] 2012). A dissolution system specifically developed for DPI release profile studies (Copley Scientific, Nottingham, UK) was used, with the method adapted from the "paddle disc method" (Eur. Ph. 7). To study the release profile of particles deposited in the lungs, the insulin formulation was first fractionated using NGI. A cup for stage 3 was selected to be equipped with a removable disc insert for particle collection. Of particular interest was the cutoff diameter of 2.18–3.42 μm for stage 3 at the selected inhalation rate (100 L/min, 2.4 s), which allowed for the selection of particles targeting the lungs. Capsules containing an appropriate amount of the formulation according to Example 13 were weighed to collect approximately 3 mg of insulin in stage 3. The disc insert was then covered with a polycarbonate membrane (0.4 μm pore size) (Merck Millipore) and placed in a paddle dissolution apparatus (Erweka DT6; ERWEKA GmbH, Heusenstamm, Hesse, Germany) filled with 400 mL of 0.01 mM phosphate-buffered saline (PBS) pH 7.4.

Dissolution tests were conducted at 37±0.2℃ and pH 7.35±0.05, depending on the tank conditions. The paddle rotation speed (with a distance of 25±2 mm between the paddle and the center of the disc assembly) was set to 50±4 rpm. Samples of 5.0 mL were taken at predetermined intervals between 2 minutes and 24 hours, and replaced with 5.0 mL of free, preheated PBS.

At the end of the dissolution assay, the disc assembly is opened into the dissolution vessel and sonicated for 30 minutes to establish a 100% insulin dissolution value.

The sample (Christ Epsilon 1-6) was lyophilized in the presence of 3% w/v trehalose. The lyophilized product was dissolved in 500 μL HCl 0.02N and injected into an HPLC system as described in European Pharmacopoeia 9.2. (2017).

Figure 6 shows the release curves of insulin from the inhaled portion of the DPI formulation prepared according to Example 13 (up to 240 minutes, mean ± SD, n = 2) compared to the comparative formulation described by Depreter et al. (up to 180 minutes) (B).

Example 13 shows that the cumulative release values at different time points are lower than those of the two formulations by Depreter et al., for example, at 1 hour, they are about 60% and about 100%, respectively. This indicates that the insulin composition disclosed in this invention is superior to the insulin microcrystals and lipid-coated insulin microcrystals described by Depreter et al. (F1 and F2 in Figure 6, respectively) in terms of controlling insulin release.

Example 16. Preparation of cisplatin dry powder formulation n°3 and comparative examples

First, cisplatin (Umicore, Hanau-Wolfgang, Germany) was suspended in 50 mL of ethanol to achieve a concentration of 5% w/v. It was then homogenized using an EmulsiFlex-C3 high-pressure homogenizer (Avestin Inc., Ottawa, Canada) at high speed (10 minutes, 24,000 rpm) (X620 motor and T10 dispersion shaft, Ingenieurbüro CAT M. Zipperer GmbH, Staufen, Germany) and high pressure for 40 cycles at 20,000 PSI to reduce its size. These cycles were performed by directly recirculating the treated suspension into the sample cell (closed loop). Because HPH causes a rise in sample temperature, all operations were performed using a heat exchanger placed before the homogenizer valve, maintaining the sample temperature at 15 ± 1 °C.

At the end of the process, equal portions of the sample were removed from the suspension to measure the PSD of cisplatin crystals by laser diffraction (see below).

Then, hydrogenated castor oil (BASF, Ludwigshafen, Germany) and TPGS (Sigma-Aldrich, St. Louis, USA) dissolved in heated isopropanol (or tristearin for comparative examples) were added to the microcrystalline suspension to obtain a final concentration of 2.0% w/v cisplatin and 2.0% w/v triglyceride/TPGS (99:1 w/w) mixture, which was then spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain the human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air 800 L/min, and drying air flow rate 35 ^m³ /h. The unit is equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

The geometric PSD of bulk cisplatin (cisplatin microparticles from the size reduction process and cisplatin dry powder formulations) was measured as suspended and individualized particles, and within the scope of this invention. As described in Example 1, this was performed using a Mastersizer 3000 laser diffractometer (Malvern Instruments Ltd., Worcestershire, UK) connected to a Hydro MV dispenser (Malvern Instruments Ltd.) equipped with a 40W ultrasonic probe.

