COMPOSITION THAT FORMS LIQUID CRYSTALLINE PARTICLES
20210369623 · 2021-12-02
Inventors
Cpc classification
A61K9/0056
HUMAN NECESSITIES
A61K9/2072
HUMAN NECESSITIES
International classification
Abstract
The invention provides an oral disintegrating tablet (ODT) for administration to oral mucosa comprising an amphiphilic compound capable of self-assembling into liquid crystalline particles when contacted with a hydrophilic solvent, a therapeutically effective amount of a 1 to 10 kDa active ingredient and a pharmaceutically acceptable disintegrant, wherein the 1 to 10 kDa active ingredient is orally bioavailable. The water channels of the liquid crystalline particle may be increased in size. ODTs with enhances water channels may also be useful for delivery of active ingredients of less than 1 kDa.
Claims
1. An oral disintegrating tablet, suitable for administration to the oral mucosa, comprising an amphiphilic compound capable of self-assembling into liquid crystalline particles when contacted by a hydrophilic solvent; a therapeutically effective amount of a 1 kDa to 10 kDa active ingredient; and a pharmaceutically acceptable disintegrant.
2. The tablet according to claim 1, wherein the active ingredient is a low molecular weight heparin.
3. The tablet according to claim 1, wherein the active ingredient is ardeparin, bemiparin, nadroparin, reviparin, enoxaparin, parnaparin, certoprain, dalteparin and/or tinzaparin, preferably enoxaparin and/or dalteparin.
4. The tablet according to claim 1, further comprising a water channel enhancer.
5. The tablet according to claim 4, wherein the water channel enhancer increases the size of the water channels in the liquid crystalline structure formed when the amphiphilic compound self-assembles by at least about 5 Å.
6. The tablet according to claim 4, wherein the water channel enhancer increases the lattice size in the liquid crystalline structure formed when the amphiphilic compound self-assembles by at least about 10 nm.
7. The tablet according to claim 6, wherein the water channel enhancer increases the lattice size in the liquid crystalline structure formed when the amphiphilic compound self-assembles by at least about 20 nm.
8. The tablet according to claim 4, wherein (i) the amphiphilic compound, in the presence of the water channel enhancer, is capable of self-assembling into cubic Im3m liquid crystalline particles and (ii) the amphiphilic compound in the absence of the water channel enhancer is capable of self-assembling into cubic Pn3m liquid crystalline particles.
9. The tablet according to claim 4, wherein the water channel enhancer is selected from the group consisting of Tween 80, sodium cyclamate, saccharin sodium, Poloxamer 188, Poloxamer 407, Mg Trisilicate, SLS, menthol, stearic acid, butylated hydroxyl toluene and sodium bicarbonate.
10. The tablet according to claim 1, wherein when the ODT is administered to the oral mucosa, the ODT facilitates systemic administration of the active ingredient across the oral mucosa.
11. The tablet according to claim 1, wherein when the tablet contacts a hydrophilic solvent the amphiphilic compound self-assembles into liquid crystalline particles.
12. The tablet according to claim 1, wherein the oral disintegrating tablet disintegrates in less than 2 minutes following contact with a hydrophilic solvent.
13. The tablet according to claim 1, wherein the amphiphilic compound has a critical packing parameter (CPP) of >½ and/or a hydrophilic lipophilic balance (HLB) of 0 to <10.
14. The tablet according to claim 1, wherein the amphiphilic compound has the structure of Formula (I):
X−T Formula (I) wherein X has at least 2 hydrogen bond forming functional groups and is selected from the group consisting of an ester, ether, anhydride, amide, amine, carbamide, glycerol, biuret, phenyl, pyridine or phosphate; and T is selected form the group consisting of: (i) a single C12 to C18 alkyl, alkenyl and alkynyl terminally attached to X optionally comprising: a. one or more double bonds (preferably cis and at about C7 to C11); or b. three or more methyl branches (preferably isoprenoid branching); and (ii) two C12 to C18 alkyl, alkenyl and alkynyl both terminally attached to X.
15. The tablet according to claim 1, wherein the active ingredient has a log P of −0.5 to 6.4.
16. The tablet according to claim 1, wherein the disintegrant is about 10 to about 50% w/w of the tablet.
17. (canceled)
18. The tablet according to claim 1, wherein the disintegrant is sodium starch glycolate, crosslinked polyvinylpyrrolidone or both.
19. The tablet according to claim 1, wherein the amphiphilic compound is a glycerol monoleate.
20. The tablet according to claim 19, wherein the amphiphilic compound, in the absence of a water channel enhancer, forms cubic Pn3m liquid crystalline particles.
21. The tablet according to claim 1, wherein the amphiphilic compound is 1 to 20% w/w of the tablet and/or the active ingredient is 0.5 to 10% w/w of the tablet.
22-47. (canceled)
Description
BRIEF DESCRIPTION OF THE DRAWINGS
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DETAILED DESCRIPTION OF THE EMBODIMENTS
[0140] It will be understood that the invention disclosed and defined in this specification extends to all alternative combinations of two or more of the individual features mentioned or evident from the text or drawings. All of these different combinations constitute various alternative aspects of the invention.
[0141] Reference will now be made in detail to certain embodiments of the invention. While the invention will be described in conjunction with the embodiments, it will be understood that the intention is not to limit the invention to those embodiments. On the contrary, the invention is intended to cover all alternatives, modifications, and equivalents, which may be included within the scope of the present invention as defined by the claims.
[0142] Further aspects of the present invention and further embodiments of the aspects described in the preceding paragraphs will become apparent from the following description, given by way of example.
[0143] All of the patents and publications referred to herein are incorporated by reference in their entirety.
[0144] For purposes of interpreting this specification, terms used in the singular will also include the plural and vice versa.
[0145] One skilled in the art will recognize many methods and materials similar or equivalent to those described herein, which could be used in the practice of the present invention. The present invention is in no way limited to the methods and materials described.
[0146] The inventors of the present invention have developed an ODT comprising a large API for the delivery of active ingredients via the oral mucosa of a subject. The ODT disintegrates and forms liquid crystalline particles when it contacts a hydrophilic solvent. The liquid crystalline particles that form when the ODT contacts a hydrophilic solvent optionally also have increased water channel size. The ODT comprising an amphiphilic compound capable of self-assembling into liquid crystalline particles when contacted with a hydrophilic solvent, a therapeutically effective amount of an active ingredient, a water channel enhancer, and a pharmaceutically acceptable disintegrant that results in the formation of liquid crystalline particles with increased lattice size/water channel size are optionally be prepared with either large or small molecule active ingredients.
[0147] The term “therapeutically effective amount” as used throughout the specification is understood to mean the amount or dose of a compound or composition thereof that will lead to one or more desired effects, for example, the reduction of cholesterol synthesis. A therapeutically effective amount of an active ingredient will vary according to factors such as the disease state, age, sex, and weight of a subject, and the ability of the substance to elicit a desired response in the subject.
[0148] The term “dalteparin” refers to a low molecular weight heparin having a molecular weight of 5.6-6.4 kDa. An example is marketed as Fragmin by Pfizer Inc.
[0149] The term “water channels” refers to the channels of hydrophilic solvent through a complex liquid crystalline particle or bulk phase, such as a cubic or hexagonal particle/phase, assembled in hydrophilic solvent.
[0150] The term “water channel enhancer” refers to an ingredient that increases the size of the repetitive lattice structure and/or water channel size within a complex liquid crystalline particle or bulk phase, such as a cubic or hexagonal particle/phase. The size of the usual water channel/lattice for a specific amphiphile in certain conditions (eg temperature) can be determined by combining that amphiphile with a hydrophilic solvent so that it self-assembles into liquid crystalline particles and then analysing the particles on the SAX/WAX beamline of a synchrotron and analysing the results as described below and in the examples. A compound can be tested for water channel enhancing effect by combining the compound with the amphiphile and the hydrophilic solvent and then testing on the SAX/WAX beamline of a synchrotron and analysing the results as described below and in the examples to determine if the water channel and/or lattice size has increased. Many water channel enhancers have a threshold effective concentration. This concentration can also be determined experimentally. When reference is made to an increase in the water channel and/or lattice size for the liquid crystalline structure formed by an amphiphile, this refers to an increase in comparison to the size of the water channel and/or lattice size for the same amphiphile in the same conditions except without the presence of the water channel enhancer.
[0151] As used herein, the terms “amphiphilic compound” and “amphiphile” are used interchangeably.
[0152] The term “enoxaparin” and “enoxaparin sodium” refer to a low molecular weight heparin and its sodium salt having a molecular weight of 3.8-5.0 kDa. Examples are marketed as Clexane and Xaparin.
[0153] The term “low molecular weight heparin” refers to fractionated heparin where at least 60% of all chains have a molecular weight less than 8 kDa. Optionally, at least 70%, at least 80% or at least 90% of all chains have a molecular weight less than 8 kDa. Examples include ardeparin, bemiparin, nadroparin, reviparin, enoxaparin, parnaparin, certoprain, dalteparin and tinzaparin. The low molecular weight heparin can be a heparin-related molecule.
