AU601712B2

AU601712B2 - Stabilised formulations of alpha-interferon

Info

Publication number: AU601712B2
Application number: AU68292/87A
Authority: AU
Inventors: Helmut Dr. Franz
Original assignee: Dr Karl Thomae GmbH
Current assignee: Boehringer Ingelheim Pharma GmbH and Co KG
Priority date: 1986-02-05
Filing date: 1987-02-04
Publication date: 1990-09-20
Anticipated expiration: 2007-02-04
Also published as: DK164202B; NO169638C; NZ219169A; ATE63823T1; FI86144B; PH24377A; IE870295L; HU196560B; AU6829287A; ES2028796T3; NO870441L; EP0231816A3; NO169638B; NO870441D0; ZA87793B; FI870457A0; IE59697B1; DK58387D0; GR3002270T3; FI86144C

Abstract

The pharmaceutical forms are intended for topical administration and employ as vehicle for and stabiliser of the active ingredient a composition which specifically confers special stability on the active ingredient IFN alpha. The forms are essentially composed of hydroxyethylcellulose and gelatin and preferably contain highly purified IFN alpha which has been prepared by DNA recombination as active ingredient.

Description

COMMONWE A-L TH OF A -S T R A L IA PATENT ACT 1952 COMPLETE SPECIFICATION J (Original) FOR OFFICE USE Class Int. Class Application Number: Lodged: zcrz7,/?7 Complete Specification Lodged: Accepted: Published: Priority: Related Art: 4' 4; S t f i Name of Applicant: Address of Applicant: Actual Inventor(s): Address for Service: DR. KARL THOMAE GESELLSCHAFT MIT BESCHRANKTER HAFTUNG D-7950 Biberach an der Riss, FEDERAL REPUBLIC OF GERMANY Helmut FRANZ DAVIES COLLISON, Patent Attorneys, 1 Little Collins Street, Melbourne, 3000.

Complete Specification for the invention entitled: "STABILISED FORMULATIONS OF e-INTERFERON" The following statement is a full description of this invention, including the best method of performing it known to us -I ~e la- 16T 51 334 "Stabilised formulations of a-interferon" The present invention relates to stabilised formulations of a-interferon for topical use.

Human interferons, and interferons in general, are divided into three groups, depending on their biological and chemical properties and the site of their synthesis, which are called alpha-interferon, beta-interferon and gamma-interferon, corresponding to a-IFN, B-IFN and -IFN. The site of synthesis of a-IFN, to which the present invention relates, Sis the leucocytes which, after stimulation or induction If 15 by appropriate inducers (for example viruses or S\ poly I' poly C) synthesize and release a-IFN.

4 e Bacteria transformed with suitable DNA sequences Sare able to synthesise a-IFN which has a biological activity analogous to that of the genuine human a-IFN, with the advantage that highly purified proteins which are essentially free of contamination can be obtained in comparatively large amounts and can be used for clinical administration, for example against various viral infections and malignant tumours.

All known interferons have antiviral, antiproliferative and, but to varying extents, immunomodulating effects.

S.Initially recognised as antiviral agents, their therapeutic uses now include their use as antineoplastic agents.

The application route is of particular importance with respect to the nature of the interferon effects and the rate of side effects attached thereto.

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I t 2 Systemic administration of interferons has particularly noticeable disadvantages since they reach and affect not only the target tissue but also healthy, uninfected or non-transformed cells or organs and thus induce side effects which may prevent intravenous or intramuscular use. In addition, the efficacy of systemic, administration has not always proved to be that desired.

In a few cases, although direct injection may be possible, the agent of choice is, for example for viral or malignant diseases or other related diseases which can be treated with interferon and which are accessable to external and local treatment (for example, skin, eyes, nose), topical administration of interferon, or of a-IFN to which this invention relates. Very 15 recently there have been reports from various sources of the topical application of human leucocyte interferon (a synonym for a-IFN) and it was surprisingly found that the success rate for various malignant diseases was high (for example Ikic et al, Lancet, 1022-24, 1981, Vol.1; Ikic et al., Lancet 1025-27, 1981, Vol.l).

