BE1023653B1 - PROTEIN BASED EXCIPIENT FOR ACTIVE PHARMACEUTICALS - Google Patents

Abstract

In this application there is provided a pharmaceutical formulation comprising a protein-based excipient in combination with an active pharmaceutical ingredient (API), said formulation being substantially amorphous; and further a method for producing said pharmaceutical formulation, and said formulation for use as a medicament.

Description

PROTEIN BASED EXCIPIENT FOR ACTIVE PHARMACEUTICALS. FIELD OF THE INVENTION

In this application, a pharmaceutical formulation is provided comprising a protein-based excipient in combination with an active pharmaceutical ingredient (API), wherein said formulation is substantially amorphous; and further a method of producing said pharmaceutical formulation; and said pharmaceutical formulation for use as a medicament.

BACKGROUND

To be able to perform its full effect, an "active pharmaceutical ingredient" (API) must overcome a number of obstacles to reach its (biological) target. Poor solubility and long dissolution rates (i.e., disintegration times) can impede the application rate of an API. A particularly poorly soluble API can have a greatly reduced bioavailability (i.e., the administered dose of the unchanged API that enters the bloodstream).

Because of their chemical properties, most API candidates are subject to solution problems during drug development. In certain cases, these solubility problems are solved in the research centers and the API candidates can become the active ingredient of a medication. However, for a large group of API candidates that are difficult to formulate, the low absorption rates for oral administration are so extensive that the API candidates eventually need a different way of ingestion or, in the worst case, do not end up on the market at all.

The generally preferred dosage form for an API is a solid dosage form, such as a tablet or a capsule. Typically, the solid dosage form contains not only the API, which is the drug itself, but at least one component other than the API. Traditionally, said additional component is pharmaceutically inert and does not react biologically with the API itself, apart from stabilizing the formulation. These inactive ingredients are generally referred to in the pharmaceutical context as excipients. Thus, unlike the API, which is chosen primarily because of its active properties, the excipient is mainly chosen because of its non-active properties. Various synthetic polymers (e.g., polymethyl-methacrylate-sorbitol) are currently being developed to fulfill the role of excipient and to create a soluble formulation. For example, polymers used in amorphous solid dispersions exhibited solubility-enhancing properties. However, the applicability of polymers in the pharmaceutical industry has also created new challenges associated with their development, processing and production.

Polymers have the specific disadvantage that they do not come from a biologically relevant source. Since many of the intermolecular interactions and complex formations (e.g. H-bridges, ionic and / or van der Waals interactions) of hydrophilic polymers are adjusted with the API to find a balance between the solubility and stability of the formulation, it is often unpredictable whether the presence of polymers may or may not have additional undesirable effects on the drug, or that certain desired interactions may be missing or lost.

Moreover, the costs for developing, testing and subsequently producing said synthetic polymer excipients may even be higher than the production costs of the API itself, especially for standard drugs.

Certain APIs also lose their active properties or are completely lost as a result of the physical conditions (eg temperature, humidity, flow, etc.) that are present during the development and / or production phase of the formulation. Exposure to extreme temperatures and humidity causes structural degradation and changes in chemical behavior of an API. For example, certain APIs have shown that they are not suitable for hot melt extrusion, a general production method for polymers.

In addition, certain APIs also exhibit chemical interaction with residual solvents commonly used for polymer processing, or in certain cases the APIs may even react with the polymer itself when coupled together; in particular for C-O and C-N bonds and double bonds. With C-O bonds, for example, oxidation, reduction, cleavage, addition and elimination can easily occur. All of these situations lead to impurities in the API formulation that have no therapeutic value and may even be harmful (i.e., genotoxic). For example, the use of alcohols such as methanol or ethanol, which are commonly used solvents during the manufacture of polymers, will cause sulfonate esters to exhibit genotoxic properties in certain APIs.

The solubility and dissolution level of the API used in the formulation directly affects the bioavailability and the supersaturation state. It is therefore very important to increase the solubility and dissolution levels of the API, especially for APIs that have low solubility and / or bioavailability. Achieving a state of oversaturation, and subsequently maintaining said oversaturation state for as long as possible, leads to even more favorable results for the API. In addition, an improvement in bioavailability and the maintenance of supersaturation will increase the absorption rate of the API, which could lead to a lower required total weight / volume (dosage) of the API in a formulation.

Accordingly, there arises a need to obtain an effective and stable excipient that can be combined in a formulation to improve the solubility and dissolution level for an API. In addition, said auxiliary substance preferably also increases the bioavailability of said APIs, and furthermore makes a state of oversaturation possible and maintains it. Specifically, there is a need for such an adjuvant for APIs that have a low solubility and dissolution rate and / or levels. Excipients from the prior art do not achieve the combined safety and biological relevance / interaction, improved oversaturation and bioavailability, and process scale, costs and times in a manner achieved by the present invention.

SUMMARY OF THE INVENTION

The present invention relates to an excipient as described in this application, a method for producing said excipient, and therefore a formulation comprising said excipient in combination with an API.

Although many APIs are known in the art, various classes of APIs can be distinguished based on their solubility and / or bioavailability. APIs that exhibit low solubility and / or bioavailability are described in the art and are known to those skilled in the art. In a particular embodiment of the present invention, the (protein-based) excipient of the present invention is combined with an API. In particular, said API is an API that exhibits poor solubility and / or bioavailability, and therefore benefits from the properties of the excipient of the present invention that improve solubility and bioavailability. In particular embodiments, said API is selected from Flubendazole, Carbamazepine, Griseofulvine, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, Jopanoic Acid, Itraconazole or Naproxen, and especially Flubendazole.

Accordingly, the present invention relates to a formulation comprising the following: - a protein-based excipient obtained from a protein composition or a hydrolyzate thereof comprising proteins with a length of at least 10 amino acids; and - an active pharmaceutical ingredient (API); characterized in that said protein-based excipient and said API are both substantially amorphous.

In particular, the formulation as described in this application provides that the protein-based excipient is substantially undenatured and / or retains at least a portion of its biological activity.

In particular, the formulation as described in this application provides that at least one protein of the protein composition or a hydrolyzate thereof is selected from soy protein, pea protein, gelatin, keratin, corn, wheat, hemp, rye, oats, peanut, barley, casein, albumin, whey protein (lactalbumin), hydrolyzed whey protein isolate (HWPI), serum albumin, bovine serum albumin (BSA), egg albumin, fish albumin, elastin, collagen, hydrolyzed collagen, plasma proteins or a combination thereof, preferably BSA and / or gelatin. The aforementioned list includes proteins obtained from natural sources by extraction methods and is also known as "native proteins", but also variants optimized using recombination technology, also known as "recombined proteins".

In particular, the formulation as described in this application provides that the API exhibits low solubility, dissolution rate or level, poor oversaturation state and / or bioavailability.

In particular, the formulation as described in this application provides for the API to be selected from the following list: Flubendazole, Carbamazepine, Griseofulvin, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, Jopanoic Acid, Itraconazole or Naproxen, preferably Flubendazole.

In particular, the formulation as described in this application provides that said API is Flubendazole and wherein said protein-based excipient is obtained from a protein composition or a hydrolyzate thereof comprising BSA and / or gelatin.

In particular, the formulation as described in this application provides for said formulation to be dosed in a solid dosage form, preferably a tablet, pill or capsule.

In particular, the formulation as described in this application provides that said formulation is characterized in that it has a particle size between 1 µm and 1 mm; preferably between 5 pm and 50 pm; most preferably between 10 µm and 20 µm.

In another aspect, the present invention relates to the use of a protein composition or a hydrolyzate thereof comprising proteins with a length of at least 10 amino acids as a protein-based excipient.

In another aspect, the present invention relates to a method for producing a pharmaceutical formulation, comprising - a protein-based excipient obtained from a protein composition or a hydrolyzate thereof comprising proteins of at least 10 amino acids in length; and - an active pharmaceutical ingredient (API); characterized in that said protein-based excipient and said API are both substantially amorphous; wherein said method comprises at least the steps of: (a) dissolving said API using a solvent to obtain a solution; and (b) drying the solution from step (a) to obtain a powder that is substantially amorphous.

In particular, the method as described in this application provides for the protein-based excipient to be prepared through the steps of: (i) dissolving a protein composition or hydrolyzate thereof using a solvent to obtain a solution; and (ii) drying the solution of step (i) to obtain said protein-based excipient.

In particular, the method as described in this application provides for the solutions of steps (a) and (i) to be dissolved using the same or a different solvent.

In particular, the method as described in this application provides that the API and the protein-based excipient: - be dissolved and dried together in the same solvent to form said pharmaceutical formulation; - separately dissolved in the same solvent or different solvents and then dried together to form the pharmaceutical formulation; - are dissolved in the same solvent or different solvents and dried separately and then mixed to form the pharmaceutical formulation.

In particular, the method as described in this application provides that the solvent is an organic acid, preferably formic acid or acetic acid.

In another aspect, the present invention relates to a formulation as described in this application for use as a medicament.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1: Graph of a solution profile showing the average solution (%) of various protein sources as a function of the dissolution time (min) and the pH value. The results are further discussed in Example 1 and the legend is as follows: square - gelatin; circle - BSA (albumin); triangle - pea; lozenge - soy; stripe - whey; cross - zein (maize).

FIG. 2: Gel filtration chromatograms showing the absorbance (AU) of bovine serum albumin (BSA) as a function of the elution time (min). The results are further discussed in Example 2 and the legend is as follows: gray line - BSA dissolved in H2O; black broken line - BSA dissolved in formic acid.

FIG. 3: Gel filtration chromatograms showing the absorbance (AU) of bovine serum albumin (BSA) as a function of time (min) sampled from a solution in formic acid at different time intervals. The results are further discussed in Example 2 and the legend is as follows: black - 0 o'clock; black interrupted - 4 hours; gray - 8 hours; black dotted - 24 hours.

FIG. 4: Binding curves showing the level of tryptophan quenching (relative Δ fluorescence units) of bovine serum albumin (BSA) at different pH values, derived from pH-regulated redissolved films cast from a 5% solution in formic acid, as a function of the molar concentration of Flubendazole (FLU) (μΜ). The results are further discussed in Example 3 and the legend is as follows: round - pH 7.0; square - pH 4.0; triangle - pH 1.0.

FIG. 5: Graph of a supersaturation profile representing the average concentration of dissolved Flubendazole (FLU) (pg / ml) from a 225 pg / ml amorphous FLU (FLU stock dissolved in formic acid) in FaSSGF medium for each series comprising a other gelatin-based excipient as a function of the dissolution time (min). The results are further discussed in Example 4.

