RU2816232C1 - Injectable pharmaceutical compositions and use thereof - Google Patents

Abstract

FIELD: medicine; veterinary science.

SUBSTANCE: group of inventions refers to medicine, particularly to veterinary science, and discloses an injectable veterinary pharmaceutical composition containing (a) microspheres containing (a1) from approximately 1 to approximately 40 wt./wt.% in terms of weight of moxidectin microspheres, and (a2) from about 60 to about 99 wt./wt.% in terms of weight of microspheres of polycaprolactone (PCL), and (b) particles of an isoxazoline compound selected from the group consisting of fluralaner, afoxolaner, sarolaner and lotilaner, where the D50 of the volume-weighted average particle size of the isoxazoline compound of formula (I) is from about 65 to about 150 mcm; and c) an aqueous carrier; where microspheres (a) and particles of compound (b) are suspended in aqueous carrier (c). Also disclosed are a method for preparing an injection veterinary composition and a kit.

EFFECT: providing long-term activity, as well as favorable tolerance in the area of injection, storage stability, acceptable viscosity and good bioavailability.

15 cl, 9 dwg, 3 tbl, 10 ex

Description

The present invention relates to injectable veterinary pharmaceutical compositions containing a combination of an isoxazoline compound and a physiologically active macrocyclic lactone.

BACKGROUND OF THE INVENTION

A number of parasites can infect or infect pets, especially companion animals such as cats and dogs. These parasites cause significant concern to both animals and their owners.

Isoxazoline compounds are known in the art, and the preparation of these compounds and their use as antiparasitic agents are described, for example, in US patent application US 2007/0066617 and international patent applications WO 2005/085216, WO 2007/079162, WO 2009/002809, WO 2009/024541, WO 2009/003075, WO 2009/080250, WO 2010/070068, WO 2010/079077 and WO 2011/124998.

Injectable formulations of isoxazoline compounds are described. WO 2015/048371 describes long-acting injectable compositions containing spirocyclic isoxazoline compounds, one biopolymer and at least one carrier, solvent or excipient. WO 2016/138339 describes long-acting injectable formulations containing at least one isoxazoline active substance, a poloxamer and a co-solvent. WO 2016/164487 describes sustained release injectable veterinary formulations containing at least one isoxazoline active ingredient, a pharmaceutically acceptable polymer and a solvent for use against parasites.

US Pat. No. 9,609,869 discloses insecticidal compounds based on isoxazoline derivatives for use in the control of parasites associated with agriculture, horticulture, livestock and companion animals.

US Patent Application Publication No. 2017/0239218 describes long-acting injectable parasite control compositions containing at least one isoxazoline active ingredient, a liquid PEG and/or a neutral oil.

In addition, physiologically active macrocyclic lactones are known to combat parasitic infestations. For example, moxidectin is useful for the prevention and treatment of infections and infestations caused by helminths, nematodes and ectoparasitic arthropods.

Moxidectin is described in US Pat. No. 4,916,154 and EP 0 525 307, and EP 1 197 207 describes microspheres and injectable compositions of moxidectin and their preparation and use.

The essence of the invention

In one aspect, the present invention relates to an injectable veterinary pharmaceutical composition containing

(a) microspheres containing

(a1) from about 1% to about 40% wt./mass. based on the weight of the microspheres, one or more physiologically active macrocyclic lactone(s), and

(a2) from about 60% to about 99% wt./mass. based on the weight of polycaprolactone (PCL) microspheres, and

(b) particles of the isoxazoline compound of formula (I)

formula (I),

Where

R ¹ =halogen, CF ₃ , OCF ₃ , CN,

n=integer from 0 to 3, preferably 1, 2 or 3,

R ² =C ₁ -C ₃ -haloalkyl, preferably CF ₃ or CF ₂ Cl,

T=5-12 membered mono- or bicyclic ring system, which is optionally substituted by one or more Y radicals,

Y=methyl, halomethyl, halogen, CN, NO ₂ , NH ₂ -C=S, or two adjacent Y radicals together form a chain, especially a three- or four-membered chain;

Q=X-NR³R⁴ or a 5-membered N-heteroaryl ring which is optionally substituted by one or more radicals;

X=CH ₂ , CH(CH ₃ ), CH(CN), CO, CS,

R ³ =hydrogen, methyl, haloethyl, halopropyl, halobutyl, methoxymethyl, methoxyethyl, halomethoxymethyl, ethoxymethyl, haloethoxymethyl, propoxymethyl, ethylaminocarbonylmethyl, ethylaminocarbonylethyl, dimethoxyethyl, propynylaminocarbonylmethyl, N-phenyl-N-methylamino, haloethylaminocarbonylmethyl, haloethyls nocarbonylethyl, tetrahydrofuryl, methylaminocarbonylmethyl, ( N,N-dimethylamino)carbonylmethyl, propylaminocarbonylmethyl, cyclopropylaminocarbonylmethyl, propenylaminocarbonylmethyl, haloethylaminocarbonylcyclopropyl,

where Z ^A =hydrogen, halogen, cyano, halogenmethyl (CF ₃ );

R ⁴ =hydrogen, ethyl, methoxymethyl, halogenethoxymethyl, ethoxymethyl, haloethoxymethyl, propoxymethyl, methylcarbonyl, ethylcarbonyl, propylcarbonyl, cyclopropylcarbonyl, methoxycarbonyl, methoxymethylcarbonyl, aminocarbonyl, ethylaminocarbonylmethyl, ethylaminocarbonylethyl, dimethoxyethyl, propynylaminocarbonylmethyl, haloethyl aminocarbonylmethyl, cyanomethylaminocarbonylmethyl or haloethylaminocarbonylethyl; or

R ³ and R ⁴ together form a substituent selected from the group consisting of:

And ; And

c) aqueous carrier;

wherein microspheres (a) and compound particles (b) are suspended in an aqueous vehicle (c).

In another aspect, the present invention relates to a method for preparing such an injectable veterinary composition, comprising the steps of:

i) preparing microspheres (a) from the macrocyclic lactone by solvent evaporation, spinning disk atomization or spray drying, and optionally sieving the resulting product,

ii) obtaining particles (b) of the isoxazoline compound of formula (I),

iii) obtaining an aqueous carrier by dissolving one or more suspending agents and/or one or more wetting agents in water, and

iv) mixing the particles obtained in step i) and the microspheres obtained in step ii) with the aqueous carrier obtained in step iii).

In another aspect, the present invention provides a kit, wherein the kit contains:

(A) a first container containing a mixture of microspheres and particles of the isoxazoline compound of formula (I) as defined above,

(B) a second container containing an aqueous carrier as defined above; And

(C) instructions for reconstituting the microspheres and particles in an aqueous vehicle prior to injection into the animal.

In another aspect, the present invention relates to an injectable veterinary composition, as defined above, for use in the treatment and/or prevention of parasitic infestation in an animal.

Description of drawings

Figure 1: Scanning electron microscope image of microspheres containing moxidectin and polycaprolactone (PCL) obtained by solvent evaporation.

Figure 2: Scanning electron microscope image of microspheres containing moxidectin and polycaprolactone (PCL) obtained by spinning disk atomization (using dichloromethane).

Figure 3: Scanning electron microscope image of microspheres containing moxidectin and polycaprolactone (PCL) obtained by spinning disk atomization (using acetone).

Figure 4: Scanning electron microscope image of microspheres containing moxidectin and polycaprolactone (PCL) obtained by spray drying.

Figure 5: Scanning electron microscope image of microspheres containing 50% fluralaner and PLGA.

Figure 6: Average plasma concentration of fluralaner in dogs after administration of the injectable veterinary composition of the present invention.

Figure 7: Average plasma concentration of moxidectin in dogs after administration of the injectable veterinary composition of the present invention.

Figure 8: Mean plasma fluralaner concentrations in dogs following administration of various fluralaner and moxidectin suspensions.

Figure 9: Average plasma moxidectin concentration in dogs following administration of various fluralaner and moxidectin suspensions.

Detailed description

There is an unmet veterinary need to protect animals, especially companion animals, over the long term with a single injection against most important parasites.

This means long-lasting protection against ectoparasites such as fleas, ticks and mites, and helminth parasites such as heartworms and certain gastrointestinal worms. To maintain or alternatively enhance/improve the prophylactic and/or therapeutic effect of an injectable veterinary pharmaceutical composition, it would be desirable to have a long-acting dosage form containing one or more active substances from different classes of antiparasitic agents.

In particular, what is desirable is an advantageous injectable pharmaceutical composition for veterinary use that allows a single injection to provide effective concentration levels of both classes of active compounds: isoxazoline compounds as well as a macrocyclic lactone, such as moxidectin, in the blood plasma of treated animals over an extended period of time. . In particular, what is desirable is an acceptable injectable composition that allows the effective and safe release of an effective amount of an isoxazoline compound as described above, as well as a macrocyclic compound such as moxidectin, in the combination composition safely and effectively over an extended period of time.

For high efficacy and bioavailability of pharmaceuticals, it may be necessary to achieve high serum levels of active substances in a very short period of time, requiring dosage forms that are released as quickly as possible. However, rapid release is often hampered by the low solubility of the active substances in water.

In addition to the duration of action, there are other desirable features for such injectable veterinary compositions, such as: ease of use (ability to draw into a syringe and ability to resuspend), the ability to sterilize the composition and/or the absence of side effects such as local reaction at the injection site and systemic side effects after introduction.

Another object is to provide a composition that provides stable levels of active ingredients in such dosage forms, especially a macrocyclic compound such as moxidectin.

The present inventors have identified novel injectable compositions in the form of an aqueous suspension, as described herein, with beneficial properties that address one or more of these desirable attributes.

As part of the present invention, the inventors have discovered injectable compositions of a combination of an isoxazoline compound of formula (I) or a compound of formula (II) and one or more macrocyclic lactones, which have long-lasting activity and provide plasma levels in vivo over a long period of time, and which have favorable properties such as injection tolerance, storage stability, acceptable viscosity, syringability over a wide temperature range and good bioavailability.

Aqueous suspension means a composition that contains particles that are mixed, but not dissolved, in an aqueous vehicle (also called a liquid aqueous vehicle or inert medium) containing water used as a solvent (or diluent) in which the active substance constitutes and/or enter. The aqueous carrier may contain some excipients, for example, one or more suspending agents and/or one or more wetting agents.

