CA2899746A1 - Pharmaceutical composition comprising leflunomide - Google Patents

Abstract

The present invention relates to a pharmaceutical composition according to the invention containing 14 to 17.5 mg leflunomide prepared as a single dose and uses thereof.

Description

Alfred E. Tiefenbacher (GmbH & Co. KG), Hamburg; medac GmbH, Wedel Pharmaceutical Composition comprising Leflunomide Technical Field:
The present invention relates to an inventive pharmaceutical composition comprising 14 to 17.5 mg leflunomide arranged in a single dose and the uses thereof.
Prior Art:
The pharmaceutically active agent leflunomide (formula I) with the chemical name 5-methyl-N-[4-(trifluoromethyl)phenyI]-1,2-oxazole-4-carboxamide is for some time now being used as a disease modifying antirheumatic drug (also abbreviated as: DMARDs = disease modifying antirheumatic drugs) from the group of isoxazoles. Besides the use as an antirheumatic drug in the treatment of rheumatoid arthritis and/or psoriasis arthritis the efficacy of leflunomide is presently being exam-ined in the treatment of further autoimmune diseases, in particular lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (Wegener's granulomatosis); in the course of solid organ transplantation, in particular transplantation of kidney and liver; oncologic diseases, in particular melanoma, sarcoma, glioma, prostate carcinoma, brain-and/or CNS tumors; and/or of diseases correlated with the human immune deficiency virus (HIV).
,0 =
N\icx H

N
N
. 101 Leflunomide itself represents a prodrug, which is after oral application metabolized in the intestinal wall, but also in the liver by opening the isoxazole ring to the active malono nitril amide metabolite and active agent teriflunomide (formula II, also known as A77 1726), with immunomodulatory properties. Concentrations of leflunomide following oral application are detectable in vivo only in very low concentrations. The mode of action of leflunomide is essentially based on the effective-

2 ness of the active metabolite teriflunomide, which according to the present state of art on the one hand inhibits dihydroorotate dehydrogenase and on the other hand shows anti proliferative and/or anti-inflammatory properties.
The maximum blood concentration of teriflunomide can be measured 6 to 12 hours after oral appli-cation. Due to the relatively long half-life of teriflunomide (appr. 2 weeks) steady state blood con-centrations can be measured after approximately 2 months for a once daily application of 10 or 20 mg leflunomide respectively. As reliable propositions on the individual efficacy of le-flunomide/teriflunomide and a possible adaptation by increase or reduction of the dose can only be conducted after entering the steady state concentration, attempts have been made to reduce the time until entering the steady state concentration.
By oral application of 100 mg leflunomide once daily for 3 days (loading dose) and subsequent application of 20 mg or 10 mg leflunomide once daily (maintenance dose) respectively, the time span until entering the steady state concentration can be reduced from approximately 2 months to approximately 2 weeks, wherein the therapeutic efficacy can generally be expected after 4 to 6 weeks and can even increase during the following 4 to 6 months.
According to the prescribing information of Arava , a commercially available pharmaceutical prepa-ration of leflunomide tablets with 10 mg, 20 mg and 100 mg leflunomide, it is thus recommended to start the respective treatments of active rheumatoid arthritis or active psoriasis arthritis with a loading dose of 100 mg once daily for 3 days. In case of treating active rheumatoid arthritis, the maintenance dose shall subsequently range from 10 to 20 mg leflunomide once daily depending on the severity (activity) of the disease; when treating active psoriasis arthritis 20 mg leflunomide shall be applied as the subsequent maintenance dose. Side effects have been observed for the respective dose regime, wherein the following side effects have been observed often: increase of blood pressure, leukopenia, paresthesia, headaches, dizziness, gastrointestinal disorders, such as diarrhoea, nausea, vomitus, diseases of the oral mucosa, and/or abdominal pain, increased hair loss, eczema, skin eruption, pruritus, dry skin, tenosynovitis, increase of creatinine kinase (CK), loss of appetite, weight loss, asthenia, light allergic reactions and increased liver values (transami-nases, gamma-GT, alkaline phosphatases, bilirubin). Based on one or more of these side effects, in particular with hematotoxic and/or hepatotoxic and/or gastrointestinal effects the therapy may be discontinued or the active agent may be changed within 12 months following start of therapy.
In addition, some publications discuss the influence of inflammatory reactions, which are present, e.g., with rheumatoid arthritis on the pharmacokinetic of the active agents.
As an example for being influenced by inflammatory reactions, metabolising enzymes, transporter proteins and/or plasma binding proteins are mentioned. For chronic inflammatory reactions the metabolic system of the

3 liver can be overloaded or saturated with inflammatory products, so that this can amend the me-tabolism of an active agent and can, e.g., result in a reduction of the metabolism of the active agent and in an increase of the plasma level of the active agent.
It is now the aim of the present invention to provide a pharmaceutical composition comprising leflunomide, which in comparison to an application of 20 mg or 10 mg leflunomide once daily re-spectively = shows a comparable or enhanced efficacy in the prophylaxis and/or treatment of autoimmune diseases, preferably of rheumateid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (Wegener's granulomatosis); and/or in the prophylax-is and/or treatment in the course of organ transplantations, preferably of kidney transplanta-tion and/or liver transplantation; and/or in the prophylaxis and/or treatment of oncologic dis-eases, preferably melanomas, sarcomas, gliomas, prostate carcinomas, brain and/or CNS
tumors; and/or in the prophylaxis and/or treatment of diseases correlated with the human immune deficiency virus (HIV) and/or = provides a reduction in one or more of the aforementioned side effects, preferably one or more hematotoxic and/or hepatotoxic and/or gastrointestinal side effects and/or = provides a reduction of discontinuation of leflunomide treatment or a change to alternative ac-tive agents.
Short Description of the Invention:
One or more of the aforementioned advantages of the inventive problem are solved by the in-ventive subject matter of the claims. Advantageous embodiments are disclosed in the dependent claims as well as in the following description.
Accordingly, the first inventive aspect relates to a pharmaceutical composition comprising leflunomide, characterized in that the composition is arranged as a single dose with 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide.
A second inventive aspect relates to the use of leflunomide in the prophylaxis and/or treatment of of an autoimmune disease, preferably of rheumatoid arthritis, psoriasis arthritis, arthritis as compli-cation of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (VVegener's granulomatosis); and/or in the prophylaxis and/or treatment in the course of an organ transplantation, preferably of kidney transplantation