FIG. 6: Graph of a dissolution profile representing the average solution (%) of Flubendazole (FLUB) mixed with a whey protein-based excipient as a function of the dissolution time (min). The results are further discussed in Example 4 and the legend is as follows: square - whey protein FLUB physical mixture; round - whey protein FLUB film.

FIG. 7: Graphs of dissolution profiles (in 0.1 N HCl, pH 1.5) showing the following: (a) the dissolved Flubendazole (FLU) CF | Ub (pg / ml) or (b) the dissolved percentage of the total amount of FLU (%) for each set of protein-based formulations as a function of the dissolution time (min); the values shown represent the calculated means for each experiment. The results are further discussed in Example 7 and the legend is as follows: the solid line marked with a circle corresponds to a spray-dried formulation comprising 80% pig skin gelatin (Bloom = 50 g) and 20% FLU; the solid line marked with a square corresponds to a spray-dried formulation comprising 90% pig skin gelatin (Bloom = 225 g) and 10% FLU; a dashed line corresponds to a formulation comprising a physical mixture of 80% pig skin gelatin (Bloom = 50 g) and 20% FLU.

FIG. 8: Graph of solution profiles (first 90 minutes in 0.1 N HCl, pH 1.5, then 250 minutes in P042 'buffer, pH 6.8 (addition of Na3 PO4)) showing the following: (a) the concentration dissolved Flubendazole (FLU) CRub (pg / ml) or (b) the dissolved percentage of the total amount of FLU (%) for each series of BSA-based formulations as a function of the dissolution time (min); the values shown represent the calculated means for each experiment. The results are further discussed in Example 7 and the legend is as follows: the solid line marked with a circle corresponds to a spray-dried formulation comprising 90% BSA and 10% FLU; the solid line marked with a triangle corresponds to a spray-dried formulation comprising 80% BSA and 20% FLU; the solid line marked with a square corresponds to a spray-dried formulation comprising 70% BSA and 30% FLU; a dashed line corresponds to a formulation comprising a physical mixture of 80% BSA and 20% FLU.

FIG. 9: Graph of X-ray diffraction patterns of the formulations comprising: (1) 80% Soluplus®; 20% Flubendazole (FLU); (2) 40% polymer: 40% gelatin: 20% FLU; (3) 10% polymer: 70% gelatin: 20% FLU; (4) 80% gelatin: 20% FLU (4). The results are further discussed in Example 8.

DETAILED DESCRIPTION OF THE INVENTION

Before describing the present system and method of the invention, it should be understood that this invention is not limited to specific systems and methods or combinations described, since such systems and methods and combinations may, of course, vary. It should also be understood that the terminology used in this application is not intended to be limiting, since the scope of the present invention is limited solely by the appended claims.

As used in this application, the singular forms "the", "it", and "one" include both the singular and the plural references, unless the context clearly indicates otherwise.

The terms "comprising", "includes" and "consisting of" are, as used in this application, synonymous with "including", "including" or "containing", "contains", and include or with an open end and close no additional unlisted members, parts or process steps. It will be understood that the terms "comprising", "includes" and "consisting of" as used in this application include the terms "consisting of" and "consists of".

The indication of numerical ranges with end points includes all numbers and fractions that fall within the respective ranges, as well as the stated end points.

The term "approximately" or "approximate" as used in this application when reference is made to a measurable value such as a parameter, a quantity, a duration and the like, is intended to include variations of +/- 10% or less, preferably +/- 5% or less, more preferably +/- 1% or less, and even more preferably +/- 0.1% or less at and from the specified value, to the extent that such variations are suitable in the described invention to be implemented. It should be understood that the value to which the modifier "roughly" or "approximate" refers is also specific and preferably described.

Although the terms "one or more" or "at least one", such as one or more members or at least one member of a group of members are per se clear, the terms include, by way of further clarification, a reference to any member from said members, or to two or more of said members, such as any £ 3,> 4,> 5,> 6 or> 7 etc. of said members, and up to and including all said members.

All references mentioned in the present description are hereby incorporated by reference in their entirety. In particular, the doctrine of all references to which this application specifically refers is incorporated by reference.

Unless defined otherwise, all terms used in describing the invention, including technical and scientific terms, have the meaning that is generally understood by one of ordinary skill in the art to which this invention belongs. By way of further guidance, the definitions of the terms are included to better understand the teachings of the present invention.

Various aspects of the invention are defined in more detail in the following passages. Each of the aspects thus defined can be combined with one or more other random aspects, unless the contrary is clearly indicated. In particular, any feature that is indicated to be preferred or advantageous may be combined with one or more other random features that are indicated to be preferred or advantageous.

Reference throughout this description to "one embodiment" or "an embodiment" means that a specific feature, specific structure or specific feature described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, appearances of the phrases "in one embodiment" or "in an embodiment" at different places in this description do not necessarily all refer to the same embodiment, but it could. Furthermore, specific features, structures or features may be combined in any suitable manner in one or more embodiments, as would be apparent from this disclosure to one skilled in the art. In addition, combinations of features of different embodiments, although certain embodiments described in this application include some features included in other embodiments but others not, are intended to fall within the scope of the present invention and to form different embodiments as clearly would be to experts in the field. For example, in the appended claims, any of the claimed embodiments can be used in any combination.

In the present description of the invention, reference is made to the accompanying drawings which form a part thereof, and in which specific embodiments are shown by way of explanation only, in which the invention can be carried out. Reference numbers in brackets or in bold print which are attached to respective parts only illustrate the parts by way of example, which is not intended to limit the associated parts. It should be understood that other embodiments can be used and that structural or logical changes can be made without departing from the scope of the present invention. The following detailed description, therefore, is not to be construed as limiting, and the scope of the present invention is defined by the appended claims.

Each drug generally consists of two components or aspects. The first component is the actual API, which is the most important component. The second component is known as an aid. This refers to the substance in the medicine or tablet. If the medicine is in syrup form, the excipient is the liquid used.

As used herein, the term "active pharmaceutical ingredient" (API) generally refers to a substance in a pharmaceutical formulation that is biologically active and intended to produce the desired effect in the body. Other terms such as "active ingredient", "active ingredient" and "active ingredient" have the same meaning and can be used interchangeably.

Traditionally, the term adjuvant refers to a biologically inactive substance used in a pharmaceutical formulation that acts as a medium for the substance that is active. For the purposes of the present invention, however, the protein-based excipient is preferably not biologically active, but instead interacts with the API, thereby maintaining it in solution or oversaturation. Thus, a distinction is made between the "protein-based excipient as used in the present invention, which is obtained from a natural source, in particular protein-based, comprising a protein composition or hydrolysates thereof, and" traditional excipients, which are usually are derived from a natural or synthetic substance, for example polymers. Traditional excipients can also be referred to by other terms such as "non-active substance," non-active ingredient "and" non-active ingredient ", which have the same meaning and can be used interchangeably. As used in this application, the term "excipient in general refers to" protein-based excipient unless otherwise indicated.

In a first aspect, the present invention relates to a formulation comprising a protein-based excipient and an API, both of which are substantially amorphous.

In particular, the present invention relates to a formulation comprising: - a protein-based excipient obtained from a protein composition or a hydrolyzate thereof comprising proteins with a length of at least 10 amino acids; and - an active pharmaceutical ingredient (API); characterized in that said protein-based excipient and said API are both substantially amorphous.

As used in this application, the term "formulation" generally refers to a material or mixture prepared according to a formula. For the purposes of the present invention, the formulation comprises at least one (protein-based) excipient of the present invention and at least one API, in particular an API that exhibits low solubility, low dissolution rate and / or bioavailability, so as to To form a "pharmaceutical formulation". Other terms, such as "mixture" or "combination" generally have the same meaning, unless otherwise indicated, and can be used interchangeably.

As used in this application, the term "protein" generally refers to a polymer chain, or multiple polymer chains, made from amino acids linked to each other through peptide bonds.

For the purposes of the present invention, the proteins contain at least 10 amino acids in their primary structure. In certain embodiments of the present invention, the protein-based excipient is characterized in that it is obtained from a protein composition or a hydrolyzate thereof that contains proteins of at least 10 amino acids in length; preferably with a length of at least 20 amino acids; more preferably with a length of at least 50 amino acids; most preferably with a length of at least 100 amino acids.

As used in this application, the terms "amorphous or" non-crystalline "generally refer to a state or solid lacking a defined or clear shape, which would be a characteristic of an ordered crystalline structure. For the purposes of the present invention, a powder or a solid dispersion is referred to as "substantially amorphous if it contains virtually no trace of crystallinity, i.e., less than 10%; and is referred to as "completely amorphous" if it contains no traces of crystallinity, i.e., less than 1%. The amorphous state can be experimentally verified for most powders or solid dispersions (e.g., X-ray powder diffraction spectroscopy (XRD), differential scanning calorimetry (DSC).

To date, the biggest problem in increasing saturation has been that when the concentrations of the free AP1 become higher than the equilibrium solubility, this leads to precipitation or crystallization of the drug. The inventors have unexpectedly discovered a formulation that is essentially amorphous and prevents said crystallization. In fact, this formulation showed higher levels of API oversaturation, and maintained these higher oversaturation levels for a long time. Given the large number of APIs in current drug research with poor solubility in water, the concept of supersaturation can serve as an effective formulation approach for increasing bioavailability. The formulation is intended to provide significantly high concentrations of the AP1 in the gastrointestinal tract by achieving a state of oversaturation and furthermore to maintain the state of oversaturation for a long period of time. In this way the state of supersaturation can improve the absorption of the AP1 in the intestines, which can then lead to an improved bioavailability of said API. This oversaturation effect can further be controlled by means of tailor-made (i.e. improved or selective) dissolution rates and / or levels, as described below.

As used in this application, the term "supersaturation" generally refers to a state of a solution containing more of the dissolved material than can be dissolved by the solvent under normal conditions. Specifically for the present invention, the state of oversaturation refers to a state in which the formulation dissolves more absorbable material, ie the API, than could be dissolved under normal conditions, ie without the excipient as described by the present invention, preferably in the stomach gut region. As used in this application, the term "bioavailability" generally refers to the dose of the unchanged API that reaches the circulatory system; and is usually exposed to digestive processes and absorption in the gastrointestinal tract. Thus, the bioavailability for a formulation indicates the systemically available fraction of the API. The bioavailability can be measured experimentally (e.g. in vivo) and is generally known for most APIs in the art.

In general, the formulation of the present invention exhibits at least one of the following measurable improvements over a control formulation comprising the API without the protein-based excipient as described by the present invention: (a) an increase in the maximum API concentration of at least about 25%; (b) an increase in the dissolution rate of at least about 25%; (c) an increase in the period of time during which a state of oversaturation is reached and maintained of at least about 25%; (d) an increase in the bioavailability of the API of at least about 25%. The experimental data that support said improvements are presented in the examples.