A particle is a mobile, undissolved, solid that is suspended in a liquid. As used herein, the term covers both crystalline or (alternatively) amorphous solid forms of isoxazoline compounds as particulates and microspheres as particulates.

In one embodiment, the particles of the isoxazoline compound or compound of formula (II) are crystalline.

Preferably, the isoxazoline compound particles are produced in accordance with the method described in WO 2019/091940 A1 or described in the examples.

In another embodiment, the isoxazoline compound particles are present as microspheres, especially PLGA.

A microsphere is a small spherical particle that has a size in the range of 0.1-200 microns and consists of biodegradable and non-biodegradable material and active ingredients, and can be injected.

It has been found that it is advantageous to include a macrocyclic lactone in microspheres prepared from polycaprolactone.

One important aspect of the present invention is an injectable composition comprising (a) microspheres containing (a1) from about 1% to about 40% by weight of one or more physiologically active macrocyclic lactone(s), and (a2) from about 60% to approximately 99% by weight of polycaprolactone (PCL), based on the weight of microspheres with (b) particles of an isoxazoline compound of formula (I) and/or a compound of formula (II), wherein microspheres (a) and compound (b) are suspended in an aqueous vehicle (c) containing one or more wetting agents and/or one or more suspending agents and water.

The injectable veterinary composition contains component (a): microspheres containing (a1) from about 1% to about 40% by weight of one or more physiologically active macrocyclic lactone(s) and (a2) from about 60% to about 99% by weight of polycaprolactone (PCL), both based on the mass of microspheres.

In a preferred embodiment of the invention and/or embodiments thereof, microspheres (a) contain from about 1% by weight to about 40% by weight, preferably from about 2% by weight to about 35% by weight, more preferably from about 3% by weight by weight to about 30% by weight, in particular from about 5% by weight to about 20% by weight, especially from about 8% to 12%, especially 10% by weight of one or more physiologically active macrocyclic lactone(s) (a1 ) based on the mass of microspheres (a).

In a preferred embodiment of the invention and/or embodiments thereof, the present composition contains one or more physiologically active macrocyclic lactone(s) in an amount of from about 0.01% by weight to about 1.0% by weight, more preferably from about 0.05 % by weight to about 0.85% by weight, even more preferably from about 0.1% by weight to about 0.7% by weight, especially preferably from about 0.15% by weight to about 0.5% by weight , especially about 0.17% by weight based on the total weight of the composition.

In one embodiment of the invention and/or embodiments thereof, the one or more physiologically active macrocyclic lactone(s) (a1) include, but are not limited to, avermectins or milbemycins. Such avermectins or milbemycins are known.

In one embodiment of the invention and/or embodiments thereof, the physiologically active macrocyclic lactone(s) (a1) is selected from ivermectin, abamectin, milbemycin oxime, moxidectin, milbemectin, nemadectin, milbemycin-D, doramectin, selamectin, eprinomectin, emamectin and their mixtures.

In a preferred embodiment of the invention and/or its embodiments, the physiologically active macrocyclic lactone(s) (a1) is moxidectin.

In a preferred embodiment of the invention and/or embodiments thereof, microspheres (a) contain from about 60% by weight to about 99% by weight, preferably from about 65% by weight to about 98% by weight, more preferably from about 70% by weight by weight to about 97% by weight, in particular from about 80% by weight to about 95% by weight, especially about 90% by weight (a2) of polycaprolactone (PCL) based on the weight of the microspheres (a).

In a preferred embodiment of the invention and/or embodiments thereof, microspheres (a) contain about 60% by weight, about 65% by weight, about 70% by weight, about 72% by weight, about 75% by weight, about 78% by weight by weight, approximately 80% by weight, approximately 81% by weight, approximately 82% by weight, approximately 83% by weight, approximately 84% by weight, approximately 85% by weight, approximately 86% by weight, approximately 87% by weight, approximately 88% by weight, approximately 90% by weight, approximately 91% by weight, approximately 92% by weight, approximately 93% by weight, approximately 94% by weight, approximately 95% by weight, approximately 96% by weight, approximately 97 % by weight, approximately 98% by weight, or approximately 99% (a2) polycaprolactone (PCL) based on the weight of microspheres (a).

Without wishing to be bound by any theory, it appears that the volume weighted average particle size of the microspheres (a) and the particle size of the isoxazoline compound are important for administration of the present composition. The particle size of any ingredient, particularly the active ingredient, in suspension can affect resuspensibility and syringability.

For this purpose, the particle size must be small enough to prevent compaction or caking and to facilitate resuspension and reconstitution without causing any adverse effect when administered to an animal.

As used herein, particle size data reported herein refers to volume-weighted average particle size as determined by conventional particle sizing techniques well known to those skilled in the art, such as static light scattering (also known as laser diffraction ), image analysis or sifting. Additional discussion of particle size measurement is provided below.

Volume-weighted average particle size can be determined by sieving, microscopy or laser diffraction (Malvern or Sympatec). Volume-weighted average particle size measurements can be performed using a Malvern Mastersizer 2000 with a Hydro 2000G measuring cell or a Horiba LA-910 Laser Scattering Particle Size Distribution Analyzer. Volume-weighted average particle size can be determined using a Sympatec Helos device. Preferably, a Malvern Mastersizer 2000 is used with a Scirocco 2000 dry particle sizing module for particle size analysis.

Particle size distribution describes the relative amounts of particles present depending on size.

The average particle size (D10), which is also referred to as the D10 value of the integral volume distribution, is defined in the context of the present invention as the particle diameter in which 10 percent of the particles by volume have a diameter smaller than the diameter corresponding to the D10 value. Likewise, 90 percent of particles by volume have a diameter greater than the D10 value.

The average particle size (D50), which is also referred to as the D50 value of the integral volume distribution, is defined in the context of the present invention as the particle diameter in which 50 percent of the particles by volume have a diameter smaller than the diameter corresponding to the D50 value. Likewise, 50 percent of particles by volume have a diameter larger than the D50 value.

The average particle size (D90), which is also referred to as the D90 value of the integral volume distribution, is defined in the context of the present invention as the particle diameter in which D90 percent of the particles by volume have a diameter smaller than the diameter corresponding to the D90 value. Likewise, 10 percent of particles by volume have a diameter larger than the D90 value.

In one embodiment or embodiments thereof, the D10 of the volume weighted average particle size distribution of the microspheres (a) is from about 2 μm to about 130 μm, more preferably from about 3 μm to about 115 μm, even more preferably from about 5 μm to about 100 μm, in particular from about 10 μm to about 50 μm.

In one embodiment or embodiments thereof, the D10 of the volume-weighted average particle size distribution of the microspheres (a) is from about 2 μm to about 10 μm, more preferably from about 3 μm to about 8 μm, in particular from about 4 μm to approximately 6 µm.

In one embodiment or embodiments thereof, the D50 of the volume weighted average particle size distribution of the microspheres (a) is from about 8 μm to about 250 μm, more preferably from about 20 μm to about 200 μm, even more preferably from about 40 μm to about 80 μm, in particular from about 50 μm to about 70 μm.

In one embodiment or embodiments thereof, the D90 of the volume-weighted average particle size distribution of the microspheres (a) is from about 25 μm to about 325 μm, more preferably from about 35 μm to about 300 μm, even more preferably from about 50 μm to about 280 μm, in particular from about 100 μm to about 150 μm.

The injectable veterinary composition contains component (b), namely, particles of the isoxazoline compound of formula (I)

Formula (I),

Where

R ¹ = halogen, CF ₃ , OCF ₃ , CN,

n=integer from 0 to 3, preferably 1, 2 or 3,

R ² =C ₁ -C ₃ -haloalkyl, preferably CF ₃ or CF ₂ Cl,

T=5-12 membered mono- or bicyclic ring system, which is optionally substituted by one or more Y radicals,

Y=methyl, halomethyl, halogen, CN, NO ₂ , NH ₂ -C=S, or two adjacent Y radicals together form a chain, in particular a three- or four-membered chain,

Q=X-NR³R⁴ or A 5-membered N-heteroaryl ring which is optionally substituted by one or more radicals,

X=CH ₂ , CH(CH ₃ ), CH(CN), CO, CS,

R ³ =hydrogen, methyl, haloethyl, halopropyl, halobutyl, methoxymethyl, methoxyethyl, halomethoxymethyl, ethoxymethyl, haloethoxymethyl, propoxymethyl, ethylaminocarbonylmethyl, ethylaminocarbonylethyl, dimethoxyethyl, propynylaminocarbonylmethyl, N-phenyl-N-methylamino, haloethylaminocarbonylmethyl, haloethyls nocarbonylethyl, tetrahydrofuryl, methylaminocarbonylmethyl, ( N,N-dimethylamino)carbonylmethyl, propylaminocarbonylmethyl, cyclopropylaminocarbonylmethyl, propenylaminocarbonylmethyl, haloethylaminocarbonylcyclopropyl,

where Z ^A =hydrogen, halogen, cyano, halogenmethyl (CF ₃ );

R ⁴ =hydrogen, ethyl, methoxymethyl, halogenethoxymethyl, ethoxymethyl, haloethoxymethyl, propoxymethyl, methylcarbonyl, ethylcarbonyl, propylcarbonyl, cyclopropylcarbonyl, methoxycarbonyl, methoxymethylcarbonyl, aminocarbonyl, ethylaminocarbonylmethyl, ethylaminocarbonylethyl, dimethoxyethyl, propynylaminocarbonylmethyl, haloethyl aminocarbonylmethyl, cyanomethylaminocarbonylmethyl or haloethylaminocarbonylethyl; or

R ³ and R ⁴ together form a substituent selected from the group consisting of:

And

and/or

particles of a compound of formula (II)

Formula (II).

In a preferred embodiment of the invention and/or embodiments thereof, the isoxazoline compounds of formula (I) and/or the compound of formula (II) also include physiologically acceptable salts, esters, solvents and/or N-oxides thereof. In addition, isoxazoline compounds of formula (I) and/or compounds of formula (II) refer equally to any of their polymorphic forms, enantiomers or stereoisomers.

In a preferred embodiment of the invention and/or embodiments thereof, the isoxazoline compounds of formula (I) and/or the compound of formula (II) may exist in different isomeric forms. Reference to a compound of formula (I) and/or a compound of formula (II) includes all possible isomeric forms of such compound.

In a preferred embodiment of the invention and/or its embodiments, a racemic mixture of isoxazoline compounds of formula (I) and/or a compound of formula (II) can be used as isoxazoline compounds of formula (I) and/or compounds of formula (II), i.e. it contains equal amounts of enantiomers of the corresponding compounds.