[0154] The term “heparin” refers to a linear sulfated glycosaminoglycan polymer made up of the following structure:
##STR00001##
[0155] The most common disaccharide unit of heparin is depicted below:
##STR00002##
[0156] Other common substitution patterns present in the disaccharide units of heparin include GlcA-GlcNAc, GlcA-GlcNS, IdoA-GlcNS, IdoA(2S)-GlcNS, IdoA-GlcNS(6S) and IdoA(2S)-GlcNS(6S); wherein GlcA is β-D-glucuronic acid, IdoA is α-L-iduronic acid, IdoA(2S) is 2-O-sulfo-α-L-iduronic acid, GlcNAc is 2-deoxy-2-acetamido-α-D-glucopyranosyl, GlcNS is 2-deoxy-2-sulfamido-α-D-glucopyranosyl and GlcNS(6S) is 2-deoxy-2-sulfamido-α-D-glucopyranosyl-6-O-sulfate. Heparin also includes disaccharide units containing a 3-O-sulfated glucosamine (GlcNS(3S,6S)) or a free amine group (GlcNH.sub.3.sup.+) in low concentrations.
[0157] Unfractionated heparin is heparin having a molecular weight of 12 to 15 kDa.
[0158] The term “heparin-related molecule” refers to an oligomer or polymer substantially related to heparin or a fragment of heparin. Examples include fondaparinux, heparan sulfate and fractions of heparan sulfate.
[0159] The term “self-assembled particles” as used throughout the specification is understood to mean an aggregate of amphiphiles that possess some degree of internal organisational order, for example, a liquid crystalline or colloidal particle or colloidosome or a solid lipid particle. The particles can be either nanoparticles or microparticles depending on their average size, typically less than about 1 μm, preferably in a range of about 10 nm to about 500 nm, more commonly about 200 nm. The self-assembled particles are formed by contacting the amphiphile with solvent (ie lyotropic liquid crystalline particles). In some embodiments, the self-assembled particles themselves aggregate into a bulk lyotropic phase.
[0160] The term “liquid crystalline particles” as used throughout the specification is understood to mean self-assembled particles including liquid crystalline structure.
[0161] The term “bulk phase” as used throughout the specification is understood to mean a lyotropic phase that includes but is not limited to: micellar cubic (I1); normal hexagonal (H1); bicontinuous cubic (V1); lamellar (L); reversed bicontinuous cubic (V2); reversed hexagonal (H2); reversed micellar cubic (I2) and sponge (L3) phases.
[0162] The term “cubic phase” as used throughout the specification is understood to refer to two main classes of phases: micellar cubic (11) and bicontinuous cubic (V1). ‘Micellar cubic phase’ refers to a phase consisting of spherical micelles arranged in a cubic array. A ‘normal micellar cubic phase’ or ‘II phase’ (ie (11)) consists of spherical normal micelles arranged in a cubic array, whilst an ‘inverse micellar cubic phase’ or ‘III phase’ (ie (I2)) consists of spherical inverse micelles arranged in a cubic array. ‘Bicontinuous cubic phase’ refers to a family of closely related phases that consist of a single curved lipid bilayer that forms a complex network that separates the polar solvent space into two continuous, but non-intersecting volumes. Bicontinuous cubic phases possess long range order based upon a cubic unit cell. Bicontinuous cubic phases have zero mean curvature; that is, at all points on surface of the amphiphile bilayer, the surface is as convex as it is concave. Bicontinuous cubic phases include the normal (‘vI phase’ or ‘V1 phase’) or reverse (‘vII phase’ or ‘V2 phase’) type. Several types of long range orientational orders have been observed for bicontinuous cubic phases; the orientational order in these phases correspond to space groups Ia3d, Pn3m, and Im3m. When a colloidosome possesses the internal structure of a bulk cubic phase the colloidosome is referred to as a ‘cubosome’.
[0163] The term “hexagonal phase” as used throughout the specification is to be understood to mean an amphiphile phase consisting of long, rod-like micelles packed into a hexagonal array. A “normal hexagonal phase” is a hexagonal phase consisting of long, rod-like normal micelles, whilst an ‘inverse hexagonal phase’ is a hexagonal phase consisting of long, rod-like inverse micelles. The normal hexagonal phase is also referred to as the “HI phase” or “H1 phase” and the inverse hexagonal phase is also referred to as the “HII phase” or “H2 phase”. When a colloidosome possesses the internal structure of a bulk hexagonal phase the colloidosome is referred to as a ‘hexosome’.
[0164] The term “lamellar phase” or “L phase” as used throughout the specification is to be understood to mean a stacked bilayer arrangement, where opposing monolayers of the hydrophilic portion of amphiphile molecules are separated by a polar solvent domain, while the hydrophobic portion of the amphiphile molecule of the back-to-back layers are in intimate contact to form a hydrophobic layer. The planar lamellar phase is referred to as the “Lα phase”. There are three lamellar phases, (1) the fluid lamellar phase (Lα) where the chains are melted, (2) the gel lamellar phase (Lβ) where the chains are mostly melted but some degree of short range order and (3) the lamellar crystalline phase (Lc), where the chains are crystalline with very short range order.
[0165] The term “sponge phase” or “L3 phase” as used throughout the specification refers to a phase that resembles a bicontinuous cubic phase, in that it possesses an amphiphile bilayer that separates the polar solvent space into two unconnected volumes, but it does not possess long range order. Accordingly, these phases are analogous to a ‘melted cubic phase’.
[0166] The term “prodrug” as used throughout the specification refers to a biologically active agent including structural modifications thereto, such that in vivo the prodrug is converted, for example, by hydrolytic, oxidative, reductive or enzymatic cleavage to the biologically active agent by one or more reactions or steps. It includes an agent that requires one or more chemical conversion steps or steps of metabolism to produce the active molecule.
[0167] The term “disintegrant” as used throughout the specification refers to a tabletting excipient that is incorporated into the formulation of tablets or capsules to promote their disintegration when they come into contact with liquid or fluid matter, specifically aqueous liquid. The general purpose of incorporating one or more disintegrants in the product formulation is to increase the surface area of the product and soften the binding matter that holds together the solid particles that make up the product. The rate of dissolution in the media increases as the particle size reduces and is greatest when the tablets or capsules reduced to fine particles, as shown schematically in Figure. 1. Rapid dissolution increases the rate of absorption of the active ingredient by the body, producing the desired therapeutic action. Tablets that are labelled as chewable generally do not require a disintegrant to be incorporated in the formulation.
[0168] The terms “oral disintegrating tablet”, “orally dispersible tablet”, “fast dissolving tablet”, or “ODT” as used throughout the specification refer to tablets that disintegrate rapidly in the oral cavity of a subject after administration. ODTs do not need to be swallowed. No liquid is required to be consumed by the subject when taking medication from an ODT either. The ODTs of the invention are for delivery of an API via the oral mucosa. However, traditional ODTs were designed to ease swallowing of an API.
[0169] The term “substantially disintegrates” as used throughout the specification refers to a disintegration of the tablet largely into constituent particles which were previously compressed into whole tablets.
[0170] The term “drug”, “active” or “active pharmaceutical ingredient” as used herein includes a pharmaceutically acceptable and therapeutically effective compound suitable for treating or preventing one or more diseases, symptoms of diseases, and medical conditions, as well as pharmaceutically acceptable salts, stereoisomers and mixtures of stereoisomers, solvates (including hydrates), and/or esters thereof.
[0171] The liquid crystalline particles of the present invention are optionally self-assembled into bulk phase including an active ingredient. Typically, a bulk material having a certain phase will form from an amphiphile, that is, a molecule that possesses both a hydrophilic portion and a hydrophobic portion. The self-assembly behaviour of amphiphiles in solvent arises because of the preferential interaction between the solvent and either the hydrophilic or hydrophobic portion of the amphiphilic molecule. When an amphiphile is exposed to a polar solvent, the hydrophilic portion of the amphiphile tends to preferentially interact with the polar solvent, resulting in the formation of hydrophilic domains. The hydrophobic portion of the amphiphile molecules tend to be excluded from this domain, resulting in the de facto formation of a hydrophobic domain.
[0172] It is in a self-assembled form (ie as liquid crystalline particles or liquid crystalline bulk phase) that amphiphiles are capable of acting as an inert carrier or matrix into which biologically active molecules, such as an active ingredient, are incorporated. The nanoscale porosity of the self-assembled materials provides a high internal and external surface area. An active ingredient that is distributed within a region of this material is believed to be distributed in an ordered arrangement, and at a high loading concentration due to the large internal and external liquid crystal surface area. Self-assembled bulk phases exist with a variety of orientational orders. If long-range orientational order is observed within the self-assembled bulk phase at equilibrium, the self-assembled bulk phase is termed a ‘mhesophase’, a ‘lyotropic liquid crystalline phase’, a ‘lyotropic phase’ or, as used herein, simply a ‘phase’.
[0173] There are 2 principal types of liquid crystalline phases: thermotropic liquid crystals and lyotropic liquid crystals. Thermotropic liquid crystals can be formed by heating a crystalline solid or by cooling an isotropic melt of an appropriate solute. Lyotropic liquid crystals can be formed by addition of a solvent to an appropriate solid or liquid amphiphile. The manipulation of parameters such as amphiphile concentration and chemical structure, solvent composition, temperature and pressure can result in the amphiphile-solvent mixture adopting lyotropic phases with distinctive characteristics.
[0174] Examples of particular phases that can be formed by self-assembled particles are set out above. It is possible to disperse the bulk phases described above to form colloidal particles (so-called ‘colloidosomes’) that retain the internal structure of the non-dispersed bulk phase. When these particles possess the internal structure of a reversed bicontinuous cubic phase, the particles are colloquially referred to as cubosomes. Similarly, when the particles possess the internal structure of a reversed hexagonal phase, they are referred to as hexosomes. When the particles possess the internal structure of a lamellar phase, they are referred to as liposomes.
