WO2015071837A1

WO2015071837A1 - Complexes of cyclosporine a and its derivatives, process for the preparation thereof and pharmaceutical compositions containing them

Info

Publication number: WO2015071837A1
Application number: PCT/IB2014/065988
Authority: WO
Inventors: Erzsébet Réka ANGI; Tamás SOLYMOSI; Richard Balázs KÁRPÁTI; Zsófia FENYVESI; Zsolt ÖTVÖS; László MOLNÁR; Hristos Glavinas; Genovéva FILIPCSEI; Katalin Ferenczi; Gábor HELTOVICS
Original assignee: Druggability Technologies Holdings Ltd
Current assignee: Tavanta Therapeutics Inc
Priority date: 2013-11-12
Filing date: 2014-11-12
Publication date: 2015-05-21
Anticipated expiration: 2016-05-12
Also published as: HUP1300647A2

Abstract

The present invention relates to pharmaceutically accepted complex formulae comprising complexes of Cyclosporine A or derivatives thereof and complexation agents and pharmaceutically accepted excipients, process for the preparation thereof and pharmaceutical compositions containing them. The complex formulae of the present invention have improved physicochemical properties and decreased variability and food effect and are useful in the treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis and severe psoriasis.

Description

COMPLEXES OF CYCLOSPORINE A AND ITS DERIVATIVES, PROCESS FOR THE PREPARATION THEREOF AND PHARMACEUTICAL COMPOSITIONS

CONTAINING THEM

FIELD OF THE INVENTION

The invention is directed to a stable complex with increased apparent solubility and increased dissolution rate comprising as active compound Cyclosporine A or derivatives thereof, which is useful in the treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis. More specifically, the complex of the present invention possesses increased apparent solubility, permeability and enhanced biological performance including significantly decreased fed/fasted effect. The invention also relates to methods of formulating and manufacturing complex according to the invention, pharmaceutical compositions containing it, its uses and methods of treatment using the complex and its compositions.

BACKGROUND OF THE INVENTION

Background Regarding Cyclosporine A

Cyclosporine is a cyclic polypeptide immunosuppressant agent consisting of 11 amino acids. It is produced as a metabolite by the fungus species Beauveria nivea. Chemically, cyclosporine is designated as [R-[R*,R*-(E)]]-cyclic(L-alanyl-D-alanyl-N-methyl-L-leucylN- methyl-L-leucyl-N-methyl-L-valyl-3-hydroxy-N,4-dimethyl-L-2-amino-6-octenoyl-L-a- amino-butyrylN-methylglycyl-N-methyl-L-leucyl-L-valyl-N-methyl-L-leucyl).

O \ Cyclosporine A exhibits very poor solubility in water, therefore suspension and emulsion forms of the drug have been developed for oral administration and injection. Cyclosponne A was originally brought to market under the brand name Sandimmune.

Cyclosporine A is taken by orally or given by injection. The oral medication is available either as a liquid or in capsule form. Liquid cyclosporine is available in 50-ml (5000-mg) bottles (100 mg is equal to 1 ml). Cyclosporine A capsules are available in 100-mg, 50-mg, and 25-mg doses. Since 2002, a topical emulsion of Cyclosporine A for treating inflammation caused by keratoconjunctivitis sicca (dry eye syndrome) has been marketed under the trade name Restasis (0.05%). Inhaled Cyclosporine A formulations are in clinical development, and include a solution in propylene glycol and liposome dispersions.

Mechanism of Action: Cyclosporine A is a potent immunosuppressive agent which in animals prolongs survival of allogeneic transplants involving skin, heart, kidney, pancreas, bone marrow, small intestine, and lung. Cyclosporine A has been demonstrated to suppress some humoral immunity and to a greater extent, cell-mediated reactions such as allograft rejection, delayed hypersensitivity, experimental allergic encephalomyelitis, Freund's adjuvant arthritis, and graft vs. host disease in many animal species for a variety of organs.

The exact mechanism of action of Cyclosporine A is not known. Experimental evidence suggests that the effectiveness of Cyclosporine A is due to specific and reversible inhibition of immunocompetent lymphocytes in the GO- or Gl -phase of the cell cycle. T-lymphocytes are preferentially inhibited. The T-helper cell is the main target, although the T-suppressor cell may also be suppressed. Cyclosporine A also inhibits lymphokine production and release including interleukin-2 or T-cell growth factor (TCGF).

