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WO1994019001A1 - Micro-emulsions comprenant des peptides therapeutiques - Google Patents

Micro-emulsions comprenant des peptides therapeutiques Download PDF

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Publication number
WO1994019001A1
WO1994019001A1 PCT/US1994/001678 US9401678W WO9419001A1 WO 1994019001 A1 WO1994019001 A1 WO 1994019001A1 US 9401678 W US9401678 W US 9401678W WO 9419001 A1 WO9419001 A1 WO 9419001A1
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WO
WIPO (PCT)
Prior art keywords
medium
fatty acid
microemulsion
hlb surfactant
die
Prior art date
Application number
PCT/US1994/001678
Other languages
English (en)
Inventor
Panayiotis Pericleous Constantinides
Original Assignee
Smithkline Beecham Corporation
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Smithkline Beecham Corporation filed Critical Smithkline Beecham Corporation
Priority to JP6519110A priority Critical patent/JPH08507066A/ja
Priority to EP94909654A priority patent/EP0684833A4/fr
Publication of WO1994019001A1 publication Critical patent/WO1994019001A1/fr

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
    • A61K9/1075—Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/25—Growth hormone-releasing factor [GH-RF], i.e. somatoliberin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/39—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin, cold insoluble globulin [CIG]

Definitions

  • This invention relates to pharmaceutically acceptable water-in-oil (w/o) self-emulsifying microemulsions containing therapeutic agents, processes for their preparation and their use.
  • Microemulsions can be defined in general as thermodynamically stable, isotropically clear dispersions of two immiscible liquids stabilized by interfacial films of surface- active molecules.
  • the formation of microemulsions usually involves a combination of three to five components, namely, an oil, water, a surfactant, a cosurfactant and an electrolyte.
  • the tendency towards a water-in-oil (w/o) or an oil-in-water (o/w) microemulsion is dependent on the properties of the oil and the surfactant.
  • Surfactants are conveniently classified on an empirical scale known as the hydrophilic-lipophilic balance (HLB) which runs from 1 to about 45.
  • HLB hydrophilic-lipophilic balance
  • microemulsions are formed using surfactants (or emulsifiers) which have an HLB value in the range of about 3 to 6 while (o/w) microemulsions are formed using surfactants which have an HLB value in the range of about 8 to 18.
  • the surfactant should preferably exhibit low solubility in both the oil and water phases, and be preferentially absorbed at the water-oil interface with concomitant lowering of interfacial tension. When interfacial tension is less than 2 x 10" 2 dyn/cm, a stable microemulsion can form.
  • Microemulsions are typically substantially non-opaque, i.e. are transparent or opalescent when viewed by optical microscopic means. In the undisturbed state, they are optically isotropic (non-birefringent) when examined under polarized light.
  • the dispersed phase typically comprises particles or droplets which are normally between 5 and 200 nm in size and this gives rise to their optical transparency. These particles may be spherical although other structures are feasible.
  • the role of the cosurfactant is to increase the interfacial fluidity by penetrating the surfactant film and consequently creating a disordered film due to the void space among surfactant molecules.
  • the co-surfactant is usually a short-chain alcohol.
  • the use of a cosurfactant in microemulsions is, however, optional and alcohol-free self-emulsifying emulsions and microemulsions have been described in the literature (see for instance, Pouton et al., Int. Journal of Pharmaceutics, 27, 335-348, 1985 and Osborne et al., J. Disp. Sci. Tech., 9, 415-423, 1988).
  • Microemulsions form spontaneously, without the need for a high input of energy and are therefore easy to prepare and scale up for commercial applications; they have thermodynamic stability due to their small particle size and therefore have a long shelf life; they have an isotropically clear appearance so that they may be monitored by spectroscopic means; they have a relatively low viscosity and are therefore easy to transport and mix; they have a large interfacial area which accelerates surface reactions; they have a low interfacial tension which permits flexible and high penetrating power and, lastly, they offer the possibility of improved drug solubiUzation and protection against enzymatic hydrolysis.
  • microemulsions may undergo phase inversion upon addition of an excess of the dispersed phase or in response to a temperature change and this is a property of these systems that can affect drug release from microemulsions both in vitro and in vivo. The reasons for this improved drug delivery are not however well understood.
  • GB 2 222770-A (Sandoz Ltd) describes microemulsions and corresponding microemulsion "pre-concentrates" for use with the highly hydrophobic cyclosporin peptides.
  • a suitable pre-concentrate comprises 1,2- propylene glycol as the hydrophilic component, a caprylic-capric acid triglyceride as the lipophilic component and a mixture of a polyoxyethylene glycolated hydrogenated castor oil and glycerin monooleate (ratio 11:1) as the surfactant-cosurfactant.
  • Such formulations may then be diluted with water but to give an oil-in-water rather than a water-in-oil microemulsion.
  • GB 2098 865A (Sandoz Ltd) describes topical compositions in the form of microemulsions comprising a water-immiscible organic solvent, an emulsifier, a co- emulsifier, water and a (non-peptide) therapeutic agent. These formulations are said to have improved skin penetrating properties.
  • Suitable organic solvents include (C 1 0-22)- fatty acid esters of (C3-i8)-alcohols such as hexyl laurate, (Ci2-32)-hydrocarbons such as squalene and mono- or diesters of glycerol with a (C6-22) carboxylic acid such as glyceryl caprylate (which may also act as a co-emulsifier).
  • C3-i8)-alcohols such as hexyl laurate
  • (Ci2-32)-hydrocarbons such as squalene and mono- or diesters of glycerol with a (C6-22) carboxylic acid such as glyceryl caprylate (which may also act as a co-emulsifier).
  • C6-22 carboxylic acid
  • glyceryl caprylate which may also act as a co-emulsifier
  • US 4712239 (Muller et al.) describes multi-component systems for pharmaceutical use comprising an oil, a nonionic surfactant with an HLB value above 8 and a co-surfactant which is a partial ether or ester of a pol hydroxyl alcohol and a (C ⁇ -22) fatty alcohol or acid, which components form a "single phase" on mixing.
  • the special properties of the system are attributed to the particular blend of surfactant and co ⁇ surfactant selected.
  • An aqueous phase is an optional extra and the therapeutic agent may be lipophilic or hydrophilic.
  • Such systems are said to give enhanced transdermal delivery characteristics.
  • one (example 1, formulation I) has PEG 20 Ethylene Oxide (20-EO)-oleic acid glycerol partial esters (40%), caprylic-capric acid glycerol partial esters (42% monoglyceride content, 24%), medium-chain triglycerides (16%) and water (20%). It is to be noted that the ratio of the medium-chain triglyceride to the caprylic-capric acid glycerol partial esters is 1:1.5. In comparison example 1, this is formulated with the drug arecaidine /z-propyl ester HC1 salt.
  • a further example incorporates a macromolecule, the polypeptide hirudin but in this, the oil is wo-propyl palmitate.
  • DSS Dioctyl sodium sulfosuccinate
  • Docusate Sodium or Aerosol OT is widely used as a solubilizing, wetting, emulsifying or dispersing agent.
  • Known pharmaceutical uses of DSS include: a) as a therapeutic agent, alone or as an adjuvant in the prevention or treatment of constipation, b) as a tablet formulation adjuvant to facilitate tablet coating and improve tablet disintegration and dissolution characteristics, and c) as an absorption enhancer.
  • DSS has been reported to increase the small intestinal absorption of heparin (Engel, R.H. et ah; J. Pharm. Sci. 58, 706-710, 1969), insulin (Dupont, A.
  • EP-387647 (Matouschek, R.) describes pharmaceutical microemulsion compositions containing acidic or basic drug and compound forming ion-pairs for nasal, rectal or transdermal delivery.