It is conceivable that the topical administration of a-interferon will become of increasing medical importance in the future for the treatment of various viral or tumour-related diseases.

There are considerable difficulties, from various aspects, associated with the topical administration of a-IFN, as well as with interferons in general.

From the aspect of the pharmaceutical formulation, 30 a requirement is that sufficiently high stability is conferred on the biologically active a-IFN in order for it to act over an adequately long period both at room temperature and at body temperature or to remain effective "in situ" over a suitable period of hours. It is known that a-IFN is a protein which is very unstable to heat. Considerable losses of activity, of up to 80%, have been observed within 0.1 0 0 1411

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'It i L- l-, i L.1 -LIL- i -I 3 a few weeks at 4 0 C in a gel which contained human leucocyte interferon and, as gel component, carboxyrnethyl cellulose (Moller B.R. et al., Obstetrics Gynecology 62, 625-629, 1983) used for topical administration in the treatment of cervical carcinoma in situ.

The stability-maintaining components which are therefore necessary must be of such a nature that they neither cause intolerance reactions at the site of administration, conceivably due to the effect of biologically active foreign substances, nor impair the penetration or diffusion of a-IFN.

Recently, various components have been used to S: 15 stabilise interferons. Estis, Evans and Testa have used hydroxyethyl cellulose as an excipient for a the preparation of interferon-containing gels or ointments but have only been able to prevent considerable losses of a-IFN, B-IFN or 8-IFN activity under 20 realistic conditions of administration by the addition of a protease inhibitor (EP-A-0 142 345).

it Those skilled in the art know that hydroxyethyl cellulose is extremely well suited for the preparation of gels or other matrices, for example because of its stability to heat, and its ready commercial availability.

The particular advantages of hydroxyethyl cellulose 30 compared with other excipients for the preparation of a matrix containing a-IFN is that it does not coagulate or gel at the boiling point of water, it is very soluble in water, and the viscosity of such a solution increases with decreasing temperature.

It is not possible to stabilise interferon or a- IFN at physiological temperatures by hydroxyethyl i;lil 4 cellulose alone. It is also well known that protein products such as interferons, which have not been highly purified and are thus contaminated, are subject to hydrolysis by proteases under suitable conditions.

Other attempts to confer suitable stability on interferons contained in pharmaceutical preparations have been made by Akagi, Miura and Hoshino (EP-A-0 150 067) using polysaccharides differing chemically from hydroxyethyl cellulose, such as dextran and hydroxyethyl starch with the addition of human serum albumin, mono- or disaccharides, or amino acids, the preparations described being chiefly for the parenteral, systemic administration route via intravenous or intramuscular injection.

4 Various sugar alcohols such as glycerol, erythritol, Sarabitol, xylitol, sorbitol and mannitol, as well as various sugar acids such as iduronic acid, galacturonic acid and other uronic acids, and ascorbic acid and 20 their salts, and various organic buffer systems such as citrate buffer, succinate buffer, tartrate buffer, fumarate buffer, glyconate buffer, oxalate buffer, 4 lactate buffer and acetate buffer, in combination with various pharmaceutical excipients such as waxes, sodium carboxymethyl cellulose, carboxyvinyl derivatives Sand water have also been disclosed for the stabilisation of interferon in gels, ointments, suppositories, sprays etc. for topical administration (EP-A-0 080 879).

«i All the above-mentioned interferon-stabilising components, and especially the compositions of the pharmaceutical formulations and their methods of administration, are fundamentally different from the present invention and cannot be compared thereto.

There has therefore been a need for a carrier matrix for highly purified a-interferon, specifically

I

is 5 for recombinant-prepared a-IFN, which would comply with the requirements listed above as well as having a composition which is as favourable as possible, in terms of both the preparation technique and economics, i.e. a simple composition for topical administration which does not cause loss of the requisite optimal efficacy of the biologically active substance a-IFN contained in the excipient matrix. We have now found that by combining together a suitable carrier, especially hydroxyethyl cellulose, and gelatine in appropriate proportions we obtain a carrier and stabiliser for a-IFN which not only meets the requirements described above, at least in large measure, but also, surprisingly, O4t, 15 confers a particular stability on the biological r activity of a-IFN, and thus is extremely suitable for the topical administration of highly purified a-IFN, especially that prepared by recombinant fit DNA technology.