No functional effects on phagocytic (changes in enzyme secretions not altered, chemotactic migration of granulocytes, macrophage migration, carbon clearance in vivo) or tumor cells (growth rate, metastasis) can be detected in animals. Cyclosporine A does not cause bone marrow suppression in animal models or man.

Pharmacokinetics: The absorption of Cyclosporine A from the gastrointestinal tract is incomplete and variable. Peak concentrations (C_max) in blood and plasma are achieved at about 3.5 hours. C_max and area under the plasma or blood concentration/time curve (AUC) increase with the administered dose; for blood, the relationship is curvilinear (parabolic) between 0 and 1400 mg. As determined by a specific assay, C_max is approximately 1.0 ng/mL/mg of dose for plasma and 2.7-1.4 ng/mL/mg of dose for blood (for low to high doses). Compared to an intravenous infusion, the absolute bioavailability of the oral solution is approximately 30% based upon the results in 2 patients. The bioavailability of Cyclosporine A in Soft Gelatin Capsules (cyclosporine capsules, USP) is equivalent to Cyclosporine A Oral Solution, (cyclosporine oral solution, USP).

Distribution: Cyclosporine is distributed largely outside the blood volume. In blood, the distribution is concentration dependent. Approximately 33%-47% is in plasma, 4%-9% in lymphocytes, 5%-12% in granulocytes, and 41%-58% in erythrocytes. At high concentrations, the uptake by leukocytes and erythrocytes becomes saturated. In plasma, approximately 90% is bound to proteins, primarily lipoproteins.

Elimination: The disposition of cyclosporine from blood is biphasic with a terminal half-life of approximately 19 hours (range: 10-27 hours). Elimination is primarily biliary with only 6% of the dose excreted in the urine.

Metabolism: Cyclosporine is extensively metabolized but there is no major metabolic pathway. Only 0.1% of the dose is excreted in the urine as unchanged drug. Of 15 metabolites characterized in human urine, 9 have been assigned structures. The major pathways consist of hydroxylation of the Cy-carbon of 2 of the leucine residues, Cn-carbon hydroxylation, and cyclic ether formation (with oxidation of the double bond) in the side chain of the amino acid 3-hydroxyl-N,4-dimethyl-L-2-amino-6-octenoic acid and N demethylation of N-methyl leucine residues. Hydrolysis of the cyclic peptide chain or conjugation of the aforementioned metabolites do not appear to be important biotransformation pathways.

Cyclosporine A, unformulated, is characterized by low water solubility, low bioavailability and significant food effect when administered orally. The drug is marketed under the trade name Neoral. Neoral is a self-forming emulsion concentrate containing organic solvents and emulsifying agents developed to increase bioavailability and reduce food effect. Yet, significant food effect can still be observed for Neoral.

Another unfavorable pharmacokinetic property of the current Cyclosporine A formula is high inter- and intra-individual variability of blood levels following oral administration. This can become crucial in case of this active, as prolonged high plasma concentrations (repeated high Cmax) can result in renal toxicity, while too low trough plasma concentrations increase the frequency of organ transplant rejection. Part of the variability has been attributed to the food effect.

A variety of strategies have been used to attempt to overcome these issues, see for example WO/2001/017546, US20020016290, US5798333, JP2000229878, CN1625391, EP2079456, US6306434, and CN101810560.

DESCRIPTION OF THE INVENTION

Disclosed herein is a stable complex comprising as active compound selected from the group of Cyclosporine A or derivatives thereof; at least one agent selected from the group of a polyvinylcaprolactam-polyvinyl acetate-polyethylene-glycol graft copolymers, D-a- Tocopherol polyethylene glycol 1000 succinate (TPGS); poloxamers; polyvinylpyrrolidone; copolymers of vinylpyrrolidone and vinyl-acetate; and poly(maleic acid-co-methyl-vinyl- ether) (PMAMVE), preferably d-alpha tocopheryl polyethylene glycol 1000 succinate (TPGS). a) is instantaneously redispersable in physiological relevant media

b) is stable in solid form and in colloid solution and/or dispersion;

c) apparent solubility in water of at least 1 mg/mL;

d) shows X-ray amorphous character in the solid form;

e) has a PAMPA permeability of at least 0.1 * 10^"6 cm/s when dispersed in FaSSIF or FeSSIF biorelevant media, which does not decrease in time at least for 12 months; and f) is characterized by infrared (ATR) spectrum having main/characteristic absorption peaks at 1754 cm^"1, 1736 cm^"1, 1628 cm-¹ 1466 cm^"1, 1455 cm^"1, 1412 cm^"1, 1360 cm^" 1342 cm^"1, 1279 cm^"1, 1241 cm^"1, 1216 cm^"1, 1147 cm^"1, 1102 cm^"1, 1060 cm^"1, 962 cm^"1, 947 cm^"1 841 cm^"1, 588 cm^"1 , 529 cm^"1 , 509 cm^"1 preferable at 1628 cm^"1.

