Compositions and methods of their use

Abstract

There are provided compositions comprising two particular cannabinoids and a third active ingredient selected from a particular third cannabinoid and beta-caryophyllene. Methods for making the compositions and using them to control the growth of a particular bacterium are also disclosed.

Description

[0001] Compositions and Methods of Their Use

[0002] Related Case

[0003] This application claims Paris Convention priority from, and the US benefit of, US Provisional application serial no. 63 / 627,438, filed January 31, 2024 and entitled "Compositions and Methods of Their Use"; the contents of this provisional application are incorporated herein by reference.

[0004] Background

[0005] Periodontal disease represents a group of oral inflammatory infections initiated by oral pathogens which exist as a complex biofilm on the tooth surface and cause destruction to tooth supporting tissues. The severity of this disease ranges from mild and reversible inflammation of the gingiva (gingivitis) to chronic destruction of connective tissues, the formation of periodontal pocket and ultimately result in loss of teeth. While human subgingival plaque harbors more than 500 bacterial species, considerable research has shown that Porphyromonas gingivalis (PG), a Gramnegative anaerobic bacterium, is the major etiologic agent which contributes to chronic periodontitis. This black-pigmented bacterium produces a myriad of virulence factors that cause destruction to periodontal tissues either directly or indirectly by modulating the host inflammatory response (How et al., "Porphyromonas gingivalis: an overview of periodontopathic pathogen below the gum line", Frontiers in Microbiology, 2016, vol. 7: p. 53.). PG can colonize in the subgingival pocket and be incorporated into a subgingival biofilm that initiates periodontitis.

[0006] Brief Description

[0007] There is provided, in accordance with an embodiment of the invention, a composition comprising a mixture of CBDA, CBC, and at least one of CBDV and BCP, which combination is substantially free of THC, THCA and THCV.

[0008] In some embodiments, the composition is substantially free of CBG. In some embodiments, the composition is substantially free of CBD. In some embodiments, the composition is substantially free of cannibinoids other than CBDA, CBC, and CBDV.

[0009] In some embodiments, the composition comprises a mixture of CBDA, CBC, and CBDV. In some embodiments, the ratio by weight of each one of CBDA, CBC, and CBDV to the others of CBDA, CBC, and CBDV is not more than 10:1. In some embodiments, the ratio by weight of each one of CBDA, CBC, and CBDV to the others of CBDA, CBC, and CBDV is in the range of 1:3 to 3:1. In some embodiments, the ratio by weight of each one of CBDA, CBC, and CBDV to the others of CBDA, CBC, and CBDV is in the range of 1.1:1 to 1:1.1. In some embodiments, the ratio by weight of each one of CBDA, CBC, and CBDV to the others of CBDA, CBC, and CBDV is 1:1. In some embodiments, the composition is substantially free of BCP. In some embodiments, the composition also comprises BCP. In some embodiments, the total concentration of CBDA, CBC and CBDV taken together in the composition is at least 0.5 microgram / ml composition. In some embodiments, the total concentration of CBDA, CBC and CBDV taken together in the composition is at least 3.75 microgram / ml composition. In some embodiments, the total concentration of CBDA, CBC and CBDV taken together in the composition is at least 0.5 microgram / g composition. In some embodiments, the total concentration of CBDA, CBC and CBDV taken together in the composition is at least 3.75 microgram / g composition.

[0010] In some embodiments, the composition comprises a mixture of CBDA, CBC, and BCP. In some embodiments, the ratio by weight of each one of CBDA, CBC, and BCP to the others of CBDA, CBC, and BCP is not more than 10:1. In some embodiments, the ratio by weight of each one of CBDA, CBC, and BCP to the others of CBDA, CBC, and BCP is in the range of 1:3 to 3:1. In some embodiments, the ratio by weight of each one of CBDA, CBC, and BCP to the others of CBDA, CBC, and BCP is in the range of 1.1:1 to 1:1.1. In some embodiments, the ratio by weight of each one of CBDA, CBC, and BCP to the others of CBDA, CBC, and BCP is 1:1. In some embodiments, the composition also comprises CBDV. In some embodiments, the total concentration of CBDA, CBC and BCP taken together in the composition is at least 0.5 microgram / ml composition. In some embodiments, the total concentration of CBDA, CBC and BCP taken together in the composition is at least 3.75 microgram / ml composition. In some embodiments, total concentration of CBDA, CBC and BCP taken together in the composition is at least 0.5 microgram / g composition. In some embodiments, the total concentration of CBDA, CBC and BCP taken together in the composition is at least 3.75 microgram / mg composition.

