HK1174567A - Combination preparations comprising exosomes and corticosteroid - Google Patents

Description

Combination preparation with exosomes and corticosteroids

Technical Field

The present invention relates to pharmaceutical compositions for use in combination therapy with corticosteroids and exosomes. Diseases such as osteoarthritis, arthritis and/or degenerative spinal diseases can be treated with this combination therapy, wherein preferably the treatment is performed locally.

Technical Field

In germany, osteoarthritis (Arthrose) describes "joint wear" to a degree beyond age. It is accompanied by cartilage defects in the affected joints, thereby producing pain and functional decline. Causes caused by overload, congenital or trauma, such as joint dislocation, or bone deformation due to bone diseases such as osteoporosis are considered to be causes. It can also lead to other diseases such as bone inflammation or exudation accompanied by excessive load.

In principle, all joints may be affected by osteoarthritis changes. In germany, the disease is usually localized to the knee joint. Osteoarthritis is the most frequent counseling cause in common hospital affairs. Approximately 10% of the population in western countries suffer from osteoarthritis. If this is accounted for by small vertebra osteoarthritis and degenerative disc disease, the affected population is even about 15% -20%. The risk of developing osteoarthritis increases with increasing age. Approximately two-thirds of people over 65 years of age suffer from the disease, however not all affected people suffer from their symptoms.

For the treatment of osteoarthritis, several treatment modalities are known. Conservative (e.g., pharmaceutical) therapy and surgical intervention are included up to the replacement of an intact joint with a prosthesis. In order to avoid such massive and irreversible interventions, in principle an effective drug treatment is more desirable in order to delay the point of complete joint replacement as far as possible.

However, many drug treatments also have disadvantages. On the one hand, this is due to side effects of the drug itself, but is also partly only provided to a limited extent due to its effect.

A common drug substance used in the treatment of osteoarthritis and rheumatoid arthritis is cortisone (also applied topically) and related corticosteroids. This is administered systemically in the case of rheumatoid arthritis and locally as an injection in the affected joints in the case of osteoarthritis. Here, however, the positive effect of corticosteroid administration is significantly reduced after only one week, both in the case of osteoarthritis and in the case of rheumatoid arthritis. This was confirmed clinically from randomized studies and clinical experience. In the case of rheumatoid arthritis, attempts are made to bring the systemic effect level to a high level by continuous administration of cortisone, which is, however, very problematic due to the intensification of side effects and the reduction of the therapeutic effect in the case of prolonged administration.

Exosomes are small, lipid membrane-enclosed vesicles found, for example, in the extracellular space of the human body. They are formed and secreted from cells by separation from the cytoplasmic membrane. Typically, these exosomes also contain proteins, which they receive from their source cells. Methods for obtaining and administering exosomes are described, for example, in patent application WO 2006/007529a 2. In the case of in vitro induction of prophylactically or therapeutically effective proteins, such as IL-1Ra, exosomes are formed by culturing blood samples in a suitable container, such as a syringe. Thus, for example, Orthokin contains exosomes. The formation of exosomes may be increased by adding additives that promote the formation of exosomes. The concentration of exosomes may be increased, for example, by centrifugation at high centrifugal acceleration. The use of exosomes in the treatment of rheumatoid arthritis is known per se.

Another drug which can be used for the treatment of osteoarthritis is the endogenous protein IL-1Ra or its isoforms (isoform) or fragments with similar activity. Like interleukin-1 (IL-1), interleukin-1 receptor antagonist (IL-1 Ra) binds to the same receptor on the cell surface, but does not trigger the signaling cascade there that is normally caused by IL-1 binding. By binding to the IL-1-receptor, IL-1Ra blocks IL-1 binding and thus prevents signal transduction and thus the pro-inflammatory effects of IL-1 on target cells.

It is known in the prior art to treat patients with endogenous sera in which IL-1Ra is enriched. IL-1Ra as used herein is also known as Orthokin. In contrast, a recombinant IL-1Ra fragment, Anakinra (Anakinra), did not show any significant effect in treating osteoarthritis as compared to placebo treatment. Anakinra is a subtype of human interleukin-1 receptor antagonist shortened to amino acids 26-177 and aminoacylated at the terminal L-methionine and has a sequence length of 153 amino acids. This preparation is carried out, for example, by recombinant methods using E.coli (Escherichia coli) strains.

In view of the prior art, the task has arisen to provide a medication for osteoarthritis which has better efficacy and, in particular, good long-term efficacy. Furthermore, preferably, the treatment should have good efficacy, preferably long-term efficacy, in the case of arthritis, in particular rheumatoid arthritis.

Summary of The Invention

Surprisingly, it has now been determined that in the case of osteoarthritis, arthritis and degenerative spinal diseases, the efficacy, in particular the long-term efficacy and the rapidity of onset of action, of corticosteroids, such as cortisone, can be significantly or synergistically improved by additional administration of exosomes. This is particularly indicated when the therapeutic agent is applied topically directly to the joint to be treated. One surprising recognition is that, for example, the use of corticosteroids (e.g., corticosteroid with Orthokin) in combination with exosomes (e.g., obtained from blood samples in Orthokin syringes) appears to be superior to treatments that do not contain active-substance exosomes in terms of rapidity of onset in the case of osteoarthritis. If exosomes are added as additional active substances to an already effective osteoarthritis treatment (e.g. a corticosteroid together with anakinra), a fast-acting effect can also be observed.