The invention is a complex formula having increased apparent solubility, permeability and decreased variability along with decreased fed/fasted effect.

We have found that only the selected combinations of complexation agents and pharmaceutically accepted excipients disclosed in the present invention result in a stable complex formula having improved physicochemical characteristics and decreased variability along with decreased fed/fasted effect.

The expression Cyclosporine A is generally used for Cyclosporine A, or its derivatives. In an embodiment, said complexation agent is selected from polyvinylcaprolactam-polyvinyl acetate-polyethylene-glycol graft copolymers, D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS); poloxamers; polyvinylpyrrolidone; copolymers of vinylpyrrolidone and vinyl-acetate; and poly(maleic acid-co-methyl-vinyl-ether) (PMAMVE), preferably d-alpha tocopheryl polyethylene glycol 1000 succinate (TPGS).

In an embodiment, said complexfurther comprises at least one additional active agent selected from the group of sodium-lauryl-sulfate, dioctyl sodium sulfosuccinate and polyoxyethylene- polyoxypropylene block copolymers, preferably sodium-lauryl-sulfate and polyoxyethylene- polyoxypropylene block copolymer terminated with primary hydroxyl groups (Mw= 10,000- 20,000).

In an embodiment, said additional active agent is selected from agents useful for the treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis.

In an embodiment, said complex has enhanced biological performance including significantly decreased fed/fasted effect. In an embodiment, said complex possesses at least two of the properties described in a) - f).

In an embodiment, said complex possesses at least three of the properties described in a) - f).

In an embodiment, said complex has an increased dissolution rate.

Further disclosed herein is a stable complex comprising an active compound selected from the group of cyclosporine A, or derivatives thereof; at least one complexation agent chosen poloxamers; polyvinylcaprolactam-polyvinyl acetate-polyethylene-glycol graft copolymers; polyvinylpyrrolidone; copolymers of vinylpyrrolidone and vinyl-acetate; poly(maleic acid- co-methyl -vinyl -ether) (PMAMVE) and D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS), and optionally at least one pharmaceutically accepted excipient selected from the group of sodium-lauryl-sulphate, dioctyl sodium sulphosuccinate, sodium-acetate and polyoxyethylene-polyoxypropylene block copolymers; wherein said complex obtainable via a mixing process.

In an embodiment, said complexation agent is a D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS). In an embodiment, said pharmaceutically acceptable excipients are sodium-lauryl-sulphate and polyoxyethylene-polyoxypropylene block copolymer terminated with primary hydroxyl groups (Mw=10,000- 20,000).

In an embodiment, said complex is obtained via a continuous flow mixing process. In an embodiment, a complex comprises a complexation agent which is a D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS) and a pharmaceutically acceptable excipients which are sodium-lauryl-sulphate and polyoxyethylene-polyoxypropylene block copolymer terminated with primary hydroxyl groups (Mw= 10,000- 20,000), in a total amount ranging from about 1.0 weight% to about 95.0 weight % based on the total weight of the complex. In an embodiment, a complex comprises a complexation agent which is a D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS) and a pharmaceutically acceptable excipients which are sodium-lauryl-sulphate and polyoxyethylene-polyoxypropylene block copolymer terminated with primary hydroxyl groups (Mw= 10,000- 20,000), in a total amount ranging from about 50.0 weight% to about 95.0 weight % based on the total weight of the complex. Further disclosed herein is a process for the preparation of the complex, comprising the steps of mixing a solution of Cyclosporine A, or derivatives thereof, and at least one complexation agent selected from poloxamers; polyvinylcaprolactam-polyvinyl acetate-polyethylene-glycol graft copolymers; polyvinylpyrrolidone; copolymers of vinylpyrrolidone and vinyl -acetate; poly(maleic acid-co-methyl-vinyl-ether) (PMAMVE) and D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS) in a pharmaceutically acceptable solvent with an aqueous solution containing at least one pharmaceutically acceptable excipient chosen from sodium- lauryl-sulphate, dioctyl sodium sulphosuccinate, sodium-acetate and polyoxyethylene- polyoxypropylene block copolymers.

