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1. WO2021064132 - TRAITEMENT DU SYNDROME DE LA MÉNOPAUSE ET/OU DES SYMPTÔMES ASSOCIÉS À LA MÉNOPAUSE

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[ EN ]

TREATMENT OF MENOPAUSAL SYNDROME AND/OR SYMPTOMS ASSOCIATED WITH MENOPAUSE

Technical Field

The present invention relates to compositions that are useful in the treatment of menopausal female subjects and/or female subjects suffering from menopausal syndrome. Furthermore, the invention relates to methods of treating perimenopausal, menopausal and/or postmenopausal female subjects and/or female subjects suffering from menopausal syndrome using said compositions.

Background of the Invention

Menopause is defined as the permanent cessation of menstrual activity resulting from the loss of follicular function. Menopause marks the transition of a woman from the reproductive to the non-reproductive state. After a woman has gone through menopause she will not ovulate and will not have the associated menstrual bleeding anymore. The main trigger for the transition is the depletion and aging of the finite amount of oocytes. The cessation of follicular activity is characterized by dramatic changes in estrogen and FSH levels.

Menopause is part of the normal aging process. On average it occurs at an age in the range of 49-52 years. Menopause can occur at an earlier age, e.g. as a consequence of (idiopathic) pathology, referred to as ‘pre-mature ovarian failure’, as a consequence of surgical interventions, such as oophorectomy, of medical treatments, such as radiotherapy or chemotherapy, and of certain life-style habits, such as smoking.

During menopause, ovarian estradiol synthesis and secretion come to an end. As a result, total estrogen production is reduced by about sixty percent. The relative insufficiency of estrogen causes atrophy of epithelia and a significant decrease in the supply of estrogen to target organs. As a direct consequence, women in menopause experience symptoms of instability of their hypothalamic thermoregulatory centers, and many suffer from recurrent symptoms (i.e. besides changes in menstruation patterns). Such symptoms are in fact experienced by the large majority of women going through menopause, typically by about 75 % or more. Symptoms can last for the whole menopausal transition and some may persist until (long) thereafter. The most common symptoms include vasomotor symptoms, symptoms associated with vulvar and vaginal

atrophy, symptoms associated with brain-nervous system changes, symptoms resulting from musculoskeletal changes, symptoms caused by urogenital changes and emotional disturbances.

Vasomotor symptoms include hot flushes. Hot flushes are by far the most common symptom; surveys have shown that about 75% of menopausal women experience repeated "hot flush" episodes over an average of 3.8 years. Hot flushes (which are immediately followed by a decrease in core body temperature) are a result of blood flow to skin and sudomotor sweat responses that raise skin temperature to reduce core body temperature. They are typically very sudden and may be accompanied by a rapid heartbeat and blood vessel dilatation. The duration of such symptoms varies, and may typically be from two to thirty minutes on each occasion. The frequency also varies, e.g. symptoms may occur a few times each week or up to a dozen times a day and more. Also the intensity may vary:

mild hot flushes include a sensation of heat without perspiration; moderate hot flushes include a sensation of heat with perspiration, but the subject is able to continue its activities;

severe hot flushes include a sensation of heat with sweating, severe enough to hamper normal activity.

Vaginal and vulvar atrophy are among the (very) common consequences of the decrease in estrogenic levels in menopause. The fall in circulating estrogen levels is associated with thinning of the vaginal walls, with reduction of elasticity and decrease in the natural secretion and lubrication, which makes sexual intercourse difficult and painful. Symptoms associated with vulvar and vaginal atrophy include vaginal pain associated with sexual activity; bleeding associated with sexual activity; dysuria; dryness, irritation, itching, and/or burning in and around the vaginal area; and combinations thereof.

Other symptoms can be experienced by women going through menopause, such as symptoms resulting from musculoskeletal changes, such as myalgia, arthralgia and back pain; symptoms caused by urogenital changes such as urinary frequency or urinary incontinence, and symptoms due to nervous system changes such as hypomnesis, depression, loss of concentration and dizziness. The menopause transition also frequently results in emotional disturbances, which may be the consequence of physical symptoms or may be associated with direct nervous system changes. Frequently reported emotional symptoms include irritability, feelings of

sadness, lack of motivation, anxiety, aggressiveness, difficulty concentrating, fatigue, mood changes and tension. When the menopausal transition is associated with symptomatology this is also often referred to as ‘menopausal syndrome’.

The severity of symptoms associated with menopause range from mild to incapacitating and it has been estimated that 20-40% of women suffer symptoms severe enough to lead to physical or mental incapacitation. The severity of symptoms can vary from month to month or year to year for a particular woman.

Symptoms can persist for some time after menopause, typically up to around 5 years from the last menses. Around 1 in every 10 women experience symptoms for up to 12 years after the last menses.

The medical community is in general agreement that, if estrogen could be administered without risk, all women in the menopausal phase of their life cycle should receive estrogen medication to counteract and abolish the symptoms typically associated with estrogen deficiency. To date, however, there is a general consensus that the administration of estrogen, in particular of synthetic estrogens, is associated with serious health-risks. A number of studies have reported that estrogen replacement therapy may increase the risks of uterine bleeding, stroke, heart attack, breast cancer, and uterine cancer (JAMA. 2013;310(13): 1353—1368, New Engl. J. Med. 1995.

332:1589-1593). Gallbladder disease is also more common among women who take estrogen, as are systemic problems such as hypertension and water retention. For these reasons, only periodic and cautious medication with estrogen is recommended. To reduce the risks associated with long-term estrogen use, it is possible to combine estrogen with a progestin, which provides endometrial protection.

To date, only 35 to 40% of women with menopausal symptoms use estrogen replacement therapy, and even among those women there is reluctance to do so continuously for long period.

One approach to mitigate the risks associated with estrogen supplementation relies on local administration of an estrogen-containing composition, so as to avoid high systemic levels. Local treatment usually involves administration of (controlled-release) intravaginal ointments, gels or tablets. With treatments relying on intravaginal (self) administration, patient compliance is relatively low, because of cultural, personal, and/or hygienic reasons.

In some cases, estrogen supplementation does not lessen menopausal symptoms unless given at very high doses. In such cases androgen supplementation

has been proposed instead of estrogen supplementation. This type of therapy, however, has also been accompanied by the risk of developing significant negative side effects, including endometriosis and hyperplasia.

In view of the drawbacks and negative side effects of conventional estrogen supplementation and estrogen and androgen medication, for many women no treatment is available, for which the therapeutic efficacy would outweigh the risks and disadvantages.

Hence, there is a strong-felt need for effective treatment and management of symptoms associated with menopause, not relying on estrogen administration. Dietary supplements effective in preventing, treating and/or alleviating symptoms of menopause would be a particularly attractive solution. In general, treatments based on dietary supplements, i.e. supplements that are taken orally, typically on a daily basis, are not perceived as burdensome and treatment compliance/adherence is generally good. Another factor to be considered in this regard is the general attitude towards the use of dietary supplements, which is often better if the supplement is perceived as ‘natural’.

It is an objective of the present invention to provide compositions, in particular in the form of dietary supplements, that can be used effectively in the treatment of perimenopausal, menopausal and postmenopausal female subjects. It is a further object of the invention to provide treatments, based on the administration of said compositions.

Summary of the Invention

In accordance with the present invention, it has been found that compositions comprising one or more proanthocyanidins in combination with hyaluronic acid, e.g. in the form of oral (dietary) supplements, can be used effectively in a method of treating perimenopausal, menopausal and/or postmenopausal female subjects, in order to, e.g., alleviate, prevent and/or suppress one or more symptoms associated with menopause and/or to improve the well-being of said subjects in any (other) way, as can be assessed using (e.g.) menopausal specific questionnaires, prospective diaries and similar self-assessment protocols.

Accordingly, the invention provides compositions, especially pharmaceutical and/or nutraceutical (unit dosage) formulations, comprising one or more proanthocyanidins in combination with hyaluronic acid as a pharmacologically and/or physiologically active ingredient combination. The invention also provides kits-of-parts comprising said compositions.

The present invention also provides a method of treating a subject in need thereof, such as a perimenopausal, menopausal or postmenopausal female subject, said method comprising administration of the compositions of the invention, especially the pharmaceutical and/or nutraceutical (unit dosage) formulations, comprising one or more proanthocyanidins in combination with hyaluronic acid.