In an alternative preferred embodiment of the invention and/or embodiments thereof, the isoxazoline compounds of formula (I) and/or the compound of formula (II) may be the isoxazoline compounds of formula (I) and/or the compound of formula (II), which include enriched stereoisomers of one of the enantiomers of the corresponding compound compared to the racemic mixture.

Also, the isoxazoline compounds of formula (I) and/or the compound of formula (II) may be substantially pure stereoisomers of the corresponding compounds. Particularly preferred is the ( S )-enantiomer of isoxazoline compounds of formula (I) or compounds of formula (II). Such enriched or purified stereoisomeric preparations of isoxazoline compounds of formula (I) and/or compounds of formula (II) can be prepared by methods known in the art. For the chemical preparation of the products of the invention, the person skilled in the art has at his disposal, among other things, the entire contents of the "Chemical Abstracts" and the documents cited therein. Examples are chemical processes using catalytic asymmetric synthesis or separation of diastereomeric salts (see, for example: WO 2009/063910 and JP 2011/051977, respectively).

In a preferred embodiment of the invention and/or embodiments thereof, the present composition contains isoxazoline compounds of formula (I) and/or a compound of formula (II) in an amount of from about 0.1% by weight to about 50.0% by weight, preferably from about 1.0% by weight to about 45% by weight, more preferably from about 2.0% by weight to about 35% by weight, even more preferably from about 3% by weight to about 25% by weight, in particular from from about 5% by weight to about 15% by weight, especially from about 9% by weight to 12% by weight, based on the total weight of the composition.

In a preferred embodiment of the invention and/or embodiments thereof, the present composition contains isoxazoline compounds of formula (I) and/or a compound of formula (II) in an amount of 0.1% by weight, about 0.25% by weight, about 0.5% by weight, approximately 1% by weight, approximately 2% by weight, approximately 3% by weight, approximately 4% by weight, approximately 5% by weight, approximately 6% by weight, approximately 7% by weight, approximately 8% by weight , approximately 9% by weight, approximately 10% by weight, approximately 11% by weight, approximately 12% by weight, approximately 13% by weight, approximately 14% by weight, approximately 15% by weight, approximately 20% by weight or approximately 25% by weight based on the weight of the composition.

In a preferred embodiment of the invention and/or embodiments thereof, in the isoxazoline compound of formula (I), T is selected from

And .

wherein in T-1, T-3 and T-4 the radical Y is preferably hydrogen, halogen, methyl, halomethyl, ethyl or haloethyl.

In a preferred embodiment of the invention and/or embodiments thereof, in the isoxazoline compound of formula (I), Q is selected from

where R ³ , R ⁴ , X and Z ^A are as defined above, and

Z ^B represents

or

Z ^D represents

or

Preferred compounds of formula (I) are given in Table 1:

Table 1:

(R ¹ ) _n R ² R ³ R ⁴ T Y Q Z X 3-Cl, 5Cl CF ₃ CH ₂ CF ₃ H T-2 - Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ CH ₃ H T-2 - Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ CH ₂ OCH ₃ H T-2 - Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-2 - Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-2 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-2 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-2 - Q-1 - C(O) 3-CF ₃ , 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-2 - Q-1 - C(O) 3-CF ₃ , 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-2 - Q-1 - C(O) 3-Cl, 5Cl CF ₃ - T-2 - Q-6 Z ^B -7 3-Cl, 5Cl CF ₃ - - T-2 - Q-7 Z ^B -7 3-Cl, 5Cl CF ₃ - - T-2 - Q-5 Z ^B -7 3-Cl, 5Cl CF ₃ - - T-2 - Q-2 Z ^D -1 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CC H T-3 CH ₃ Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CN H T-3 CH ₃ Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-3 CH ₃ Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-3 CH ₃ Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 4-Cl,
5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 4-Cl,
5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 4-F, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 4-F, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-20 - Q-1 - C(O) 3-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-20 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ CH ₃ T-20 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ CH ₃ T-20 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-20 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-20 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-21 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-21 - Q-1 - C(O) 3-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-21 - Q-1 - C(O) 3-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-21 - Q-1 - C(O) 3-Cl, 5-Cl CF ₃ CH ₂ CH ₂ SCH ₃ H T-21 - Q-1 - C(O) 3-Cl, 4-Cl, 5-Cl CF ₃ C(O) _CH3 H T-22 F Q-1 - CH ₂ 3-Cl, 4-Cl, 5-Cl CF ₃ C(O)CH(CH ₃ ) ₂ H T-22 F Q-1 - CH ₂ 3-Cl, 4-Cl, 5-Cl CF ₃ C(O)-cyclopropyl H T-22 F Q-1 - CH ₂ 3-Cl, 4-F, 5-Cl CF ₃ C(O) _CH3 H T-22 F Q-1 - CH ₂ 3-Cl, 4-Cl, 5-Cl CF ₃ C(O)CH ₂ CH ₃ H T-22 F Q-1 - CH ₂ 3-Cl, 4-F, 5-Cl CF ₃ C(O) _CH3 H T-22 Cl Q-1 - CH ₂ 3-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-1 CH ₃ Q-1 - C(O) 3-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CH ₃ H T-1 CH ₃ Q-1 - C(O) 3-Cl, 5-Cl CF ₃ R ³ -1 (Z) H T-1 CH ₃ Q-1 - C(O) 3-Cl, 5-Cl CF ₃ R ³ -1 (E) H T-1 CH ₃ Q-1 - C(O)

More preferred compounds of formula (I) are shown in Table 2.

Table 2:

(R ¹ ) _n R ² R ³ R ⁴ T Y Q Z X 3-Cl, 5Cl CF ₃ CH ₂ CF ₃ H T-2 - Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ CH ₃ H T-2 - Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ CH ₂ OCH ₃ H T-2 - Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-2 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-2 - Q-1 - C(O) 3-CF ₃ , 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-2 - Q-1 - C(O) 3-Cl, 5Cl CF ₃ - T-2 - Q-6 Z ^B -7 3-Cl, 5Cl CF ₃ - - T-2 - Q-7 Z ^B -7 3-Cl, 5Cl CF ₃ - - T-2 - Q-5 Z ^B -7 3-Cl, 5Cl CF ₃ - - T-2 - Q-2 Z ^D -1 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CC H T-3 CH ₃ Q-1 - C(O) 3-Cl, 5Cl CF ₃ CH ₂ C(O)NHCH ₂ CN H T-3 CH ₃ Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 4-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 4-F, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-3 CH ₃ Q-1 - C(O) 3-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-20 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ CH ₃ T-20 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-20 - Q-1 - C(O) 3-CF ₃ , 5-CF ₃ CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-21 - Q-1 - C(O) 3-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-21 - Q-1 - C(O) 3-Cl, 5-Cl CF ₃ CH ₂ CH ₂ SCH ₃ H T-21 - Q-1 - C(O) 3-Cl, 4-Cl, 5-Cl CF ₃ C(O) _CH3 H T-22 F Q-1 - CH ₂ 3-Cl, 4-Cl, 5-Cl CF ₃ C(O)CH(CH ₃ ) ₂ H T-22 F Q-1 - CH ₂ 3-Cl, 4-Cl, 5-Cl CF ₃ C(O)-cyclopropyl H T-22 F Q-1 - CH ₂ 3-Cl, 4-F, 5-Cl CF ₃ C(O) _CH3 H T-22 F Q-1 - CH ₂ 3-Cl, 4-Cl, 5-Cl CF ₃ C(O)CH ₂ CH ₃ H T-22 F Q-1 - CH ₂ 3-Cl, 4-F, 5-Cl CF ₃ C(O) _CH3 H T-22 Cl Q-1 - CH ₂ 3-Cl, 5-Cl CF ₃ CH ₂ C(O)NHCH ₂ CF ₃ H T-1 CH ₃ Q-1 - C(O) 3-Cl, 5-Cl CF ₃ R ³ -1 (Z) H T-1 CH ₃ Q-1 - C(O) 3-Cl, 5-Cl CF ₃ R ³ -1 (E) H T-1 CH ₃ Q-1 - C(O)

In a particularly preferred embodiment of the invention and/or embodiments thereof, the isoxazoline compound corresponds to formula (Ia)

(Formula Ia)

Where

R ^1a , R ^1b , R ^1c are independently hydrogen, Cl or CF ₃ , more preferably R ^1a and R ^1c are Cl or CF ₃ , and R ^1b is hydrogen,

T represents

or ,

where Y is methyl, Cl, Br, F, CN or C(S)NH ₂ and

Q is as described above.

In another preferred embodiment of the invention and/or its embodiments, R ³ is H and R ⁴ is -CH ₂ -C(O)-NH-CH ₂ -CF ₃ , -CH ₂ -C(O)-NH- CH ₂ -CH ₃ , -CH ₂ -CH ₂ -CF ₃ or -CH ₂ -CF ₃ .

In one embodiment of the invention and/or embodiments thereof, the isoxazoline compound of formula (I) is 4-[5-(3,5-dichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl]-2-methyl - N -[(2,2,2-trifluoroethylcarbamoyl)methyl]benzamide (CAS RN [864731-61-3]; USAN: fluralaner).

In another embodiment of the invention and/or embodiments thereof, the isoxazoline compound of formula (I) is (Z)-4-[5-(3,5-dichlorophenyl)-5-trifluoromethyl-4,5-dihydroisoxazol-3-yl] -N-[(methoxyimino)methyl]-2-methylbenzamide (CAS RN [928789-76-8]).

In one embodiment and/or embodiments thereof, the isoxazoline compound of formula (I) is 4-[5-[3-chloro-5-(trifluoromethyl)phenyl]-4,5-dihydro-5-(trifluoromethyl)-3 -isoxazolyl]-N-[2-oxo-2-[(2,2,2-trifluoroethyl)amino]ethyl]-1-naphthalenecarboxamide (CAS RN 1093861-60-9, USAN: afoxolaner), which is described in WO 2007 /079162.

In one embodiment and/or embodiments thereof, the isoxazoline compound of formula (I) is lotilaner (CAS RN: 1369852-71-0; 3-methyl-N-[2-oxo-2-(2,2,2- trifluoroethylamino)ethyl]-5-[(5S)-5-(3,4,5-trichlorophenyl)-5-(trifluoromethyl)-4H-1,2-oxazol-3-yl]thiophene-2-carboxamide).