Thus in one aspect our invention provides pharmaceutical formulations for topical administration which stabilise the a-interferon contained therein, comprising as carrier and stabiliser a mixture of 1. a cellulose derivative, polyvinyl alcohol, polyvinylpyrrolidone, polyacrylic acid polymer or a mixture of any of these, and 2. gelatine, which has been subjected to acid or alkaline digestion, and, in addition, an anti-adhesion agent.

Another advantage of this invention is that the carrier/stabilising complex which has been found contains no human serum albumin which it is known may, especially on topical administration over lengthy periods, result in local immune reactions extending up to allergies.

Hence the present invention provides improved pharmai t I 6 ceutical formulations for the topical administration of a-IFN.

In the formulations of the present invention the two individual components 1) and which function as carrier and stabiliser, together exert an activitymaintaining effect on a-IFN which is highly unexpected.

It is known that a-IFN, and interferons in general, are proteins which are unstable to, for example, physical effects both in the "naked" form and when incorporated into other gel matrices (for example, .Moller B.R. et al., see above). However the formulations o. according to the invention appear to have a remarkable and unexpected intrinsic or inherent synergistic effect on the stability of a-IFN. Thus the gelatine in the present invention does not on its own act as a significant stabilising agent; only in a complex together with component 1) does it display the astonishing t0, synergistic stabilising affect on a-IFN to which S20 this invention relates.

t4 The pharmaceutical formulations according to the invention may contain a-IFN obtained from a cell culture which has been transformed with a DNA sequence coding for human a-IFN. Preferably the pharmaceutical formulations contain a-IFN obtained from transformed cells of E.Coli.

The pharmaceutical formulations according to the invention may contain a-IFN with a specific activity of 1 x 105 to 1 x 100 Iu/100g finished product.

The pharmaceutical formulations of a-IFN according to the invention which are for topical administration are preferably administered as viralstatics, therapeutic use being dermal, nasal, ophthalmic or intravaginal, A for example, for Herpes labialis, Herpes genitalis

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0- 7 including papilloma infections, warts and for Herpes zoster infections. The range of uses also embraces topical treatment of paracanceroses and canceroses.

The pharmaceutical formulations may additionally comprise other conventional adjuvants such as buffers and salts, humectants, preservatives, penetrationpromoters, and excipients.

Components 1) of the carrier/stabiliser mixture are known substances, viz. cellulose derivatives (e.g.

O carboxymethyl cellulose, methyl cellulose, hydroxypropyl- ,0 0 methyl cellulose) and polyvinyl alcohols, polyvinyl- 0o pyrrolidone, polyacrylic acid polymers (Carbopol 0 0 a' 15 and mixtures of any of these, together with, as component gelatine which has been subjected to acid or alkaline digestion preferably having a Bloom value of 160-200. The preferred excipient and stabiliser used for a-IFN is hydroxyethyl cellulose 20 with a degree of substitution of about 2.5, in defined mixing ratios of 0.1 to 2% by weight with respectively from 0.1 to 10% by weight of gelatine, based on the finished product. When polyvinyl alcohols or polyvinyl pyrrolidone are used then up to 15% by weight of these are employed. This achieves special stabilisation, specifically of °0 a-IFN, as demonstrated for example in the stability behaviour experiments which have been carried out and as shown graphically in Figures 1 to 3, which

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t embrace an experimental period of 14 weeks with storage temperatures of -10, +4 and +21 0 C in each case.

It is possible to use anti-adhesion agents to improve the spreading of the gels, and as anti-adhesive filtration aids anionic, cationic as well as neutral l4 surface-active substances can be used, preferably polyethoxylated sorbitan estars such as Polysorbate S 20, 60 or 80 whose chemical composition is given 7-E s' 8 hereinafter. Appropriate inorganic ax buffer solutions having a pH in the ri 7.4, which is optimal for a-IFN prepai sodium phosphate buffer, potassium hyc sodium dihydrogen phosphate buffer anc phate buffer, potassium sodium glutam; buffer and succinate buffer.