In a particular embodiment of the present invention, the protein-based excipient is characterized in that said excipient is obtained by dissolving or solubilizing a protein composition or a hydrolyzate thereof in a solvent to obtain a protein solution and drying said protein solution to obtain the excipient to gain. In essence, the excipient of the invention is a protein-based excipient that comprises a protein source of animal, natural (i.e., vegetable) and / or microbial origin.

In another aspect, the present invention relates to an API that is substantially amorphous, characterized in that said API is obtained by dissolving or solubilizing an API in a solvent to obtain an API solution, and drying said API solution. solution to obtain the API that is essentially amorphous. In certain particular embodiments, the API according to the present invention is substantially amorphous, more particularly almost completely amorphous, and preferably particularly completely amorphous.

The inventors have unexpectedly observed that the adjuvant of the present invention combined with an API forms a stable, amorphous formulation after drying the solution comprising said adjuvant, an API and a solvent. This formulation achieves solubility, dissolution rates and levels, oversaturation and / or bioavailability that are higher than that obtained using traditional (e.g., polymeric) excipients (experimentally confirmed).

In particular embodiments, the formulation of the present invention is characterized in that the protein-based excipient is obtained by dissolving or solubilizing a protein composition or a hydrolyzate thereof in a solvent to obtain a protein solution and drying said protein solution to obtain a protein solution. protein-based excipient, and further characterized in that the amorphous API is obtained by dissolving or solubilizing an API in a solvent, similar to or different from the solvent used for the protein solution, to obtain an API solution and drying said API solution to obtain an API that is substantially amorphous, and further combining said dried adjuvant and said dried API to obtain a formulation that is substantially amorphous according to an embodiment of the invention.

Alternatively, in certain other embodiments, the formulation is characterized in that the formulation is obtained by dissolving or solubilizing a protein composition or a hydrolyzate thereof, together with an API, in a conventional solvent and drying said protein API solution to obtain a formulation that is substantially amorphous according to an embodiment of the invention.

Alternatively, in certain other embodiments, the formulation is characterized in that the formulation is obtained by dissolving or solubilizing an API in a solvent and drying said API solution to obtain an API that is substantially amorphous, and furthermore the combining said dried API with a provided protein-based excipient that is preferably substantially not denatured to obtain a formulation that is substantially amorphous according to an embodiment of the invention.

As used in this application, the term "protein composition" generally refers to a mixture comprising similar proteins from a similar source, similar proteins from different sources, different proteins from a similar source, or different proteins from different sources. For the purposes of the present invention, the protein composition refers to a mixture of different proteins or hydrolysates thereof.

The term "solvent" generally refers to a substance (liquid, solid or gaseous) in which a substance to be dissolved (ie a substance that is chemically different from the solvent) is dissolved to result in a solution. For the purposes of the present In this invention, the solvent is generally a liquid in which a solid (e.g., a protein composition or hydrolyzate thereof, and / or an API) is dissolved In a particular embodiment, the solvent is an organic acid, preferably formic acid or acetic acid.

After extraction from a biological source, most protein compositions are amorphous. However, during processing, the protein composition is exposed to solvent, such as organic acids, which usually denature the proteins, i.e., the process in which proteins or nucleic acids lose the quaternary structure, tertiary structure and / or secondary structure present in their native state. Since the biological activity (i.e., the interaction with foreign molecules, such as the API) of the proteins is dependent on the fully folded structure of the protein molecule, it is generally believed that this denaturation process will reduce the proteins to a biologically inactive state.

In particular, the formulation as described in this application provides that the protein-based excipient is substantially not denatured and / or retains at least part of its biological activity.

As used in this application, the term "denaturation" generally refers to the process in which proteins or nucleic acids lose their quaternary, tertiary and / or secondary structure present in their native state, by applying a certain external pressure (e.g., temperature) , radiation, etc.) or compounds (e.g., a strong acid or base, a concentrated inorganic salt, certain organic solvents, etc.). When proteins are denatured, this often results in a reduction or disruption of their biological activity. For the purposes of the invention, a protein is referred to as "substantially non-denatured" if it exhibits virtually no traces of denaturation, ie, less than 10%, and is referred to as "completely non-denatured" if it has no traces of denaturation, ie less than 1%. The denaturation state can be experimentally verified for most proteins (e.g., double-polarization interferometry, circular dichroism, quartz crystal microbalance) either directly or through associated loss of biological activity.

In certain embodiments of the present invention, the excipient is substantially not denatured. In certain embodiments of the present invention, the excipient is completely non-denatured.

In certain other embodiments of the present invention, the excipient retains at least a portion of its biological activity. In certain other embodiments of the present invention, the excipient retains almost entirely its biological activity. In certain other embodiments of the present invention, the excipient retains its full biological activity.

The inventors have unexpectedly observed that the protein-based excipient can enter into a biological interaction with the API to keep it in solution and / or oversaturation. The biological activity of a protein is related to the secondary, tertiary and quaternary structure of said protein; thus, denaturation can be linked to a reduction or loss of biological activity. The inventors have thus found that by maintaining the biological activity of the proteins comprising the protein-based excipient, the formulation comprising said protein-based excipient can exhibit various advantages over traditional (polymeric) excipients of the prior art. The advantages may include improved solubility, dissolution rate and / or levels, achieving a state of super-saturation, maintaining a state of super-saturation for a longer period of time (experimentally confirmed). As a result, the formulation can further improve the absorption of the API in the gastrointestinal tract, which can then bring about an improved bioavailability of said API.

In particular, the formulation as described in this application provides that at least one protein of the protein composition or a hydrolyzate thereof is selected from soy protein, pea protein, gelatin, keratin, corn, wheat, hemp, rye, oats, peanut, barley, casein, albumin, whey protein (lactalbumin), hydrolyzed whey protein isolate (HWPI), hydrolyzed collagen, plasma proteins, serum albumin, bovine serum albumin (BSA), egg albumin, fish albumin, elastin, collagen or a combination thereof, preferably BSA and / or gelatin. The aforementioned list includes proteins obtained from natural sources by extraction methods, also known as "native proteins," but also variants optimized using recombination technology, also known as "recombined proteins." For the purposes of the invention, the term "proteins" refers equally to the native or recombined proteins, unless otherwise stated, or alternatively, to proteins that have been improved using different pre-processing or post-processing methods to improve their physical, biological and / or or chemical properties.

Alternatively, in another embodiment of the invention, the excipient of the present invention is prepared from a protein composition or a hydrolyzate thereof extracted from animal sources, in particular selected from sour pork source, alkaline bovine source, bovine skins, soda-treated skins , sour pork skins, sour pork bones, base-treated beef bones, sour beef bones, sour beef skins, base-treated pork bones, fish, or a combination thereof. For extracted gelatin, the protein is further classified based on the Bloom value, which is a measure of the stiffness and strength of the gelatin. The value represents the weight (in grams) required for a probe to bend the surface of the gel by 4 mm without breaking it, which is usually between 30 and 300 g Bloom. Different Bloom values can be used for the purposes of the invention; Bloom values above 200 g are preferably used.

In general, protein compositions comprising proteins shorter than 10 amino acids in length do not require a solvent and can be easily dissolved in aqueous media. In addition, the inventors have unexpectedly observed that protein excipients comprising a protein composition or a hydrolyzate thereof with a length greater than 100 amino acids (e.g., BSA and gelatin) may exhibit additional beneficial biological activity in combination with certain APIs. Thus, formulations comprising a protein-based excipient comprising a protein composition or hydrolyzate thereof with a length greater than 100 amino acids may have a further improved solubility, dissolution rate and / or levels, achieving a state of supersaturation, maintaining a state of supersaturation over a longer period of time (experimentally confirmed). As a result, the formulation can further improve the absorption of the API in the gastrointestinal tract, which can then ensure improved bioavailability of said API.

As a first reference, the complete precursor protein of BSA has a length of 607 amino acids, and weighs 69324 Dalton (Da). The full length of a mature BSA protein is 583 amino acids and weighs 6,6463 Da. In certain embodiments, the BSA protein can be cleaved to achieve a shorter amino acid length, with BSA still retaining its biological activity for the purposes of the present invention. In addition, the composition of BSA can be combined to, for example, further improve the saturation values; thus, formulations that use said recombined BSA can even achieve a higher degree of supersaturation and bioavailability compared to formulations that only use native BSA.

As a second reference, the helical portion of fibrous collagens, homo- or heterotrimers in their native states, generally contains about 1000 amino acids per chain, which for a well-folded molecule amounts to 3000 amino acids in total, and weighs more than 300 kDa a significant amount of post-translational modifications. In the manufacture of gelatin, treatment of the animal starting material with diluted acid (type A process) or lye (type B process) results in partial splitting of the crosslinks between the chains that determine the thermal tolerance of collagen fibrils. The structure is degraded to such an extent that "water-soluble collagen", i.e., gelatin, is formed. In the subsequent extraction step, hot water melts the collagen fibrils into the individual chains that make them up, and at the same time serves as the solvent. Gelatin is therefore a polydisperse mixture of protein fragments with varying molecular masses, in the range of 15 to more than 400 kDa, depending on the level of chemical / thermal hydrolysis of the polypeptide chains and the level of the hydrolysis of the crosslinks between the chains. Dissolving dried gelatin in formic acid does not change its molecular weight profile for days, and after evaporation of the formic acid solvent, the gelatin retains the gel-forming / melting properties of the original product. In particular, the formulation as described in this application provides that the API exhibits low solubility, dissolution rate or level, supersaturation state and / or bioavailability.

As used in this application, the term "solubility" refers to a quantitative term that refers to the property of a solid, liquid, or gaseous chemical called a substance to be dissolved, to dissolve in a solid, liquid, or gaseous The solubility is expressed in terms of the "dissolution level", which expresses the amount of said substance that will dissolve in a given amount of solvent. In general, if more than 0.1 g of that substance dissolves in 100 ml of solvent, the substance is called soluble; if less than 0.1 g dissolves in 100 ml of solvent, the substance is considered insoluble, or more precisely, hardly soluble. The solubility can be measured experimentally and is known in the art. Explanation of how the solubility of the present invention was determined is further given in the examples of this description. A related term is the "dissolution rate", with which the solubility is expressed measured over the period of time during which said substance will dissolve in a given amount of solvent. In general, highly soluble substances also exhibit a high dissolution rate, although certain substances may have a high dissolution level while still dissolving very slowly. Selective adjustments to the dissolution level and dissolution rate may have secondary advantages with regard to the functionality of a formulation and its pharmacokinetic properties.