In one embodiment and/or embodiments thereof, the isoxazoline compound of formula (I) is sarolaner (CAS RN: 1398609-39-6; 1-(5′-((5 S )-5-(3,5-dichloro -4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′-H-spiro(azetidine-3,1′-(2)-benzofuran)-1-yl)-2 -(methylsulfonyl)ethanone).

In one embodiment and/or embodiments thereof, the isoxazoline compound of formula (I) is 5-[5-(3,5-dichlorophenyl)-4,5-dihydro-5-(trifluoromethyl)-3-isoxazolyl]-3 -methyl-N-[2-oxo-2-[(2,2,2-trifluoroethyl)amino]ethyl]-2-thiophenecarboxamide (CAS RN 1231754-09-8), which is described in WO 2010/070068.

In one embodiment and/or embodiments thereof, the isoxazoline compound of formula (I) is 4-[5-(3,5-dichlorophenyl)-5-(trifluoromethyl)-4H-isoxazol-3-yl]-2-methyl -N-(thiethan-3-yl)benzamide (CAS RN 1164267-94-0), which is described in WO 2009/0080250.

In an alternative embodiment of the invention and/or its embodiments, compound b) comprises particles of a compound of formula (II). The compound of formula (II) is 2-chloro- N- (1-cyanocyclopropyl)-5-[1'-methyl-3'-(1,1,2,2,2-pentafluoroethyl)-4'-(trifluoromethyl) [1,5'-bi- 1H- pyrazol]-4-yl]benzamide; tigolaner (CAS RN 1621436-41-6), which is described in WO 2019/012377.

In one embodiment and/or embodiments thereof, the isoxazoline compound of formula (I) is selected from the group consisting of fluralaner, afoxolaner, lotilaner, sarolaner, and mixtures thereof.

In one embodiment and/or embodiments thereof, the isoxazoline compound of formula (I) is afoxolaner or fluralaner.

In a preferred embodiment of the invention and/or embodiments thereof, the present composition contains afoxolaner or fluralaner, preferably fluralaner, in an amount of from about 0.1% by weight to about 50.0% by weight, preferably from about 1.0% by weight to about 45% by weight, more preferably from about 2.0% by weight to about 35% by weight, even more preferably from about 3% by weight to about 25% by weight, in particular from about 5% by weight to about 15 % by weight, especially from about 9% by weight to 12% by weight, based on the total weight of the composition.

In one embodiment and/or embodiments thereof, the isoxazoline compound of formula (I)) is the ( S )-enantiomer of afoxolaner (esafoxolaner) or the ( S )-enantiomer of fluralaner.

In one embodiment and/or embodiments thereof, the isoxazoline compound of formula (I) is the ( S )-enantiomer of fluralaner.

In one embodiment, the composition of the invention contains particles of a compound of formula (II)

.

It has been found that injectable compositions according to the invention containing particles of an isoxazoline compound of formula (I) and/or a compound of formula (II) with a certain particle size have particularly favorable properties.

In one embodiment of the invention and/or embodiments thereof, the volume weighted average particle size of microspheres (a) containing (a1) one or more physiologically active lactone(s), preferably moxidectin, and a polymer (a2), preferably polycaprolactone, and the weighted average By volume, the particle size of the isoxazoline compound of formula (I) or the compound of formula (II) are in similar ranges.

In one embodiment of the invention and/or embodiments thereof, the volume-weighted average particle size of microspheres (a) containing (a1) one or more physiologically active lactone(s), preferably moxidectin, and (a2), polycaprolactone, and the volume-weighted average size particles of an isoxazoline compound of formula (I) or a compound of formula (II) are not in similar ranges.

The volume-weighted average particle size of the microspheres (a) and the volume-weighted average particle size of the isoxazoline compound of formula (I) and/or the compound of formula (II) are measured in the same manner.

In one embodiment or embodiments thereof, the D10 of the volume weighted average particle size distribution of the isoxazoline compound of formula (I) and/or the compound of formula (II) is from about 5 μm to about 75 μm, more preferably from about 10 μm to about 50 μm, even more preferably from about 15 μm to about 40 μm, in particular from about 20 μm to about 35 μm.

In one embodiment or embodiments thereof, the D50 of the volume weighted average particle size distribution of an isoxazoline compound of formula (I) and/or a compound of formula (II) is from about 65 μm to about 150 μm, more preferably from about 80 μm to about 120 μm, even more preferably from about 85 μm to about 115 μm, in particular from about 90 μm to about 105 μm.

In one embodiment or embodiments thereof, the D90 of the volume weighted average particle size distribution of the isoxazoline compound of formula (I) and/or the compound of formula (II) is from about 100 μm to about 250 μm, more preferably from about 130 μm to about 200 μm, even more preferably from about 150 μm to about 180 μm, in particular from about 155 μm to about 175 μm.

The microspheres (a) and particles (b) are suspended in an aqueous carrier containing one or more suspending agents and/or one or more wetting agents.

The aqueous carrier is present as a liquid aqueous or inert medium. An aqueous carrier may be used as a solvent (or diluent) in which the active agent is formulated or administered.

In a preferred embodiment of the invention and/or its embodiments, the aqueous carrier is water, water and a water-miscible liquid, or a water-miscible liquid.

The water-miscible liquid (also referred to as a cosolvent) may include, but is not limited to, ethanol, isopropanol, benzyl alcohol, glycol ethers (e.g., including, but not limited to, diethylene glycol monoethyl ether (DGME, Transcutol®), butyl diglycol, dipropylene glycol n-butyl ether, ethylene glycol monoethyl ether, ethylene glycol monomethyl ether, dipropylene glycol monomethyl ether, propylene glycol monomethyl ether, propylene glycol monoethyl ether, etc.), liquid polyethylene glycols (PEG) (e.g. PEG 400), propylene glycol, carbonates (e.g. propylene carbonate), cyclic ethers (for example, tetrahydrofuran and dioxane), 2-pyrrolidone, N-methylpyrrolidone, dimethylisosorbitol (DMI), dimethylformamide, acetamide, dimethylacetamide, dimethyl sulfoxide, glycerol, or a mixture thereof.

In one embodiment of the invention and/or embodiments thereof, the water-miscible liquid may be a polar protic solvent, including, but not limited to, an alcohol such as ethanol, isopropanol, or glycol, or a glycol ether.

In one embodiment of the invention and/or embodiments thereof, the water-miscible liquid may be a polar aprotic solvent such as N-methylpyrrolidone, dimethylisosorbitol, dimethylacetamide, dimethyl sulfoxide, or propylene carbonate.

In one embodiment, the pharmaceutical composition is substantially free of water-miscible liquid (cosolvent).

In one embodiment and/or embodiments thereof, the aqueous carrier contains a suspending agent. For purposes of the present invention, a suspending agent can be considered a substance that can be added to a liquid to help suspend or disperse particles and reduce precipitation.

Suspending agents include, but are not limited to, sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone, gum tragacanth, gum arabic, and mixtures thereof.

In one embodiment and/or embodiments thereof, the suspending agent is selected from carboxymethylcellulose, in particular sodium carboxymethylcellulose (NaCMC), polyvinylpyrrolidone, methylcellulose and mixtures thereof.

In one embodiment and/or embodiments thereof, the suspending agent is carboxymethylcellulose, in particular sodium carboxymethylcellulose (NaCMC).

In one embodiment and/or embodiments thereof, the suspending agent is polyvinylpyrrolidone.

In one embodiment and/or embodiments thereof, the suspending agent is methylcellulose.

In one embodiment and/or embodiments thereof, the present composition contains a suspending agent in an amount of from about 0.5% by weight to about 15% by weight, preferably from about 1.0% by weight to about 12.5% by weight , more preferably from about 1.5% by weight to about 10% by weight, in particular from about 2.0% by weight to about 7.5% by weight, based on the total weight of the composition.

In one embodiment and/or embodiments thereof, the aqueous carrier includes a wetting agent.

Wetting agents (sometimes also referred to as dispersants) are compounds useful for reducing surface tension between phases such as liquid-liquid, liquid-gas and liquid-solid.

Wetting agents include, but are not limited to, naturally occurring phosphatides such as lecithin, alkylene oxide condensates of fatty acids such as polyoxyethylene stearate, ethylene oxide condensates of long chain aliphatic alcohols such as heptadecaethyleneoxycetanol, ethylene oxide condensates of partial esters formed from fatty acids and hexitol, such as polyoxyethylene sorbitan monooleate, or condensation products of ethylene oxide with partial esters formed from fatty acids, hexitol anhydrides, such as polyethylene sorbitan monooleate, sorbitan fatty acid esters (Span), polyoxyethylene esters and fatty acids (polysorbates/Tween), castor oil derivatives with polyoxyethylene (Cremaphor) and TPGS (d-α-tocopheryl polyethylene glycol 1000 succinate).

In one embodiment and/or embodiments thereof, the wetting agent is a poloxamer.

Poloxamers are nonionic triblock copolymers containing a central hydrophobic polyoxypropylene chain (poly(propylene oxide)) flanked by two hydrophilic polyoxyethylene chains (poly(ethylene oxide)). Poloxamer 124 (also known as Lutrol L44 or Kollisolv P124) is a polyethylene glycol-block-polypropylene glycol-block-polyethylene glycol. Poloxamer 188, also known as Lutrol F68 or Kolliphor P188, is another case of polyethylene glycol-block-polypropylene glycol-block-polyethylene glycol.

In one embodiment and/or embodiments thereof, the present composition comprises a wetting agent in an amount of from about 0.01% by weight to about 1.0% by weight, preferably from about 0.05% by weight to about 0.9% by weight, more preferably from about 0.1% by weight to about 0.8% by weight, in particular from about 0.2% by weight to about 0.5% by weight, based on the total weight of the composition.

In one preferred embodiment of the invention and/or its embodiments, the present composition further contains one or more physiologically acceptable excipients. Physiologically acceptable excipients are known in the art. For example, they are described in "Gennaro, Remington: The Science and Practice of Pharmacy" ( ^20th Edition, 2000). All such physiologically acceptable excipients must be substantially pure and non-toxic from a physiological and veterinary point of view in the amounts used and must be compatible with the active ingredients.

In one preferred embodiment of the invention and/or its embodiments, physiologically acceptable excipients are contained in microspheres (a) and/or in an aqueous carrier.

In one preferred embodiment of the invention and/or its embodiments, physiologically acceptable excipients are selected from surfactants, buffers, preservatives and mixtures thereof.