4 and/or liver transplantation; and/or in the prophylaxis and/or treatment of oncologic diseases, preferably melanomas, sarcomas, gliomas, prostate carcinomas, brain and/or CNS
tumors; and/or in the prophylaxis and/or treatment of diseases correlated with the human immune deficiency virus (HIV), characterized in that leflunomide is arranged or arrangeable as single dose of a pharmaceu-tical composition with 14 to 17.5, preferably 15 to 17.5, more preferably 15 mg leflunomide and is applied once, twice, three or four times daily.
The inventive aspects as disclosed hereinbefore can comprise - in case it is reasonable for a per-son skilled in the art - any combination of the preferred inventive embodiments, which are dis-closed hereinafter and in particular in the dependent claims.
Detailed Description of the invention:
The inventors of the present invention have surprisingly found out, that an inventive pharmaceuti-cal composition based on the single dose comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide per pharmaceutical formulation and a respective once, twice, three or four times daily (simultaneous or subsequent) application in the prophylaxis and/or treat-ment of autoimmune diseases, preferably of rheumatoid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (Wegener's granulomatosis);
and/or in the prophy-laxis and/or treatment in the course of an organ transplantation, preferably of kidney transplanta-tion and/or liver transplantation; and/or in the prophylaxis and/or treatment of oncologic diseases, preferably melanomas, sarcomas, gliomas, prostate carcinomas, brain and/or CNS
tumors; and/or in the prophylaxis and/or treatment of diseases correlated with the human immune deficiency virus (HIV) = shows a comparable or enhanced efficacy when compared to the standard therapy with 20 mg or 10 mg leflunomide respectively arranged as a single and/or = provides a reduction in one or more of the aforementioned side effects, in particular increase of blood pressure; hematotoxic side effects, such as, e.g., leukopenia;
paresthesia; head-aches; dizziness; gastrointestinal disorders, such as diarrhea, nausea, vomitus, diseases of the oral mucosa, and/or abdominal pain; increased hair loss; dermatologic side effects, such as, e.g., eczema, skin eruption, pruritus and/or dry skin; tenosynovitis;
increase of creatinine kinase (CK); loss of appetite; weight loss; asthenia; light allergic reactions and/or hepatotoxic side effects, preferably characterized by increased liver values (transaminases, gamma-GT, alkaline phosphatases, bilirubin), more preferably a reduction of one or more hematotoxic and/or hepatotoxic and/or gastrointestinal side effects and/or = Provides a reduction of discontinuation of leflunomide treatment or a reduction of change to alternative active agents.
The application of the inventive pharmaceutical composition can thus also lead to an enhanced patient compliance.

5 Although the underlying mechanisms are presently not fully understood, the aforementioned ad-vantageous properties of the inventive subject matter are somewhat surprising to a person skilled in the art, as until now he assumed that the dose-response-relationship would be linear, in particu-lar for the autoimmune diseases, such as for rheumatoid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (Wegener's granulomatosis);
i.e. that a higher leflunomide dose correlates with a higher efficacy and, thus, it would be expected that a dose reduction from 20 mg to 14 to 17.5 mg, preferably 15 to 17.5 mg, most preferably 15 mg leflunomide would result in a reduction of efficacy.
Inventive pharmaceutical compositions, which comprise 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide in a single dose and are applied once daily, surprisingly show a similar or enhanced efficacy in particular in the prophylaxis and/or treatment of autoimmune dis-eases, preferably rheumatoid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (Wegener's granulomatosis) when compared to pharmaceutical compositions, which comprise 20 mg leflunomide in a single dose and are applied once daily.
Inventive pharmaceutical compositions, which comprise 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide in a single dose and which are applied once, twice, three times or four times daily dependent on the severity of the disease, show in addition surprisingly the same or an enhanced efficacy in comparison to the presently known recommendations for therapy in particular for the prophylaxis and/or treatment in the course of an organ transplantation, preferably kidney transplantation or liver transplantation; and/or of oncologic diseases, preferably melanomas, sarcomas, gliomas, prostate carcinomas, brain and/or CNS tumors; and/or of diseases correlated with the human immunodeficiency virus (HIV).
Cumulatively or alternatively to the aforementioned advantages the number of aforementioned side effects in particular hematotoxic and/oi' hepatotoxic and/or gastrointestinal side effects and/or the severity of side effects can be reduced by once daily application of the inventive pharmaceutical single dose composition of 14 to 17.5 mg, preferably 15 to 17.5 mg more preferably 15 mg leflunomide e.g. in the prophylaxis and/or treatment of autoimmune diseases, preferably rheuma-toid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (VVegen-

6 er's granulomatosis) in comparison to the single dose of 20 mg or 10 mg leflunomide respectively, whereby also the number of discontinuations of therapy or change of therapy can be reduced. The reduction of side effects and/or the severity of side effects is all the more amazing, as the prior art shows that an increase from 10 mg leflunomide single dose to 20 mg leflunomide single dose leads to a significant increase in (severe) side effects and thus also to an increase in discontinua-tions in therapy (Po6r, G õEfficacy and safety of leflunomide 10 mg versus 20 mg once daily in patients with active rheumatoid arthritis: multinational double-blind, randomized trial" Rheumatolo-gy 2004; 43: 744-749).
Surprisingly it can additionally be observed that an inventive pharmaceutical composition compris-ing 14 to 17.5 mg, preferably 15 to 17.5, more preferably 15 mg leflunomide single dose can lead to a faster adjustment of the steady state concentration in the prophylaxis and/or treatment of autoimmune diseases, preferably rheuratoid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or of granulomatosis with polyangiitis (Wegener's granulomatosis); and/or in the course of an organ transplantation, preferably kidney transplantation and/or liver transplantation; and/or oncologic diseases, preferably melanomas, sarcomas, gliomas, prostate carcinomas, brain and/or CNS
tumors; and/or diseases correlated with human immune deficiency virus (HIV) in comparison to the single dose comprising 20 mg and/or 10 mg leflunomide, so that an assessment of the therapeutic efficacy can be taken earlier and, thus, if necessary the respective dose can be adapted earlier, i.e.
dose reduction or increase can be conducted earlier and under doses and/or side effects can be prevented.
The provision of an inventive pharmaceutical composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide single dose can facilitate an (enhanced) personal-ized therapy in the prophylaxis and/or treatment of autoimmune diseases, preferably rheumatoid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or of granulomatosis with polyangiitis (Wegener's granulomatosis); and/or in the course of an organ transplantation, preferably kidney transplantation and/or liver transplantation; and/or oncologic diseases, preferably melanomas, sarcomas, gliomas, prostate carcinomas, brain and/or CNS tumors; and/or diseases correlated with human immune deficiency virus (HIV).
The application of an inventive pharmaceutical composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide single dose according to the first aspect of the invention can in addition be of advantage for special patient populations in comparison to the commercially available single dose of 20 mg leflunomide, in case an individual disadvantageous risk-benefit profile exists based on a side effect or toxicity of leflunomide.
Such a disadvantageous risk-benefit profile in the treatment of rheumatoid arthritis and in particular in a therapy regime with

7 one or more further autoimmune active agents can be based on polymorphisms of genes, which code for proteins, which in particular respectively intervene in metabolism reactions, preferably in the transport or the uptake from the gastro intestinal tract, the distribution in the body (plasma level) or the excretion of leflunomide and/or its metabolites, preferably its active metabolite teriflunomide.
According to a preferred inventive embodiment the inventive composition of the first inventive aspect is applied to patients, preferably humans, which show a polymorphism at enzyme cyto-chrome P 450 2C19 (CYP2C19) and are in particular poor to intermediate metabolizers (see Wiese et al.: Polymorphisms in cytochrome P450 2C19 enzyme and cessation of leflunomide in patients with rheumatoid arthritis. Arthritis Research & Therapy 2012 14:R163). Poor CYP2C19 metaboliz-ers are generally characterized in accordance with Wiese et al. in that they have 2 alleles with loss of function, whereas intermediate metabolizers have 1 allele with loss of function and one wild type allele. Patients, who are poor or intermediate CYP2C19 metabolizers have a disadvantageous risk-benefit profile concerning the toxicity of leflunomide and discontinue with a higher likelihood the leflunomide therapy, in particular for rheumatoid arthritis and in particular in a therapy regime with further autoimmune active agents. An application of an inventive composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide single dose according to the first aspect of the invention can, in particular when treating rheumatoid arthritis, lead to a reduction of toxicity and, thus, a reduction of discontinuations of therapy in particular when applying further autoimmune active agents, which is based on the reduction of the leflunomide content.
According to a further cumulatively or alternative preferred inventive embodiment the inventive composition of the first inventive aspect is applied to patients, preferably humans, which show a polymorphism at the ATP binding cassette sub-family G member 2 (ABCG2), also known as breast cancer resist protein (BCRP) (see Wiese et al.: Polymorphisms in cytochrome P450 2C19 enzyme and cessation of leflunomide in patients with rheumatoid arthritis. Arthritis Research & Therapy 2012 14:R163). Patients with such a respective ABCG2 genotype or BCRP genotype have a dis-advantageous risk-benefit-profile concerning the side effect diarrhea and discontinue with a higher likelihood the leflunomide therapy, in particular when treating rheumatoid arthritis and in particular in therapy with further active agents. An application of an inventive composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide single dose according to the first aspect of the invention can in particular when treating rheumatoid arthritis lead to a reduction of the incidence and/or severity of diarrhea and, thus, a reduction of discontinuation of therapy in particular when applying leflunomide and further autoimmune active agents.
The application of an inventive pharmaceutical composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide single dose according to the first aspect of the invention can furthermore be advantageous in comparison to the commercially available 20 mg