Even in the treatment of rheumatoid arthritis, the combination of corticosteroids with exosomes shows a remarkably good efficacy, not only in terms of rapidity of onset of action, but also in terms of long-term efficacy. The particularly advantageous effects shown by the present invention depend on a surprisingly high potency (depending on different pathophysiological starting points), and also on their particularly advantageous side-effect profile for the patient. This advantageous side effect profile is particularly important because long-term corticosteroid administration is associated with significant side effects such as metabolic disorders, osteoporosis, and other side effects. By combining exosomes with corticosteroids, such as cortisone, the adverse effects of corticosteroids (cortisone) are reduced and the efficacy of the treatment is improved.

Independent of the indication, it can be observed that in some patients exosomes in combination with corticosteroids show a therapeutic effect that exceeds the sum of the individual therapeutic effects of both exosomes and corticosteroid active substances, especially if both active substances are not therapeutically effective when administered alone.

Thus, in a first aspect, the present invention provides a pharmaceutical composition comprising a corticosteroid and an exosome in the presence or absence of a cytokine antagonist and/or a growth factor.

The expression "without cytokine antagonist" means within the scope of the present invention that no cytokine antagonist is present. Accordingly, this is also for growth factors.

According to a first aspect, the pharmaceutical composition thus comprises in each case a corticosteroid and exosomes. Furthermore, a choice of the presence or absence of cytokine antagonist is given; furthermore, a choice for the presence or absence of growth factors is given.

Likewise, the therapeutic agents may be administered in two different pharmaceutical compositions simultaneously or sequentially in time. Accordingly, in a second and third aspect, the present invention provides a pharmaceutical composition comprising exosomes, in the presence or absence of cytokine antagonist and/or growth factor, for use in combination therapy with a corticosteroid; and a pharmaceutical composition comprising a corticosteroid in the presence or absence of a cytokine antagonist and/or a growth factor for use in combination therapy with exosomes.

In a fourth aspect, the invention provides a pharmaceutical composition comprising a cytokine antagonist and/or a growth factor for use in combination therapy with a corticosteroid and exosomes.

In a fifth aspect according to the present invention, there is provided a kit comprising (i) a pharmaceutical composition comprising exosomes in the presence or absence of cytokine antagonist and/or growth factor, and (ii) a pharmaceutical composition comprising corticosteroid in the presence or absence of cytokine antagonist and/or growth factor.

Furthermore, in a sixth aspect, the present invention relates to the use of exosomes in the presence or absence of cytokine antagonists and/or growth factors for the preparation of a pharmaceutical composition for use in combination therapy with a corticosteroid; and, in a seventh aspect, to the use of a corticosteroid in the presence or absence of a cytokine antagonist and/or a growth factor for the preparation of a pharmaceutical composition for use in combination therapy with exosomes.

The combination therapy mentioned above in relation to the second, third, fourth, sixth and seventh aspects is preferably a combination therapy with a cytokine antagonist and/or a growth factor, especially in cases where the pharmaceutical composition itself is absent of a cytokine antagonist and/or a growth factor.

In an eighth aspect, the present invention relates to a process for preparing a pharmaceutical composition comprising a corticosteroid and exosomes, comprising the steps of: a blood sample containing exosomes is provided, preferably concentrated, and mixed with a corticosteroid.

Other embodiments of the invention are set forth in the following detailed description and in the claims.

Description of the invention

The present invention is based on the surprising recognition that joint and spinal diseases such as osteoarthritis, arthritis and degenerative spinal diseases, as well as in the case of autoimmune diseases such as neurodermatitis and alopecia areata, can be significantly improved by the additional administration of exosomes by means of the treatment with corticosteroids. The present invention therefore focuses on the combination treatment of such diseases with corticosteroids and exosomes. This may be done in the presence or absence of cytokine antagonists and/or growth factors. The presence of cytokine antagonists and/or growth factors is preferred.

The different active substances can be administered simultaneously-in the same formulation or in different formulations-or else one after the other. Thus, the pharmaceutical composition according to the invention, which comprises only one of the two active substances exosome and corticosteroid, as well as the kit according to the invention, can be destined for simultaneous administration of exosomes and corticosteroid on the one hand, and for their sequential administration on the other hand. However, simultaneous administration is preferred, in particular in only one formulation. Thus, for example, the two pharmaceutical compositions of the kit of the invention may be mixed in an appropriate ratio prior to administration to a patient and then administered as a single formulation. In case of sequential administration of exosomes and corticosteroid, the respective active ingredients are administered over a period of one week, preferably within 5 days, 3 days, 1 day or within 12 hours.