Surfactants can be considered substances that reduce the interfacial tension between two phases. Common surfactants include alkyl sulfates (eg, sodium lauryl sulfate), alkyl trimethyl ammonium salts, alcohol ethoxylates, and the like.

Again, these compounds, as well as their amounts, are well known in the art.

In another embodiment of the invention and/or embodiments thereof, the compositions of the invention may include polyoxyl 35 castor oil (Kolliphor® EL) as a surfactant. In other embodiments, the compositions of the invention may include polyoxyl 40 hydrogenated castor oil (Kolliphor® RH 40) or polyoxyl 60 hydrogenated castor oil as a surfactant. The compositions of the invention may also include a combination of surfactants.

Preferably, the surfactant is present in an aqueous carrier.

Buffers are substances that maintain/adjust the pH value of a product. Again, such compounds are well known to one skilled in the art, as well as how to use these compounds. Buffer systems include, but are not limited to, for example, systems selected from the group consisting of acetic acid/acetate, malic acid/malate, citric acid/citrate, tartaric acid/tartrate, lactic acid/lactate, phosphoric acid/phosphate, phosphoric acid/hydrogen phosphate, glycine/glycimate, glutamic acid/glutamates and sodium carbonate/bicarbonate, especially phosphoric acid/sodium phosphate or citric acid/sodium citrate. Preferably, the buffer is present in an aqueous vehicle.

A preservative is a substance that is added to a mixture, in particular to a mixture containing a physiologically active substance, to prevent degradation through microbial growth or through undesirable chemical changes. Benzyl alcohol is preferred. The preservative is present in the aqueous vehicle or microspheres (a).

In one embodiment and/or embodiments thereof, the composition comprises eprinomectin as (a1) the physiologically active macrocyclic lactone, and fluralaner, preferably ( S )-fluralaner, as (b) the isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises milbemycin oxime as (a1) the physiologically active macrocyclic lactone, and fluralaner, preferably ( S )-fluralaner, as (b) the isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises selamectin as (a1) the physiologically active macrocyclic lactone, and fluralaner, preferably ( S )-fluralaner, as (b) the isoxazoline compound of formula (I).

In a particularly preferred embodiment of the invention and/or embodiments thereof, the composition contains moxidectin as (a1) the physiologically active macrocyclic lactone, and fluralaner, preferably ( S )-fluralaner, as (b) the isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises moxidectin in an amount of from about 0.01% by weight to about 1.0% by weight, more preferably from about 0.05% by weight to about 0.85% by weight by weight, even more preferably from about 0.1% by weight to about 0.7% by weight, especially preferably from about 0.15% by weight to about 0.5% by weight, especially about 0.17% by weight, and fluralaner in an amount of from about 0.1% by weight to about 50.0% by weight, preferably from about 1.0% by weight to about 45% by weight, more preferably from about 2.0% by weight to about 35 % by weight, even more preferably from about 3% by weight to about 25% by weight, in particular from about 5% by weight to about 15% by weight, especially from about 9% by weight to 12% by weight calculated for the total weight of the composition.

In one embodiment and/or embodiments thereof, the composition comprises eprinomectin as (a1) the physiologically active macrocyclic lactone, and afoxolaner, preferably ( S )-afoxolaner, as (b) the isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises milbemycin oxime as (a1) the physiologically active macrocyclic lactone, and afoxolaner, preferably ( S )-afoxolaner, as (b) the isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises selamectin as (a1) the physiologically active macrocyclic lactone, and afoxolaner, preferably ( S )-afoxolaner, as (b) the isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises moxidectin as (a1) the physiologically active macrocyclic lactone, and afoxolaner, preferably ( S )-afoxolaner, as (b) the isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises eprinomectin as (a1) a physiologically active macrocyclic lactone, and sarolaner as (b) an isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises milbemycin oxime as (a1) a physiologically active macrocyclic lactone, and sarolaner as (b) an isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises selamectin as (a1) a physiologically active macrocyclic lactone, and sarolaner as (b) an isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises moxidectin as (a1) a physiologically active macrocyclic lactone, and sarolaner as (b) an isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises eprinomectin as (a1) a physiologically active macrocyclic lactone, and lotilaner as (b) an isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises milbemycin oxime as (a1) a physiologically active macrocyclic lactone, and lotilaner as (b) an isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises selamectin as (a1) a physiologically active macrocyclic lactone, and lotilaner as (b) an isoxazoline compound of formula (I).

In one embodiment and/or embodiments thereof, the composition comprises moxidectin as (a1) a physiologically active macrocyclic lactone, and lotilaner as (b) an isoxazoline compound of formula (I).

Injectable compositions generally must be sterilized before administration to an animal. In a preferred embodiment of the invention and/or its embodiments, the microspheres are sterilized, for example, by gamma radiation or electron beam irradiation. Although physiologically active macrocyclic lactones are described to degrade and lose significant biological activity upon irradiation, microspheres (a) can be sterilized for injection by irradiation without negatively affecting the stability of the active ingredients.

In one embodiment and/or embodiments thereof, the present composition may contain an additional therapeutic agent.

In one embodiment and/or embodiments thereof, the present composition may be administered in combination with an additional therapeutic agent. Administration of the additional therapeutic agent may be in the same composition or in a different composition.

The additional therapeutic agent may preferably be a parasiticide or a vaccine, preferably another parasiticide.

The additional therapeutic agent may be selected from the group consisting of probenzimidazoles (eg, febantel, netobimine and thiophanate); benzimidazole derivatives such as thiazole benzimidazole derivatives (eg thiabendazole and cambendazole), carbamate benzimidazole derivatives (eg fenbendazole, albendazole (oxide), mebendazole, oxfendazole, parbendazole, oxybendazole, flubendazole and triclabendazole); imidazothiazoles (eg levamisole and tetramizole); tetrahydropyrimidine (eg morantel and pyrantel), salicylanilides (eg closantel, oxyclozanide, rafoxanide and niclosamide); nitrophenol compounds (for example, nitroxynyl and nitroscanate); benzene disulfonamides (eg clorsulon); pyrazinoisoquinolines (eg praziquantel and epsiprantel); cyclooctadipsipeptides (for example, emodepside); paraherquamides (eg derquantel) and aminoacetonitrile compounds (eg monepantel, AAD 1566); amidine compounds (for example, amidantel and tribendimidine and mixtures thereof.

In one embodiment of the invention and/or its embodiments, the injectable veterinary pharmaceutical composition must be reconstituted before injection. For example, a mixture of (a) microspheres and (b) particles of isoxazoline compound(s) of formula (I) and/or compounds of formula (II) can be reconstituted in an aqueous vehicle prior to injection.

A reconstituted formulation is one wherein a liquid carrier (eg, an aqueous carrier) is contained in one container (eg, a vial) and one or more solid active ingredients (eg, microspheres and particles) are contained in another container, and the contents of the two containers are combined with obtaining a liquid final composition at some point in time before administration to the animal. In the case of the composition of the present invention, an aqueous suspension is formed which includes macrocyclic lactone microspheres and particulate fluralaner.

In another embodiment of the invention and/or its embodiments, the injectable veterinary composition is a ready-to-use composition ready for injection into an animal.

It appears that the present invention relates to injectable compositions containing particles of isoxazoline compound(s) of formula (I) and/or compounds of formula (II), as well as microspheres containing one or more physiologically active macrocyclic lactone(s), such as moxidectin, wherein the compositions provide safety, physical and chemical stability of the composition and active substances and/or a reduced risk of irritation at the injection site.

The physical stability of the injection suspension is particularly important to ensure correct dosing by injecting a homogeneous suspension containing the correct amount of both one or more physiologically active macrocyclic lactone(s) and isoxazoline compound of formula (I) and/or compound of formula (II) and in one general composition. Here, the inventors solve the problem that providing a homogeneous suspension of two different solid components is difficult due to their different densities.

In addition, it is also contemplated to provide a stable composition. In particular, the stability of one or more physiologically active macrocyclic lactone(s) is contemplated, in particular moxidectin, stable compositions of which are particularly problematic.

Moreover, a stable suspension is provided which can be readily resuspended in an aqueous vehicle by gentle shaking without causing foaming or floating or settling of suspended particles which would adversely affect dosing accuracy. Moreover, the final composition to be injected remains physically (and chemically) stable throughout the entire period of use after resuspension/reconstitution.

The microspheres (a) and isoxazoline particles (b) and the present injectable veterinary pharmaceutical composition can preferably be sterilized by gamma radiation or electron beam without significant degradation, i.e. maintaining shelf life without significant loss of biological activity.

Production of injectable veterinary composition

Another aspect of the invention relates to a method for preparing an injectable veterinary composition in accordance with the present invention, comprising the steps of:

i) obtaining particles of an isoxazoline compound of formula (I) and/or a compound of formula (II),

ii) obtaining microspheres (a) by evaporation of the solvent, atomization using a rotating disk or spray drying, and sieving the resulting product,

iii) obtaining an aqueous carrier by dissolving one or more suspending agents and/or one or more wetting agents in water,

iv) adding the particles obtained in step i) and the microspheres obtained in step ii) to the aqueous carrier and vice versa.

With regard to components such as, for example, the physiologically active macrocyclic lactone (a1), the isoxazoline compound of formula (I) and/or the suspending agent, their contents and properties, the same as described above applies.

Particles of isoxazoline compounds of formula (I) and/or compounds of formula (II)

In step i) particles of an isoxazoline compound of formula (I) and/or a compound of formula (II) are obtained. The particles can be obtained, for example, by the method described in WO 2019/091940 A1.

In a preferred embodiment of the invention and/or embodiments thereof, the method comprises initiating crystallization and then maintaining the crystallization temperature in the metastable region by removing, reheating and returning a portion of the solvent, thereby allowing existing crystals to grow larger while minimizing the formation of new smaller crystals.

Solvent includes, but is not limited to, methanol, ethanol, isopropanol (isopropyl alcohol), acetone, ethyl acetate, acetonitrile, dimethyl acetamide (DMA), N-methylpyrrolidone (NMP), dimethyl sulfoxide (DMSO), N,N-diethyl-m-toluamide (DEET), 2-pyrrolidone, g-hexalactone, glycofurol (tetraglycol), methyl ethyl ketone, diethylene glycol monoethyl ether (Transcutol®), dimethyl isosorbitol, macrogol glycerol capryl caprate (Labrasol®), dipropylene glycol monomethyl ether (Dowanol ^TM DPM), glycerol formaldehyde, benzyl alcohol, methanol, polyethylene glycol 200, propylene carbonate, 1-methoxy-2-propyl acetate (Dowanol ^TM PMA), isopropylidene glycerol (Solketal), ethyl alcohol, glycerol triacetate (triacetin), propylene glycol, medium chain triglycerides (Miglyol® 812), ethyl oleate, toluene or mixtures thereof, preferably isopropanol (isopropyl alcohol).

