TREATMENT OF SEBACEOUS GLAND DISORDERS

Abstract

The present invention provides a topical formulation for use in a method of treating or preventing a sebaceous gland disorder selected from acne, seborrhea, rosacea, perioral dermatitis, corticosteroid-induced acneiform lesions and oily skin and/or for use in a method of inhibiting growth of riopionibacterium acnes, said method comprising topically administrating a formulation comprising (i) cannabidiol and (ii) triterpenes selected from asiaticoside, madecassoside, asiatic acid, madecassic acid and combinations thereof.

The invention further relates to a topical formulation comprising: cannabidiol; triterpenes selected from asiaticoside, madecassoside, asiatic acid, madecassic acid and combinations thereof; flavonolignans selected silibinin, isosilibinin, silicristin, silidianin and combinations thereof, the concentrations of these flavonolignans being calculated as silibinin.

Claims

1. A method of treating or preventing a sebaceous gland disorder selected from acne, seborrhea, rosacea, perioral dermatitis, corticosteroid-induced acneiform lesions and oily skin and/or of inhibiting of growth of Priopionibacterium acnes, the method comprising topically administrating a formulation comprising: (i) cannabidiol; and (ii) triterpenes selected from asiaticoside, madecassoside, asiatic acid, madecassic acid and combinations thereof.

2. The method according to claim 1, wherein the formulation comprises 0.1-3 wt. % cannabidiol.

3. The method according to claim 1, wherein the formulation comprises 0.025-3 wt. % of the triterpenes.

4. The method according to claim 1, wherein the formulation comprises cannabidiol and the triterpenes in a weight ratio of 10:1 to 1:10, respectively.

5. The method according to claim 1, wherein the formulation comprises an extract of Centella asiatica.

6. The method according to claim 1, wherein the formulation further comprises 0.05-2 wt. % of flavonolignans selected from silibinin, isosilibinin, silicristin, silidianin and combinations thereof, the concentrations of these flavonolignans being calculated as silibinin.

7. The method according to claim 6, wherein the formulation comprises cannabidiol and the flavonolignans in a weight ratio of 1:5 to 5:1.

8. The method according to claim 6, wherein the formulation comprises an extract of S. marianum.

9. The method according to claim 1, wherein the sebaceous gland disorder is acne.

10. The method according to claim 1, the method being a method of inhibiting of growth of Priopionibacterium acnes.

11. A topical formulation, comprising: (a) cannabidiol; (b) triterpenes selected from asiaticoside, madecassoside, asiatic acid, madecassic acid and combinations thereof; (c) flavonolignans selected silibinin, isosilibinin, silicristin, silidianin and combinations thereof, the concentrations of these flavonolignans being calculated as silibinin.

12. The topical formulation according to claim 11, wherein the formulation comprises 0.1-3 wt. % cannabidiol.

13. The topical formulation according to claim 11, the formulation comprises 0.025-3 wt. % of the triterpenes.

14. The topical formulation according to claim 11, wherein the formulation comprises 0.05-2 wt. % of the flavonolignans.

15. The topical formulation according to claim 11, wherein the formulation comprises cannabidiol and the triterpenes in a weight ratio of 10:1 to 1:2, respectively.

16. The topical formulation according to claim 11, wherein the formulation comprises cannabidiol and the flavonolignans in a weight ratio of 1:5 to 5:1, respectively.

17. The topical formulation according to claim 11, wherein the formulation comprises an extract of Centella asiatica.

18. The topical formulation according to claim 11, wherein the formulation comprises an extract of Silybum marianum.

19. A method of preparing topical formulation according to claim 11, the method comprising combining an extract of Cannabis, an extract of Centella asiatica and an extract of Silybum marianum.

Description

DETAILED DESCRIPTON OF THE INVENTION

[0029] A first aspect of the invention relates to a topical formulation for use in a method of treating or preventing a sebaceous gland disorder selected from acne, seborrhea, rosacea, perioral dermatitis, corticosteroid-induced acneiform lesions and oily skin or for use in a method of inhibiting growth of Priopionibacterium acnes, said method comprising topically administrating a formulation comprising (i) cannabidiol and (ii) triterpenes selected from asiaticoside, madecassoside, asiatic acid, madecassic acid and combinations thereof.

[0030] The present treatment is particularly suitable for treating acne vulgaris, including adolescence acne and adult acne.

[0031] The method present treatment preferably comprises topical administration of the formulation to the skin of a human subject. According to a particularly preferred embodiment, the formulation the treatment comprises topical administration to the face, the upper part of the chest or the back of a human subject.