Example 17 - Based on Example 6 Deposition rate analysis of the cisplatin dry powder formulation prepared in 16.

The in vitro lung deposition pattern and FPF value of the preparation based on Example 16 were analyzed.

The FPF (Frequency Productivity) – which is the percentage of the recovery dose of cisplatin-based particles with a diameter less than 5 μm – and the aerodynamic PSD, characterized by MMAD, were determined using an NGI (Copley Scientific, Nottingham, UK) as described in European Pharmacopoeia 8.0. (2014). 20 mg of DPI formulation (according to Example 17), pre-sieved through a 355 mm stainless steel sieve, was weighed into HPMC capsules (Quali-V-I, Qualicaps, Madrid, Spain) and deposited into the NGI using an RS.01 dry powder inhaler (RPC Plasciape, Osnago, Italy), which was mounted on the inhaler via its adapter (n=3).

A deposition rate of 100 ± 5 L/min was obtained by connecting two HCP5 air pumps (Copley Scientific, Nottingham, UK) in series with a TPK critical flow controller (Copley Scientific, Nottingham, UK) and measuring it using a DFM3 flow meter (Copley Scientific, Nottingham, UK).

At this flow rate, the cutoff diameters between the stages of NGI are 6.12, 3.42, 2.18, 1.31, 0.72, 0.40, and 0.24 μm. As required by European Pharmacopoeia 8.0. (2014), a critical flow controller is used to ensure a deposition time of 2.4 s and a critical flow rate of <0.5 for the P3/P2 ratio at 100 L/min.

Following impact, cisplatin material deposited in the capsule, device, inhalation port, pre-separator, seven stages, and NGI MOC was collected using DMF as a diluent and sonicated for 30 minutes. Impact mass in each stage was determined by quantifying cisplatin content using the empirically validated ETAAS method described by Levet et al. (Levet, Int JPharm [International Journal of Pharmaceutics] 2016).

The results were then plotted in Copley Inhaler Trial Data Analysis Software 1 (Copley Scientific, Nottingham, UK) to obtain an FPD below 5 μm. This was done by interpolating the recovery mass with the cutoff diameter of the corresponding stage. FPF is expressed as a percentage of the nominal dose.

The results obtained, when combined with those obtained in Example 3, indicate that the cisplatin composition disclosed in this invention is superior to other triglyceride-based cisplatin DPI formulations in terms of lung deposition.

Example 18. Analysis of the dissolution rate of cisplatin from a cisplatin dry powder formulation prepared according to Example 16.

The dissolution properties of DPI formulations were established by applying a modified method described by Pilcer et al. (Pilcer et al., J Pharm Sci [Pharmaceutical Science Journal] 2013). A dissolution system specifically developed for DPI release profile studies (Copley Scientific, Nottingham, UK) was used, with the method adapted from the "paddle disc method" (Eur. Ph. 7). To study the release profiles of particles deposited in the lungs, the cisplatin formulation was first fractionated using NGI. A third-stage cup was selected and equipped with a removable disc insert to collect particles. Of particular interest was the cutoff diameter of 2.18–3.42 μm in the third stage at a selected inhalation rate (100 L/min, 2.4 s), allowing selection of lung-targeting particles from capsules filled with an appropriate amount of the formulation according to Example 16 to deposit approximately 2 mg of cisplatin in the third stage. The disc insert was then covered with a polycarbonate membrane (0.4 μm pore size) (Merck Millipore) and placed in a paddle dissolution apparatus (Erweka DT6; ERWEKA GmbH, Heusenstamm, Hesse, Germany) filled with 400 mL of mSLF (Son and McConville, 2009) – a medium simulating a combination of lung electrolytes and surfactants.