The inventors have unexpectedly observed that the adjuvant of the present invention is a stable powder after drying the solution comprising said protein composition or a hydrolyzate thereof and a solvent, preferably an organic acid. This protein-based excipient retains solubility and dissolution rates higher than that obtained by traditional (e.g., polymeric) excipients (experimentally confirmed). Therefore, formulations are designed to make use of said properties that improve solubility and dissolution rate.

In particular, the formulation as described in this application provides that the API is selected from the following list: Flubendazole, Carbamazepine, Griseofulvin, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, Jopanoic Acid, Itraconazole or Naproxen, preferably Flubendazole.

For the purposes of the invention, the API can be selected from any API category. Preferably, the API truly exhibits low solubility, dissolution rate and / or bioavailability, thereby fully benefiting from the properties of said formulation that improve solubility, dissolution rate and dissolution level, oversaturation and / or bioavailability, to enhance the active properties of said API possible or to improve. In a particular embodiment of the present invention, said formulation comprises an API selected from Flubendazole, Carbamazepine, Griseofulvin, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, Jopanoic Acid, Itraconazole, or Naproxen, more particularly Flubendazole.

The inventors have unexpectedly observed that the adjuvant of the present invention in combination with an API forms a stable amorphous formulation after drying the solution comprising said adjuvant, an API and an acid. This process results in a formulation that achieves solubility and dissolution rates above those obtained using traditional polymer excipients (experimentally confirmed).

Often such a crystalline state exhibits very poor solubility and dissolution rates, which necessitates the use of multiple solvents or a high temperature to dissolve into a solution. By dissolving a crystalline protein composition in an acid solvent (e.g. formic acid, acetic acid), the protein composition is transformed into a substantially amorphous state; said acid solvents are also capable of disrupting protein-protein interactions that cannot be disrupted using a neutral solvent (e.g., H 2 O). Typically, such an amorphous state exhibits improved solubility and dissolution rates relative to the crystalline states. In certain cases, the protein composition becomes completely soluble in an amorphous state, even at low temperatures.

In certain embodiments, the formulation of the present invention comprises a ratio of API to excipient between about at least 10% API and at most 90% excipient (w / w), and at least 90% API and at most 10% excipient (w / w w). In certain other embodiments, the formulation may include an API to excipient ratio of about 50% API and about 50% excipient (w / w).

The inventors have discovered that these concentrations offer an optimum balance between the properties of the excipient (e.g. improved solubility, dissolution rates and bioavailability) and the dosage of the API. Moreover, these concentrations of materials offer an optimum balance between the aforementioned properties and production costs and times of the formulation.

In certain particular embodiments, the formulation comprises a ratio of API to excipient (w / w) between at least 10% API and at most 90% excipient, up to at most 30% API and at least 70% excipient. In certain other preferred embodiments, the formulation comprises a ratio of API to excipient (w / w) between at least 10% API and at most 90% excipient to at most 20% API and at least 80% excipient.

In certain more particular embodiments, the formulation may comprise an API to excipient ratio of about 30% API and about 70% excipient (w / w). In more preferred embodiments, the formulation may comprise an API to excipient ratio of about 20% API and about 80% excipient (w / w). In more preferred embodiments, the formulation may comprise an API to excipient ratio of about 10% API and about 90% excipient (w / w).

In a particular embodiment, the formulations of the present invention comprise 5% API, 10% API, 15% API, 20% API, 25% API, 30% API, 35% API, 40% API, 45% API or 50% API and 95% auxiliary, 90% auxiliary, 85% auxiliary, 80% auxiliary, 75% auxiliary, 70% auxiliary, 65% auxiliary, 60% auxiliary, 55% auxiliary or 50% auxiliary.

The inventors have discovered that such preferred ratios can result in an amorphous state of the formulation of the present invention and thereby exhibit improved solubility, dissolution rates and levels, achieve supersaturation state, and / or better bioavailability. compared to formulations that include a ratio other than the preferred ratios.

In particular, the formulation as described in this application provides that said API is Flubendazole and wherein said protein-based excipient is obtained from a protein composition or a hydrolyzate thereof comprising BSA and / or gelatin.

In particular, the formulation as described in this application provides for said formulation to be dosed in a solid dosage form, preferably a tablet, pill or capsule.

In certain embodiments, the formulation further comprises a solid dosage form, preferably a solid dosage form suitable for oral administration. In certain preferred embodiments, the formulation further comprises the form of a tablet, pill or capsule.

Having the API substantially amorphous in a solid dosage form can allow immediate oversaturation and can further help maintain this degree of oversaturation. Consequently, substantially amorphous having the formulation in a solid dosage form can exhibit similar advantages. For oral administration, a solid dosage form is the preferred industry and commercial form. All steps for processing and producing a solid dosage form are also known in the art. The excipient and formulations as described by the present invention have demonstrated suitability in obtaining and maintaining the structural and chemical properties expected from such a solid dosage form.

Having the API or formulation in a solid dosage form substantially amorphous can allow beneficial interactions with traditional (polymeric) excipients. The presence of at least one excipient such as a polymer that interacts, for example, may allow immediate oversaturation, and / or may also help maintain this degree of oversaturation. This process can be easily scalable and directly adaptable to create a commercial product. Traditional (polymeric) excipients do not reach the amorphous state observed for the protein-based excipient as described by the present invention. However, by mixing a protein vehicle with a traditional (polymeric) vehicle, the amorphous state of the formulation can be improved accordingly. In particular, amounts of gelatin or albumin can significantly increase the amorphous state of the formulation (experimentally verified). In particular embodiments, the formulation of the present invention further comprises at least one stabilizer. Administering a stable solid dosage can often be a challenge in producing a solid dosage form. Stabilizers as used in this application may include, but are not limited to, antioxidants, sequestering agents, emulsifiers, and surfactants, UV absorbers, extinguishers, scavengers, and the like. The choice of the stabilizer depends primarily on the properties of the API (e.g., sensitivities, activity) and the processing of the formulation.

In particular embodiments, the formulation of the present invention further comprises at least one additional excipient that is not described by the present invention. By combining different excipients an improved (synergistic) effect can be observed, which would further improve the solubility, dissolution rate and / or bioavailability so that they exceed those of the excipients when used separately. Typically, non-protein-based excipients are synthetic polymer excipients. The use of polymeric adjuvants is well known in the art, and may include, but is not limited to, polyvinylpyrrolidone (PVP), polyethylene oxide (PEO), hydroxypropylcellulose (HPC), hydroxypropylmethylcellulose acetate succinate (HPMCAS), ethylcellulose (EC), cellulose acetatebutyrate CAB), Cellulose acetate phthalate (CAP), Polyvinyl alcohol (PVA), Poly (ethylene glycol) (PEG), Poly (vinyl acetate) (PVAc), Methacrylates, Polylactide (PLA), Polyglycolide (PGA), Copolymers of PLA / PGA, Polycaprolactone (PCL) ), Ethylene vinyl acetate (EVA), Polyurethanes (TPU), Polyethylene (PE), Soluplus®, and the like.

Having the API or formulation in a solid dosage form substantially amorphous can allow beneficial interactions with traditional (polymeric) excipients. For example, the presence of at least one excipient such as a polymer interaction may allow immediate oversaturation, and / or may further help maintain this degree of oversaturation. This process can easily be scaled up and immediately adaptable to create a commercial product. Traditional (polymeric) excipients do not reach the amorphous state observed for the protein-based excipient as described by the present invention. However, by mixing a protein-based excipient with a traditional (polymeric) excipient, the amorphous state of the formulation can be improved accordingly. In particular, amounts of gelatin or albumin can significantly increase the amorphous state of the formulation (experimentally verified).

In certain particular embodiments, the formulation of the present invention further comprises at least one taste-masking agent. Providing effective protection against bitter tastes or unpleasant odors can often be a challenge when formulating a formulation. The formulation as described in this application can guarantee effective taste masking without compromising the release times of the API once administered (orally). Characteristic flavor masking ingredients may include, but are not limited to, Aspartame, Acesulfame Potassium, Sucralose, Citric Acid, Zinc Sulphate, Cyclodextrin (e.g. Beta, Gamma, Hydroxyl Propyl, etc.), flavor additives (e.g. lemon, peppermint, etc.) and such.

In certain particular embodiments, the formulation of the present invention further comprises at least one envelope layer. Providing effective protection against external influences can be an important requirement for a formulation. Multiple coating layers can further improve the chemical properties of the formulation; for example, by providing a catalytic effect on solubility; or can further prevent the formulation from dissolving before the desired target is achieved, for example by protecting a formulation for the intestines against gastric juices. In addition, certain enveloping layers can also act as a moisture-protecting component. If a solid dosage form (eg tablets) is exposed to moisture, it can swell and burst. A moisture protector can counterbalance excessive intake of moisture and thus prevent the solid dosage form from bursting. The use of an envelope layer is known in the art and examples may include, but are not limited to, methacrylate formulations, hydroxypropyl methylcellulose formulations (HPMC), polyvinyl alcohol formulations (PVA), Kollicoat® and the like.

In certain particular embodiments, the formulation according to the invention further comprises at least one surfactant. A surfactant can be used to reduce the surface tension of a liquid in which the formulation is dissolved. The surfactants used in this application may include, but are not limited to, pH modifiers, fillers, complexing agents, solubilizers, pigments, lubricants, lubricants, flavoring agents, plasticizers, taste masking agents, release modifying polymers, and the like.

In particular, the formulation as described in this application provides that said formulation is characterized in that it has a particle size between 1 µm and 1 mm; preferably between 5 pm and 50 pm; most preferably between 10 µm and 20 µm.

As used in this application, the term "particle size" refers to the size of the individual particles that are covered by a powder. Said particles have a diameter in the range between a nanometer and several millimeters. For the present invention, the particle size is preferably in the order of micrometers. Related to this is the "particle size distribution" (PSD) of a powder or particles dispersed in a liquid; which includes a list of values or a mathematical function that represents the relative amount, usually by weight, of particles present based on their size. The PSD can influence the reactivity of solids that participate in chemical reactions, and must be precisely regulated in pharmaceutical products.

The following usually applies to rapid release: the smaller the particle size, the faster the API goes into solution. However, for APIs that exhibit low solubility and / or bioavailability, the sooner they dissolve, the higher the local temporary concentration and the greater the pressure to precipitate as crystals. The inventors have thus found that for certain APIs it is better to have a larger particle size of up to a millimeter instead. Certain processing techniques, such as the drying processes, make it possible for the particle size to be controlled by means of the process parameters. The advantage of a customized particle size is that in this way the flow properties of the formulation can also be manipulated, for example by incorporating additives.