8 leflunomide single dose, in case the steady state level of leflunomide and/or its metabolites, pref-erably teriflunomide shall be reached earlier. A respective advantageous risk-benefit profile can preferably be advantageous in the treatment of rheumatoid arthritis and in particular in a therapy regime with one or more further autoimmune active agents and can, e.g., be based on the satura-tion of transporter proteins, which in particular respectively intervene in the uptake from the gastro-intestinal tract, the distribution in the body (plasma level) and/or in the excretion (e.g. biliary) of leflunomide and/or its metabolites, preferably teriflunomide.
In accordance with a further preferred inventive embodiment the inventive composition of the first inventive aspect is preferably applied in the treatment of rheumatoid arthritis and in particular in io therapy regimes with one or more further autoimmune active agents to patients, preferably hu-mans, who show a polymorphism of the ABCG2/BCRP transporter. For simplification reasons, it is essentially referred to BCRP, wherein this is used synonymously for ABCG2 in the context of the present invention. Such transporter proteins play an important role in the absorption, distribution, metabolism and/or excretion of leflunomi0e and/or its metabolites, preferably teriflunomide.
BCRP is one of the most important efflux transporters of the cells of the endo-and epithelium. In vitro studies show, that in particular leflunomide and its metabolite teriflunomide show as sub-strates a high binding affinity to the BCRP transporter [Bartlett, R.R., et al., Effects of leflunomide on immune responses and models of inflammation. Springer Semin Immunopathol, 1993. 14(4): p.
381-94]. Thus, the saturation of the BCRP transporter plays an important role i) in reaching thera-peutically active plasma levels of leflunomide and/or its metabolites, preferably teriflunomide, in particular in view of the induction phase as well as ii) in the time span for reaching the steady state levels of leflunomide and/or its metabolites, preferably teriflunomide, as leflunomide and/or its metabolites, preferably teriflunomide represent a substrate for the BCRP
transporter.
An application of an inventive composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide single dose according to the first aspect of the invention can lead preferably when treating rheumatoid arthritis and in particular in a therapy regime with one or more further autoimmune active agents to a = modified ratio of the plasma level of leflunomide in comparison to the plasma level of its me-tabolites, preferably teriflunomide and/or = modified time window within which the steady state level of leflunomide and/or its metabolites preferably teriflunomide is reached and thereby can lead to a more rapid onset of the therapeutic active effects and/or to a reduction of the toxicity of leflunomide and/or its metabolites, preferably teriflunomide.

