Spinosyn formulations for treating ocular and facial disease caused by demodex mites

Abstract

The present invention relates to materials and methods for treating ocular Demodex mite infestations of the eye of a human or animal subject, or for treating eye or skin conditions associated with ocular Demodex infestations, the methods comprising topically administering to the ocular surface (conjunctiva and / or cornea) or other anatomical structure of the eye or area adjacent to the eye a composition comprising one or more spinosyn compounds.

Description

[0001] Cross-references to related applications

[0002] This application claims the benefit of U.S. Provisional Application No. 62 / 711,309, filed July 27, 2018, which is incorporated herein by reference in its entirety, including any figures, tables, nucleic acid sequences, amino acid sequences or figures. Background Technology

[0003] Demodex mites are a genus of ectoparasitic mites belonging to the class Arachnida, which live in or near the hair follicles of mammals. Demodex mites have a symbiotic relationship with humans, parasitizing the hair follicles of human hair follicles without symptoms. Therefore, these mites are part of the normal microbiome in the sebaceous gland region of human skin (Grice EA et al., “The skinmicrobiome: potential for novel diagnostic and therapeutic approaches tocutaneous disease,” Semin Cutan Med Surg., 2014, 33(2):98-103). In healthy skin, mite density is usually low, and this low density does not cause skin diseases. When the density of these mites increases, they can induce facial skin conditions. Patients with ocular Demodex mite infestations often complain of itching, burning, redness, crusting at the base of the eyelashes, blurred vision, and dry eyes.

[0004] Of the more than 100 identified Demodex species, *Demodex folliculorum* and *Demodex brevis* are two of the most common species in humans (Lacey N et al., “Under the lash: Demodex mites in human diseases,” Biochem (Lond.), August 1, 2009, 31(4):2-6). Most individuals settle in childhood and their numbers increase in and around the pilosebaceous unit during puberty as sebum levels increase, and increase with age (Elston DM, “Demodex mites: facts and controversies”, Clin Dermatol, 2010, 28(5):502-504). This may be due to lipases produced by Demodex mites, which allow the mites to use sebum as a food source (Jimenez-Acosta F et al., “Demodex mites contain immunoreactive lipase”, Arch Dermatol, 1989, 125(10):1436-1437). Other sources of nutrition include cell debris and bacteria present in and around the hair follicle sebaceous unit.

[0005] Demodex folliculorum, with its relatively long body, proliferates at the base of the eyelashes (cylindrical dandruff), causing anterior blepharitis. Specifically, Demodex folliculorum consumes the epithelial cells at the hair follicle, causing eyelash dilatation, hypoplasia, and reactive hyperkeratosis, which is often observed as trichiasis and eyelash loss. Demodex brevis, smaller in size, burrows into the sebaceous glands and meibomian glands, causing posterior blepharitis. Demodex brevis obstructs the openings of the meibomian glands, leading to insufficient tear lipid secretion, which may be an inflammatory trigger for dry eye syndrome.

[0006] Demodex mites also carry their own bacterial reservoir, which can cause inflammation of the ocular surface. The bacteria on the mite's surface have been shown to compete with known staphylococcal species, generating an immune response in the human host that leads to inflammation of the eyelids and periorbital epidermis, extending to subepidermal reactions. If the disease is in a chronic and progressive stage, the inflammation may spread to the conjunctiva and cornea, potentially causing infiltrative keratoconjunctivitis, nodular scarring, and corneal neovascularization. Summary of the Invention

[0007] This invention relates to materials and methods for treating ocular Demodex mite infections in humans or animals, or for treating eye or skin diseases associated with ocular Demodex mite infections (e.g., Demodex folliculorum, Demodex brevis, or both).

[0008] One aspect of the invention relates to a method for treating ocular Demodex mite infection in a human or animal subject, or for treating eye or skin diseases associated with ocular Demodex mite infection, comprising topically applying a composition comprising one or more spinosad compounds to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye or areas adjacent to the eye. In some embodiments, the disease is selected from one or more of the following: Demodex mite-induced blepharitis (also known as Demodex blepharitis), Demodex mite-induced ocular erythema, Demodex mite-induced facial erythema, dry eye syndrome, meibomian gland dysfunction, meibomian gland cysts, meibomianitis, follicular inflammation, nonspecific facial dermatitis, infiltrative keratoconjunctivitis, nodular scarring, or corneal neovascularization. This treatment can alleviate one or more signs or symptoms in subjects selected from the following groups: itching, burning, foreign body sensation, crusting and redness of the eyelid margins, blurred vision, cylindrical scales, misaligned eyelashes, trichiasis, eyelash loss, inflammation of the eyelid margins, meibomian gland dysfunction, blepharokeratitis, and blepharokeratitis.

[0009] In some embodiments, the composition is applied topically to an area adjacent to the eye, said area including one or more of the eyelid, eyelid margin, eyelash, eyelash follicle, eyebrow, or eyebrow follicle. In some embodiments, the composition is applied topically to an area adjacent to the eye, said area including sebaceous gland openings of the eyelid (e.g., one or more of the gland of Zeis, gland of Moll, or meibomian gland).

[0010] Another aspect of the invention relates to a topical composition comprising 0.1% to 10% (w / v) of one or more spinosad compounds (also referred to herein as the "topical composition" or "spinosad composition" or "topical spinosad composition") and a method for preparing the topical composition. In some embodiments, the composition is a solution, suspension, ointment, spray, lotion, gel, paste, balm, foam, mousse, scrub, or detergent (e.g., shampoo or soap), cream, or ointment. The soap may be solid, such as in bars, liquid, or semi-solid.

[0011] Another aspect of the invention relates to an article comprising the said topical composition; and a container therein containing the topical composition. In some embodiments, the container is a collapsible or non-collapsible tube, bag, pouch, blister pack, strip, ampoule, medicine bottle, bottle, jar, or wide-mouth jar.

[0012] Another aspect of the invention relates to an eye or face applicator pretreated with or containing a topical composition. In some embodiments, the applicator is a swab, makeup pad, wiping cloth, wiping stick, small towel, sponge, gauze, powder puff, stick, brush, or comb.

[0013] Another aspect of the invention relates to a kit for treating ocular Demodex mite infestations in human or animal subjects, or for treating eye or skin diseases associated with ocular Demodex mite infestations (e.g., Demodex folliculorum, Demodex brevis, or both). The kit comprises a topical composition; and an ocular or facial applicator. Optionally, the applicator may be pretreated with or contain the topical composition. The kit may further include a container holding the composition. The kit may further include instructions (e.g., printed or digital instructions) for treating ocular Demodex mite infestations in human or animal subjects by applying the composition to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye, or to areas adjacent to the eye, or for treating eye or skin diseases associated with ocular Demodex mite infestations. Attached Figure Description

[0014] Figure 1 The chemical structures (Formula (I)) of some embodiments of spinosad compounds that can be used in the compositions and methods of the present invention are shown. Detailed Implementation

[0015] This invention relates to materials and methods for treating ocular Demodex mite infections in human or animal subjects, or for treating eye or skin diseases associated with ocular Demodex mite infections (e.g., Demodex folliculorum, Demodex brevis, or both).

[0016] One aspect of the invention relates to a topical composition for treating ocular Demodex mite infection in human or animal subjects, or for treating eye or skin diseases associated with ocular Demodex mite infection, the composition comprising 0.1% to 10% (w / v) of one or more spinosad compounds.

[0017] The fermentation product identified as A83543 in U.S. Patent No. 5,362,634 is a series of related compounds produced by *Saccharopolyspora spinosa*. These compounds have been referred to as factors or components A, B, C, D, E, F, G, H, J, K, L, M, N, O, P, Q, R, S, T, U, V, W, Y, etc. (see also published international patent applications WO 93 / 09126 and WO 94 / 20518), and are called spinosad A, B, C, etc. Spinosads are a class of macrocyclic lactones with insecticidal activity against a variety of pests. Early identification of spinosad revealed that it has a 5,6,5-tricyclic system, a 12-membered macrocyclic lactone, a neutral sugar (rhamnose) and an amino sugar (forosamine) fusion (see Kirst, HA et al. (1991) "A83543A-D, Unique Fermentation-Derived Tetracyclic Macrolides" Tetrarahedron Letters 32(37):4839-4842). Natural spinosad can be produced by fermentation of cultures from the original culture collection of the Midwest Area Northern Regional Research Center, Agricultural Research Service, United States Department of Agriculture, 1815 North University Street, Peoria, Illinois, 61604, USA, with accession numbers NRRL 18719, 18537, 18538, 18539, 18743, 18395, and 18823. Spinosad is also disclosed in U.S. Patent Nos. 5,496,931, 5,670,364, 5,591,606, 5,571,901, 5,202,242, 5,767,253, 5,840,861, 5,670,486, and 5,631,155. Spinosad has now been found to be useful for controlling arachnids, nematodes, and insects.

[0018] Examples of spinosyns and spinosoids (semi-synthetic analogs) are disclosed in Kirst HA, “The spinosyn family of insecticides: realizing the potential of natural products research,” The Journal of Antibiotics, 2010, 63: 101-111; Legocki J et al., “Contemporary trends in development of active substances possessing the pesticidal properties: spinosyn insecticides,” Pestycydy / Pesticides, 2010, 1-4: 59-71; Sparks TC et al., “Natural products as insecticides: the biology, biochemistry and quantitative structure-activity relationships of spinosyns and spinosoids,” Pest Manag Sci, 2001, 57: 896-905; and Salgado VL et al., “Studies on the Mode of Action of Spinosad: The Internal Effective Concentration and the Concentration Dependence of Neural Excitation, “Pesticide Biochemistry and Physiology,” 1998, 60: 103-110, each of which is incorporated herein by reference in its entirety.

[0019] In various aspects and embodiments of the invention, useful spinosad compounds refer to spinosads and spirocyclospora spinosads that have acaricidal activity against members of the genus Demodex. The terms “acaricidal” and “miticidal” are used interchangeably herein and refer to the ability to kill Demodex mites at any stage of their life cycle, or the ability to interfere with their growth or life cycle in any way that results in an overall reduction of the Demodex mite population. In some embodiments, the Demodex species are *Demodex folliculorum*, *Demodex brevis*, or both. For example, the term “miticidal” includes inhibiting or eliminating the reproductive capacity of the pest, as well as inhibiting the development of the pest from one form to a more mature form, such as the transition between various larval forms or from larva to first nymph or from first nymph to nymph, or from nymph to adult. Furthermore, the term “miticidal” is intended to encompass all stages of the mite life cycle. Therefore, for example, the term includes larval, ovic, and adult killing effects (e.g., see Rather PA et al. Human Demodex Mite: The Versatile Mite of Dermatological Importance, Indian J Dermatol, 2014, Jan-Feb, 59(1):60-66). Techniques for assessing in vitro and in vivo Demodex killing activity are known in the art and can be utilized (see, for example, U.S. Patent No. 8,128,968 (Gao et al.); Gao YY et al., “In vitro and in vivo killing of ocular Demodex by tea tree oil,” Br J Ophthalmol, 2005, 89:1468-1473; and Tighe S et al., “Terpinen-4-ol is the most active ingredient of tea tree oil to kill Demodex mites,” Trans Vis Sci Tech., 2013, 2(7):2).

[0020] In some embodiments, one or more spinosad compounds have Figure 1 The chemical structure of formula (I) shown or its pharmaceutically acceptable salt, wherein R 1 R 2 R 3 R 4 R 5 R 6 R 7 R 8 R 9 R 10 R 11 R12 R 13 R 14 R 15 and R 16 It can be independently selected from the group consisting of the following groups: unsubstituted (null); H; F; Cl; Br; I; OH; CN; (C 1-4 Alkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl; (C 2-4 Alkenyl, such as vinyl, propenyl, butenyl, wherein the double bond may be located at any position on the alkenyl carbon chain, and includes any alkenyl conformational isomer; alkynyl; aralkyl; alkylaryl; haloalkyl; heteroalkyl; aryl; heterocyclic; cycloalkyl; cycloalkenyl; cycloalkynyl; hydroxyalkyl; aminoalkyl; amino; alkylamino; arylamino; dialkylamino; alkylarylamino; diarylamino; acylamino; hydroxyl; thiol; thioalkyl; alkoxy; alkylthio; alkoxyalkyl; aryloxy; arylalkoxy; acyloxy; nitro; carbamoyl; trifluoromethyl; phenoxy; benzyloxy; phosphonic acid; phosphate ester; sulfonic acid (-SO3H); sulfonate; sulfonamide; alkylaryl; arylalkyl; carbamoyl; amino; alkylamino; arylamino; dialkylamino; alkylarylamino; diarylamino; alkylthio; heteroalkyl; alkyltriphenylphosphine; heterocyclic; ketone (=O); ether (-OR) 17 ); and esters (-COOR) 18 and -OC(=O)R 18 );

[0021] Where R 5 and R 7 It could be a double bond within a cyclopentane ring;

[0022] Where R 11 and R 13 It could be a double bond within the cyclohexane ring;

[0023] Where R 17 It can be independently selected from the group consisting of the following groups: (C 1-4 Alkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl; (C 2-4 Alkenyl groups, such as vinyl, propenyl, butenyl, wherein the double bond may be located at any position on the alkenyl carbon chain and includes any alkenyl conformational isomer; and ynyl groups;

[0024] Where R 18 It can be independently selected from the group consisting of the following groups: (C 1-4 Alkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl; (C 2-4Alkenyl groups, such as vinyl, propenyl, butenyl, wherein the double bond may be located at any position on the alkenyl carbon chain and includes any alkenyl conformational isomer; and ynyl groups.