  • Suitable oils included isopropyl myristate, 2-octyl dodecanol and paraffin oil with the surfactant comprising polyoxyethylene fatty acid ester and/or polyoxyethylene fatty alcohol ether.
  • the cosurfactant used was polyoxyethylene glycerol fatty acid ester with water being the aqueous phase.
  • the compound forming an ion-pair was sulphate.
  • compositions which have improved solubility, stability and/or particle size for the oral delivery of therapeutic agents, in particular for peptides, is still needed.
  • the present invention provides a pharmaceutically acceptable, stable, self- emulsifying microemulsion comprising:
  • a lipophilic phase having a medium-chain fatty acid triglyceride and a low HLB surfactant and in which the ratio of the medium-chain fatty acid triglyceride to the low HLB surfactant is from 5:1 to 1.5:1;
  • Figure 1 illustrates a pseudo-ternary phase diagram reading of a microemulsion system containing an oil and a low HLB surfactant in a fixed ratio X, a high HLB surfactant and an aqueous phase.
  • Figure 2 illustrates a pseudo-ternary phase diagram of the microemulsion system
  • CAPTEX8000/CAPMUL C8 as the oil/low HLB surfactant in a ratio of 2 to 1, labeled as component (1); DSS/PEG 400 (1:1) as the high HLB surfactant, labeled as component
  • component (2) water as the aqueous phase, labeled as component (2).
  • the present invention provides for a stable, self-emulsifying water-in-oil (w/o) comprising: (a) a lipophilic phase having a medium-chain fatty acid triglyceride and a low HLB surfactant and in which the ratio of the medium-chain fatty acid triglyceride to the low HLB surfactant is from 5:1 to 1.5:1; (b) an admixture of a polyol and at least one high HLB surfactant which is i) a sulfate or pharmaceutically acceptable salt thereof, which is an aliphatic sulfate, an aryl sulfate, an aliphatic-aryl sulfate, or mixtures thereof; ii) a sulfonate or pharmaceutically acceptable salt thereof, which is an aliphatic sulfonate, or
  • the present invention provides for a pharmaceutically acceptable, stable, self -emulsifying microemulsion comprising each of (a), (b) and (c) above and (d) a water-soluble biologically active agent.
  • microemulsion of an aliphatic, preferably a medium chain alkyl or dialkyl, sulfate or sulfonates or sulfosuccinate is believed to enhance the absorption of a biologically active agent when administered in the above formulations, particularly in reference to field A in Figure 2.
  • the pharmaceutically acceptable, stable, self- emulsifying microemulsion comprises: (a) a lipophilic phase having i) a medium-chain fatty acid triglyceride of 50 to 100% (w/w) of caprylic acid and from 0 to 50% (w/w) of capric acid triglycerides; and ii) a low HLB surfactant which is a medium-chain fatty acid monoglyceride or diglyceride or a mixture thereof, optionally comprising a small amount by weight of free medium-chain fatty acid; and in which the ratio of the medium-chain fatty acid triglyceride to the low HLB surfactant is from 5:1 to 1.5:1; (b) an admixture of a polyol and at least one high HLB surfactant which is i) a medium chain alkyl or dialkyl sulfate, an aryl sulfate, or a medium chain alkyl-aryl s
  • the high HLB surfactant is a polyoxyethylene fatty acid ester, polyoxyetheylene-sorbitan fatty acid ester, polyethylene glycol long-chain alkyl ether or a polyethylene glycol long-chain alkyl ester.
  • Suitable medium-chain fatty acid triglycerides for use in the present invention may be of natural, semi-synthetic or synthetic origin and may include blends of different medium chain fatty acid triglycerides, and/or medium and long fatty acid triglycerides. Such blends include not only physical blends of medium- and long-chain fatty acyl triglycerides but also triglycerides which have been chemically modified, by, for instance interesterification, to include a mixture of medium- and long-chain fatty acyl moieties. Suitable such triglycerides and diesters are readily available from commercial suppliers.
  • the fatty acid composition comprises caprylic (Cs) acid optionally admixed with capric (Cio) acid, for instance from 50 to 100% (w/w) of caprylic acid and from 0 to 50% (w/w) of capric acid triglycerides.
  • Suitable examples include tiiose available under the trade names MYRTTOL; CAPTEX (Karlshams Lipid Specialties, Columbus OH), for instance CAPTEX 300, CAPTEX 350, CAPTEX 355, CAPTEX 850 and CAPTEX 8000; MIGLYOL (BASF), for instance the grades MIGLYOL 808, MIGLYOL 810, MIGLYOL 812 and MIGLYOL 818 (which also comprises a linoleic acid triglyceride) and MAZOL 1400 (Mazer Chemical, Gumee, ⁇ .).
  • MYRTTOL trade names
  • CAPTEX Kerlshams Lipid Specialties, Columbus OH
  • MIGLYOL BASF
  • MIGLYOL 810, MIGLYOL 812 and MIGLYOL 818 which also comprises a linoleic acid triglyceride
  • MAZOL 1400 Mel Chemical, Gumee, ⁇ .
  • Suitable long-chain fatty acid triglycerides may be conveniendy obtained from neutral plant, vegetable and fish oils such as shark oil, coconut oil, palm oil, olive oil, sesame oil, peanut oil, castor oil, safflower oil, sunflower oil and soybean oil, which oils may be in their natural state or partially or fully hydrogenated.
  • Soybean oil consists of oleic acid (25%), linoleic acid (54%), linolenic acid (6%), palmitic acid (11%) and stearic acid (4% triglycerides while safflower oil consists of oleic acid (13%), linoleic acid (76%), stearic acid (4%) and palmitic acid (5%) triglycerides.
  • die major fatty acid components are Cis-saturated, monounsaturated or polyunsaturated fatty acids, preferably Cis-monounsaturated or polyunsaturated fatty acids, such as oleic, linoleic and liolenic acid.
  • Cis-saturated, monounsaturated or polyunsaturated fatty acids preferably Cis-monounsaturated or polyunsaturated fatty acids, such as oleic, linoleic and liolenic acid.
  • Other suitable triglycerides include interesterified triglycerides which may be derived synthetically by chemically reacting blends of medium- and long-chain triglycerides, for instance triglycerides containing caprylic and capric acid moieties and vegetable oils rich in oleic or linoleic acids.
  • Suitable such interesterified triglycerides include d e products available from Karlshams Lipid Specialties, as CAPTEX 810A - D and 910A - D, which typically contain from 30 to 80% capric and caprylic acids, 10 to 50% linoleic acid (810 series) or 10 to 60% oleic acid plus up to 5% linoleic acid (910 series), and up to 25% of oti er acids.
  • intermediate-chain such as in “medium-chain fatty acid”
  • fatty acid saturated, mono-unsaturated or poly-unsaturated, having from 6 to 12, preferably from 8 to 10 carbon atoms, which may be branched or unbranched, preferably unbranched, and in which fatty acyl chain may be optionally substituted.
  • Optional substituents include for instance, halogen, hydroxy, alkoxy, thioalkyl or halo substituted alkyl.
  • Halogen as used herein includes F, Cl, Br and I.
  • long-chain such as in “long-chain fatty acid”, and as used herein refers to a carbon chain, or a fatty acid which may be saturated, mono-unsaturated or poly ⁇ unsaturated, having from 14 to 22, preferably 14 to 18, carbon atoms which may be branched or unbranched, preferably unbranched, and which may be optionally substituted as noted above for medium chain fatty acids.
  • a non-ionic high HLB surfactant may be usefully included as an auxiliary high HLB surfactant to produce microemulsions which can solubilize a larger amount of aqueous phase.
  • Such microemulsions are, however, generally more viscous than those in which the non-ionic high HLB surfactant is absent.
  • Suitable low HLB surfactants for use in the present invention include fatty acyl monoglycerides, fatty acyl diglycerides, sorbitan medium or long-chain fatty acyl esters and medium-chain free fatty acids, as well as mixtures thereof.
  • Suitable mono- and di ⁇ glycerides may each include blends of different fatty acyl mono- and di-glycerides and the fatty acid moieties may be medium- or long-chain or a mixture thereof.
  • Suitable medium chain fatty acyl mono- and di-glycerides are formed from caprylic and capric acids. Suitable blends comprise from about 50 to 100% caprylic acid and from 0 to 50% capric acid. Mixtures of mono-and di-glycerides preferably comprise at least 50, more preferably at least 70% by weight of monoglycerides.