[0032] The formulation that is employed in the treatment according to the present invention preferably contains 0.1-3 wt. % cannabidiol, more preferably 0.3-2 wt. % cannibidiol and most preferably 0.5-1 wt. % cannabidiol.

[0033] The triterpenes selected from asiaticoside, madecassoside, asiatic acid, madecassic acid and combinations thereof are preferably present in the formulation in a concentration of 0.025-3 wt. %, more preferably in a concentration of 0.05-2 wt. % and most preferably in a concentration of 0.1-1 wt. %.

[0034] In a preferred embodiment, the formulation contains the triterpene asiaticoside. More preferably, the formulation contains 0.01-1.5 wt. % asiaticoside, even more preferably 0.02-0.6 wt. % asiaticoside and most preferably 0.03-0.45 wt. % asiaticoside.

[0035] The triterpenic genins selected from asiatic acid, madecassic acid and combinations thereof are preferably present in the formulation in a concentration of 0.01-2 wt. %, more preferably in a concentration of 0.03-1.5 wt. % and most preferably in a concentration of 0.05-1 wt. %.

[0036] In a preferred embodiment, the formulation of the present invention contains asiaticoside and triterpenic genins selected from asiatic acid, madecassic acid and combinations thereof in a weight ratio asiaticoside : triterpenic genins of 1:4 to 4:1, more preferably in a weight ratio asiaticoside:triterpenic genins of 1:1 to 1:2

[0037] Cannabidiol and the triterpenes are typically present in the formulation in a weight ratio of 10:1 to 1:10, more preferably in a weight ratio of 6:1. to 1:4, most preferably in a weight ratio of 4:1 to 1:1.

[0038] The effectiveness of the present formulation against sebaceous gland disorders may be further enhanced by the additional inclusion of flavonolignans selected from silibinin, isosilibinin, silicristin, silidianin and combinations thereof. Preferably, the formulation contains 0.05-2 wt. %, more preferably 0.05-1 wt. % and most preferably 0.25-1 wt. % of flavonolignans selected silibinin, isosilibinin, silicristin, silidianin and combinations thereof, the concentrations of these flavonolignans being calculated as silibinin.

[0039] Silibinin is preferably contained in the formulation in a concentration of 0.01-1.5 wt. %, more preferably of 0.03-1.2 wt. % and most preferably of 0.1-1 wt. %.

[0040] In another preferred embodiment, the formulation contains 0.001-1.5 wt. %, more preferably 0.01-1 wt. % and most preferably 0.05-0.5 wt. % of silicristin and/or silidianin, the concentrations of these flavonolignans being calculated as silibinin.

[0041] Cannabidiol and the aforementioned flavonolignans (concentration of the flavonolignans being calculated as silibinin) are preferably present in a weight ratio of 1:6 to 6:1, more preferably in a weight ratio of 1:4 to 4:1.

[0042] In accordance with another preferred embodiment, the formulation additionally contains hemp seed oil. More preferably, the formulation contains 0.01-5 wt. %, more preferably 0.03-3 wt. %, most preferably 0.1-1 wt. % of hemp seed oil.

[0043] Hemp seed oil is obtained by pressing hemp seeds. Hemp seed oil is manufactured from varieties of Cannabis sativa that do not contain significant amounts of tetrahydrocannabinol. Typically around 50% of the weight of hempseed is an edible oil that contains omega-6 fatty acids including linoleic acid (appr. 54%) and gamma-linolenic acid (appr. 3%), as well as the omega-3 fatty acid alpha-linolenic acid (appr. 17%) in addition to monounsaturated fatty acids and stearidonic acid (appr. 2%). Hemp seed oil typically contains 5% to 7% saturated fatty acids.

[0044] The formulation of the present invention typically comprises 5-90 wt. % water. More preferably, the water content of the formulation is in the range of 10 to 85 wt. %, most preferably in the range of 20 to 80 wt. %.

[0045] The formulation of the present invention can be provided in different forms. Preferably, the formulation is a gel, a cream, a lotion, a soap or a spot treatment product.

[0046] Another aspect of the invention relates to a topical formulation comprising: [0047] cannabidiol; [0048] triterpenes selected from asiaticoside, madecassoside, asiatic acid, madecassic acid and combinations thereof; [0049] flavonolignans selected silibinin, isosilibinin, silicristin, silidianin and combinations thereof, the concentrations of these flavonolignans being calculated as silibinin.