9 Patients showing a c.421CA genotype and a polymorphism of the BCRP transporter protein gen-erally show in comparison to patients without such a polymorphism a higher plasma level of leflunomide metabolites, preferably teriflunomide, which is why this polymorphism leads to an inter-individual variability of the plasma level of leflunomide metabolites in the treatment of rheumatoid arthritis and in particular in the therapy regime with one or more further autoimmune active agents [Williams, J.W., et al., lmmunosuppressive effects of leflunomide in a cardiac allograft model.
Trans-plant Proc, 1993. 25(1 Pt 1): p. 745-6]. This factor is in particular then particularly relevant, in case one or more of the further autoimmune active agents themselves and/or its metabolites rep-resent in the combination therapy regime substrates for this BCRP transporter protein.
Thus, saturation and polymorphism of the BCRP transporter proteins play an important role, pref-erably when treating rheumatoid arthritis and in particular in a therapy with one or more further autoimmune active agents, as due to the saturation or the polymorphism of the BCRP transporter protein the plasma level and/or the time span, which is necessary to reach the steady state level of leflunomide and/or its metabolites, preferably teriflunomide, and/or the time span until reaching therapeutic efficacy and/or the toxicit'l level of leflunomide and/or its metabolites, preferably teriflunomide can be altered. This factor is in particular then particularly relevant, in case in the combination therapy regime one or more of the further autoimmune active agents themselves and/or its metabolites represent substrates for this PCRP transporter protein.
Thus, due to an altered ratio of the plasma level of leflunomide in comparison to its metabolites an application of an inventive composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide single dose according to the first aspect of the invention and in par-ticular when treating rheumatoid arthritis can lead to a reduction of toxicity and accordingly to a reduction of therapy discontinuations under leflunomide in particular in combination with one or more further autoimmune active agents.
According to the first inventive aspect the pharmaceutical composition comprising leflunomide is characterized in that the composition is arranged or arrangeable as single dose with 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide.
According to a preferred embodiment the inventive pharmaceutical compositions of the first in-ventive aspect are preferably characterized in that the compositions are in solid or liquid form and are suitable for oral application. Preferred solid inventive pharmaceutical compositions for oral application are selected from the group consisting of powders, pellets, globuli, granulates, tablets and/or capsules, in particular preferred are tablets and further particularly preferred are film coated tablets. Preferred liquid inventive pharmaceutical composition for oral application are selected from the group consisting of elixirs, solutions, suspensions, emulsions, mixtures, syrups, juices and/or drops. The inventive pharmaceutical compositions of the first inventive aspect can be inventively used in retarded or non-retarded formulation.
According to the present invention for the production of inventive compositions according to the first inventive aspect all of the pharmaceutically acceptable excipients generally known for the 5 respective pharmaceutical formulations can be used.
When producing tablet formulations as inventive pharmaceutical compositions according to first inventive aspect, pharmaceutical acceptable excipients can depending on the mode of forming tablets (direct tableting, dry or wet granulation) be selected from the group of respectively suitable excipients consisting of i) filler materials, preferably starch or starch derivatives, in particular pref--io erably potato, wheat and/or maize starch (derivatives), lactose, lactose monohydrate, mannitol, sorbitol, calcium carbonate, laevulose, glucose, cellulose, microcrystalline cellulose, hydroxypropylcellulose, and/or calcium phosphates, in particular preferred dicalcium phosphate, particularly preferred filler materials are selected from the group consisting of lactose, lactose monohydrate, dicalcium phosphate, and/or hydroypropylcellulose, preferably lactose monohydrate;
ii) disintegrants, preferably starch or starch derivatives, in particular potato, and/or maize starch (derivatives), pectin, alginates, alginic acid, microcrystalline cellulose, crosslinked sodium carboxymethylcellulose, crosslinked polyvinylpyrrolione, such as crospovidone (polyplastone) and/or croscaramellose; iii) binding agents or adhesives (in particular for wet granulation), prefera-bly sugar, starch, starch paste and/or starch derivatives, in particular preferred are potato, wheat and/or maize starch (derivatives), gelatin, cellulose derivatives, in particular preferred (low substi-tuted) hydroxpropylcellulose, hydroxyethylcellulose and/or hydroxyporpylmethylcellulose, micro-crystalline cellulose, Arabic gum, tragacanth, polyethylene glycol, and/or polyvinylpyrrolidone, and/or collidone, a particular preferred binding agent or adhesive is (low substituted) hydroxypropylcellulose; iv) dry binding agents (in particular for dry granulation), preferably micro-crystalline cellulose, lactose, lactose monohydrate, saccharose, mannitol, sorbitol, calcium car-bonate, polyvinylpyrrolidone, collidone, and/or polyethylene glycol v) glidants or drying agents, preferably highly dispersed silicium dioxide; and/or vi) lubricants, preferably talc, siliconized talc, stearic acid, palmitic acid, calcium, aluminum and/or magnesium stearate, calcium behenate (mix-ture of calcium salts of higher fatty acids, preferably behenic acid), starch, aerosol, (colloidal) silicium dioxide, polyethylene glyclol, hydroxypropylcellulose, stearyl, cetyl and/or myristyl alcohol, paraffin, hydrated fats and/or magnesium trisilicate, preferred lubricants are in particular selected from the group consisting of magnesium stearate and/or hydroxypropylcellulose, further preferred is magnesium stearate; and/or further suitable excipients, such as wetting agents, such as sodium lauryl sulfate or polysorbate, particularly preferred is sodium lauryl sulfate, and excipients for the production of film coatings, e.g., film binding agents, pigments, filler materials, softeners and/or stabilizers, preferably selected from th group consisting of polydextrose (E1200), hypromellose, triacetin, macrogol, iron oxides (e.g., red and/or yellow), polyvinyl alcohol, titan dioxide, talc, leci-thin (preferably derived from soy beans), xanthan gum and/or suitable polyacrylic acid derivatives and/or (soluble) collidones.
The inventive pharmaceutical compositions preferably as solid or liquid oral formulation preferably further comprise according to a cumulative or alternative preferred embodiment of the first in-ventive aspect a pH regulating agent, in particular preferred are monobasic organic or inorganic acids or organic or inorganic polyacids, wherein the inorganic acids are preferably selected from the group consisting of phosphoric acid, sulfuric acid, their acidic reacting salts, such as dihydrogen phosphates and hydrogen sulfates as well as their condensates, such as polyphosphoric acid, and/or inorganic buffer systems, which react in aqueous solution to a pH of 7 or lower; and wherein the organic acids are preferably selected from the group consisting of car-bonic acids, preferably tartaric acid, malic acid, fumaric acid, citric acid, malonic acid, maleic acid as well as methane, ethane and/or p-toluene sulfonic acid, further preferred tartaric acid, malic acid, fumaric acid, citric acid, malonic acid, maleic acid and/or organic buffer systems, which react in aqueous solution to a pH of 7 or lower (e.g., citric acid/citrate buffer and/or acetic acid/acetate buffer). Respective inventive pharmaceutical compositions are in particular preferred, as they can increase the shelf life of leflunomide, LEI. reduce the transformation of leflunomide to teriflunomide during storage, in the inventive pharmaceutical composition in comparison to pharmaceutical compositions comprising leflunomide without acidic addition.
zo According to an in particular preferred embodiment of the first inventive aspect the inventive com-position as tablet comprises in addition to the content of 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide the following excipients in suitable amounts, namely a) lactose monohydrate, low substituted hydroxypropylcellulose, a pH regulating agent, preferably selected from organic acids, particularly preferred tartaric acid or citric acid, sodium lauryl sulfate and mag-nesium stearate or b) maize starch, povidone (1201), crospovidon (1202), highly dispersed silicium dioxide, magnesium stearate (E470b) and lactose monohydrate. In case of film coated tablets the inventive pharmaceutical compositions comprise preferably suitable amounts of the following excipient, namely a) polyvinyl alcohol, titan dioxide, talc, lecithin, xanthan gum or b) talc (E553b), hypromellose (E464), titan dioxide (E171), macrogol 8000 and optionally iron (III) hydroxide oxides (E172).