[0175] Whilst the bulk materials can be of use in some circumstances, the use of bulk materials having cubic phases in drug administration is limited by their high viscosity making them difficult to administer. In these cases, colloidal dispersions of particles of these phases can optionally be used in drug delivery. More preferred phases for use as drug delivery vehicles when prolonged release of the active ingredient is desired are bicontinuous cubic phase or reversed hexagonal phase. The inverse cubic phase affords distinct aqueous regions that form two continuous water networks (or channels) throughout the cubic phase that more readily allow diffusion of an active ingredient. The inverse cubic liquid crystal phase is thermodynamically stable and co-exists in equilibrium with excess water over a broad temperature range. Alternatively, if the bicontinuous cubic phase is viscous and difficult to administer it could be possible to administer a lamellar phase material that converts into the cubic phase upon dissolution with aqueous, water rich, body fluids (thus facilitating the conversion of one phase to another). For example, a suitable material is a phospholipid such as 1,2-dioleoyl-sn-glycero-3-phosphocholine. The cubic phase in situ provides a viscous depot from which an active ingredient can slowly be released. An inverse cubic liquid crystal phase provides an appropriate scaffold in which to distribute or load the active ingredient owing to the high surface area of the internal liquid crystal structure (up to 400 m.sup.2/g).
[0176] Suitable pharmaceutical carriers, excipients, diluents, additives and vehicles are known to those skilled in the art.
[0177] The formulation optionally includes one or more binders such as hydroxypropylmethylcellulose (HPMC), ethyl cellulose, acacia, polyvinyl alcohol (PVA), and polyvinylpyrrolidone (Povidone). Povidone is a preferred binder. Optionally, the binder is about 0.5 to about 5% w/w of the ODT. Optionally, the binder is about 1 to about 3% w/w or about 1.5 to about 2.5% w/w of the ODT.
[0178] The formulation optionally includes one or more glidants such as talc, magnesium trisilicate and colloidal silicon dioxide.
[0179] The formulation optionally includes one or more fillers such as lactose, mannitol, sorbitol, starch, maltodextrin, acacia and silicon dioxide.
[0180] The formulation optionally includes one or more lubricants such as glyceryl behenate, stearic acid, talc, zinc stearate, calcium stearate, magnesium stearate, aluminum stearate and sodium stearyl fumarate.
[0181] If the formulation is prepared by thermoplastic granulation the formulation optionally includes thermoplastic granulation agents such as glycerol monostearate and glyceryl behenate.
[0182] The presence of liquid crystalline phase or a colloidal dispersion of liquid crystalline particles in a self-assembled structure of the invention or following contact of a composition or ODT of the invention with a hydrophilic solvent can be determined using the SAX/WAX beamline of a synchrotron, cross polarised light microscopy (CPLM) or Cry-Em. In certain circumstances, such as a low proportion of amphiphilic compound in the ODT, liquid crystalline phase may not be identified using the SAX/WAX beamline of the synchrotron and an alternative, such as, CPLM may be preferred. CPLM can identify LC structures but does not provide information on the internal phase.
[0183] Properties of liquid crystals have been extensively characterized using small angle x-ray scattering (SAXS). SAXS is a non-destructive method to evaluate the shape, size, internal structure, crystallinity and porosity of a nanostructured sample. An x-ray beam is fired at the sample and the intensity of the scattered x-rays are recorded according to Bragg's law;
2d sin θ=nλ
[0184] Where n is an integer, λ is the wavelength, θ is the scattering and d is the interplanar distance. The scattering pattern is converted to a plot of intensity versus scattering vector, q, by equation:
q=4πλ sin θ/2
[0185] The scattering vector where peaks are located can be used to calculate d by equation:
d=2π/q
[0186] Through integration, the pattern intensity vs. scattering vector (q) plot is generated. The unique spacing of peaks act as a ‘fingerprint’ to accurately determine the liquid crystalline structure present.
[0187] Using Bragg's law, the lattice parameter, a, can be calculated.
[0188] While lattice parameter acts as an indication for water channel size, accurate calculations can be performed using the following equations:
(Pn3m)r=0.391α−I
(Im3m)r=0.305α−I
[0189] Where, r=the radius of the water channel, a=the lattice parameter (determined via SAXS) and I=lipid chain length. The lipid chain length (I) of GMO is about 18 Angstrom.
[0190] The release of an active ingredient depends on both the liquid crystalline structure and the water channel size. For a specific amphiphile, the water channel size varies between the different liquid crystalline structures as follows:
H2<Ia3d<Pn3m<Im3m.
[0191] The cubic structures (Ia3d, Pn3m, Im3m) have bicontinuous water channels (ie 2 water channels in a maze-like configuration) whereas H2 has smaller discontinuous water channels. Most of the water channels in H2 liquid crystalline particles are closed. The closed water channels is one of the reasons for the slow release of active ingredient from this structure.
[0192] The CPP of an amphiphilic compound can be determined by quantum mechanics molecular simulations to determine geometrical and quantitative structure-activity relationship (QSAR) values. See, Fong 2016. The HLB of an amphiphilic compound is calculated based on the number and identity of hydrophilic/lipophilic groups. The skilled person understand these calculations.
[0193] The CPP and HLB of some amphiphilic compounds are in Table A.
TABLE-US-00001 TABLE A CPP and HLB for various amphiphilic compounds Amphiphile CPP V A.sub.0 L.sub.C HLB Phytantriol 0.650 303.5 27.9 16.8 6.36 Monolinolein 1.016 341.0 22.6 14.8 1.02 Glucose stearate 0.456 315.3 31.2 22.2 9.28 Fructose stearate 0.421 315.3 33.8 22.2 9.28
[0194] The active ingredients melatonin and atenolol have been shown to load and release from a monoolein-water liquid crystalline system previously and are expected to be compatible with the ODT of this invention. Atropine, haloperidol, levofloxacin, indomethacin, diazepam, trans retinol, prednisolone, progesterone, hydrocortisone and dexamethasone have been shown to load an release from monoolein and/or phytantriol liquid crystals previously and are expected to be compatible with the ODT of this invention. Irinotecan and paclitaxel has also been released from inverse hexagonal phase previously and are expected to be compatible with the ODT of this invention. Niacin has been shown to load and release from inverse hexagonal phase (see WO 2014/179845). The log P and molecular weight for some active ingredients are in Table B below.
TABLE-US-00002 TABLE B LogP and molecular weight for various active ingredients API MW (g/mole) logP Atorvastatin 558.6 6.36 Oxycodone 315.4 1.03 Adrenaline 333.3 −0.5 to −1.4 Heparin — −13.2 Rosuvastatin 521.6 1.92 Niacin 123.1 0.36
[0195] The molecular weight for some large active ingredients are in Table C below.
TABLE-US-00003 TABLE C Molecular weight of large active ingredients API MW (g/mole) Enoxaparin 3.8-5.0 kDa Dalteparin 5.6-6.4 kDa Low molecular weight Up to 8 kDa heparin
[0196] Palatability Requirements
[0197] Because ODT compositions disintegrate in the mouth of the patient, it is highly preferable for ODT compositions to be palatable, that is, have acceptable organoleptic attributes such as taste, after-taste, “mouthfeel”, disintegration time, and Flavour-Sweetener Balance (FSB). The FSB scale ranges from 4 (low, need to increase), 7 (optimum), and 10 (over-powered, need to reduce).
[0198] Taste and after-taste attributes are defined using a bitterness scale of 1 to 10, wherein 1 is extremely bitter; taste-masking required, 5 is acceptable, and 10 is pleasant tasting. The mouthfeel scale ranges from 1 (very gritty, unacceptable), 5 (non-gritty), and 10 (creamy, smooth).
[0199] The ODTs of the present invention have taste and aftertaste attributes at least about 5, at least about 6, at least about 7, at least about 8, at least about 9, or about 10.
[0200] The ODTs of the present invention have mouthfeel attributes of at least about 5, at least about 6, at least about 7, at least about 8, at least about 9, or about 10. The ODT compositions of the present invention have FSB attributes of about 5 to about 9, or about 6 to about 8, or about 7.
[0201] Optionally, the compositions, self-assembled particles and ODTs of the invention do not include non-API proteins and/or tocopherol. Optionally, the compositions, self-assembled particles and ODTs of the invention do not include poloxamers and/or surfactants other than the amphiphilic compound.
[0202] Preparation of the ODTs of the Invention
[0203] The ODTs of the invention can be prepared in accordance with the methods described in the examples and those in International patent application no PCT/AU2018/050364. The skilled person is able to determine when in the processing the water channel enhancer should be added based on the nature of the ODT (ie active ingredient or sweetener etc) and the properties of the water channel enhancer.
[0204] Compositions of the invention can be prepared by blending the amphiphile with the active ingredient and then the water channel enhancer or vice versa or blending all three ingredients at the same time. If the amphiphile is viscous or semi-solid the amphiphile may require heating before blending.
[0205] A self-assembled structure of the invention can be prepared by contacting an ODT or composition of the invention with a hydrophilic solvent or by adding an amphiphile to hydrophilic solvent containing the active ingredient and water channel enhancer.