[0025] In some embodiments, the composition comprises spinosad A, spinosad D, or a combination thereof. The chemical structures of these and many other spinosad compounds have been known for some time. Spinosad A has the following substituted atoms or groups in the structure of formula (I): R 1 R 2 =(C 1-4 )alkyl, i.e. methyl; R 3 =Ketone (=O); R 4 = No substitution; R 5 and R 7 It is a double bond within the cyclopentane ring; R 8 R 9 R 10 =-OR 17 That is, -OCH3; R 11 and R 13 It is a double bond within the cyclohexane ring; R 12 and R 14 For H; R 15 = No substitution; and R 16 =Ketone (=O). Spinosad D has the following substituent atoms or groups in the structure of formula (I): R 1 R 2 =(C 1-4 )alkyl, i.e. methyl; R 3 =Ketone (=O); R 4 = No substitution; R 5 and R 7 It is a double bond within the cyclopentane ring; R 8 R 9 R 10 =-OR 17 That is, -OCH3; R 11 and R 13 It is a double bond within the cyclohexane ring; R 12 For (C) 1-4 )alkyl, i.e. methyl; R 14 For H; R 15 = No substitution; and R 16 =Ketone (=O).

[0026] In some embodiments, spinosad A and spinosad D are present in the composition at a ratio of about 18:2 to about 16:4. In some embodiments, spinosad A and spinosad D are present in the composition at a ratio of about 17:3.

[0027] The composition can be formulated in any dosage form suitable for topical application to the ocular surface (conjunctiva and / or cornea), other anatomical structures of the eye, and / or areas adjacent to the eye, such as eyelids, eyelid margins, eyelashes, eyelash follicles, eyebrows, or eyebrow follicles, or areas surrounding the openings of eyelid sebaceous glands (e.g., one or more of Zei's glands, Mohr's glands, or meibomian glands). In some embodiments, the composition is a solution, suspension, ointment, spray, lotion, gel, paste, balm, foam, mousse, scrub, or cleanser (e.g., shampoo or soap), cream, or ointment. Soap can be solid, such as in bars, liquid, or semi-solid forms.

[0028] One or more spinosad compounds can be mixed with one or more other components in the composition using methods for preparing dermatologically acceptable and / or ophthalmologically acceptable compositions. The components of the composition can be pre-sterilized, or the composition can be sterilized during or at the end of production.

[0029] In some embodiments, the composition can be formulated as a cosmetic product (e.g., a cosmetic product), such as foundation, mascara, eyeshadow, or eyeliner. The composition can also be in the form of a face mask or makeup remover.

[0030] In various embodiments, the compositions covered herein comprise pharmaceutically acceptable excipients, such as those listed in Remington: the Science and Practice of Pharmacy 866-885 (Alfonso R. Gennaro, ed., 19th ed., 1995; Ghosh, TK et al., Transdermal and Topical Drug Delivery Systems, which are incorporated herein by reference in their entirety), including but not limited to protective agents, adsorbents, modifiers, emollients, preservatives, antioxidants, humectants, buffers, solubilizers, skin penetrants, and surfactants.

[0031] Preferably, the spinosad composition comprises a dermatologically acceptable and / or ophthalmologically acceptable base. As used herein, a "dermatologically acceptable base" refers to one or more non-lotion excipients that, when applied to the skin near the eyes at effective concentrations, will not cause irritation, inflammation, pain, or other harm to the skin. As used herein, an "ophthalmologically acceptable base" refers to one or more non-lotion excipients that, when applied topically to the ocular surface at effective concentrations, will not irritate or otherwise harm the ocular surface.

[0032] Protective agents and adsorbents include, but are not limited to, powders, zinc stearate, PVC foam, dimethicone, silicone, zinc carbonate, aloe vera gel and other aloe vera products, vitamin E oil, allantoin, glycerin, petrolatum, and zinc oxide. Soothing agents include, but are not limited to, benzoin, hydroxypropyl cellulose, hydroxypropyl methylcellulose, and polyvinyl alcohol. Emollients include, but are not limited to, animal and vegetable oils, myristol, alum, and aluminum acetate. Preservatives include, but are not limited to, chlorine dioxide, quaternary ammonium compounds such as benzalkonium chloride, benzyl chloride, bromophthalium trimethylammonium, dequinoline chloride, and cetylpyridinium chloride; mercury agents such as phenylmercuric nitrate, phenylmercuric acetate, and thimerosal; alcohol reagents such as chlorobutanol, phenethyl alcohol, and benzyl alcohol; antimicrobial esters such as esters of p-hydroxybenzoic acid; and other antimicrobial agents such as chlorhexidine, chlorocresol, benzoic acid, and polymyxin. Suitable antioxidants include, but are not limited to, ascorbic acid and its esters, sodium bisulfate, butylated hydroxytoluene, butylated hydroxyanisole, tocopherol, and chelating agents such as EDTA and citric acid. Suitable moisturizers include, but are not limited to, glycerin, sorbitol, polyethylene glycol, urea, and propylene glycol. Suitable solubilizers include, but are not limited to, quaternary ammonium chloride, cyclodextrin, benzyl benzoate, lecithin, and polysorbate. Suitable skin penetrants include, but are not limited to, ethanol, isopropanol, octylphenyl polyethylene glycol, oleic acid, polyethylene glycol 400, propylene glycol, N-decylmethyl sulfoxide, fatty acid esters (e.g., isopropyl myristate, methyl laurate, glyceryl monooleate, and propylene glycol monooleate); and N-methylpyrrolidone.

[0033] The composition may contain one or more excipients. In some embodiments, the composition contains one or more excipients selected from petrolatum, mineral oil, propylparaben, methylparaben, lanolin, chlorobutanol, water, lanolin alcohol, sodium thiosulfate, sodium dihydrogen phosphate, phenylmercuric acetate, mannitol, zinc chloride, sodium phosphate, potassium acetate, hydroxypropyl methylcellulose, gentamicin sulfate, boric acid, sodium hydroxide, lanolin oil, carbomer homopolymer type B (allyl pentaerythritol crosslinked), or benzalkonium chloride. In some embodiments, the composition does not contain parabens.

[0034] The composition may contain a gelling agent. Examples of gelling agents include gums, agars, carrageenan, petrolatum, or cellulose polymers (e.g., hydroxyethyl cellulose or hydroxymethyl cellulose).

[0035] The composition may further comprise one or more solvents, cosolvents, modifiers, emollients, preservatives, antimicrobial agents, antioxidants, humectants, stabilizers, or solubilizers.

[0036] In some embodiments, the composition comprises one or more excipients selected from water, sodium hydroxide, polysorbate 80, glycerin, castor oil, sodium acetate, boric acid, sorbic acid, disodium EDTA, carbomer copolymer type A or B (crosslinked with allyl pentaerythritol), or silicone oil or silicone polymer gel (e.g., dimethicone or cyclomethicone).

[0037] Mucosal adhesion polymers can be included, which deliver active ingredients locally to anatomically targeted sites and are particularly useful in gels. Such polymers possess properties known as bioadhesion, which refers to the ability of a drug carrier to adhere to specific biological tissues such as mucous membranes. These polymers can prolong the contact time between the active agent and the biological tissue, thereby improving bioavailability. Examples of bioadhesive polymers with various qualities of mucosal adhesion performance include carboxymethyl cellulose, carbomer, polycarbofil, and sodium alginate.

[0038] The pH of the spinosad composition should be within a dermatologically and / or ophthalmologically acceptable range. Normal tears have a pH of approximately 7.4 and possess some buffering capacity. Applying the composition to the eye stimulates tear flow and rapidly neutralizes any excess hydrogen or hydroxide ions within the tear's buffering capacity. If only 1 or 2 drops of the liquid composition containing the active agent are added to the eye, the buffering effect of the tears is usually sufficient to raise the pH and prevent noticeable discomfort.

[0039] While it is desirable for ophthalmic compositions to have the same pH and isotonicity as tears, this is generally not possible. A buffering system that is closest to the physiological pH of 7.4 and does not impair the acaricidal efficacy of the composition should be selected. Depending on its chemical properties, the pH of the mixture may be validated and adjusted with a solution of buffer or neutralizing agent, and optionally, the incorporation of additives, in one step of the preparation process. In some embodiments, the spinosad composition comprises one or more dermatologically and / or ophthalmologically acceptable pH adjusters or buffers, including but not limited to acids such as acetic acid, boric acid, citric acid, lactic acid, phosphoric acid, and hydrochloric acid; bases such as sodium hydroxide, sodium phosphate, sodium borate, sodium citrate, sodium acetate, sodium lactate, and tris(hydroxymethyl)aminomethane; and buffers such as citrate / glucose, sodium bicarbonate, and ammonium chloride. The amounts of these acids, bases, and buffers required to maintain the pH of the composition within a dermatologically and / or ophthalmologically acceptable range include those required for this purpose.

[0040] In some embodiments, the composition does not contain other agents with activity against Demodex mites. In some embodiments, the composition does not contain any active agent other than one or more spinosad compounds.

[0041] In some embodiments, where possible, the composition may include a pharmaceutically acceptable salt of a compound, such as spinosad compounds and other present compounds. In some embodiments, the composition comprises an acid addition salt of a compound. In some embodiments, the composition comprises a base addition salt of a compound. As used herein, the term pharmaceutically acceptable salt or complex refers to a salt or complex (e.g., solvate, polymorph) that retains the desired biological activity of the parent compound and exhibits minimal (if any) undesirable toxicological effects.

[0042] In some embodiments, the composition further comprises other agents with acaricidal activity against Demodex mites, such as one or more agents selected from the following: ivermectin, pyrethroids, pyrethroids (e.g., permethrin, cypermethrin, or D-benzylpyrethrin), tea tree oil (TTO), TTO components (e.g., 4-terpene alcohol (T4O)), metronidazole, hypochlorous acid (HOCl), essential oils (e.g., peppermint oil or sage), alkali metal salts (e.g., lithium salts), thiophosphates (e.g., non-volatile, fat-soluble thiophosphates (e.g., phosmet) or formamidin (e.g., amitraz).

[0043] In some embodiments, the composition further comprises one or more antibiotic agents. In some embodiments, the antibiotic agents have antibacterial activity against one or more bacteria (e.g., Streptococci, Staphylococci, or Bacillus oleronius) found on or within Demodex mites. In some embodiments, the antibiotic agents also have acaricidal activity, such as metronidazole.

[0044] In some embodiments, the composition further comprises one or more anti-inflammatory agents. Examples of anti-inflammatory agents that may be included are glucocorticoids or other steroids (e.g., prednisone, cortisone acetate, prednisolone, methylprednisolone, dexamethasone, betamethasone, triamcinolone, beclomethasone, flucortisone acetate, deoxycorticosterone acetate, aldosterone), nonsteroidal anti-inflammatory drugs (e.g., salicylates, aryl fatty acids, 2-arylpropionic acid, N-aryl-o-aminobenzoic acid, oxacin, coxib, or sulfanilide), and Cox-2 specific inhibitors (e.g., vardicoxib, ...). Celecoxib or roficoxib), leflunomide, glucosinolate, auroflavin, sulfasalazine, hydroxychloroquine, minocycline, TNF-α binding protein (e.g., infliximab, etanercept, or adalimumab), abatacept, anaerobicin, interferon-β, interferon-γ, interleukin-2, allergy vaccine, antihistamines, antileukotriene, β-agonists, theophylline or anticholinergics, antibiotics, tacrolimus or retinoids.

[0045] In some embodiments, the anti-inflammatory agent is formulated for topical application. In some embodiments, the topical anti-inflammatory agent is a topical steroid. In the United States, topical steroids are classified into groups I-VII based on their ability to constrict capillaries and cause skin whitening, with group I being the strongest or most effective and group VII being the weakest and least potent. Some examples of topical steroids that may be used include clobetasol, betamethasone, diflubenzuron, fluocinolone acetonide, fluocinolone acetonide, halometasol, amcinonide, desoxymethasone, halcinonide, fluticasone, triamcinolone, fluocinolone acetonide, hydrocortisone, mometasone, triamcinolone, aclomethasone, desonide, and prednisone.

[0046] In some embodiments, the anti-inflammatory agent is formulated for ophthalmic use. Some examples of ophthalmic formulations of steroids that may be used include dexamethasone, difluprednisolone, fluorometholone, cloprednisolone (loteprednol etabonate), and limexolone.

[0047] Optionally, the composition may contain an abrasive for application to areas of skin and hair adjacent to the eyes. Containing an abrasive can mechanically agitate tissue areas near the eyes, eyelids, eyelashes, and hair follicles to improve the ability of one or more spinosad compounds in the composition to approach Demodex mites. This treatment can be used to agitate or move unwanted materials, such as Demodex mite eggs, larvae, or mites, and remove them from eyelash follicles or other areas of the eyes or eyelids, or bring them into contact with the spinosad composition. The abrasive can also eliminate organisms by directly killing or destroying them. Abrasives may include any abrasive particles, powders, or crystals, including but not limited to one or more of the following: alumina (e.g., aluminum oxide, aluminum trioxide, corundum powder), barium sulfate, boron nitride, calcium carbonate, cellulose acetate, ceramics, diamond, diatomaceous earth, emerald, ethylene / acric acid copolymer, fiber, garnet, glass, kaolin, lauroyl lysine, lava, magnesium oxide, mica, modified starch, nylon, other metals, other polymers, other silica or silicon-containing materials, polyethylene, polymethyl methacrylate polypropylene, polystyrene, polytetrafluoroethylene (PTFE), pumice, ruby, sand, sapphire, shell, sericite, silica, silica, silicon carbide, sodium bicarbonate, sodium chloride crystals, starch, silk, talc, topaz, zeolite, or polymer particles.

[0048] Abrasive particles can be of any shape and can have any number of sides. Abrasive particles can be integrally rhomboid (triangular), elliptical, Marquise-shaped, octagonal, pear-shaped, rectangular, circular, square, or combinations or variations of these shapes (e.g., rounded square). Abrasive particles can have one surface or more surfaces (e.g., sides or front). Abrasive particles can have 2, 3, 4, 5, 6-10, 11-20, 21-30, up to 40, up to 50, up to 60, or more sides. The surface of the abrasive particle can be substantially smooth, regular, textured, or irregular. Abrasive particles can have one or more sharp edges or tips. The longest dimension of the abrasive particle can be from about 1 to about 600 micrometers.