  • Suitable commercial sources of these include die products available under the trade name CAPMUL (Karlsham Lipid Specialties), for instance the products CAPMUL MCM which comprises monoglycerides (77%), diglycerides (21%) and free glycerol (1.6%), with a fatty acid composition which comprises caproic acid (3%), caprylic acid (67%) and capric acid (30%) and CAPMUL C8 which has monoglycerides (70 - 90%), diglycerides (10 - 3 ⁇ %) and free glycerol (2 - 4%), with a fatty acid composition which comprises at least 98% caprylic acid
  • the low HLB surfactant contains a mixture of mono- and di-glycerides having at least about 80% by weight, preferably at least about 90% by weight, and more preferably at least about 95% by weight of a caproic, caprylic, capric monoglyceride or mixtures thereof, preferably a caprylic, capric monoglyceride or mixtures thereof.
  • these surfactants include Imwitor 308TM (Huls America, Inc.) which has about 80-90% wt. caprylic monoglycerides; and Glycerol Monocaprylin, manufactured as 1-monoctanoyl-rac- glycerol (Sigma Chemicals) having about 99% wt. caprylic monoglycerides; and Glycerol Monocaprate, manufactured as 1-monodecanoyl-rac-glycerol (Sigma Chemicals) having about 99% wt. capric monoglycerides.
  • Suitable long-chain fatty acyl monoglycerides include glycerol monooleate, glycerol monopalmitate and glycerol monostearate.
  • Suitable commercially available examples of such include the products available under die trade names MYVEROL, such as MYVEROL 18-92 (a sunflower oil monoglyceride) and 18-99 (a rapeseed oil monoglyceride), MYVATEX and MYVAPLEX, respectively, from Eastman Kodak Chemicals, Rochester, New York.
  • a further useful long-chain fatty acyl monoglyceride- containing product is ARLACEL 186 (available from ICI Americas Inc.) which includes, in addition to glycerol monooleate, propylene glycol (10%).
  • the main fatty acids of MYVEROL 18-92 are oleic acid (19%), linoleic acid (68%) and palmitic acid (7%) while those of MYVEROL 18-99 are oleic acid (61%), linoleic acid (21%), linolenic acid (9%) and palmitic acid (4%).
  • the major fatty acid component is a Ci8-saturated, monounsaturated or polyunsaturated fatty acid, preferably a Ci 8 -monounsaturated or polyunsaturated fatty acid.
  • diacetylated and disuccinylated versions of the monoglycerides such as the product available under die trade name MYVATEX SMG are also useful.
  • Suitable sorbitan long-chain esters for use in the present invention include sorbitan monooleate, available commercially under die trade names SPAN 80 and ARLACEL 80 and sorbitan sesquioleate, available commercially under die trade names SPAN 83 and ARLACEL 83.
  • Suitable sorbitan medium-chain esters for use in the present invention include sorbitan caprylate, sorbitan caprate, sorbitan laurate.
  • Suitable medium-chain fatty acids for use in die present invention include caprylic and capric acids and mixtures thereof.
  • the low HLB surfactant will have an HLB value in the range of about 2 to 8.
  • the HLB values of the products CAPMUL MCM, MYVEROL 18-99, ARLACEL 80, ARLACEL 83 and ARLACEL 186 are respectively about 5.5 to 6, 3.7, 4.3, 3.7 and 2.8.
  • the estimated HLB of 1-monocaprylin is about 8.0.
  • microemulsions comprise maxim - chain fatty acyl components, such as those derived from caprylic and capric acids, especially ti ose derived from caprylic acid.
  • preferred microemulsions include blends of CAPTEX 355, 810, CAPTEX 8000, particularly CAPTEX 8000; CAPMUL MCM or CAPMUL C8, particularly CAPMUL C ⁇ -
  • a self-emulsifying (w/o) microemulsion according to die present invention of a low HLB surfactant which is a medium-chain fatty acid monoglyceride and/or a medium- chain fatty acid diglyceride as hereinbefore defined and which is a component of the lipophilic phase provides for reduced droplet size and this is believed to aid in d e absorption of d e therapeutic agent.
  • the present invention provides for a microemulsion in which the medium-chain fatty acid triglyceride is a caprylic acid or a blend of caprylic and capric acids triglycerides as hereinbefore defined and in which the low HLB surfactant is a medium-chain fatty acid monoglyceride or diglyceride or a mixture thereof in which the medium-chain fatty acid is caprylic acid or a mixture of caprylic and capric acids as hereinbefore defined, optionally admixed witi a small amount of medium-chain fatty acid, in particular, a blend of caprylic acid triglyceride and caprylic acid monoglyceride or diglyceride or mixture thereof.
  • the high HLB surfactant for use in the present invention is an aliphatic, aryl or an aliphatic-aryl sulfate, sulfonate or sulfosuccinate.
  • the aliphatic moiety is a medium or long chain alkyl or dialkyl group.
  • the medium chain moiety contains C o 12. preferably 8 to 10 carbons, and die long chain moiety contains from C12 to 24 » preferably 14 to 18 carbons as previously defined.
  • the aliphatic chain may be branched or unbranched and may be optionally substituted and may be saturated, mono-unsaturate or poly-unsaturated.
  • the aliphatic moiety is of medium chain length.
  • Theie are two forms of linkages possible with the sulfate, sulfonate and sulfosuccinate groups herein, one is designated as a fatty acyl ester linkage and die other an ester linkage. If the aliphatic or aryl moiety forms an fatty acyl ester linkage with d e sulfate, sulfonate or sulfosuccinate, i.e. contains an additional carbonyl [C(O)] group, the aliphatic (or aryl) group becomes known as the fatty acid portion of the ester linkage.
  • a fatty acyl ester linkage is for instance, a C -12-C (O) - O S(O)3Na+ .
  • This is in contras to the generally recognized linkage of C6-12 C-O - S(O)3Na+, referred to herein as the ester linkage.
  • This terminology stems from the reaction of a C -12 OH moiety with the sulfonic acid O-S(O)2 " Na + group forming a sulfonic ester.
  • the linkage is an ester linkage and not a fatty acyl ester linkage. More preferably the linking group is a medium chain alkyl or dialkyl moiety.
  • aryl sulfate, sulfonate or sulfosuccinate means a phenyl or naphdiyl moiety.
  • Suitable aryl sulfonates include, for instance, benzene sulfonate.
  • the sulfates, sulfonates and sulfo- succinates tiiey may be mixtures of bom aliphatic and aryl linkages formed, such as do- decyl benzene sulfate.
  • This combination as used herein is referred to as "aliphatic-aryl”.
  • the aliphatic moiety in tiiis instance is a medium chain alkyl or dialkyl group.
  • Suitable long chain alkyl or dialkyl sulfates, sulfonates and sulfosuccinates include, but are not limited to myristic (C14), palmitic (C16), palmitoleic (C16:l), stearic (C18), oleic (C18:l), vaceenic (trans oleic acid) or linoleic (C18:2) .
  • the alkyl and dialkyl moieties may be mixtures of different medium and long chain moieties, such as a dialkyl sulfate having a C12 and a C16 group.
  • the sulfates, sulfonates and sulfosuccinates are medium chain alkyl or dialkyl derivatives.
  • Suitable medium chain moieties include, but are not limited to octyl (Cs), decyl (Cio), dodecyl (C12), iso-octoanoic, or di-octyl, di-decyl, or di-dodecyl.
  • the medium chain moiety is octyl, decyl, dodecyl, or dioctyl.
  • the high HLB is a octyl, decyl, dodecyl or di-octyl sulfosuccinate, or is a octyl, decyl, or dodecyl sulfate.
  • Yet another aspect of the instant invention is the combination of mixtures of the different aliphatic, aryl or aliphatic-aryl sulfates, sulfonates and sulfosuccinates as the high HLB surfactant used herein.
  • the sulfates, sulfonates, or sulfosuccinates will be pharmaceutically acceptable water-soluble salts, for instance alkali metal salts, such as sodium and potassium salts, or ammonium or quaternary ammonium salts also referred to as N(R)4 wherein R is an alkyl derivative, or is a primary and secondary (protonated) amine salt, such as etiianolamine or triethanolamine.
  • the salts are the alkali metal salts.
  • die salts are of the medium chain alkyl or dialkyl sulfates, such as octyl, decyl or dodecyl sulfate; or die salts of dialkyl sulfosuccinate, of which die salts sodium dioctyl sulfosuccinate and sodium dodecyl sulfate are preferred.
  • Sodium dioctyl sulfosuccinate (DSS) and sodium dodecyl sulfate (SDS) have estimated HLB values of 41 and 40 respectively.
  • die term "polyol” is a polyhydric alcohol, i.e. containing two or more hydroxyl groups. Such as but not limited to, ethylene glycol, propylene glycol or polyethylene glycol (PEG). PEG is also referred to as a polyglycol witii eti ylene glycol as a polymerized unit. Preferably, the polyol has tiiree or more alcoholic units and is glycerol or a PEG. More preferably, the polyol is a PEG having a molecular weight of 400 (PEG-400). Other suitable polyhydric alcohol's for use herein include, but not limited to, etiiylene glycol (2-OH units), sorbitol (6-OH), and mannitol (6-OH).