REFERENCES
[0206] The text of each of the following references is incorporated by reference into this specification. [0207] Caffrey, M.; Cheng, A., Kinetics of lipid phase changes. Curr. Opin. Struct. Biol. 1995, 5, 548-555. [0208] Chang, C.-M.; Bodmeier, R., Low viscosity monoglyceride-based drug delivery systems transforming into a highly viscous cubic phase. Int. J. Pharm. 1998, 173, 51-60. [0209] Clogston, J.; Rathman, J.; Tomasko, D.; Walker, H.; Caffrey, M., Phase behavior of a monoacylglycerol (Myverol 18-99K)/water system. Chem. Phys. Lipids 2000, 107, 191-220. [0210] Drummond, C. J.; Fong, C., Surfactant self-assembly objects as novel drug delivery vehicles. Current Opinion in Colloid & Interface Science 1999, 4, 449-456. [0211] Fong, W et al, Dynamic formation of nanostructured particles from vesicles via invertase hydrolysis for on-demand delivery, The Royal Society of Chemistry: Electronic Supplementary Material (ESI) for RSC Advances, 2016, S1-S22. [0212] Hyde, S. T., Bicontinuous structures in lyotropic liquid crystals and crystalline hyperbolic surfaces. Current Opinion in Solid State and Materials Science 1996, 1, 653-662. [0213] Israelachvili, J., The science and applications of emulsions—an overview. Colloids Surf. Physico chem. Eng. Aspects, 1994, 91, 1-8. [0214] Kaasgaard, T.; Drummond, C. J., Ordered 2-D and 3-D nanostructured amphiphile self-assembly materials stable in excess solvent. Phys. Chem. Chem. Phys. 2006, 8, 4957-4975) [0215] World Intellectual Property Office publication, WO 2014/179845. [0216] International patent application nos PCT/AU2018/050366, PCT/AU2018/050367 and PCT/AU2018/050364.
[0217] It will be understood that the invention disclosed and defined in this specification extends to all alternative combinations of two or more of the individual features mentioned or evident from the text or drawings. All of these different combinations constitute various alternative aspects of the invention.
EXAMPLES
Example 1—Testing Mixtures of GMO and Excipients/Active Ingredients for Effect on Liquid Crystal Particle Size
[0218] GMO and excipient or active ingredient at various weight proportions were added into small HPLC vials. To ensure adequate mixing, the samples were initially heated above the melting temperature of GMO (>40° C.) and mixed vigorously with a metal spatula. The samples were then mixed via a roller mixer at ˜10 RPM at 40° C. for at least 3 days.
[0219] One week prior to SAXS experiment, 100 mg samples were loaded into a transparent polystyrene 96 well plate (Nunc™) and immersed in 200 μL of PBS buffer (pH 6.8). The samples were stored away from light at ambient temperature, to allow the samples to reach equilibrium.
[0220] Small Angle x-Ray Scattering (SAXS) Setup
[0221] The SAXS/WAXS beamline at Australian Synchrotron, Melbourne, Australia was used to determine the liquid crystalline nanostructure in the samples.
[0222] A custom-designed plate holder was used to mount the samples plate directly onto the SAXS/WAXS beamline. Scans were automated using a pre-loaded set of position variables based on the well positions within the plate, the exposure time was 5 sec.
[0223] Data were obtained at ambient temperature (˜22° C.). The experiments used a beam of wavelength λ=1.033 Å (12.0 keV) and a typical flux of 1.2×1013 photons/s. The 2-D diffraction images were recorded on a Pilatus 1M detector and radially integrated using the in-house software “ScatterBrain”.
[0224] The liquid crystal phase structures were determined by indexing the Bragg peaks according to their corresponding reflection laws (see Hyde, S. T., Bicontinuous structures in lyotropic liquid crystals and crystalline hyperbolic surfaces. Current Opinion in Solid State and Materials Science 1996, 1, 653-662).
[0225] The results in tables 1 to 3 below show GMO forming a bulk phase of complex crystalline particles (ie cubic or hexagonal or a mix of the two) following mixing with a number of ingredients and then emersion of the mix in a hydrophilic solvent (PBS buffer). When the ratio of ingredient to GMO increases, size of the lattice increases, which means the water channel size increases. The percentage w/w of the ingredient to GMO at which the water channel size increases is the “threshold” for the ingredient or the minimum amount of the ingredient required to increase the water channel size.
[0226] For some ingredients, further increases in the proportion of ingredient to GMO also resulted in a change in liquid crystalline structure to a structure with larger water channels eg a V2(Pn3m) to V2(Im3m) change. Other ingredients, did not increase the water channel size of the original liquid crystalline structure but at a threshold concentration did cause a change in liquid crystalline structure to a structure with larger water channels eg a V2(Pn3m) to V2(Im3m) change.
[0227] The inventors have also tested many ingredients that do not increase water channel size or result in a change in liquid crystalline structure.
[0228] Results
1.1 Ingredients Increasing Pn3m Lattice Size
[0229]
TABLE-US-00004 TABLE 1 Tween 80 - V2(Pn3m) lattice size increased at 10-20%. Tested in SBT126 and shown to increase Pn3m lattice size from an ODT. A threshold of 8-10% or about 10%. Transformation in structure by about 20%. Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) 91.9 5 V2(Pn3m) 97.9 10 V2(Pn3m) 106.3, 138.8 20 V2(Pn3m) + V2(Im3m) 137.5, 172.5
TABLE-US-00005 TABLE 2 Sodium cyclamate - V2(Pn3m) lattice size increased at 5-10%. A threshold of 3-5% or about 5%. Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) 93.1 5 V2(Pn3m) 103.3 10 V2(Pn3m) + other 112.7
TABLE-US-00006 TABLE 3 Rosuvastatin - V2(Pn3m) lattice size increased at 5-15%. A threshold of 3-5% or about 5%. Transformation in structure by about 15%. Rosuvastatin was also tested in SBT177 and shown to increase Pn3m lattice size of liquid crystalline particles self-assembled from an ODT. Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) 84.2 5 V2(Pn3m) 109.4 10 V2(Pn3m) 109.4, 130.7 15 V2(Pn3m) + V2(Im3m) 121.8, 148.1
TABLE-US-00007 TABLE 4 Saccharin sodium - V2(Pn3m) lattice size increased at 5-10%. A threshold of 3-5% or about 5%. Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) 95.4 5 V2(Pn3m) 107.1 10 V2(Pn3m) + others 120.1
TABLE-US-00008 TABLE 5 Mg Trisilicate - V2(Pn3m) lattice size increased at 35-50%. A threshold of 30-35% or about 35%. Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) 90.8 5 V2(Pn3m) 95.6 10 V2(Pn3m) + H2? 95.5 20 V2(Pn3m) + H2? 95.5, 55.6 35 V2(Pn3m) + H2? 114.2, 50.5 50 V2(Pn3m) + H2? 116.9, 50.7
TABLE-US-00009 TABLE 6 Ibuprofen lysine - V2(Pn3m) lattice size increased at 5-10%. A threshold of 3-5% or about 5%. Transformation in structure by about 10%. Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) 88.9 5 V2(Pn3m) 104.8 10 V2(Pn3m) + V2(Ia3d) 130.7, 189.4 15 Lα + V2(Ia3d) 80.69, 198.1
TABLE-US-00010 TABLE 7 Atorvastatin - V2(Pn3m) lattice size increased at 10-35%. A threshold of 8-10% or about 10%. Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) 93.1 5 V2(Pn3m) 99.2 10 V2(Pn3m) 104.8 15 V2(Pn3m) 100.6 20 V2(Pn3m) 103.3 25 V2(Pn3m) 101.9 35 V2(Pn3m) 104.8
TABLE-US-00011 TABLE 8 SLS - V2(Pn3m) lattice size increased at 5-10%. A threshold of 3-5% or about 5%. SLS was also tested in SBT137 and shown to increase Pn3m lattice size of liquid crystalline particles self-assembled from an ODT. In ODT SBT125 - 5 mg SLS to 5.4 mg GMO or 48% SLS (based on the weight of SLS and GMO) also caused a shift to Im3m structure of 144-148 nm lattice size. Im3m is a cubic structure with large water channels. Transition at about 50%. Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) 95.4 5 V2(Pn3m) 116.2 10 V2(Pn3m) 132.6
1.2 Ingredients Causing Transition to V2(Im3m) without Simple Water Channel Increase
[0230] Some ingredients caused transition to V2(Im3m) at certain concentrations relative to the GMO, without first expanding the V2(Pn3m) structure.
TABLE-US-00012 TABLE 9 Poloxamer 188 - transition to Im3m by 5% Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) + V2(Im3m) 91.9, 126.0 5 V2(Im3m) 128.1 10 V2(Im3m) 125.9
TABLE-US-00013 TABLE 10 Poloxamer 407 - transition to Im3m by 5% Concentration in GMO (%) Phase structure Lattice size 1 V2(Pn3m) + V2(Im3m) 92.0, 123.8 5 V2(Im3m) 123.8 10 V2(Im3m) 123.8 20 V2(Im3m) + Lα 123.8, 161.1
1.3 Ingredients Having No Effect on the Liquid Crystal Particle Size.
[0231] The tablet excipients Pharmaburst, crospovidone XL, sodium starch glycolate and sorbitol were also tested and found to have minimal impact on the GMO cubic structure formed.
[0232] Pharmaburst 1 to 50% in GMO resulted in V2(Pn3m) of 89-92 nm lattice size.
[0233] Crospovidone 1 to 50% in GMO resulted in V2(Pn3m) of 90-94 nm lattice size.
[0234] Sodium starch glycolate 1 to 50% in GMO resulted in V2(Pn3m) of 87-92 nm lattice size and some V2(Ia3d) of 131-141 nm at 5-20%.