In the pharmaceutical compositions of the invention comprising exosomes and/or corticosteroids, cytokine antagonists and/or growth factors may or may not be present. The presence of cytokine antagonists and/or growth factors is preferred. Since in the case of joint and spinal disorders cytokine antagonists such as Orthokin (native IL-1Ra) and anakinra (recombinant IL-1Ra) have a clearly favourable effect on the efficacy, in particular the long-term efficacy, of corticosteroids, especially if the therapeutic agent is administered directly topically to the joint to be treated, if additionally. Similar, surprisingly good efficacy of these active substance combinations is also observed for autoimmune diseases, wherein in particular an anti-inflammatory effect plays a role. In the case of joint and spinal disorders such as osteoarthritis, arthritis and degenerative spinal disorders, the efficacy of the combination therapy with corticosteroids and exosomes is also greatly improved by the additional administration of cytokine antagonists. In the case of treatment with recombinant IL-1Ra, anakinra, surprisingly, efficacy was only produced by combination with corticosteroids. In the case of native IL-1Ra, ortho, a significant improvement in efficacy was observed just in the case of inflammatory diseases. One possible explanation for this is that cytokine antagonists have anabolic effects and may counteract or even reverse the catabolic effects of corticosteroids. Thus, in the treatment of, for example, osteoarthritis, corticosteroids are alternatively or additionally combined with anabolic growth factors in addition to cytokine antagonists to achieve similar effects. In general, growth factors may be used in place of or in combination with cytokine antagonists.

The pharmaceutical composition according to the invention (according to the fourth aspect of the invention) comprising a cytokine antagonist and/or a growth factor for use in a combination therapy with a corticosteroid and exosomes may be specified for simultaneous administration of the cytokine antagonist and/or growth factor, corticosteroid and exosomes on the one hand and for their sequential administration on the other hand. Simultaneous administration is preferred, in particular in only one formulation. Thus, for example, a cytokine antagonist and/or growth factor may be mixed with a corticosteroid and exosomes in appropriate proportions prior to administration to a patient and then administered as a formulation. Alternatively, in the case of simultaneous administration, the cytokine antagonist and/or growth factor, exosome or corticosteroid may be administered separately, with the other ingredients being mixed as one formulation. However, it is also possible to administer the cytokine antagonist and/or the growth factor, the exosomes and the corticosteroid sequentially in any order, or one of these active substances at a time point different from the other two. In case of sequential administration, these different active substances are preferably administered within a period of one week, preferably within 5 days, 3 days, 1 day or within 12 hours.

In the case of further pharmaceutical compositions according to the invention and in the case of kits according to the invention and uses according to the invention (first, second, third, fifth, sixth and seventh aspects of the invention), the presence or absence of a cytokine antagonist in a pharmaceutical composition is possible. The presence of a cytokine antagonist is preferred. Furthermore, the combination therapy according to the invention is preferably a combination therapy with a cytokine antagonist, especially in the case of the pharmaceutical composition itself without a cytokine antagonist (see above).

The cytokine antagonist used according to the invention may be each substance or each mixture of substances which reduces or inhibits at least one, preferably substantially all, of the biological activity of one or more cytokines in the body of the patient. Here, the antagonistic effect can be achieved directly or indirectly by the antagonist, for example by activating or inhibiting other signaling pathways, which in turn have an effect on the biological activity of the cytokine. Preferably, the biological activity of the cytokine is inhibited by preventing its interaction with one or more receptors to which it can bind. This can be achieved, for example, by competitive binding of the antagonist to the corresponding receptor or by binding of the antagonist to the cytokine itself. Preferably, the cytokine antagonist inhibits the effect of cytokine IL-1.

The cytokine antagonist may be, for example, a protein, peptide, nucleic acid, lipid, or organic compound. The cytokine antagonist may also consist of a mixture of two or more cytokine antagonists as described herein. In particular, the cytokine antagonist may be a naturally occurring peptide or protein, or may also be a recombinantly produced peptide or protein. In addition, the cytokine antagonist may be or comprise an antibody or antigen-binding fragment of an antibody, which may bind to the cytokine or cytokine receptor involved. Examples of suitable cytokine antagonists are interleukin antagonists, particularly IL-1 antagonists such as IL-1Ra, Tumor Necrosis Factor (TNF) antagonists, particularly TNF- α -antagonists such as anti-TNF- α -antibodies, interferon antagonists, and chemokine antagonists. Particularly preferred are naturally occurring or recombinant IL-1Ra proteins, preferably human IL-1 Ra. IL-1Ra preferably comprises or consists of the amino acid sequence: a subtype or homologue of human IL-1Ra according to SEQ ID NOs:1, 2, 3, 4 or 5, a subtype of IL-1Ra according to SEQ ID NOs:6 or 7 in a horse, or IL-1Ra according to SEQ ID NO:8 in a dog.