Dissolution tests were conducted at 37±0.2℃ and pH 7.35±0.05, depending on the tank conditions. The paddle rotation speed (with a center-to-center distance of 25±2 mm between the paddle and the disc assembly) was set at 50±4 rpm. A 2.0 mL sample volume was filtered through a 0.22 mm cellulose acetate injection filter (VWR, Leuven, Belgium) at predetermined intervals between 2 minutes and 24 hours, and replaced with 2.0 mL of free, preheated mSLF.

At the end of the dissolution assay, the disc assembly was opened into the dissolution vessel and sonicated for 30 minutes to establish a 100% cisplatin dissolution value.

Figure 7 shows the release curves (mean ± SD, n = 3) of cisplatin from the inhalable portion of the DPI formulation prepared according to Example 16, compared to a comparative formulation composed of cisplatin microparticles.

The two dissolution profiles were compared using a similarity factor f2 (Shah et al., Pharm Res, 1998). The profiles were significantly different (f2 < 50). Furthermore, the cumulative release values of Example 16 at all time points were significantly lower than those of cisplatin microcrystals (p < 0.05, t-test), for example, at 4 hours, they were 57 ± 4% and 76 ± 5%, respectively (p < 0.01), indicating a controlled release profile of cisplatin from the compositions of the present invention.

Example 19. Preparation of Budesonide dry powder formulation for inhalation n°4

Under magnetic stirring, 0.1500% w/v budesonide, 2.8215% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany) (or tristearin for comparative example BUD-TS4), and 0.0285% w/v TPGS (Sigma-Aldrich, St. Louis, USA) were dissolved in hot isopropanol (65°C), and the hot solution was spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain the human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air 800 L/min, and drying air flow rate 35 ^m³ /h. The unit is equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

Example 20. Preparation of Budesonide dry powder formulation for inhalation n°5

Under magnetic stirring, 0.600% w/v budesonide, 2.376% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany) (or tristearin for comparative example BUD-TS5), and 0.024% w/v TPGS (Sigma-Aldrich, St. Louis, USA) were dissolved in hot isopropanol (65°C), and the hot solution was spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain the human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air 800 L/min, and drying air flow rate 35 ^m³ /h. The unit is equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

Example 21. Preparation of Budesonide dry powder formulation for inhalation n°6

Under magnetic stirring, 1.500% w/v budesonide, 1.485% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany), and 0.015% w/v TPGS (Sigma-Aldrich, St. Louis, USA) were dissolved in hot isopropanol (65°C), and the hot solution was spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain a human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air supply of 800 L/min, and drying air flow rate of 35 ^m³ /h. The unit was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain a relative humidity of 50% HR during spray drying.

Example 22. - Deposition rate analysis of budesonide dry powder formulations prepared according to Examples 19 and 20.

The in vitro lung deposition patterns and fine particle fractions of the formulations according to Examples 19 and 20 were analyzed. Comparative examples of formulations 19 and 20 (i.e., BUD-TS4 and BUD-TS5, respectively) were produced using the same corresponding protocol, replacing hydrogenated castor oil with tristearin (TS) (Tokyo Chemical Company, Tokyo, Japan).

The FPF (Frequency Productivity) – which is the percentage of budesonide-based particles with a diameter less than 5 μm – and the aerodynamic PSD, characterized by MMAD, were determined using an NGI (Copley Scientific, Nottingham, UK) as described in European Pharmacopoeia 8.0. (2014). 10 mg of DPI formulation, pre-sieved through a 355 mm stainless steel sieve, was weighed into HPMC capsules (Quali-V-I, Qualicaps, Madrid, Spain) (according to Examples 19 and 20 and comparative powders BUD-TS4 and BUD-TS5) and deposited in the NGI using an RS.01 dry powder inhaler (RPC Plasciape, Osnago, Italy), which was mounted on the inhaler via its adapter (n=3).