In an embodiment, said process is performed in a continuous flow instrument. In an embodiment, said continuous flow instrument is a microfluidic flow instrument.

In an embodiment, said pharmaceutically acceptable solvent is chosen from methanol, ethanol, i-propanol, n-propanol, acetone, acetonitrile, dimethyl-sulfoxyd, or tetrahydrofuran, or combinations thereof. embodiment, said pharmaceutically acceptable solvent is methanol. In an embodiment, said pharmaceutically acceptable solvent and said aqueous solvent are miscible with each other.

In an embodiment, said aqueous solvent comprises 0.1 to 99.9% weight of the final solution.

In an embodiment, said aqueous solvent comprises 50 to 90% weight of the final solution. In an embodiment, said aqueous solvent comprises 50 to 80% weight of the final solution.

In an embodiment, said aqueous solvent comprises 50 to 70% weight of the final solution.

In an embodiment, said aqueous solvent comprises 50 to 60% weight of the final solution.

In an embodiment, said aqueous solvent comprises 50 % weight of the final solution.

In an embodiment, a pharmaceutical composition comprising the complex together with pharmaceutically acceptable carrier.

In an embodiment, said composition is suitable for oral, pulmonary, rectal, colonic, parenteral, intracisternal, intravaginal, intraperitoneal, ocular, otic, local, buccal, nasal, or topical administration.

In an embodiment, said composition is suitable for oral administration. In an embodiment, said complex is for use in the manufacture of a medicament for the treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis.

In an embodiment, said complex is used for the treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis. In an embodiment, a method of treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis comprises administration of a therapeutically effective amount of a complex or a pharmaceutical composition as described herein.

Further disclosed herein is a stable complex comprising a. 5 - 30%) by weight of Cyclosporine A, or derivatives thereof;

b. 10 - 50% by weight of a D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS); c. 1 - 35 % by weight of a sodium -lauryl-sulphate; and d. 30 - 80 % by weight of polyoxyethylene-polyoxypropylene block copolymer terminated with primary hydroxyl groups (Mw=l 0,000-20,000). wherein said complex has a controlled particle size in the range between 50 nm and 600 nm; and wherein said complex is not obtained via a milling process or by high pressure homogenization process, encapsulation process and solid dispersion process, but it is obtained by a mixing process, preferable continuous flow mixing process.

In an embodiment, said complex shows reduced fed/fasted effect based on in vivo studies.

In an embodiment, said complex shows slower and more prolonged absorption delivering the necessary blood levels for longer time which results in lower inter- and intra-patient variability of Cyclosporine A pharmacokinetics based on in vivo studies.

In an embodiment, said complex is instantaneously redispersable in physiological relevant media.

In an embodiment, said complex is stable in solid form and in colloid solution and/or dispersion.

In an embodiment, said complex has apparent solubility in water.

In an embodiment, said complex has a PAMPA permeability of at least 0.1 * 10^"6 cm/s when dispersed in FaSSIF or FeSSIF biorelevant media, which does not decrease in time at least for 12 months.

In an embodiment, said complex is characterized by infrared (ATR) spectrum having main/characteristic absorption peaks at least at 1628 cm^"1.

In an embodiment, said complex is characterized by infrared (ATR) spectrum having main/characteristic absorption peaks at least at 1754 cm^"1, 1736 cm^"1, 1628 cm^"1 1466 cm^"1, 1455 cm^"1, 1412 cm^"1, 1360 cm^"1, 1342 cm^"1, 1279 cm^"1, 1241 cm^"1, 1216 cm^"1, 1147 cm^"1, 1102 cm^"1, 1060 cm^"1, 962 cm^"1, 947 cm^"1, 841 cm^"1, 588 cm^"1, 529 cm^"1, 509 cm^"1, preferable at 1628 cm^"1.