Furthermore, according to the invention, fragments or derivatives of IL-1Ra can be used as cytokine antagonists, as long as they can produce the desired function, i.e.reduce or inhibit one or more of the biological functions of IL-1. Preferably, fragments of IL-1Ra comprise at least 20, more preferably 40, 60, 80 or 100 amino acids of the native IL-1Ra sequence. Preferably, the fragment is a naturally occurring secreted fragment of IL-1 Ra. In one embodiment, the IL-1Ra comprises amino acids 26-177 of human IL-1Ra, preferably amino acids 26-177 of the sequence according to SEQ ID NO: 1. Derivatives of IL-1Ra are preferably homologous to native IL-1Ra, and preferably have at least 60%, more preferably at least 70%, 75%, 80%, 85%, 90%, 95% and most preferably at least 98% homology or identity to native IL-1Ra over at least 20 contiguous amino acids of IL-1Ra, preferably at least 40, 60, 80 or at least 100 contiguous amino acids, most preferably over the entire length of IL-1 Ra. Particularly preferred is the isolation of IL-1Ra (also known as Orthokin), and IL-1Ra fragments having amino acids 26-177 of human IL-1Ra (also known as anakinra), from natural biological samples such as blood. The obtaining of ortho is described in particular in patent applications WO 00/46249 a1 and WO 03/080122 a 1. Anakinra and other IL-1 antagonists which can be used in the present invention are described, inter alia, in patent application EP 0343684A 1.

In case the IL-1Ra is obtained from a natural biological sample, such as blood, for example in the case of Orthokin, the IL-1Ra solution obtained preferably also comprises growth factors. Thus, according to the present invention, a cytokine antagonist is also present in combination with or replaced by one or more growth factors. According to the invention, the growth factor preferably has an anabolic effect. Examples of suitable growth factors are TGF-beta, IGF, BMP, HGF and VEGF. Also included are analogs, derivatives and fragments of these growth factors, provided that they have the desired effect, i.e., in particular as growth factors.

The corticosteroid used according to the present invention may be any naturally occurring and synthetically prepared corticosteroid. In particular, it may be a glucocorticoid, mineralocorticoid or androgen, wherein preferably a glucocorticoid is used. Likewise, mixtures consisting of two or more corticosteroids as described herein may also be used. Examples of suitable glucocorticoids are cortisone, hydrocortisone, prednisone, prednisolone, deflazacort, fluocortin, triamcinolone, dexamethasone, methylprednisolone, fluprednisolone, beclomethasone, clobetasone, alclomethasone, flumethasone, fluprednide, fludroxycorsone, betamethasone, beclomethasone, mometasone, fluticasone, halomethasone, fluocinolone (fluocinolone), diflorasone, desoximetasone, fluocinonide, amcinonide, halcinonide, diflucortolone, clobetasol and paramethasone. Examples of suitable mineralocorticoids are aldosterone, deoxycorticosterone and fludrocortisone, and examples of suitable androgens are Dehydroepiandrosterone (DHEA) and estrogens. Corticosteroids may be used as the free compound or in the form of salts, esters or prodrugs. In a preferred embodiment, the corticosteroid used is triamcinolone, cortisone, hydrocortisone, prednisolone or prednisone.

Preferably, the exosomes in the pharmaceutical composition comprising exosomes according to the invention or in the kit according to the invention are preparable by a method comprising the following steps: providing a blood sample comprising exosomes and preferably concentrating the exosomes. The concentration is preferably performed by a centrifugation step at least 100000 g, since such high centrifugation accelerations are particularly suitable for concentrating exosomes. This step is preferably carried out for at least 30 minutes, in particular at least 60 minutes, since concentration is particularly effective thereafter.

The step of providing a blood sample comprising exosomes preferably comprises the steps of: providing a blood sample obtained from a patient, optionally adding an additive that promotes exosome formation, and incubating the blood sample in a container suitable for exosome preparation. This incubation results in a treated blood composition. Suitable containers for exosome preparation are, for example, syringes, cuvettes such as vacuum cuvettes, microtiter plates and infusion bags. The surface intended to come into contact with the blood sample preferably comprises, and preferably consists of, in the case of a container suitable for exosome preparation, glass, synthetic materials (such as polystyrene, polyvinyl chloride, polyethylene and polypropylene), corundum and quartz. Preferably, for the preparation of the exosomes, surface-enlarging additives consisting of glass, synthetic materials, corundum or quartz, such as pellets, gels, wool, flours, granules or microparticles, are added to these containers. The additive that promotes the formation of exosomes is preferably IL-1 Ra. The additive for promoting the formation of exosomes is preferably used in an amount of 1 to 20. mu.g/ml of whole blood.

In case of a pharmaceutical composition according to the invention for use in combination therapy with exosomes, it is suitable that the exosomes are obtained from a blood sample. The exosomes are preferably autologous or exogenous to the patient to be treated. For the obtention of exosomes, see the discussion of preparation of exosomes made above. Preferably, the centrifugation step described above with at least 100000 g (preferably, at least 30 minutes, in particular at least 60 minutes) is carried out in the treatment of a disease where it is reasonable to increase the concentration of exosomes, preferably in the treatment of rheumatoid arthritis. Particularly preferably, such centrifugation step is typically performed for cases where the combination therapy involves exosomes obtained from a blood sample of the patient.

Thus, the detailed discussion of the preparation of exosomes made above is considered to be a process for preparing a pharmaceutical composition comprising a corticosteroid and exosomes according to the present invention, comprising the steps of: a blood sample containing exosomes is provided, preferably concentrated, and mixed with a corticosteroid. The mixing can be carried out in any manner customary to the expert.