[0049] The abrasive particles in a group can be similarly shaped to each other or can be shaped differently from each other. The abrasive particles in a group can all have the same size, or can be within a size range. The particles in a group can be larger than a minimum size or smaller than a maximum size. A group of abrasive particles can include particles spanning from about 1 to about 15 micrometers, from about 15 micrometers to about 25 micrometers, from about 25 micrometers to about 100 micrometers, from about 100 to about 300 micrometers, or from about 300 to about 600 micrometers. In one embodiment, a group of abrasive particles can include particles spanning from greater than about 25 micrometers to less than about 300 micrometers. Different sizes and shapes of particles can be selected for various reasons. Different sizes and shapes can be selected for different eye area diseases, different skin types or sensitivities, and / or different application methods. Particles with a rough surface can be applied using an applicator such as a stick or a small towel, while substantially round particles can be pushed onto a surface under pressure.

[0050] Abrasives may comprise a set of individual particles, or may comprise a substrate having an abrasive surface, or may be any combination or variation. The substrate may have multiple abrasive particles attached thereto to provide a grinding surface, or it may be a material with a rough textured surface. The rough textured surface may have pore sizes on its surface ranging from about 1 to about 15 micrometers, from about 15 micrometers to about 25 micrometers, from about 25 micrometers to about 100 micrometers, from about 100 to about 300 micrometers, or from about 300 to about 600 micrometers.

[0051] ointment

[0052] In some embodiments, dermatologically and / or ophthalmologically acceptable matrices include pharmaceutically acceptable ointment matrices. Examples of suitable ointment matrices include, but are not limited to, oily ointment matrices such as petrolatum (e.g., liquid petrolatum or white petrolatum), plastibase (a composite ointment matrices of liquid paraffin and polyethylene), hard paraffin, white soft paraffin, yellow soft paraffin, liquid paraffin, emulsified wax, microcrystalline wax, white beeswax, yellow beeswax, carnauba wax, lanolin (lanolin fat), mineral oil, olive oil, purified lanolin, anhydrous lanolin, and water-soluble ointment matrices such as polyethylene glycol (e.g., polyethylene glycol 400 or polyethylene glycol 3350), propylene glycol, polyethylene oxide, polypropylene oxide, or any combination thereof.

[0053] In some embodiments, the composition is an ointment and includes one or more excipients selected from: petrolatum, mineral oil, propylparaben, methylparaben, lanolin, chlorobutanol, water, lanolin alcohol, sodium thiosulfate, sodium dihydrogen phosphate, phenylmercuric acetate, mannitol, zinc chloride, sodium phosphate, potassium acetate, hydroxypropyl methylcellulose, gentamicin sulfate, boric acid, sodium hydroxide, lanolin oil, carbomer homopolymer type B (allyl pentaerythritol crosslinked) or benzalkonium chloride.

[0054] In some embodiments, the spinosad composition is an ointment having a petrolatum matrix. Petrolatum is a semi-solid mixture of hydrocarbons, offering certain advantages for dermatological and ophthalmic applications. Petrolatum is a pseudoplastic body, providing solid-state behavior that increases stability and prevents phase separation and sedimentation of suspended particles. Petrolatum is thixotropic, which is excellent for holding the drug in suspension while maintaining spreadability. For ophthalmic applications, it can be dispersed in the eye, and the product becomes more fluid and retains its flowability when shear forces are applied (during blinking), thus aiding in eye coverage. Because petrolatum is non-aqueous, it spreads and softens but is not washed out of the eye by tears (unlike aqueous solutions). As a non-aqueous and non-hygroscopic excipient, the problems of component hydrolysis and hydrolytic degradation are avoided. As an insoluble excipient, petrolatum limits treatment to local administration with minimal systemic absorption. Petrolatums used in pharmaceutical products are classified as petrolatum USP and white petrolatum USP. White petrolatum is generally preferred for ophthalmic applications. Petrolatums vary in color and transparency, as well as rheological properties (consistency, flowability, and yield stress). Compared to their concentration in the formulation, other excipients (such as mineral oil, surfactants, and preservatives) can reduce apparent viscosity and yield stress.

[0055] lotion

[0056] The compositions of the present invention can be formulated into emulsions, such as oil-in-water emulsions. The topical oil-in-water emulsion compositions can be in liquid, paste, or solid form, or in the form of ointments, creams, gels, sprays, foams, suspensions, lotions, shampoos, or detergent bases. The compositions can also be suspensions of microspheres or nanospheres, lipid or polymer vesicles, polymer patches, and hydrogels for controlled release. These compositions for topical application can be in anhydrous, aqueous, or emulsion form.

[0057] In some embodiments of the invention, the composition is in the form of an emulsion, gel, or solution of the type of cream or lotion, and more particularly in the form of a cream.

[0058] In some embodiments, the emulsion contains one or more excipients selected from water, sodium hydroxide, polysorbate 80, glycerin, castor oil, carbomer copolymer type A, sodium acetate, boric acid, sorbic acid, disodium EDTA, or silicone oil or silicone polymer gel (e.g., dimethicone or cyclomethicone).

[0059] Conventional emulsions are typically relatively unstable, nearly homogeneous systems of two immiscible liquids, one dispersed in the other as fine droplets (micelles). This dispersion is stabilized by a surfactant-emulsifier, which alters the structure and force ratios at the interface, thus increasing the dispersion's stability by reducing interfacial tension energy. Surfactant-emulsifiers are amphiphilic compounds containing a hydrophobic component with an affinity for oil and a hydrophilic component with an affinity for water, thereby forming a bond between the two phases. Therefore, ionic or nonionic emulsifiers stabilize oil / water emulsions by adsorbing onto the interface and forming a liquid crystal film.

[0060] The compositions of the present invention may comprise one or more nonionic surfactant emulsifiers. The emulsifying ability of a nonionic surfactant is closely related to the polarity of its molecules. This polarity is defined by HLB (hydrophilic / lipophilic balance). Conventional emulsions are typically stabilized by a mixture of surfactants, which may have completely different HLBs, but their proportions in the mixture correspond to the HLB required for the fatty phase to be emulsified.

[0061] Suitable nonionic surfactant emulsifiers can be selected from the group consisting of: cetearyl alcohol, cetyl alcohol, cocamide DEA, cocamide MEA, isocetyl alcohol polyether-20, oleyl alcohol, sorbitol monostearate, sorbitol tristearate, stearyl alcohol, tetrabutylphenol aldehyde, styrosinase, Solutol HS 15, poloxamer, and Proponic F-68LF. TM Or Lutrol F68, Prnik L-62LF TM and Prnik L62DTM (BASF Wyandotte Corp., Parsippony, New Jersey, USA), polysorbates such as polysorbate 20 and polysorbate 80, polyoxyethylene fatty acid esters such as Emulphor TM (GAF Corp., Wayne, New Jersey, USA).

[0062] In some embodiments, the emulsion includes a carbopol homopolymer, which can emulsify oil-in-water suspensions, thereby allowing for use in non-turbid / non-sticky formulations.

[0063] In some embodiments, the compositions of the present invention are oil-in-water emulsions comprising: (a) one or more spinosad compounds; (b) an oil phase comprising fatty substances and / or emollients; (c) water; and optionally (d) a surfactant emulsifier.

[0064] Examples of fatty substances include vegetable oils, mineral oils, animal or synthetic oils, silicone oils, guerbert alcohols or other substances, and mixtures thereof.

[0065] Examples of mineral oils include paraffin oils of various viscosities, such as Primol 352, Marcol 82, or Marcol 152 sold by Esso. Examples of vegetable oils include sweet almond oil, palm oil, soybean oil, sesame oil, and sunflower oil. Examples of animal oils include lanolin oil, squalene oil, fish oil, and mink oil. Examples of synthetic oils include esters, such as cetearyl isononanoate sold by Cognis France under the trade name Cetiol SN; diisopropyl adipate, such as the product sold by ISF under the trade name Ceraphyl 230; isopropyl palmitate, such as the product sold by Croda under the trade name Crodamol IPP; or caprylic / capric triglycerides, such as Miglyol 812 sold by Huls / Lambert Riviere. Examples of silicone oils include polydimethylsiloxane, such as products sold under the trade name Dow Corning 200 fluid, and cyclomethicone, such as products sold by Dow Corning under the trade name Dow Corning 244 fluid, and products sold by SACI-CFPA under the trade name Mirasil CM5.

[0066] Other fatty substances may include fatty acids, such as white petrolatum, stearic acid, fatty alcohols such as stearyl alcohol, cetearyl alcohol and cetyl alcohol or their derivatives, waxes such as beeswax, carnauba wax or candelilla wax, and gums, especially silicone rubber.

[0067] The amount of one or more fatty substances in the composition may be from about 20% to about 40% by weight of the composition. In some embodiments, the amount of one or more fatty substances is from about 20% to about 35% by weight of the composition.

[0068] The oil phase of the composition may include one or more emollients, such as vegetable oil, mineral oil, animal oil or synthetic oil, silicone oil, isopropyl palmitate, 1-decene polymer (hydrogenated), C 12 -C 15 Alkyl benzoate, C 12 -C 15 Alkyl benzoate esters, lanolin alcohol, and isopropyl myristate; and mixtures thereof. The emollient in the composition may be present in an amount from about 10% to about 20% by weight of the composition.

[0069] In some embodiments, the oil phase of the composition comprises a mixture of fatty substances and emollients, and in some embodiments, at least two emollients. In some embodiments, the fatty substances are white petrolatum and mineral oil, while the emollients are lanolin alcohol and isopropyl myristate.

[0070] The oil phase of the emulsion may be present in an amount of 15% to 45% by weight relative to the total weight of the composition, preferably 20% to 40% by weight.

[0071] The composition may contain up to 15% by weight of a suitable surfactant emulsifier. In some embodiments, the surfactant emulsifier is present in the composition in an amount of about 5% to about 15% by weight of the composition.

[0072] The composition may contain 0.1% to 10% by weight of one or more spinosads relative to the total weight of the composition. In some embodiments, the composition contains 0.5% to 10% by weight of one or more spinosads relative to the total weight of the composition.

[0073] The compositions of the present invention may also contain 50% to 95% water or a buffer. In some embodiments, the composition contains 55% to 80% water or a buffer by weight relative to the total weight of the composition. The water used in the compositions according to the present invention is preferably purified water or distilled water.

[0074] The composition may also contain inert additives or combinations of additives, such as flavor enhancers, preservatives, stabilizers, humidity regulators, pH adjusters, osmotic pressure regulators, UV-A and UV-B shielding agents, and antioxidants. In some embodiments, the spinosad oil-in-water emulsion composition does not contain preservatives.

[0075] These additives may be present in the composition at a concentration of 0.001% to 20% by weight relative to the total weight of the composition.

[0076] In some embodiments, the composition is an oil-in-water emulsion comprising: (a) one or more spinosads in an amount of about 0.01% to about 10% by weight; and (b) an oil phase comprising about 20% to about 40% by weight of one or more fatty substances and about 10% to about 20% by weight of one or more emollients; (c) about 5% to about 15% by weight of one or more surfactant emulsifiers; and (d) water.

[0077] Exemplary formulations of the composition

[0078] In some embodiments, the composition is a cream comprising, in addition to one or more spinosad compounds (e.g., spinosad), one or more of the following ingredients: water or a buffer, mineral oil, acetylated lanolin alcohol, stearyl alcohol, cetearyl alcohol, and propylene glycol. In some embodiments, the cream contains each of the aforementioned ingredients.

[0079] In some embodiments, the composition is a cream comprising, in addition to one or more spinosad compounds (e.g., spinosad), one or more of the following ingredients: water or a buffer, propylene glycol, mineral oil, acetylated lanolin alcohol, cetyl alcohol, stearyl alcohol, cetearyl alcohol, and polyoxyethylene 35 castor oil. In some embodiments, the cream contains each of the aforementioned ingredients.

[0080] In some embodiments, the composition is a detergent comprising, in addition to one or more spinosad compounds (e.g., spinosad), one or more of the following ingredients: water or a buffer, isopropyl myristate, castor oil, polyoxyethylene 40 stearate, cetearyl alcohol, and cetyl wax. In some embodiments, the detergent comprises each of the foregoing ingredients.

[0081] In some embodiments, the composition is an ointment comprising, in addition to one or more spinosad compounds (e.g., spinosad), one or more of the following ingredients: water or a buffer, propylene glycol, castor oil, acetylated lanolin alcohol, cetyl alcohol, stearyl alcohol, cetearyl alcohol, cetyl wax, povidone K 90, and stearic acid. In some embodiments, the ointment comprises each of the aforementioned ingredients.

[0082] In some embodiments, the composition is an ointment comprising, in addition to one or more spinosad compounds (e.g., spinosad), one or more of the following ingredients: propylene glycol, mineral oil, acetylated lanolin alcohol, stearyl alcohol, glyceryl stearate / PEG-100 stearate, polyhydroxy40 stearate, and stearic acid. In some embodiments, the ointment comprises each of the aforementioned ingredients.

[0083] In some embodiments, the composition is described in one of Tables 1-5. In some embodiments, for a given composition, the sum of the percentage concentrations of each component listed in the table is 100%.

[0084] In some embodiments, the composition is described in one of Tables 1-5, but one or more of the listed excipients are used instead of one or more different excipients having the same listed function.

[0085] In some embodiments, in each of Tables 1-5, 0.1% spinosad is used as a reference point. For every decrease in spinosad concentration by w:w%, the dominant component in the formulation (i.e., the component with the largest and smallest percentages) also increases by the same w:w%. Conversely, for every increase in spinosad concentration by w:w%, the dominant excipient in the formulation decreases by the same w:w%. For example, in Table 1, for every 0.1% increase in spinosad concentration, distilled water or buffer is reduced by 0.1%. As a further embodiment, in Table 5, for every 1.0% increase in spinosad concentration from 0.1%, mineral oil is reduced by 1%.