The complexation agents and pharmaceutically acceptable excipients of the Cyclopsorine A complex formulae of the invention are selected from the group of pharmaceutically acceptable nonionic, anionic, cationic, ionic polymers, surfactants and other types of excipients. The complexation agents themselves or together with the pharmaceutically accepted excipients have the function to form a complex structure with an active pharmaceutical ingredient through non-covalent secondary interactions. The secondary interactions can form through electrostatic interactions such as ionic interactions, H-bonding, dipole-dipole interactions, dipole-induced dipole interactions, London dispersion forces, π-π interactions, and hydrophobic interactions. The complexation agents, pharmaceutically accepted excipients and active ingredients are selected from the group of complexation agents, pharmaceutically accepted excipients and active ingredients which are able to form such complex structures through non-covalent secondary interactions. In some embodiments, the compositions may additionally include one or more pharmaceutically acceptable excipients, auxiliary materials, carriers, active agents or combinations thereof. In some embodiments, active agents may include agents useful for the treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis. Another aspect of the invention is the complex formulae of the Cyclopsorine A with complexation agents and pharmaceutically acceptable excipients in which the complexation agents and pharmaceutically acceptable excipients preferably are associated or interacted with the Cyclosporine A especially as the results of the mixing process, preferably continuous flow mixing process. In some embodiment, the structure of the complex Cyclosporine A formula is different from the core-shell type milled particle, precipitated encapsulated particles, micelles and solid dispersions.

The pharmaceutical composition of the invention can be formulated: (a) for administration selected from the group consisting of oral, pulmonary, rectal, colonic, parenteral, intracisternal, intravaginal, intraperitoneal, ocular, otic, local, buccal, nasal, and topical administration; (b) into a dosage form selected from the group consisting of liquid dispersions, gels, aerosols, ointments, creams, lyophilized formulations, tablets, capsules; (c) into a dosage form selected from the group consisting of controlled release formulations, fast melt formulations, delayed release formulations, extended release formulations, pulsatile release formulations, and mixed immediate release and controlled release formulations; or (d) any combination of (a), (b), and (c). The compositions can be formulated by adding different types of excipients for oral administration in solid, liquid, local (powders, ointments or drops), or topical administration, and the like.

The compositions can be formulated by adding different types of pharmaceutically acceptable excipients for oral administration in solid, liquid, local (powders, ointments or drops), or topical administration, and the like.

A preferred dosage form of the invention is a solid dosage form, although any pharmaceutically acceptable dosage form can be utilized.

Solid dosage forms for oral administration include, but are not limited to, capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active agent is admixed with at least one of the following excipients: (a) one or more inert excipients (or carriers), such as sodium citrate or dicalcium phosphate; (b) fillers or extenders, such as starches, lactose, sucrose, glucose, mannitol, microcrystalline cellulose and silicic acid; (c) binders, such as cellulose derivatives, alginates, gelatin, polyvinylpyrrolidone, sucrose and acacia; (d) humectants, such as glycerol; (e) disintegrating agents, such as crospovidon, sodium starch glycolate, effervescent compositions, croscarmellose sodium,, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates and sodium carbonate; (f) solution retarders, such as acrylates, cellulose derivatives, paraffin; (g) absorption accelerators, such as quaternary ammonium compounds; (h) wetting agents, such as polysorbates, cetyl alcohol and glycerol monostearate; (i) adsorbents, such as kaolin and bentonite; and j) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. For capsules, tablets, and pills, the dosage forms may also comprise buffering agents.

Besides such inert diluents, the composition can also include adjuvants, such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents. Advantages of the complex Cyclosporine A formulae of the invention include, but are not limited to (1) physical and chemical stability, (2) instantaneous redispersibility, (3) stability in colloid solution or dispersion in the therapeutic time window, (4) decreased variability, (5) decreased fed/fasted effect and (6) good processability.

Beneficial features of the present invention are as follows: the good/instantaneous redispersibility of solid complex formulae of Cyclosporine A in water, biologically relevant media, e.g.; physiological saline solution, pH=2.5 HC1 solution, FessiF and FassiF media and gastro intestinal fluids and adequate stability in colloid solutions and/or dispersion in the therapeutic time window.

One of the preferred characteristics of the complex Cyclosporine A formulae of the present invention is their increased apparent solubility and permeability. In some embodiments, the permeability of the complex Cyclosporine A formulae is at least 0.1 * 10^"6 cm/s, respectively.

Another preferred characteristic of the complex Cyclosporine A formulae of the present invention relates to the enhanced pharmacokinetic performance of the complex Cyclosporine formulae. In some embodiments, the complex Cyclosporine formulae have slower and more prolonged absorption delivering the necessary blood levels for longer time when compared to the conventional Cyclosporine formulation.