[0235] Sorbitol 1 to 50% in GMO resulted in V2(Pn3m) of 83-92 nm lattice size (the small decrease in size possibly due to a dehydration effect) and at 1% some V2(Ia3d) blended with V2(Pn3m) of about 138 nm.
[0236] Silicon dioxide 1 to 10% in GMO resulted in V2(Pn3m) of 91-93 nm lattice size and some V2(Ia3d) of 138-145 nm blended with the V2(Pn3m) at 2.5-7.5%.
[0237] Magnesium stearate can self-assemble on its own. Some lamella phase was observed at 5-30% with a 50 nm lattice size. The GMO V2(Pn3m) was formed at 1-20% with a 93-95 nm lattice size and H2 formed at 30% with a 62 nm lattice size.
[0238] Sodium chloride was not tested but is expected not to impact lattice size.
[0239] Povidone (poly vinyl pyrrolidone) has been tested in blends of 3 to 40% by weight.
Example 2—Formulations with 5:1 and 3:1 w/w Ratio of GMO to Enoxaparin (SBT265/SBT266) and 5:1 and 7:1 w/w Ratio of GMO to Dalteparin
[0240] Each tablet had 1 mg of active ingredient (ie enoxaparin sodium or dalteparin sodium). Neither tablet includes a water channel enhancer demonstrating that an enhancer is not required to formulate these larger active ingredients.
TABLE-US-00014 TABLE 11 5:1 and 3:1 w/w ratio of GMO to enoxaparin (SBT265/SBT266) Ingredient SBT265 % w/w SBT266 % w/w Function Pharmaburst - (co- 69.87 71.91 Filler, Taste masking, processed mixture of Disintegration agent. Mannitol, Sorbitol Crospovidone & Silicon dioxide) Crospovidone XL 9.88 9.88 Disintegration agent Sodium Chloride 0.25 0.25 Osmotic agent Enoxaparin sodium 1.02 1.02 Drug substance/API Povidone (Poly vinyl 1.50 2.50 Binder pyrrolidone) Glyceryl Monooleate 5.10 3.06 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 4.88 4.88 Disintegration agent Glycolate Colloidal Silicon 1.50 1.50 Glidant Dioxide Magnesium Stearate 1.00 1.00 Lubricant Ethanol N/A * N/A * Solvent * Evaporated during the drying process.
TABLE-US-00015 TABLE 12 5:1 and 7:1 w/w ratio of GMO to dalteparin (SBT267/SBT268) Ingredient SBT265 % w/w SBT266 % w/w Function Pharmaburst - (co- 69.87 80.95 Filler, Taste masking, processed mixture of Disintegration agent. Mannitol, Sorbitol Crospovidone & Silicon dioxide) Crospovidone XL 9.83 10.50 Disintegration agent Sodium Chloride 0.25 0.25 Osmotic agent Dalteparin sodium 1.04 1.04 Drug substance/API Povidone (Poly vinyl 1.50 1.75 Binder pyrrolidone) Glyceryl Monooleate 5.18 7.25 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 4.83 5.50 Disintegration agent Glycolate Colloidal Silicon 1.50 1.66 Glidant Dioxide Magnesium Stearate 1.00 1.10 Lubricant Ethanol N/A * N/A * Solvent * Evaporated during the drying process.
Example 3—Manufacture of the Formulations in Example 2
[0241] Process for wet granulation manufacturing: [0242] Enoxaparin sodium or dalteparin sodium was mixed with melted GMO [0243] Povidone was dissolved in ethanol (granulation solution) [0244] Pharmaburst, Crospovidone and Sodium chloride were granulated with the GMO:enoxaparin sodium or GMO:deltaparin sodium paste and the binder solution [0245] The granules were dried and milled. [0246] The milled granules were blended with the remaining excipients to form the final blend for compression.
[0247] The GMO used melted at about 35° C. Drying after the granulation step often occurred at 40° C., 5° C. above the GMO melting point.
Example 3A—Manufacture of the Formulations in SBT176 (Table 22), SBT227, SBT226, SBT177 (Table 23), SBT233 (Table 23A), SBT 232, SBT122 (Table 14), SBT123 (Table 24)
[0248] Process for wet granulation manufacturing: [0249] Rosuvastatin calcium was dispersed in melted GMO [0250] Povidone, Menthol & Sacharin Sodium was dissolved in ethanol [0251] Pharmaburst, Sodium chloride, Sodium cyclamate Crospovidone were granulated with the Rosuvastatin:GMO suspension and the Povidone solution [0252] The granules were dried and milled. [0253] The milled granules were blended with the remaining excipients to form the final blend for compression.
[0254] The GMO used melted at about 35° C. Drying after the granulation step often occurred at 40° C., 5° C. above the GMO melting point.
[0255] The tablets prepared by this method achieve 100% dissolution within 5 minutes (Dissolution apparatus II, paddles, 50 rpm, 900 ml, Citrate buffer pH 6.6).
[0256] Manufacture of the atorvastatin calcium trihydrate, oxycodone hydrochloride and epinephrine (adrenaline) ODTs also used this method.
Example 4—Disintegration Time for the Formulations in Example 2
[0257] The formulations in Examples 2 were subjected to in vitro disintegration testing. Dissolution testing is not sufficiently precise for this purpose. An active ingredient can take about 5 minutes to be homogeneously dissolved in a 900 ml dissolution testing vessel, therefore, dissolution results taken earlier than 5 minutes are not reliable.
[0258] Disintegration testing was conducted in a basket-rack assembly and in accordance with Appendix XII A. Disintegration of the European Pharmacopoiea edition 9.0 (Ph. Eur. Method 2.9.1). The solvent was water at 37° C.
[0259] The enoxaparin sodium 5:1 formulation (SBT265) in Example 2 disintegrated within 25 to 40 seconds. The enoxaparin sodium 3:1 formulation (SBT266) in Example 2 disintegrated within 25 to 32 seconds.
[0260] The dalteparin sodium 5:1 formulation (SBT267) in Example 2 disintegrated within 30 to 50 seconds. The dalteparin sodium 7:1 formulation (SBT268) in Example 2 disintegrated within 35 to 80 seconds.
[0261] Without being bound by theory, it is thought that the larger size of the dalteparin sodium was responsible for the slightly slower disintegration of the dalteparin tablets, when compared to the enoxaparin tablets. The ODTs with more GMO to API were also slower to disintegrate.
Example 5—Synchrotron Testing to Confirm Formation of Liquid Crystalline Particles from ODTs
[0262] Preparation of Samples Tablets for Simulated Dissolution Study
[0263] For equilibrium samples, the tablets were loaded into a transparent polystyrene 24 well plate (Nunc™) and immersed in PBS buffer (pH 6.8). Three different plates were tested. The first was hydrated the night before testing and stored away from light at ambient temperature overnight prior to SAXS experiment. The second was hydrated about 30 minutes before testing and the third hydrated 1½ to 2 hours before testing.
[0264] Small Angle x-Ray Scattering (SAXS) Setup
[0265] The SAXS/WAXS beamline at Australian Synchrotron, Melbourne, Australia was used to determine the liquid crystalline nanostructure in the samples.
[0266] A custom-designed plate holder was used to mount the samples plate directly onto the SAXS/WAXS beamline. Scans were automated using a pre-loaded set of position variables based on the well positions within the plate, the exposure time was 5 seconds.
[0267] Data were obtained at ambient temperature (˜22° C.). The experiments used a beam of wavelength λ=1.033 Å (12.0 keV) and a typical flux of 1.2×1013 photons/s. The 2-D diffraction images were recorded on a Pilatus 1M detector and radially integrated using the in-house software “ScatterBrain”.
[0268] The liquid crystal phase structures were determined by indexing the Bragg peaks according to their corresponding reflection laws (see Hyde, S. T., Bicontinuous structures in lyotropic liquid crystals and crystalline hyperbolic surfaces. Current Opinion in Solid State and Materials Science 1996, 1, 653-662).
[0269] Each sample was tested in triplicate at 125 times/locations. The results are in table 13 below.
TABLE-US-00016 TABLE 13 Results of synchrotron testing of hydrated ODTs from Example 2 Plate 1 - hydrated the night before synchrotron testing Control - 1 mg Rosuvastatin tablet with no GMO Lalpha 1 mg Daltaparin tablet (5:1) Pm3m, Lalpha 1 mg Daltaparin tablet (7:1) Pm3m, Lalpha 1 mg Enoxaparin tablet (5:1) Pm3m, Lalpha 1 mg Enoxaparin tablet (3:1) Pm3m, Lalpha Plate 2 - hydrated 30 minutes before synchrotron testing Control - Rosuvastatin tablet with no GMO Lalpha Deltaparin tablet (5:1) Pm3m, Lalpha Deltaparin tablet (7:1) Pm3m, Lalpha Enoxaparin tablet (5:1) Pm3m, Lalpha Exnoxaparin tablet (3:1) Pm3m, Lalpha Plate 3 - hydrated 1 to 1½ hours before synchrotron testing Control - Rosuvastatin tablet with no GMO Lalpha Daltaparin tablet (5:1) Pm3m, Lalpha Daltaparin tablet (7:1) Pm3m, Lalpha Enoxaparin tablet (5:1) Pm3m, Lalpha Enoxaparin tablet (3:1) Pm3m, Lalpha
[0270] Each tablet was tested in triplicate.