[0050] Advantageous embodiments of this formulation have already been described above.

[0051] Yet another aspect of the invention relates to a method of preparing topical formulation as described herein before, said method comprising combining an extract of Cannabis, an extract of C. asiatica and an extract of S. marianum.

[0052] The extract of Cannabis that is employed in this method preferably contains at least 20%, more preferably at least 30% and most preferably at least 40% cannabidiol by weight of dry matter.

[0053] The extract of C. asiatica used in the method preferably contains at least 20%, more preferably at least 30% and most preferably at least 40% of the triterpenes (asiaticoside, madecassoside, asiatic acid, madecassic acid and combinations thereof) by weight of dry matter.

[0054] The extract of S. marianum that is used in the method preferably contains at least 20%, more preferably at least 30% and most preferably at least 40% of the flavonolignans (silibinin, isosilibinin, silicristin, silidianin and combinations thereof) by weight of dry matter.

[0055] The invention is further illustrated by the following non-limiting examples.

EXAMPLES

Example 1

[0056] The impact of cannabidiol, extract of Centella asiatica (leaf) and combinations of these components on the growth of P. acnes and on the production of cytokines IL-1β and TNFα by a U937 cell line were investigated.

[0057] The specifications of the aforementioned components are provided in Table 1.

TABLE-US-00001 TABLE 1 Supplier Characteristics Cannabidiol Echo Pharmaceuticals BV, Extracted Cannabidiol (98%) the Netherlands C. asiatica Indena S.p.a., Italy Powder (bulk density 0.40 g/ml) extract Product code 3022060 80 wt. % ≤ 50 μm Extraction solvent: ethanol Asiaticoside: 36.0-44.0% (HPLC assay) Sum of Asiatic acid and Madecassic aid: 56.0-64.0% (HPLC assay) Proteins: 1.4 wt. % Fat: 0.4 wt. % Moisture 1.0 wt. %

[0058] Propionibacterium acnes Growth Inhibition

[0059] A streak plate isolation technique was performed to isolate a single homogenous colony of P. acnes. A pinch of one colony was inoculated from agar plate into U-shape falcon tube containing 4 ml of broth media. Cultures were incubated at 37° C. in reduced oxygen-conditions for 72 hours. The cultures were then diluted and grown to mid-log phase (O.D. 0.5; 600 nm) for 3 hr.

[0060] The bacteria were incubated with the test components (cannabidiol and/or extract of C. asiatica) at different concentrations in triplicate at a final volume of 200 μl in an N-pre-flushed U shape 96-well plates for 3 hr. An additional blank control group was included (media w/o bacteria). Also, a negative (bacteria without test samples) control groups was included in the assay. In addition, combinations of the these components were tested (see Table 2).

[0061] The absorbance was recorded at 600 nm. The absorbance of the blank control was subtracted from all measurements. The change in bacterial number in response to increasing test sample concentrations was plotted.

[0062] The results show (see Table 2) that: [0063] neither cannabidiol nor extract of C. asiatica significantly inhibits growth of P. acnes [0064] the combination of cannabidiol and extract of C. asiatica has a significant inhibitory effect on growth of P. acnes

[0065] Inhibition of Secretion of IL-1β and TNFα

[0066] U937 cells, were thawed and treated according to the manufacturer's instructions. The cultures were incubated for recovery at 37° C. with 5% CO.sub.2 until 70-80% confluency was reached (visual estimation). Then, approx. 0.3×10.sup.6 ¢/mL (determined by counting) were seeded in 96 well plates containing 200 μL/well of complete growth medium. The cells were incubated at 37° C. with 5% CO.sub.2 until 60% confluency was reached (visual estimation, typically—after 48 hr).

[0067] After the cells had reached the required confluency, the medium was replaced with pre-prepared growth medium supplemented with 1 μg/mL of lipopolysaccharides, and the test components were added, at a final volume of 200 μL/well. The following control groups were also added: Naïve cells, vehicle control, stimulation control, stimulation vehicle control. A proper blank control was subtracted from the measurements.

[0068] The cells were incubated for 24 hr. at 37° C. with 5% CO.sub.2. After the incubation, the conditioned medium of the different treatment groups was collected under standardized conditions and centrifuged at 250×g for 5 min to remove particulates. Clear supernatants were frozen at −70° C. until cytokines analyses. The secretion levels of IL-1β and TNFα (a major inflammatory marker) were quantified by commercial ELISA.