In a preferred embodiment of the invention and/or its embodiments, step b) of the method can be carried out at a temperature of from 60°C to 75°C, preferably at about 65°C.

Alternatively, the particles may be microspheres, such as PLGA microspheres.

Macrocyclic lactone microspheres

In step ii), microspheres (a) are prepared by evaporation of the solvent, atomization using a rotating disk or spray drying, and optionally sieving the resulting product.

In general, methods for solvent evaporation, spinning disk atomization, spray drying, and screening methods are known to one skilled in the art.

Solvent evaporation is one of the most commonly used methods for producing microspheres. It is usually carried out by dissolving the polymer in a volatile organic solvent in which the drug is dissolved. The resulting solution is then added to the aqueous phase containing the surfactant under high homogenization to obtain an emulsion. Finally, the organic solvent is evaporated either by increasing the temperature under reduced pressure or by continuous stirring to obtain a droplet dispersion.

Spinning disk atomization is an encapsulation method that uses the mechanical energy of compressing a liquid film or increasing its kinetic energy for eventual disintegration into droplet form.

Spray drying is a method of producing dry powder from a liquid or slurry by rapid drying with hot gas.

In other words, microspheres (a) can be considered microspheres obtained by incorporating a physiologically active macrocyclic lactone (a1) and optionally other excipients into the polymer (a2), and then forming microparticles from the resulting mixture by various methods, such as the methods indicated above. Alternatively, a mixture of physiologically active macrocyclic lactone (a1) and optionally other excipients with polymer (a2) can be cooled to form a solid, which can then be processed by methods such as grinding, trituration, and the like.

In a preferred embodiment of the invention and/or its embodiments, microspheres (a) are obtained by evaporating the solvent and optionally sieving the resulting product.

To this end, it is preferred that the physiologically active macrocyclic lactone (a1), polymer (a2) and optionally additional physiologically acceptable excipient can be dissolved in a solvent, preferably dichloromethane, and added, preferably to an aqueous solution, preferably polyvinyl alcohol in water. Preferably, the above addition in step ii) is carried out at a temperature of from about 20°C to about 30°C, preferably about 23°C, to obtain a mixture. Preferably, the above addition in step ii) is carried out by mechanical processing such as stirring. Furthermore, step ii) preferably involves isolating the microspheres (a) by sifting the resulting mixture through a sieve, preferably a 75 μm sieve, and then drying, preferably at 23°C and/or under reduced pressure.

In another preferred embodiment of the invention and/or its embodiments, microspheres a) are obtained using a rotating disk and sieving the resulting product.

To this end, it is preferred that the physiologically active macrocyclic lactone (a1), polymer (a2) and optionally additional physiologically acceptable excipient can be dissolved in a solvent, preferably dichloromethane, and applied to a specialized disk rotating at standard parameters known to one skilled in the art. Preferably, the above feeding is carried out at a temperature of from about 0°C to about 20°C. The resulting particles are preferably sieved through at least one sieve, preferably a 420 μm and 250 μm sieve, to obtain microgranules (a). It turns out that "spinning disk" is a way to produce uniform spherical particles with a low particle distribution range, for example, by controlling process parameters such as melting point, flow rate and disk speed.

In another preferred embodiment of the invention and/or its embodiments, microspheres a) are obtained by spray drying and sieving.

To this end, it is preferred that the physiologically active macrocyclic lactone (a1), polymer (a2) and optionally additional physiologically acceptable excipient can be dissolved in a solvent, preferably acetone, and atomized by means of a conventional laboratory spray dryer rotating at standard parameters known to one skilled in the art. . Preferably, the above atomization is carried out at a temperature of from about 15°C to about 25°C, preferably at about 20°C.

Water carrier

In step iii), the aqueous carrier is prepared by dissolving one or more suspending agents and/or one or more wetting agents and optionally other excipients in water. Preferably, one or more suspending agents and/or one or more wetting agents and optionally additional physiologically acceptable excipient(s) are suspended in water or in water and a water-miscible liquid, preferably water. As for water-miscible liquids, the same as described above applies. It is preferable that the above-described dissolution step iii) is carried out at a temperature of from about 20°C to about 30°C, preferably at about 23°C, to obtain a mixture. Preferably, the above-described dissolution in step iii) is carried out by mechanical processing such as stirring.

In step iii), the particles obtained in step i) and the microspheres obtained in step ii) are mixed with the aqueous carrier obtained in step iii). Particles and microspheres can be added to the aqueous carrier or vice versa. Preferably, the particles obtained in step i) and the microspheres obtained in step ii) are added to the aqueous carrier. Preferably, the above addition in step iv) is carried out at a temperature of from about 20°C to about 30°C, preferably at about 23°C, to obtain a mixture. Preferably, the above-described addition in step iv) is carried out by mechanical processing such as stirring.

Another aspect of the invention relates to a kit containing:

(A) the first container containing

- a mixture of microspheres, as described in any of claims 1-14 of the formula, and particles of an isoxazoline compound of formula (I), as described in any of claims 1-14, and/or a compound of formula (II), as described in any of claims 1-14 of the claims,

(B) a second container with an aqueous carrier containing one or more wetting agents and/or one or more suspending agents and water, and

(C) instructions for reconstituting microspheres as described in any of claims 1 to 14, and particles of an isoxazoline compound of formula (I) as described in any of claims 1 to 14, or a compound of formula (II), as described in any of claims 1 to 14, with an aqueous vehicle before injection.

With regard to components such as, for example, the physiologically active macrocyclic lactone (a1), the isoxazoline compound of formula (I) and/or the suspending agent, their contents and properties, the same as described above applies.

In one embodiment of the invention and/or embodiments thereof, the kit further comprises a device for reconstituting and parenterally administering a mixture of the composition from the first and second container to the animal, in particular a syringe.

Method of treatment

Another aspect of the present invention is an injectable veterinary composition of the present invention for use in the treatment and/or prevention of parasitic infestation in an animal. The composition of the present invention is administered parenterally by injection.

Parasitic infestation refers to the presence of parasites in quantities that pose a risk to humans or animals.

In the example, it was shown that the composition according to the invention leads to effective concentrations of moxidectin and fluralaner in the blood plasma over a prolonged period, for more than 6 months. This means that the present injectable composition achieves a sustained release effect of a physiologically active macrocyclic lactone (a1) and an isoxazoline compound of formula (I) or a compound of formula (II) against ectoparasites (such as fleas, ticks and mites), and endoparasites such as worms ( helminths).

The invention also relates to a method of administering and maintaining blood levels of a physiologically active macrocyclic lactone (a1) and an isoxazoline compound of formula (I), especially fluralaner, and/or a compound of formula (II), especially moxidectin and fluralaner, preferably (S)-fluralaner, in animals for an extended period of time and to a method for preventing or treating infections and infestations caused by helminths, nematodes and ectoparasitic arthropods in animals.

In addition, the present injectable composition, which is safe for the animal after injection, has been found to exhibit the desired bioavailability and duration of effectiveness, and does not cause unacceptable side effects, especially not cause unacceptable irritation at the injection site, where the present composition can be administered by subcutaneous or intramuscular injection. In one embodiment, the suspension is administered to the animal via subcutaneous or intramuscular injection.

In addition, it has been found that single administration of such compositions typically provides high activity against one or more parasites (e.g., ectoparasites such as fleas, ticks, or mites), also tending to provide rapid onset of activity, long duration of activity, and /or desired security profiles.

Finally, the new injectable formulation allows the use of these advanced compounds in settings where single injections and repeated administration are undesirable.

The present invention relates to an injectable composition demonstrating at least one of the above-mentioned advantageous properties.

Another aspect of the invention relates to an injectable veterinary composition of the present invention for use in the treatment and/or prevention of parasitic infestation in an animal.

With regard to the injectable veterinary composition, the same as described above applies. The same applies to parasites and parasitic infestations. The term "treatment" as used herein refers to reversing, alleviating, inhibiting parasitic infestation. Prevention/protection is the stopping of the development of a new or impending infestation or infection.

For in vivo administration of a composition according to the invention, an effective amount is synonymous with a "therapeutically or prophylactically effective amount", which is a dose or amount that prevents or treats/alleviates the symptoms and/or signs of a parasitic infection or infection of a treated animal or prevents or reduces parasitic infection number of parasites in and/or on the animal and/or inhibit the development of parasitic infestation in or on the animal, in whole or in part, with an acceptable benefit/risk ratio applicable to any drug treatment.

An endoparasite that seriously harms animals is Dirofilaria immitis , also known as heartworm. The most common hosts are dogs and cats, but other animals such as ferrets and raccoons can also be infected. The parasitic worm is transmitted through the bite of a mosquito that carries heartworm larvae. Adult worms live in the large blood vessels of the lung, causing inflammation of the blood vessels and potentially causing heart damage and early death. With advanced infections, the worms also penetrate the heart.

In a particularly preferred embodiment of the invention, the compositions of the invention are used for the treatment or prevention of Dirofilaria immitis infection. In another embodiment, the compounds and compositions of the invention are used to treat or prevent Dirofilaria repens infection.

Macrocyclic lactones are particularly useful for the control of heartworm ( Dirofilaria spp.) infestations in animals, especially pets or companion animals, especially dogs or cats.

The concentration of the active ingredients in the composition must be sufficient to provide the desired therapeutically or prophylactically effective amount in a volume that is acceptable for injection depending on the animal being treated.

The injectable veterinary composition is administered parenterally, in particular by subcutaneous or intramuscular injection to the animal. Subcutaneous or intramuscular injection can be carried out, for example, by means of a syringe.

The compositions according to the invention have good syringability. The term "syringable" describes a suspension that can be readily withdrawn from an ampoule/vial/container by a needle into a syringe and then injected from the syringe through a needle (e.g., 18-gauge needle) intramuscularly (IM) or subcutaneously (SC). /To).

