Use of protease inhibitors as antiexudative, antiphlogistic and antimicrobial agents

Abstract: A medcamentous form with antiexudation, antiphlogistic and antimicrobial effects in an aqueous or ointment base, suitable above all as an ophthalmologic, otolaryngologic, or dermatologic drug, which contains inhibitors or proteases, as are aprotinin, soyabean trypsine inhibitor, and elastatinal, either single or in a combination, in the amount of 0.1 mg to 20 mg per 1 ml of physiological saline or buffer solution, which is preferably ionically balanced and has pH 6.5 to 7.5, pr per 1 g of ointment base. The medicamentous form may further contain 0.05 to 1.5 wt.-% of steroidal antiphlogistics, for example, dexamethasone, and/or 0.05 to 5 wt.% of nonsteroidal antiphlogistics, for example, indomethacin, and/or 0.2 to 1 wt.-% of antibiotics suppressing the growth of microbes, as are, for example, neomycin, bacitracin, chloramphenicol, tetracycline.

Description: The invention pertains to a pharmaceutically active composition and more particularly to a medicamentous form of the composition for external use and having strong antiexudation, antiphlogistic, and antimicrobial effects. The composition is provided in a suitable aqueous or ointment base and is applicable, for example, as an ophthalmologic, otolaryngologic, or dermatologic drug. Arachidic acid is liberated in damaged, wounded or inflamed tissues from phospholipids of cytoplasmatic membranes by the action of phospholipase enzyme, and may be then metabolized by the cyclooxygenase cycle (by cyclooxygenase enzyme) or the lipooxygenase cycle (by lipooxygenase enzyme) to prostanoids and eicosanoids. Antiphlogistics of both the steroid and nonsteroid nature, antibiotics, and sulfonamides are often used for therapeutic purposes. The antibiotics, which specifically suppress pathogenic microbes and which are most often used in ophthalmology, include tetracycline, chloramphenicol, bacitracin, and neomycin. Therapeutic substances which prevent the development of inflammation (antiphlogistics) are known in both steroidal and nonsteroidal forms. The steroidal antiphlogistics (for example dexamethasone) block in particular phospholipase. The antiinflammatory drugs of nonsteroid nature (e.g., indomethacin, flurbiprofen, pirprofen) block in particular cyclooxygenase. The blockade of these enzymes is important, because the products formed in metabolic cycles have a strong chemotactic effect (they cause accumulation of leucocytes (e.g., some leucotrienes) in the sites of origin) and increase the vascular permeability. This contributes to an excessive development of inflammation. Inflammations (both of infectious and noninfectious origin) are very dangerous for the anterior and posterior segments of the eye. Thus, scars formed in the cornea as the final stage of healing processes cause the loss of an exceptional function of this tissue, i.e. of its transparency. The loss of transparency of optical media of the eye (cornea, lens) then leads to a reduction in or even loss of sight. Disadvantages of locally applied conventional antiphlogistics are their relatively low efficiency, retardation of healing, and their contribution to the development of infection. The local effect of antibiotics has been found to be limited and, moreover, there is a danger of the development of an allergic reaction. A higher concentration of antibiotics, which is necessary for obtaining the healing effect in many cases, can have toxic effects on the tissue. For this reason, a local treatment with antibiotics is commonly supplemented by a general administration of antibiotics, which has disastrous consequences with respect to the suppression of antibody formation and the ability of the organism to combat infection. This is why new methods of treatment are desirable. One of the possibilities for use in the treatment of inflammation is the inhibition of plasmin and other destruction proteases (e.g., collagenase or elastase) with specific inhibitors. The said enzymes either directly develop the destruction processes (e.g., plasmin) or enable or activate these processes by their own activity (e.g., collagenase, elastase). However, plasmin is effective not only as an initiator of the development of the chain of degeneration processes, but also contributes to an excessive development of inflammation by several other mechanisms, of which at least chemotaxis should be mentioned. Amongst others, plasmin may be inhibited with aprotinin. This substance when administered in an aqueous solution and low concentrations has been successfully used in the treatment of some lesions of the anterior segment of eye. See EP-A-0223254 which discloses the use of 5 to 200 IU/ml (0.0007 to 0.028 mg/ml) aprotinin for such use. The authors of this invention found, moreover, that aprotinin inhibits not only plasmin but also several other proteolytic enzymes (for example, leucocytic elastase), which contribute to the destruction processes. Accordingly, the present invention seeks to provide a pharmaceutically active composition which can be put into a medicamentous form, in an aqueous or ointment base, particularly suitable as an