In certain particular embodiments, the protein-based excipient of the present invention is characterized by having a particle size of at least 0.1 µm to at most 1.0 mm; or at least 1 pm to a maximum of 50 pm; or at least 5 pm to a maximum of 40 pm. In a particular embodiment, the protein-based excipient of the present invention is characterized by having a particle size of at least 5 µm to at most 25 µm; more preferably at least 7 µm to at most 15 µm; most preferably about 10 µm.

In general, a smaller particle size can be associated with a better dissolution level and a better dissolution rate, which can further improve the attainment of a state of super-saturation and can help to maintain said state of super-saturation. It will be appreciated that processing steps attempting to reduce the particle size, e.g., by changing the solvent, the drying method or the mixing method, may also further contribute to the advantages described by the present invention. Additional post-processing steps that can cause physical stress on the native protein, leading to a loss of the quaternary, tertiary and / or secondary structure present in their native state, or a loss of the biological activity of the protein, will occur in the generally harmful to the purposes of the present inventions. Examples may include: further cutting, grinding, slicing, pressing, and the like.

In a particular embodiment, the BSA-based excipient is characterized by having a particle size of at least 0.1 µm to a maximum of 1.0 mm; or at least 1 pm to a maximum of 50 pm; or at least 5 pm to a maximum of 40 pm; preferably about 10 µm.

In a particular embodiment, the gelatin-based excipient is characterized by having a particle size of at least 0.1 µm to at most 1.0 mm; or at least 1 pm to a maximum of 50 pm; or at least 5 pm to a maximum of 40 pm; preferably about 15 µm.

The particle size described can be controlled to control the processing scale of the formulation in powder form. By obtaining a sufficiently small particle size that is essentially amorphous, higher total solids contents can be processed more easily in comparison to traditional (polymeric) excipients. In particular, an increase of 25% in additional cost efficiency and time savings can be achieved.

In another aspect, the present invention relates to the use of a protein composition or a hydrolyzate thereof, comprising proteins with a length of at least 10 amino acids as a protein-based excipient.

In another aspect, the present invention relates to a method for producing a pharmaceutical formulation comprising - a protein-based excipient obtained from a protein composition or a hydrolyzate thereof comprising proteins with a length of at least 10 amino acids; and - an active pharmaceutical ingredient (API); characterized in that said protein-based excipient and said API are both substantially amorphous; wherein said method comprises at least the steps of: (a) dissolving said API using a solvent to obtain a solution; and (b) drying the solution of step (a) to obtain a powder that is substantially amorphous.

In a particular embodiment of the present invention, said protein composition or hydrolyzate thereof is selected from soy protein, whey protein, pea protein, hydrolyzed whey protein isolate (HWPI), albumin, serum albumin, bovine serum albumin (BSA), hydrolyzed collagen, plasma proteins , gelatin, keratin, or a combination thereof, preferably in particular albumin or gelatin.

In certain particular embodiments, the solvent used to dissolve the protein composition or its hydrolyzate and / or the API is an organic acid; preferably the solvent is formic acid or acetic acid.

In essence, any acid suitable for dissolving a protein composition or hydrolyzate thereof without unduly affecting the protein structure (i.e., denaturation, caking, disintegration, combustion, etc.) may be suitable for the purposes of the present invention. However, the inventors have discovered that formic acid and acetic acid exhibit highly desirable properties for the purposes of the present invention. Both acids dissolve the proteins into a desirable protein solution without substantially denaturing said proteins (i.e., they retain their biological activity), thereby providing a protein solution that can be dried to obtain the protein-based excipient as described by the present invention. In particular, formic acid and acetic acid were found to exhibit highly desirable properties for dissolving albumin (e.g., BSA) and gelatin.

Similar considerations were made for solving the API; that is, essentially any acid suitable for dissolving APIs without adversely affecting the chemical structure or biological activity of the API may be suitable for the purposes of the present invention. In particular, formic acid and acetic acid have been found to exhibit desirable properties for dissolving Flubendazole, Carbamazepine, Griseofulvin, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, Jopanoic Acid, Itraconazole or Naproxen.

The term "drying" refers to any method or technique that promotes mass transfer of a liquid medium, such as water or other solvent, by evaporation. Specifically for the purposes of the present invention, drying includes any process that converts the liquid state of a solution (e.g., protein solution, API solution, protein API solution) into a solid, preferably powdered, state (e.g., dried on protein) excipient, dried API, dried formulation). It may be necessary to ensure that the drying process does not cause undesirable damage to the excipient or API, which can be caused by physical conditions such as excessive heat. Preferably, the drying method is selected from spray drying, freeze drying, vacuum drying, flash drying, agitator drying, air drying, condensation drying and the like.

In essence, any drying method suitable for drying a protein solution, or an API solution or a protein API solution without unduly affecting the chemical structure of the solution (s) may be suitable for the purposes of the present invention invention. However, the inventors have found that spray drying exhibits very desirable properties for the purposes of the present invention.

Spray drying dries said protein solution without substantially denaturing the dissolved proteins (i.e., they retain their biological activity), thereby obtaining a dried protein-based excipient as described by the present invention. In particular, spray drying has been found to exhibit highly desirable properties for drying albumin (e.g., BSA) and gelatin. Spray drying also dries said API solution without unduly affecting the chemical structure and biological activity of the API, thereby obtaining a dried API that is substantially amorphous, as described by the present invention. In particular, spray drying has been shown to have very desirable properties for drying Flubendazole, Carbamazepine, Griseofulvin, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, Jopanoic Acid, Itraconazole or Naproxen. In certain embodiments, spray drying is carried out at a temperature of at least 60 ° C to at most 240 ° C; or at least 110 ° C to at most 160 ° C; preferably at least 130 ° C to at most 150 ° C. In a most preferred embodiment, spray drying is carried out at a temperature of about 140 ° C.

The inventors have discovered that a temperature of 140 ° C provides an optimum temperature for effective spray drying of the protein solution without causing undesirable chemical or structural changes to the excipient.

The term "spray drying" as used herein refers to a preferred method of drying for the present invention, wherein a hot gas is used to produce a dry powder from a liquid solution (or suspension or slurry). The standard laboratory arrangement for putting spray drying into practice by a person skilled in the art usually comprises (1) a solution or suspension to be dried, (2) gas used for drying, (3) spray nozzles for spraying said solution or suspension; (4) a drying chamber, (5) a whirl chamber (6) connecting parts between the drying chamber and the whirl chamber and (7) a collection vessel. Other components or components that are not described in detail may also be included, depending on the production parameters (i.e., the types of solution or gas, types of nozzles, production scale and times, etc.). Examples of alternative components include other nozzles, e.g., high-pressure fluid nozzles for a single liquid, ultrasonic nozzles; or other parts such as turntables, atomizing wheels. In general, the spray drying arrangement is well known in the art and may include various models and techniques as is apparent to those skilled in the art.

In certain preferred embodiments, the protein-based excipient is characterized in that the excipient is obtained by dissolving or solubilizing a protein composition or a hydrolyzate thereof in an organic solvent to obtain a protein solution and (spraying) drying said protein solution to the protein-based solution based excipient. In certain other preferred embodiments, the protein-based excipient is characterized in that the excipient is obtained by dissolving or solubilizing a protein composition or a hydrolyzate thereof in formic acid to obtain a protein solution and (spray) drying said protein solution around the protein-based protein solution excipient. In certain other preferred embodiments, the protein-based excipient is characterized in that the excipient is obtained by dissolving or solubilizing a protein composition or a hydrolyzate thereof in acetic acid to obtain a protein solution and (spray) drying said protein solution around the protein-based protein solution excipient. .

It is generally believed that dissolving a protein in an organic acid at pH 1 followed by drying would affect its tertiary and secondary structure, thereby producing powders consisting of denatured protein. To their surprise, the inventors have observed that proteins retain their structure and biological activity after the process; in particular gelatin and albumin. Since their biological activity (i.e., the interaction with foreign molecules, such as the API) is dependent on the fully folded structure of the protein molecule, it was observed that the excipients obtained from acidic solutions of dried protein remained unaffected for at least a number of hours. These findings using gel permeation analysis are demonstrated in the examples section.

By drying, the amorphous state of the protein composition dissolved in an organic acid is preserved, thereby forming a powder that is substantially amorphous and substantially not denatured; in particular completely amorphous and not denatured. The resulting powder retains many preferred properties for use as an excipient; in particular a protein-based excipient. The same effects are observed for a powder obtained from a single protein source (e.g. BSA or gelatin), but also for a protein composition comprising multiple protein sources (e.g. BSA and gelatin).

In certain particular embodiments, the process for making the formulation of the present invention further comprises a stabilization process for producing a solid dosage form, such as a tablet, pill, or capsule with the desired size parameters. A stabilization step includes casting, compression and the like.

In particular, the method as described in this application provides for the protein-based excipient to be prepared according to the steps of: (i) dissolving a protein composition or hydrolyzate thereof using a solvent to obtain a solution; and (ii) drying the solution of step (i) to obtain said protein-based excipient.

In particular, the method as described in this application provides for the solutions of steps (a) and (i) to be dissolved using a common or different solvent.

In particular, the method as described in this application provides that the API and the protein-based excipient are: - dissolved and dried together in the same solvent, thereby forming said pharmaceutical formulation; - separately dissolved in the same solvent or different solvents and then dried together, thereby forming said pharmaceutical formulation; - dissolved in the same solvent or different solvents and dried separately and then mixed to form said pharmaceutical formulation.

In particular, the method as described in this application provides that the solvent is an organic acid, preferably formic acid or acetic acid.

In particular embodiments, the method of the present invention is characterized in that the protein-based excipient is obtained by dissolving or solubilizing a protein composition or a hydrolyzate thereof in a solvent to obtain a protein solution and drying said protein solution to a protein-based excipient and is further characterized in that the amorphous API is obtained by dissolving or solubilizing an API in a solvent, similar to or different from the solvent used for the protein solution, to obtain an API solution and drying said API solution to obtain an API that is substantially amorphous, and further combining said dried adjuvant and said dried API to obtain a formulation that is substantially amorphous according to an embodiment of the invention.

Alternatively, in certain embodiments, the method of the present invention is characterized in that the formulation is obtained by dissolving or solubilizing a protein composition or a hydrolyzate thereof in a solvent to obtain a protein solution, dissolving or solubilizing an API and a common or different solvent and then mixing the protein solution with the API and drying said mixture to obtain a formulation that is substantially amorphous according to an embodiment of the invention.