A deposition rate of 100 ± 5 L/min was obtained by connecting two HCP5 air pumps (Copley Scientific, Nottingham, UK) in series with a TPK critical flow controller (Copley Scientific, Nottingham, UK) and measuring it using a DFM3 flow meter (Copley Scientific, Nottingham, UK).

At this flow rate, the cutoff diameters between the stages of NGI are 6.12, 3.42, 2.18, 1.31, 0.72, 0.40, and 0.24 μm. As required by European Pharmacopoeia 8.0. (2014), a critical flow controller is used to ensure a deposition time of 2.4 s and a critical flow rate of <0.5 for the P3/P2 ratio at 100 L/min.

Following impaction, budesonide deposits in the capsules, apparatus, inlet, pre-separator, seven stages, and NGI's MOC were collected as a diluent phase using a solution of 0.5% w/v poloxamer 407 in an ultrapure water:isopropanol 60:40 (v/v) mixture and sonicated at 60 °C for 30 min. The solutions were then filtered through a 0.45 μm pore size regenerated cellulose Minisart syringe filter (Sartorius Stedim Biotech GmbH, Germany). Budesonide content was determined by quantifying the impaction mass in each stage using a validated HPLC method. The chromatographic system (HP 1200 series, Agilent Technologies, Diegem, Belgium) was equipped with a quaternary pump, autosampler, and diode array detector. Separation was performed on a reversed-phase Alltima C18 column (5 mm, 150 mm x 4.6 mm) (Hichrom, Theale, UK). The mobile phase consisted of pH 3.20 phosphate buffer:acetonitrile (65:35 v/v), delivered at a flow rate of 1.5 mL/min. Quantification was performed at 245 nm. The injection volume was 100 μL, the temperature was set at 40 °C, and the analysis run time was 22 minutes.

The results were then plotted in Copley Inhaler Trial Data Analysis Software 1 (Copley Scientific, Nottingham, UK) to obtain an FPD below 5 μm. This was done by interpolating the recovery mass with the cutoff diameter of the corresponding stage. FPF is expressed as a percentage of the nominal dose.

Figure 8 shows the FPF values (mean ± SD, n = 3) of Examples 19 and 20 compared to Comparative Examples BUD-TS4 and BUD-TS5. (***) p < 0.001, t-test, indicating that the budesonide composition disclosed in this invention is superior to other triglyceride-based budesonide DPI formulations in terms of lung deposition.

Example 23. Dissolution rate of budesonide from budesonide dry powder formulations prepared according to Examples 19, 20 and 21 Rate analysis.

The dissolution properties of DPI formulations and comparative budesonide powder (i.e., micronized budesonide) according to Examples 19, 20, and 21 were established by applying modifications to the method described by Pilcer et al. (Pilcer et al., J Pharm Sci [Pharmaceutical Science Journal] 2013). Comparative micronized budesonide powder was prepared by spray drying (Mini-Spray Dryer B-290, Büchi Labortechnik AG, Flavell, Switzerland) of a 3% w/v budesonide isopropanol solution to obtain human DPI formulations.

A dissolution system specifically developed for DPI release profile studies (Copley Scientific, Nottingham, UK) was used, with the method adapted from the "paddle disc method" (Eur. Ph. 7). To study the release profiles of particles deposited in the lungs, the budesonide formulation was first fractionated using NGI. A stage 2 cup was selected, equipped with a removable disc insert to collect particles. Of particular interest was the stage 2 cutoff diameter of 6.12 μm–3.42 μm at the selected inhalation rate (100 L/min, 2.4 s), allowing selection of lung-targeting particles from capsules filled with appropriate amounts of the formulation (according to micronized budesonide and Examples 20, 21, and 22) to deposit approximately 500 μg of budesonide in stage 3. The disc insert was then covered with a polycarbonate membrane (0.4 μm pore size) (Merck Millipore) and placed in a paddle dissolution apparatus (Erweka DT6; ERWEKA GmbH, Heusenstamm, Hesse, Germany) filled with 400 mL of 0.01 mM phosphate-buffered saline (PBS) pH 7.4.