[0271] The lamella alpha phase for the control in A1 to A3 is expected to have been the result of the magnesium stearate, which forms lamella phase. All tablets with GMO present included the GMO cubic phase. Both the enoxaparin and daltaparin had Pn3m cubic phase with increased lattice size to above 110 nm (ie about 120 to 125 nm).
TABLE-US-00017 TABLE 13A lattice and water channel size for dalteparin and enoxaparin Formulation Pn3m lattice size (nm) Water channel size (Å) Dalteparin 5:1 121.6 59.09 Dalteparin 7:1 119.9 57.76 Enoxaparin 5:1 119.9 57.76 Enoxaparin 3:1 125.2 61.91
Example 6—Effect of Tween 80 on ODT
[0272] Using the process of Example 3 and the ingredients in Table 14 below, an ODT with and without Tween 80 has been developed. The GMO has retained the ability to form cubic liquid crystalline structures. SBT126 and SBT122 are the same 5 mg rosuvastatin formulations with a 1:1 ratio of rosuvastatin to GMO. However, SBT126 includes Tween 80 and has a Pn3m cubic lattice size of 143 nm (water channel size of 76 Å) rather than 123 nm. This indicates that Tween 80 can increase the size of water channels in liquid crystalline particles (ie a colloid dispersion) formed by contacting an ODT with a hydrophilic solvent.
TABLE-US-00018 TABLE 14 SBT122 and SBT126 % w/w for % w/w for Ingredient SBT122 SBT126 Function Mannitol 75.3 75.2 Filler/Carrier Rosuvastatin Calcium 7.3 5.4 Drug substance/API Povidone (Poly vinyl 1.7 — Binder pyrrolidone) Glyceryl Monooleate 7.3 5.4 Bioadhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 5.0 5.0 Disintegration agent Glycolate Colloidal Silicon 2.0 2.0 Glidant Dioxide Magnesium Stearate 1.5 1.5 Lubricant Tween 80 — 2.3 Ethanol N/A * N/A * Solvent * Evaporated during the drying process.
[0273] Disintegration testing was conducted in a basket-rack assembly and in accordance with Appendix XII A. Disintegration of the European Pharmacopoiea edition 9.0 (Ph. Eur. Method 2.9.1). The solvent was water at 37° C.
[0274] The SBT122 disintegrated within 8-10 minutes and achieved 100% dissolution in water within 15 minutes (Dissolution apparatus II, paddles, 50 rpm, 900 ml, Citrate buffer pH 6.6).
Example 7—Effect of SLS on ODT
[0275] Using the process of Example 3 and the ingredients in Table 15 below, an ODT with and without SLS has been developed. The GMO has retained the ability to form cubic liquid crystalline structures. SBT130, SBT 136 and SBT137 are 5 mg rosuvastatin formulations with a 7:1 ratio of GMO to rosuvastatin. However, SBT136 includes SLS and SBT137 includes SLS as well as replacing mannitol with Pharmaburst. SBT 136 has a Im3m cubic lattice (lattice: 211.6 nm; water channels: 93.1 Å.) and SBT137 has a Pn3m cubic lattice size of 143 nm (water channels of 76 Å) rather than the SBT130 V2(Pn3m) cubic lattice size of 123 nm (water channels of 65 Å). This indicates that SLS can increase the size of water channels.
TABLE-US-00019 TABLE 15 SBT130, SBT136 and SBT137 % w/w for % w/w for % w/w for Ingredient SBT130 SBT136 SBT137 Function Pharmaburst — — 60.5 Filler, Taste masking, disintegration agent. Mannitol 61.0 60.5 — Filler/Carrier Rosuvastatin Calcium 1.6 1.6 1.6 Drug substance/API Povidone (Poly vinyl 1.7 1.7 1.7 Binder pyrrolidone) Glyceryl Monooleate 10.5 10.5 10.5 Bioadhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 20.0 20.0 20.0 Disintegration agent Glycolate Colloidal Silicon 2.0 1.5 1.5 Glidant Dioxide Magnesium Stearate 1.5 1.0 1.0 Lubricant SLS — 1.5 1.5 Surfactant, Penetration enhancer Ethanol N/A * N/A * Solvent * Evaporated during the drying process.
[0276] Using the process of Example 3 and the ingredients in Table 16 below, an ODT with SLS has been developed (SBT125). SBT 125 has 5 mg SLS and 5.4 mg of GMO which is equivalent to 48% SLS. The Im3m lattice size identified at 10-60 minutes of testing was 145 nm. At 60-120 min Im3m structure was identified (lattice: 148 nm; water channels 54.3 Å).
TABLE-US-00020 TABLE 16 SBT125 % w/w for Ingredient SBT125 Function Mannitol 73.94 Filler, carrier. Rosuvastatin Calcium 5.4 Drug substance/API Povidone (Poly vinyl 1.7 Binder pyrrolidone) Glyceryl Monooleate 5.4 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 5.0 Disintegration agent Glycolate Colloidal Silicon 2.0 Glidant Dioxide Magnesium Stearate 1.5 Lubricant SLS 5.0 Surfactant, Penetration enhancer. Ethanol N/A * Solvent
[0277] Using the process of Example 3 and the ingredients in Table 17 below, an ODT with SLS and Tartaric Acid has been developed (SBT134). SBT 134 has 5.0 mg SLS, 45 mg Tartaric acid and 36 mg of GMO. The Pn3m lattice size identified at 10-60 minutes of testing was 170 nm (97.57 Å).
TABLE-US-00021 TABLE 17 SBT134 % w/w for Ingredient SBT134 Function Mannitol 46.4 Filler, carrier. Rosuvastatin Calcium 1.6 Drug substance/API Povidone (Poly vinyl 1.7 Binder pyrrolidone) Glyceryl Monooleate 10.5 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 20.0 Disintegration agent Glycolate Colloidal Silicon 2.0 Glidant Dioxide Magnesium Stearate 1.5 Lubricant SLS 1.5 Surfactant, Penetration enhancer. Tartaric acid 13.2 Acidifying agent Ethanol N/A * Solvent
Example 8—Effect of Rosuvastatin on ODT
[0278] Using the process of Example 3 and the ingredients in Table 18 below, an ODT with and without rosuvastatin has been developed. The GMO has retained the ability to form cubic liquid crystalline structures. SBT177 has 5 mg rosuvastatin formulations with a 4:1 ratio of GMO to rosuvastatin. SBT189 is a placebo of SBT176 ie it has 5 mg GMO but no rosuvastatin present. However, SBT177 has a Pn3m cubic lattice size of 143 nm (76 Å) rather than the 126 nm (water channels of 62 Å) for the placebo. This indicates that rosuvastatin can increase the size of water channels.
[0279] Using the process of Example 3 and the ingredients in Table 23 below, an ODT with 20% rosuvastatin to GMO has been developed.
TABLE-US-00022 TABLE 18 SBT177 and SBT189 % w/w for % w/w for Ingredient SBT177 SBT189 Function Pharmaburst - (co- 64.5 74.11 Filler, Taste masking, processed mixture of Disintegration agent. Mannitol, Sorbitol Crospovidone & Silicon dioxide) Crospovidone XL 15.0 Disintegration agent Sodium Chloride 0.3 0.25 Osmotic agent Sodium Cyclamate 0.6 0.60 Sweetener Saccharin Sodium 0.4 0.4 Sweetener Menthol 0.2 0.2 Flavouring agent Rosuvastatin Calcium 1.8 — Drug substance/API Povidone (Poly vinyl 2.5 1.5 Binder pyrrolidone) Glyceryl Monooleate 7.3 5.4 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 5.0 5.0 Disintegration agent Glycolate Colloidal Silicon 1.5 1.5 Glidant Dioxide Magnesium Stearate 1.0 1.0 Lubricant Ethanol N/A * N/A * Solvent * Evaporated during the drying process.
[0280] SBT177 preparation is according to Example 3 except mixing of the rosuvastatin calcium and GMO is for 1-2 minutes.
[0281] The SBT177 tablets weighed 300 mg, had a hardness of 2.4-3.5 kp, friability of 0.1%, 2.65 mm thickness and disintegrated in 60-90 seconds following emersion in a hydrophilic solvent.
[0282] The ODTs of Table 18 achieve 100% dissolution within 5 minutes (Dissolution apparatus II, paddles, 50 rpm, 900 ml, Citrate buffer pH 6.6).
[0283] The tablets were administered sublingually to three different human subjects (one Caucasian male, one Caucasian female and one Asian male) and the speed of tablet disintegration monitored. The SBT177 disintegrated within 20 to 40 seconds of administration for all three subjects.
[0284] The manufacturing of tablets involved mixing of Rosuvastatin with the GMO at its melting point for a short period, until a homogenous dispersion was obtained (approximately 5 minutes) and then mixed with other excipients using a high shear mixer. When combined with the other excipients the temperature of GMO returned to below the GMO melting point and the GMO returned to its semi-solid form.
[0285] The formulation of SBT122 disintegrated within 20-40 seconds of contact with oral mucosa (Basket-rack assembly, Ph. Eur. Method 2.9.1, water at 37° C.). The formulation of Table 4 disintegrated within 40-90 seconds of contact with oral mucosa.