  • the ratio of d e sulfates, sulfonates, or sulfosuccinates to the polyol in die admixture is from about 10:1 to 1:1, preferably about 5:1 to 1:1, more preferably from 5:1 to 2:1, and most preferably about 1:1.
  • DSS Commercially available DSS products, includes .[all in USP grades] DSS (100%), DSS 85% surfactant (with 15% sodium benzoate) or DSS (50%) in PEG 400.
  • die admixture for use herein is a polyol with at least one high HLB surfactant which is a sulfate, or a sulfonate, or a sulfosuccinate or pharmaceutically acceptable salts thereof.
  • the sulfate moiety, or a pharmaceutically acceptable salt thereof may be an aliphatic sulfate, an aryl sulfate, an aliphatic-aryl sulfate, or a mixture thereof.
  • the sulfonate or pharmaceutically acceptable salt thereof may be an aliphatic sulfonate, aryl sulfonate, an aliphatic-aryl sulfonate, or a mixture thereof.
  • the sulfosuccinate or pharmaceutically acceptable salt thereof may be an aliphatic sulfosuccinate, an aryl sulfosuccinate, an aliphatic-aryl sulfosuccinate, or a mixture thereof.
  • the high HLB surfactant may be a mixture of any of the above noted sulfates, sulfonates or sulfosuccinates and tiieir pharmaceutically acceptable salts thereof. Such a mixture could be referred to as binary or ternary mixture of the sulfate groups, the sulfonate groups or sulfosuccinate groups.
  • the preferred high HLB is a medium chain alkyl or dialkyl sulfate, sulfonate or sulfosuccinate or pharmaceutically acceptable salts thereof.
  • the total amount of high HLB surfactant which is a sulfate, sulfonate and sulfosuccinate should comprise at least 50% of die total amount, by weight, of the high HLB surfactants required to be present in the formulation, and more preferably closer to a 100%. It is recognized tiiat other high HLB surfactants may be present in the formulation and are further described herein.
  • the total amount of high HLB surfactant present shoul range from about 5% to about 30% and from about 70% to less tiian 100% (w/w). More preferably the total amount of high HLB surfactant should range from 5% to about 25% (w/w), and from 80% to less dian 100% w/w.
  • any additional non-ionic, ionic and zwitterionic surfactants should comprise no more than 50% of the total high HLB surfacta present in the formulation, preferably less.
  • die high HLB surfactant may contain additional excipients or co-surfactants and include, but are not limited to the 1) non-ionic surfactants, such as
  • polyoxyethylene fatty acid esters for example polyoxyethylene stearic acid esters of the type available under die trade name MYRJ (ICI Americas, Inc.), for instance the product MYRJ 52 (a polyoxyethylene 40 stearate);
  • polyoxyetheylene-sorbitan fatty acid esters for example the mono- and tri-lauryl, palmityl, stearyl and oleyl esters, for instance the polyoxyed ylene sorbitan monooleates available under die trade name of TWEEN (ICI Americas Inc.), such as TWEEN 20, 21, 40, 60, 61, 65, 80, 81 and 85, of which class TWEEN 80 is especially preferred;
  • PEG glycerol etiiers such as the polyeti ylene glycol long-chain alkyl etiiers, which include polyetiiylated glycol lauryl ether; and PEG fatty alcohol ethers; and
  • PEG glycerol esters and PEG fatty acid esters such as die long-chain alkyl esters, which include PEG-monostearate.
  • the surfactant system may contain additional surfactants and include, but not limted to: 1) cationic surfactants, such as cetyl ammonium bromide (CTAB) or benzalkonium bromide;
  • CTAB cetyl ammonium bromide
  • benzalkonium bromide cationic surfactants, such as cetyl ammonium bromide (CTAB) or benzalkonium bromide
  • anionic surfactants such as bile salts and d e alkali metal salts tiiereof, including but not limited to cholate, deoxycholate, taurocholate, etc., sodium taurocholate and Co- 18 fatty acyl carnitines; or
  • low HLB surfactants such as other lipids, i.e., phospholipids which may be anionic, cationic or zwitterionic, in particular lecithins, such as soya bean lecitiiins, egg lecithin or egg phosphatide, cholesterol or long-chain fatty acids such as oleic acid.
  • lecithins such as soya bean lecitiiins, egg lecithin or egg phosphatide, cholesterol or long-chain fatty acids such as oleic acid.
  • die term "therapeutic agent” or “biologically active agent” refers to any compound which has biological activity, is soluble in d e hydrophilic phase and has an HLB value of at least that of die high HLB surfactant used in the formulation, to ensure that the drug is preferentially dissolved in the hydrophilic ratiier than the lipophilic phase.
  • drug biologically active agent
  • Suitable peptides include not only small peptides but also larger peptides/polypeptides and proteins. Suitable such peptides preferably have a molecular weight from about 100 to 10,000, more preferably from about 100 to about 6,000.
  • peptides having from 2 to 35 amino acid moieties are especially preferred.
  • Suitable small peptides have from about 2 to about 10, more preferably from about 2 to about 6 amino acid moieties.
  • Preferred small peptides include die RGD containing peptides, such as die fibrinogen receptor antagonists which are tetrapeptides having an average molecular weight of about 600. These peptide antagonists are highly potent platelet aggregation inhibitors at plasma levels as low as 1 pmol/ml.
  • Preferred fibrinogen antagonists include d e peptide cyclo(S,S)-N a -acetyl-Cys-(N a -methyl)Arg-Gly-Asp-Pen- NH 2 (Ali etal, EP 0 341 915, whose disclosure is herein incorporated by reference in its entirety) and die peptide cyclo(S,S)-(2-mercapto)benzoyl-(N a -methyl)Arg-Gly-Asp-(2- mercapto)phenylamide (EP 0423 212, whose disclosure is herein incorporated by reference in its entirety).
  • fibrinogen antagonists useful in the present invention are tiiose peptides disclosed by Pierschbacher et al, WO 89/05150 (US/88/04403); Marguerie, EP 0275 748; Adams et al, U.S. 4,857,508; Zimmerman et al, U.S.
  • the RGD peptide may be usefully included in the microemulsion formulation in an amount up to about 600mg/g of the hydrophilic phase or from 0.1 to 60 mg/g of the formulation.
  • peptides useful in die present invention include, but are not limited to, other RGD containing peptides such as tiiose disclosed by Momany, U.S. 4,411,890 and U.S. 4,410,513; Bowers et al, U.S. 4,880,778, U.S. 4,880,777, U.S. 4,839,344; and WO 89/10933 (PCT/US89/01829); the peptide Ala-His-D-Nal-Ala-Trp-D-Phe-Lys-NH 2 (in which Nal represents ⁇ -naphthylalanine) and die peptides disclosed by Momany, U.S.
  • Otiier suitable peptides include hexapeptides such as d e growth hormone releasing peptide (GHRP) His-D-Trp-Ala-Trp-D-Phe-Lys-NH 2 , (Momany, US 4,411,890, the disclpsure of which is herein incorporated by reference in its entirety), and analogs or homologs diereof. This may usefully be included in an amount up to about 250mg/g of the hydrophilic phase or from 0.1 to 25mg/kg of the formulation.
  • GHRP growth hormone releasing peptide
  • Suitable larger polypeptides and proteins for use in microemulsions of die present invention include insulin, calcitonin, elcatonin, calcitonin-gene related peptide and porcine somatostatin as well as analogs and homologs diereof.
  • Otiier suitable larger polypeptides include tiiose disclosed by Pierschbacher et al, U.S. 4,589,881 (>30 residues); Bittie et al, U.S. 4,544,500 (20-30 residues); and Dimarchi et al, EP 0204 480 (>34 residues).
  • Otiier type of compounds useful in die present invention include analogs or homologs of LHRH which display potent LH releasing activity or inhibit the activity of LHRH; analogs or homologs of HP5 (hemopoietic factor 5) which possesses hematopoetic activity; analogs or homologs of endothelin, some of which possess hypotensive activity amonst other activities; analogs or homologs of enkephalin which have antinociceptive activity; analogs or homologs of chlorecystokinin; analogs or homologs of cyclosporin A which have immunosuppressive activity; analogs or homologs of atrial natriuretic factor; peptidergic antineoplastic agents; analogs or homologs of gastrin releasing peptide; analogs or homologs of somatostatin; gastrin antagonists; bradykinin antagonists; neurotensin antagonists; bombesin antagonists; oxytocin agonists and antagonists;