In a preferred embodiment, the pharmaceutical composition according to the invention or the kit according to the invention is intended for the treatment of joint diseases such as osteoarthritis, arthritis, joint inflammation and inflammatory cartilage defects, degenerative spinal diseases, joint pain and autoimmune diseases. Osteoarthritis to be treated may arise as a result of excessive loading, have congenital or traumatic causes or be the result of other diseases such as inflammation. The osteoarthritis to be treated is preferably in-onset osteoarthritis or inflammatory osteoarthritis. The pharmaceutical composition according to the present invention can be used for the treatment of osteoarthritis and arthritis in each of any joints such as knee joint, hip joint, ankle joint, shoulder joint, spinal joint, finger joint, elbow joint, toe joint, jaw joint, wrist joint. The arthritis to be treated may be infection-induced arthritis such as bacterial arthritis or non-infection-induced arthritis such as rheumatoid arthritis, psoriatic arthritis or gouty arthritis. Alternatively, the pharmaceutical composition according to the invention and/or the kit according to the invention are also intended for the treatment of diseases other than the one or more mentioned diseases (e.g. rheumatoid arthritis). The degenerative spinal disease to be treated may be, for example, a disc herniation. Autoimmune diseases include, in particular, autoimmune diseases of the joints such as, for example, Behcerew's disease (Morbus Bechterew), rheumatoid arthritis and systemic lupus erythematosus, and also other autoimmune diseases such as, in particular, neurodermatitis and alopecia areata (alopecia areata).

The pharmaceutical composition according to the invention or the kit according to the invention is preferably suitable for topical administration. They are preferably intended for topical administration. Thus, in a preferred embodiment they are specified for injection, in particular for injection into the body region to be treated, in particular into the affected joint, at the affected nerve root or in the affected intervertebral disc, or in its local environment. Thus, the pharmaceutical composition is specified for intra-articular and/or peri-radicular injection in particular. Alternatively, the pharmaceutical composition according to the invention may be formulated in particular as a cream or gel for topical administration, or in the form of tablets, capsules or lozenges for systemic administration. The mode of administration therefore depends inter alia on the disease to be treated. Thus, in the case of local osteoarthritis or degenerative spinal diseases, local administration of the pharmaceutical composition according to the invention is preferred. In a preferred embodiment, the pharmaceutical composition according to the invention or the kit according to the invention is only intended for or suitable for administration other than systemic administration.

The pharmaceutical compositions according to the invention are always suitably formulated for different administration routes in a manner known to the skilled person. Thus, pharmaceutical compositions suitable for injection are preferably present as a solution or dispersion, or even as a dry form such as a powder or a lyophilisate, which, prior to injection, must be dissolved in a suitable solvent such as water. The pharmaceutical composition according to the invention comprises a therapeutically effective amount of exosomes and/or corticosteroids. The corticosteroid is preferably present in the pharmaceutical composition comprising the corticosteroid at a concentration of 1-80 mg/dose, more preferably 5-40 mg/dose. If desired, the cytokine antagonist present is preferably present in the pharmaceutical composition comprising the cytokine antagonist at a concentration of 0.5-150 mg/dose, but may also be present at very small concentrations, e.g., 1 ng/dose or more, e.g., 1-1000 ng/dose. Small agent concentrations can be used in particular in combination with growth factors and/or in the case of natural IL-1Ra preparations, for example compositions with Orthokin. High agent concentrations are preferred, for example, in the case of recombinantly produced cytokine antagonists such as anakinra. Furthermore, the pharmaceutical composition according to the invention may also comprise one or more carrier substances and/or one or more excipients.

It may also be specified that the pharmaceutical composition of the invention is used for the treatment of patients who have already experienced an additional treatment of the relevant disease, i.e. e.g. osteoarthritis, arthritis and/or degenerative spinal diseases, in particular if this additional treatment has not been successful or the disease symptoms have at least partially relapsed despite the initial success of the treatment. In a preferred embodiment, the other treatment is treatment with exosomes but no corticosteroid; or treatment with corticosteroids, particularly corticosteroids such as those described above, but without exosomes.

Within the scope according to the invention, the patient may be a human or an animal suffering from a disease described herein. Thus, the pharmaceutical composition according to the invention may be suitable for the treatment of humans and/or animals, such as dogs, cats, horses, cows, pigs, goats, camels, etc.

Brief Description of Drawings

Fig. 1A to 1C show photographs of the feet of a patient with neurodermatitis, a case of which is described below as case IX (example 4).

Figure 1A again shows the dermatological changes in both feet immediately prior to treatment with exosomes and triamcinolone.

Fig. 1B shows the condition for the right foot one week after treatment.

Fig. 1C shows the condition for the left foot one week after treatment.

Examples

Various pathological studies of patients with advanced osteoarthritis are described below. It is treated with a cytokine antagonist (e.g., recombinant IL-1Ra or IL-1Ra obtained from an autologous blood sample) and a corticosteroid.