According to a further cumulatively or alternatively preferred embodiment of the first inventive aspect the inventive pharmaceutical compositions are applied in the prophylaxis and/or treatment of autoimmune diseases, preferably selected from the group consisting of rheumatoid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis, granulomatosis with polyangiitis (Wegener's granulomatosis); in the course of organ transplantations, preferably selected of the group consist-ing of kidney transplantation and/or liver transplantation; of oncologic diseases, preferably selected of the group consisting of melanomas, sarcomas, prostate carcinomas, brain and/or CNS tumors;
and/or of diseases correlated with the human immune deficiency virus (HIV); in particular preferred in the prophylaxis and/or treatment of rheumatoid arthritis, psoriasis arthritis and/or arthritis as complication of cystic fibrosis. Generally, the inventive pharmaceutical compositions comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, in particular preferred 15 mg leflunomide is applied once daily or depending on the kind and/or severity of diseases (simultaneously or sequentially) twice, three or four time daily in the prophylaxis and/or treatment of the aforementioned diseases.
In the context of the present invention the term "simultaneous" application refers according to the lo first inventive aspect to a twice, three or four times daily application of an inventive pharmaceutical composition comprising leflunomide arranged as single dose of 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide and means that the inventive pharmaceutical compo-sition is taken by a patient simultaneously, i.e. in narrow time sequence, generally within a time period of up to 10 minutes, preferably up to 5 minutes, more preferably up to 1 minute.
In the context of the present invention the term "sequential" application refers according to the first inventive aspect to a twice, three or four times daily application of an inventive pharmaceutical composition comprising leflunomide arranged as single dose of 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunumide and means that the inventive pharmaceutical compo-sition is taken by a patient sequentially, i.e. in a temporary offset, generally with a time lag in par-ticular for a twice daily application in a sequence of approximately 12 hours, in particular for a three times daily application in a sequence of approximately 8 hours and/or in particular for a four times daily application in a sequence of approximately 6 hours.
Generally, the intake of the inventive pharmaceutical compositions according to the first inventive aspect can be conducted at all day and night times as well as prior, during and after meals, prefer-ably in a time sequence prior to the meal.
In the context of the prophylaxis and/or treatment of autoimmune diseases, preferably selected from a group consisting of rheumatoid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, and/or myasthenia gravis, and/or the prophylaxis and/or treatment of diseases correlated with the human immune deficiency virus (HIV) the inventive pharmaceutical composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, in particular preferred 15 mg leflunomide single dose is generally applied once or twice (simul-taneously or sequentially) daily, more preferably once daily to achieve the inventive advantages, in particular an optimized effect-side effect profile.
In the context of the prophylaxis and/or treatment of granulomatosis with polyangiitis (Wegener's granulomatosis) the inventive pharmaceutical composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide arranged as single dose is generally applied once, twice or three times (simultaneously or sequentially) daily, more preferred twice or three time (sim-ultaneously or sequentially) daily, to achieve the inventive advantages, in particular an optimized effect-side effect profile.
In the context of the prophylaxis and/or treatment of organ transplantations, preferably selected from the group consisting of kidney transplantation and/or liver transplantation; and/or oncologic diseases, preferably selected from the group consisting of melanoma, sarcoma, glioma, prostate carcinomas and/or brain and/or CNS tumors, the inventive pharmaceutical composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, in particular preferred 15 mg leflunomide single dose is generally applied once, twice, three or four times daily (simultaneously or sequentially), more preferred twice, three or four time daily (simultaneously or sequentially), in particular preferred three or four times daily (simultaneously or sequentially), to achieve the inventive advantages, in particular an optimized effect-side effect profile.
According to a further advantageous embodiment of the present invention the inventive pharma-ceutical composition according to the first inventive aspect is preferably applied to a patient of equal or more than 45 years of age, more preferably equal or more than 55 years of age, even more preferred equal or more than 65 years of age, even more preferred equal or more than 70 years of age, to achieve optimized results concerning the advantageous inventive effects in par-ticular concerning the equal or enhan, .-d efficacy, the reduced side effects and the reduction in therapy discontinuations or the change to other active agents.
The second inventive aspect relates to the use of leflunomide in the prophylaxis and/or treatment of autoimmune diseases, preferably of rheumatoid arthritis, psoriasis arthritis, arthritis as complica-tion of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (Wegener's granulomatosis); and/or in the prophylaxis and/or treatment in the course of an organ transplantation, preferably kidney transplantation and/or liver transplantation; and/or in the prophylaxis and/or treatment of oncologic diseases, preferably melanomas, sarcomas, gliomas, prostate carcinomas, brain and/or CNS tumors;
and/or in the prophylaxis and/or treatment of diseases correlated with the human immune deficiency virus (HIV), characterized in that leflunomide is arranged or arrangeable as single dose of an pharmaceutical composition with 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide and is applied once, twice, three or four times daily. The preferred embodiments according to the inven-tion, which have been discussed in the present application with respect to the first inventive aspect are to be applied alternatively or cumulatively also to the second inventive aspect.
According to the present invention the term "leflunomide is arranged as single dose of an pharma-ceutical composition with 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide" means that the commercially available inventive pharmaceutical composition is al-ready present in such a separated form, that one or more single dose formulations with 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide are comprised.
According to the present invention the term "leflunomide is arrangeable as single dose of an phar-maceutical composition with 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide" means that the commercially available inventive pharmaceutical composition does not already comprise the single dose with 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide in separated form, however can be separated in particular by the patients in such a way, that an inventive pharmaceutical single dose formulation with 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide can be arranged. In case the inventive pharmaceuti-cal compositions for oral consumption is present in liquid form, such a single dose formulation can, e.g., be arranged by measuring the liquid by means of a measuring cup or measuring spoon or by means of counting of drops or any other suitable method. In case the inventive pharmaceutical compositions for oral consumption is present in solid form, such a single dose formulation can, e.g., be arranged by breaking a (film coated) tablet, preferably a (film coated) tablet with a score line, counting of pellets, globuli or granulates or any other suitable method.
The present invention will be described in the following by use of exemplary embodiments, which shall rather be regarded as examples and shall not limit the scope of protection of the present IP
right.