[0011] In some embodiments, the composition also comprises at least one pharmaceutically acceptable carrier or excipient. In some embodiments, the composition is in the form of a tablet or caplet. In some embodiments, the composition is in the form of a powder. In some embodiments, the composition is in the form of a liquid. In some embodiments, the liquid is formulated as an oral rinse. In some embodiments, the liquid is formulated for aerosolization or nebulization for use for oral or nasal inhalation. In some embodiments, the composition is in the form of a paste. In some embodiments, the composition is in the form of a lozenge. In some embodiments, the composition is in the form of a gelcap. In some embodiments, the composition is contained in a capsule.

[0012] There is also provided, in accordance with an embodiment of the invention, a method for controlling the growth of Porphyromonas gingivalis (PG), comprising contacting PG with a composition as described hereinabove or hereinbelow, in an amount and for a time efficacious to control such growth. In some embodiments, the PG is in a biofilm. In some embodiments, the PG is not in a biofilm.

[0013] There is also provided, in accordance with an embodiment of the invention, a method of reducing the likelihood of infection by Porphyromonas gingivalis (PG) in the oral cavity, or of reducing the severity of a PG infection in the oral cavity, comprising rinsing the oral cavity with a composition in liquid form as described hereinabove or hereinbelow.

[0014] There is also provided, in accordance with an embodiment of the invention, a composition as described and / or claimed herein for use as a medicine for controlling the growth of Porphyromonas gingivalis (PG) or reducing the likelihood of infection by PG.

[0015] Definitions:

[0016] BCP: (-)-beta-caryophyllene, CAS registry number 118-65-0

[0017] CBD: (-)-trans-Cannabidiol, CAS registry number 13956-29-1

[0018] CBDA: cannabidiolic acid, CAS registry number 1244-58-2

[0019] CBG: Cannabigerol, CAS registry number 25654-31-3

[0020] CBC: Cannabichromene, CAS registry number 20675-51-8

[0021] THC: (-)-trans-A9-Tetrahydrocannabinol, CAS registry number 1972-08-3

[0022] THCA: Tetra hydrocannabinolic acid (with the -COOH group at either the 2- or the 4-position, viz. ortho or para to the hydroxy group), CAS registry number 23978-85-0 or 23978-84-9 THCV: Tetrahydrocannabivarin, CAS registry number 31262-37-0

[0023] CBDV: Cannabidivarin, CAS registry number 24274-48-4

[0024] Pharmaceutical Compositions

[0025] Compositions in accordance with embodiments of the invention include pharmaceutical compositions, which can be administered to a patient to reduce the likelihood of infection by Porphyromonas gingivalis (PG) in the oral cavity, or to reduce the severity of a PG infection in the oral cavity. Such pharmaceutical compositions will thus comprise, in addition to CBDA, CBC, and at least one of CBDV and BCP, one or more pharmaceutically acceptable carriers or excipients. The carrier(s) or excipients must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof.

[0026] It is expected that pharmaceutical compositions in accordance with embodiments of the invention will commonly be formulated for administration in a patient's mouth, for example in liquid form for use as a rinse (mouthwash or gargle) or spray, or as a paste or cream that can be placed topically in the mouth, such as on the cheek, or under the tongue, or on the gums. The ability to place a topical formulation directly on the gums is expected to allow placement of the composition directly on the site of a PG infection. It is contemplated that a rinse may be utilized as needed, and that formulations for topical oral use may be applied multiple times per day, but sustained release oral topical formulations, which will require application once-daily or less, are also contemplated.

[0027] Pharmaceutical formulations are discussed more extensively, inter alia, in Remington: The Science and Practice of Pharmacy, 23rdEdition, Academic Press an imprint of Elsevier, 2021, and pages 359-380, 381-393, and 633-643 therein are incorporated herein by reference.

[0028] Pharmaceutical compositions in accordance with embodiments of the invention may also be formulated as suitable for oral, parenteral (including subcutaneous, intradermal, intramuscular, intravenous and intraarticular), rectal and topical (including dermal, buccal, sublingual and intraocular) administration. The most suitable route may depend upon the condition of the recipient and the particular disorder from which the recipient suffers. The formulations may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy. All methods include the step of bringing into association with CBDA, CBC, and at least one of CBDV and BCP ("active ingredients") the carrier which constitutes one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association the active ingredients with liquid carriers or finely divided solid carriers or both and then, if necessary, shaping the product into the desired formulation.

[0029] Formulations suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active ingredients; as a powder or as granules; as a solution or a suspension in an aqueous liquid or a non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion. The active ingredients may also be presented as a bolus, electuary or paste.

[0030] A tablet may be made by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder, lubricant, inert diluent, lubricating, surface active or dispersing agent. Molded tablets may be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide sustained, delayed or controlled release of the active ingredient therein.

[0031] Formulations for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient. Formulations for parenteral administration also include aqueous and non-aqueous sterile suspensions, which may include suspending agents and thickening agents. The formulations may be presented in unit-dose of multidose containers, for example sealed ampoules and vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of a sterile liquid carrier, for example saline, phosphate-buffered saline (PBS) or the like, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.