Alternatively, the method of the present invention is characterized in certain other embodiments in that the formulation is obtained by dissolving or solubilizing a protein composition or a hydrolyzate thereof together with an API in a common solvent and drying said protein API solution to obtain a formulation that is substantially amorphous according to an embodiment of the invention.

Alternatively, in certain other embodiments, the method of the present invention is characterized in that the formulation is obtained by dissolving or solubilizing an API in a solvent and drying said API solution to obtain an API that is substantially amorphous, and furthermore combining said dried API with a provided protein-based excipient that is preferably substantially not denatured to obtain a formulation that is substantially amorphous according to an embodiment of the invention.

In certain particular embodiments, the formulation further comprises a co-solubilizer. Certain APIs can prove difficult to dissolve in an acid or protein solution. A co-solubilizer can be used to make this possible. In particular, said co-solubilizer is selected from a cyclodextrin, sorbitan monostearate, a polyoxyethylene-polyoxypropylene block copolymer, polyoxyethylene glycerol triricinoleate 35, dimethylformamide, and the like.

The solubility and dissolution rate of the API used in the formulation of the present invention directly affects the bioavailability of the API. It is therefore very important to increase the solubility and the dissolution rate of the API, especially for APIs that exhibit low solubility and / or bioavailability. Achieving a state of oversaturation and subsequently maintaining said state of oversaturation for as long as possible results in even more favorable results of the API. In addition, an improvement in bioavailability and oversaturation increases the uptake speed of the API; resulting in a lower total weight / volume (dosage) required from the API in a formulation.

Alternatively, the excipient of the present invention as described in this application may be combined with at least one synthetic polymer excipient to obtain a formulation or formulation that may even have the properties to improve the solubility and / or bioavailability of both types of excipients separately. It can also further facilitate reaching a state of oversaturation, and then aids in maintaining said state of oversaturation for as long as possible. The formulation can thus form a solid dispersion.

The present invention encompasses various embodiments for obtaining the formulation, and those skilled in the art will understand further variations of the formulation and method of preparation with which similar effects as described in this application may be obtained. Examples of variations on the development of the formulation may include: (i) adding additional ingredients to the formulation, such as at least one additional (non-protein based) excipient, stabilizer, taste-masking component, coating layer, moisture-protecting component, surfactant, etc .; (ii) controlling the rate and extent of oversaturation of the formulation by optimizing said properties of the protein-based excipient; e.g., choosing other proteins, other protein sources, other protein compositions, etc .; (iii) implementing selective adaptations to the protein excipient to control the pharmacokinetic properties of the formulation, e.g., to control absorption (ie, control the rate and concentration of the API in the circulatory system), release (ie, control) of the time and place where the API is released from the formulation), the spread (ie stimulating or preventing the spread of the API through the body's fluids and tissues), the metabolism (ie protecting against or promoting the degradation of the API in body fluids and tissues) and excretion (ie, the safe removal of non-absorbed API from the body for certain cases where the API can irreversibly accumulate in body tissue); (iv) adjusting the properties and processing costs, time and / or scale of the formulation by optimizing the processing method, such as changing solvents, dissolving and drying methods, etc.

In another aspect, the present invention relates to a formulation as described in this application for use as a medicament.

In particular, said formulation is used to treat problems with the gastrointestinal tract (digestive system), problems with the heart vascular system, problems with the central nervous system, problems with the musculoskeletal system, problems with the respiratory system, problems with the endocrine system, problems with the reproductive system, problems with the urinary tract, problems with the immune system, obstetric problems and problems with gynecology (contraception) and / or for problems with the eyes, ears, nose, oropharynx or skin. The formulation as described by the present invention can be a product for treating infections and attacks (antibiotic, antifungal, antiparasitic), pain and consciousness (analgesics), allergic disorders, nutritional disorders, and / or neoplastic disorders. The formulation as described by the present invention can be a product for diagnostic use.

EXAMPLES

Example 1: Determining Preferred Protein Sources Various proteins from all natural sources (e.g. of animal, vegetable and microbial origin) are suitable for dissolving in an organic acid followed by drying and redissolving in a liquid system.

In the following experiment, 5% solutions of various proteins were dissolved in formic acid and cast into protein layers. The proteins were selected from gelatin, BSA (albumin), pea, soy, whey and zein (maize). The protein layers were then dissolved in 0.1 N HCl with a pH of 1.5 for 90 minutes at 37 ° C, after which the pH was adjusted to 6.8 for a maximum period of 330 minutes.

The solution of the protein layers was evaluated using spectrophotometry (extinction at 280 nm), and plotted as a function of time.

The solution profiles are set forth in FIG. 1, in which the average solution (%) of the different protein sources is displayed as a function of the dissolution time (minutes) and pH value. The legend is as follows: square - gelatin; circle - BSA (albumin); triangle - pea; lozenge - soy; stripe - whey; cross - zein (maize).

Summarizing the results, the protein layers of gelatin and BSA achieve a (nearly) complete solution, both at low and high pH values. Protein layers of pea, soy and whey show an intermediate solution level between 40 and 60%. Protein layers of zein show the lowest solution level of the series, which reaches about 10%.

Based on the data presented here, the protein layers of gelatin and BSA can be considered as the preferred proteins to act as protein-based adjuvants for rapid release formulations. While various other protein sources, such as pea and soy, have also demonstrated suitability for such purposes, these proteins and very poorly water-soluble layers such as those obtained with zein could be used for formulations that target delayed or controlled release profiles. In such specific embodiments, where a formulation with a lower dissolution rate or a lower dissolution level would be desired, the latter protein sources may even be preferred.

Example 2: Determining the effects of the processing technique on the structure of the protein.

The effects of the processing technique (i.e., the solvent and the drying method) on the native structure of the protein used to obtain a protein-based excipient were verified.

For the purposes of the present invention, various solvents may be suitable for dissolving a protein composition or a hydrolyzate thereof and / or an API; although an organic acid such as formic acid or acetic acid may be preferred. Similarly, different drying techniques may be suitable for drying a protein solution, an API solution or an API protein solution; however, standard techniques in the art such as spray drying or freeze drying may be preferred. For the purposes of this experiment, BSA was the protein source, formic acid acted as the solvent, and spray drying was chosen as the drying method.

The native structure of the BSA-based excipient was evaluated using gel permeation chromatography, which was able to separate the predominant monomer fraction from the dimer and trimer molecules. A raised portion of the latter molecules (i.e., dimers and trimers) acts as an indicator for structural loss of BSA.

First, the effect of formic acid as a solvent relative to a neutral solvent was studied, namely H2O. BSA was dissolved using the respective two solvents and spray dried under identical conditions. Subsequently, the resulting powder of the BSA-based excipient was dissolved in a phosphate buffer and analyzed using HPLC-based gel filtration chromatography.

The results are shown in FIG. 2, in which the extinction (AU) of BSA (albumin) is displayed as a function of time (min). The gray line represents BSA dissolved in H20; while the interrupted black line represents BSA dissolved in formic acid. There are no significant differences between the two samples, indicating that formic acid does not cause denaturation of BSA compared to H2 O. Similar results are expected for acetic acid.

The effect of formic acid was then compared for different incubation periods; namely 0, 4, 8 and 24 hours. BSA was dissolved using formic acid and the amounts of pH were neutralized at the indicated time intervals, followed by an analysis using HPLC-based gel filtration chromatography.

The results are shown in FIG. 3, showing the extinction (AU) of BSA (albumin) dissolved in formic acid as a function of time (min). The black line represents 0 hours, the black dashed line 4 hours, the gray line 8 hours and the dotted black line 24 hours. It can be concluded that incubation in formic acid for 0, 4 and 8 hours does not appear to significantly affect the quaternary structure of BSA. After about 24 hours of incubation, some signs of degradation of BSA are visible, as indicated by a higher fraction of larger aggregates, i.e., fewer monomers are present.

In conclusion, the formic acid solvent does not affect the native structure of BSA up to an 8-hour incubation period at room temperature. Similar results were observed for acetic acid and can be expected for similar organic acids.

Example 3: Verifying the effects of the processing technique on the biological activity of the protein.

Example 2 has already shown that up to 8 hours of incubation in formic acid did not appear to significantly affect the quaternary structure of BSA. For the purposes of the present invention, however, it is preferred that the protein-based excipient at least partially retains its biological activity. More preferably, it at least partially retains its biological activity under low pH conditions such as those present in the gastrointestinal tract. This biological activity can be beneficial for achieving a state of oversaturation and maintaining said state of oversaturation for a long period.

For the following experiment, the binding strength of a protein-based excipient was verified on a model API. For this experiment, BSA was the protein source, Flubendazole was used as the model API, formic acid acted as the solvent, and spray drying was chosen as the drying method. BSA was first dissolved in formic acid together with different concentrations of Flubendazole (FLU), then the solvent of the BSA-API solutions was evaporated and the dried formulations were dissolved in buffers with one of the pH values, pH 7.0 (neutral ), pH 4.0 (acid) and pH 1.0 (extremely acid). The fraction of FLU-bound BSA was determined on the basis of tryptophan quenching using a spectrofluorometer, a method for measuring equilibrium constants for BSA API formulations. The results are shown in FIG. 4, which shows the level of FLU-dependent tryptophan quenching as a function of the molar FLU concentration (μΜ). The circles represent the data obtained at pH 7.0; the squares represent pH 4.0; and the triangles represent pH 1.0. It was observed that the binding affinity decreased by a factor of two from pH 7.0 to pH 1.0, as derived by comparison of the binding strength, dissociation constant Kd, from 116 μΜ at pH 7, with the Kd from 234 μΜ at pH 1. At however, all pH values note that BSA retains its biological activity (ie binding activity), hence its native conformation at pH 1.0.

Although it is seen that the binding affinity of a BSA-based excipient decreases in an extremely acidic environment similar to that found in the stomach (ie pH 1.0), it can be concluded that its biological activity is maintained in all cases studied . Similar results can be observed for other protein sources (eg gelatin) and other APIs (eg Carbamazepine, Griseofulvin, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, jopanoic acid, Itraconazole, Naproxen).

Example 4: Determining the effects of a protein-based excipient on the supersaturation state of the formulation.

Various proteins or protein compositions were prepared from various protein sources at various concentrations to evaluate the extent of the effects of a protein-based excipient included in a formulation with regard to achieving a supersaturation state and maintaining said supersaturation state. Achieving high supersaturation can be considered as an indicator for improved bioavailability of an API included in said formulation.

The protein-based excipients were all based on gelatin extracted from various sources; the raw materials for the gelatin protein composition were extracted from pig skin or beef bone and / or a combination thereof; all produced by Rousselot®. Flubendazole was again chosen as the model API.