  • Suitable drugs include non-peptide tiierapeutic agents such as antibotics, antimicrobial agents, antineoplastic agents, cardiovascular and renal agents, antiinflammatory, immunosuppressive and immunostimulatory agents and CNS agents.
  • the drug is a peptide such as a fibrinogen receptor antagonist peptide (an RGD peptide), a Growth Hormone Releasing Peptide (GHRP) such as (His-D-Trp-Ala-Trp-D- Phe-Lys-NH2), a vasopressin, a calcitonin or an insulin, more preferably the fibrinogen receptor antagonist peptides cyclo(S,S)-N a -acetyl-Cys-(N a -methyl)Arg-Gly-Asp-Pen-NH2 or cyclo(S,S)-(2-mercapto)benzoyl-(N a -methyl)Arg-Gly-Asp-(2-mercapto)phenylamide o GHRP.
  • GHRP Growth Hormone Releasing Peptide
  • the present invention provides microemulsions comprising a peptide which may be orally administered and which will retain biological activity, diereby overcoming the disadvantages of earlier formulations in which die bioavailability of the peptide has been less tiian satisfactory.
  • the present invention provides formulations which by their nature permit the preparation and administration of a peptide in sufficiently high concentration to allow not only convenient oral administration but also adequate bioavailability of the peptide.
  • die degree of incorporation into d e (w/o) microemulsions of the present invention is limited only by its solubility in the hydrophilic phase.
  • isotonic aqueous phase in the physiological pH range (3 - 8) may be used to aid drug dissolution by die proper modification of die PEG (400) and die high HLB surfactant salt ratio, in particular the fatty acyl or diacyl derivatives of the aliphatic sulfate or sulfonate or sulfosuccinates, without compromising the integrity of the active ingredient and stability of the composition.
  • the aqueous hydrophilic phase suitably comprises water or an isotonic saline solution and may also include a pharmaceutically acceptable solvent which is non-miscible with the selected lipophilic phase, such as polyetiiylene glycol, propylene glycol, sorbitol, mannitol and otiier mono- or di-saccharides.
  • a pharmaceutically acceptable solvent which is non-miscible with the selected lipophilic phase, such as polyetiiylene glycol, propylene glycol, sorbitol, mannitol and otiier mono- or di-saccharides.
  • (1) represents the mixture of the oil and die low HLB surfactant, at a fixed ratio X, (2) die hydrophilic (aqueous) phase and (3) die high HLB surfactant.
  • the point "A” represents a mixture 50% oil plus low HLB surfactant, 20% aqueous phase and 30% high HLB surfactant.
  • the regions of the phase diagram in which microemulsions according to die present invention exist may be determined by titrating a mixture of the oil and low HLB surfactant (in a fixed ratio) against die high HLB surfactant and d e hydrophihc phase, noting points of phase separation, turbidity and transparency. Clear, transparent formulations are indicative of d e formation of a stable microemulsion. Liquid and gel formulations may be obtained at room temperature according to d e specific nature of die components employed.
  • phase diagram From this phase diagram, appropriate percentages may then be read off. The process may then be repeated for other ratios of oil to low HLB surfactant so that an overall picture may be obtained.
  • Representative pseudo-ternary phase diagram of systems containing a medium-chain oil (CAPTEX 8000TM) and low HLB surfactant (CAPMUL CsTM) in the ratios of 2:1, high HLB surfactant (DSS/PEG 400) in a ratio of 1:1, and water is shown in Figure 2.
  • the mixture of oil plus the low HLB surfactant is indicated as component (1), water as component (2) and die high HLB surfactant as component (3).
  • This system produces a two fields of clear, transparent microemulsions which are shown in the phase diagram as die microemulsion field whose fields may be usefully be sub-divided into regions (A) and (C).
  • Ald ough unstable formulations were produced in the region between fields (A) and (C) it is conceivable tiiat, using otiier ratios of PEG 400 to DSS and/or otiier oils/low HLB surfactants, stable microemulsions can be obtained producing a continuous field from region (A) to region (C).
  • This sub-division is based primarily on differences in conductance, viscosity and dilutability in the presence of excess water. Both die viscosity and conductance increase in region (C). In the presence of excess of the dispersed phase (saline or water), microemulsions of regions (A) are inverted to turbid emulsions (o/w) indicative of tiieir original (w/o) nature. In contrast, microemulsions from region (C) remain clear, characteristic of an oil-in-water isotropic system (microemulsion).
  • Microemulsions within the scope of the present invention are those falling within regions (A) and (C) of die pseudo-ternary phase diagram.
  • the present invention provides for stable, self- emulsifying microemulsions as hereinbefore defined in which the relative proportions of the various components lie within regions (A) and (C) of die pseudo-ternary phase diagram in Figure 2.
  • stable clear, transparent liquid microemulsions are obtained when d e oil plus low HLB surfactant is present in the range from about 5% to about 25% , and from 70% to less than 100%, the high HLB surfactant from about 5% to about 30% and from about 70% to less tiian 100% and the aqueous hydrophilic phase, i.e. water or saline is less than 20% (w/w) of the microemulsion.
  • the medium-chain fatty acid triglyceride and die low HLB surfactant together comprise from about 70 to about 100%, (w/w) of the microemulsion.
  • the medium-chain fatty acid triglyceride and the low HLB surfactant together comprise from about 5 to about 25%, (w/w) of the microemulsion.
  • the medium-chain fatty acid triglyceride and the low HLB surfactant may be combined and mixed at various ratios.
  • Useful (w/o) microemulsions may be obtained when die ratio of medium-chain fatty acid triglyceride to low HLB surfactant is in the range of about 5:1 to about 1.5:1, preferably about 4:1 to about 2:1. It may be found that as the ratio of medium-chain fatty acid triglyceride to low HLB surfactant is increased towards 5:1, region (C) of the microemulsion existence field becomes increasingly predominant
  • the high HLB surfactant is present in the range of about 5 to about 30%, and in anodier instance from about 70% to less than 100%.
  • d e high HLB surfactant will be admixed with a polyol in a 10:1 to 1:1 ratio, preferably in a 1 : 1 ratio.
  • the hydrophilic phase comprises from just greater than 0 to about 20%, preferably from about 0.1 to 15% (w/w) of the microemulsion.
  • the ratio of medium-chain fatty acid triglyceride to low HLB surfactant is preferably between 4:1 and 2:1.
  • the microemulsions of the present invention are substantially non-opaque, that is they are transparent or opalescent when viewed by optical microscopic means. In their undisturbed state, they are optically isotropic (non-birefringent) when examined under polarized light. They exhibit excellent stability at low and ambient temperatures, without phase separation, clouding or precipitation, even over prolonged periods of time.
  • the formulations may be stored in a stable form at various temperatures, such as at 4°C, ambient temperature, 37°C and at 50°C, preferably at 4°C or ambient temperatures.
  • Peptide-containing microemulsions of die present invention exhibit a similar stability (shelf life) profile to mat of the corresponding peptide-free microemulsions.
  • Stable (w/o) microemulsions may be formed when d e pH of the aqueous phase varies from a pH of approximately 3 to about 8, a property that can be beneficial for drugs exhibiting higher solubility at low or high pH.
  • the microemulsions are of varying viscosity.
  • Microemulsions with a relatively higher amount of a high HLB surfactant, field (C) DSS/PEG 400 1:1 tend to be more viscous due to the greater viscosity of this material.
  • the diameter of droplets or particles of the microemulsions of the present invention measured, for instance, as the number-average diameter by laser light scattering techniques, is less than 150 nm, more preferably less than 100 nm, yet more preferably less than 50 nm and most preferably in the range 5 to 35 nm.