The invention also provides compositions, especially pharmaceutical and/or nutraceutical (unit dosage) formulations, comprising one or more proanthocyanidins in combination with hyaluronic acid for use in the treatment of subjects in need thereof, such as a perimenopausal, menopausal or postmenopausal female subject.

The invention also provides the use of compositions comprising one or more proanthocyanidins in combination with hyaluronic acid in the manufacture of a pharmaceutical or nutraceutical product for use in a method of treating a subject in need thereof, such as a perimenopausal, menopausal or postmenopausal female subject.

Good results are attained with compositions comprising, besides the combination of proanthocyanidins and hyaluronic acid, one or more further pharmacologically and/or physiologically active ingredients, especially one or more further pharmacologically and/or physiologically active ingredients selected from the group consisting of Nw-methylserotonin, ascorbic acid, Ginkgolides, valerenic acid, apigenin, and magnesium.

Particularly good results are attained when the treatment comprises the (alternate) administration of two distinct compositions, each comprising proanthocyanidins and hyaluronic acid in combination with one or more further pharmacologically and/or physiologically active ingredients, which two distinct compositions are adapted to the female subject’s circadian rhythm. More in particular, in accordance with the invention, a treatment is provided comprising the administration of a first composition to be administered in the morning, which first composition is specifically adapted to aid in the alleviation, prevention and/or suppression of one or more symptoms of menopause, while at the same supporting typical day-time physiological activity levels, and a second composition to be administered in the evening, which second composition is specifically adapted to aid in the alleviation, prevention and/or suppression of one or more symptoms of menopause, while at the same supporting night-rest.

The invention also provides compositions and kits-of-parts suitable and/or intended for use in such methods of treatment.

In particularly preferred embodiments of the invention, pharmacologically and/or physiologically active ingredients are provided in the form of herbal preparations, such as in the form of a plant extract. The use of such extracts from a naturally occurring source is particularly advantageous as it aids in general acceptance and treatment compliance by the subject to be treated.

These and other aspects of the invention will become apparent to those skilled in the art on the basis of the following description.

Detailed Description of the Invention

The compositions

A first aspect of the invention provides compositions comprising hyaluronic acid and one or more proanthocyanidins, such as pharmaceutical formulations and/or food supplement products comprising hyaluronic acid and one or more proanthocyanidins in association with one or more further ingredients, typically one or more further active ingredients and/or one or more excipients.

Hyaluronic acid

Hyaluronic acid, also known as hyaluronan or hyaluronate, is a naturally occurring unsulphated glycosaminogycan composed of repeating disaccharide units of N-acetylglucosamine (GlcNAc) and glucuronic acid (GlcUA) linked together by alternating beta-1 ,4 and beta-1 ,3 glycosidic bonds.

Hyaluronic acid may be obtained by isolation from a natural source or by chemical synthesis. Natural sources of hyaluronic acid include rooster combs and (optionally recombinant) microorganisms. In a preferred embodiment of the invention, the composition comprises hyaluronic acid obtained from a bacterium, preferably a Gram-positive bacterium from the class Bacillus, preferably a bacterium from the genus Streptococcus, most preferably Streptococcus zooepidemicus.

In a preferred embodiment of the invention, the hyaluronic acid has a number average molecular weight of more than 0.01 MDa, preferably of more than 0.1 MDa, preferably of more than 0.5 MDa, preferably of more than 1 MDa and/or a number

average molecular weight of less than 20 MDa, preferably of less than 10 MDa, preferably of less than 5 MDa, preferably of less than 1 .5 MDa. The molecular weight may be determined by any method known in the art, preferably the molecular weight is determined by size-exclusion chromatography as described in Soltes, L, et al. "Molecular characteristics of some commercial high-molecular-weight hyaluronans." Biomedical Chromatography 16.7 (2002): 459-462.

Without wishing to be bound by any theory, the present inventors believe that the molecular weight values recited herein are advantageous as hyaluronic acid characterized by said molecular weight values may be small enough to provide adequate absorption and bioavailability, while still being of sufficient size to provide desirable therapeutic and/or physiological effects.

In accordance with the invention, hyaluronic acid may be provided in the form of a pharmaceutically acceptable salt or hydrate, preferably in the form of an alkali metal salt, most preferably in the form of sodium hyaluronate.

Proanthocyanidins

Proanthocyanidins, also known as procyanidins, are a class of polyphenols which are oligomeric flavan-3-ols and are found in a variety of plants. Proanthocyanidins are polymers consisting of the monomer units catechin and/or epicatechin, which are connected either by 4-8 or 4-6 linkages. Typically, the proanthocyanidins oligomers have a chain length of 2-12 monomer units. A great variety of proanthocyanidins exist.

Proanthocyanidins occur naturally in various plants. In accordance with the invention, the one or more proanthocyanidins may be obtained from a natural source or by chemical synthesis. In a preferred embodiment, the compositions of the invention comprise one or more proanthocyanidins in the form of a herbal preparation, such as an extract.

As used throughout this document, the term “herbal preparation” covers comminuted or powdered plant materials, as well as extracts thereof. Herbal preparations are typically produced by extraction, fractionation, purification, concentration or other ways of physically and/or biologically processing plant materials.

In a preferred embodiment, the compositions of the invention comprise one or more proanthocyanidins in the form of a herbal preparation, preferably an extract, from a plant material selected from the group consisting of grapes (such as grape seeds or grape skin), pine bark, ginkgo leaves, peanuts, cocoa beans, tamarind, tomato, peanut, almond, apple, cranberry, blueberry, tea leaves and combinations thereof, preferably pine bark.

In a particularly preferred embodiment, the compositions of the invention comprise one or more proanthocyanidins in the form of a bark extract from Pinus radiata, Pinus thunbergii, Pinus densiflora, Pinus roxburghii, Pinus Siberian, Pinus brutia, Pinus caribaea, Pinus morrisonicola, Pinus koraiensis, Pinus cembra or Pinus pinaster, most preferably Pinus pinaster. Pinus pinaster and Pinus maritima are generally understood to refer to the same species commonly called “French Maritime Pine”.

In accordance with a preferred embodiment of the invention, the composition comprises one or more proanthocyanidins in the form of an extract as described herein, having a proanthocyanidin content of more than 40 wt.%, based on the dry weight of the proanthocyanidin-containing plant extract, i.e. the plant derived components after removal of the extraction solvents, as determined by gel-permeation chromatography, preferably more than 60 wt.%, more preferably more than 65 wt.%.

In accordance with a preferred embodiment of the invention, the composition comprises one or more proanthocyanidins in the form of an extract as described herein before having a total polyphenol content of more than 80 wt.% based on the dry weight of the proanthocyanidin-containing plant extract, i.e. the plant derived components after removal of the extraction solvents, as determined by gel-permeation chromatography, preferably more than 90 wt.%, more preferably more than 95 wt.%.

The extracts as described herein may typically comprise one, two, three or four additional active ingredients selected from the group consisting of monomeric phenols, cinnamic acid, arabinogalactan sulphate and taxifolin.

Suitable extracts can be produced with polar solvents, such as with water, ethyl acetate, ethanol and mixtures thereof. In preferred embodiments the extract is produced with an extraction solvent having a relative polarity of more than 0.2, such as more than 0.3 or more than 0.5. The meaning of relative polarity values is commonly understood by those skilled in the art. Preferably, the relative polarity is determined as a normalized value from measurements of solvents shifts of adsorption spectra, wherein water has a relative polarity value of 1 , as explained in Christian Reichardt, Solvents and Solvent Effects in Organic Chemistry, Wiley-VCH Publishers, 3rd ed., 2003. Suitable extracts can be produced with water, with alcohols, in particular

methanol, ethanol, propanol or mixtures thereof. In preferred embodiments, the extraction solvent is a mixture of water and alcohol, preferably a mixture with a level of alcohol within the range of 10-95 %(v/v), 30-90 %(v/v) or 60-80 %(v/v). In preferred embodiments the extract is produced with a water-ethanol mixture. In more preferred embodiments, the extract is produced with a water-ethanol mixture.

In other embodiments of the invention, the extracts are produced with ethyl acetate or water ethyl acetate mixtures, preferably mixtures with a level of ethyl acetate within the range of 10-95 %(v/v), 30-90 %(v/v) or 60-80 %(v/v) as the extraction solvent.