[0032] Formulations for rectal administration may be presented as a suppository with the usual carriers such as cocoa butter or polyethylene glycol.

[0033] Formulations for topical administration in the mouth, for example buccally or sublingually, include lozenges comprising the active ingredient in a flavoured basis such as sucrose and acacia or tragacanth, and pastilles comprising the active ingredients in a basis such as gelatin and glycerin or sucrose and acacia.

[0034] Preferred unit dosage formulations are those containing an effective dose, as hereinbelow recited, or an appropriate fraction thereof, of the active ingredient.

[0035] It should be understood that in addition to the ingredients particularly mentioned above, the formulations of this invention may include other agents conventional in the art having regard to the type of formulation in question, for example those suitable for oral administration may include flavoring agents. Liquid formulations for use as a mouthwash may also contain, for example, other anti-bacterial agents (such as cetyl pyridinium chloride, chlorhexidine, saline, menthol, thymol, clove, eucalyptus oil, cinnamon oil, methyl salicylate), humectants (such as glycerin, sorbitol, and polyethylene glycols), surfactants (such as sodium lauryl sulphate), fluoride, alcohol, coloring agents, and / or sweetening agents.

[0036] Experimental

[0037] The efficacy of the compositions disclosed herein against growth of Porphyromonas gingivalis (PG) and biofilm formation by PG were demonstrated as follows.

[0038] Methods

[0039] Inhibition of bacterial growth: Dilutions of compounds in Wilkins broth were prepared in a polystyrene flat bottomed 96-well microplate. Wells with no compounds and with bacteria served as positive controls. Wells with no bacteria and with compounds served as blanks. An equal volume (100 pl) of the bacterial suspension at optical density (OD) at 600 nm=0.1 was added to each well. After a 24-h incubation at 37°C under anaerobic conditions, growth was monitored by recording the OD at 600 nm using a Tecan plate reader Spectrophotometer. Each assay was performed in triplicate. The minimum inhibitory concentration (MIC) was defined as the lowest compound's concentration that resulted in at least 90% (MIC90) inhibition of the bacterial growth compared with that in the untreated controls. Inhibition of biofilm formation: each assay was performed as described above for inhibition of bacterial growth. After incubation for 24 h, spent media and free-floating bacteria were removed by aspiration and the wells were washed carefully three times with doubly distilled water (DDW), prior to quantification of biofilm by crystal violet staining. The minimum biofilm inhibition concentration (MBIC) was defined as the lowest compound's concentration that resulted in at least 50% (MBIC50) or 90% (MBIC90) inhibition of the formation of biofilms compared with that in the untreated controls.

[0040] Agents tested individually were the cannabinoids CBC, CBDA, CBDV, and the terpenoid BCP. Different combinations of three agents were also tested on PG growth and biofilm formation as described above. In the combinations, each agent was present in the same concentration; the numbers shown in Tables 1, 2 and 3 for the combinations indicate the combined concentration of the agents.

[0041] Crystal violet test: 0.02% crystal violet was added to wells and left in each well for 45 min, which then was washed twice with DDW to remove unbound dye. After adding 200 pl of 30% acetic acid into each well, the plate was shaken for 10 min to release the dye, and the biofilm was quantified by measuring the absorbance at 595 nm using a Tecan plate reader Spectrophotometer. Each assay was performed in triplicate.

[0042] Results

[0043] Table 1 shows the results expressed in pl / ml for inhibition of bacterial growth (MIC90) for CBDA, CBC, CBDV and BCP individually, as well as for the combinations of CBDA+CBC+CBDV and CBDA+CBC+BCP. Table 2 shows the results expressed in pl / ml for inhibition of biofilm growth (MBIC90) for these same individual compounds and combinations. Table 3 shows the results expressed in pl / ml for inhibition of biofilm growth (MBIC50) for these same individual compounds and combinations. It will be noted that in these combinations, no other cannabinoids or terpenoids (e.g. sesquiterpenes, diterpenes, triterpenes) were present.

[0044] TABLE 1

[0045] TABLE 2 TABLE 3

[0046] The combinations of CBDA+CBC+CBDV and CBDA+CBC+BCP thus show synergistic effects in the control of both bacterial growth and biofilm formation. Without wishing to be bound by theory, it appears that since inhibitory growth activity was similar to anti-biofilm effect, these combinations of compounds prevent biofilm formation due to their bacteriostatic effect. The combinations may thus be suitable for controlling PG growth and associated infections, such as gingivitis and periodontitis, with reduced risk of adverse effects as well as lower cost as compared to use of each agent alone.

Claims

CLAIMS1. A composition comprising a mixture of CBDA, CBC, and at least one of CBDV and BCP, which combination is substantially free of THC, THCA and THCV.