Dissolution tests were conducted at 37±0.2℃ and pH 7.35±0.05, depending on the tank conditions. The paddle rotation speed (with a distance of 25±2 mm between the paddle and the center of the disc assembly) was set to 50±4 rpm. A 2.0 mL sample volume was filtered through a 0.22 mm cellulose acetate injection filter (VWR, Leuven, Belgium) at predetermined intervals between 2 minutes and 24 hours, and replaced with 2.0 mL of free, preheated PBS.

A 100% budesonide dissolution value corresponds to the mass of deposits in stage 3.

Figure 9 shows the release profiles (n=1) of budesonide from the inhalable portion of micronized budesonide comparative examples and DPI formulations prepared according to Examples 19, 20 and 21, demonstrating the controlled release profiles of budesonide from the compositions of the present invention and the possibility of adjusting the release profile by adjusting the drug/lipid ratio.

Example 24. Preparation of Paclitaxel Dry Powder Formulation for Inhalation n°1

Under magnetic stirring, 0.1500% w/v paclitaxel, 2.8215% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany), and 0.0285% w/v TPGS (Sigma-Aldrich, St. Louis, USA) were dissolved in hot ethanol (50°C). The hot solution was then spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain a human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air supply of 800 L/min, and drying air flow rate of 35 ^m³ /h. The apparatus was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain a relative humidity of 50% HR during spray drying.

Example 25. Preparation of Paclitaxel Dry Powder Formulation for Inhalation n°2

Under magnetic stirring, 0.600% w/v paclitaxel, 2.376% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany), and 0.024% w/v TPGS (Sigma-Aldrich, St. Louis, USA) were dissolved in hot ethanol (50°C). The hot solution was then spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain a human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air supply of 800 L/min, and drying air flow rate of 35 ^m³ /h. The apparatus was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain a relative humidity of 50% HR during spray drying.

Example 26. Preparation of inhaled paclitaxel dry powder formulation n°3

Under magnetic stirring, 1.500% w/v paclitaxel, 1.485% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany), and 0.015% w/v TPGS (Sigma-Aldrich, St. Louis, USA) were dissolved in hot ethanol (50°C), and the hot solution was spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain a human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air flow rate of 800 L/min, and drying air flow rate of 35 ^m³ /h. The apparatus was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain a relative humidity of 50% HR during spray drying.

Example 27. Preparation of Paclitaxel Dry Powder Formulation for Inhalation n°4

Under magnetic stirring, 2.700% w/v paclitaxel, 0.297% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany), and 0.003% w/v TPGS (Sigma-Aldrich, St. Louis, USA) were dissolved in hot ethanol (50°C), and the hot solution was spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain a human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air flow rate of 800 L/min, and drying air flow rate of 35 ^m³ /h. The unit was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain a relative humidity of 50% HR during spray drying.

Example 28. Preparation of Paclitaxel Dry Powder Formulation for Inhalation n°5

Under magnetic stirring, 1.50% w/v paclitaxel, 1.35% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany), and 0.15% w/v TPGS (Sigma-Aldrich, St. Louis, USA) were dissolved in hot ethanol (50°C), and the hot solution was spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain a human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air supply of 800 L/min, and drying air flow rate of 35 ^m³ /h. The unit was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain a relative humidity of 50% HR during spray drying.