Abbreviations:

re right side

left side of li

bds double sided

IRO internal rotation

ARO external rotation

VAS pain visual simulation score (0 to 10)

WOMAC patient questionnaire for osteoarthritis

CRP c-reactive protein, inflammation marker detectable in blood

OSG hip joint

Treatment with orthiokin and cortisone in the case of osteoarthritis:

the number of patients is N =129

Average review time: 3 months old

Mean pain reduction 71% (i.e. from 100% pain before treatment to 29% after treatment)

Dramatic fast onset of action

1. Topical application of anakinra and cortisone and exosomes

Case I, T.56 years old, female

And (3) diagnosis: clinically by radiology methods, medial and postpatellar left gonarthritis, IV degrees, exists. From an external perspective, a full left knee replacement has been planned;

treatment: exosomes combining anakinra and 10mg triamcinolone were injected 3 times in the left knee (2x weekly) to avoid knee surgery.

As a result: at the third injection time point, 100% pain improved. The function is obviously improved. The scheduled period of the operation is declared cancelled, and the patient still has no pain after 5 months after the treatment is finished.

2. Topical application of anakinra and cortisone and Orthokin

Case II, L.57 years old, male

And (3) diagnosis: from 6 months onwards, intense left shoulder pain (VAS 8); from which sleep is significantly disturbed. Patients have little ability to sleep during the last 6 months, thereby also interfering with general health. Injection of cortisone in the left shoulder several times was not successful. The appointment for left shoulder surgery has been agreed. Attempts should be made here to avoid surgery. Radiologic and clinical findings of partial swivel cuff rupture and subacromial stenosis, complete left shoulder stiffness; the left arm feels uncomfortable, the hand and left forearm strength are insufficient, 4 degrees.

Treatment: injections were applied ventrally and laterally in the left shoulder. In the left shoulder, 2ml of Orthokin was applied by syringe together with 10mg anakinra and 10mg triamcinolone. Treatment was continued for 4 consecutive days.

As a result: on the second treatment day, the patient had developed an extreme improvement in pain with a reduction in pain of approximately 90%. The VAS drops from 8 to 1, the shoulder is free and can move normally. Patients may sleep continuously for the first time since 6 months. The patient is thereby significantly improved in his general health. Treatment continued on day 4. In case the review at 6 months after treatment gave positive examination results which were unchanged, still showed a significant improvement as unchanged on the 2 nd treatment day. The operation is cancelled and the mobility is free and the patient can lift the case and book over the shoulder height again without problems.

Case III, F., age 45, female

And (3) diagnosis: from about 8 months onwards, complete right shoulder stiffness. All treatments to date have had no effect and are planned for surgery. The patient is willing to try another conservative treatment. Since several weeks, nighttime sleep is no longer possible. Shoulder pain started on the left side, but the main examination result was the right shoulder, where VAS was 9, with severe acute attacks up to 10, whereby general health was overall reduced.

Treatment: on 6 consecutive days, the right shoulder was treated with 2ml of Orthokin applied separately in combination with another syringe consisting of a combination of 150mg anakinra and 5mg triamcinolone.

As a result: from day 5, 85% of pain improved. Starting from treatment 2, continuous sleep overnight is possible, thereby significantly improving general health. At the end of treatment, in the 1 st review 8 months post-treatment, VAS continued to show very good unchanged results; the procedure is cancelled.

3. Exosomes incubated with IL-1Ra and triamcinolone/prednisone

Case IV, S.S., 25 years old, male

And (3) diagnosis: severe juvenile rheumatoid arthritis has started around age 15.

Treatment was with 25mg Enbrel (2 times/week), 10mg of mdroprion, 5mg of deoxycorticosterone and naproxen (2X 1/day). Two OSGs and shoulders severe synovitis and pain. The shoulders are abducted by 60 degrees before treatment. Laboratory CRP value 5.35 (to 0.5mg is normal); leukocytosis.

Treatment: blood was drawn to prepare exosomes in 6ml syringes (Orthokin syringes). An incubation was then performed at 37 degrees for 24 hours, wherein the syringes were filled with blood before 1mg anakinra (IL-1 Ra) and 2mg prednisone were applied in the syringes. After different centrifugation steps (up to 100000 g), the mixture is applied to the patient at OSG and shoulder.

As a result: the joint started to subside significantly after 3 days. Clinical and laboratory examinations after 9 days showed an 80% improvement in pain, normal OSG, no swelling. The CRP value is now up to 1.93. And also improved in other affected joints that were not injected topically. The general quality of life is significantly improved. An unchanged stable condition was shown in the examination after 3 months. Pre-treatment VAS 9, VAS 3 from the first week post-injection. The patient is very satisfied and can work again.