2. The composition of claim 1, wherein the composition is substantially free of CBG.

3. The combination of claim 1 or claim 2, wherein the composition is substantially free of CBD.

4. The composition of any one of claims 1 to 3, wherein the composition is substantially free of cannibinoids other than CBDA, CBC, and CBDV.

5. The composition of any one of claims 1 to 4, wherein the composition comprises a mixture of CBDA, CBC, and CBDV.

6. The composition of claim 5, wherein the ratio by weight of each one of CBDA, CBC, and CBDV to the others of CBDA, CBC, and CBDV is not more than 10:1.

7. The composition of claim 6, wherein the ratio by weight of each one of CBDA, CBC, and CBDV to the others of CBDA, CBC, and CBDV is in the range of 1:3 to 3:1.

8. The composition of claim 7, wherein the ratio by weight of each one of CBDA, CBC, and CBDV to the others of CBDA, CBC, and CBDV is in the range of 1.1:1 to 1:1.1.

9. The composition of claim 8, wherein the ratio by weight of each one of CBDA, CBC, and CBDV to the others of CBDA, CBC, and CBDV is 1:1.9.

1. The composition of any one of claims 1 to 9, wherein the composition is substantially free of BCP.

10. The composition of any one of claims 5-9, which also comprises BCP.

11. The composition of any one of claims 5-10, wherein the total concentration of CBDA, CBC and CBDV taken together in the composition is at least 0.5 microgram / ml composition.

12. The composition of claim 11, wherein the total concentration of CBDA, CBC and CBDV taken together in the composition is at least 3.75 microgram / ml composition.

13. The composition of any one of claims 5-10, wherein the total concentration of CBDA, CBC and CBDV taken together in the composition is at least 0.5 microgram / g composition.

14. The composition of claim 13, wherein the total concentration of CBDA, CBC and CBDV taken together in the composition is at least 3.75 microgram / g composition.

15. The composition of any one of claims 1 to 4, wherein the composition comprises a mixture of CBDA, CBC, and BCP.

16. The composition of claim 15, wherein the ratio by weight of each one of CBDA, CBC, and BCP to the others of CBDA, CBC, and BCP is not more than 10:1.

17. The composition of claim 16, wherein the ratio by weight of each one of CBDA, CBC, and BCP to the others of CBDA, CBC, and BCP is in the range of 1:3 to 3:1.

18. The composition of claim 17, wherein the ratio by weight of each one of CBDA, CBC, and BCP to the others of CBDA, CBC, and BCP is in the range of 1.1:1 to 1:1.1.

19. The composition of claim 18, wherein the ratio by weight of each one of CBDA, CBC, and BCP to the others of CBDA, CBC, and BCP is 1:1.

20. The composition of any one of claims 15-19, which also comprises CBDV.

21. The composition of any one of claims 15-20, wherein the total concentration of CBDA, CBC and BCP taken together in the composition is at least 0.5 microgram / ml composition.

22. The composition of claim 21, wherein the total concentration of CBDA, CBC and BCP taken together in the composition is at least 3.75 microgram / ml composition.

23. The composition of any one of claims 15-18, wherein the total concentration of CBDA, CBC and BCP taken together in the composition is at least 0.5 microgram / g composition.

24. The composition of claim 23, wherein the total concentration of CBDA, CBC and BCP taken together in the composition is at least 3.75 microgram / mg composition.

25. The composition of any one of claims 1-24, which also comprises at least one pharmaceutically acceptable carrier or excipient.

26. The composition of claim 25 which is in the form of a tablet or caplet.

27. The composition of claim 25 which is in the form of a powder.

28. The composition of claim 25 which is in the form of a liquid.

29. The composition of claim 25 which is in the form of a paste.

30. The composition of claim 25 which is in the form of a lozenge.

31. The composition of claim 25 which is in the form of a gelcap.

32. The composition of claim 25 which is contained in a capsule.

33. The composition of claim 28, wherein the liquid is formulated as an oral rinse.

34. The composition of claim 28, wherein the liquid is formulated for aerosolization or nebulization for use for oral or nasal inhalation.

35. A method for controlling the growth of Porphyromonas gingivalis (PG), comprising contacting PG with a composition according to any one of claims 1-24 in an amount and for a time efficacious to control such growth.

36. The method of claim 35, wherein the PG is in a biofilm.

37. The method of claim 35, wherein the PG is not in a biofilm.

38. A method of reducing the likelihood of infection by Porphyromonas gingivalis (PG) in the oral cavity, or of reducing the severity of a PG infection in the oral cavity, comprising rinsing the oral cavity with a composition according to claim 28 or 33.

40. A composition according to any one of claims 1-34 for use as a medicine for controlling the growth of Porphyromonas gingivalis (PG) or reducing the likelihood of infection by PG.

Images