  • the various phases may optionally contain, some in minor amounts, further ingredients, such as, but not limited to: i) antioxidants such as n-propyl gallate, butylated hydroxyanisole (BHA) and mixed isomers diereof, d-a-tocopherol and mixed isomers thereof, ascorbic acid, propylparaben, methylparaben and citric acid (monohydrate); ii) stabilizers, such as hydroxypropyl cellulose; iii) antimicrobials, such as benozoic acid (sodium salt); and iv) protease inhibitors such as aprotinin.
  • antioxidants such as n-propyl gallate, butylated hydroxyanisole (BHA) and mixed isomers diereof, d-a-tocopherol and mixed isomers thereof, ascorbic acid, propylparaben, methylparaben and citric acid (monohydrate); ii) stabilizers, such as hydroxy
  • microemulsions of the present invention form spontaneously or substantially spontaneously when their components are brought into contact, that is without die application of substantial energy supply, for instance in the absence of high shear energy such as imparted by homogenization and/or microfluidization or other mechanical agitation.
  • die microemulsions may be readily prepared by die simple process of admixing appropriate quantities, witii gende hand mixing or stirring if necessary to ensure thorough mixing.
  • die drug is dissolved in the hydrophilic phase, eitiier directiy or by dilution of a stock solution thereof and this may then be added to a pre-mixed combination of the oil and die low HLB surfactant with mixing, followed by the high HLB surfactant or vice versa.
  • a drug- free microemulsion may be initially prepared by admixing the oil, d e low HLB surfactant, the high HLB surfactant and drug-free hydrophilic phase; to which may en be added further hydrophihc phase in which die drug is dissolved. While higher temperatures (40-60°C) may be needed to solubilize all components during the preparation of the microemulsion, d e preferred systems may be formulated at room temperature. Formulation at ambient temperature is particularly advantageous for thermolabile active ingredients such as peptides.
  • Microemulsions of the present invention are pharmaceutical compositions which comprise a therapeutic agent and are therefore intended for use in therapy, for administration to animals, including man.
  • the present invention provides a metiiod of treatment which comprises administering an effective amount of a microemulsion as hereinbefore defined comprising a therapeutic agent to a patient in need diereof.
  • the amount of drug required for tiierapeutic effect on administration will, of course, vary with the agent chosen, the nature and severity of the condition and die animal undergoing treatment, and is ultimately at the discretion of the physician.
  • the optimal quantity and spacing of individual dosages of a drug will be determined by the nature and extent of the condition being treated, the form, route and site of administration, the particular patient being treated and that such optima can be determined by conventional techniques. It will also be appreciated tiiat die optimal course of treatment, that is, the number of doses given, can be ascertained by those skilled in die art using conventional course of treatment determination tests.
  • the present invention also provides for the use of a medium-chain fatty acid triglyceride, a low HLB surfactant, a high HLB surfactant, a therapeutic agent and a hydrophilic phase as hereinbefore defined in die manufacture of a medicament.
  • Microemulsions of the present invention may be used for oral, topical, rectal, intra- vaginal or other forms of systemic administration and accordingly will be presented in forms suitable for such.
  • microemulsions intended for oral administration may be presented in soft gelatin capsules while the viscosity characteristics of some of the microemulsions make them suitable for direct topical application.
  • Compositions suitable for oral or topical administration are especially prefered.
  • a Pseudo-ternary phase diagram was constructed for the representative system comprising: medium-chain fatty acid triglyceride (oil) CAPTEX 8000 low HLB surfactant CAPMUL C8 high HLB surfactant DSS/PEG 400 (1:1) aqueous phase water in which the ratio of the oil to d e low HLB surfactant was 2: 1.
  • pseudo- ternary phase diagrams for die above noted system can be made wherein the oil to the low HLB surfactant will be 5:1, 4:1 or 3:1.
  • phase diagrams may be made for the representative system above at the ratios from 2:1 to 5:1 wherein the high HLB surfactant may be DSS pure or DSS plus 15% sodium benzoate solublized in PEG 400 or another polylol in ratios ranging from 10: 1 to 1:1.
  • the high HLB surfactant may be DSS pure or DSS plus 15% sodium benzoate solublized in PEG 400 or another polylol in ratios ranging from 10: 1 to 1:1.
  • phase diagram The regions of the phase diagram in which microemulsions according to die present invention exist were determined by titrating a mixture of the oil and low HLB surfactant (in a fixed ratio) against die high HLB surfactant and die aqueous phase, noting points of phase separation, turbidity and transparency.
  • the resultant phase diagram is shown as figure 2. When examined under polarized light, non-birefringent behaviour was observed.
  • Example additional formulations may be prepared but substituting for PEG 400 other polyol's such as propylene glycol or glycerol.
  • the DSS high HLB surfactant may be substituted by sodium dodecyl sulfate (SDS).
  • Formulations having the following compositions can be prepared in accordance with the description below or as described herein.
  • the aqueous phase can be an isotonic solution at pH ranging from 3 to 8.
  • microemulsions were formulated by adding the hydrophilic phase to the solution of dioctyl sodium sulfosuccinate in PEG. To this mixture the oil plus die low HLB surfactant mixture was subsequendy added and thoroughly mixed by hand shaking to produce a clear and transparent microemulsion.
  • the hydrophillic phase can be added to die appropriate amounts by weight of a mixture of the oil and d e low HLB surfactant, to which was then added die high HLB surfactant, with gentle stirring (magnetic hot plate).
  • Active ingredient incorporating microemulsions can be prepared in a similar manner to that shown above, by either dissolving die appropriate amoung of the drug in die appropriate amount of the aqueous phase or preferably, using a stock solution which was then further diluted if so required, and witii vortex stirring if necessary to obtain complete diss solution.
  • the levels of die compound in die plasma samples are determined using fluorescence spectroscopy. After an i.d. dosing at approximately a 1.25 ⁇ Mol/kg or l.Oml kg microemulsion, the bioavailability is determined and compared to the bioavailability when adminstered as an isotonic Tris buffer.
  • Intravenous (i.v.) or intraduodenal (i.d.) administration of the compound (in this instance Calcein) eitiier from a solution of a microemulsion is carried out using conventional methods.
  • fasted rats are anesthetized witii an intraperitoneal injection of a mixture of Rompun (5mg/kg) and Ketset (35mg/kg) and a jugular catheter is implanted. Rats are allowed to recover from surgery for 1 day. Catiierized rats were fasted for 18 hr prior to the administration of die compound. Each compound is administered by lateral tail-vein administration. Blood samples of 0.5ml aliquots were collected at 0, 1, 3, 5, 15, 30, 45, 60, 90, 120, 150 and 180 minutes. The 0 min sample is taken 15 min prior to administration of die dose.
  • Plasma is removed from whole blood by centrifugation at 1600x g for 5 min, and tiien stored at -20C in 250 ⁇ l aliquots per sample.
  • the blood pellet is reconstituted with 12.5 units heparinized saline and returned to die appropriate rat via the jugular catheter. After the experiment, rats are euthanized with i.v. administration of pentobarbital.
  • duodenal cad eters are surgically implanted in anestiietized rats and die animals allowed to recover from surgery for 4-5 days.
  • the compound is administered eitiier from a solution or microemulsion via the duodenal cadieter.
  • Blood samples of 0.5 ml aliquots are collected via jugular catheter in heparinized eppendorf tubes at 0, 10, 30, 60, 120, 180, 240 and 1440 min. The 0 min sample is taken 15 min prior to administration of the dose.
  • Plasma is collected for analysis and die blood returned to rats as described for i.v. administration protocol.