In other, highly preferred, embodiments of the invention, the extract is produced with water as the sole extraction solvent. Hence, in accordance with a preferred embodiment of the invention, the composition comprises one or more proanthocyanidins in the form of an aqueous extract, preferably an extract produced with water as the sole extraction solvent (referred to herein as a ‘100% water extract’), of a proanthocyanidin comprising plant material as described above. In an embodiment of the invention, a composition as defined herein is provided comprising an aqueous extract, preferably a 100% water extract of a proanthocyanidin comprising plant material as described above. In accordance with a preferred embodiment of the invention, the extraction with water as the sole extraction solvent is performed at a temperature of more than 60°C, preferably more than 80°C, most preferably more than 95°C.

As is known to the skilled person, a crude extract may further be purified employing solvents other than the solvent used for producing the crude extract, in order to separate (extract) one or more components of interest from one or more unwanted components. Such additional steps of extracting components from a crude extract is referred to herein as fractionation or partitioning of the original crude extract. In certain preferred embodiments of the invention, the extracts described herein before have been subjected to one or more additional extraction steps, employing one or more solvents, such as an alcohol or ethyl acetate, preferably ethyl acetate, which is different from the solvent used to produce the crude extract,.

In an embodiment of the invention, a composition as defined herein is provided comprising one or more proanthocyanidins in the form of an extract which is the alcohol or ethyl acetate fraction, preferably the ethyl acetate fraction obtained by partitioning an aqueous extract, preferably a 100% water extract of the proanthocyanidin comprising plant material.

In a preferred embodiment of the invention, the composition comprises a proanthocyanidin-containing plant extract obtainable by aqueous extraction, preferably a 100% water extraction, performed in such a way that the weight : weight ratio of plant material : extract is within the range of 800:1 to 1200:1 , wherein the amount of extract is determined after substantially all solvent has been removed and before addition of any excipient or carrier.

In a preferred embodiment of the invention, the composition comprises a proanthocyanidin-containing plant extract obtainable by a method comprising the steps of:

a) providing pine bark plant material, preferably Pinus pinaster bark;

b) macerating or grinding the plant material provided in step a);

c) performing an aqueous extraction, preferably a 100% water extraction, preferably at a temperature of more than 95°C;

d) concentrating the extract, preferably by evaporation or freeze-drying, preferably by freeze-drying.

In a preferred embodiment of the invention, the composition comprises a proanthocyanidin-containing plant extract obtainable from step f) of a method comprising the steps of:

a) providing pine bark plant material, preferably Pinus pinaster bark;

b) macerating or grinding the plant material provided in step a);

c) performing an aqueous, preferably a 100% water extraction, preferably at a temperature of more than 95°C;

d) optionally concentrating the extract, preferably by evaporation or freeze-drying, preferably by freeze-drying;

e) partitioning the extract obtained in step c) or step d) using an alcohol or ethyl acetate, preferably ethyl acetate, resulting in a water fraction and an alcohol or ethyl acetate fraction; and

f) concentrating the alcohol or ethyl acetate fraction.

Proanthocyanidin-containing extracts suitable for use in the context of the present invention and production thereof have been described in the prior art, e.g. in: • Cretu, Elena, et al. "In vitro study on the antioxidant activity of a polyphenol-rich extract from Pinus brutia bark and its fractions." Journal of medicinal food 16.11 (2013): 984-991 ;

• Kim, Sang Min, et al. "A comparison of pycnogenol and bark extracts from Pinus thunbergii and Pinus densiflora: Extractability, antioxidant activity and proanthocyanidin composition." Journal of Medicinal Plants Research 6.14 (2012): 2839-2849.

Proanthocyanidin-containing extracts suitable for use in the context of the present invention are available commercially, e.g. from the company “DRT” in France under the reference “Cosmythic® Pine bark extract (Pinus pinaster)”.

In a preferred embodiment of the pharmaceutical composition, the weight to weight ratio of hyaluronic acid to proanthocyanidins is within the range of 10: 1 to 1 : 10, preferably within the range of 5:1 to 1 :5, preferably within the range of 5:1 to 1 :2, most preferably within the range of 3: 1 to 1 :1.

In a preferred embodiment of the invention, the pharmaceutical composition comprises one or more proanthocyanidins in the form of a proanthocyanidin-containing extract as described herein before, wherein the weight to weight ratio of hyaluronic acid to proanythocyanidin-containing extract is within the range of 10:1 to 1 :10, preferably within the range of 5:1 to 1 :5, preferably within the range of 5:1 to 1 :2, preferably within the range of 2:1 to 1 :1 , most preferably within the range of 1.5:1 to 1 :1.

Nw-meth ylserotonin

In particularly preferred embodiments of the invention, compositions as defined herein are provided, further comprising Nw-methylserotonin. Nw-methylserotonin is a tryptamine alkaloid which has the following molecular structure:


Nw-methylserotonin

Nw-methylserotonin occurs naturally in various plants, animals and fungi. Nw-methylserotonin may be obtained from a natural source or by chemical synthesis. In a preferred embodiment, the compositions of the invention comprise Nw-methylserotonin in the form of a herbal preparation, such as an extract. In a preferred embodiment, the compositions of the invention comprise Nw-methylserotonin in the form of a herbal preparation, preferably an extract, from a plant material derived from a plant species in a genus selected from the group consisting of Zanthoxylum, Citrus, Collema and Cimicifuga, preferably Cimicifuga. As will be understood by the skilled person, although the (former) genus Cimicifuga is now generally included in Actaea, for the purposes of this disclosure it is still considered as a separate genus. In a preferred embodiment, the compositions of the invention comprise Nw-methylserotonin in the form of a herbal preparation, preferably an extract, from plant material derived from Zanthoxylum piperitum, Zanthoxylum simulans, Zanthoxylum bungeanum, Zanthoxylum schinifolium, Zanthoxylum nitidum, Zanthoxylum rhetsa, Zanthoxylum alatum, Zanthoxylum acanthopodium, Zanthoxylum americanum, Citrus bergamia, Citrus medica, Citrus sinensis, Citrus reticulata, chinotto, Citrus limon, Collema cristatum, Collema callopismum, Collema flaccidum, Collema fuscovirens, or Cimicifuga racemosa, preferably Cimicifuga racemosa (also known as black cohosh).

The plant material may comprise fruit, leave and/or root, preferably the plant material comprises or consists of root. Hence, in a particularly preferred embodiment of the invention, the composition comprises Nw-methylserotonin in the form of a Cimicifuga extract, preferably a Cimicifuga root extract, more preferably a Cimicifuga racemosa extract, most preferably a Cimicifuga racemosa root extract. In preferred embodiments said Cimicifuga racemosa extract is made from sun-dried roots which were harvested in September or October.

Suitable extracts can be produced with polar solvents, such as with water, methanol, ethanol and mixtures thereof. In preferred embodiments the extract is produced with a solvent having a relative polarity of more than 0.5, such as more than 0.6 or more than 0.7.

Preferred extracts can be produced with water, with alcohols, in particular methanol, ethanol, propanol or mixtures thereof, or with mixtures of water and alcohol, preferably mixtures with a level of alcohol within the range of 10-95 %(v/v), 30-90 %(v/v) or 60-80 %(v/v) as the extraction solvent. In preferred embodiments the extract is produced with water, methanol, ethanol, water-ethanol or water-methanol mixtures.

In more preferred embodiments, the extract is produced with water, ethanol, or water-ethanol mixtures. Highly preferred extracts can be produced with water as the sole/main extraction solvent.

Hence, in accordance with a preferred embodiment of the invention, the composition comprises Nw-methylserotonin in the form of an aqueous extract, preferably a 100% water extract of a Nw-methylserotonin comprising plant material as described above. In an embodiment of the invention, a composition as defined herein is provided comprising an aqueous extract, preferably a 100% water extract of a Nw-methylserotonin comprising plant material. In accordance with a preferred embodiment of the invention, the aqueous extraction or the 100% water extraction, is performed at a temperature of more than 60°C, preferably more than 80°C, most preferably more than 95°C.

In a preferred embodiment of the invention, the composition comprises a Nw-methylserotonin-containing plant extract having a Nw-methylserotonin level of at least 0.1 wt.%, based on the dry weight of the Nw-methylserotonin-containing plant extract, i.e. based on the total weight of plant-derived components (excluding extraction solvent), e.g. at least 0.5 wt.%, at least 1 wt.%, at least 2 wt.%, at least 3 wt.%, at least 4 wt.% or at least 5 wt.%.