ophthalmologic, otolaryngologic, and dermatologic drug, which contains, inhibitors of proteases. According to the present invention there is provided a pharmaceutically active composition suitable for use in an ophthalmologic, otolaryngologic or dermatologic medicament having anti-exudation, antiphlogistic and/or anti-microbial properties, which composition comprises an inhibitor of proteases in physiological saline or buffer solution or in an ointment base characterised in that the inhibitor of proteases is selected from elastatinal, elastatinal and aprotinin, and elastatinal and soya bean trypsin inhibitor and is present in a concentration of 0.1 to 20mg of inhibitor per 1ml of physiological saline or buffer solution or per 1g of ointment base. There is also provided a medicament containing the composition of the present invention. The inhibitors may be applied either singly or in combination, dissolved in physiological saline or buffer solution with pH 6.5 to 7.5, which is advantageously ionically balanced (e.g., phosphate or borax buffer) or present in an ointment base. The ionically balanced buffer solution means that sodium chloride is added to the buffer solution in such a way, that the resulting solution is ionically balanced, for example, a precise formulation for a borax buffer with pH 7.4 is as follows: solution A - 1.9 g Na 2 P 4 O 7 per 100 ml H 2 O pro injectione B - 1.25 g H 3 BO 3 + 0.3 g NaCl per 100 ml H 2 O pro injectione - 10 ml of solution A + 90 ml of solution B. The composition and its medicamentous form according to the invention in liquid state may further advantageously contain 0.05 to 15 wt.-% of thickeners selected from the group comprising hydroxypropyl methyl cellulose, methyl cellulose, polyvinylpyrrolidone, polyvinyl alcohol, poly(alklene glycols), poly/hydroxyalkyl(meth)acrylates or poly(meth)acrylamides. High concentrations of elastatinal alone, or also with the further specified inhibitors, locally applied can act not only curably with respect to the advanced stage of disease but also preventively, i.e. they can prevent the formation or onset of tissue destruction processes if administered in time. The vehicles (thickeners) with protracted effect then enable a longer contact of the remedy (e.g. aprotinin) with the tissue. The combination of elastatinal with other inhibitors, e.g. with aprotinin or inhibitor from soya-beans, enhances the therapeutic effect. The pharmaceutically active composition according to the invention may advantageously further comprise 0.05 to 1.5 wt.-% of steroidal antiphlogistics, for example, indomethacin, and/or 0.2 to 1 wt.-% of antibiotics killing microbes, for example, bacitracin, neomycin, tetracycline, or chloramphenicol. The combination of the protease inhibitor elastatinal alone or with the other specified inhibitors with antiphlogistics or antibiotics or all the substances together increases the anti-inflammatory and anti-microbial effect (the inhibitors block some products of microbes, e.g. elastase and other proteases). This permits the use of antibiotics only locally and in smaller doses. The concentration of antiphlogistics may also be reduced and, at the same time, the therapeutic effect is greater and the time of treatment may be shorter, which is of particular importance for the healing of tissue. The composition or its medicamentous form is most often applied by instillation or as an ointment into the conjunctival sac. However, it can be also used for irrigation or lubrication of the eye, facial sinuses, and external auditory meatus, and it may be injected into the anterior eye chamber, and the like. The composition or its medicamentous form in the liquid state may also be presented in the form of a hydrophilic three-dimensional polymer matrix, advantageously in the form of a strip, contact lens, and the like, from which the active components are released in use. The incorporation of the composition or its medicamentous form into a hydrophilic matrix may be performed according to the invention, for example, by conditioning of the matrix in a solution of the composition or its medicamentous form in order to attain the required concentration of inhibitors, or also antiphlogistics and antibiotics, in the polymer matrix. Pharmaceutically active composition according to the invention have a strong antiexudative, antiphlogistic and antimicrobial effect. Their therapeutic effects consist above all in the inhibition of plasmin, leucolytic elastase, and other serine proteases, and in the inhibited activation of latent forms of some endoproteases and several further subsequent reactions such as chemotaxis and vasculatisation of the cornea. The pharmaceutically active composition according to the present invention prevents in many cases the development of some diseases and, in other cases, stops the development of disease. The invention is further illustrated in the following exemplary examples, without being limited in its scope to them. Preparation of medicamentous form in liquid state: Each substance is separately dissolved in a small amount (10 to 40 ml) of buffer or physiological saline. Ointment base and ointment preparation: 10 g