[0086] Table 1. Cream

[0087]

[0088] Table 2 Creams

[0089]

[0090]

[0091] Table 3 Detergents

[0092]

[0093] Table 4 Ointments

[0094]

[0095] Table 5 Ointments

[0096]

[0097]

[0098] Treatment

[0099] One aspect of the invention is a method for treating ocular Demodex mite infection in the eyes of a human or animal subject or for treating eye or skin diseases associated with ocular Demodex mite infection, comprising topically applying a composition comprising one or more spinosad compounds to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye or areas adjacent to the eye.

[0100] Not limited by theories about the underlying mechanisms of action, for existing Demodex mite infections, as a treatment approach, the goal is generally to reduce the number of Demodex mites in anatomical sites such as the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye or areas adjacent to the eye, so as to alleviate or eliminate any or one of the signs of Demodex-related disease. Because Demodex mites are part of the normal skin microbiome, and susceptibility to Demodex mites and levels of mite-associated immune response factors can vary among subjects, the goal of treatment may be to reduce the number of surviving Demodex mites to a normal or healthy level for the individual (below the threshold for parasite overdose), or to eradicate all Demodex mites from the target anatomical site or locus (LOC).

[0101] The composition can also be applied topically preventively to subjects without Demodex mite infection to prevent or delay the onset of Demodex mite infection.

[0102] By administering the compound once or multiple times, an amount of spinosad compound sufficient to reduce the number of Demodex mites in the corresponding anatomical site (acaricidal effective amount) is delivered compared to the amount or concentration of spinosad or other acaricides used without the use of spinosad or other acaricides.

[0103] The composition may be applied topically to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye, or to areas adjacent to the eye, either by a subject (i.e., self-application) or by another person.

[0104] In some embodiments, for formulations intended for application to the ocular surface, depending on their viscosity, the composition may be applied drop by drop (e.g., using an eye dropper) or placed on the corner of the eye or eyelid or inside the upper or lower eyelid. For example, one or more drops may be applied (e.g., about 30 microliters per drop). The subject may blink to facilitate the distribution of the composition on the ocular surface. In some embodiments, the composition is applied topically to the ocular surface.

[0105] In some embodiments, the composition is applied topically to the ocular surface or other anatomical structures of the eye, or to areas adjacent to the eye, just before falling asleep.

[0106] In some embodiments, the composition is applied topically to an area adjacent to the eye, wherein the area includes the sebaceous gland openings of the eyelid (e.g., one or more of Zeiss glands, Mohr's glands, or meibomian glands).

[0107] In some embodiments, the composition is applied topically to one or more epidermal invaginations called pilosebaceous units (e.g., hair, hair follicle, sebaceous gland).

[0108] The composition can be applied topically using an eye or face applicator as described herein or by using a part of the body, such as a finger or knuckle (by wiping, rubbing, massaging, etc.).

[0109] Examples of applicators include cotton swabs, cosmetic pads, wiping cloths, wiping sticks, small towels, sponges, gauze, powder puffs, sticks, brushes, or combs. In some embodiments, the applicator includes at least a portion that contacts the target anatomical area and may be made of woven or non-woven materials such as cotton, polyester, or synthetic fibers. The applicator may be disposable or reusable.

[0110] The spinosad composition can be gently applied topically to one or more anatomical sites of a subject, including the ocular surface or other anatomical structures of the eye, or areas adjacent to the eye, such as the eyelids, eyelid margins, eyelashes, eyelash follicles, eyebrows or eyebrow follicles, or sebaceous gland openings of the eyelids (e.g., one or more of Zei's glands, Moll's glands, or meibomian glands). The spinosad composition can be massaged into the skin. In some embodiments, the spinosad composition is left on the area until the next treatment. In other embodiments, excess spinosad composition is wiped off or washed away after application.

[0111] Optionally, one or more anti-inflammatory agents may be administered to the subject before, during, or after the application of the spinosad composition. This may help suppress or reduce inflammation caused by some embodiments of the spinosad composition or by any mechanical stimulation that may be part of treatment. The anti-inflammatory agent may also suppress or reduce potential inflammation associated with decaying mites. The anti-inflammatory agent may be administered to the subject via a route suitable for the formulation. For example, the anti-inflammatory agent may be administered topically, including in the spinosad composition or as a separate composition. In other embodiments, the anti-inflammatory agent may be administered to the subject systemically (e.g., orally).

[0112] Examples of applicable anti-inflammatory agents include glucocorticoids or other steroids (e.g., prednisone, cortisone acetate, prednisolone, methylprednisolone, dexamethasone, betamethasone, triamcinolone, beclomethasone, flucortisone acetate, deoxycorticosterone acetate, aldosterone), nonsteroidal anti-inflammatory drugs (e.g., salicylates, aryl fatty acids, 2-arylpropionic acid, N-aryl-o-aminobenzoic acid, oxacin, coxib, or sulfanilide), and Cox-2 specific inhibitors (e.g., vardicoxib). Celecoxib or roficoxib), leflunomide, glucosinolate, gold thiomalate, aurofin, sulfasalazine, hydroxychloroquine, minocycline, TNF-α binding protein (e.g., infliximab, etanercept, or adalimumab), abatacept, anaerobicin, interferon-β, interferon-γ, interleukin-2, allergy vaccine, antihistamines, antileukotriene, β-agonists, theophylline or anticholinergics, antibiotics, tacrolimus or retinoids.

[0113] Optionally, as a pretreatment or preparatory step prior to treatment with the spinosad composition, the anatomical site may be scrubbed with an agent such as a cleaning substance. The abrasive may be applied to areas adjacent to the eyes to remove skin, skin layers, skin debris, microorganisms, eyelashes, oil, and / or other related or unrelated substances. The abrasive may mechanically agitate tissue areas near the eyes, eyelids, eyelashes, and hair follicles to enhance the ability of the spinosad compounds in the composition to access Demodex mites. This treatment may be used to agitate or move unwanted materials, such as Demodex mite eggs, larvae, or mites, and remove them from eyelash follicles or other areas of the eye or eyelid, or bring them into contact with the spinosad composition. The abrasive may also eliminate organisms by directly killing or destroying them. Abrasives may include any abrasive particles, powders, or crystals, including but not limited to one or more of the following: alumina (e.g., aluminum oxide, aluminum trioxide, corundum powder), barium sulfate, boron nitride, calcium carbonate, cellulose acetate, ceramics, diamond, diatomaceous earth, emerald, ethylene / acrylic acid copolymer, fiber, garnet, glass, kaolin, lauroyl lysine, lava, magnesium oxide, mica, modified starch, nylon, other metals, other polymers, other silica or silicon-containing materials, polyethylene, polymethyl methacrylate polypropylene, polystyrene, polytetrafluoroethylene (PTFE), pumice, ruby, sand, sapphire, shell, sericite, silica, silica, silicon carbide, sodium bicarbonate, sodium chloride crystals, starch, silk, talc, topaz, zeolite, or polymer particles.

[0114] Optionally, other agents active against Demodex mites may be applied before, during, or after the application of the spinosad composition. In some embodiments, the reagent has activity against Demodex mites, such as ivermectin, pyrethroids, pyrethroids (e.g., permethrin, benzalkonium chloride, or D-phenylephrine), tea tree oil (TTO), TTO components (e.g., 4-terpene alcohol (T4O)), metronidazole, hypochlorous acid (HOCl), essential oils (e.g., peppermint oil or sage), alkali metal salts (e.g., lithium salts), thiophosphates (e.g., non-volatile, fat-soluble thiophosphates (e.g., phosmet, also identified as O,O-diethyl-O-(3-chloro-4-methylcoumarin-7)thiophosphate) or formamidin (e.g., amitraz, also identified as N,N'-[(methylimino)dimethyldiyne)]di-2,4-dimethylamine). The other reagents may be applied as a component of the spinosad composition, or applied topically as a separate composition or via another route.

[0115] Dosage regimens for treating, preventing, or improving a disease for which remission is sought can be modified based on a variety of factors. These factors include the disease suffered by the subject, as well as the subject's age, weight, sex, diet, and medical condition. Therefore, the actual dosage regimen used can vary considerably and may deviate from the dosage regimen described in this article.

[0116] Detection of Demodex mites

[0117] The method of the present invention may further include confirming the presence of Demodex mites on the subject before administering the spinosad composition. Several clinical signs can alert one to the presence of Demodex mites in the eyelashes. Identifying these clues allows one to perform one of two simple examination techniques that will confirm the presence of mites.

[0118] 1. Cylindrical scales (CD). This is the most obvious clinical sign and an excellent indicator of mite infestation. Cylindrical scales can be seen under a slit lamp, which is different from the scaly, dandruff-like fragments seen in traditional anterior blepharitis.

[0119] 2. Changes in the skin around the eyelash follicles. The skin near the follicle opening becomes swollen and raised, which may give the skin around the follicle a greasy, oily appearance.

[0120] 3. Changes in the appearance of eyelashes. With prolonged infection, eyelashes may become thinner and more brittle, or they may begin to lose their color. Incorrect eyelash direction and reduction may also occur.

[0121] 4. Eyelid congestion / telangiectasia. Because the presence of mites is associated with increased levels of interleukins and different types of pro-inflammatory cytokines, increased eyelid inflammation levels may be significant, potentially leading to increased angiogenesis along the eyelid margin.

[0122] 5. Patient history / association with other diseases. The prevalence of Demodex mites increases with age, and strong associations have been found between patients with rosacea, seborrheic dermatitis, and other forms of inflammatory skin diseases (including allergic symptoms such as itching).

[0123] These clinical symptoms can indicate the presence of an overpopulation of Demodex mites. However, apart from the cylindrical dandruff, none of these items are diagnostic in themselves. The presence of mites and the classification of the degree of mite infestation can be determined by one of two simple steps: eyelash removal (e.g., two to four distinct eyelashes on the eyelid and associated fragments / cylindrical dandruff, observed as a complex under an optical microscope) and eyelash rotation (transferring the fragments from the hair follicle to a slide for examination under an optical microscope or observation at high magnification (25-40x) under a slit lamp).

[0124] Therefore, the presence of Demodex mites can be confirmed by one or more techniques, such as direct mite identification and mite density measurement, for example, the presence of cylindrical dandruff at the base of the eyelashes, changes in the skin around the eyelash follicles, changes in the eyelashes, and eyelash sampling (sampling hair loss with an optical microscope or rotating eyelashes with a slit lamp). In addition to ex vivo hair removal and microscopy, in vivo confocal microscopy can also be used (Randon M et al., “In vivo confocal microscopy as a novel and reliable tool for the diagnosis of Demodex eyelid infestation,” Br JOphthalmol, 2015, 99(3):336-341, which is incorporated herein by reference in its entirety).

[0125] Molecular diagnostic techniques can be used to detect Demodex mite-related molecules at the genetic and / or protein levels in biological samples obtained from subjects in areas where mites may be found (e.g., eyelashes, skin biopsy tissue). For example, polymerase chain reaction (PCR) or immunoassays (e.g., enzyme-linked immunosorbent assay (ELISA)) can be used to detect and quantify target Demodex folliculorum nucleic acid or protein molecules, such as extraskeletal chitin, chitin synthase, and 62-kDa and 83-kDa antigens of Bacillus thuringiensis, from skin and / or hair samples from subjects (e.g., see Zhao Ya-e et al., “Cloning and sequence analysis of chitin synthase gene fragments of Demodex mites,” J Zheijang Univ Sci B, 2012, 13(10): 763-768; Zhao Ya-e et al., “Discrimination between Demodex folliculorum (Acari:Demodicidae) isolates from China and Spain based on mitochondrial cox1 sequences,” J Zheijang Univ Sci B, 2013, 14(9): 829-836; Hu L et al., “Molecular identification of four phenotypes of human Demodex folliculorum”). inChina," Exp Parisitol, 2014,142:38-42; Li J et al., "Correlation between ocularDemodex infestation and serum immunoreactivity to Bacillus proteins inpatients with facial rosacea," Ophthalmology, 2010,117(5):870-877; Szkaradkiewicz A et al., "Bacillus oleronius and Demodex mite infestation inpatients with chronic blepharitis,” Clinical Microbiology and Infection, 2012,18(10):1020-1025;Kasetsuwan N et al., “Prevalence of ocular demodicosis among patients at Tertiary Care Center, Bangkok, Thailand,” Int J Ophthalmol, 2017, 10(1): 122-127; Liu D et al., “Demodex (Hair Follicle Mite),” Molecular Detection of Human Parasitic Pathogens, Chapter 72, CRC Press, 2012, pp. 741-750, the entire contents of which are incorporated herein by reference.

[0126] Therefore, in some embodiments of the method of the present invention, confirming the presence of Demodex mites includes identifying Demodex mites in in vivo or ex vivo biological samples (e.g., hair or skin samples), or detecting Demodex-associated molecules (e.g., chitin, chitin synthase, lipase, Demodex-associated bacterial antigens (e.g., Bacillus antigens, such as Bacillus vegetableis antigen), Demodex 16S rDNA, Demodex 18S rDNA). For example, Demodex mites have been shown to produce immunoreactive lipases, which can be used as targets for mite detection (Jimenez-Acosta F. et al., “Demodex mites contain immunoreactive lipase,” Arch Dermatol., 1989, 125: 1436–7). Target DNA and RNA can be extracted and amplified and qualitatively used as molecular barcodes for identification and interspecies and intraspecies differentiation, and quantitatively used to assess the extent of infection. Analysis based on gene or protein microarrays can also be used. Biological samples from subjects, such as skin surface, hair, tears, and peripheral blood, can be used to analyze the presence and levels of inflammatory markers consistent with Demodex mite infection.

[0127] To monitor the progress of treatment or confirm its efficacy, samples from the subjects can be studied after a period of treatment to determine whether the subjects have Demodex mites or threshold levels of Demodex mites, in order to determine whether the Demodex mites have been reduced to normal levels or eradicated through treatment.