Examples: , A: Production of inventive pharmaceutical compositions The production described in the following discloses one possibility of producing the inventive oral pharmaceutical compositions, in particular as inventive film coated tablets comprising leflunomide 5 listed in the following table:
Ingredients (in mg/ tablet) Fl F2 F3 F4 F5 Tablet Core Leflunomide (Ph. Eur.) 14.000 15.000 16.000 17.000 17.500 Lactose Monohydrate (Ph. Eur.) 50.000 - 45.000 215.000 106.500 Microcrystalline Cellulose (Ph. Eur.) 45.000 25.000 85.000 55.000 35.000 Dicalcium phosphate, anhydrous - 80.000 - - 10.500 (DI-CAFOS, Ph. Eur.) Copovidone (Ph. Eur.) - - 10.000 -9.000 Maize starch (Ph. Eur.) 15.000 - -Crospovidone (Ph. Eur.) - 12.000 5.500 3.000 -Croscarmellose (Ph. Eur.) - 15.000 5.500 -8.500 Tartaric Acid (Ph. Eur.) - 3.350 - - .
Citric acid, anhydrous (Ph. Eur.) 3.500 . - - --Sodium lauryl sulfate (Ph. Eur.) - 0.650 - - -Polysorbate 80 (Ph. Eur.) 0.500 - - -Highly dispersed silicon dioxide (Ph.
- - 3.000 - 5.000 Eur.) Magnesium stearate (Ph. Eur.) 2.000 3.000 - 3.000 2.000 ' Calcium behenate (Ph. Eur.) - 2.500 -Purified water (Ph. Eur.) q.s. q.s. - - -Weight Tablet Core (mg) 130.000 154.000 172.500 293.000 194.000 Filmcoating Polydextrose (E1200) 0.960 0960 - -Hypromellose (Ph. Eur.) 1.461 1.461 5.200 0.250 silia - - -2.650 Triacetin (Ph. Eur.) 0.240 0.240 - -Macrogol (Ph. Eur.) 0.080 0.080 0.600 0.150 Iron oxide yellow (E172) 0.009- N/A - -Iron oxide red (E172) - 0.009 - -Polysorbate 80 (Ph. Eur.) - - 0.090 -Titanium dioxide (Ph. Eur.) 1.250 1.250 2.110 -Ethanol 96 % (Ph. Eur.) - - -q.s.
Purified water (Ph. Eur.) q.s. q.s. q.s.
q.s.
Weight Film Coated Tablet (mg) 134.000 158.000 N/A
301.000 197.050 __________________________ , _________________________________________ Ingredients (in mg/ tablet) F6 F7 F8 F9 F10 Tablet Core Leflunomide (Ph. Eur.) 14.000 15.000 16.000 17.000 17.500 Lactose Monohydrate (Ph. Eur.) 80.000 120.000 55.000 - 36.500 Microcrystalline Cellulose (Ph. Eur.) - - 25.000 180.00 63.000 Mannitol (Ph. Eur. - - - 155.00 -Hypromellose (Ph. Eur.) - - - - 10.500 Hydroxypropylcellulose, low substi-15.000 7.500 - - -tuted (NF) Maize starch (Ph. Eur.) 15.000 - 9.000 --Sodium carboxyl nnethylstarch (Ph.
- - 18.500 --Eur.) Croscaramellose (Ph. Eur.) 10.000- - - 12.000 Tartaric acid (Ph. Eur.) 5.000 4.500 - - -Citric acid, anhydrous (Ph. Eur.) - - 2.300 2.000 -Sodium lauryl sulfate (Ph. Eur.) 1.500 0.750 - - -Polysorbate 80 (Ph. Eur.) 2.000 - 0.750 --Highly dispersed silicon dioxide - -1.200 -(Ph. Eur.) Magnesium stearate (Ph. Eur.) - 2.250 0.500 - 1.500 Calcium behenate (Ph. Eur.) 2.500 - 1.200 --Purified water (Ph. Eur.) q.s. q.s. q.s. q.s. -Sodium stearyl fumarate (Ph. Eur.) - - 9.000 q.s.
Weight Tablet Core (mg) 145.000 150.000 129.450 363,000 141.000 Film Coating Polydextrose (E1200) - - -Hypromellose (Ph. Eur.) - - 2.500 -Triacetin (Ph. Eur.) - - - -Macrogol (Ph. Eur.) 0.200- 0.810 0.215 Iron oxide yellow (E172) - - 0.040 0.035 Iron oxide red (E172) - - 0.050 N/A -Polyvinyl alcohol (Ph. Eur.) 1.500 2.048 - 1.460 Titanium dioxide (Ph. Eur.) 2.300 1.440 1.200 1.540 Talc (Ph. Eur.) - 0.900 0.450 -Lecithin (from soy beans) (NF) - 0.090 - -Xanthan gum (Ph. Eur.) - 0.022 - -Purified water (Ph. Eur.) q.s. q.s. q.s. q.s.
Weight Film Coated Tablet (mg) 149.000 154.500 134.500 N/A
144.250 Ingredients (in mg/ tablet) F11 F12 F13 F14 Tablet Core Leflunomide (Ph. Eur.) 14.000 15.000 15.000 17.500 Lactose Monohydrate (Ph. Eur.) 80.000 105.000 118.000 100.000 Microcrystalline Cellulose (Ph. Eur.) - - - 51.500 Hydroxypropylcellulose, low substi-- - 8.000 -tuted (NF) Maize starch (Ph. Eur.) - 18.000 6.300 25.000 Povidone (Ph. Eur.) 6.000 4.750 - 4.000 Crospovidone (Ph. Eur.) - 12.000 - 6.000 Sodium carboxyl methylstarch (Ph.
12.000 - 5.000 -Eur.) Tartaric acid (Ph. Eur.) - - 4.000 -Citric acid, anhydrous (Ph. Eur.) - - - -Sodium lauryl sulfate (Ph. Eur.) - - - 3.500 Polysorbate 80 (Ph. Eur.) - - - -Highly dispersed silicon dioxide 1.500 2.000 - 4.500 (Ph. Eur.) Talc (Ph. Eur.) - 1.000 - -Magnesium stearate (Ph. Eur.) - 1.250 2.200 3.500 Calcium behenate (Ph. Eur.) 1.500 - - -Purified water Ph. Eur.) q.s. q.s. q.s. q.s.
Weight Tablet Core (mg) 115.000 159.000 158.500 215.500 Film Coating Polydextrose (E1200) - - - -Hypromellose (Ph. Eur.) - 2.500 2.200 -Triacetin (Ph. Eur.) - - - -Macrogol (Ph. Eur.) - 0.810 0.700 -Iron oxide yellow (E172) - 0.040 - 0.050 Iron oxide red (E172) - - - 0.050 Polyvinyl alcohol (Ph. Eur.) 2.200 - - 2.800 Titanium dioxide (Ph. Eur.) 1.320 1.200 1.650 1.400 Talc (Ph. Eur.) 0.300 0.450 0.450 0.400 Lecithin (from soybeans) (NF) 0.090 - - 0.100 Xanthan gum (Ph. Eur.) 0.090 - - 0.200 Purified water (Ph. Eur.) q.s. q.s. q.s. q.s.
Weight Film Coated Tablet (mg) 119.00 164.000 163.500 220.500 According to the invention the individual features of the exemplary embodiments of the inventive pharmaceutical composition as disclosed hereinbefore can respectively be separately combined with features of the further exemplary embodiments or with the general description of the invention.
The advantages of the application of the inventive pharmaceutical composition comprising 14 to 17.5 mg, preferably 15 to 17.5 mg, more preferably 15 mg leflunomide single dose according to the first inventive aspect as well as its inventive use according to the second inventive aspect, prefera-bly in the treatment of rheumatoid arthritis and in particular in therapy regimes with one or more further autoimmune active agents can be demonstrated in comparison to commercially available single doses of 10 mg and 20 mg leflunomide in animal models. Such animal models allow exami-nations of autoimmune diseases, in particular of chronic diseases such as rheumatoid arthritis (RA). Respective animal models comprise in particular, but are not limited to, the collagen-induced arthritis (CIA) animal model, the adjuvant-induced-arthritis (AIA) animal model and the experi-mental allergic encephalomyelitis (EAE) animal model as well as animal models concerning graft-versus-host disease or reactions concerning graft rejection. In addition animal models suitable for the examination of other autoimmune diseases, such as, e.g., psoriasis arthritis, arthritis as com-plication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (Wegener's granulomatosis) can be used. Alterna-tively, animals models can be used, which allow the examination of organ transplantations, prefer-ably of kidney transplantations and/or liver transplantations, and/or the examination of oncologic diseases, preferably melanomas, sarcomas, gliomas, prostate carcinomas, brain and/or CNS
tumors; and/or the examination of diseases correlated with the human immune deficiency virus (HIV).
Safety and toxicity studies with leflunomide in naive, untreated animals (dogs, rats) showed that side effects occurred, which also are present in patients under leflunomide treatment. Such are, e.g., hematotoxic side effects, gastrointestinal side effects, creatinine kinase (CK) increase, body weight reduction and/or hepatotoxic side effects.
Animal models for chronic autoimmune diseases, in particular animal models for rheumatoid arthri-tis showed similar side effects such as altered metabolization and/or pharmacokinetic and/or pharmacodynamics of the active agents in the treatment of patients with autoimmune diseases, such as rheumatoid arthritis, psoriasis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (Wegener's disease).
Therefore, the animal models discussed hereinbefore are suitable to study the risk-benefit ratio of the respective active agents, preferably leflunomide and/or its metabolites, such as preferably teriflunomide. Safety studies treating naïve, untreated animals with leflunomide have been pub-lished up to now. Aforementioned animal models can represent patient situations and allow draw-ing conclusions concerning the risk-benefit-ratio without impairing the safety of humans.
The inventors of the present invention have surprisingly discovered that due to the content of 14 to 17.5 mg, preferably 15 to 17.5 mg, in particular preferably 15 mg leflunomide per pharmaceutical formulation of an inventive pharmaceut:cal composition according first inventive aspect as well as its use according to the second inventive aspect show in one or more animal models of these diseases, e.g. but not limited to, the collagen induced arthritis (CIA) RA
animal model = a comparable or enhanced efficacy in the prophylaxis and/or treatment of autoimmune dis-eases, preferably of rheumatoid arthritis, psoriasis arthritis, arthritis as complication of cyst-ic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis and/or granulomatosis with polyangiitis (Wegener's granulomatosis); and/or in the prophylaxis and/or treatment in the course of an organ transplantation, preferably kidney transplantation and/or liver transplantation; and/or in the prophylaxis and/or treatment of oncologic diseas-es, preferably melanomas, sarcomas, gliomas, prostate carcinomas, brain and/or CNS tu-mors; and/or in the prophylaxis and/or treatment of diseases correlated with human im-mune deficiency virus (HIV) in comparison to the standard therapy with 20 mg and/or 10 mg leflunomide arranged as single dose and/or = a reduction of one or more of the side effects, preferably one or more hematotoxic and/or hepatotoxic and/or gastro intestinal side effects and, thus, an enhanced safety in compari-son to the standard therapy with 20 mg and/or 10 mg leflunomide arranged as single dose and/or = a reduction of therapy discontinuations with leflunomide or changes to other therapeutic ac-tive agents in comparison to the standard therapy with 20 mg and/or 10 mg leflunomide ar-ranged as single dose.
The inventive film coated tablets can be produced in accordance with common production pro-cesses (direct tableting, dry or wet granulation). According to a preferred embodiment all or a part of the constituents of the tablet core can be grouted in the wet granulation process to a tablet core.
In a suitable equipment the tablet cores can subsequently be coated with the constituents of the film coating and can optionally subsequently be dried.
In the following a choice of assessment possibilities are described, to proof the efficacy and safety of leflunomide and/or its active metabolite (A77 1726) teriflunomide according to the inventive pharmaceutical composition in particular rheumatoid arthritis.