[0069] The results show (see Table 3) that: [0070] extract of C. asiatica does not significantly attenuate secretion of IL-1β and TNFα [0071] cannabidiol (CBD) attenuates secretion of IL-1β and TNFα [0072] extract of C. asiatica enhances the inhibitory effect of cannabidiol on the secretion of IL-1β and TNFα

[0073] Results

[0074] The results of the aforementioned investigations are summarised in Tables 2 and 3.

TABLE-US-00002 TABLE 2 μg/ml Inhibition (in %) Sample Cannabidiol C. asiatica P. acnes 1 1.25 0 −10.9% 2 2.5 0    5.7% 3 0 0.3125    8.3% 4 0 0.625  −1.4% 5 0 1.25    0.3% 6 1.25 0.3125   51.0% 7 2.5 0.625   45.5%

TABLE-US-00003 TABLE 3 μg/ml Inhibition (in %) Sample Cannabidiol C. asiatica IL-1β TNFα 1 10 0 28.4 48.0 2 0 10 0.6 0.0 3 10 10 45.8 55.2

Example 2

[0075] The impact of cannabidiol, extract of Centella asiatica (leaf), extract of Silybum marianum (fruit without pappus) on the secretion of prostaglandin E2 (PGE.sub.2) was investigated.

[0076] The cannabidiol and C. asiatica extract used were the same as in Example 1. The specification of the S. marianum extract is shown in Table 4.

TABLE-US-00004 TABLE 4 Supplier Characteristics S. marianum Indena S.p.a., Italy Powder (bulk density 0.44 g/ml) extract Product code 9065110 91 wt. % ≤ 50 μm Extraction solvent: ethyl acetate Flavonoids (as monohydrate Silibinin) ≥ 80.0% (UV assay) Silibinin and Isosilibinin ≥ 30.0% (HPLC assay) Silymarin (as Silibinin): 50.0-60.0% (HPLC assay) Silicristin and Silidianin (as Silibinin), on the total content of Silymarin: 20.0-45.0% (HPLC assay) Silibinin A and Silibinin B (as Silibinin), on the total content of Silymarin: 40.0-65.0% (HPLC assay) Isosilibinin A and Isosilibinin B (as Silibinin), on the total content of Silymarin: 10.0- 20.0% (HPLC assay) Proteins: 1.7 wt. % Fat: 2.3 wt. % Dietary fibre: 3.5 wt. % Moisture 1.4 wt. %

[0077] Secretion of PGE.sub.2

[0078] RAW 264.7 cells (approx. 2.5×10.sup.5 ¢/ml, by counting) were seeded in 96 well plates containing 170 μl/well of complete growth medium. The cells were incubated at 37° C. with 5% CO.sub.2 for 24 hr. Then, the medium was aspirated and replaced by LPS-containing medium (12.5 ng/ml) without or with the test components. In addition, naïve cells, vehicle-treated cells, Stimulated Control, and Stimulated Vehicle Control served as negative controls. Dexamethasone served as a positive control for anti-inflammatory assay. A Blank control group was included in the assay.

[0079] The spent media from all test groups were collected under standardized conditions and centrifuged at 250 g for 5 min to remove particulates. Clear supernatants were frozen at −70° C. until analysis. The secretion levels of PGE.sub.2 (an important inflammatory marker) were determined.

[0080] The results show (see Table 5) that: [0081] extract of C. asiatica does not significantly attenuate secretion of PGE.sub.2 [0082] cannabidiol (CBD) attenuates secretion of PGE.sub.2 at a concentration of 2.5 μg/ml, but not at 1.25 μg/ml [0083] extract of C. asiatica enhances the inhibitory effect of CBD on the secretion of PGE.sub.2 [0084] extract of S. marianum does not significant attenuate secretion of PGE.sub.2 [0085] extract of S. marianum enhances the inhibitory effect of the combination of CBD and extract of C. asiatica on the secretion of PGE.sub.2

[0086] Results

[0087] The results of the aforementioned investigation are summarised in Table 5.

TABLE-US-00005 TABLE 5 μg/ml Inhibition (in %) Sample Cannabidiol C. asiatica S. marianum PGE.sub.2 1 1.25 0 0 1.9 2 2.5 0 0 24 3 0 1.25 0 −2.3 4 0 2.5 0 2.4 5 0 0 2.5 5.0 6 1.25 1.25 0 29 7 2.5 1.25 0 34 8 2.5 2.5 0 40 9 2.5 2.5 2.5 46