In a preferred embodiment of the invention and/or its embodiments, the dosage regimen of the injectable veterinary composition is once a month, once every six months or once a year. Preferably, the injectable pharmaceutical compositions can be administered every month, every two months, every three months, every four months, every five months, every six months, every seven months, every eight months, every nine months, every ten months, every eleven months, every twelve months, every 13 months, every 14 months, every 15 months, every 16 months, every 17 months or every 18 months, in particular once every six months or once every twelve months.

Particularly preferred is administration every 6 months. Administration every 12 months is also preferred. In one embodiment, the dosing regimen is at least once every six months or once every twelve months.

This provides long-term protection for animals against both ectoparasites, especially fleas and ticks, and endoparasites, especially heartworms and/or gastrointestinal helminths. Long-term protection against heartworm infection is particularly preferred.

It is advantageous to be able to use the injectable composition of the invention in conjunction with annual vaccination against infectious diseases such as distemper, influenza, rabies, and other vaccines with traditional antigens.

In a preferred embodiment of the invention and/or embodiments thereof, the animal is a pet. A pet (also called a companion animal) can be considered an animal that usually lives in or near the owner's home. Examples of pets include, but are not limited to, dogs, cats, rabbits, guinea pigs, and birds such as budgies and parakeets.

In one embodiment, the animal is a pet. In one embodiment, the animal is a dog or cat. In a more preferred embodiment of the invention and/or embodiments thereof, the animal is a dog.

In a preferred embodiment of the invention and/or embodiments thereof, the isoxazoline compound of formula (I), preferably fluralaner, and/or the compound of formula (II) is administered in an amount of from about 0.01 to about 200 mg/kg body weight of the animal, preferably from about 0.1 to about 100 mg per animal body weight, more preferably from about 0.5 to about 50 mg per kg animal body weight, in particular from about 1 to about 30 mg per kg animal body weight. The total dose may be administered at one time or in divided doses.

In a preferred embodiment of the invention and/or embodiments thereof, the physiologically active macrocyclic lactone, preferably moxidectin, is administered in an amount of from about 0.01 to about 10 mg/kg body weight of the animal, preferably from about 0.1 to about 5 mg/kg body weight animal body.

It has been found that when the present injectable veterinary composition is used to treat and/or prevent parasitic infestation in an animal, the treated animal has minimal irritation at the injection site.

Injection site irritation is the damage that occurs in the injection site and surrounding tissues when an animal is injected with a pharmaceutical composition. This damage may include swelling, skin discoloration, or tissue necrosis. Although some irritation at the injection site is inevitable in some animals, swelling at the injection site greater than 2 x 2 cm, which also persists for more than two to three days, is generally considered unacceptable by veterinarians and pet owners.

Another aspect of the present invention relates to a method of treating and/or preventing a parasitic infestation in an animal, comprising administering to an individual in need thereof a therapeutically effective amount of an injectable veterinary composition in accordance with the present invention or a kit in accordance with the present invention.

Again, with respect to the injectable veterinary composition and kit, the same as described above applies. The same applies to parasites and parasitic infestations.

Thus, the invention relates to a method of treating and/or preventing parasitic infestation, which includes administering to an animal a therapeutically effective amount of an injectable veterinary composition of the present invention or using a kit in accordance with the present invention.

Features of the invention are described in embodiments of the present application; however, for the sake of brevity, not all feature combinations are described verbatim. However, combinations of features described above are expressly considered part of the invention.

EXAMPLES

Example 1 : Production of microspheres containing moxidectin and polycaprolactone (PCL) by solvent evaporation

11.25 g of polyvinyl alcohol was dissolved in 1500 ml of deionized water. Separately, 4.5 g of polycaprolactone and 0.5 g of moxidectin were dissolved in 50 ml of dichloromethane. While stirring the aqueous solution with an overhead stirrer at 330 rpm and 20°C, a solution containing dichloromethane was added dropwise to the aqueous solution. The resulting emulsion was stirred for four hours. Microspheres were separated using a 75-μm sieve and rinsed with deionized water and then air-dried at room temperature.

The SEM image of the resulting microspheres is shown in Figure 1 .

Particle size analysis showed the following values: D10 value: 117.66 µm; D50 value: 190.48 µm; D90 value: 273.43 µm.

Example 2 : Production of microspheres containing moxidectin and polycaprolactone (PCL) by spinning disk atomization using dichloromethane

6.0 g of moxidectin and 54.0 g of polycaprolactone were dissolved in 690 ml of dichloromethane and cooled to 4°C. The solution was applied at 150 g/min to a specialized four-inch (~10 cm) diameter disk rotating at 2500 rpm from a height of 30 feet (9.1 m). The atomization space was cooled to 19°C. The resulting particles were collected and passed through a 420 μm sieve and then through a 250 μm sieve to obtain microspheres.

The SEM image of the resulting microspheres is shown in Figure 2.

Particle size analysis showed the following values: D10 value: 41.19 µm; D50 value: 68.01 µm; D90 value: 113.89 µm.

Example 3 : Production of microspheres containing moxidectin and polycaprolactone (PCL) by spinning disk atomization using acetone

2.5 g of moxidectin and 22.5 g of polycaprolactone were dissolved in 475 ml of acetone and cooled to 4°C. A dedicated stainless steel rotating disk atomization chamber (approximately 4 ft (1.2 m) diameter cone) containing a rotating disk at the top center and a cyclone manifold connected to the bottom of the cone was heated to 35°C. The solution was pumped at 50 g/min onto a three-inch (7.6 cm) diameter stainless steel disk rotating at approximately 5000 rpm. The resulting particles were collected to obtain microspheres.

The SEM image of the resulting microspheres is shown in Figure 3 .

Particle size analysis showed the following values: D10 value: 31.23 µm; D50 value: 58.65 µm; D90 value: 105.44 µm.

EXAMPLE 4 : Production of microspheres containing moxidectin and polycaprolactone (PCL) by spray drying

37.0 g of polycaprolactone and 4.2 g of moxidectin were dissolved in 800 ml of acetone. The mixture was atomized at 6-7 g/min through a 600 µm two-fluid nozzle (3.2 bar nozzle pressure) into a Pro-C-epT 4M8 laboratory spray dryer with an inlet temperature of 25°C, a chamber temperature of 19°C and outlet temperature 19.54°C. The resulting particles were collected to obtain microspheres.

The SEM image of the resulting microspheres is shown in Figure 4 .

Particle size analysis demonstrated the following values: D10- value: 4.54 µm; D50 value: 11.16 µm; D90 value: 33.94 µm.

EXAMPLE 5 : Sterilization of microspheres containing moxidectin and polycaprolactone (PCL)

Samples of microspheres containing moxidectin and polycaprolactone (PCL) and samples of the drug substance moxidectin were placed in 20-ml serum bottles. Microspheres containing moxidectin and polycaprolactone were then irradiated at 15, 20 and 25 kGy via both gamma irradiation and electron beam for sterilization. Microspheres were sterilized at either 5°C or ambient temperature (23°C) with or without nitrogen. Samples were assessed for changes in the assay. % in the analysis is presented as % relative to the analysis without irradiation. The results are presented in Table 1.

CONCLUSION: Moxidectin and PCL microspheres can be sterilized using both irradiation methods. The loss of moxidectin was higher in microspheres sterilized by gamma radiation than in microspheres exposed to E-beam. E-beam irradiation involved the use of a higher dose level, which provided shorter exposure times and reduced potential drug degradation. It was also found that degradation was dose dependent. Flushing the microspheres with nitrogen showed no effect on stability.

Table 1: Results of analysis of samples exposed to electron beam and gamma radiation

Electron beam Gamma radiation Lot of microspheres Dose (kGy) Temperature Purging Analysis Lot of microspheres Dose (kGy) Temperature Purging Analysis 10% moxidectin in PCL microspheres 15 Environment Without purging 95.39 10% moxidectin in PCL microspheres 15-18 Environment Without purging 69.74 Nitrogen in headspace 96.80 Nitrogen in headspace 70.30 5°C Without purging 92.33 5°C Without purging 73.57 Nitrogen in headspace 91.59 Nitrogen in headspace 74.62 20 Environment Without purging 90.09 20-24 Environment Without purging 51.64 Nitrogen in headspace 88.97 Nitrogen in headspace 51.86 5°C Without purging 89.30 5°C Without purging 56.24 Nitrogen in headspace 90.99 Nitrogen in headspace 57.59 25 Environment Without purging 87.85 25-30 Environment Without purging 45.97 Nitrogen in headspace 88.04 Nitrogen in headspace 51.59 5°C Without purging 87.61 5°C Without purging 49.48 Nitrogen in headspace 88.22 Nitrogen in headspace 58.84 The drug substance moxidectin 15 Environment Without purging 96.08 The drug substance moxidectin 15-18 Environment Without purging 97.18 Nitrogen in headspace 94.98 Nitrogen in headspace 97.69 5°C Without purging 93.42 5°C Without purging 95.59 Nitrogen in headspace 93.06 Nitrogen in headspace 95.76 20 Environment Without purging 95.31 20-24 Environment Without purging 95.51 Nitrogen in headspace 95.91 Nitrogen in headspace 96.67 5°C Without purging 94.07 5°C Without purging 94.30 Nitrogen in headspace 94.03 Nitrogen in headspace 93.64 25 Environment Without purging 94.94 25-30 Environment Without purging 95.23 Nitrogen in headspace 95.11 Nitrogen in headspace 96.03 5°C Without purging 93.15 5°C Without purging 93.13 Nitrogen in headspace 93.48 Nitrogen in headspace 95.06

EXAMPLE 6 : Preparation of a liquid aqueous carrier (medium)

Exemplary aqueous media (reconstitution/resuspension media):

Ingredients % w/w Na CMC 2.2 Poloxamer 124 0.11 Sodium phosphate (dibasic dihydrate) 0.77 Benzyl alcohol 2.2 HCl 0.17 WFI QS

Approximately 50% of the water for injection was placed in a container and heated to approximately 70-80°C, and the suspending agent sodium carboxymethylcellulose (NaCMC), Poloxamer 124 was added and homogenized until dissolved. The other ingredients were slowly added and stirred using a magnetic stirrer until dispersion was achieved. Heating was stopped and cold water for injection was added to bring the volume to 10 liters. The pH was adjusted to 4.5-5.5 by adding HCl.

The vehicle was sterilized in an autoclave and the vehicle solution was stored in sterile containers.