B: Non clinical studies for the analysis of efficacy of leflunomide Rheumatoid arthritis is a chronic inflammatory disease, which is characterized by a symmetrical polyarticular synovitis with defects in cartilage and bone. Generally, many rat models imitating RA
can be used, including the collagen type II induced arthritis (CIA), the pristane (2, 6, 10, 14-5 tetramethyl pentadecane) induced arthritis (PIA), adjuvant induced arthritis, oil induced arthritis, avridine induced arthritis, collagen type XI induced arthritis and the oligomer matrix protein induced arthritis of the cartilage.
CIA is one of the most used models of the RA and can easily be induced in sensitive rats and mice by intradermal injection of autologous or heterologous collagen type II (CII) with incomplete

10 Freund's adjuvant. Another commonly used model for RA is PIA, which is induced by a single injection of pristane. The course of disease is followed over 28 days in the PIA model, wherein the symptoms generally begin 14 days post pristane injection. The peak of the disease systems is generally reached 21 days post disease induction. In contrast thereto the disease symptoms gen-erally start in the CIA 18 days post CII application, wherein the peak of the disease symptoms is 15 reached approximately 28 days post disease induction.
Materials and methods All human and animal materials of the models described hereinbefore underlie the ethical require-ments or will be gained for the respective use after permission of the ethical commission.
Animals In accordance with the models described hereinbefore Dark Argouti (DA) rats of Taconic Europa (DK) can be used as animals, wherein the animals are preferably kept in a specific pathogen free condition and in a 12 h light-dark cycle.
Preparation of CII
CII will generally be produced by the pepsin digestion process from the cartilage of the xiphoid process of the DA rat (s. LU et al: Die Immunisierung von Ratten mit homologen Typ XI Kollagen fLihrt zu chronischer und rezidivierender Arthritis, mit unterschiedlicher Genetik und Gelenkpathologie, als mit homologen Typ-II-Kollagen induzierte Arthritis. J
Autoimmun 2002; 18:
199-211).
Generally the preparation will be extracted in buffer (1.0 M NaCl/ 50 mM Tris / pH 7.5) after cleav-age by pepsin. Subsequently, 0.9 M NaCI can be used to precipitate C II. The collagen will gener-ally be lyophilized, weighted and then dissolved and kept in 0.1 M acetic acid until use. The purity of the CII can be proved by Coonnassie Blue Staining after SDS PAGE analysis.
Induction of arthritis In the CIA model, rats are generally immunized, i.e. the disease is induced by a single intradermal injection of 150 pl emulsion comprising 10 pg C 11 dissolved in 75 pL 0.1 mo1/1 acetic acid and 75 pL incomplete Freund's adjuvant (Sigma-Aldrich, USA).
In the PIA model, 150 pL pristane (Arcos Organics, Belgium) is generally injected at the tail bases of the rats, to induce the disease pattern.
io Rats of the control group generally receive a subcutaneous injection of 150 pL
phosphate buffered sodium chloride solution.
Clinical evaluation of the arthritis Generally a macroscopic evaluation system is used, to monitor the development of the arthritis in all 4 extremities. Within this evaluation system generally (i) one point is awarded for each swollen Or red metacarpophalangeal/metatarsophalangeal joint, (ii) One point is awarded for swollen or red interphalangeal (IP) joints of each single toe, and (iii) Five points at maximum are awarded for each wrist or ankle (1, slight red; 2, swollen joint at inside, outside or central; 3, moderate swelling / redness; 4, strong, but incom-plete joint swelling; 5, complete joint swelling / redness).
The maximum point number for each paw is 15. Rats are generally examined 3 times per week post induction of the disease.
Pathological evaluation of the arthritis The left hindpaw of the rats are generally removed and fixed and then discalcified for 4 weeks in 12.5 % EDTA (ethylenediaminetetraacetic acid) solution. During this time the solution is generally changed every second day.
The discalcified samples are subsequently embedded in paraffin and sliced in 6 pm big tissue segments, which are then stained with hematoxylin and eosin (H & E).

A pathologic evaluation system is generally used to evaluate the severity of the arthritis. Synovitis is thereby evaluated according to the number of synovial cell layer areas, areas of the pannus bearing area, infiltration of synovial inflammatory cells and novel vascularization. For each index 0 to 3 points are awarded. Destruction of the joint by degradation of the cartilage, bone erosion, synarthrophysis and joint structure are rated with 0 to 4 points. Damage repair by building of new cartilage and bone is rated with 0 to 3 points.
Pathological evaluation of further organs Gastrointestinal tract, liver, inguinal lymph nodes and popliteal lymph nodes can be sampled from the rat and can be weighted after sampling. The pathological modifications in the sampled tissues of both, the control and the PIA animals can be identified and evaluated after fixation with para-formaldehyde and H & E staining.
Measuring the A77 1726 level The measuring of the A77 1726 level can be conducted as described in Rakhila et al. (2011) J.
Pharm. Biomed. Anal. 5; 55 (2), S. 325-31.
Interaction of leflunomide and/or its metabolite with cytochrome P450 enzymes The cytochrome P450 level can be measured by commercially available ELISA kits specific for rats.
Measuring the active agent transporter protein level The amount on mRNA of the ATP binding cassette sub family G member 2 (ABCG2), also known as breast cancer resistance protein (BCRP), can be measured as described in Takara et al. (2003) Drug Metab Dispos 31:1235-1239 und Takara et al. (2007) EXCLI Journal 6:138-144 Takara et al.
(2003) Drug Metab Dispos 31:1235-1239 und Takara et al. (2007) EXCLI Journal 6:138-144.
Measuring the DHODH level / activity The DHODH level can be measured by use of a commercially available ELISA kit specific for rats.

Measuring the kinase level / activity Identifying kinases, which have been activated by leflunomide and/or A77 1726 A kinase scan can be conducted as described in Fabian, M.A. et al. (A small molecule-kinase interaction map for clinical kinase inhibitors. Nat. Biotechnol. 23, 329-336;
2005) and Wodicka, L.M. et al. (Activation state-dependent binding of small molecule kinase inhibitors: structural in-sights from biochemistry. Chem. Biol. 17, 1241-1249; 2010). Kinases fused to T7 phages are produced in an E. coli host, which is generally produced from the strain BL21.
E. coli is accordingly generally cultivated up to the log phase and then infected with T7phages and incubated under agitation at 32 C until lysis. The lysates are centrifuged and filtrated to remove the cell fractions.
The remaining kinases are expressed in fusion with NF KB in HEK-293 cells. The constructs are subsequently marked with DNA for a PCR analysis.
Streptavidin coated magnetic pearls can be treated for 30 minutes at room temperature with small molecule ligands comprising biotin, to generate affinity resins for the kinase assay. Here, the pearls comprising ligands are generally blocked with excess biotin and are washed with blocking buffer (Seablock (Pierce), 1% BSA, 0,05% Tween 20, 1 mM DTT), to remove unbound ligands and to avoid nonspecific bindings.
Status / activity of phosphorylation In use of standard techniques freshly isolated tissue is generally washed and homogenized in ice cold homogenization buffer.
Lysed tissue is generally incubated with commercially available primary antibodies, which specifi-cally bind the respective kinases. These are activated by leflunomide and/or teriflunomide. The presence of phosphorylated and complete kinases are measured by use of cell based ELISA in accordance to the producer instructions.
Localizing leflunomide and/or A77 1726 activated kinases in situ The phosphorylation of kinases in situ ,:;an be examined in use of standard immunohistochemical (INC) techniques and in use of commercially available phosphor antibodies (BD
Biosciences).
Monoclonal antibodies for kinases, which are activated by leflunomide and/or teriflunomide (kinase scan), are generally used here, to localize the activity on cellular base.