Example 29. - Preparation of Paclitaxel Dry Powder Formulation for Inhalation n°6

Under magnetic stirring, 1.5% w/v paclitaxel, 1.2% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany), and 0.3% w/v TPGS (Sigma-Aldrich, St. Louis, USA) were dissolved in hot ethanol (50°C), and the hot solution was spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain a human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air flow rate of 800 L/min, and drying air flow rate of 35 ^m³ /h. The unit was equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain a relative humidity of 50% HR during spray drying.

Example 30. Preparation of Paclitaxel Dry Powder Formulation for Inhalation n°7

Under magnetic stirring, 1.500% w/v paclitaxel, 1.485% w/v hydrogenated castor oil (BASF, Ludwigshafen, Germany), 0.015% w/v TPGS (Sigma-Aldrich, St. Louis, USA), and 0.3% w/v L-leucine (Sigma-Aldrich) were dissolved in hot ethanol (50°C), and the hot solution was spray-dried using a Mini-Spray Dryer B-290 (Büchi Labortechnik AG, Flavell, Switzerland) to obtain the human DPI formulation. The operating parameters used during spray drying were as follows: feed rate 3.0 g/min, inlet temperature 70°C, 0.7 mm nozzle, 1.5 mm nozzle cap, compressed air 800 L/min, and drying air flow rate 35 ^m³ /h. The unit is equipped with a B-296 dehumidifier (Büchi Labortechnik AG) to maintain the relative humidity at 50% HR during spray drying.

Example 31. Preparation of Budesonide dry powder formulation for inhalation n°7

First, budesonide dry powder formulation n°4 for inhalation was prepared according to Example 19. Then, 10 g of budesonide dry powder formulation n°4 for inhalation and 30 g of lactose (SV003, DFE Pharma, Goch, Germany) were mixed in a mixer (Willy A. Bachofen AG, Muttenz, Switzerland) until homogeneous to obtain a carrier-based dry powder formulation for inhalation.

It should be understood that the present invention is not limited to the described embodiments, and variations may be applied without departing from the scope of the appended claims.

Claims (51)