Another aspect of the invention is a pharmaceutical composition comprising a stable complex Cyclosporine A formulae or composition of them according to the invention and optionally pharmaceutically accepted auxiliary materials formulated for oral administration having decreased variability and eliminated fed/fasted effect compared to the commercial formulation of Cyclosporine A.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1. Complexation agent screening for formula selection

Figure 2. Particle size of redispersed solid complexes in colloid solution

Figure 3. Comparative PAMPA assays of complex Cyclosporine A formula, the marketed drug and the unformulated compound

Figure 4. SEM photo of complex Cyclosporine A Figure 5. ATR spectra of crystalline Cyclosporine A (A), amorphous Cyclosporine A (B), complex Cyclosporine A (C), placebo sample (D), and D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS)(E), polyoxyethylene-polyoxypropylene block copolymer terminated with primary hydroxyl groups (Mw=10,000-20,000) (Lutrol F108) (F) and Sodium-lauryl- sulfate (G)

Figure 6. Plasma concentration of Cyclosporine A following the oral administration of the marketed drug (Neoral) and Cyclosporine A complex to rats under the fasting conditions (n=4, dose: 15 mg/kg).

Figure 7. Pharmacokinetic parameters following the oral administration of the marketed drug (Neoral) and the Cyclosporine A complex to rats under the fasting conditions (n=4, dose: 15 mg/kg).

EXAMPLES

1. Selection of complexation agents and pharmaceutically accepted excipients for the production of complex Cyclosporine A formulae

Several complexation agents and pharmaceutically accepted excipients and their combinations were tested in order to select the formulae having instantaneous redispersibility.

2. Process for producing stable colloid solution and or dispersion of solid complex Cyclosporine A formula

Colloid solution of Cyclosporine A complex formula of the present invention was prepared by continuous flow mixing process in a flow instrument. As a starting solution, 2g Cyclosporine A and 3g TPGS dissolved in 100 mL methanol was used. The prepared solution was passed into the instrument with 4 mL/min flow rate. Meanwhile, antisolvent containing 10 g sodium- lauryl-sulphate and 2.25 g polyoxyethylene-polyoxypropylene block copolymer terminated with primary hydroxyl groups (Mw=10,000-20,000) in 500 mL water was passed into the instrument with 16 mL/min flow rate, where Cyclosporine A was mixed to form complex Cyclosporine A composition. The colloid solution of the complex Cyclosporine A is continuously produced at atmospheric pressure. The produced colloid solution was frozen on dry-ice and then it was lyophilized using Scanvac CoolSafe 110-8 freeze drier equipped with - 110°C ice condenser, with a Vacuubrand RZ6 vacuum pump. Particle size of complex Cyclosporine A was measured in reconstituted colloid solution right after the lyophilization. 3. Comparative in vitro PAMP A assays

PAMPA permeability measurements were performed as described by M. Kansi et al. (Journal of medicinal chemistry, 41, (1998) pp 1007) with modifications based on S. Bendels et al (Pharmaceutical research, 23 (2006) pp 2525). Sample containing the reference compound was a suspension of crystals visible by the naked eye, while samples of the marketed drug and the novel complex were opalescent colloid solutions. Permeability was measured in a 96-well plate assay across an artificial membrane composed of dodecane with 20% soy lecithin supported by a PVDF membrane (Millipore, USA). The receiver compartment was phosphate buffered saline (pH 7.0) supplemented with 1% sodium dodecyl sulfate. The assay was performed at room temperature; incubation time was 1-24 hours. The concentration in the receiver compartment was determined by HPLC.

PAMPA permeability of complex Cyclosporine A formula of Example 2 and the marketed drug was in the 2-3 * 10^"6 cm/s range, while PAMPA permeability of the reference compound was in the 0.5-1 * 10^"6 cm/s range with higher apparent permeability measured in the fed state simulation than in the fasted state simulation in all cases.

4. Structural analysis

Morphology of complex Cyclopsorine A was investigated using FEI Quanta 3D scanning electron microscope. Complex Cyclosprine A of the present invention consists of spherical particles (Figure 4.).