C: In vitro studies with leflunomide and its metabolites Example 1: Kinome Scan of leflunomide an A77 1726 Binding reactions can be conducted by combining kinases and ligand affinity pearls for leflunomide and its active metabolite A77 1726 (teriflunomide). The kinase concentration in the eluates are generally measured with quantitative PCR.
Example 2: proliferation assay in use of mitogen induced peripheral blood cells (human and rat) Heparinized full blood, buffy coat or leukocyte filter fraction of healthy volunteers (human) are used for the isolation of human peripheral blood mononuclear cells. Heparinized full blood or spleen are removed of female or male DA rats (Taconic Europa, DK).
Human peripheral blood mononuclear cells (PBMCs) can be isolated from heparinized venous blood of healthy volunteers by centrifugation on a Ficoll Paque gradient (Amersham Biosciences AB). Here the white cells are sampled from the Ficoll plasma intermediate phase and are washed twice with fresh phosphate buffered sodium chloride solution (PBS).
Erythrocytes remaining in the cellular pellet are lysed with distilled H20 and the reaction is stopped by the addition of the same volume of 1.8 % NaCI, to restore the isotonicity.
Isolated cells are resuspended in sterile complete media R10 (RPM! 1640 with HEPES buffer 25 mM, Glutamax I, 10% fetal bovine sera and penicillin streptomycin (5 ml penicillin 5000IU/mL +
streptomycin 5000pg/mL). The cells are generally plated on 96 well plates with a density of 1 x 106 cells / ml and are incubated for an hour with A771726 (concentration 1 x 10-9¨ 1 x 10-4 M).
Subsequently the cells are generally stimulated either with anti CD3 (10 ng/ml), PHA (1pg/m1) or with ConA (10 pg/ml) and are cultivated at 27 C, 5% CO2, 90% relative humidity (RH) for 24, 48 or 72 hours.
The cells are generally pulsed one hour prior to the harvest with BrdU (30 pM) (for measurement of the proliferative capacity). At time of harvest the supernatants are sampled and stored at ¨ 80 C
for further analysis.
The cells are generally stained with surface marking agents for identification of the relevant popula-tions of leukocytes and T cells with anti CD3, anti CD4, anti CD8, B cells with anti CD19 and mon-ocytes as well as neutrophils. The cells are generally subsequently fixed, permeabilized and stained for presence of nuclear BrdU. Subsequently, the cells are generally analyzed in use of FACS Calibur and BrdU positive cells and are recorded within the respective leucocyte populations as indication of proliferative activity.
The cells, which have been incubated with teriflunomide and then stimulated with mitogen, are generally analyzed on DHODH and selected tyrosine kinase activity, to study the concentration-5 efficacy relationship.
White blood cells of the rat are isolated from the spleen of the control and the pristane treated rats and are gently homogenized to generate a single cell suspension. The spleen cells are generally obtained in PIA rats at peak of the inflammation and are set on Ficoll Paque.
Subsequently, the spleen cells can be prepared in the same manner as described for human PBMC
only in use of rat 10 specific antibody for leukocyte subpopulation. The cells, which have been incubated with teriflunomide and the stimulated as described with mitogen can also be analyzed on DHODH and selected tyrosine kinase activity, to study the concentration efficacy relationship.
D: In vivo studies with leflunomide 15 Example 3: Influence of the Inflammation on the dose exposure relationship leflunomide Male DA rats at the age of 8 to 12 weeks are randomly divided in 2 groups: One PIA group and one control group. Each group is further divided into dose groups (n = 3-4) and receive a single dose formulation of leflunomide (e.g., 5 different doses). To determine pharmacokinetic profiles, blood samples are generally taken at 7 to 10 time points after application for measuring the plasma 20 concentration of an active leflunomide metabolite, (A77 1726) teriflunomide. Animals showing an inflammation, the dosing and blood sample taking follows at the peak of the inflammation.
Preferably each group contains a subgroup, which is not treated.
These animals are generally used to confirm the influence of the inflammation on systems under involvement of the liver active agent clearance, e.g., expression / activity of the cytochrome P450 25 (CYP) and of influx / efflux transporters, and are sacrificed at the peak of the inflammation. The livers are taken from the animals. One part of the liver is prepared and used for immunohistochemical analysis, to study the expression level of the transporter proteins. The re-maining liver tissue is homogenized and plasmosomes are isolated by use of differential centrifu-gation.
The complete CYP expression in plasmosomes can be measured (carbon monoxide differential spectra) and activities of the CYP isoforms can be determined by quantifying the metabolization of testosterone.

Example 4: Effect of inflammation on the efficacy and toxicity dose relationship of leflunomide Male DA rats at the age of 8 to 12 weeks are randomly divided in 2 groups: One PIA group and one control group.
Each group is further divided in treatment groups (preferably n = 10), which receive leflunomide once daily (e.g., 4 different doses or vehicles) over 3-4 weeks.
To establish the toxic kinetic, blood samples are generally taken at 7 to 10 time points on day 1 and on the day of the peak of the disease for animals with inflammation for measuring the plasma concentrations of active leflunomide m'Otabolite, (A77 1726) teriflunomide.
Blood samples, which are taken at the peak of the disease, can further be analyzed for clinical chemistry, hematology and plasma marker, e.g., cytokines. During the study disease scores and general health parameter (weight, behavior) are measured. After taking the second blood sample, the animals are generally sacrificed and the efficacy, toxicity parameter as well as histopathology of the relevant tissue (joints, liver, GI tract, lymphatic organs) is obtained. The pathological evaluation of leflunomide treated PIA and healthy control rats enables the determination of the dose toxicity and the dose efficacy relationship for leflunomide.
E: Ex vivo studies with leflunomide Example 5: Ex vivo measurement of DHODH activity Tissue samples, which have been taken in example 4 for the ex vivo measurement, can be used to establish the DHODH activity ex vivo.
Example 6: Ex vivo measurement of the kinase activity Tissue samples, which have been taken in example 4 for the ex vivo measurement, can be used to establish the kinase activity ex vivo.
The status / activity of phosphorylation selected kinases, which have been identified in the kinase scan (example 1), can be quantified in homogenized tissue by use of an ELISA
based assay and its cellular base in the tissue can be localized by use of immunohistochemistry.

Claims (8)

Claims:

1. Pharmaceutical composition comprising leflunomide, characterized in that the composition is arranged as single dose with 14 to 17.5 mg leflunomide.

2. Pharmaceutical composition according to claim 1, characterized in that the composition is arranged as a single dose with 15 mg leflunomide.

3. Pharmaceutical composition according to claim 1 or 2, characterized in that the composition is solid or liquid and is suitable for oral application.

4. Pharmaceutical composition according to claim 3, characterized in that the solid composition is selected from the group consisting of powders, pellets, globuli, granulates, tablets and/or capsules or that the liquid composition is selected from the group consisting of elixirs, solu-tions, suspensions, emulsions, mixtures, syrups, juices and/or drops.

5. Pharmaceutical composition according to any one of claims 1 to 4 in the prophylaxis and/or treatment of autoimmune diseases, in the course of an organ transplantation;
of oncologic diseases; and/or of diseases correlated with the human immune deficiency virus (HIV).

6. Pharmaceutical composition according to claim 5, whereby the single dose with 14 to 17.5 mg leflunomide is applied once, twice, three or four times daily simultaneously or sequential-ly.

7. Use of leflunomide in the prophylaxis or treatment of an autoimmune disease; in the course of an organ transplantation; of oncologic diseases; and/or of diseases correlated with the human immune deficiency virus (HIV), characterized in that leflunomide is arranged or arrangeable as a single dose of a pharmaceutical composition with 14 to 17.5 mg leflunomide and is applied once, twice, three or four times daily.

8. Pharmaceutical composition according to claim 5 or use according to claim 5, characterized in that the a. Autoimmune disease is selected from the group consisting of rheumatoid arthritis, psoria-sis arthritis, arthritis as complication of cystic fibrosis, lupus nephritis, systemic lupus erythematosus, uveitis, myasthenia gravis, and/or granulomatosis with polyangiitis (We-gener's granulomatosis); and/or b. Organ transplantation is selected from kidney transplantation and/or liver transplantation and/or c. Oncologic diseases are selected from the group consisting of melanoma, sarcoma, glioma, prostate carcinoma, brain and/or CNS tumors.