1. A dry powder inhalation formulation comprising an active pharmaceutical ingredient and a lipid matrix, wherein the lipid matrix comprises a triglyceride, and the triglyceride is hydrogenated castor oil; Wherein, relative to the weight of the dry powder inhalation formulation, the weight of the active pharmaceutical ingredient is 0.1 to 90 wt%, and The weight ratio of the active pharmaceutical ingredient to the triglyceride is from 10/90 to 60/40. 2. The dry powder inhalation formulation according to claim 1, wherein the weight ratio of the active pharmaceutical ingredient to the triglyceride is 40/60 to 60/40 relative to the total weight of the dry powder inhalation formulation. 3. The dry powder inhalation formulation according to claim 1, wherein the weight ratio of the active pharmaceutical ingredient to the triglyceride is 50/50 relative to the total weight of the dry powder inhalation formulation. 4. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is a small chemical molecule with a solubility of at least 0.1 w%/v in alcohol. 5. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is a small chemical molecule with a solubility of at least 0.5 w%/v in alcohol. 6. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is a small chemical molecule with a solubility of at least 1 w%/v in alcohol. 7. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is a small chemical molecule with a solubility of at least 5 w%/v in alcohol. 8. The dry powder inhalation formulation according to claim 4, comprising 1 wt% to 85 wt% of the active pharmaceutical ingredient relative to the weight of the dry powder inhalation formulation. 9. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is a small chemical molecule with a solubility in alcohol of less than 0.5 w%/v. 10. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is a small chemical molecule with a solubility in alcohol of less than 0.1 w%/v. 11. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is a small chemical molecule with a solubility in alcohol of less than 0.01 w%/v. 12. The dry powder inhalation formulation according to claim 9, comprising 1 wt% to 90 wt% of the active pharmaceutical ingredient relative to the weight of the dry powder inhalation formulation. 13. The dry powder inhalation formulation of claim 9, comprising 1 wt% to 85 wt% of the active pharmaceutical ingredient relative to the weight of the dry powder inhalation formulation. 14. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is a macromolecule. 15. The dry powder inhalation formulation of claim 14, comprising 1 wt% to 90 wt% of the active pharmaceutical ingredient relative to the weight of the dry powder inhalation formulation. 16. The dry powder inhalation formulation of claim 14, comprising 1 wt% to 85 wt% of the active pharmaceutical ingredient relative to the weight of the dry powder inhalation formulation. 17. The dry powder inhalation formulation according to any one of claims 1 to 16, further comprising a lung retention prolonging excipient. 18. The dry powder inhalation formulation according to claim 17, wherein the excipient is a polyethylene glycol-modified excipient or a polysaccharide. 19. The dry powder inhalation formulation according to claim 17, wherein the excipient is chitosan or dextran. 20. The dry powder inhalation formulation of claim 17, wherein the extended lung retention excipient is a polyethylene glycol-modified excipient and is present in an amount of 0.1 wt% to 20 wt% relative to the total weight of the dry powder inhalation formulation. 21. The dry powder inhalation formulation of claim 17, wherein the extended lung retention excipient is a polyethylene glycol-modified excipient and is present in an amount of 0.2 wt% to 10 wt% relative to the total weight of the dry powder inhalation formulation. 22. The dry powder inhalation formulation of claim 17, wherein the extended lung retention excipient is a polyethylene glycol-modified excipient and is present in an amount of 0.5 wt% to 5 wt% relative to the total weight of the dry powder inhalation formulation. 23. The dry powder inhalation formulation of claim 18, wherein the polyethylene glycol excipient is derived from vitamin E or phospholipids. 24. The dry powder inhalation formulation of claim 18, wherein the PEGylated excipient is derived from tocopheryl polyethylene glycol succinate or distearate phosphate ethanolamine polyethylene glycol 2000. 25. The dry powder inhalation formulation according to claim 1, wherein the formulation is in the form of fine particles having a geometric particle size distribution d ₅₀ of less than or equal to 30 μm. 26. The dry powder inhalation formulation according to claim 1, wherein the formulation is in the form of fine particles having a geometric particle size distribution d ₅₀ of less than 15 μm. 27. The dry powder inhalation formulation according to claim 1, wherein the formulation is in the form of fine particles having a geometric particle size distribution d ₅₀ of less than or equal to 10 μm. 28. The dry powder inhalation formulation according to claim 1, wherein the formulation is in the form of fine particles having a geometric particle size distribution d ₅₀ of less than or equal to 5 μm. 29. The dry powder inhalation formulation according to claim 1, wherein the formulation is in the form of fine