.1 id organic ange 6.5 to rations, are drogen phosphated citric acid/phosate phosphate i f f

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I r It iic t Suitable humectants for the formulations containing a-IFN, according to the invention are glycerol, 1,2-propylene glycol, sorbitol, butylene glycol and polyols polyethylene glycol 400).

Examples of suitable preservatives which are tolerated 15 by skin are e.g. p-hydroxybenzoic esters (Nipaester), preferably methyl p-hydroxybenzoate; also benzoic acid, sorbic acid, chlorhexidine digluconate, benzalkonium chloride and hexadecyltrimethylammonium bromide (cetrimonium bromide).

A suitable penetration-promoter which is compatible with a 2 -IFN is up to 50% by weight of dimethyl sulfoxide (DMSO).

The most suitable vehicle for non-alcoholic hydrogels is water.

Suitable pharmaceutical forms for topical administration on the skin, in the eye or in the nose or other 30 body cavities are, for example, hydrogels. Hydrogels generally contain 0.1 to 2.0% by weight of both gelatin and cellulose derivatives and/or polyvinyl alcohols, polyvinylpyrrolidones or polyacrylic acids, it being necessary to adjust the mutual ratios depending on the desired viscosity. When polyvinyl alcohols or polyvinylpyrrolidone is used, q up to 15% by weight thereof may be necessary.

)i t-' I Itr iv 8a In the case of the manufacture of vaginal suppositories the proportion of gelatine should be increased to as much as 10% by weight, likewise depending on the desired viscosity.

According to another aspect, the invention provides a process for the preparation of pharmaceutical formulations for topical use which stabilise the a-interferon contained therein, wherein: a. gelatine in an amount of 0.1 to by weight, based on the finished product, is dissolved in water at up to 80 0

C,

4I and an anti-adhesion agent is added t to the solution, b. optionally preservatives, humectants, o 15 salts, buffer systems and penetrationpromoters are introduced into this solution, c. into this solution at up to 80 0 C is introduced hydroxethyl cellulose or ~carboxymethyl cellulose, methyl cellulose, .o 20 hydroxypropylmethyl cellulose, polyvinyl alcohol, polyvinylpyrrolidone, polyacrylic acid or a mixture of any of these, in an amount of 0.1 to 2.0% by weight, based on finished product, and of up S 25 to 15% by weight in the case of polyvinyl St alcohols or polyvinylpyrrolidone, and d. a-interferon, in an amount of 1 x 10 to 1 x 10 IU/100 g of finished product Sis homogeneously dispersed therein, and this mixture is filled into sterile containers.

According to a further aspect, the invention provides a process for the preparation of pharmaceutical formulations for intra-vaginal administration, S wherein: 8b a. hydroxyethyl cellulose in an amount of 0.1 to 2.0% by weight is dissolved together with gelatine in an amount of up to 10% by weight, in a water/glycerol mixture at temperatures up to 60 0 C and b. at a temperature of 40 to 50 0 C, an a- IFN solution which contains polysorbate and hydrochloric acid is cautiously stirred in, and c. this solution is immediately poured into suitable moulds.

S; In the following Examples there are detailed descriptions 0 0 9 °of the preparation of pharmaceutical formulations oa 15 for topical administration which stabilise the #Oft 064: s aoo t 9 S -n i 9 a-IFN contained therein, the formulations being made according to the present invention mainly of cellulose derivatives and gelatine, anti-adhesion agents and the active substance a-IFN contained therein, and various additives. Obviously many modifications and alterations can be made, which will include the subject-matter of the invention, without departing from the scope of the invention, and the listed examples do not limit the invention.

In the Examples: Nipagin M Polysorbate 20 methyl p-hydroxybenzoate, Polyoxyethylene(20) sorbitan

R

monolaurate (Tween Polyoxyethylene(20) sorbitan monostearate (Tween Polyoxyethylene(20) sorbitan

R

monooleate (Tween 4 0 4 4 Polysorbate 60 Polysorbate 80

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0 4r I IL 44 4 44 4 44 4 Benzalkonium chloride Cetrimonium bromide 3.2 x 10 I.U.

mixture of alkyldimethylbenzylammonium chlorides with alkyl groups having 8 to 18 carbon atoms, hexadecyltrimethylammonium bromide, a 2 -Arg interferon is equivalent to 1 mg of this substance.