The dissolution profile and the supersaturation state (i.e. the available concentration) were tested using a dissolution medium that mimics the juice present in the human stomach for eating food (i.e., fasting state); namely FaSSGF (pH 1.6) available from Bio-relevant®. A suitable method for determining the concentration is reverse phase high performance liquid chromatography (RP-HPLC), in which molecules are separated on the basis of hydrophobicity, which causes different retention times as they flow out of the column. The other operating principles of RP-HPLC are known in the art. The RP-HPLC set-up used an Eclipse Zorbax Agilent column of 5 µm (4.6 x 150 mm) with a flow rate of 1 ml / min. The injection volume was 20 µl with the mobile phase ACN / TFA containing 0.1% (55:45) and the elution time of Flubendazole was about 1.9 minutes. The wavelength for the measurements was set at 280 nm.

Prior to the experiments, the solubility of Flubendazole in FaSSGF was determined to serve as a baseline reference value. The reference solution was prepared by introducing an excess of Flubendazole into 8 ml of FaSSGF solution. This solution was rotated for 72 hours and a 1 ml sample was taken every 24 hours. These reference samples were passed through a 0.45 µm Polytetrafluoroethylene filter (PTFE) to determine the Flubendazole concentration by the RP-HPLC method tested. The equilibrium concentration was achieved at any time when it was observed that the Flubendazole concentration remained unchanged. Said reference value was 11.2 pg / ml Flubendazole in FaSSGF medium.

The effect of the gelatin-based excipients on the supersaturation state of the formulation was then determined. Therefore, a gelatin-based excipient from various gelatin sources was added to obtain 0.1% protein solutions (8 mg w / v); namely gelatine from pig skin characterized by a Bloom value of 50, 75 and 225 g, gelatine from bovine bone characterized by a

Bloom value of 150 and 225 g, and gelatin peptides obtained from either pig skin or beef bone collagen characterized by an average molecular weight of 5,000 Da. With respect to equilibrium solubility, a twenty-fold excess of Flubendazole (final concentration 225 pg / ml) from a concentrated stock solution dissolved in formic acid was added to each solution to investigate the time window of supersaturation. All solutions were rotated and 1 ml samples were taken at the following time intervals: after 5, 15, 30, 60 and 120 minutes. Each sample was filtered through the 0.45 µm PTFE filter, diluted 1: 100 with the mobile phase and analyzed using RP-HPLC.

The results are shown in FIG. 5, wherein the average concentration of Flubendazole (pg / ml) as a function of the incubation time (min) in FaSSGF medium is shown for each formulation containing a different gelatin-based excipient. The high concentrations indicate that a state of oversaturation has been reached, and a plateau indicates that said oversaturation state has been maintained for a long time.

In FaSSGF medium a degree of supersaturation is maintained for all excipients for the duration of the experiment, and is characterized by a time-dependent exponential decrease within two hours after stirring, whereby the highest concentrations of Flubendazole are maintained in the presence of the gelatine from pig skin with low (50 and 75 g) Bloom values, and the gelatin from beef bone with high (225 g) Bloom value with 32 pg / ml (3x Cmax), 29 pg / ml (2.6x Cmax), and 32 pg / ml (3x Cmax).

In general, the results showed a significant improvement in the available concentrations of the poorly soluble API Flubendazole. All formulations comprising a gelatin-based excipient and Flubendazole achieved higher Flubendazole concentrations relative to the concentrations observed for the reference samples comprising Flubendazole alone. In addition, certain gelatin-based excipients achieved even significantly higher Flubendazole concentrations and maintained said high concentrations for a longer period of time.

The above results were verified using a protein-based excipient obtained from a whey protein composition. For the following experiment, two samples of a formulation comprising a whey protein-based excipient and Flubendazole were prepared under identical conditions. One sample was kept as a physical mixture and one sample was poured as a solid layer. Upon completion, both samples were released in 0.1 N HCl at 37 ° C to verify and maintain a state of supersaturation of Flubendazole. Samples were taken at the following time intervals: after 2, 5, 10, 20, 30, 60 and 120 minutes, and analyzed using RP-HPLC.

The results are shown in FIG. 6, wherein the average solution (%) of Flubendazole (FLUB) mixed with a whey protein-based excipient as a function of the dissolution time (min) is shown. The legend is as follows: square - whey protein - Flubendazole physical mixture; round - whey protein - Flubendazole layer.

These results clearly demonstrate that formulations comprising a whey protein-based excipient and Flubendazole create and preserve Flubendazole supersaturation in 0.1 N HCl at 37 ° C. In addition, the beneficial effects are shown in particular for solvent cast formulations.

As a general conclusion, the results show that protein-based excipients are actually able to promote a state of oversaturation in an environment of the gastrointestinal tract, and furthermore may help to maintain said oversaturation state for a lower period of time. These findings suggest that the bioavailability of poorly soluble APIs (e.g. Flubendazole, Carbamazepine, Griseofulvine, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, Jopanoic Acid, Itraconazole, Naproxen) prepared in at least one protein-based excipient (e.g., BSA, gelatin) may increase accordingly.

Example 5: Structural properties of formulations that include a gelatin-based excipient.

Three gelatin-based excipients were kept separate to further evaluate the structural properties (e.g., solid state and powder particle size) of formulations comprising said excipients after processing; in particular gelatin from pig skin with Bloom = 50 g, gelatin from pig skin with Bloom = 75 g, and gelatin from beef bone with Bloom = 225 g. Flubendazole was again chosen as the model API, formic acid acted as the solvent and spray drying was chosen as the drying method.

The three gelatin samples were all dissolved together with Flubendazole in formic acid, for four different ratios (% /%); i.e., [90-10], [80-20], [70-30] and [60-40] excipient / API (%). The protein API solutions were then spray dried under identical conditions to obtain very high-yield powder formulations. The reference sample for setting the standard calibration parameters included gelatin dissolved in formic acid without an API.

For the spray-drying device, both spray-drying with a bifluid and with an ultrasonic nozzle were tested, and they yielded comparable results. The solid state (i.e., crystalline or amorphous) of the formulations was evaluated using X-ray powder diffraction technology (XPRD). This technique uses the structural interference of X-rays with the crystallographic arrangement to determine the structure and phase formulation of the spray-dried powder formulation. The operating principles of XRD or XRPD are known in the art. Accordingly, the XRD experiments were performed using an automated X'pert PROdiffractometer (PANalystical, Netherlands) equipped with a Cu tube (Κα λ = 1.5418 A) with the generator set at 45 kV and 40 mA. Samples were applied to rotating sample containers with a background of zero. The measurements were performed in a continuous scanning mode from 4 ° to 40 ° with a step size of 0.0167 ° and a counting time of 400 s per step.

A suitable method for determining the particle size distribution (PSD) of the spray-dried powder formulation is dry powder laser diffraction technology. The particle size distributions were determined by measuring the angle variation in the intensity of the light that is scattered as a laser beam passes through the powder sample. The operating principles of dry powder laser diffraction are known in the art. Accordingly, the excipient powders were dispersed with compressed air at 3 bar by means of a RODOS dry dispersing device for size sorting with a HELOS laser diffraction sensor (Sympatec, the Netherlands) with a measuring range of: 0.9 -175 µm. The data on the angular scattering intensity were then analyzed to calculate the size of the particles responsible for creating the scattering pattern. The particle size is stated as a volume equivalent sphere diameter.

The results of the solid state analysis and particle size (PS) determination for formulations comprising different ratios of a gelatin-based excipient (conc. Auxiliary) and Flubendazole (conc. API) are shown in Table 1 below.

Table 1: Structural properties of formulations that include a gelatin-based excipient

A summary of Table 2: The average particle size (PS) of the formulations comprising a 50PS30-based excipient and Flubendazole is approximately 14.03 µm; that of a 75PS18-based excipient and Flubendazole is approximately 13.1 µm; and that of an adjuvant based on 225LB30 and Flubendazole is about 20.2 µm; thus, the overall average PS of the types of gelatin-based excipients is approximately 15.8 µm.

All formulations comprising gelatin with a relative excipient ratio of 90% were found to be completely amorphous. However, if the relative adjuvant ratio was lowered to 70% or lower, at least a portion of the powder particles was found to be semi-crystalline.

In general, a lower PS and a substantially amorphous state can be associated with better solubility and dissolution rates, and as a result can help to achieve a state of supersaturation more easily. In conclusion, the results teach that formulations comprising a higher ratio of excipient to API (% /%) may prove even more beneficial for the purposes of the present invention. Only the formulations that show a completely amorphous state were kept separate for further testing of their solubility and dissolution profiles.

Example 6: Structural properties of formulations comprising an excipient based on BSA.

Similar to Example 5, the structural properties of formulations comprising BSA-based excipients were evaluated after processing. The starting materials for the BSA protein composition were extracted from a bovine source. Flubendazole was again chosen as the model API, formic acid acted as the solvent and spray drying was chosen as the drying method. The BSA samples were all dissolved together with Flubendazole in formic acid for four different ratios (% /%); i.e., [90-10], [80-20], [70-30] and [60-40] excipient / API (%). The protein API solutions were then spray dried under identical conditions to obtain powder formulations with a very high yield. The reference sample for setting the standard calibration parameters consisted of BSA dissolved in formic acid without an API.

The properties were again evaluated using XPRD and dry powder laser diffraction technology, using the same parameters as set forth for Example 5. The result of both measurements is set forth in Table 2 below.

Table 2: Structural properties of formulations comprising an excipient based on BSA

A summary of Table 2: The average particle size (PS) of the formulations comprising a BSA-based excipient and Flubendazole is approximately 9.1 µm. All formulations were found to be completely amorphous, regardless of the relative ratio. In conclusion, the results teach that formulations comprising a BSA-based excipient and an API may prove particularly beneficial to the objects of the present invention.

Example 7: Explosion profiles of formulations comprising an excipient based on BSA or gelatin.

The formulations of Examples 5 and 6 that showed an amorphous state were tested for their solubility and dissolution levels; viz. pork gelatin with Bloom 50 g or 225 g: FLU (ratios of 80: 20% or 90: 10%, respectively) and BSA (70-90%): FLU (30-10%).