[0286] 12 mm round tablets of this formulation were stability tested at 5° C. for 6 months. Assay of the 5 mg rosuvastatin showed 100.1% at t=0, 100.4% at t=3 months and 97.9% at t=6 months. The formulation was also stability tested at 25° C./60% RH for 6 months. Assay of the 5 mg rosuvastatin showed 100.1% at t=0, 98.6% at t=3 months and 97.6% at t=6 months. In addition, the assay of the tablets showed 0.05% at t=0, 0.31% at t=3 months and 0.49% at t=6 months of rosuvastatin in the lactone form and 0.3% at t=0, 0.31% at t=3 months and 0.36% at t=6 months of 5-oxo-rosuvastatin calcium (TP-13 impurity 1) at following storage at 25° C./60% RH. The assay of the tablets also showed 0.05% at t=0, 0.08% at t=3 months and 0.05% at t=6 months of rosuvastatin in the lactone form and 0.3% at t=0, 0.23% at t=3 months and 0.24% at t=6 months of the 5-oxo-rosuvastatin calcium at both 3 and 6 months at 5° C.
Example 9—Effect of Sodium Bicarbonate on ODT
[0287] Using the process of Example 3 and the ingredients in Table 19 below, an ODT with sodium bicarbonate has been developed (SBT127). SBT 127 has 5 mg Sodium carbonate and 5.4 mg of GMO which is equivalent to 48% Sodium Carbonate. The Pn3m lattice size identified at 10-60 minutes of testing was 175.9 nm (101.6 Å water channel).
TABLE-US-00023 TABLE 19 SBT127 % w/w for Ingredient SBT127 Function Mannitol 73.9 Filler, Carrier Rosuvastatin Calcium 5.4 Drug substance/API Povidone (Poly vinyl 1.7 Binder pyrrolidone) Glyceryl Monooleate 5.4 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 5.0 Disintegration agent Glycolate Colloidal Silicon 2.0 Glidant Dioxide Magnesium Stearate 1.0 Lubricant Sodium bicarbonate 4.7 Alkalizing/Basifying agent Ethanol N/A * Solvent
Example 10—Effect of Menthol on ODT
[0288] Using the process of Example 3 and the ingredients in Table 20 below, an ODT with sodium bicarbonate has been developed (SBT131). SBT 131 has 4 mg menthol and 5.4 mg of GMO which is equivalent to 42.5% menthol. The Pn3m lattice size identified at 10-60 minutes of testing was 141 nm.
TABLE-US-00024 TABLE 20 SBT131 % w/w for Ingredient SBT131 Function Mannitol 74.9 Filler, Carrier Menthol 3.5 Flavouring agent Rosuvastatin Calcium 5.4 Drug substance/API Povidone (Poly vinyl 1.7 Binder pyrrolidone) Glyceryl Monooleate 5.44 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 3.5 Disintegration agent Glycolate Colloidal Silicon 5.4 Glidant Dioxide Magnesium Stearate 1.7 Lubricant Menthol 3.5 Flavouring agent Ethanol N/A * Solvent
Example 11—In Vitro Release Testing
[0289] Release of an active ingredient from and ODT through a mucosal membrane can be tested in vitro.
[0290] Porcine buccal mucosa was freshly isolated from pigs cheeks, mounted between modified Ussing chambers with a donor chamber, receptor chamber and the porcine buccal mucosa in between with a diffusional area of 0.64 cm.sup.2, and incubated in Krebs bicarbonate Ringer buffer (KBR, pH 7.4) for 30 min. The tablet was applied to the porcine buccal mucosa (ie in the donor chamber) and, when necessary, Parafilm was applied to cover the formulation (ie for tablets and for mixtures containing glyceryl monooleate (GMO) and rosuvastatin). The Parafilm prevented the various formulations from detaching from the buccal mucosa. KBR buffer (1.5 mL) was then added to both the donor and receptor chambers, and receptor samples (200 μL) were collected from the receptor chamber at various time points up to 4-5 hours to determine the amount of rosuvastatin that passed through the porcine buccal mucosa to the receptor chamber. 200 μL of fresh KBR was dispensed into the receptor chamber after each collection (to ensure volume balance). Receptor chamber samples were quantified by HPLC.
[0291] Positive control was tested by making solutions of 0.4 and 0.8 mg/1.5 ml active ingredient in KBR solution, equivalent to 1:1 and 7:1 ODT's.
[0292] Rosuvastatin Release Testing
[0293] When a mixture was tested, GMO and rosuvastatin were manually mixed on the day of the experiment. The amount of rosuvastatin in the receptor chamber is in
TABLE-US-00025 TABLE 21 effect of ratio of GMO to API Highest average peak Formulation area result (HPLC) GMO:API 1:1 mixture 22 GMO:API 5:1 mixture 27 GMO:API 7:1 mixture 43
[0294] These results show that including more GMO than API improves the quantity of permeation of the API through the buccal mucosa.
[0295] When the tablets were tested, the tablet of Table 22 was applied to the porcine buccal mucosa whole but the tablet of Table 23 was halved before application to the mucosa to better enable the parafilm to hold the tablet in place. The amount of rosuvastatin in the receptor chamber during testing of the tablets is in
TABLE-US-00026 TABLE 22 ODT with 1:1 GMO to statin (SBT176) Ingredient % w/w Function Pharmaburst - (co- 68.67 Filler, Taste masking, processed mixture of Disintegration agent. Mannitol, Sorbitol Crospovidone & Silicon dioxide) Crospovidone XL 15.00 Disintegration agent Sodium Chloride 0.25 Osmotic agent Sodium Cyclamate 0.60 Sweetener Saccharin Sodium 0.40 Sweetener Menthol (Optional) 0.20 Flavouring agent Rosuvastatin Calcium 5.44 Drug substance/API (micronized) Povidone (Poly vinyl 1.50 Binder pyrrolidone) Glyceryl Monooleate 5.44 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 5.00 Disintegration agent Glycolate Colloidal Silicon 1.50 Glidant Dioxide Magnesium Stearate 1.00 Lubricant Ethanol N/A * Solvent * Evaporated during the drying process.
[0296] A further ODT (SBT227) was prepared using 5% w/w of oxycodone hydrochloride and the same amount of GMO. The crospovidone was also reduced to 10% w/w and the Pharmaburst increased to 69.55% w/w. The ODT (SBT226) had a 1:1 ratio of GMO to oxycodone hydrochloride and 5 mg oxycodone hydrochloride.
TABLE-US-00027 TABLE 23 ODT with 4:1 GMO to statin (SBT177) Ingredient % w/w Function Pharmaburst - (co- 64.47 Filler, Taste masking, processed Disintegration agent. mixture of Mannitol, Sorbitol Crospovidone & Silicon dioxide) Crospovidone XL 15.00 Disintegration agent Sodium Chloride 0.25 Osmotic agent Sodium Cyclamate 0.60 Sweetener Saccharin Sodium 0.40 Sweetener Menthol 0.20 Flavouring agent Rosuvastatin Calcium 1.81 Drug substance/API (micronized) Povidone (Poly vinyl 2.50 Binder pyrrolidone) Glyceryl Monooleate 7.26 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 5.00 Disintegration agent Glycolate Colloidal Silicon 1.50 Glidant Dioxide Magnesium Stearate 1.00 Lubricant Ethanol N/A * Solvent * Evaporated during the drying process.
[0297] A further ODT (SBT233) containing 10 mg atorvastatin was prepared with a 4:1 ratio of GMO to atorvastatin. The formulation is in Table 23A below.
TABLE-US-00028 TABLE 23A ODT with 4:1 GMO to atorvastatin (SBT233) Ingredient % w/w Function Pharmaburst - (co- 58.86 Filler, Taste masking, processed mixture of Disintegration agent. Mannitol, Sorbitol Crospovidone & Silicon dioxide) Crospovidone XL 13.82 Disintegration agent Sodium Chloride 0.22 Osmotic agent Sodium Cyclamate 0.53 Sweetener Saccharin Sodium 0.41 Sweetener Menthol 0.21 Flavouring agent Atorvastatin Calcium 3.19 Drug substance/API Trihydrate Povidone (Poly vinyl 2.50 Binder pyrrolidone) Glyceryl Monooleate 12.76 Bio adhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 5.00 Disintegration agent Glycolate Colloidal Silicon 1.50 Glidant Dioxide Magnesium Stearate 1.00 Lubricant Ethanol N/A * Solvent
[0298] A further ODT (SBT232) was prepared in accordance with Table 23A but using 1.67% w/w of oxycodone hydrochloride and four times as much GMO (6.67% w/w). The crospovidone was also reduced to 10% w/w and the Pharmaburst increased to 65.22% w/w, when compared to the rosuvastatin formulation in Table 23A. The ODT (SBT232) had a 4:1 ratio of GMO to oxycodone hydrochloride and 5 mg oxycodone hydrochloride.
TABLE-US-00029 TABLE 24 SBT123 Ingredient % w/w Function Mannitol 76.0 Filler/Carrier Rosuvastatin Calcium 1.6 Drug substance/API (micronized) Povidone (Poly vinyl 3.4 Binder pyrrolidone) Glyceryl Monooleate 10.5 Bioadhesive/ (GMO) Mucoadhesive agent, Gelling agent, nonionic surfactant, sustained release agent Sodium Starch 5.0 Disintegration agent Glycolate Colloidal Silicon 2.0 Glidant Dioxide Magnesium Stearate 1.5 Lubricant Ethanol N/A * Solvent
TABLE-US-00030 TABLE 25 effect of tablet formulation Highest average peak Formulation area result (HPLC) GMO:API 1:1 mixture 22 GMO:API 7:1 mixture 43 Formulation of Table 14 93 (1:1 ratio) (SBT122) Formulation of Table 24 355 (7:1 ratio) (SBT123)
[0299] The results of Table 25 show that permeation through the buccal mucosa is better with a tablet than with a simple mixture. Also, an increased ratio of GMO to API still improves permeation in the tablet form. The results are shown in
[0300] When tested in the permeability cell, SL tablets were well hydrated and disintegrated within the experimental timeframe. 0.17% or 1.34 ng/ml/min of rosuvastatin in the 1:1 SL tablet used in the clinical trial achieving 125-132% (GS & VO) of the oral Crestor tablet, passed into the receiving chamber. Interestingly 1.41% or 6.1 ng/ml/min more of the 7:1 rosuvastatin passed into the receiving chamber.