In accordance with a preferred embodiment of the invention, the composition comprises Nw-methylserotonin in the form of an extract, preferably an aqueous extract, more preferably a 100% water extract, wherein the extract comprises less than 10 wt.% of triterpene glycosides, based on the dry weight of the Nw-methylserotonin-containing plant extract, preferably less than 5 wt.%, preferably less than 3 wt.%.

Aqueous extracts, such as 100% water extracts, from Nw-methylserotonin-containing plant extracts may typically comprise one, two, three, four or five additional active ingredients selected from the group consisting of caffeic acid, ferulic acid, isoferulic acid, cimicifugoside H-1 , cimiracemoside A, cimicifugoside H-2, (26R)-actein, 26-deoxycimicifugoside, (26S)-actein, 23-epi-26-deoxyactein, 23-OAc-shengmanol-3-O-b-d-xyloside, 26-deoxyactein, 25-OAc-cimigenol-3-0-a-l-arabinoside, 25-OAc-cimigenol-3-0-p-d-xyloside, cimigenol-3-O-a-l-arabinoside, cimigenol-3-0-p-d-xyloside, kaempferol and formononetin.

In a preferred embodiment of the invention, the composition comprises a Nw-methylserotonin-containing plant extract obtainable by aqueous extraction, preferably a 100% water extraction, performed in such a way that the weight : weight ratio of (dry) plant material : extract is within the range of 4:1 to 6: 1 , wherein the weight of the extract is the dry weight, i.e. the total weight of plant derived components, excluding any extraction solvent.

In a preferred embodiment of the invention, the composition comprises a Nw-methylserotonin-containing plant extract obtainable by a method comprising the steps of:

a) providing Cimicifuga racemosa plant material, most preferably a Cimicifuga racemosa root;

b) macerating or grinding the plant material provided in step a);

c) performing an aqueous extraction, preferably a 100% water extraction, preferably at a temperature of more than 95°C;

d) concentrating the extract, preferably at a temperature not exceeding 95°C, more preferably not exceeding 85°C; and

e) drying, preferably spray-drying, the concentrated extract.

Nw-methylserotonin-containing extracts suitable for use in the context of the present invention, and the production thereof, have been described in the prior art, e.g. in Schmid, Diethart, et al. "Aqueous extracts of Cimicifuga racemosa and phenolcarboxylic constituents inhibit production of proinflam matory cytokines in LPS-stimulated human whole blood." Canadian journal of physiology and pharmacology 87.11 (2009): 963-972.

Nw-methylserotonin-containing extracts suitable for use in the context of the present invention are available commercially, e.g. from the company “NATURAL” in France under the reference “Cimicifuga root extract 4/1”.

In a preferred embodiment of the pharmaceutical composition, the weight to weight ratio of hyaluronic acid to Nw-methylserotonin is within the range of 100:1 to 1 :1 , preferably within the range of 60:1 to 1 :1 , preferably within the range of 40:1 to 5: 1 , most preferably within the range of 40: 1 to 9: 1 .

In a preferred embodiment of the invention, the pharmaceutical composition comprises Nw-methylserotonin in the form of a Nw-methylserotonin-containing extract as described herein before, wherein the weight to weight ratio of hyaluronic acid to Nw-methylserotonin-containing extract is within the range of 10:1 to 1 :10, preferably within the range of 5:1 to 1 :5, preferably within the range of 2:1 to 1 :2, preferably within the range of 1 .5: 1 to 1 : 1 .5, most preferably within the range of 1 .2: 1 to 1 : 1 .2.

Day-time composition

In certain preferred embodiments of the invention, compositions as defined herein are provided, further comprising one or two additional ingredients selected from a ginkgolide and vitamin C. Such compositions have proven to be particularly useful in the alleviation, prevention and/or suppression of one or more symptoms of menopause, while at the same time supporting day-time physiological activity levels. For ease of reference, compositions, formulations or products comprising these combinations of ingredients may be referred to as the ‘day-time composition’, ‘day-time formulation’ and ‘day-time product’ respectively throughout this document.

Ginkgolides

In particularly preferred embodiments of the invention, a composition as defined herein is provided which is a day-time composition further comprising one or more ginkgolides. Ginkgolides are terpene trilactones present in Ginkgo biloba and have the following molecular structure, wherein R1, R2 and R3 independently represent H or OH:


Ginkgolides

Ginkgolides occur naturally in ginkgo biloba, also known as the maidenhair tree, the only living species in the division Ginkgophyta. The one or more ginkgolides may be obtained from a natural source or by chemical synthesis. In a preferred embodiment, the day-time compositions of the invention comprise one or more ginkgolides in the form of a herbal preparation, such as powdered herbal material or an extract. In a preferred embodiment, the day-time compositions of the invention comprise one or more ginkgolides in the form of a herbal preparation from a plant material derived from Ginkgo biloba (interchangeably referred to herein as Ginkgo biloba L). The plant material may comprise fruit, leaf and/or root, preferably the plant material comprises or consists of leaf.

Hence, in a particularly preferred embodiment, the day-time compositions of the invention comprise one or more ginkgolides in the form of powdered Ginkgo biloba plant material, preferably powdered Ginkgo biloba leaf. In preferred embodiments said powdered Ginkgo biloba plant material comprises powdered, dried, e.g. sun-dried, leaves. Preferably, a germ reduction technique common in the art, such as steam treatment, is applied to the powdered plant material.

In an embodiment of the invention, the day-time composition comprises one or more ginkgolides in the form of a Ginkgo biloba extract, preferably a Ginkgo biloba leaf extract. In preferred embodiments said Ginkgo biloba extract is made from dried, e.g. sun-dried leaves.

In a preferred embodiment of the invention, the composition comprises a ginkgolide-containing plant extract having a ginkgolide level of at least 0.01 wt.%, based on the dry weight of the extract, i.e. based on the total weight of plant derived components excluding any extraction solvent, e.g. at least 0.005 wt.%, at least 0.01 wt.%, at least 0.05 wt.%, at least 0.1 wt.%, at least 0.2 wt.% or at least 0.3 wt.%.

In a preferred embodiment of the invention, the composition comprises a ginkgolide-containing plant extract having a ginkgolide level of at least 0.1 wt.%, based on the weight of the one or more ginkgolides-containing plant extract essentially free of extraction solvent, e.g. at least 0.5 wt.%, at least 1 wt.%, at least 2 wt.% or at least 3 wt.%.

In accordance with a preferred embodiment of the invention, the extract comprises less than 10 wt.%, based on the dry weight of the extract, of ginkgolic acid, preferably less than 5 wt.%, more preferably less than 3.5 wt.%.

Suitable extracts can be produced with polar solvents, such as with water, methanol, ethanol and mixtures thereof. In preferred embodiments the extract is produced with a solvent having a relative polarity of more than 0.5, such as more than 0.6 or more than 0.7. Preferred extracts can be produced with water, acetone, alcohols (in particular ethanol), or mixtures thereof.

In some preferred embodiment of the invention, the ginkgolide-containing extract is a water-acetone or a water-ethanol extract of Ginkgo biloba, preferably Ginkgo biloba leaf. Preferred extraction solvents are mixtures of water with acetone or ethanol, wherein the level of acetone or ethanol is within the range of 10-95 %(v/v), 30-90 %(v/v) or 60-90 %(v/v).

In other, particularly preferred embodiments, the extract is produced with water as the sole/main extraction solvent. Hence, in accordance with a preferred embodiment of the invention, the day-time composition comprises one or more ginkgolides in the form of an aqueous extract, preferably an extract produced with water as the sole extraction solvent (a ’100% water extract’), of Ginkgo biloba, preferably Ginkgo biloba leaf. In an embodiment of the invention, a day-time composition as defined herein is provided comprising an aqueous extract, preferably a 100% water extract of Ginkgo biloba, preferably of Ginkgo biloba leaf.

In accordance with a preferred embodiment of the invention, the aqueous extraction or the 100 % water extraction is performed at a temperature of more than 60°C, preferably more than 80°C, most preferably more than 95°C.