Example 7 : Preparation of fluralaner particles

Fluralaner was dissolved at 73-77°C in isopropanol. The solution was filtered and transferred to a crystallizer reactor. The mixture was cooled to 48-52°C and sieved. After standing for 30 min after seeding, the mixture was cooled to 20°C (seed layer). One part of the resulting suspension was transferred to a second reactor (dissolution device) and heated to 70°C until complete dissolution was achieved. The remaining suspension in the crystallizer was heated to 54°C. The contents of the crystallizer were transferred into the solvent and back into the crystallizer with a constant recirculation flow rate of 0.40 BV/h (batch volume (BV) = 10 V → 4.0 V/h), for 2.5 h. After completion of the transfer of the solution from device for dissolution into the crystallizer, the slurry was kept at 54°C for 5 hours. Then very slow stepwise cooling began: from 54°C to 45°C for 6 hours (0.025°C/min) and from 45°C to 0 °C for 15 h (3°C/h). The suspension was kept at -10°C for 1 hour, then the product was isolated by filtration without washing. The wet product was then dried.

Example 8 : Preparation of fluralaner particles in PLGA microspheres

16.0 g fluralaner and 16.0 g 50:50 lactide-glycolide copolymer were dissolved in 285.0 g dichloromethane. The solution was applied at approximately 100 g/min to a customized 4 inch (~10 cm) diameter disk rotating at approximately 3000 rpm. The disk was placed in a specialized polyethylene square (4 × 4 × 4 ft (1.2 × 1.2 × 1.2 m)) box with a lower cone at an angle of 60°. The box was heated with dry air to 54.2-54.7°C and 36.3-38.1°C from the bottom of the box during atomization.

The SEM image of the resulting microspheres is shown in Figure 5.

Example 9 : Preparation of an injectable composition according to the invention and determination of dose accuracy

The vial containing the moxidectin microspheres from Example 3 and the fluralaner particles from Example 7 was reconstituted with 17 ml of aqueous vehicle according to Example 6. During reconstitution, the vial containing the moxidectin microspheres and fluralaner particles was rotated and held horizontally while the vehicle was added.

After all of the vehicle was added, the vial was shaken for 1 minute.

A dose accuracy study was conducted to demonstrate that the correct amount of moxidectin and fluralaner were distributed when dosing 1 ml of the reconstituted composition containing moxidectin and fluralaner.

In this case, the bottle containing the reconstituted composition, as described above, was subjected to one of the following manipulations:

- vigorous shaking by hand for 1, 2 or 3 minutes before taking samples;

- vigorous shaking by hand for 3 minutes and mixing using a syringe 5 times before taking samples; or

- Shake vigorously by hand for 3 minutes, mix using a syringe 5 times and vortex for 20 seconds before taking samples.

After complete suspension of the solid material, six samples were taken at a dose of 1 ml from the vial. Before each dose was taken, the vial was shaken vigorously for 15 seconds to avoid settling between samples.

Each of the six doses was transferred into a separate 50-ml volumetric flask with 10 ml of water: the samples were then diluted to the required volume with a 50/50 mixture of acetonitrile/isopropanol and sonicated for 20 minutes.

These samples were analyzed to determine the concentration of moxidectin.

Additionally, 1 mL of samples were transferred into 25 mL volumetric flasks and diluted with a 40/40/20 acetonitrile/isopropanol/water mixture. Fluralaner concentrations were determined in these diluted samples.

Table 2 below presents the concentration results of moxidectin and fluralaner and shows that any manipulation of the samples resulted in a uniform concentration of the active ingredient in the sample aliquots, thus the injectable composition is easily resuspended and uniform.

Table 2: Moxidectin and fluralaner concentration results

Recovery technique Dose No. Percentage of declared moxidectin content Percentage of declared fluralaner content Shaking by hand for 1 minute 1 103.25 102.68 2 101.25 101.83 3 103.84 106.97 4 101.70 103.52 5 103.45 104.38 6 89.32 90.80 Shaking by hand for 2 minutes 1 101.80 80.44 2 102.26 103.67 3 100.54 102.82 4 101.89 84.95 5 96.62 99.01 6 104.02 107.40 Shaking by hand for 3 minutes 1 99.92 95.36 2 98.29 102.59 3 98.52 95.41 4 98.71 99.43 5 101.21 108.06 6 85.54 88.68 Shake by hand for 3 minutes and mix using a syringe 1 98.43 105.95 2 98.31 104.31 3 95.62 89.74 4 98.17 105.87 5 97.67 104.26 6 c95.90c 103.83 Shake by hand for 3 minutes, mix using a syringe and shake with a vortex mixer 1 93.87 100.86 2 97.78 106.80 3 97.72 105.14 4 99.89 105.86 5 98.19 106.37 6 95.12 104.61

Example 10 : Pharmacokinetic evaluation of microspheres containing moxidectin microspheres in polycaprolactone and microspheres containing fluralaner and lactic-glycolic acid copolymer

Example 10.1:

A ready-to-use injectable suspension of moxidectin-containing microspheres produced according to Example 1 and fluralaner in PLGA microspheres produced as shown in Example 8 was administered subcutaneously as a single dose to three Beagle dogs at a dose of 10 mg/kg body weight (BW) of fluralaner and 0 .17 mg/kg BW moxidectin. Local tolerance of the test sample was assessed at intervals up to 84 days.

Blood samples were taken to determine plasma concentrations of fluralaner and moxidectin on days 1, 3, 4, 5, 7, 10, 14, 21, 28, 35, 43, 49, 56, 70, 84, 98, 112, 126. 140, 154, 168 and 182 trials.

Plasma concentrations of fluralaner and moxidectin are presented in Figures 6 and 7, respectively.

A favorable pharmacokinetic profile demonstrating sustained plasma concentrations of moxidectin was achieved following subcutaneous administration of 10% moxidectin in PCL microspheres prepared by solvent evaporation.

Example 10.2:

Three injection suspensions of moxidectin-containing microspheres produced in Examples 2 and 3 and fluralaner particles produced in Example 7 were reconstituted with 17 ml of aqueous vehicle according to Example 6 and administered once to eight Beagle dogs at a dose of 15 mg/kg BW fluralaner and 0. 17 mg/kg BW moxidectin.

Group 1: Microspheres according to example 3.

Groups 2 and 3 are microspheres according to example 2, in which case group 2 was irradiated at 15 kGy and group 3 was irradiated at 25 kGy.

Local tolerance of the tested samples was assessed over 54 days.

Blood samples were taken to determine plasma concentrations of fluralaner and moxidectin on days 1 (pre-dose and 8 hours post-dose), 2, 4, 6, 8, 11, 15, 22, 29, 36, 43, 50. 57, 71, 85, 99, 113, 127, 141, 155, 169 and 183 trials.

Plasma concentrations of fluralaner and moxidectin are presented in Figures 8 and 9, respectively.

Moxidectin concentration decreased more rapidly for moxidectin in PCL microspheres prepared by spinning disk atomization compared to solvent evaporation.

However, favorable pharmacokinetic profiles demonstrating long-term plasma concentrations of fluralaner were obtained for all formulations.

CONCLUSION: The combination of 10% moxidectin in solvent evaporated PCL microspheres with fluralaner particles provides long-lasting plasma concentrations in dogs for up to six months.

Claims (28)

1. Injectable veterinary pharmaceutical composition containing (a) microspheres containing (a1) from about 1 to about 40% wt./mass. based on the mass of moxidectin microspheres, and (a2) from about 60 to about 99% wt./mass. based on the weight of polycaprolactone (PCL) microspheres, and (b) particles of an isoxazoline compound selected from the group consisting of fluralaner, afoxolaner, sarolaner and lotilaner, wherein the D50 of the volume weighted average particle size distribution of said isoxazoline compound is from about 65 to about 150 μm; And c) water carrier; wherein microspheres (a) and compound particles (b) are suspended in an aqueous vehicle (c). 2. The injectable veterinary composition according to claim 1, wherein the D50 of the volume-weighted average particle size distribution of said isoxazoline compound is from about 80 to about 120 microns. 3. Injectable veterinary composition according to claim 1 or 2, wherein the D50 of the volume-weighted average particle size distribution of the microspheres (a) is from about 80 to about 250 μm. 4. Injectable veterinary composition according to claim 1 or 2, wherein the D50 of the volume-weighted average particle size distribution of the microspheres (a) is from about 20 to about 200 μm. 5. The injectable veterinary composition according to claim 4, wherein the D50 of the volume-weighted average particle size distribution of the microspheres (a) is from about 40 to about 80 μm. 6. The injectable veterinary composition according to claim 4, wherein the D50 of the volume-weighted average particle size distribution of the microspheres (a) is from about 50 to about 70 μm. 7. Injectable veterinary composition according to any one of claims 1 to 6, where the aqueous carrier contains a suspending agent and/or a wetting agent and wherein the suspending agent is selected from sodium carboxymethylcellulose, polyvinylpyrrolidone, methylcellulose and mixtures thereof. 8. Injectable veterinary composition according to claim 7, where the wetting agent is a poloxamer. 9. A method for producing an injectable veterinary composition according to any one of claims 1-8, including the stages: i) producing microspheres of (a) moxidectin by solvent evaporation, spinning disk atomization or spray drying, and optionally sieving the resulting product, ii) obtaining particles (b) of said isoxazoline compound according to claim 1, iii) obtaining an aqueous carrier, in accordance with claims 7, 8, by dissolving one or more suspending agents and/or one or more wetting agents in water, and iv) mixing the particles obtained in step i) and the microspheres obtained in step ii) with the aqueous carrier obtained in step iii). 10. Set, where the set contains: (A) a first container containing a mixture of moxidectin microspheres and particles of said isoxazoline compound according to any one of claims 1 to 6, (B) a second container containing an aqueous carrier as defined in any one of claims 7, 8, and (C) instructions for reconstituting moxidectin microspheres and isoxazoline compound particles in an aqueous vehicle prior to injection into the animal. 11. Injectable veterinary composition according to claims 1-8 for use in the treatment and/or prevention of parasitic infection in an animal. 12. Injectable veterinary composition for use according to claim 11, wherein the injectable veterinary composition is administered by subcutaneous or intramuscular injection to the animal. 13. An injectable veterinary composition for use according to claim 11 or 12, wherein the dosage regimen of the injectable veterinary composition is once every six months or once every twelve months. 14. Injectable veterinary composition for use according to any one of claims 11-13, where the animal is a pet. 15. Injectable veterinary composition for use according to claim 14, where the animal is a dog.

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