[0128] This detection procedure can be used for any purpose requiring the detection of Demodex mites (e.g., Demodex folliculorum and / or Demodex brevis), including diagnostic and prognostic applications, such as in laboratory and clinical settings. Suitable samples include any biological sample in which Demodex mites can be found, including clinical samples obtained from medical subjects (humans) or veterinary subjects (non-human animals).

[0129] When the target molecules associated with Demodex mites in a sample are molecules targeted by antibodies, such as proteins or carbohydrates (e.g., chitin, chitin synthase, lipase), or Demodex-associated bacterial antigens (e.g., Bacillus antigens, such as Bacillus vegetableis antigen), immunochemical techniques utilizing antibodies (polyclonal or monoclonal) or antigen-binding fragments of such antibodies are used. These immunochemical techniques may involve radioimmunoassay or other recognized assays, such as enzyme-linked immunosorbent assay (ELISA). Target molecules can also be determined using standard non-immunochemical techniques, such as gas chromatography.

[0130] Detecting target nucleic acids (e.g., Demodex folliculorum and / or Demodex brevis, or Demodex-associated bacterial nucleic acids, or subject immune molecules) in a sample using gene-based molecular diagnostic techniques typically involves mixing the sample with at least one probe capable of hybridizing with the target nucleic acid, such as Demodex folliculorum and / or Demodex brevis nucleic acid, under very stringent conditions, and being able to detect the hybridization between the target nucleic acid and the probe. Detection of hybridization between the probe and the target nucleic acid indicates the presence of the target nucleic acid in the sample (and therefore the presence of an organism, such as Demodex folliculorum and / or Demodex brevis).

[0131] In some implementations, the nucleic acids of Demodex folliculorum and / or Demodex brevis present in the sample are amplified before detection using a hybridization probe. For example, it may be advantageous to amplify a subset of Demodex folliculorum and / or Demodex brevis nucleic acids and then detect the presence of the amplified Demodex folliculorum and / or Demodex brevis nucleic acids.

[0132] Detection of amplified products typically involves using labeled probes that are sufficiently complementary and hybridize with the amplified target nucleic acid sequence. Therefore, the presence, quantity, and / or characteristics of the amplified product can be detected by hybridizing a labeled probe complementary to the amplified product, such as a fluorescently labeled probe. In one embodiment, detection of a target nucleic acid sequence of interest, such as Demodex folliculorum and / or Demodex brevis nucleic acid, includes the combined use of PCR amplification and a labeled probe, thereby measuring the product using real-time PCR. In another embodiment, detection of an amplified target nucleic acid sequence of interest includes transferring the amplified target nucleic acid to a solid support, such as a blot, for example a Northern blotting, and detecting the blot with a probe, such as a labeled probe, which is complementary to the amplified target nucleic acid sequence. In yet another embodiment, detection of an amplified target nucleic acid sequence of interest includes hybridization of the labeled amplified target nucleic acid with a probe disclosed herein, the probe being arranged in a predetermined array at addressable locations and complementary to the amplified target nucleic acid.

[0133] Any nucleic acid amplification method can be used to detect the presence of Demodex mites in a sample. In one specific non-limiting embodiment, polymerase chain reaction (PCR) is used to amplify the target nucleic acid sequence. In other specific non-limiting embodiments, real-time PCR, reverse transcriptase-polymerase chain reaction (RT-PCR), real-time reverse transcriptase-polymerase chain reaction (rt RT-PCR), ligase chain reaction, or transcription-mediated amplification (TMA) are used to amplify the target nucleic acid. In one specific embodiment, real-time PCR, for example, real-time... PCR was used to amplify the nucleic acids of Demodex folliculorum and / or Demodex brevis. The techniques for nucleic acid amplification are well-known to those skilled in the art.

[0134] Typically, at least two primers are used in the amplification reaction. Amplification of Demodex mite nucleic acid involves contacting the Demodex mite nucleic acid with one or more primers that are capable of hybridizing with the Demodex mite nucleic acid and directing its amplification (e.g., primers that are capable of hybridizing with Demodex folliculorum and / or Demodex brevis under very stringent conditions).

[0135] The amplified Demodex mite nucleic acid can be detected in real time, for example, by real-time PCR, to determine the presence and / or quantity of Demodex mite-specific nucleic acids (e.g., Demodex folliculorum and / or Demodex brevis) in a biological sample. In this way, the amplified nucleic acid sequence, such as the amplified Demodex folliculorum and / or Demodex brevis nucleic acid sequence, can be detected using probes specific to the products amplified from the sequence of the Demodex mite of interest.

[0136] A fluorophore is a chemical compound that, when excited by exposure to a specific stimulus (e.g., light of a defined wavelength), emits light (fluorescence) of a different wavelength (e.g., longer wavelengths of light). Fluorophores are part of a large class of luminescent compounds. Luminescent compounds include chemiluminescent molecules, which do not require specific wavelengths of light to emit light but instead use chemical energy. Therefore, the use of chemiluminescent molecules (e.g., luminescent proteins in jellyfish) eliminates the need for an external electromagnetic radiation source (e.g., a laser).

[0137] The probe contains isolated nucleic acid capable of hybridizing with target nucleic acids, such as Demodex mite nucleic acid, for example, Demodex folliculorum and / or Demodex brevis ribosomal nucleic acid molecules. A detectable label or reporter molecule can be attached to the probe. Typical labels include radioactive isotopes, enzyme substrates, cofactors, ligands, chemiluminescent or fluorescent agents, haptens, and enzymes.

[0138] For example, Sambrook et al., *Molecular Cloning: A Laboratory Manual*, Cold Spring Harbor Laboratory Press (1989), and Ausubel et al., *Current Protocols in Molecular Biology*, Greene Publishing Associates and Wiley-Intersciences (1987), discuss guidelines for selecting appropriate markers for various purposes and methods for using markers.

[0139] Those skilled in the art will know suitable methods for extracting nucleic acids, such as RNA and / or DNA, from samples. Such methods will depend on the type of sample, for example, where Demodex mite nucleic acids are found. For example, nucleic acids can be extracted using guanidine isothiocyanate, for instance, by a one-step separation using acidic guanidine isothiocyanate-phenol-chloroform extraction as described by Chomczynski et al. (Anal. Biochem., 1997, 162: 156-59). Samples can be used directly or processed, for example, by adding solvents, preservatives, buffers, or other compounds or substances. Nucleic acids can be extracted using standard methods. For example, commercially available kits (e.g.) can be used. DNA Mini Kit Roche MagNA Purification and Compressed Nucleic Acid Isolation Kit I or Mini reagent kit NASBA diagnostic procedure (bioMérieux); or MASTERPURE TM The Complete DNA and RNA Purification Kit (EPICENTRE) enables rapid nucleic acid preparation.

[0140] Arrays can be used for the rapid detection of Demodex mite nucleic acids in samples. An array is an arrangement of addressable sites on a substrate, each address containing a nucleic acid, such as a probe (e.g., a Demodex mite probe). In some implementations, each address corresponds to a single type or category of nucleic acid, such as a single probe, although a particular nucleic acid may be redundantly contained in multiple addresses. “Microarrays” are miniaturized arrays that require microscopic examination to detect hybridization. Larger “macroarrays” allow each address to be identified with the naked eye, and in some implementations, hybridization signals can be detected without additional magnification. These addresses may be labeled, typed with individual guides, or otherwise identified by location.

[0141] The use of the term "array" includes arrays found in DNA microchip technology. As a non-limiting embodiment, probes may be contained on a DNA microchip similar to those commercially available from Affymetrix, Inc. (Santa Clara, CA). Products and related products. In short, a DNA microchip is a miniature, high-density array of probes on a glass wafer substrate. Specific probes are selected, and photolithographic masks are designed for use in processes based on solid-phase chemical synthesis and photolithography techniques similar to those used in the semiconductor industry. The mask is used to isolate exposed sites on the chip, where probes are chemically synthesized, each probe located at a predetermined position in the array. Once fabricated, the array is ready for hybridization. Probes or nucleic acids in the sample can be labeled, for example, with fluorescent labels, and after hybridization, the hybridization signal can be detected and analyzed.

[0142] Combination therapy

[0143] Other agents may be administered to the subject simultaneously or continuously with the spinosad composition. Other agents may be administered before, during, or after the topical application of the spinosad composition. Additional agents may be administered in the same composition as one or more spinosad compounds, or as a separate composition to the subject. If administered as a separate composition, the other agents may be administered topically or via any local or systemic route suitable for having the desired effect on the subject. In some embodiments, the other agents are administered topically. In other embodiments, the other agents are administered orally.

[0144] In some embodiments, the other reagents are acaricidal agents against Demodex mites, such as one or more reagents selected from the following: ivermectin, pyrethroids, pyrethroids (e.g., permethrin, cypermethrin, or D-benzylpyrethrin), tea tree oil (TTO), TTO components (e.g., 4-terpene alcohol (T4O)), metronidazole, hypochlorous acid (HOCl), essential oils (e.g., peppermint oil or sage), alkali metal salts (e.g., lithium salts), thiophosphates (e.g., non-volatile, fat-soluble thiophosphates (e.g., phosmet) or formamidin (e.g., amitraz, also identified as N,N'-[(methylimino)dimethyldiyne)]di-2,4-dimethylamine).

[0145] Benzyl chlorpyrifos is a member of the pyrethroid family, which is a class of synthetically derived insecticides. Pyrethroids are structurally related to naturally occurring pyrethroids, pyrethroid I, and pyrethroid II. Synthetic pyrethroids include benzyl chlorpyrifos (US Patent No. 4,113,968; US Patent Publication No. 20160361297), benzylfuran, and ethoxysulfuron (US Patent Nos. 3,934,023 and 2,348,930), each of which is incorporated herein by reference in its entirety. Other examples of pyrethroids include bifenthrin, cyhalothrin, deltamethrin, cypermethrin, phenacetin, D-phenethrin, deltamethrin, cypermethrin, fenprropathrin, fenpropathrin, γ-trifluorocypermethrin, cyproterin, laminarin, momfluorothrin, permethrin, pyrethrin, pyrethrin, pyrethrin, pyrethrin, cypermethrin, and pyrethrin.

[0146] In some embodiments, the other agent is an antibiotic. In some embodiments, the antibiotic has antibacterial activity against one or more bacteria found on or within Demodex mites, such as Streptococcus, Staphylococcus, or Bacillus subtilis. In some embodiments, the antibiotic also has acaricidal activity, such as metronidazole.

[0147] In some embodiments, other agents are one or more anti-inflammatory agents. Examples of anti-inflammatory agents that may be included are glucocorticoids or other steroids (e.g., prednisone, cortisone acetate, prednisolone, methylprednisolone, dexamethasone, betamethasone, triamcinolone, beclomethasone, flucortisone acetate, deoxycorticosterone acetate, aldosterone), nonsteroidal anti-inflammatory drugs (e.g., salicylates, aryl fatty acids, 2-arylpropionic acid, N-aryl-o-aminobenzoic acid, oxacin, coxib, or sulfanilide), and Cox-2 specific inhibitors (e.g., vardicoxib, ...). Celecoxib or roficoxib), leflunomide, glucosinolate, auroflavin, sulfasalazine, hydroxychloroquine, minocycline, TNF-α binding protein (e.g., infliximab, etanercept, or adalimumab), abatacept, anaerobicin, interferon-β, interferon-γ, interleukin-2, allergy vaccine, antihistamines, antileukotriene, β-agonists, theophylline or anticholinergics, antibiotics, tacrolimus or retinoids.

[0148] In some embodiments, the anti-inflammatory agent is formulated for topical application. In some embodiments, the topical anti-inflammatory agent is a topical steroid. In the United States, topical steroids are classified into groups I-VII based on their ability to constrict capillaries and cause skin whitening, with group I being the strongest or most effective and group VII being the weakest and least potent. Some examples of topical steroid formulations that may be used include betamethasone, diflubenzuron, fluocinolone acetonide, fludroxil, halometasone, amcinonide, desoxymethasone, halcinonide, fluticasone, triamcinolone, fluocinolone acetonide, hydrocortisone, mometasone, triamcinolone, aclomethasone, desonide, and prednisone.

[0149] In some embodiments, the anti-inflammatory agent is formulated for ophthalmic use. Some examples of ophthalmic formulations of steroids that may be used include dexamethasone, difluprednisolone, fluorometholone, cloprednisolone (loteprednol etabonate), and limexolone.

[0150] In some embodiments, the other reagents are one or more alkali metal salts that can be used as acaricides. Lithium salts are preferred alkali metal salts. Lithium chloride is the most preferred alkali metal salt.

[0151] It should be understood that whenever alkali metal salts or lithium salts are mentioned, these alkali metal salts or lithium salts include any organic and inorganic salts of alkali metals or lithium that are suitable for and capable of exerting acaricidal effects (preferably against Demodex mites). Organic salts of alkali metals such as lithium include salts such as citrates, carbonates, lactates, formates, acetates, trifluoroacetates, maleates, tartrates, orotates, etc. Inorganic acid salts of alkali metals include salts such as fluorides, chlorides, bromides, sulfates, phosphates, etc. For the purposes of this specification, inorganic salts may be preferred over organic salts. Among inorganic salts, halides may be preferred over other inorganic salts. Among halides, chlorides are particularly preferred. Regardless of the type of salt used (organic or inorganic, etc.), water-soluble salts are preferred because they can be provided in the form of compositions that are not easily digested by bees (e.g., sucrose solutions) and fed to bees, thus indirectly absorbing the Varroa mite via this route. Whenever a particular salt is mentioned, it is always included in its hydrated or anhydrous form unless otherwise specified. The group of alkali metals includes lithium, sodium, potassium, rubidium, and cesium. Among the alkali metals, lithium is particularly preferred.