Aqueous extracts, including 100% water extracts, may typically comprise one, two, three, four, five, six or seven additional active ingredients selected from the group consisting of glucose, rhamnose, D-glucaric acid, ginkgolic acid, quercetin, kaempferol and isorhamnetin. In accordance with a preferred embodiment of the invention, the extract comprises more than 5 wt.%, based on the dry weight of extract, of flavone glycosides, preferably flavone glycosides selected from the group consisting of quercetin, kaempferol, isorhamnetin and combinations thereof, as can determined by the colorimetric method of Zhuang and Yu (Chinese Trad. Herbal Med. (1992) 23(3), 122-124), preferably more than 10 wt.%, more preferably more than 20 wt.%

In a preferred embodiment of the invention, the composition comprises an extract obtainable by aqueous extraction, preferably a 100% water extraction, which is performed in such a way that the weight : weight ratio of plant material : extract is within the range of 40: 1 to 60: 1 .

In a preferred embodiment of the invention, the composition comprises an extract obtainable by a method comprising the steps of:

a) providing Ginkgo biloba plant material, most preferably Ginkgo biloba leaf; b) macerating or grinding the plant material provided in step a);

c) performing an aqueous extraction, preferably a 100% water extraction, preferably at a temperature of more than 95°C;

d) concentrating the extract, preferably at a temperature not exceeding 95°C, more preferably not exceeding 85°C; and

e) drying, preferably spray-drying, the concentrated extract.

Ginkgolide-containing extracts suitable for use in the context of the present invention, and the production thereof, have been described in the prior art, e.g. in DING, ZHIEN. (2007). Studies on extraction and isolation of flavonoids from Ginkgo leaves. Journal of Food Quality 22(6):693 - 700 May 2007.

Ginkgolide-containing extracts suitable for use in the context of the present invention are available commercially, e.g. as the standardized extract EGb 761 or from the company “MB Med S.r.l.” in Italy under the reference “Ginkgo Biloba Foglie 3-464”.

L-Ascorbic acid

In particularly preferred embodiments of the invention, a composition as defined herein is provided which is a day-time composition further comprising vitamin C, interchangeably referred to herein as L-ascorbic acid.

Night time composition

In other preferred embodiments of the invention, compositions as defined herein are provided, further comprising one, two or three additional ingredients selected from valerenic acid, magnesium citrate and apigenins. Such compositions have proven to be particularly useful in the alleviation, prevention and/or suppression of one or more symptoms of menopause, while at the same time supporting night-rest. For ease of reference, any composition, formulation or product comprising these combinations of ingredients may be referred to as the ‘night-time composition’, ‘night-time formulation’, and ‘night-time product’, throughout this document.

Valerenic acid

In particularly preferred embodiments of the invention, night-time compositions as defined herein are provided comprising valerenic acid.

Valerenic acid is naturally occurring hydrophilic sesquiterpenoid. Valerenic acid has the CAS Registry Number 3569-10-6 and systematic (lUPAC) name (2E)-3-[(4S,7R,7aR)-3,7-dimethyl-2,4,5,6,7,7a-hexahydro-1 FI-inden-4-yl]-2-methylacrylic acid. Valerenic acid has the following molecular structure:

Valerenic acid

Valerenic acid occurs naturally in various plants, such as plants in the Valerianaceae family. As will be understood by the skilled person, although the (former) family Valerianaceae is now generally included in Caprifoliaceae, for the purposes of this disclosure it is still considered as a separate family. Valerenic acid occurs naturally in various plants, such as plants in the genera centranthus, fedia, nardostachys, patrinia, plectritis, valeriana and valerianella.

Plants of particular interest as a valerenic acid source are plants in the genus valeriana, such as Valeriana alypifolia, Valeriana aretioides, Valeriana aretioides, Valeriana asterothrix, Valeriana bertiscea, Valeriana buxifolia, Valeriana californica, Valeriana celtica (Alpine valerian or valerian spikenard), Valeriana cernua, Valeriana coleophylla, Valeriana dioscoridis, Valeriana edulis, Valeriana fauriei (Korean valerian), Valeriana glechomifolia, Valeriana kreyeriana, Valeriana montana, Valeriana occidentalis, Valeriana officinalis (garden valerian), Valeriana pancicii, Valeriana pauciflora, Valeriana saxatilis, Valeriana scouleri (Scouler's valerian), Valeriana secunda, Valeriana sitchensis (Sitka valerian) and Valeriana uliginosa, with Valeriana officinalis being a preferred source.

Valerenic acid may be obtained from a natural source or by chemical synthesis.

In a preferred embodiment, the night-time compositions of the invention comprise valerenic acid in the form of a herbal preparation, preferably an extract. In a preferred embodiment, the night-time compositions of the invention comprise valerenic acid in the form of a herbal preparation, preferably an extract, from a plant material derived from a plant species in a genus selected from the group consisting of centranthus, fedia, nardostachys, patrinia, plectritis, valeriana and valerianella, preferably the night-time compositions of the invention comprise valerenic acid in the form of a herbal preparation, preferably an extract, from a plant material derived from a plant species in the genus valeriana. In a preferred embodiment, the night-time compositions of the invention comprise valerenic acid in the form of a herbal preparation, preferably an extract, from plant material derived from Valeriana alypifolia, Valeriana aretioides, Valeriana aretioides, Valeriana asterothrix, Valeriana bertiscea, Valeriana buxifolia, Valeriana californica, Valeriana celtica (Alpine valerian or valerian spikenard), Valeriana cernua, Valeriana coleophylla, Valeriana dioscoridis, Valeriana edulis, Valeriana fauriei (Korean valerian), Valeriana glechomifolia, Valeriana kreyeriana, Valeriana montana, Valeriana occidentalis, Valeriana officinalis (garden valerian), Valeriana pancicii, Valeriana pauciflora, Valeriana saxatilis, Valeriana scouleri (Scouler's valerian), Valeriana secunda, Valeriana sitchensis (Sitka valerian) or Valeriana uliginosa, preferably the night-time compositions of the invention comprise valerenic acid in the form of a herbal preparation, preferably an extract, from plant material derived from Valeriana officinalis.

The plant material preferably comprises or consists of stem, leaves or root, most preferably root.

Hence, in preferred embodiments the extract is a root extract, preferably an extract of the roots of Valeriana officinalis.

Suitable extracts can be produced with polar solvents, such as with water, methanol, ethanol and mixtures thereof. In embodiments the extract is produced with a solvent having a relative polarity of more than 0.5, such as more than 0.6 or more than 0.7 or more than 0.8.

Preferred extracts can be produced with water, methanol, ethanol, propanol or mixtures thereof, preferably with a level of alcohol within the range of 10-60 %(v/v), preferably within the range of 20-40 %(v/v), preferably within the range of 25-35 %(v/v), preferably within the range of 28-32 %(v/v). In preferred embodiments the extract is produced with water, methanol, ethanol or water-ethanol or water-methanol mixtures. In more preferred embodiments, the extract is produced with water, ethanol, or water-ethanol mixtures. Most preferably, the extract is produced with water-ethanol mixtures.

In embodiments the extract is produced with a water-ethanol mixture with a level of ethanol within the range 10-60 %(v/v), preferably within the range of 20-40 %(v/v), preferably within the range of 25-35 %(v/v), preferably within the range of 28-32 %(v/v).

Hence, in accordance with a preferred embodiment of the invention, the night time composition comprises valerenic acid in the form of an extract of a valerenic acid comprising plant material produced with water, methanol, ethanol, propanol or

mixtures thereof, preferably a water-ethanol extract. In an embodiment of the invention, a night-time composition as defined herein is provided comprising a water, methanol, ethanol or water-ethanol or water-methanol extract of a valerenic acid-containing plant material, preferably a water-ethanol extract.

In accordance with a preferred embodiment of the invention, the night-time composition comprises valerenic acid in the form of a water-ethanol extract produced with a water-ethanol mixture with a level of alcohol within the range of 10-60 %(v/v), preferably within the range of 20-40 %(v/v), more preferably within the range of 25-35 %(v/v), most preferably within the range of 28-32 %(v/v), from Valeriana officinalis root.

The valerenic acid-containing plant extracts, may typically comprise one, two, three, four, five or six additional active ingredients selected from the group consisting of hydroxyvalerenic acid, acetoxyvalerenic acid, valepotriates, baldrinal, homobaldrinal, valtrate, valeranone and alkaloids.