  • the stool of each rat over time is evaluated for consistency by a rank of soft, soft/watery, or mucoid.
  • %F (AUC i /AUC iv) X (Dose i v /Dose i ) X 100
  • formulations of the present invention may be tested for GI irritation assessment with and without an active ingredient by the following method: Oral Dosing in Rats/GI Irritation Assessment
  • Suitable rats for use in this assement are male Sprague-Dawley (Caesarian Delivery - Virus Antibody Free; Charles River Laboratories). The rats are fasted overnight the day before the experiment. Dosing with the microemulsion at the desired dose is done by gavage at a volume not exceeding 10 ml/kg. Upon termination of the experiment animals are euthanized witii asphyxiation using carbon dioxide and exsanguinated. Abdominal incisions are then performed and gross observations of the gastric and duodenal mucosa are made witii naked eyes and under a microscope (Nikon model SMZ-10 binocular microscope).
  • One aspect of the present invention are the formulations of w/o self-emulsifying microemulsions with or witiiout peptide which produce littie, if any, damage along die GI tract upon oral administration.
  • the present formulations may be given orally by gavage (preferably at three rats per formulation). After 24 hrs the animals are exsanguinated and upon abdominal incisions are examined both by naked eye and under the microscope. The mucosal surface of both the stomach and duodenum of the animals are examined to see if they are free of any lesions at naked eye.
  • microemulsions formulated as described above and containing, for instance, 3mg of a peptide, such as an RGD fibrinogen receptor antagonist containing peptide, per gr of microemulsion are tested in die following manner for oral bioavailability.
  • Fasted rats are given an intraperitoneal (i.p.) injection and surgically fitted witii femoral artery catheters. Rats ware allowed to recover from the surgery for 1 day. Cadierized rats are fasted for 18 hr prior to the experiment. Each rat receives 3mg of peptide by lateral tail-vein administration from a solution prepared as follows:
  • Intraduodenal ( d.) administration of peptide in microemulsion Fasted rats are given an i.p. injection of anesthesia cocktail and surgically fitted with jugular and duodenal catheters. Rats are allowed to recover from the surgery for 4-5 days. Catherized rats are fasted 18-20 hrs. prior to the experiment. Each rat receives lOmg of peptide in either microemulsion or saline solution. Blood samples of 0.5ml aliquots are collected via jugular catheter in heparinized eppendorf tubes at 0, 10, 30, 60, 120, 180, 240 and 1440 minutes. The 0 min sample is taken 15 min prior to administration of die dose by duodenal catheter.
  • Plasma is collected for analysis and the blood returned to rats as described in the i.v. administration (part a) above. After 1440 min, rats are euthanized by iv administration of pentobarbital, exsanguinated and the GI tract removed for gross observation.
  • the area under die plasma concentration curve (AUC) from 0 to 240 minutes is determined for each rat.
  • AUC area under die plasma concentration curve
  • percentage bioavailability is determined for each animal by d e following equation with the average AUC from iv administration: [(AUq AUQ v )*(dose /v /dose ⁇ i)] * [100].
  • the oral bioavailability data for the RGD peptide in rats after intraduodenal administration of a microemulsion containing the above formulations incorporating a fibrinogen receptor antagonist of a peptide dose may en be obtained in die above noted manner.
  • the formulations of the present invention are tested for in vivo activity.
  • a fibrinogen receptor antagonist a platelet aggregation assay is employed to determine pharmacological activity of the peptide from microemulsions.
  • Dogs used in this assay are male Mongrels (i.e. from mixed breeds). The dog(s) are fasted overnight the day before the experiment.
  • the cephalic vein of choice is prepared for the indwelling catheter in the following way: the area is first shaved and cleaned with a gauze soaked in 70% alcohol. An indwelling catheter is placed in the caphalic vein and attached to a luer lock adapter filled with 3.8% sodium citrate. The catheter is securely taped down.
  • the blood samples are then assayed for platelet aggregation inhibition using die Chromo-Log whole blood aggregometer.
  • the instrument is warmed to 37°C before samples are run and die probe is cleaned with distilled water and a soft brush.
  • the probe is attached to die aggregometer and placed in a cuvette of saline solution and warmed in a side cuvette well in the aggregometer.
  • 1 ml of the 2.7 ml of blood sample mixed with the 0.3 ml 3.8% sodium citrate contained in the Venoject vacuum tube is added to a cuvette and placed in the aggregometer well.
  • a stir bar is placed in die cuvette and set at 900 rpm.
  • the probe is placed firmly into the test cuvette and die lid is shut.
  • the stirring cuvette is permitted to settle for five minutes at which point 5 ⁇ l of collagen is added to the whole blood diat is being stirred to yield to a 5 ⁇ g/ml final solution in the cuvette.
  • the reaction is monitored for two minutes once the slope change reaches the baseline of the collagen addition, calculating the change in ohms per minute using the slope of the two minutes.
  • the change in ohms per minute is calculated as a % of die control.
  • the control value is determined by the average of the -15 and the 0 time points. After each use the probe is removed and cleaned with distilled water and wiped with a soft cloth and brush.
  • a dog is considered a good model to assess the pharmacological effect of one class of peptides of interest herein, the RGD containing fibrinogen receptor antagonists.
  • Experiments are conducted as described above, witii a peptide dose of 3 mg kg or microemulsion dose of 0.5 ml/kg.
  • Control experiments where the peptide is given orally in a saline solution are independently carried out earlier and serve as a useful comparison to the effects seen witii d e microemulsion-formulated peptide.
  • die active ingredients utilized herein is a Growth Hormone Releasing Peptide die appropriate assay for in vivo activity is determined as shown below.
  • a microemulsion with a composition (w/w) in accordance witii the Examples above is made. Upon preparation, they are further stored in a stable form at ambient temperature for approximately 48 hrs before the in vivo evaluation.
  • a control solution of a GHRP peptide, His-D-Trp-Ala-Trp-D-Phe-Lys-NH2, in saline at 1.5 mg/ml is also prepared.
  • Dosing is done by single intraduodenal administration of GHRP at 3 mg/kg in male rats in saline solution (control) and in die aforementioned microemulsion using 3 rats in each case.
  • each rat Prior to actual sampling and dosing, each rat is anesthetized with Pentobarbitol at 50 mg/kg i.p, diluted with saline to a final volume of 1 ml. The rats stay anesthetized for the entire experiment. Dosing is achieved in the following way: a small incision 2-3 cm long is made on d e abdominal midline, and tiien a purse-string suture is placed on die duodenal muscle.
  • a small hole is made in d e center of the purse-string suture in which a blunt 23 G stub needle attached to a tuberculin syringe is inserted to deliver die dose.
  • die purse-string is tied to close die opening. The incision is closed with wound clips.
  • a 0.2 ml blood sample is obtained via jugular catheter at the following intervals: -15, 0, 5, 10, 15, 30, 45, 60, 90, and 120 minutes. Blood samples are stored on ice and subsequently analyzed for Growth Hormone by an RIA method.
  • the amount of active ingredient required for therapeutic systemic administration will, of course, vary with die compound chosen, the nature and severity of the condition, and the mammal, including humans, undergoing treatment, and is ultimately at the discretion of the physician.
  • the present invention also includes a metiiod of treatment which comprises administering an effective amount of a pharmaceutical composition as defined herein to a patient in need thereof.
  • the therapeutic agent is selected from a fibrinogen receptor antagonist peptide, growth hormone releasing peptide or an analog or homolog thereof, a vasopressin or analog or homolog thereof, elcatonin, a calcitonin, a calcitonin-gene releated peptide, a porcine somatostatin an analog or homolog thereof, insulin or a homolog or analog thereof.