[0152] Among alkali metal salts, organic or inorganic salts of lithium that are hydrated or anhydrous are preferred over corresponding, such as potassium salts. Inorganic lithium salts, such as LiCl, are advantageous because they are less expensive than organic lithium salts. Therefore, inorganic lithium salts may be preferred over organic lithium salts in the group of lithium salts. Preferred inorganic lithium salts include, but are not limited to, lithium chloride, lithium bromide, lithium nitrate, lithium sulfate, lithium phosphate, etc. However, organic lithium salts may be advantageous because they can complex more than one lithium ion and because they are better absorbed by bees than inorganic lithium salts. Preferred organic lithium salts include, but are not limited to, lithium citrate, lithium carbonate, lithium lactate, lithium formate, lithium acetate, lithium trifluoroacetate, lithium maleate, lithium tartrate, lithium orotate, etc. In the group of inorganic lithium salts, halides are preferred over other inorganic salts. In the group of organic lithium salts, lactates, citrates, acetates, and carbonates may be preferred over other organic salts. In all aspects and embodiments described below, a preferred alkali metal salt is lithium chloride in hydrated or anhydrous form.

[0153] Like other reagents, alkali metal salts can be administered by a variety of methods. Preferably, the alkali metal salt is applied topically to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye, or to areas adjacent to the eye. The alkali metal salt may be included in a spinosad composition or administered to the subject as a separate composition before, during, or after the application of the spinosad composition. Preferably, an effective acaricide is applied. In some embodiments, the concentration is in the range of about 0.5 mM to about 150 mM of an alkali metal salt (e.g., LiCl).

[0154] Products

[0155] Other aspects of the invention include articles of manufacture that can be used to carry out the methods of the invention, such as spinosad compositions combined with containers; eye or face applicators; and kits.

[0156] One aspect of the invention is an article comprising the spinosad composition disclosed herein; and a container therein containing the topical composition. Thus, the spinosad composition can be contained in the container and can be removed for administration to a subject. The container may have a defined interior to contain the composition. Preferably, the composition and the interior of the container are sterile. Examples of containers include collapsible or non-collapsible tubes, bags, pouches, blister packs, strips, ampoules, medicine bottles, bottles, jars, or wide-mouth flasks. The container may be composed of one or more materials suitable for its use, such as aluminum, polyethylene (e.g., low-density polyethylene (LDPE), high-density polyethylene (HDPE), or polyethylene terephthalate (PET)), polypropylene, or glass.

[0157] In some embodiments, the container has a closure such as a lid that can be opened to remove the composition inside. The closure used to cover the container after the filling process should be as inert as possible, not causing undesirable interactions between the contents and the external environment, and should provide a complete seal. In addition to its protective function, the closure should also allow for easy and safe application of the spinosad composition. In some embodiments, the container has an tamper-evident closure. In some embodiments, the container has no closure, and the spinosad composition is obtained by tearing, separating, or breaking the material of the container.

[0158] In some embodiments, the container includes a rubber cap (composed of an elastomeric material), or a cap and / or an overseal, which may be composed of aluminum or plastic (e.g., polyethylene, polypropylene).

[0159] The container may include tamper-evident features, such as a tamper-evident closure. For example, eye ointments are typically supplied in small, sterile, collapsible tubes (containing, for example, up to about 5 grams of the composition) equipped with a tamper-evident applicator. The tube may include a nozzle shaped to allow application of the ointment without contaminating any remaining product in the tube. The container may include a child-proof closure that can be resealed, such as a push-to-turn or squeeze-to-turn type.

[0160] The container can be a single-dose or multi-dose container. In some embodiments, the container is a single-dose container (containing a single dose of the composition for treating ocular Demodex mite infection in the eyes of a human or animal subject, or for treating eye or skin diseases associated with ocular Demodex mite infection). In other embodiments, the container is a multi-dose container (containing multiple doses of the composition for treating ocular Demodex mite infection in the eyes of a human or animal subject, or for treating eye or skin diseases associated with ocular Demodex mite infection).

[0161] Optionally, the container containing the composition described herein is light-resistant and has a tight seal. For example, one or more containers may comprise one of the dermatologically or ophthalmologically acceptable spinosad compositions described herein. In some embodiments, the container prevents the ingress of certain wavelengths of light and prolonged high temperatures and / or air. In some embodiments, the container is a sealed, light-resistant container.

[0162] Labels may be on or associated with a container. A label may be located on the container when the letters, numbers, or other characters that make up the label are pasted, molded, or etched into the container itself; a label may be associated with the container when it is present in a receptor or carrier that also holds the container, such as as a packaging insert. Labels can be used to indicate the contents intended for a specific therapeutic application. Labels may also indicate instructions for use of the contents, such as those described herein.

[0163] Another aspect of the invention is an ocular or facial applicator for treating ocular Demodex mite infestations in human or animal subjects, or for treating eye or skin diseases associated with ocular Demodex mite infestations. The applicator is pretreated with or contains a spinosad composition disclosed herein. The applicator can be used by a subject or another person to topically apply a composition containing one or more spinosad compounds to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye or areas adjacent to the eye. In some embodiments, the applicator is a cotton swab, cosmetic pad, wiping cloth, wiping stick, small towel, sponge, gauze, powder puff, stick, brush, or comb.

[0164] Another aspect of the invention is a kit for treating ocular Demodex mite infection in human or animal subjects, or for treating eye or skin diseases associated with ocular Demodex mite infection. The kit comprises a spinosad composition and an eye or face applicator. The kit may further include containers disclosed herein for containing the composition. Examples of applicators include cotton swabs, cosmetic pads, wiping cloths, wiping sticks, small towels, sponges, gauze, powder puffs, sticks, brushes, or combs.

[0165] In some embodiments, the applicator is pretreated with or contains the composition. In this way, when the applicator is pretreated with or contains the composition, it can also be used as a container.

[0166] The kit may include a set of printed or digital instructions for treating ocular Demodex mite infestation in human or animal subjects by topically applying the composition to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye, or to areas adjacent to the eye, or for treating ocular or skin diseases associated with ocular Demodex mite infestation. For example, the kit may include instructions for use comprising the steps of: topically applying the spinosad composition to the affected area or other target area; repeating the administration steps until sufficient relief or elimination of one or more signs or symptoms associated with Demodex mite infestation, reduction of the number of surviving Demodex mites, elimination of surviving Demodex mites in the desired area, reduction of reinfection with Demodex mites, or prevention or delay of the onset of Demodex mite infestation.

[0167] The kit may include packaging materials that are spaced apart to accommodate one or more containers, such as liquid bottles, tubes, etc., each container including one of the individual elements to be used in the methods described herein. For example, packaging materials for packaging pharmaceutical products are shown only by way of example, such as U.S. Patent Nos. 5,323,907, 5,052,558, and 5,033,252. Examples of pharmaceutical packaging materials include, but are not limited to, blister packs, bottles, tubes, pumps, bags, liquid bottles, opaque, sealed containers, syringes, vials, and any packaging materials suitable for the selected formulation and the intended route of administration and treatment.

[0168] The kit may include one or more additional containers, each containing one or more materials that are commercially and user-desired for the treatment of Demodex mites described herein. Non-limiting embodiments of such materials include, but are not limited to, buffers, diluents, carriers, packaging, containers, labels on vials and / or tubes listing the contents and / or instructions for use, and packaging inserts with instructions for use.

[0169] Labels may be on or associated with containers. A label may be on the container when the letters, numbers, or other characters that make up the label are pasted, molded, or etched into the container itself; a label may be associated with the container when it is present in a receiver or carrier that also holds the container, such as as a packaging insert. Labels may be used to indicate the contents intended for a specific therapeutic application. Labels may also indicate instructions for use of the contents, such as those described herein.

[0170] In some embodiments of the kit, the spinosad composition may be present in a packaging or dispenser device that may contain one or more unit dosage forms of the spinosad composition disclosed herein. The packaging may, for example, contain metal or plastic foil, such as blister packs. The packaging or dispenser device may be accompanied by instructions for administration. The packaging or dispenser may also be accompanied by a notification associated with the container, in the form prescribed by a government agency regulating the manufacture, use, or sale of the drug, reflecting that agency’s approval of the form of the drug for human or veterinary use. For example, such a notification may be a prescription drug label approved by the U.S. Food and Drug Administration or an approved product insert. A composition containing the spinosad composition provided herein may also be prepared in a compatible drug carrier, placed in an appropriate container, and labeled for the treatment of a specified disease.

[0171] definition

[0172] As used herein, the term "amplification" refers to increasing the copy number of a target nucleic acid molecule. The resulting amplification product is called an "amplifier." Amplification of nucleic acid molecules (e.g., DNA or RNA molecules) refers to the use of techniques to increase the copy number of nucleic acid molecules in a sample, such as the copy number of nucleic acid in Demodex mites (e.g., Demodex folliculorum or Demodex sebaceous gland). One embodiment of amplification is polymerase chain reaction (PCR), in which a sample is contacted with a pair of oligonucleotide primers under conditions that allow primers to hybridize with a nucleic acid template in the sample. The primers are extended under suitable conditions, dissociated from the template, annealed, extended again, and dissociated again to amplify the copy number of the nucleic acid. This cycle can be repeated. Amplification products can be characterized by techniques such as electrophoresis, restriction endonuclease cleavage patterns, oligonucleotide hybridization or ligation, and / or nucleic acid sequencing.

[0173] Primer pairs can be used to amplify target nucleic acid sequences, for example, by PCR, real-time PCR, or other nucleic acid amplification methods known in the art. The “upstream” or “forward” primer is the 5’ primer relative to a reference point on the nucleic acid sequence. The “downstream” or “reverse” primer is the 3’ primer relative to a reference point on the nucleic acid sequence. Typically, at least one forward primer and one reverse primer are included in the amplification reaction. PCR primer pairs can be used, for example, by using a computer program (version 0.5) designed for this purpose (e.g., Primer(primer)). Whitehead Institute for Biomedical Research, Cambridge, Massachusetts) or The software (AB Inc. of Foster City, California, Applied Biosystems) is derived from known sequences (e.g., Demodex mite sequences).

[0174] Methods for preparing and using primers are described, for example, in Sambrook et al. (1989) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor, NY; and Ausubel et al. (1987) Current Protocols in Molecular Biology, Greene Publ. Assoc. & Wiley-Intersciences. In one embodiment, the primer includes a marker.

[0175] Other embodiments of in vitro amplification techniques include quantitative real-time PCR; reverse transcriptase PCR (RT-PCR); real-time PCR; real-time reverse transcriptase PCR (rt RT-PCR); nested PCR; strand displacement amplification (see US Patent No. 5,744,311); transcription-free isothermal amplification (see US Patent No. 6,033,881); repair chain reaction amplification (see WO 90 / 01069); ligase chain reaction amplification (see European Patent Publication EP-A-320 308); gap-filling ligase chain reaction amplification (see US Patent No. 5,427,930); coupled ligase detection and PCR (see US Patent No. 6,027,889); and NASBA. TM RNA amplification without transcription (see U.S. Patent No. 6,025,134), etc.

[0176] As used herein, the term "detection" refers to determining the presence of a reagent (e.g., a signal, a specific nucleotide, amino acid, nucleic acid molecule, and / or organism), such as Demodex mites, such as *Demodex folliculorum* and / or *Demodex sebaceous*. In some embodiments, this may further include quantification. For example, the use of probes allows for the detection of fluorophores, such as the detection of signals from fluorophores, which can be used to determine the presence of nucleic acids corresponding to the nucleic acids of Demodex mites (*Demodex folliculorum* and / or *Demodex sebaceous*). Detection of nucleic acid molecules of *Demodex folliculorum* and / or *Demodex sebaceous* indicates the presence of *Demodex folliculorum* and / or *Demodex sebaceous* in a sample, e.g., *Demodex folliculorum* and / or *Demodex sebaceous* infection.

[0177] The terms “acaricidal” and “miticidal” are used interchangeably herein to refer to the ability to kill mites of the genus *Demodex* at any stage of their life cycle, or the ability to interfere with the growth or life cycle of *Demodex* in any way that results in an overall reduction in the *Demodex* population. In some embodiments, the *Demodex* species are *Demodex folliculorum*, *Demodex sebaceous gland*, or both. For example, the term “acaricidal” includes inhibiting or eliminating the reproductive capacity of pests, as well as inhibiting the development of pests from one form to a more mature form, such as the transition between various larval forms or from larva to first nymph or from first nymph to nymph, or from nymph to adult. Furthermore, the term “acaricidal” is intended to encompass all stages of the mite life cycle. Thus, for example, the term includes larval, ovicidal, and adult-killing effects.

[0178] As used herein, the term "mite-effective" refers to an amount or concentration of spinosad or other acaricide that is sufficient to reduce the number of Demodex mites at a anatomical site compared to a corresponding anatomical site in which no amount or concentration of spinosad or other acaricide is present.

[0179] As used herein, the terms “treatment,” “treating,” and their grammatical variations refer to the improvement, prevention, or reversal of a disease, condition, or at least one of its signs or symptoms. In some embodiments, “treatment” or “treating” refers to the improvement, prevention, or reversal of at least one measurable bodily parameter associated with the disease or condition being treated, not necessarily in the subject’s body or as identified by the subject. In yet another embodiment, “treatment” or “treating” refers to inhibiting or slowing the progression of a disease or condition, whether physically, for example, by stabilizing identifiable signs or symptoms, or physiologically, for example, by stabilizing physical parameters, or both. In yet another embodiment, “treatment” or “treating” refers to delaying the onset of a disease or condition.

[0180] Exemplary Implementation

[0181] 1. A method for treating ocular Demodex mite infection in the eyes of a human or animal subject, or for treating eye or skin diseases associated with ocular Demodex mite infection, said method comprising topically applying a composition comprising one or more spinosad compounds to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye, or to areas adjacent to the eye.