In a preferred embodiment of the invention, the composition comprises a valerenic acid-containing plant extract obtainable from a water-ethanol extraction employing a water-ethanol mixture with a level of alcohol within the range of 10-60 %(v/v), preferably within the range of 20-40 %(v/v), preferably within the range of 25-35 %(v/v), preferably within the range of 28-32 %(v/v) as the extraction solvent, wherein the extraction is performed in such a way that an extract is produced that, excluding any extraction solvent, is from 1/6 - 1/3 of the (dry) weight of the plant material before extraction.

In a preferred embodiment of the invention, the night-time composition comprises a valerenic acid-containing plant extract, having a total amount of valerenic acid of at least 0.01 wt.%, based on the weight of the valerenic acid-containing plant extract essentially free of extraction solvent, e.g. at least 0.05 wt.%, at least 0.1 wt.%, at least 0.15 wt.%, at least 0.2 wt.% or at least 0.3 wt.%, preferably as determined in accordance with the method described in the EP hydroalcoholic Valerian dry extract monograph.

In a preferred embodiment of the invention, the night-time composition comprises a valerenic acid-containing plant extract obtainable by a method comprising the steps of:

a) providing Valeriana officinalis plant material, preferably Valeriana officinalis roots;

b) macerating or grinding the plant material provided in step a);

c) performing an extraction with a water-ethanol mixture with a level of alcohol within the range of 10-60 %(v/v), preferably within the range of 20-40 %(v/v), more preferably within the range of 25-35 %(v/v), even more preferably within the range of 28-32 %(v/v), most preferably at a temperature of less than 90°C;

d) concentrating the extract, preferably at a temperature not exceeding 70°C, more preferably not exceeding 60°C, most preferably not exceeding 45°C; and

e) drying, preferably spray-drying, the concentrated extract.

Extracts suitable for use in the context of the present invention, and the production thereof, have been described in the prior art, e.g. in:

• Boyadzhiev, L, et al. "Extraction of valerenic acids from valerian (Valeriana officinalis L.) rhizomes." Die Pharmazie-An International Journal of Pharmaceutical Sciences 59.9 (2004): 727-728.;

• EP1235583A1 .

Valerenic acid-containing extracts suitable for use in the context of the present invention are available commercially, e.g. from the company “Evear extraction solutions” in France under the reference “Valerian dry extract >0.3% E2123ST”.

Magnesium citrate

In a preferred embodiment of the invention, the composition further comprises magnesium. Suitable magnesium sources may comprise any pharmaceutically acceptable inorganic or organic compound containing magnesium such as magnesium carbonate, magnesium hydroxide, magnesium gluconate, magnesium oxide, magnesium citrate or magnesium lactate; preferably magnesium citrate or magnesium lactate, most preferably magnesium citrate. Magnesium citrate is preferably provided in the form of trimagnesium citrate, most preferably in the form of anhydrous trimagnesium citrate. Current guidelines in the US provide a recommended daily amount of magnesium of 300 - 400 mg (for adult women). It has been well established that magnesium contributes to increased hepatic metabolism. The present inventors believe that the increased hepatic effect of magnesium increases the sympathetic effects of the proanthocyanins and allows for increased adsorption. Hence, in a highly preferred embodiment of the invention, the composition comprises magnesium in the form of trimagnesium citrate, preferably anhydrous trimagnesium citrate.

Magnesium sources suitable for use in the context of the present invention are available commercially, e.g. from the company “Quimdis S.A.S.” in France under the reference “trimagnesium citrate anhydrous”.

Apiaenin(s)

In particularly preferred embodiments of the invention, night-time compositions as defined herein are provided comprising apigenins.

Apigenin, 4',5,7-trihydroxyflavone, is a naturally occurring compound belonging to the class of flavones. Apigenin has the CAS Registry Number 520-36-5 and systematic (lUPAC) name 5,7-Dihydroxy-2-(4-hydroxyphenyl)-4H-1-benzopyran-4-one. Common synonyms include Apigenine; Chamomile; Apigenol; Spigenin; Versulin; 4',5,7-Trihydroxyflavone and C.l. Natural Yellow 1. Apigenin has the following molecular structure:


Apigenin

In natural conditions, apigenin predominantly occurs in the form of an apigenin glycoside, wherein apigenin is bound to a sugar through a glycosidic bond. As used herein, the term “apigenin” refers to the aglycone of apigenin glycosides. The plural form, i.e. “apigenins”, is used herein to generically denote apigen and the apigenin glycosides, as will be understood by those skilled in the art.

Apigenin and apigenin glycosides occur naturally in various plants, such as plants in the genera artemisia, perilla, urtica, passiflora, camelia, cayaponia, colocasia, desmodium, hordeum, origanum, ocimum, jatropha, parkinsonia, peperomia, piheranthos, centaurea, indigo, bomba, lychnophera, asplenium, chinotto, citrus, viola, trigonella, lamiacea, labiatae, lavandula, nipponanthemum, abrus, santalum, oryza, scleropyrum, tulsi, indigofera, bombax, glinus, lychnophora, urticaceae, rosales, malpighiales, thymus, prunus, camellia, olea, brassica, apium, petroselinum, matricaria, chamaemelum, and chrystanthemum.

Plants of particular interest as an apigenin or apigenin glycoside source are vitex agnus-castus, petroselinum crispum, apium graveolens, citrus paradisi, citrus maxima, perilla frutescens, Matricaria chamomilla, chrysanthemum morifolium and chamaemelum nobile, and in particular matricaria recutita (also known as Matricaria chamomilla); chamaemelum nobile; and chrysanthemum morifolium. Apigenin or apigenin glycosides are particularly abundant in the flowers of chamomile plants, constituting up to 68% of total flavonoids. In most camomille plant varieties, apigenin-7-O-glucoside is the dominant form and is present in about 100-500mg/100g flowers, while apigenin is present in an amount of about 5-25mg/100g flowers.

In preferred embodiments, the night-time compositions of the invention comprise apigenin or an apigenin glycoside, preferably an apigenin glycoside selected from the group consisting of apigenin 7-O-apioglucoside (apiin), apigenin 7-0-glucoside (apigetrin), apigenin 8-C-glucoside (vitexin), apigenin 6-C-glucoside (isovitexin), apigenin 7-O-neohesperidoside (rhoifolin), or apigenin 6-C-glucoside 8-C-arabinoside (schaftoside), preferably apigenin 7-O-apioglucoside (apiin) or apigenin 7-O-glucoside (apigetrin), most preferably apigenin 7-O-glucoside.

In natural conditions, the apigenin or apigenin glycoside commonly exists in an acetylated form, such as a monoacetylated or diacetylated form. In embodiments, the night-time compositions and extracts described herein comprise acetylated apigenin or apigenin glycoside.

Apigenin and/or apigenin glycosides may be obtained from a natural source or by chemical synthesis.

In a preferred embodiment, the night-time compositions of the invention comprise apigenin or an apigenin glycoside in the form of a herbal preparation, preferably an extract. In a preferred embodiment, the night-time compositions of the invention comprise apigenin or an apigenin glycoside in the form of a herbal preparation, preferably an extract, from a plant material derived from a plant species in a genus selected from the group consisting of artemisia, perilla, urtica, passiflora, camelia, cayaponia, colocasia, desmodium, hordeum, origanum, ocimum, jatropha, parkinsonia, peperomia, piheranthos, centaurea, indigo, bomba, lychnophera, asplenium, chinotto, citrus, viola, trigonella, lamiacea, labiatae, lavandula, nipponanthemum, abrus, santalum, oryza, scleropyrum, tulsi, indigofera, bombax, glinus, lychnophora, urticaceae, rosales, malpighiales, thymus, prunus, camellia, olea, brassica, apium, petroselinum, matricaria, chamaemelum, and chrystanthemum,

preferably the night-time compositions of the invention comprise apigenin or an apigenin glycoside in the form of a herbal preparation, preferably an extract, from a plant material derived from a plant species in a genus selected from the group consisting of thymus, prunus, camellia, olea, brassica, apium, petroselinum, matricaria, chamaemelum and chrystanthemum. In a preferred embodiment, the night-time compositions of the invention comprise apigenin or an apigenin glycoside in the form of a herbal preparation, preferably an extract, from plant material derived from vitex agnus-castus, petroselinum crispum, apium graveolens, citrus paradisi, citrus maxima, perilla frutescens, Matricaria chamomilla, chrysanthemum morifolium or chamaemelum nobile, preferably the night-time compositions of the invention comprise apigenin or an apigenin glycoside in the form of a herbal preparation, preferably an extract, from plant material derived from matricaria recutita (also known as Matricaria chamomilla), chamaemelum nobile, or chrysanthemum morifolium.