lanolin, 10 g liquid paraffin, and 80 g white petroleum jelly is melted in a water bath, the mixture is strained through hydrophilic gauze and then sterilized. If the therapeutic ingredient is easily soluble in water, it is dissolved in the necessary amount of distilled water for the preparation of injections, mixed with the ointment base in part molten form in a water bath and stirred until complete cooling is effected. If the therapeutic ingredient is insoluble in water, it is used for this preparation in its finest powdered form and first triturated in a smaller amount of liquid paraffin and then mixed with the ointment base. Example 1 Aprotinin 0.025 g elastatinal 0.005 g hydroxypropyl methyl cellulose 2.5 g ionically balanced borax buffer up to 100 g The drops applied into the conjunctival sac every 4 hours were successfully used for the treatment of corneal infiltrates which disappeared during a week. Example 2 Aprotinin 0.004 g inhibitor from soya beans 0.1 g hydroxypropyl methyl cellulose 1.5 g elastatinal 0.01 g ionically balanced phosphate buffer up to 100 g The drops were used for the treatment of a cornea etched with concentrated acids and hydroxides. The cornea healed during a month and the transparency was recovered either completely or in part. Example 3 Aprotinin 0.1 g elastatinal 0.01 g prednisolone acetate 0.02 g polyvinylpyrrolidone (mol. weight 360,000) 1 g ionically balanced phosphate buffer of pH 7.4 up to 100 g The drops were instilled 4 times a day for the treatment of deep corneal infiltrates. Healing occurred within a week. In cases complicated with a secondary inflammation of the iris, also this inflammation disappeared during a week. Example 4 Aprotinin 1 g dexamethasone sodium phosphate 0.1 g indomethacin 1 g elastatinal 0.1 g A contact lens comprising HEMA - DEGMA (the crosslinked copolymer of 2-hydroxyethyl methacrylate with diethylene glycol methacrylate containing 55 wt.-% of equilibrium water) was applied to an eye etched with strong alkalies and the drops were instilled over a contact lens at intervals of 4 h. The lesion healed during 3 weeks. Example 5 Aprotinin 1 g tetracycline 0.3 g elastatinal 0.2 g flurbiprofen 0.5 g hydroxypropyl methyl cellulose 3 g ionically balanced phosphate buffer (pH 7.4) up to 100 g The drops were instilled at intervals of 3 h into eyes etched with concentrated hydroxides. The cornea did not break down and healed with a scar during a month. Example 6 Elastatinal 0.5 g dexamethasone sodium phosphate 0.1 g polyvinylalcohol 2 g physiological saline up to 100 g The drops were instilled into the conjunctival sac at intervals of 3 h. Perforation of the cornea healed during a week. Example 7 Elastatinal 0.3 g soya-bean inhibitor of trypsin 0.2 g hydroxypropyl methyl cellulose 2 g ionically balanced phosphate buffer (pH 7.4) up to 100 g The drops were instilled into the conjunctival sac 4 times a day. Defects of the corneal epithelium healed during a week. Example 8 Aprotinin 0.1 g elastatinal 0.05 g dexamethasone sodium phosphate 0.1 g chloramphenical 0.5 g physiological saline up to 100 g The solution was successfully used in the treatment of rhinal allergoses and allergoses of meatus acusticus externus. Examples 9 Elastatinal 0.1 g aprotinin 0.1 g dexamethasone sulfate 0.15 g flurbiprofen 0.1 g chloramphenicol physiological saline up to 100 g A contact lense made from the crosslinked copolymer of 2-hydroxyethyl methacrylate and diethylene glycol methacrylate (HEMA - DEGMA) (55 wt.-% of equilibrium water content) was swelled in this solution for 24 h. The contact lens was used for the treatment of corneae burned with alkalis and lime. The healing ad integram occurred in some cases; in more severe cases, where the cornea usually had broken down, the tissue after the application of lenses healed with a scar. In other cases, the contact lens proved very suitable for the treatment of corneal ulcers. Example 10 Aprotinin 1 g elastatinal 0.1 g flurbiprofen 0.1 g dexamethasone sodium phosphate 0.1 g neomycin sulphate physiological saline up to 100 g A contact lens from the crosslinked polymer of 2-hydroxyethyl methacrylate (38 wt.-% of equilibrium water) was swelled in the given solution for 24 h. The contact lens was used for the treatment of non-healing corneal erosions. Within a week after application, the reepithalization was accelerated. Extra-ordinary results were attained after severe etching and burning of the anterior segment of eye. An intraocular inflammation did not develop, the cornea did not exhibit ulceration and healed with a scar during a month. The transparency recovered in the periphery of corneae. Example 11 Aprotinin 0.02 g elastatinal 0.05 g dexamethasone sulphate 0.1 g chloramphenicol 0.2 g ionically balanced phosphate buffer (pH 7.4) up to 100 g A strip from HEMA - DEGMA material (55 wt.-% of equilibrium water content) was immersed for 24 h into the solution of the above given composition and then applied into the lower fornix of the eye. After 24 h the strip was removed from eye, immersed into the given solution overnight, and applied again. Corneal ulcers of various origin were healed after a week of treatment (application).