[0182] 2. The method according to embodiment 1, wherein the subject suffers from the disease, and the disease is one or more of the following: Demodex blepharitis (also known as Demodex blepharitis), ocular erythematous acne caused by Demodex mites, facial erythematous acne caused by Demodex mites, dry eye syndrome, meibomian gland dysfunction, meibomian gland cysts, meibomianitis, follicular inflammation, nonspecific facial dermatitis, infiltrative keratoconjunctivitis, nodular scar deposition, or corneal neovascularization.

[0183] 3. The method according to embodiment 1 or 2, wherein the treatment alleviates one or more signs or symptoms of a subject selected from the following: itching, burning, foreign body sensation, crusting and redness of the eyelid margin, blurred vision, cylindrical scales, eyelash misalignment, trichiasis, eyelash loss, eyelid margin inflammation, meibomian gland dysfunction, conjunctivitis, and blepharitis.

[0184] 4. The method according to any one of embodiments 1 to 3, wherein the composition is applied locally to a region adjacent to the eye, wherein the region includes one or more of the following: eyelid, eyelid margin, eyelash, eyelash follicle, eyebrow, or eyebrow follicle.

[0185] 5. The method according to any one of embodiments 1 to 3, wherein the composition is applied locally to a region adjacent to the eye, wherein the region includes the sebaceous gland openings of the eyelid (e.g., one or more of Zeiss glands, Mohr's glands, or meibomian glands).

[0186] 6. The method according to any of the foregoing embodiments, wherein the Demodex mite comprises Demodex folliculorum, Demodex sebaceous gland mite, or both.

[0187] 7. The method according to any of the foregoing embodiments, wherein the subject is symptomatic at the time of administration.

[0188] 8. The method according to any one of embodiments 1 to 6, wherein the subject is asymptomatic at the time of administration.

[0189] 9. The method according to any of the foregoing embodiments further includes confirming the presence of Demodex mites prior to the application.

[0190] 10. The method according to embodiment 9, wherein the confirmation includes identifying Demodex mites in vivo or in ex vivo biological samples (e.g., hair or skin samples), or detecting molecules associated with the presence of Demodex mites (e.g., chitin, chitin synthase, lipase, Bacillus antigen).

[0191] 11. The method according to any of the foregoing embodiments, wherein an effective amount of the composition is applied topically to reduce the number of surviving Demodex mites in the eyes and adjacent skin and hair.

[0192] 12. The method according to any of the foregoing embodiments, wherein an effective amount of the composition is applied topically to eliminate the presence of surviving Demodex mites from the eyes and adjacent skin and hair.

[0193] 13. The method according to any of the foregoing embodiments, wherein the one or more spinosad compounds have the chemical structure of formula (I) or a pharmaceutically acceptable salt thereof:

[0194]

[0195] Where R 1 R 2 R 3 R 4 R 5 R 6 R 7 R 8 R 9 R 10 R 11 R 12 R 13 R 14R 15 and R 16 It can be independently selected from: unsubstituted; H; F; Cl; Br; I; OH; CN; (C 1-4 Alkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl; (C 2-4 Alkenyl, such as vinyl, propenyl, butenyl, wherein the double bond may be located at any position on the alkenyl carbon chain, and includes any alkenyl conformational isomer; alkynyl; aralkyl; alkylaryl; haloalkyl; heteroalkyl; aryl; heterocyclic; cycloalkyl; cycloalkenyl; cycloalkynyl; hydroxyalkyl; aminoalkyl; amino; alkylamino; arylamino; dialkylamino; alkylarylamino; diarylamino; acylamino; hydroxyl; thiol; thioalkyl; alkoxy; alkylthio; alkoxyalkyl; aryloxy; arylalkoxy; acyloxy; nitro; carbamoyl; trifluoromethyl; phenoxy; benzyloxy; phosphonic acid; phosphate ester; sulfonic acid (-SO3H); sulfonate; sulfonamide; alkylaryl; arylalkyl; carbamoyl; amino; alkylamino; arylamino; dialkylamino; alkylarylamino; diarylamino; alkylthio; heteroalkyl; alkyltriphenylphosphine; heterocyclic; ketone (=O); ether (-OR) 17 ); and esters (-COOR) 18 and -OC(=O)R 18 );

[0196] Where R 5 and R 7 It could be a double bond within a cyclopentane ring;

[0197] Where R 11 and R 13 It could be a double bond within the cyclohexane ring;

[0198] Where R 17 It can be independently selected from the group consisting of the following groups: (C 1-4 Alkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl; (C 2-4 Alkenyl groups, such as vinyl, propenyl, butenyl, wherein the double bond may be located at any position on the alkenyl carbon chain and includes any alkenyl conformational isomer; and ynyl groups;

[0199] Where R 18 It can be independently selected from the group consisting of the following groups: (C 1-4 Alkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl; (C 2-4 Alkenyl groups, such as vinyl, propenyl, butenyl, wherein the double bond may be located at any position on the alkenyl carbon chain and includes any alkenyl conformational isomer; and ynyl groups.

[0200] 14. The method according to any of the foregoing embodiments, wherein the one or more spinosad compounds are selected from spinosad A, spinosad B, spinosad C, spinosad D, spinosad E, spinosad F, spinosad G, spinosad H, spinosad I, spinosad J, spinosad K, spinosad L, spinosad M, spinosad N, spinosad O, spinosad P, spinosad R, spinosad S, spinosad T, spinosad U, spinosad V, spinosad W, spinosad Y, spinosad N-demethyl D, spinosad Spinosad N-demethyl K, spinosad N, N-dimethyl K, spinosad N-demethyl P, spinosad 2'-HA, spinosad 2'-HD, spinosad 2'-O-ethyl A, spinosad 3'-HA, spinosad 3'-O-ethyl A, spinosad 3'-On-propyl A, spinosad 3'-On-butyl A, spinosad 3'-O-allyl A, spinosad-O-CH2CF3A, spinosad 4'-HA, spinosad 4'-O-ethyl A, or spinosad 2',3',4'-tri-O-ethyl A.

[0201] 15. The method according to any of the foregoing embodiments, wherein the one or more spinosad compounds comprise a combination of spinosad A and spinosad D.

[0202] 16. The method according to embodiment 15, wherein spinosad A and spinosad D are present in the composition in a ratio of spinosad A to spinosad D of about 18:2 to about 16:4.

[0203] 17. The method according to embodiment 15, wherein spinosad A and spinosad D are present in the composition in a ratio of approximately 17:3 spinosad A to spinosad D.

[0204] 18. The method according to any of the foregoing embodiments, wherein the composition does not contain any active agent other than the one or more spinosad compounds.

[0205] 19. The method according to any one of embodiments 1 to 18, wherein the composition does not contain any other reagents that have acaricidal activity against the Demodex mites.

[0206] 20. The method according to any one of embodiments 1 to 17, wherein the method does not include topical application of any active agent other than one or more spinosad compounds to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye, or areas adjacent to the eye.

[0207] 21. The method according to any one of embodiments 1 to 17, wherein the method does not include topical application of an agent having acaricidal activity against Demodex mites, other than one or more spinosad compounds, to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye, or to areas adjacent to the eye.

[0208] 22. The method according to any one of embodiments 1 to 17, wherein the composition further comprises other agents having acaricidal activity against the Demodex mites.

[0209] 23. The method according to embodiment 22, wherein the other agent having acaricidal activity against Demodex mites is one or more agents selected from the following substances: ivermectin, pyrethroids, pyrethroids (e.g., permethrin, cypermethrin, or D-phenylephrine), tea tree oil (TTO), TTO components (e.g., 4-terpene alcohol (T4O)), metronidazole, hypochlorous acid (HOCl), essential oils (e.g., peppermint oil or sage), alkali metal salts (e.g., lithium salts), thiophosphates (e.g., non-volatile, fat-soluble thiophosphates (e.g., phosmet) or formamidin (e.g., amitraz).

[0210] 24. The method according to any one of embodiments 1 to 17, further comprising applying other agents having acaricidal activity against the Demodex mites before, during, or after the topical application of the composition having one or more spinosad compounds, wherein the other agents are applied in the form of a separate composition.

[0211] 25. The method according to embodiment 24, wherein the other agent having acaricidal activity against Demodex mites is one or more agents selected from the group consisting of: ivermectin, pyrethroids, pyrethroids (e.g., permethrin, cypermethrin, or D-phenylephrine), tea tree oil (TTO), TTO components (e.g., 4-terpene alcohol (T4O)), metronidazole, hypochlorous acid (HOCl), essential oils (e.g., peppermint oil or sage), alkali metal salts (e.g., lithium salts), thiophosphates (e.g., non-volatile, fat-soluble thiophosphates (e.g., phosmet) or formamidin (e.g., amitraz).

[0212] 26. The method according to any of the foregoing embodiments further includes administering an antibiotic to the subject, wherein the antibiotic is present in the composition together with the one or more spinosad compounds or in a separate composition.

[0213] 27. The method according to embodiment 26, wherein the antibiotic has antibacterial activity against one or more of streptococci, staphylococci, or Bacillus subtilis.

[0214] 28. The method according to any of the foregoing embodiments further comprises administering an anti-inflammatory agent to the subject, wherein the anti-inflammatory agent is present in the composition together with the one or more spinosad compounds or in a separate composition.

[0215] 29. The method according to embodiment 28, wherein the anti-inflammatory agent is selected from glucocorticoids or other steroids (e.g., prednisone, cortisone acetate, prednisolone, methylprednisolone, dexamethasone, betamethasone, triamcinolone, beclomethasone, flucortisone acetate, deoxycorticosterone acetate, aldosterone), nonsteroidal anti-inflammatory drugs (e.g., salicylates, aryl fatty acids, 2-arylpropionic acid, N-aryl-o-aminobenzoic acid, oxacin, coxib, or sulfanilide), Cox-2 specific inhibitors (… Examples include vardicoxib, celecoxib, or roficoxib; leflunomide; glucosinolate, gold thiocarbamate, aurofin; sulfasalazine; hydroxychloroquine; minocycline; TNF-α binding protein (e.g., infliximab, etanercept, or adalimumab); abatacept; anaglycinol; interferon-β; interferon-γ; interleukin-2; allergy vaccines; antihistamines; antileukotrienes; β-agonists; theophylline or anticholinergics; antibiotics; tacrolimus or retinoids.

[0216] 30. The method according to any of the foregoing embodiments, wherein the composition is a solution, suspension, ointment, spray, lotion, gel, paste, balm, foam, mousse, scrub or detergent (e.g., shampoo or soap), cream or ointment.

[0217] 31. The method according to any of the foregoing embodiments, wherein the composition is applied topically using an applicator (e.g., cotton swab, makeup pad, wiping cloth, wiping stick, small towel, sponge, gauze, powder puff, stick, brush or comb).

[0218] 32. The method according to any of the foregoing embodiments, wherein the composition is a cosmetic.

[0219] 33. The method according to embodiment 32, wherein the cosmetic is mascara, eyeshadow, or eyeliner.

[0220] 34. The method according to any of the foregoing embodiments, wherein the composition further comprises one or more excipients selected from petrolatum, mineral oil, propylparaben, methylparaben, lanolin, chlorobutanol, water, lanolin alcohol, sodium thiosulfate, sodium dihydrogen phosphate, phenylmercuric acetate, mannitol, zinc chloride, sodium phosphate, potassium acetate, hydroxypropyl methylcellulose, gentamicin sulfate, boric acid, sodium hydroxide, lanolin oil, carbomer homopolymer A or B (allyl pentaerythritol crosslinked) or benzalkonium chloride.

[0221] 35. The method according to any one of embodiments 1 to 34, wherein the composition is an emulsion and further comprises one or more excipients selected from water, sodium hydroxide, polysorbate 80, glycerin, castor oil, carbomer copolymer type A, sodium acetate, boric acid, sorbic acid, disodium EDTA, or silicone oil or silicone polymer gel (e.g., dimethicone or cyclomethicone).

[0222] 36. The method according to any of the foregoing embodiments, wherein the composition further comprises a gelling agent (e.g., gum, agar, carrageenan, petrolatum, or cellulose polymer (e.g., hydroxyethyl cellulose or hydroxymethyl cellulose)).

[0223] 37. The method according to any of the foregoing embodiments further comprises one or more of a solvent, cosolvent, modifier, emollient, preservative, antioxidant, humectant or solubilizer.

[0224] 38. The method according to any of the foregoing embodiments, wherein the composition comprises 0.1% to 10% (w / v) of one or more spinosad compounds.

[0225] 39. The method according to any of the foregoing embodiments, wherein the composition is removed and dispensed from a container with or without a closure (e.g., a lid) prior to application.

[0226] 40. The method according to embodiment 39, wherein the container is a foldable or non-foldable tube, bag, pouch, blister pack, strip, ampoule, medicine bottle, bottle, jar, or wide-mouth bottle.

[0227] 41. The method according to embodiment 39 or 40, wherein the container comprises one or more materials selected from aluminum, polyethylene (e.g., low-density polyethylene (LDPE), high-density polyethylene (HDPE) or polyethylene terephthalate (PET)), polypropylene or glass.

[0228] 42. The method according to any one of embodiments 39 to 41, wherein the container is a single-dose container (containing a single dose of the composition for treating ocular Demodex mite infection in the eyes of a human or animal subject or for treating eye or skin diseases associated with ocular Demodex mite infection).

[0229] 43. The method according to any one of embodiments 39 to 41, wherein the container is a multi-dose container (containing multiple doses of the composition for treating ocular Demodex mite infection in the eyes of human or animal subjects or for treating eye or skin diseases associated with ocular Demodex mite infection).

[0230] 44. The method according to any one of embodiments 39 to 43, wherein the container has an interior for containing the composition, and wherein the interior of the container and the composition are sterile.

[0231] 45. The method according to any of the foregoing embodiments, wherein the composition is a composition from one of Table 1, Table 2, Table 3, Table 4 or Table 5.

[0232] 46. ​​A topical composition comprising 0.1% to 10% (w / v) of one or more spinosad compounds.