[0301] The appearance of oxycodone in the receptor chamber over time is depicted in
[0302] The appearance of atorvastatin in the receptor chamber over time is depicted in
[0303] Adrenaline degrades very quickly upon exposure to air or light. The in vitro release of adrenaline has not yet been tested as a suitable method accounting for the instability of adrenaline needs to be developed. However, in view of the results to date, adrenaline is expected to pass through the porcine oral mucosa in a Ussing chamber. The permeation of the active ingredient from the ODT of the invention was tested for ODTs containing oxycodone and ODTs containing atorvastatin to establish that the ODT functioned to deliver active ingredients having varied Log P values and varied dosages. This time samples were taken from the receiving chamber of the Ussing chamber repeatedly at 0.5, 1, 1.5, 2, 3 and 4 hours to establish not only that the active ingredient permeated the mucosa but that release of the active ingredient was prolonged.
[0304] Enoxaparin In Vitro Release Testing
[0305] KBR buffer (pH 7.4) was used for enoxaparin in vitro release testing. It was prepared according to Table 26.
TABLE-US-00031 TABLE 26 KBR buffer components Compound mM mOsm NaCl 115.385 230.769 KCl 4.155 8.311 NaHCO.sub.3 21.905 43.810 Glucose 12.209 12.209 Hepes 3.987 3.987 MgSO.sub.4•7H.sub.2O 1.163 2.325 NaH.sub.2PO.sub.4•2H.sub.2O 1.596 3.192 CaCl.sub.2•2H.sub.2O 2.503 7.510 Total mOsm 304.602
[0306] In the standard Ussing chambers, pig cheek skin was mounted in between the donor and receptor chambers. The porcine buccal mucosa had a diffusional area of 0.64 cm.sup.2, and was incubated in Krebs bicarbonate Ringer buffer (KBR, pH 7.4) for 30 min. In the modified Ussing Chamber model, enoxaparin and GMO solutions were added to the donor chamber to test if and how much drug would permeate the membrane. Tris or KBR buffer (1.5 mL) was then added to both the donor and receptor chambers, and receptor samples (200 μL) were collected from the receptor chamber at various time points up to 4 hours to determine the amount of enoxaparin that passed through the membrane to the receptor chamber. 200 μL of fresh KBR was dispensed into the receptor chamber after each collection (to ensure volume balance). Receptor chamber samples were quantified by activity-based quantification kits. Test kits from 5-Diagnostics determined the anti-coagulant efficacy of the enoxaparin. Specifically anti-clotting factors II and X were tested. Briefly, frozen samples from each time point were defrosted and shaken gently to ensure homogeneous dispersion. They were then analysed using a 96-well flat bottom well ELISA plate pre-warmed at 37° C. and a Perkin Elmer Ensight Multi-mode plate reader set at an absorbance of 405 nm and at room temperature, with sample absorbance compared to a calibration curve prepared from standards of enoxaparin at pH 7.4.
[0307] The results for the blends were as below:
TABLE-US-00032 TABLE 27 Starting concentrations in each cell Formula 1 (GMO:Enoxaparin 2:1) Cell 1 - Total mix concentration: 0.1341 g/1.5 ml Cell 2 - Total mix concentration: 0.0903 g/1.5 ml Cell 3 - Total mix concentration: 0.0935 g/1.5 ml Formula 2 (GMO:Enoxaparin 4:1) Cell 1 - Total mix concentration: 0.0601 g/1.5 ml Cell 2 - Total mix concentration: 0.0686 g/1.5 ml Cell 3 - Total mix concentration: 0.0631 g/1.5 ml
Formula 1:
[0308]
TABLE-US-00033 TABLE 28 Concentration (mg/mL) in receptor chamber at various time points for Formula 1 1 hr 2 hr 3 hr 4 hr Aver- Aver- Aver- Aver- age Stdev age Stdev age Stdev age Stdev Cell 1 0.000 0.000 3.930 0.156 3.190 0.235 3.485 0.205 Cell 2 0.085 0.007 2.350 0.217 0.475 0.078 2.125 0.177 Cell 3 2.210 0.000 4.305 0.191 4.813 1.167 6.220 0.636
TABLE-US-00034 TABLE 29 Mass (mg) in receptor chamber based on total volume (1.5 mL) in the receptor chamber (mg/mL × 1.5 mL) for Formula 1 1 hr 2 hr 3 hr 4 hr Aver- Aver- Aver- Aver- age Stdev age Stdev age Stdev age Stdev Cell 1 0.000 0.000 5.895 0.233 4.485 0.353 5.228 0.308 Cell 2 0.128 0.011 3.525 0.325 0.713 0.117 3.188 0.265 Cell 3 3.315 0.000 6.458 0.286 7.220 1.750 9.330 0.955
TABLE-US-00035 TABLE 30 Adjusted mass (mg) in receptor chamber, correcting for decrease in mass due to sampling (0.2 mL out of 1.5 mL) at each time point (Mass from previous time point × 0.2/1.5 + current measured mass) for Formula 1 1 hr 2 hr 3 hr 4 hr Aver- Aver- Aver- Aver- age Stdev age Stdev age Stdev age Stdev Cell 1 0.000 0.000 5.895 0.233 5.571 0.353 5.970 0.308 Cell 2 0.128 0.011 3.542 0.325 1.185 0.117 3.345 0.265 Cell 3 3.315 0.000 6.900 0.286 8.140 1.750 10.415 0.955
TABLE-US-00036 TABLE 31 Percentage of enoxaparin in receptor chamber based upon the total enoxaparin sodium added to the donor chamber at the start of the experiment for Formula 1 1 hr 2 hr 3 hr 4 hr Aver- Aver- Aver- Aver- age Stdev age Stdev age Stdev age Stdev Cell 1 0.00% 0.00% 4.40% 0.17% 4.15% 0.26% 4.45% 0.23% Cell 2 0.14% 0.01% 3.92% 0.36% 1.31% 0.13% 3.70% 0.29% Cell 3 3.55% 0.00% 7.38% 0.31% 8.71% 1.87% 11.14% 1.02%
The results of Table 31 are plotted in
Repeated analysis of the results shown in Tables 28-31 indicated that the concentrations observed in cell 2 at 2 h and 3 h were real and reproducible results.
For Formula 1, 4-11% of total Enoxaparin sodium in the donor chamber was measured in the receptor chamber after 4 hours indicating the release and permeability of drug through the membrane.
Formula 2:
[0309]
TABLE-US-00037 TABLE 32 Concentration (mg/mL) in receptor chamber at various time points for Formula 2 1 hr 2 hr 3 hr 4 hr Aver- Aver- Aver- Aver- age Stdev age Stdev age Stdev age Stdev Cell 1 0.000 0.000 0.798 0.145 5.343 0.430 4.850 0.014 Cell 2 0.900 0.099 0.840 0.151 0.808 0.056 2.050 0.071
TABLE-US-00038 TABLE 33 Mass (mg) in receptor chamber based on total volume (1.5 mL) in the receptor chamber (mg/mL × 1.5 mL) for Formula 2 1 hr 2 hr 3 hr 4 hr Aver- Aver- Aver- Aver- age Stdev age Stdev age Stdev age Stdev Cell 1 0.000 0.000 1.196 0.218 8.015 0.646 7.275 0.021 Cell 2 1.350 0.148 1.260 0.227 1.211 0.083 3.075 0.106
TABLE-US-00039 TABLE 34 Adjusted mass (mg) in receptor chamber, correcting for decrease in mass due to sampling (0.2 mL out of 1.5 mL) at each time point (Mass from previous time point × 0.2/1.5 + current measured mass) for Formula 2 1 hr 2 hr 3 hr 4 hr Aver- Aver- Aver- Aver- age Stdev age Stdev age Stdev age Stdev Cell 1 0.000 0.000 1.196 0.218 8.175 0.646 8.365 0.021 Cell 2 1.350 0.148 1.440 0.227 1.403 0.083 3.262 0.106
TABLE-US-00040 TABLE 35 Percentage of enoxaparin in receptor chamber based upon the total enoxaparin sodium added to the donor chamber at the start of the experiment for Formula 2 1 hr 2 hr 3 hr 4 hr Aver- Aver- Aver- Aver- age Stdev age Stdev age Stdev age Stdev Cell 1 0.00% 0.00% 1.99% 0.36% 13.60% 1.07% 13.92% 0.04% Cell 2 1.97% 0.22% 2.10% 0.33% 2.05% 0.12% 4.76% 0.15%
The results of Table 35 are plotted in
After 4 hours, between 4-14% of the Enoxaparin sodium was found in the receptor chamber.
For Formula 2, about 5-14% of total Enoxaparin sodium in the donor chamber was measured in the receptor chamber after 4 hours indicating the release and permeability of drug through the membrane.
Results for both Formula 1 and Formula 2 indicate that a substantial amount of enoxaparin is able to cross the membrane with the GMO enhancer.