The plant material preferably comprises or consists of leaves and flowers, preferably flowers, preferably the ray florets.

Hence, in preferred embodiments the extract is a flower extract, preferably the ray florets of matricaria recutita (also known as Matricaria chamomilla), chamaemelum nobile, or chrysanthemum morifolium, preferably an extract of the flowers, preferably the ray florets of matricaria recutita (also known as Matricaria chamomilla).

Suitable extracts can be produced with polar solvents, such as with water, methanol, ethanol and mixtures thereof. In embodiments the extract is produced with a solvent having a relative polarity of more than 0.5, such as more than 0.6 or more than 0.7 or more than 0.8.

Preferred extracts can be produced with water, methanol, ethanol, propanol or mixtures thereof, preferably with a level of alcohol within the range of 10-60 %(v/v), preferably within the range of 20-40 %(v/v), preferably within the range of 25-35 %(v/v), preferably within the range of 28-32 %(v/v). In preferred embodiments the extract is produced with water, methanol, ethanol or water-ethanol or water-methanol mixtures. In more preferred embodiments, the extract is produced with water, ethanol, or water-ethanol mixtures. Most preferably, the extract is produced with water-ethanol mixtures.

In embodiments the extract is produced with a water-ethanol mixture with a level of ethanol within the range 10-60 %(v/v), preferably within the range of 20-40 %(v/v), preferably within the range of 25-35 %(v/v), preferably within the range of 28-32 %(v/v).

Hence, in accordance with a preferred embodiment of the invention, the night time composition comprises apigenin or an apigenin glycoside in the form of a water, methanol, ethanol, propanol or mixtures thereof of an apigenin or an apigenin glycoside-comprising plant material, preferably a water-ethanol extract. In an embodiment of the invention, a night-time composition as defined herein is provided comprising a water, methanol, ethanol or water-ethanol or water-methanol extract of an apigenin or an apigenin glycoside-containing plant material, preferably a water-ethanol extract.

In embodiments, the extract is rich in apigenin-7-O-glucoside and contains more apigenin-7-O-glucoside than apigenin. In embodiments the extract is rich in apigenin-7-O-glucoside and more than 20%, preferably more than 30%, more than 50%, or more than 70% of the apigenin and apigenin-glycoside content is apigenin-7-O-glucoside. Such apigenin-7-O-glucoside-rich extracts can be produced with water, ethanol or methanol as the extraction medium, preferably water or ethanol.

In embodiments the extract is rich in apigenin and contains more apigenin than apigenin-7-O-glucoside. In embodiments the extract is rich in apigenin and more than 10%, preferably more than 20%, more than 50%, more than 70% of the apigenin and apigenin-glycoside content is apigenin. Such apigenin rich extracts can be produced with water-ethanol or water-methanol mixtures, preferably water-ethanol mixtures, preferably with a level of alcohol lower than 50 %(v/v), preferably lower than 40 %(v/v), preferably lower than 30 %(v/v). Although the flavonoid yield of such extracts rich in apigenin may be lower than the flavonoid yield of extracts rich in apigenin-7-O-glucoside, the relative increase of apigenin content may be desirable since apigenin is believed to be more physiologically active than apigenin glycosides.

In accordance with a preferred embodiment of the invention, the night-time composition comprises apigenin or apigenin-glycoside in the form of a water-ethanol extract produced with a water-ethanol mixture with a level of alcohol within the range of 10-60 %(v/v), preferably within the range of 20-40 %(v/v), more preferably within the range of 25-35 %(v/v), most preferably within the range of 28-32 %(v/v), from a flower, preferably a ray floret, of matricaria recutita (also known as Matricaria chamomilla), chamaemelum nobile, or chrysanthemum morifolium, preferably of matricaria recutita.

The apigenin or apigenin-glycoside-containing plant extracts may typically comprise one, two, three, four or five additional active ingredients selected from the group consisting of hyperoside, herniarin, umbelliferone, chlorogenic acid, caffeic acid, luteolin, luteolin-7-O-glucoside, quercetin, rutin, and naringenin.

The apigenin or apigenin-glycoside-containing plant extracts may typically be prepared by milling, crushing or grinding plant material as described herein earlier, and suspending the plant material in the extraction medium, for example at a concentration of 0.01-50 %(w/v) of fresh plant material, preferably 0.1-20 %(w/v), more preferably 0.5-10 %(w/v), yet more preferably 1-7 %(w/v), most preferably 2-6 %(w/v).

In a preferred embodiment of the invention, the night-time composition comprises an apigenin or apigenin-glycoside-containing plant extract, having a total amount of apigenin and apigenin-glycosides of at least 0.1 wt.%, based on the weight of the apigenin or apigenin-glycoside-containing plant extract essentially free of extraction solvent, e.g. at least 0.5 wt.%, at least 1 wt.%, at least 1.5 wt.%, at least 2 wt.% or at least 5 wt.%, preferably as determined in accordance with the method described in the USP chamomile monograph.

In a preferred embodiment of the invention, the night-time composition comprises an apigenin or apigenin-glycoside-containing plant extract obtainable by a method comprising the steps of:

a) providing Matricaria recutita (also known as Matricaria chamomilla) plant material, preferably Matricaria recutita flowers, preferably Matricaria recutita ray florets; b) preferably macerating or grinding the plant material provided in step a);

c) performing an extraction with a water-ethanol mixture with a level of alcohol within the range of 10-60 %(v/v), preferably within the range of 20-40 %(v/v), more preferably within the range of 25-35 %(v/v), even more preferably within the range of 28-32 %(v/v), most preferably at a temperature of less than 90°C;

d) concentrating the extract, preferably at a temperature not exceeding 70°C, more preferably not exceeding 60°C, most preferably not exceeding 45°C; and

e) drying, preferably spray-drying, the concentrated extract.

Extracts suitable for use in the context of the present invention, and the production thereof, have been described in the prior art, e.g. in

• Srivastava JK, Gupta S. Extraction, Characterization, Stability and Biological Activity of Flavonoids Isolated from Chamomile Flowers. Molecular and cellular pharmacology. 2009; 1 (3): 138;

• EP2444061A1 ;

• EP0496230A1 ;

• EP0330240A1 ;

• Chamomile: Industrial Profiles - Rolf Franke, Heinz Schilcher (2005).

Apigenin-containing extracts suitable for use in the context of the present invention are available commercially, e.g. from the company “Evear extraction solutions” in France under the reference “Chamomile dry extract 5% apigenin E2423A5”.

It is to be understood that many or most of the pharmacologically and/or physiologically active ingredients as described herein can exist in the form of a pharmaceutically acceptable salt or solvate. Such forms will typically be equally suitable for use in the present invention. It is to be understood that it is within the scope of the present invention to include pharmaceutically acceptable salt forms, solvates or esters of any of the active ingredients recited herein in the compositions in addition to or instead of the corresponding ingredient in free acid or unsolvated form. Whenever, in this document, the amount of a certain ingredient is quantified, this refers to the amount of the active ingredient as such, excluding the weight of any counterion or solvated compounds that may be present (e.g. in a given extract), unless a specific salt is explicitly referred to (as is e.g. the case for magnesium carbonate or sodium hyaluronate).

In accordance with preferred embodiments of the invention compositions as defined herein are provided comprising:

• a combination of hyaluronic acid and Pinus pinaster bark extract; or

• a combination of hyaluronic acid, Pinus pinaster bark extract and Cimicifuga racemosa root extract.

In accordance with preferred embodiments of the invention, day-time compositions as defined herein are provided comprising:

• a combination of hyaluronic acid, Pinus pinaster bark extract, Cimicifuga racemosa root extract and vitamin C;

• a combination of hyaluronic acid, Pinus pinaster bark extract, Cimicifuga racemosa root extract and powdered Ginkgo biloba leaf; or

• a combination of hyaluronic acid, Pinus pinaster bark extract, Cimicifuga racemosa root extract, vitamin C and powdered Ginkgo biloba leaf.