[0233] 47. The topical composition according to embodiment 46, wherein the one or more spinosad compounds have the chemical structure of formula (I) or a pharmaceutically acceptable salt thereof:

[0234]

[0235] Where R 1 R 2 R 3 R 4 R 5 R 6 R 7 R 8 R 9 R 10 R 11 R 12 R 13 R 14 R 15 and R 16 It can be independently selected from: unsubstituted; H; F; Cl; Br; I; OH; CN; (C 1-4 Alkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl; (C 2-4 Alkenyl, such as vinyl, propenyl, butenyl, wherein the double bond may be located at any position on the alkenyl carbon chain, and includes any alkenyl conformational isomer; alkynyl; aralkyl; alkylaryl; haloalkyl; heteroalkyl; aryl; heterocyclic; cycloalkyl; cycloalkenyl; cycloalkynyl; hydroxyalkyl; aminoalkyl; amino; alkylamino; arylamino; dialkylamino; alkylarylamino; diarylamino; acylamino; hydroxyl; thiol; thioalkyl; alkoxy; alkylthio; alkoxyalkyl; aryloxy; arylalkoxy; acyloxy; nitro; carbamoyl; trifluoromethyl; phenoxy; benzyloxy; phosphonic acid; phosphate ester; sulfonic acid (-SO3H); sulfonate; sulfonamide; alkylaryl; arylalkyl; carbamoyl; amino; alkylamino; arylamino; dialkylamino; alkylarylamino; diarylamino; alkylthio; heteroalkyl; alkyltriphenylphosphine; heterocyclic; ketone (=O); ether (-OR) 17); and esters (-COOR) 18 and -OC(=O)R 18 );

[0236] Where R 5 and R 7 It could be a double bond within a cyclopentane ring;

[0237] Where R 11 and R 13 It could be a double bond within the cyclohexane ring;

[0238] Where R 17 It can be independently selected from the group consisting of the following groups: (C 1-4 Alkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl; (C 2-4 Alkenyl groups, such as vinyl, propenyl, butenyl, wherein the double bond may be located at any position on the alkenyl carbon chain and includes any alkenyl conformational isomer; and ynyl groups;

[0239] Where R 18 It can be independently selected from the group consisting of the following groups: (C 1-4 Alkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl; (C 2-4 Alkenyl groups, such as vinyl, propenyl, butenyl, wherein the double bond may be located at any position on the alkenyl carbon chain and includes any alkenyl conformational isomer; and ynyl groups.

[0240] 48. The topical composition according to embodiment 46 or 47, wherein the one or more spinosad compounds comprise a combination of spinosad A and spinosad D.

[0241] 49. The topical composition according to embodiment 48, wherein spinosad A and spinosad D are present in the composition in a ratio of spinosad A to spinosad D of about 18:2 to about 16:4.

[0242] 50. The topical composition according to embodiment 48, wherein spinosad A and spinosad D are present in the composition in a ratio of spinosad A to spinosad D of about 17:3.

[0243] 51. The topical composition according to any one of embodiments 46 to 50, wherein the composition does not contain an active agent other than one or more spinosad compounds.

[0244] 52. The topical composition according to any one of embodiments 46 to 50, wherein the composition does not contain any other agents having acaricidal activity against Demodex mites.

[0245] 53. The topical composition according to any one of embodiments 46 to 50, wherein the composition further comprises other agents having acaricidal activity against Demodex mites.

[0246] 54. The topical composition according to embodiment 53, wherein the other agent having acaricidal activity against Demodex mites is one or more agents selected from the group consisting of: ivermectin, pyrethroids, pyrethroids (e.g., permethrin, cypermethrin, or D-benzylpyrethrin), tea tree oil (TTO), TTO components (e.g., 4-terpene alcohol (T4O)), metronidazole, hypochlorous acid (HOCl), essential oils (e.g., peppermint oil or sage), alkali metal salts (e.g., lithium salts), thiophosphates (e.g., non-volatile, fat-soluble thiophosphates (e.g., phosmet) or formamidin (e.g., amitraz).

[0247] 55. The topical composition according to any one of embodiments 46 to 54 further comprises an antibiotic agent.

[0248] 56. The topical composition according to embodiment 55, wherein the antibiotic has antibacterial activity against one or more of streptococci, staphylococci, or Bacillus subtilis.

[0249] 57. The topical composition according to any one of embodiments 46 to 56, further comprising an anti-inflammatory agent.

[0250] 58. The topical composition according to embodiment 57, wherein the anti-inflammatory agent is selected from glucocorticoids or other steroids (e.g., prednisone, cortisone acetate, prednisolone, methylprednisolone, dexamethasone, betamethasone, triamcinolone, beclomethasone, flucortisone acetate, deoxycorticosterone acetate, aldosterone), nonsteroidal anti-inflammatory drugs (e.g., salicylates, aryl fatty acids, 2-arylpropionic acid, N-aryl-o-aminobenzoic acid, oxacin, coxib, or sulfanilide), Cox-2 specific inhibitors. Agents (e.g., vardicoxib, celecoxib, or roficoxib), leflunomide, glucosinolate, gold thiomalate, aurofin, sulfasalazine, hydroxychloroquine, minocycline, TNF-α binding protein (e.g., infliximab, etanercept, or adalimumab), abatacept, anaerobicin, interferon-β, interferon-γ, interleukin-2, allergy vaccines, antihistamines, antileukotrienes, β-agonists, theophylline or anticholinergics, antibiotics, tacrolimus or retinoids.

[0251] 59. The topical composition according to any one of embodiments 46 to 58, wherein the composition is a solution, suspension, ointment, spray, lotion, gel, paste, balm, foam, mousse, scrub or detergent (e.g., shampoo or soap), cream or ointment.

[0252] 60. The topical composition according to any one of embodiments 46 to 59, wherein the composition is a cosmetic.

[0253] 61. The topical composition according to embodiment 60, wherein the composition is mascara, eyeshadow, or eyeliner.

[0254] 62. A topical composition according to any one of embodiments 46 to 61, wherein the composition is an ointment and further comprises one or more excipients selected from petrolatum, mineral oil, propylparaben, methylparaben, lanolin, chlorobutanol, water, lanolin alcohol, sodium thiosulfate, sodium dihydrogen phosphate, phenylmercuric acetate, mannitol, zinc chloride, sodium phosphate, potassium acetate, hydroxypropyl methylcellulose, gentamicin sulfate, boric acid, sodium hydroxide, lanolin oil, carbomer homopolymer type B (allyl pentaerythritol crosslinked) or benzalkonium chloride.

[0255] 63. A topical composition according to any one of embodiments 46 to 61, wherein the composition is an emulsion and further comprises one or more excipients selected from water, sodium hydroxide, polysorbate 80, glycerin, castor oil, carbomer copolymer type A, sodium acetate, boric acid, sorbic acid, disodium ethylenediaminetetraacetate, carbomer copolymer type A (allyl pentaerythritol crosslinked), or silicone oil or silicone polymer gel (e.g., dimethicone or cyclomethicone).

[0256] 64. A topical composition according to any one of embodiments 46 to 63, wherein the composition further comprises a gelling agent (e.g., gum, agar, carrageenan, petrolatum, or cellulose polymer (e.g., hydroxyethyl cellulose or hydroxymethyl cellulose)).

[0257] 65. The topical composition according to any one of embodiments 46 to 64 further comprises one or more of a solvent, a cosolvent, a modifier, a skin emulsifier, a preservative, an antioxidant, a humectant, or a solubilizer.

[0258] 66. A topical composition according to any one of embodiments 46 to 64, wherein the composition is a composition in one of Table 1, Table 2, Table 3, Table 4 or Table 5.

[0259] 67. An article comprising the topical composition of any one of embodiments 46 to 66; and a container therein containing the topical composition.

[0260] 68. The article of manufacture according to embodiment 67, wherein the container has a closure (e.g., a lid) that can be opened to remove the composition inside the container.

[0261] 69. The article of manufacture according to embodiment 67 or 68, wherein the container is a foldable or non-foldable tube, bag, pouch, blister pack, strip, ampoule, medicine bottle, bottle, jar or wide-mouth bottle.

[0262] 70. The article of any one of embodiments 67 to 69, wherein the container comprises one or more materials selected from aluminum, polyethylene (e.g., low-density polyethylene (LDPE), high-density polyethylene (HDPE) or polyethylene terephthalate (PET)), polypropylene or glass.

[0263] 71. The article of any one of embodiments 67 to 70, wherein the container is a single-dose container (containing a single dose of the composition for treating ocular Demodex mite infection in the eyes of a human or animal subject or for treating eye or skin diseases associated with ocular Demodex mite infection).

[0264] 72. The article of any one of embodiments 67 to 70, wherein the container is a multi-dose container (containing multiple doses of the composition for treating ocular Demodex mite infection in the eyes of human or animal subjects or for treating eye or skin diseases associated with ocular Demodex mite infection).

[0265] 73. The article of any one of embodiments 67 to 72, wherein the container has an interior for containing the composition, and wherein the interior of the container and the composition are sterile.

[0266] 74. An eye or face applicator pretreated with or containing the topical composition of any one of embodiments 46 to 66.

[0267] 75. An eye or face applicator according to embodiment 74, wherein the applicator is a cotton swab, makeup pad, wiping cloth, wiping stick, small towel, sponge, gauze, powder puff, stick, brush or comb.

[0268] 76. A kit comprising the topical composition of any one of embodiments 46 to 66; and an eye or face applicator.

[0269] 77. The kit according to embodiment 76, wherein the kit further comprises a container for containing the composition.

[0270] 78. The kit according to embodiment 76, wherein the applicator is pretreated with or contains the composition.

[0271] 79. The kit according to any one of embodiments 76 to 78, wherein the applicator is a cotton swab, a makeup pad, a wiping cloth, a wiping stick, a small towel, a sponge, gauze, a powder puff, a stick, a brush, or a comb.

[0272] 80. The kit according to any one of embodiments 76 to 79, further comprising instructions for treating Demodex mite infection of the eyes of human or animal subjects by applying the composition topically to the ocular surface (conjunctiva and / or cornea) or other anatomical structures of the eye, or to areas adjacent to the eyes, or for treating eye or skin diseases associated with ocular Demodex mite infection.

[0273] All patents, patent applications, provisional applications, and application publications mentioned or cited herein, including all figures and tables, are incorporated herein by reference in their entirety, provided that they do not contradict the express teachings of this specification.

[0274] It should be understood that the embodiments and implementations described herein are for illustrative purposes only, and various modifications or changes thereto will be suggested to those skilled in the art and will be included within the spirit and scope of this application and the scope of the appended claims. Furthermore, any element or limitation of any invention or implementation thereof disclosed herein may be combined with any and / or all other elements or limitations disclosed herein (alone or in any combination) or any other invention or implementation thereof, and all such combinations are to be understood as being within, but not limited to, the scope of this invention.

Claims

1. Use in the preparation of a topical composition comprising one or more spinosad compounds for treating ocular Demodex mite infestation in a human subject, wherein the composition is applied topically to the ocular surface or other anatomical structures of the eye, or to an area adjacent to the eye, wherein, The subject suffers from a disease, and the disease is one or more of the following: blepharitis caused by Demodex mites, ocular erythematous acne caused by Demodex mites, dry eye syndrome, meibomian gland dysfunction, meibomian gland cysts, meibomianitis, infiltrative keratoconjunctivitis, nodular scar deposition, corneal neovascularization, palpebral conjunctivitis, and blepharokeratitis. The composition does not contain any active agent other than the one or more spinosad compounds, and the composition does not contain any other agents with acaricidal activity against the Demodex mites. The one or more spinosad compounds are a combination of spinosad A and spinosad D, and the spinosad A and spinosad D are present in the composition in a ratio of 18:2 to 16:

4.

2. The use according to claim 1, wherein the composition is applied topically to an area adjacent to the eye, wherein the area includes one or more of the following: eyelid, eyelid margin, eyelash, eyelash follicle, eyebrow, or eyebrow follicle.

3. The use according to claim 1, wherein the composition is applied topically to an area adjacent to the eye, wherein the area includes the sebaceous gland openings of the eyelid.

4. The use according to claim 1, wherein the Demodex mite infection is caused by Demodex folliculorum.

5. The use according to claim 1, wherein the Demodex mite infection is caused by Demodex sebaceous gland.

6. The use according to claim 1, wherein the Demodex mite infection is caused by Demodex folliculorum and Demodex sebaceous gland.

7. The use according to claim 1, wherein spinosad A and spinosad D are present in the composition in a ratio of 17:3 spinosad A to spinosad D.

8. The use according to claim 1, wherein the composition is a solution, suspension, ointment, spray, lotion, gel, paste, balm, foam, scrub or detergent, or ointment.

9. The use according to claim 1, wherein the composition further comprises one or more excipients selected from petrolatum, mineral oil, propylparaben, methylparaben, lanolin, chlorobutanol, water, lanolin alcohol, sodium thiosulfate, sodium dihydrogen phosphate, phenylmercuric acetate, mannitol, zinc chloride, sodium phosphate, potassium acetate, hydroxypropyl methylcellulose, gentamicin sulfate, boric acid, sodium hydroxide, lanolin oil, carbomer homopolymer A or B, or benzalkonium chloride.

10. The use according to claim 1, wherein the composition is an emulsion and further comprises one or more excipients selected from water, sodium hydroxide, polysorbate 80, glycerin, castor oil, carbomer copolymer type A, sodium acetate, boric acid, sorbic acid, disodium EDTA, or silicone oil or silicone polymer gel.

11. The use according to claim 1, wherein the composition further comprises a gelling agent.

12. The use according to claim 1, wherein the composition further comprises one or more of a solvent, cosolvent, modifier, emollient, preservative, antioxidant, humectant, or solubilizer.

13. The use according to claim 1, wherein the composition comprises 0.1% to 10% (w / v) of one or more spinosad compounds.

14. The use according to claim 1, wherein the composition is a composition from any one of Table 1, Table 2, Table 3, Table 4, or Table 5. Table 1 Table 2 Table 3 Table 4 Table 5 。

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