Structural analysis was performed by using Bruker Vertex 70 FT-IR spectrometer with Bruker Platinum diamond ATR unit. Continuous flow mixing of Cyclosprine A in the presence of selected pharmaceutically accepted excipients, such as D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS), sodium-lauryl-sulphate and polyoxyethylene-polyoxypropylene block copolymer terminated with primary hydroxyl groups (Mw=l 0,000-20, 000) resulted in a stable complex of Cyclosporine A. In a preferred embodiment the complex or the pharmaceutical composition according to the invention characterized by at least one of the following characteristic Raman or ATR band/peak (Figure 8):

1754 cm^"1, 1736 cm^"1, 1628 cm^"1 1466 cm^"1, 1455 cm^"1, 1412 cm^"1, 1360 cm^"1, 1342 cm^"1, 1279 cm^"1, 1241 cm^"1, 1216 cm^"1, 1147 cm^"1, 1102 cm^"1, 1060 cm^"1, 962 cm^"1, 947 cm^"1, 841 cm^"1, 588 cm^"1, 529 cm^"1, 509 cm^"1, preferable at 1628 cm^"1. 5. Comparative in vivo pharmacokinetic tests

Male Wistar rats (220-270 g) were treated orally with test articles one time via esophageal gavage technique following overnight fasting. Whole blood was sampled from the saphenous vein into heparinized tubes (approx. 0.1 mL to each tube) before and at 0.5, 1, 2, 3, 5, 7, 9, 12, 24, 48, and 72 hours after the oral administration of the test articles. Whole blood samples were stored at minus 20°C until analysis. Cyclosporine A concentrations were determined from whole blood samples by LC-MS/MS technique.

Pharmacokinetic data were expressed as measured values and as normalized values. Measured values were normalized to the extraction recovery of the standard solution from whole blood. Extraction recovery of the standard solution from whole blood varied between 88.83% and 111.67%) (93.99%) in average). C_max, AUCo-72, AUCo-inf, Frelo-72 and F_reio-inf of complex Cyclosporine composition of the present invention following oral administration were significantly decreased (p<0.05), however, t_max was not changed as compared to same pharmacokinetic parameters of Neoral solution. Relative bioavailability of complex Cyclosporine A was approximately 30%> less than that of Neoral solution, i.e. Freio-72 and Freio-inf were both between 0.70 and 0.71 as calculated from both normalized and not normalized data.

The absorption rate of the complex Cyclosporine A of the present invention was lower resulting in longer absorption phase with lower blood concentrations in the early phases, yet, trough concentrations measured at 24 hours were similar indicating that despite of the lower absorption rate the therapeutically relevant concentrations are maintained by both formulae in the same time window. The longer absorption phase, however, might result in less variability.

Claims

1. A stable complex with increased apparent solubility and increased dissolution rate comprising as active compound selected from the group of Cyclosporine A or derivatives thereof; optionally an additional active agent; at least one complexation agent selected from the group of poloxamers; polyvinylcaprolactam-polyvinyl acetate-polyethylene-glycol graft copolymers; polyvinylpyrrolidone; copolymers of vinylpyrrolidone and vinyl-acetate; poly(maleic acid-co-methyl-vinyl-ether) (PMAMVE) and D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS) said complex characterized in that it possesses at least one of the following properties: a) is instantaneously redispersable in physiological relevant media;

b) has increased dissolution rate;

c) is stable in solid form and in colloid solution and/or dispersion;

d) increased apparent solubility in water;

e) has a PAMPA permeability of at least 0.1 * 10^"6 cm/s when dispersed in FaSSIF or FeSSIF biorelevant media, which does not decrease in time at least for 12 months;

f) is characterized by infrared (ATR) spectrum having main/characteristic absorption peaks at least at 1628

absorption peaks; and g) said complex has slower and more prolonged absorption delivering the necessary blood levels for longer time when compared to the conventional Cyclosporine formulation; h) said complex has decreased variability and eliminated fed/fasted effect compared to the commercial formulation of Cyclosporine A.

2. The complex according to Claim 1, wherein said complexation agent is selected from the group of poloxamers; polyvinylcaprolactam-polyvinyl acetate-polyethylene- glycol graft copolymers; polyvinylpyrrolidone; copolymers of vinylpyrrolidone and vinyl-acetate; poly(maleic acid-co-methyl-vinyl-ether) (PMAMVE) and D-a- Tocopherol polyethylene glycol 1000 succinate (TPGS), preferably D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS).

3. The complex according to Claims 1 or 2, wherein said complex further comprises at least one pharmaceutically acceptable excipient selected from the group of sodium- lauryl-sulphate, dioctyl sodium sulphosuccinate, sodium-acetate and polyoxyethylene- polyoxypropylene block copolymers, preferably the pharmaceutically accepted excipients are sodium-lauryl-sulfate and polyoxyethylene-polyoxypropylene block copolymers terminated with primary hydroxyl groups (Mw=10,000- 20,000).