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10 Example 1 (Hydrogel) 100 g of gel suh tance contains: a2-IFN Nipagin M Gelatine, Bloom 160-200 NaC1 KC1 NaH 2

PO

4 2H 2 0

K

2

HPO

4 3H 2 0 Na glutamate H 2 0 Polysorbate 20 Hydroxyethyl cellulose (e.g.

NatrosolR 250 HX) Water, deionised ad 100 g 1 x 108 IU 0.20 g 0.50 g 0.48 g 0.012 g 0.0593 g 0.0407 g 0.0636 g 0.1 g 1.75 96.7944 The a 2 -IFN hydrogel is prepared by dissolving Nipagin M in water at 80°C and then cooling the solution to room temperature (RT) and, durinn the cooling period, introducing the gelatine into the solution.

Once the gelatine has completely dissolved, at RT NaCI, KC1, NaH 2

PO

4 2H 2 0, K 2

HPO

4 3H 2 0, Na glutamate H 2 0 and Polysorbate 20 are successively added to the solution. After subsequent sterilisation by filtration, microbiologically pure hydroxyethyl cellulose is sprinkled in and the solution is again heated to 80°C while stirring. After having been cooled to PT, and while stirring cautiously, an acidic 1% polysorbate 80 containing a 2 -IFN solution consisting of 0.01 N HC1 and lyophilised a 2

-IFN,

is added and homogeneously dispersed. This mixture is placed into sterile aluminium tubes under laminar air flow conditions.

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11 Example 2 (Hydrogel) 100 g of gel substance contains: a2-IFN Hydroxyethyl cellulosp (e.g.

Natrosol R 250 HX) Gelatine, Bloom 160-200 NaC1 Na2HPO 4 .2H 2 0

KH

2

PO

4 1,2-propylene glycol Polysorbate 80 Benzalkonium chloride 90% Water, deionised ad.

1 x 1010 IU 2.0 g 0.1 q 0.35 g 0.16 g 0.09 g 10.00 g 0.1 g 0.02 g 100 g It t I It 4

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I 14 It 'f 4 ii 444( Ills The a 2 -IFN hydrogel is prepared by dissolving benzalkonium chloride in water at 30'C, adding the propylene glycol at RT, and then introducing the gelatine into the solution. Once the gelatine has completely dissolved, at RT, NaCl, Na 2

HPO

4 .2H 2 0, KH 2

PO

4 and Polysorbate 80 are successively added. After sterilisation by filtration, microbiologically pure hydroxyetyyl cellulose is sprinkled in, and the solution is 25 heated at 80 0 C for 20 min with stirring- After having been cooled to RT, while stirring cautiously an acidic polysorbate a 2 -IFN solution, consisting of lyophilised a 2 -IFN dissolved in 0.01 N HC1 and 0.1% polysorbate 80, is placed and dispersed homogeneously. This mixture is placed into sterile aluminium tubes under laminar air flow conditions.

L .0 i 6: ~llllllL ~-W 12 Example 3 (Hydrogel) 100 g of gel substance contains: a 2

-IFN

Hydroxyethyl cellulose (e.g.

NatrosolR 250 HX) Gelatine, Bloom 160-200 Polysorbate 20 Glycerol Succinic acid Chlorhexidine digluconate 1N-NaOH (ad. pH Water, Oeionised ad.

1 x 106 IU 1.0 g 1.0 g 0.1 g 5.0 g 0.24 g 0.05 g 100.0 g '44 '4 o 4 I 4 e0 44 #4 4 4L 41 44 4..

'4tP 4,c~ The a 2 -IFN hydrogel is prepared by dissolving chlorhexidine gluconate, glycerol and gelatine at about 30*C. Succinic acid is then added, the pH is adjusted to 6.0 with IN NaOH, and polysorbate 20 is added.

20 The microbiologically pure hydroxyethyl cellulose is then sprinkled in, and the solution is heated at 80*C for 20 min, with stirring. After having been cooled to RT, with cautious stirring the acidic polysorbate a2-IFN solution is added and homogeneously dispersed. Filling is carried out as in Examples 1 and 2.