For the gelatin-based formulations, the solutions were run in 400 ml HCl 0.1M at 37 ° C for a period of 80 minutes. For the BSA-based formulations, the solutions were run in 400 ml of 0.1 M HCl at 37 ° C for 90 minutes, at which time the pH of the medium was adjusted using solid Na3 PO4 to a value of 6.8. The complete solution lasted 5 hours and 30 minutes. All tests were performed in duplicate and samples were taken after 5 "; 15 "; 30 "; 60 "; 80 "(gelatin-based formulations) or after 5"; 15 "; 30 "; 60 "; 80 "; 120 "; 180 "; 240 "and 330" (BSA-based formulations). Afterwards, the samples were filtered using 0.45 µm PTFE filters. The stock solution contained 350 pg / ml Flubendazole. The standards were prepared from the stock solution, diluted in ACN / TFA 0.1% (55:45) and linearity was observed between 1 pg / ml and 350 pg / ml. All tests were performed in plural for statistical purposes; the first set comprised a total of firstly 625 mg of amorphous solid dispersion comprising 20% Flubendazole and 80% pig skin gelatin with Bloom = 50 g; secondly, 625 mg of amorphous solid dispersion containing 20% Flubendazole and 80% pig skin gelatin with Bloom = 225 g; and third, 625 mg of powder containing 20% Flubendazole and 80% pig skin gelatin with Bloom = 50 g (i.e., a physical mixture). The second series comprised a total of, firstly, 1250 mg of amorphous solid dispersion containing 10% Flubendazole and 90% BSA; secondly, 625 mg of amorphous solid dispersion containing 20% Flubendazole and 80% BSA; thirdly 416 mg of powder containing 30% Flubendazole and 70% BSA; and fourth, 625 mg of powder containing 20% Flubendazole and 80% BSA (i.e., a physical mixture).

The dissolution profiles were determined using RP-HPLC, similar to Example 4. The RP-HPLC set-up used a 5 µm (4.6 x 150 mm) Eclipse Zorbax Agilent column with a flow rate of 1 ml / min . The injection volume was 20 µl with the mobile phase ACN / TFA containing 0.1% (55:45) and the elution time of Flubendazole being approximately ± 1.9 minutes. The wavelength for measurements was set at 280 nm.

The dissolution profiles are set forth in FIG. 7 and 8. FIG. 7, graph (a), is a graph of a dissolution profile showing the Flubendazole concentration CF | Ub (pg / ml) for each set of gelatin-based formulations as a function of the dissolution time (min); the values shown represent the calculated means for each experiment. FIG. 7, graph (b), is a graph of a dissolution profile showing the average percentage of release of Flubendazole (%) for each set of gelatin-based formulations as a function of the dissolution time (min); the values shown represent the calculated means for each experiment. FIG. 8, graph (a), is a graph of a dissolution profile in which for each set of BSA-based formulations the Flubendazole concentration CF | Ub (pg / ml) is displayed as a function of the dissolution time (min); the values shown represent the calculated means for each experiment. FIG. 8, graph (b), is a graph of a dissolution profile showing the average percent release of Flubendazole (%) for each set of BSA-based formulations as a function of the dissolution time (min); the values shown represent the calculated means for each experiment.

The legend of FIG. 7 is as follows: The solid line marked with a circle corresponds to a spray-dried formulation comprising 80% pig skin gelatin (Bloom = 50 g) and 20% FLU; the solid line marked with a square corresponds to a spray-dried formulation comprising 90% pig skin gelatin (Bloom = 225 g) and 10% FLU; a dashed line corresponds to a formulation comprising a physical mixture of 80% pig skin gelatin (Bloom = 50 g) and 20% FLU. The legend of FIG. 8 is as follows: the solid line marked with a circle corresponds to a spray-dried formulation comprising 90% BSA and 10% FLU; the solid line marked with a triangle corresponds to a spray-dried formulation comprising 80% BSA and 20% FLU; the solid line marked with a square corresponds to a spray-dried formulation comprising 70% BSA and 30% FLU; a dashed line corresponds to a formulation comprising a physical mixture of 80% BSA and 20% FLU.

In general, the results of the dissolution tests for all protein-based excipients showed a significant improvement in the solution of the poorly soluble API Flubendazole in a spray-dried amorphous solution. A plateau indicates that a state of oversaturation is reached and maintained for a longer period of time. Thus, an improved solubility of Flubendazole was observed as characterized by a faster dissolution rate, as indicated by the initial slope, and a higher dissolution level, as indicated by the higher maximum Flubendazole concentration.

For all three ratios (90-80-70%) tested in the set of BSA-dependent formulations, all Cma values (i.e. the maximum

Flubendazole concentration) observed that they increased to peak at low pH values, and decreased at higher pH, although they remained high.

More specifically, it was observed that the formulations comprising high ratios of excipient based on BSA, i.e. 90 and 80%, achieved very high release rates of about 80% and a Cmax of 240 pg / ml. The formulation comprising a lower, 70% ratio of BSA-based excipient showed slightly reduced results with a maximum release rate of approximately 70% and Cmax of 220 pg / ml; the latter results are still several times, almost 20 times higher than any comparable reference value.

The cause for the relative improvement appears to be that BSA binds with Flubendazole with a binding strength Kd of about ~ 234 pM at a pH of 1, about ~ 133 pM at a pH of 4, and about -116 pM at a pH value of 7, values that generally correspond to the pH profile associated with passage of an oral dosage form through the gastrointestinal tract. This is a strong indication that BSA will interact with Flubendazole in both the stomach and small intestine, causing Flubendazole to dissolve nearly 20 times better than the reference values.

In conclusion, it is thus disclosed that protein-based excipients are actually capable of promoting a state of supersaturation in the gastrointestinal tract environment, and could further assist in maintaining said state of supersaturation for a long time. These findings suggest that the bioavailability of poorly soluble APIs (e.g. Flubendazole, Carbamazepine, Griseofulvine, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, Jopanoic Acid, Itraconazole, Naproxen) prepared in at least one protein-based excipient can be increased accordingly.

Example 8: Combinations of the protein-based excipient with polymer-based excipients.

The structural properties of various formulations comprising firstly an API and secondly a polymer-based excipient, or a protein-based excipient, or a combination of a protein and a polymer-based excipient were analyzed and compared.

The polymer-based excipient chosen was Soluplus®; produced by Badische Anilin und Soda Fabrik (BASF). The protein-based excipients were all based on gelatin extracted from various sources; the starting materials for the gelatin protein composition were extracted from pig skin or beef bone and / or a combination thereof; all produced by Rousselot®. Flubendazole was again chosen as the model API.

Various solid dispersions were produced using the methods described above and obtained by spray drying. Afterwards, their properties were again evaluated using XRD using the same parameters as set forth for Example 5.

The results are set forth in FIG. 9, showing a graph of XRD patterns of the formulations comprising (1) 80% Soluplus®: 20% Flubendazole (FLU); (2) 40% polymer: 40% gelatin: 20% FLU; (3) 10% polymer: 70% gelatin: 20% FLU; (4) 80% gelatin: 20% FLU (4).

It appears that the formulation comprising only the polymer-based excipient (cf. 1) is in fact not completely amorphous, in contrast to the formulation comprising only the protein-based excipient (cf. 4), which is completely amorphous. However, by combining the polymer-based excipient with a protein-based excipient in a common formulation, the degree of amorphous state can be increased, and it is also observed that it improves with the increased relative ratio of protein-based excipient to polymer based excipient.

In conclusion, it thus appears that protein-based aids are actually able to further promote the amorphous state in combination with an API and a polymer-based aid. The combined formulations comprising a BSA-based excipient, a polymer-based excipient and an API may prove particularly advantageous for the purposes of the present invention.

Claims (15)

CONCLUSIONS (changes indicated) A formulation comprising: - a protein-based excipient obtained from a protein composition or a hydrolyzate thereof comprising proteins with a length of at least 10 amino acids; and - an active pharmaceutical ingredient (API); characterized in that said protein-based excipient and said API are both substantially amorphous and form a solid dispersion. A formulation according to claim 1, wherein the protein-based excipient is substantially non-denatured and / or retains at least part of its biological activity. A formulation according to any one of the preceding claims, wherein at least one protein of the protein composition or a hydrolyzate thereof is selected from soy protein, pea protein, gelatin, keratin, corn, wheat, hemp, rye, oats, peanut, millet, casein, albumin, whey protein (lactalbumin), hydrolyzed whey protein isolate (HWPI), hydrolyzed collagen, plasma proteins, serum albumin, bovine serum albumin (BSA), egg albumin, fish albumin, elastin, collagen or a combination thereof, preferably BSA and / or gelatin. A formulation according to any one of the preceding claims, wherein the API exhibits low solubility, dissolution level, supersaturation state and / or bioavailability. A formulation according to any one of the preceding claims, wherein the API is selected from the following list: Flubendazole, Carbamazepine, Griseofulvin, Phenytoin, Nifedipine, Verapamil, Azithromycin, Nitrofurantoin, Jopanoic Acid, Itraconazole or Naproxen, preferably Flubendazole. A formulation according to any one of the preceding claims, wherein said API is Flubendazole and wherein said protein-based vehicle is derived from a protein composition or a hydrolyzate thereof comprising BSA and / or gelatin. A formulation according to any one of the preceding claims, wherein said formulation is characterized by having a particle size between 1 μm and 1 mm; preferably between 5 μm and 50 μm; most preferably between 10 μm and 20 μm. A formulation according to any one of the preceding claims, wherein said formulation is dosed as a solid dosage form, preferably a tablet, pill or capsule. Use of a protein composition or a hydrolyzate thereof comprising proteins of at least 10 amino acids in length as a protein-based excipient. A method for producing a pharmaceutical formulation comprising: - a protein-based excipient obtained from a protein composition or a hydrolyzate thereof comprising proteins with a length of at least 10 amino acids; and - an active pharmaceutical ingredient (API); characterized in that said protein-based excipient and said API are both substantially amorphous and form a solid dispersion; wherein said method comprises at least the steps of: (a) dissolving said API using a solvent to obtain a solution; and (b) drying the solution of step (a) to obtain a powder that is substantially amorphous. The method for producing a pharmaceutical formulation according to claim 10, wherein the protein-based excipient is prepared according to the steps of: (i) dissolving a protein composition or hydrolyzate thereof using a solvent to obtain a solution; and (ii) drying the solution of step (i) to obtain said protein-based excipient. A method of producing a pharmaceutical formulation according to claims 10 and 11, wherein the solutions of steps (a) and (i) are dissolved using the same or a different solvent. A method according to any of claims 10 to 12, wherein the API and the protein-based excipient: are dissolved and dried together in the same solvent to form said pharmaceutical formulation; - separately dissolved in the same solvent or different solvents and then dried together to form the pharmaceutical formulation; - are dissolved in the same solvent or different solvents and dried separately and then mixed to form the pharmaceutical formulation. The method of any one of claims 10 to 13, wherein the solvent is an organic acid, preferably formic acid or acetic acid. The formulation of any one of claims 1 to 8 for use as a medicament.