particles having a geometric particle size distribution d ₉₀ of less than or equal to 60 μm. 30. The dry powder inhalation formulation according to claim 1, wherein the formulation is in the form of fine particles having a geometric particle size distribution d ₉₀ of less than or equal to 30 μm. 31. The dry powder inhalation formulation according to claim 1, wherein the formulation is in the form of fine particles having a geometric particle size distribution d ₉₀ of less than or equal to 15 μm. 32. The dry powder inhalation formulation according to claim 1, wherein the formulation is in the form of fine particles having a geometric particle size distribution d ₉₀ of less than or equal to 10 μm. 33. The dry powder inhalation formulation according to claim 1, wherein the formulation is in the form of fine particles having a geometric particle size distribution d ₉₀ of less than or equal to 7 μm. 34. The dry powder inhalation formulation according to claim 1, wherein the formulation is in particulate form having a volume average diameter D[4,3] less than or equal to 40 μm. 35. The dry powder inhalation formulation according to claim 1, wherein the formulation is in particulate form having a volume average diameter D[4,3] less than or equal to 20 μm. 36. The dry powder inhalation formulation according to claim 1, wherein the formulation is in particulate form having a volume average diameter D[4,3] less than or equal to 15 μm. 37. The dry powder inhalation formulation according to claim 1, wherein the formulation is in particulate form having a volume average diameter D[4,3] less than or equal to 10 μm. 38. The dry powder inhalation formulation according to claim 1, wherein the formulation is in particulate form having a volume average diameter D[4,3] less than or equal to 6 μm. 39. The dry powder inhalation formulation of claim 1, further comprising an excipient that improves physicochemical and/or aerodynamic properties, present in an amount of 0.1 wt% to 80 wt% relative to the total weight of the dry powder inhalation formulation. 40. The dry powder inhalation formulation according to claim 1, wherein it is in the form of fine particles having a median aerodynamic diameter of less than or equal to 6 μm. 41. The dry powder inhalation formulation according to claim 1, wherein it is in the form of fine particles having a median aerodynamic diameter of less than or equal to 5 μm. 42. The dry powder inhalation formulation according to claim 1, wherein it is in the form of fine particles having a median aerodynamic diameter of less than or equal to 4 μm. 43. The dry powder inhalation formulation according to claim 1, which is in a blister pack or capsule for use in a dry powder inhaler or a sealed and/or disposable dry powder inhaler. 44. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is a small chemical molecule having bronchodilatory activity, glucocorticoid activity, anti-inflammatory activity, or anti-infective activity. 45. The dry powder inhalation formulation of claim 1, wherein the active pharmaceutical ingredient is any active pharmaceutical ingredient that is absorbed by the lungs for systemic or local treatment. 46. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is budesonide, salbutamol, fluticasone, beclomethasone, mometasone, cicsolone, formoterol, aforterol, indacaterol, olodaterol, salmeterol, ipratropium, adetamine, glycopyrronium, tiotropium bromide, fumedoxomil bromide, mometasone, ibuprofen, tobramycin, vancomycin, tetrahydroliprestain, clarithromycin, isoniazid, rifampin, pyrazinamide, itraconazole, voriconazole, aztreonam, ethambutol, Streptomycin, kanamycin, amikacin, colistin, polymyxin E sodium mesylate, cyclomycin, ciprofloxacin, rifapentine, doxycycline, cycloserine E, ethionamide, gatifloxacin, levofloxacin, moxifloxacin, ofloxacin, fosfomycin, para-aminosalicylic acid, denofossodium tetrasodium, lancoviride, ribavirin, zanamivir, laminavir, rubellavir, pentamivir, amphotericin B, posaconazole, isaconazole, capprofen, micafungin, anisfenone, iloprost, or levothyroxine. 47. The dry powder inhalation formulation according to claim 14, wherein the macromolecule is a peptide, protein, antibody, antibody fragment, nanobody, or nucleic acid. 48. The dry powder inhalation formulation according to claim 14, wherein the macromolecule is insulin, proinsulin, synthetic insulin, semi-synthetic insulin, bevacizumab, pembrolizumab, atezolizumab, nivolumab, ipilimumab, Toll-like receptor agonist, auxin-releasing peptide, IgG monoclonal antibody, small interfering RNA, alfa chain enzyme, cyclosporine A, α-1 antitrypsin, interleukin antagonist, interferon-α, interferon-β, interferon-γ, interferon-ω, interleukin-2, anti-IgEmAb, catalase, calcitonin, parathyroid hormone, human growth hormone, insulin-like growth factor-I, heparin, rhG-CSF, GM-CSF, Epo-Fc, FSH-Fc, sFc-γRIIb, or mRNA. 49. The dry powder inhalation formulation according to claim 1, wherein the active pharmaceutical ingredient is an antitumor agent. 50. The dry powder inhalation formulation according to claim 49, wherein the antitumor agent is cisplatin, carboplatin, oxaliplatin, docetaxel, paclitaxel, pemetrexed, etoposide, or vinorelbine. 51. The dry powder inhalation formulation according to claim 1, wherein it is in the form of an oral dry powder inhalation formulation.