I, i «u 13 Example 4 (Hydrogel as nasal gel) 100 g of gel substance contains: 1 x 108 IU a -IFN Hydroxypropylmethyl cellulose Methocel R Premium) Polysorbate 60 Gelatine, Bloom 160-200 Sorbitol solution 70% Citric acid H 2 0 Na 2

HPO

4 2H 2 0 NaC1 Benzalkonium chloride 90% Water, deionised ad.

0.1 1 0.1 g 0.5 g 2.0 g 0.130g 0.30 g 0.5 g 0.02 g 100 g t 4

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The a 2 -IFN hydrogel is prepared by dissolving the hydroxypropylmethyl cellulose with water at with stirring. The solution is then cooled to 20 RT, and during the cooling period, gelatine is introduced. Once it has dissolved the following are introduced successively: benzalkonium chloride, sorbitol solution, citric acid, di-Na-hydrogen phosphate, NaC1 and Polysorbate 60. The prepared 25 gel is cautiously stirred at RT and the acidic 1 polysorbate containing a 2 -IFN solution is introduced and homogeneously dispersed. The product is placed into sterile aluminium tubes under laminar air flow conditions.

L I Am i L L -I 14 Example 5 (Hydrogel as ophthalmic gel) 100 g of gel substance contains: 8 a 2 -IFN 1 x 10 IU Polyacrylic acid (MW: ca. 3 million) 0.1 g Gelatine, Bloom 160-200 0.3 g Polysorbate 20 0.1 g Sorbitol solution 70% 2.0 g Glycerol 1.0 g NaC1 0.1 g Cetrimonium bromide 0.01 g IN NaOH (ad. pH 7.4) Water, deionised ad.100 g The a 2 -IFN hydrogel is prepared by dissolving the polyacrylic acid and gelatine in water at RT with 0. stirring, and then the sorbitol solution, glycerol, 20 NaC1, cetrimonium bromide and polysorbate are succesively introduced. Once dissolution is complete, the solution is sterilised by filtration and while stirring at RT, adjusted to a pH of about 7.6 with Ssodium hydroxide solution, during which the gel forms. Into this gel at RT, the a2-IFN solution which contains hydrochloric acid (0.001 N) and S1 polysorbate is introduced, stirring cautiously, 4i and homogeneously dispersed. The product is placed into sterile aluminium tubes under laminar air 30 flow as in the previous Examples.

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t att i i~i i~ L iL~ ll 1L- X _i e~l,~ 15 Example 6 (Vaginal suppositories) 100 g of vaginal suppository mass contains: a 2

-IFN

Hydroxyethyl cellulose Gelatine, Bloom 160-200 Na 2

HPO

4 2H 2 0

KH

2

PO

4 Polysorbate 20 Glycerol Water, deionised 1 x 10 IU 0.1 g 10.0 g 0.16 g 0.09 g 0.1 g 57.0 g ad. 100 g £4 I I 4 t4 t t tl 4 4 4f i 4D 411 I( I I 44 14 4 4 The a2-IFN vaginal suppositories are prepared by dissolving thz hydroxyethyl cellulose together with the gelatine in a mixture of glycerol and water which already contains Na 2 HPO4 2H 2 0, KH 2

PO

4 and polysorbate, at about 40-50 0 C. While still viscous (at about 37-40°C), the a 2 -IFN solution 20 which contains 1 polysorbate and hydrochloric acid (0.001N) is cautiously stirred in and the solution is immediately poured into suitable cooled moulds for solidification.

25 Example 7 (Stability tests, hydrogel without gelatine vs. with gelatine) 100 g of gel substance contains: a -IFN Nipagin M Gelatine, Bloom 160-200 NaC1 KC1 Na2HPO4 2H 2 0

KH

2

PO

4 3H 2 0 Polysorbate 20 Hydroxyethyl cellulose 1 x 108 IU 1 x 108 IU 0.2 g 0.2 g 0.5 g 0.48 g 0.48 g 0.012g 0.012g 0.086g 0.086g 0.012g 0.012g 0.1 g 0.1 g 2.0 g 2.0 g i_ _i i 4: 16 (e.g NatrosolR 250 HX) Water, deionised ad. 100 g Figures 1 to 3 show the stability of an a 2

-IFN

hydrogel of the above composition without the addition of gelatine (dotted lines) and with the addition of gelatine (unbroken lines), filled into aluminium tubes. Each point represents the mean of 6 determinations. The determinations were carried out by ELISA.