4. Injection of exosomes and additional triamcinolone in injection

Case V, M.E., 64 years old, female

Severely resistant to treatment of RA despite treatment with a basis of prednisone 15mg (daily), Lantarel 20mg (weekly), Humira (every 2 weeks). Radiolytic synovectomy in the wrist joint showed only minimal effect, and intra-articular injection at doses of 10mg to 40mg triamcinolone showed only very weak (20% improvement in pain after one week) effects on pain and on inflammatory parameters. In the case of self-recommended exosome therapy, severe pain was still present in the hands and shoulders despite the 120mg/l CRP as the basal therapy described above. Both sides and shoulders were injected with exosomes (after incubation with IL-1Ra) and mixed with MCP2-5 with a total amount of 20mg of triamcinolone. Thereafter, a clinically strong improvement was initiated after 2 days (80% pain relief after 1 week), which persisted for more than 3 months. CRP was checked for 42.6mg/l after 3 months and basal treatment was unchanged, thus the effect was associated with exosomes in combination with triamcinolone. High doses of triamcinolone alone as described above do not have comparable effects.

Case VI, M.,25 years old, male

Known psoriatic arthritis; the main problem with basal treatment, 5mg prednisone and 10mg MTX, was also significant left knee swelling with synovitis, and 2cm swelling compared to the contralateral side. Additional swelling in the area of the wrist joint despite basic treatment. Intra-articular cortisone injections in the knee and wrist at doses of 20-40mg only show a very weak effect within a few days with a 10-30% reduction in pain. And performing exosome therapy. CRP5.6mg/l immediately prior to injection. After completion of exosomes, exosomes (prepared after incubation of IL-1Ra as described above) were applied in the left knee (exosomes + triamcinolone 10mg) and in both sides of MCP2+3 (exosomes each with 2mg triamcinolone/joint). Uncomplicated process, after several days, 90% of the pain is significantly relieved; CRP 3.0mg/l after 3 months, complete knee swelling after 1 week, no bilateral difference: in the context of basal therapy, 5mg prednisone may be completely stopped because the effects of intra-articular injection of exosomes and triamcinolone are long-lasting.

Case VII R, 32 years old, male

Given the years of complete alopecia, all treatment attempts include systemic and local cortisone application at high systemic and local doses (10-80mg prednisone), with no effect: treatment with exosomes is administered. Exosomes were prepared by the technique described above, and were injected intramuscularly in one shot with 10mg of triamcinolone. Normal course check, 3 months later, hair growth was detected in about half of the area of the growing hair before disease.

Case VIII h, 47 years old, male:

right-sided gonitis, with cartilage defects, was validated at 2-4 degrees under an arthroscope after Outerbridge. The patient wishes to withdraw OP and joint replacement. The previous medical history has no effect by injecting 10-40mg of triamcinolone into the joints. 1ml of exosomes prepared after incubation with IL-1Ra as described technique were injected. After a single intra-articular injection of 2ml exosomes, 4 weeks, the patient was not satisfied with clinical effect in terms of pain and function. Subsequently, it was decided to intra-articularly inject 10mg of triamcinolone in the right knee. Uncomplicated process, 95% of pain improved after 1 week, function was heavy and completely recovered, and the patient could re-play tennis again for the first time for several years.

Case IX j., age 27, male:

severe dermatologically-proven neurodermatitis, with strong changes especially in the range of hands and feet. All known dermatological treatments show no sustained improvement, in particular after intramuscular topical application of corticosteroids and systemic application of prednisone and triamcinolone at a dose of 80mg, only a short-term effect of 1 week is observed. Exosomes were prepared with the described techniques for intramuscular application; after 3 weeks, there was no satisfactory effect on skin changes and diseases. Subsequently, an intramuscular application of 2ml exosomes, combined with 20mg triamcinolone, was attempted. The improvement was significant within 2 weeks, which remained unchanged for more than 6 months. Therefore, although there is slight deterioration, there is a further improvement in comparison with the inspection result. Fig. 1A depicts the dermatological changes of both feet immediately prior to treatment, and fig. 1B and 1C are one week after right or left foot treatment.

Case X: b, 69 years old, female

Severe Behcet-Nerver arthritis with iridocyclitis of the left eye. As a basal treatment, patients received alternating doses of prednisone and MTX. CRP 20.3 mg/l; the left OSG clearly swollen +2cm relative to the other side; the MCP on the left and the distal phalanx of the 1 st left finger were markedly swollen and painful, and the swelling and pain in the right shoulder was reduced to 30 degrees compared to normal abduction. Subsequently, exosomes were injected in the affected joints with known techniques, without pain relief and CRP reduction, followed by triamcinolone injected in a total dose of 40mg in all affected joints with a slight improvement of 20% within 1 week. A total dose of 20mg of exosomes and triamcinolone was injected 4 weeks later, followed by 80% improvement in the shoulder and 50% improvement in the fingers with swelling of the left thumb ring MCP joint from 8.2cm to 6.6 cm. The ankle has no obvious effect. CRP decreased from 20.3mg/l to 2.9mg/l within 1 week.

Claims (27)

1. A pharmaceutical composition comprising a corticosteroid and exosomes in the presence or absence of a cytokine antagonist and/or growth factor.

2. A pharmaceutical composition comprising exosomes in the presence or absence of cytokine antagonist and/or growth factor for use in combination therapy with a corticosteroid.

3. A pharmaceutical composition comprising a corticosteroid in the presence or absence of a cytokine antagonist and/or a growth factor for use in combination therapy with exosomes.