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Abstract

Des compositions pharmaceutiques se présentant sous la forme de micro-émulsions comprennent une huile, un mélange de tensioactifs à équilibre hypo-hydrophile élevé et faible, le tensioactif à équilibre hydrophile élevé comprenant un sulfate alkyle/dialkyle à chaîne moyenne, un sulfonate ou un sel de sulfosuccinate dissous dans un alcool polyvalent, une phase aqueuse et éventuellement un agent biologiquement actif.
PCT/US1994/001678 1993-02-17 1994-02-17 Micro-emulsions comprenant des peptides therapeutiques WO1994019001A1 (fr)

Priority Applications (2)

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JP6519110A JPH08507066A (ja) 1993-02-17 1994-02-17 治療用ペプチド含有のミクロエマルジョン
EP94909654A EP0684833A4 (fr) 1993-02-17 1994-02-17 Micro-emulsions comprenant des peptides therapeutiques.

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US1860993A 1993-02-17 1993-02-17
US08/018,609 1993-02-17

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WO1994019001A1 true WO1994019001A1 (fr) 1994-09-01

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Cited By (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0753311A4 (fr) * 1993-04-19 1997-05-21 Inst Advanced Skin Res Inc Preparation en microemulsion contenant une substance difficilement absorbable
EP0736041A4 (fr) * 1993-11-17 1997-09-10 Ibah Inc Liquide transparent permettant l'administration de medicaments encapsules
DE19716953A1 (de) * 1997-04-22 1998-10-29 Forschungszentrum Juelich Gmbh Neue Mikroemulsionen mit Komponenten zur Unterdrückung von Flüssigkristallen, insbesondere für die in-situ-Bodensanierung
DE10013796A1 (de) * 2000-03-20 2001-09-27 Kaercher Gmbh & Co Alfred Dekontaminations-Zusammensetzung sowie Emulsionen hieraus
US7094550B2 (en) 1993-05-13 2006-08-22 Neorx Corporation Method to determine TGF-beta
EP2098230A1 (fr) 1997-03-31 2009-09-09 Boston Scientific Scimed Limited Utlisation des inhibiteurs cytosquelettiques en forme cristalline pour l'inhibition ou la prévention de la resténose
US7625410B2 (en) 2001-05-02 2009-12-01 Boston Scientific Scimed, Inc. Stent device and method
EP2292225A1 (fr) 1997-03-31 2011-03-09 Boston Scientific Scimed Limited Forme de dosage comprénant du taxol en forme cristalline
WO2018188235A1 (fr) * 2017-04-14 2018-10-18 黄月华 Groupe de signaux présentateurs d'antigène de peptide d'antigène du virus de l'hépatite b combiné à des informations d'antigène de cellules cancéreuses du foie et son application
CN109010117A (zh) * 2018-07-10 2018-12-18 山西医科大学 一种具有滋润保湿作用的微乳凝胶护唇啫喱及其制备方法

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US20110144010A1 (en) * 2007-06-01 2011-06-16 Novo Nordisk A/S Spontaneously Dispersible Preconcentrates Including a Peptide Drug in a Solid or Semisolid Carrier
JP5749155B2 (ja) 2008-03-18 2015-07-15 ノボ・ノルデイスク・エー／エス プロテアーゼ安定化アシル化インスリンアナログ
CA2870313A1 (fr) 2012-04-11 2013-10-17 Novo Nordisk A/S Formulations a base d'insuline
EP3445316B1 (fr) * 2016-04-21 2019-10-02 Unilever Plc. Procédé de production d'émulsions en petites gouttelettes à basse pression
EA037016B1 (ru) * 2016-04-21 2021-01-27 Юнилевер Н.В. Моющие композиции в виде наноэмульсии для личной гигиены и способ их получения
US20200323747A1 (en) * 2016-04-21 2020-10-15 Conopco, Inc., D/B/A Unilever Novel nanoemulsions comprising n-acyl amino acid salt and process for making
CA3046583A1 (fr) 2016-12-16 2018-06-21 Novo Nordisk A/S Compositions pharmaceutiques contenant de l'insuline

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Cited By (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0753311A4 (fr) * 1993-04-19 1997-05-21 Inst Advanced Skin Res Inc Preparation en microemulsion contenant une substance difficilement absorbable
US5948825A (en) * 1993-04-19 1999-09-07 Institute For Advanced Skin Research Inc. Microemulsion preparation containing a slightly absorbable substance
US7094550B2 (en) 1993-05-13 2006-08-22 Neorx Corporation Method to determine TGF-beta
EP0736041A4 (fr) * 1993-11-17 1997-09-10 Ibah Inc Liquide transparent permettant l'administration de medicaments encapsules
EP2098230A1 (fr) 1997-03-31 2009-09-09 Boston Scientific Scimed Limited Utlisation des inhibiteurs cytosquelettiques en forme cristalline pour l'inhibition ou la prévention de la resténose
EP2292225A1 (fr) 1997-03-31 2011-03-09 Boston Scientific Scimed Limited Forme de dosage comprénant du taxol en forme cristalline
DE19716953A1 (de) * 1997-04-22 1998-10-29 Forschungszentrum Juelich Gmbh Neue Mikroemulsionen mit Komponenten zur Unterdrückung von Flüssigkristallen, insbesondere für die in-situ-Bodensanierung
DE19716953B4 (de) * 1997-04-22 2006-02-09 Forschungszentrum Jülich GmbH Verfahren zur Sanierung von mit Schadstoff kontaminiertem Boden und bikontinuierliche Mikroemulsion
DE10013796A1 (de) * 2000-03-20 2001-09-27 Kaercher Gmbh & Co Alfred Dekontaminations-Zusammensetzung sowie Emulsionen hieraus
US7625410B2 (en) 2001-05-02 2009-12-01 Boston Scientific Scimed, Inc. Stent device and method
WO2018188235A1 (fr) * 2017-04-14 2018-10-18 黄月华 Groupe de signaux présentateurs d'antigène de peptide d'antigène du virus de l'hépatite b combiné à des informations d'antigène de cellules cancéreuses du foie et son application
CN109010117A (zh) * 2018-07-10 2018-12-18 山西医科大学 一种具有滋润保湿作用的微乳凝胶护唇啫喱及其制备方法

Also Published As

Publication number Publication date
EP0684833A1 (fr) 1995-12-06
EP0684833A4 (fr) 1996-09-11
JPH08507066A (ja) 1996-07-30

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