Figure 1 shows the stability on storage at Figure 2 shows stability on storage at and Figure 3 shows stability on storage at +21°C, in each case comparatively.

The stability behaviour was followed over a period of 14 weeks (31 months). It was found astonishingly Sand completely unexpectedly, that the differences It 20 in stability became more distinct as the storage temperature increased. On summation of the biological stability results over the entire period of 14 weeks and formation of an arithmetic mean of the values (n 11), the picture which emerges is that shown in the table which follows, in which a value of 10.0, corresponding to 108 I.U. a 2

-IFN

per 100 g, was set equal to 100% activity: .4 1 1 4 30 *4 t i, l-

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Storage Hydrogel without Hydrogel with Difference temper- addition of addition of ature gelatine gelatine -10 0 C 2 (11)=8.49 x (11)=9.72 12% 4 0 C x (11)=7.52 x (11)=9.97 +21 0 C (11)=5.67 i (11)=8.87 32% 17 The stabilising effect of the combination of hydroxyethyl cellulose and gelatine is clearly evident from this table. On storage in a refrigerator virtually no decrease in the activity of the formulation with the addition of gelatine was found after 31 months, whereas the formulation without the addition, of gelatine showed a decrease of about 25%. In this connection it is practicable to store a commercial product at refrigerator temperature. On storage at 21"C, the loss of activity of the formulation without the addition of gelatine was found to be almost one half the initial figure during the storage period of 31 months, whereas the formulation according to the invention with hydroxyethyl cellulose and gelatine showed a loss of activity of only 11% during this period, which is equivalent to a relative increase in stability of a 2 -IFN of more than Another storage test lasting 44 weeks (10 months) 41 20 of a formulation as described in Example 1, filled 1 into aluminium tubes with an inner protective lacquer (t and stored at temperatures of 4 0 C and 21*C, showed no loss of ELISA activity compared with the initial figure.

It 4i t

P

42 44244 Storage Hydrogel according to Example 1 temperature Initial Final figure figure after 10 months 4 0 C x 9.6 x 11.0 21 0 C x 9.6 x 9.6 _i i .i i 1 l-~ii-

Claims

1. Pharmaceutical formulations for topical adminis- tration which stabilise the a-interferon contained therein, comprising as carrier and stabiliser a mixture of 1. a cellulose derivative, polyvinyl alcohol, polyvinylpyrrolidone, polyacrylic acid polymer or a mixture of any of these, and

2. gelatine, which has been subjected to acid or alkaline digestion, and, in addition, an anti-adhesion agent. 2. Pharmaceutical formulations according to claim 1 in the form of a hydrogel. S3. Pharmaceutical formulations according to c claim 1 or 2 wherein said cellulose derivative 20 comprises hydroxyethyl cellulose. t 4. Pharmaceutical formulations according to claim 3 comprising hydroxyethyl cellulose and gelatine, each in an amount of 0.1 to 2.0% by weight, based on the finished product. 4 4

5. Pharmaceutical formulations according to claim 3, comprising hydroxyethyl cellulose and gelatine, the amount of gelatine being 0.1 to by weight, based on the finished product. 1

6. Pharmaceutical formulations according to any of claims 3-5, wherein the gelatine has a Bloom value of 160-200, and the hydroxyethyl cellulose has a degree of substitution of about i 19

7. Pharmaceutical formulations according to any of the preceding claims wherein said anti-adhesion agent comprises an anionic, cationic or non-ionic surfactant, preferably a polyethoxylated sorbitan ester.

8. Pharmaceutical formulations according to claim 7, wherein said anti-adhesion agent comprises the monolaurate, monostearate or monooleate of polyoxyethylene (20) sorbitan.

9. Pharmaceutical formulations according to any of the preceding claims, wherein the a-interferon contained therein has been obtained from a cell culture which has been transformed with a DNA sequence coding for human a-interferon. Pharmaceutical formulations according to Sclaim 9 wherein the a-interferon contained therein has been obtained from transformed cells of E. coli.