4. A pharmaceutical composition comprising a cytokine antagonist and/or a growth factor for use in combination therapy with a corticosteroid and exosomes.

5. A kit comprising (i) a pharmaceutical composition comprising exosomes in the presence or absence of cytokine antagonist and/or growth factor, and (ii) a pharmaceutical composition comprising corticosteroid in the presence or absence of cytokine antagonist and/or growth factor.

6. The pharmaceutical composition according to claim 2 or 3 or the kit according to claim 5, wherein the pharmaceutical composition is intended for simultaneous or sequential administration of exosomes and corticosteroid.

7. Pharmaceutical composition or kit according to one of claims 1 to 6, wherein the corticosteroid is

(a) Glucocorticoids such as cortisone, hydrocortisone, prednisone, prednisolone, deflazacort, fluocortin, triamcinolone, dexamethasone, methylprednisolone, fluprednisolone, beclomethasone, clobetasone, alclomethasone, flumethasone, fluprednide, fludrociclesonide, betamethasone, beclomethasone, flumethasone, fluticasone, halomethasone, fluocinolone, diflunisal, desoximetasone, fluocinonide, amcinonide, halcinonide, diflucortolone, clobetasol and paramethasone;

(b) mineralocorticoids such as aldosterone, deoxycorticosterone and fludrocortisone; or

(c) Androgens such as Dehydroepiandrosterone (DHEA) and estrogens;

or a salt, ester or prodrug thereof.

8. Pharmaceutical composition or kit according to one of claims 1, 3, 5, 6 or 7, wherein the corticosteroid is present in the pharmaceutical composition containing a corticosteroid at a concentration of 1 to 80mg per dose.

9. The pharmaceutical composition or kit according to one of claims 1, 2, 5, 6, 7 or 8, wherein the exosomes are prepared by a method comprising the steps of: providing a blood sample comprising exosomes and preferably concentrating the exosomes.

10. The pharmaceutical composition according to one of claims 3, 4, 6, 7 or 8 for use in combination therapy with exosomes, wherein the exosomes are obtained from blood samples.

11. The pharmaceutical composition or kit according to claim 10, wherein the exosomes are autologous or exogenous to the patient to be treated.

12. Pharmaceutical composition or kit according to one of claims 1 to 11 for use in the treatment of joint diseases such as osteoarthritis, arthritis, joint inflammation and inflammatory cartilage defects, degenerative spine diseases and/or joint pain.

13. The pharmaceutical composition or kit according to claim 12, wherein the osteoarthritis is in-onset osteoarthritis or inflammatory osteoarthritis.

14. The pharmaceutical composition or kit according to claim 12, wherein the arthritis is infection-induced arthritis such as bacterial arthritis, or non-infection-induced arthritis such as rheumatoid arthritis, psoriatic arthritis or gouty arthritis.

15. The pharmaceutical composition or kit according to claim 12, wherein the degenerative spinal disease is a disc herniation.

16. A pharmaceutical composition or kit according to one of claims 1 to 11 for use in the treatment of autoimmune diseases.

17. The pharmaceutical composition or kit according to claim 16, wherein the autoimmune disease is neurodermatitis or alopecia areata.

18. Pharmaceutical composition or kit according to one of claims 1 to 17, wherein the pharmaceutical composition is suitable for topical administration.

19. The pharmaceutical composition or kit according to claim 18, wherein the topical administration is selected from the group consisting of: injection in the affected body region, in particular in the affected joints, on the affected nerve roots or in the affected intervertebral discs or in their local environment; intra-articular injection; and external application.

20. Pharmaceutical composition or kit according to one of claims 1 to 19, wherein the pharmaceutical composition further comprises a carrier and/or an excipient.

21. Pharmaceutical composition or kit according to one of claims 1 to 20, wherein a cytokine antagonist is present and is selected from: interleukin antagonists, particularly IL-1 antagonists such as IL-1Ra, Tumor Necrosis Factor (TNF) antagonists, particularly TNF- α -antagonists such as anti-TNF- α -antibodies, interferon antagonists and chemokine antagonists.

22. Pharmaceutical composition or kit according to claim 21, wherein the cytokine antagonist is a naturally occurring or recombinant IL-1 Ra-protein, in particular Orthokin or anakinra.

23. The pharmaceutical composition or kit according to claim 21 or 22, wherein the cytokine antagonist is present in the pharmaceutical composition comprising the cytokine antagonist at a concentration of 0.5-150 mg/dose.

24. Pharmaceutical composition or kit according to one of claims 1 to 23, wherein a growth factor is present and is selected from the group consisting of: TGF-beta, IGF, BMP, HGF and VEGF.

25. Use of exosomes in the presence or absence of cytokine antagonists and/or growth factors for the preparation of a pharmaceutical composition for use in combination therapy with a corticosteroid.

26. Use of a corticosteroid in the presence or absence of a cytokine antagonist and/or a growth factor for the preparation of a pharmaceutical composition for use in combination therapy with an exosome.

27. A process for preparing a pharmaceutical composition comprising a corticosteroid and exosomes, comprising the steps of: a blood sample containing exosomes is provided, preferably concentrated, and mixed with a corticosteroid.