Claims: Claims for the following Contracting States : AT, BE, CH, DE, DK, FR, GB, IT, LI, LU, NL, SE A pharmaceutically active composition suitable for use in an ophthalmologic, otolaryngologic or dermatologic medicament having anti-exudation, antiphlogistic and/or anti-microbial properties, which composition comprises an inhibitor of proteases in physiological saline or buffer solution or in an ointment base characterised in that the inhibitor of proteases is selected from elastatinal, elastatinal and aprotinin, and elastatinal and soya bean trypsin inhibitor and is present in a concentration of 0.1 to 20mg of inhibitor per 1ml of physiological saline or buffer solution or per 1g of ointment base. A composition according to claim 1, wherein the buffer solution is ionically balanced and has a pH in the range of from 6.5 to 7.5. A composition according to any preceding claim, wherein the composition further comprises from 0.05 to 1.5 wt% of a steroidal antiphlogistic agent. A composition according to claim 3, wherein the steroidal antiphlogistic agent is dexamethasone. A composition according to any preceding claim, wherein the composition further comprises from 0.05 to 5 wt% of a non-steroidal antiphlogistic agent. A composition according to claim 5, wherein the non-steroidal antiphlogistic agent is indomethacin. A composition according to any preceding claim, wherein the composition further comprises from 0.2 to 1 wt% of an antibiotic agent. A composition according to claim 7, wherein the antibiotic agent is neomycin, bacitracin, chloramphenicol or tetracycline. A composition according to any preceding claim, wherein the composition further comprises from 0.05 to 15 wt% of thickeners selected from hydroxypropyl methyl cellulose, methyl cellulose, polyvinylpyrrolidone, polyvinylalcohol, poly(alkylene glycols), poly[alkylene glycol (meth)acrylates] and poly(meth)acrylamides. A medicament comprising a composition according to any preceding claim. A composition or medicament according to any preceding claim when present in a hydrophillic matrix, which matrix is advantageously in the form of a strip or contact lens. The use of a composition or medicament according to any preceding claim, for the manufacture of a medicament for antiexudation, antiphlogistic and/or antimicrobial ophthalmologic, otolaryngologic or dermatologic treatment. Claims for the following Contracting States : ES, GR A method of preparing a pharmaceutically active composition suitable for use in an ophthalmologic, otolaryngologic or dermatologic medicament having antiexudation, antiphlogistic and/or antimicrobal properties, which method comprises admixing at least one inhibitor of proteases and physiological saline or buffer solution or an ointment base characterised in that the inhibitor of proteases is selected from elastatinal, elastatinal and aprotinin, and elastatinal and soya bean trypsin inhibitor and is admixed in an amount of 0.1 to 20mg of inhibitor per 1ml of physiological saline or buffer solution or per 1g of ointment base. A method according to claim 1, wherein the buffer solution is ionically balanced and has a pH in the range of from 6.5 to 7.5. A method according to claim 1 or 2, which further comprises admixing in the composition from 0.05 to 1.5 wt% of a steroidal antiphlogistic agent. A method according to claim 3, wherein the steroidal antiphlogistic agent is dexamethasone. A method according to any preceding claim, which further comprises admixing in the composition from 0.05 to 5 wt% of a non-steroidal antiphlogistic agent. A method according to claim 5, wherein the non-steroidal antiphlogistic agent is indomethacin. A method according to any preceding claim, which further comprises admixing in the composition from 0.2 to 1 wt% of an antibiotic agent. A method according to claim 7, wherein the antibiotic agent is neomycin, bacitracin, chloramphenicol or tetracycline. A method according to any preceding claim, which further comprises admixing in the composition from 0.05 to 15 wt% of at least one thickener selected from hydroxypropyl methyl cellulose, methyl cellulose, polyvinylpyrollidone, polyvinylalcohol, poly(alkylene glycols), poly[alkylene glycol(meth)acrylates] and poly(meth)acrylamides. A method according to any preceding claim, which further comprises forming the composition into a medicament. A method according to any preceding claim, which further comprises impregnating the composition or medicament into a hydrophilic matrix, which matrix is advantageously in the form of a strip or contact lens. The use of a composition or medicament as formed by any preceding claim for the manufacture of a medicament for antiexudation, antiphlogistic and/or antimicrobial ophthalmologic, otolaryngologic or dermatologic treatment.

Inventors: CEJKOVA JITKA, LOJDA ZDENEK, VACIK JIRI, CEJKOVA, JITKA, LOJDA, ZDENEK, VACIK, JIRI, CEJKOVA, JITKA, C/O CZECHOSL. ACADEMY OF SCIENCES, LOJDA, ZDENEK, C/O CZECHOSL. ACADEMY OF SCIENCES, VACIK, JIRI, C/O CZECHOSL. ACADEMY OF SCIENCES
Applicants: CESKOSLOVENSKA AKADEMIE VED, ACADEMY OF SCIENCES OF THE CZECH REPUBLIC
Application year: 1990
Main classification: A61K38
IPC codes: A61K9, A61K38