11. Pharmaceutical formulations according to claim 10 wherein the a-interferon contained therein 5 has a specific activity of 1 x 10 to 1 x 10 IU/100 g finished product.

12. Pharmaceutical formulations according to any of the preceding claims which include additional adjuvants selected from buffers, humectants, pre- servatives which are tolerated by skin, penetration promoters which are compatible with a-interferon and vehicles. 4414

13. Pharmaceutical formulations according to claim 12 wherein the buffer system is sodium phosphate buffer, potassium hydrogen phosphate/sodium dihydrogen phosphate buffer, citric acid/phosphate buffer, i ~I I LIILIL L-- 20 potassium-sodium-glutamate-phosphate buffer, succinate buffer or a mixture of any of these.

14. Pharmaceutical formulations according to claim 13 wherein said buffer has a pH in solution in the range of 6.0 to 7.4. Pharmaceutical formulations according to claim 12 wherein said humectant comprises glycerol, 1,2-propylene glycol, sorbitol, butylene glycol, a polyol such as polyethylene glycol 400 or a mixture of any of these.

16. Pharmaceutical formulations according to claim 12 wherein said preservative tolerated by skin comprises, p-hydroxybenzoic esters, sorbic acid, benzoic acid, chlorhexidine digluconate, benzalkonium chloride, hexadecyltrimethylammonium o, bromide or a mixture of any of these. O

17. Pharmaceutical formulations according to claim 12 wherein said penetration promoter which is compatible with a-interferon is dimethyl sulphoxide in a proportion of up to 50% by weight.

18. Pharmaceutical formulations according to claim 12 wherein said vehicle used to form a gel *is water. 30 19. Pharmaceutical formulations according to claim 1, substantially as described herein. o. 20. Pharmaceutical formulations according to claim 1, substantially as described herein with reference to any one of the Examples. 21

21. A method of treatment for Herpes labialis, Herpes genitalis including papilloma infections, warts and for Herpes zoster infections; and for the topical treatment of paracanceroses and canceroses; comprising the administration of pharmaceutical formulations according to any of claims 1 to 20 as dermal, nasal, ophthalmic and intra-vaginal viralstatics.

22. A process for the preparation of a pharmaceutical formulation according to any of claims 1 to which comprises admixing the respective components under conditions such that said a-interferon is I e' stabilised. 15 23. Process for the preparation of pharmaceutical formulations for topical use which stabilise the a-interferon contained therein, wherein: a. gelatine in an amount of 0.1 to by weight, based on the finished product, 20 is dissolved in water at up to 80 0 C, and an anti-adhesion agent is added to the solution, Sb. optionally preservatives, humectants, salts, buffer systems and penetration- promoters are introduced into this solution, c. into this solution at up to 80 0 C is introduced hydroxethyl cellulose or carboxymethyl cellulose, methyl cellulose, hydroxypropylmethyl cellulose, polyvinyl 30 alcohol, polyvinylpyrrolidone, polyacrylic acid or a mixture of any of these, in an amount of 0.1 to 2.0% by weight, based on finished product, and of up to 15% by weight in the case of polyvinyl alcohols or polyvinylpyrrolidone, and d. a-interferon, in an amount of 1 x 105 10 to 1 x 10 1 0 IU/100 g of finished product V I ~1 22 is homogeneously dispersed therein, and this mixture is filled into sterile containers.

24. Process for the preparation of pharmaceutical formulations for intra-vaginal administration, wherein: a. hydroxyethyl cellulose in an amount of 0.1 to 2.0% by weight is dissolved together with gelatine in an amount of up to 10% by weight, in a water/glycerol o mixture at temperatures up to 60 0 C and b. at a temperature of 40 to 50 0 C, an a- o IFN solution which contains polysorbate o. 15 and hydrochloric acid is cautiously 0 stirred in, and c. this solution is immediately poured into suitable moulds. 44" Dated this 23rd day of April, 1990 0 4 DR. KARL THOMAE GMBH 004 By its Patent Attorneys DAVIES COLLISON 0 0A -Us r