4. The complex according to any of Claims 1 to 3, wherein said complex further comprises one or more additional active agents, preferable the additional active agent is selected from the group of agents useful for the treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis.

5. A stable complex according to any of Claims 1 to 4 comprising an active compound selected from the group of Cyclosporine A, or derivatives thereof; at least one complexation agent selected from the group of poloxamers; polyvinylcaprolactam- polyvinyl acetate-polyethylene-glycol graft copolymers; polyvinylpyrrolidone; copolymers of vinylpyrrolidone and vinyl-acetate; poly(maleic acid-co-methyl-vinyl- ether) (PMAMVE) and D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS); and at least one pharmaceutically acceptable excipient selected from the group of sodium-lauryl-sulphate, dioctyl sodium sulphosuccinate, sodium-acetate and polyoxyethylene-polyoxypropylene block copolymers; wherein said complex obtained via a mixing process, preferable continuous flow mixing process, more preferable microfluidic flow mixing process.

6. A complex according to any of Claims 1 to 5 comprising a complexation agent which is a D-a-Tocopherol polyethylene glycol 1000 succinate and pharmaceutically acceptable excipients which are sodium-lauryl-sulfate and polyoxyethylene- polyoxypropylene block copolymers terminated with primary hydroxyl groups (Mw=10,000- 20,000), in a total amount ranging from about 1.0 weight% to about 95.0 weight % based on the total weight of the complex.

7. A process for the preparation of the complex according to any of Claims 1 to 6, comprising the steps of mixing a solution of Cyclosporine A, or derivatives thereof, and at least one complexation agent selected from the group of poloxamers; polyvinylcaprolactam-polyvinyl acetate-polyethylene-glycol graft copolymers; polyvinylpyrrolidone; copolymers of vinylpyrrolidone and vinyl-acetate; poly(maleic acid-co-methyl-vinyl-ether) (PMAMVE) and D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS) in a pharmaceutically acceptable solvent with an aqueous solution containing at least one pharmaceutically accepted excipient selected from the group of sodium-lauryl-sulphate, dioctyl sodium sulphosuccinate, sodium-acetate and polyoxyethylene-polyoxypropylene block copolymers.

8. The process according to Claim 7, wherein said process is performed in a continuous flow instrument, preferable in microfluidic instrument.

9. The process according to Claims 7 or 8, wherein said pharmaceutically acceptable solvent is selected from the group of methanol, ethanol, isopropanol, n-propanol, acetone, acetonitrile, dimethyl-sulfoxide, tetrahydrofuran, and combinations thereof, preferable the solvent is methanol.

10. The process according to any of Claims 7 to 9, wherein the solvent and the aqueous solvent are miscible with each other and the aqueous solvent comprises 0.1 to 99.9% weight of the final solution.

11. A pharmaceutical composition comprising the complex according to any of Claims 1 to 6 together with pharmaceutically acceptable carrier.

12. A pharmaceutical composition according to Claim 11, wherein said composition is suitable for oral, pulmonary, rectal, colonic, parenteral, intracisternal, intravaginal, intraperitoneal, ocular, otic, local, buccal, nasal, or topical administration, preferable the composition is suitable for oral administration.

13. A complex according to any of Claims 1 to 6 for use in the manufacture of a medicament for the treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis.

The use of the complex according to any of Claims 1 to 6 for the treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis.

15. A method of treatment of transplant (kidney, liver, and heart) rejection, rheumatoid arthritis, severe psoriasis comprising administration of a therapeutically effective amount of the complex according to any of Claims 1 to 6 or the pharmaceutical composition according to Claim 11 orl2.

16. A method for reducing the therapeutically effective dosage of Cyclosporine A compared to orally available formulations, said method comprising oral administration of a pharmaceutical composition according to Claims 11 or 12.

17. A stable complex comprising a) 5 - 30% by weight of Cyclosporine A, or derivatives thereof;

b) 10 - 50% by weight of a D-a-Tocopherol polyethylene glycol 1000 succinate (TPGS);

c) 1 - 35 % by weight of a sodium-lauryl-sulphate; and

d) 30 - 80 % by weight of polyoxyethylene-polyoxypropylene block copolymer terminated with primary hydroxyl groups (Mw=l 0,000-20,000). wherein said complex has a controlled particle size in the range between 50 nm and 600 nm; and wherein said complex is obtained according to any of claims 7 to 10.