In accordance with preferred embodiments of the invention, night-time compositions as defined herein are provided comprising:

• a combination of hyaluronic acid, Pinus pinaster bark extract, Cimicifuga racemosa root extract and Matricaria recutita ray floret extract;

• a combination of hyaluronic acid, Pinus pinaster bark extract, Cimicifuga racemosa root extract and Valeriana officinalis root extract;

• a combination of hyaluronic acid, Pinus pinaster bark extract, Cimicifuga racemosa root extract and a magnesium source, preferably trimagnesium citrate;

• a combination of hyaluronic acid, Pinus pinaster bark extract, Cimicifuga racemosa root extract, Matricaria recutita ray floret extract and Valeriana officinalis root extract;

• a combination of hyaluronic acid, Pinus pinaster bark extract, Cimicifuga racemosa root extract, Valeriana officinalis root extract and a magnesium source, preferably trimagnesium citrate; or

• a combination of hyaluronic acid, Pinus pinaster bark extract, Cimicifuga racemosa root extract, Matricaria recutita ray floret extract, Valeriana officinalis root extract, and a magnesium source, preferably trimagnesium citrate.

Unit dosage form

Compositions as defined herein are preferably provided in the form of a pharmaceutical formulation or in the form of a nutraceutical formulation, which may also be referred to as a food supplement or food supplement formulation. In preferred embodiments, the invention provides a pharmaceutical or nutraceutical formulation in the form of a so-called ‘unit dosage form’, in particular a unit dosage form that is suitable for oral administration. The term "unit dosage form" refers to a physically discrete unit of the pharmaceutical or nutraceutical formulation comprising a fixed, pre determined dose of the pharmaceutically and/or physiologically active ingredient(s) in a form suitable for administration to a subject in need thereof.

In accordance with a preferred embodiment of the invention, a unit dosage form comprises hyaluronic acid or a pharmaceutically acceptable salt thereof, in an amount of 1 mg or more, preferably 10 mg or more, preferably 15 mg or more and/or in an amount of 200 mg or less; preferably 50 mg or less, preferably 25 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form comprises proanthocyanidins in an amount of 0.1 mg or more, as determined by gel-

permeation chromatography, preferably 1 mg or more, preferably 8 mg or more and/or 200 mg or less, preferably 50 mg or less, preferably 15 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form comprises Nw-methylserotonin in an amount of 0.01 mg or more, preferably 0.1 mg or more, preferably 0.5 mg or more and/or 50 mg or less, preferably 15 mg or less, preferably 2 mg or less

In accordance with a preferred embodiment of the invention, a unit dosage form, which is preferably a day-time composition, comprises vitamin C in an amount of 1 mg or more, preferably 50 mg or more, preferably 200 mg or more and/or in an amount of 1500 mg or less, preferably 700 mg or less, preferably 250 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form, which is preferably a day-time composition, comprises ginkgolides in an amount of 0.001 mg or more, preferably 0.01 mg or more, preferably 0.1 mg or more and/or in an amount of 100 mg or less, preferably 10 mg or less, preferably 1 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form, which is preferably a night-time composition, comprises valerenic acid in an amount of 0.001 mg or more, preferably 0.01 mg or more, preferably 0.05 mg or more, preferably 0.1 mg or more and/or in an amount of 100 mg or less, preferably 10 mg or less, preferably 1 mg or less, preferably 0.5 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form, which is preferably a night-time composition, comprises apigenins in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 5 mg or more and/or in an amount of 100 mg or less, preferably 50 mg or less, preferably 20 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form, which is preferably a night-time composition, comprises a magnesium compound, preferably trimagnesium citrate, in an amount of 1 mg or more, preferably 50 mg or more, preferably 350 mg or more and/or in an amount of 2000 mg or less, preferably 1000 mg or less, preferably 400 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form comprises Pinus pinaster bark extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 10 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 20 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form comprises Cimicifuga racemosa root extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 15 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 25 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form, which is preferably a day-time composition, comprises powdered Ginkgo biloba leaf in an amount of 1 mg or more, preferably 10 mg or more, preferably 80 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 160 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form, which is preferably a day-time composition, comprises one, two or three of the following:

• glycosylated quercetin in an amount of 0.05 mg or more, preferably 0.5 mg or more, preferably 1 mg or more and/or in an amount of 10 mg or less, preferably 5 mg or less, preferably 2 mg or less;

• glycosylated kaempferol in an amount of 0.05 mg or more, preferably 0.5 mg or more, preferably 1 mg or more and/or in an amount of 10 mg or less, preferably 5 mg or less, preferably 2 mg or less;

• glycosylated isorhamnetin in an amount of 0.01 mg or more, preferably 0.05 mg or more, preferably 0.2 mg or more and/or in an amount of 5 mg or less, preferably 1 mg or less, preferably 0.6 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form, which is preferably a night-time composition, comprises Valeriana officinalis root extract in an amount of 1 mg or more, preferably 10 mg or more, preferably 70 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 130 mg or less.

In accordance with a preferred embodiment of the invention, a unit dosage form, which is preferably a night-time composition, comprises Matricaria recutita ray floret extract in an amount of 1 mg or more, preferably 10 mg or more, preferably 80 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 140 mg or less.

In a particularly preferred embodiment of the invention a day-time unit dosage form comprises:

• hyaluronic acid or a pharmaceutically acceptable salt thereof, preferably sodium hyaluronate in an amount of 1 mg or more, preferably 10 mg or more, preferably 18 mg or more and/or in an amount of 200 mg or less; preferably 50 mg or less, preferably 20 mg or less;

• Pinus pinaster bark extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 10 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 20 mg or less, such as 12 mg, 14 mg, 15 mg, 16 mg or 18 mg;

• Cimicifuga racemosa root extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 15 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 25 mg or less, such as 17 mg, 19 mg, 20 mg, 21 mg or 23 mg;

• vitamin C in an amount of 1 mg or more, preferably 50 mg or more, preferably 200 mg or more and/or in an amount of 1500 mg or less, preferably 700 mg or less, preferably 250 mg or less; and

• powdered Ginkgo biloba leaf in an amount of 1 mg or more, preferably 10 mg or more, preferably 80 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 160 mg or less, such as 100 mg, 115 mg, 120 mg, 125 mg or 140 mg.

In a particularly preferred embodiment of the invention a night-time unit dosage form comprises:

• hyaluronic acid or a pharmaceutically acceptable salt thereof, in an amount of 1 mg or more, preferably 10 mg or more, preferably 18 mg or more and/or in an amount of 200 mg or less; preferably 50 mg or less, preferably 20 mg or less;

• Pinus pinaster bark extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 10 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 20 mg or less, such as 12 mg, 14 mg, 15 mg, 16 mg or 18 mg;

• Cimicifuga racemosa root extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 15 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 25 mg or less, such as 17 mg, 19 mg, 20 mg, 21 mg or 23 mg;

• Valeriana officinalis root extract in an amount of 1 mg or more, preferably 10 mg or more, preferably 70 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 130 mg or less, such as 80 mg, 95 mg, 100 mg, 105 or 120 mg;

• Matricaria recutita ray floret extract in an amount of 1 mg or more, preferably 10 mg or more, preferably 80 mg or more and/or in an amount of 1000 mg or less,

preferably 500 mg or less, preferably 140 mg or less, such as 90 mg, 105 mg, 110 mg, 115 mg or 130 mg; and

• a magnesium compound, preferably trimagnesium citrate in an amount of 1 mg or more, preferably 50 mg or more, preferably 350 mg or more and/or in an amount of 2000 mg or less, preferably 1000 mg or less, preferably 400 mg or less.

Pharmaceutical and/or nutraceutical formulations as defined herein are preferably provided in the form of a solid oral unit dosage form. Suitable examples include tablets, capsules, lozenges as well as particulates, such as granulates and powders, typically contained in a unit dosage packaging, such a sachet. Embodiments are also envisaged, wherein the pharmaceutical and/or nutraceutical formulation is a liquid suitable for oral administration, such as a syrup, elixir or the like, e.g. contained in a (unit dosage) vial or ampoule or in a bottle or flask (from which a single unit dose must be drawn). In particularly preferred embodiments though, the unit dosage form is a solid oral unit dosage form, more preferably a tablet or capsule suitable for oral administration. Regarding general techniques of manufacturing oral solid dosage forms that can suitably be used in accordance with the present invention, reference is made to Remington, The Science and Practice of Pharmacy, 22nd edition, 2012, chapter 45 "Oral solid dosage forms". Most preferably, the unit dosage form is a tablet for oral administration.

Tablets that can suitably be used for the purposes of the present invention typically comprise a core, comprising a combination of components as specified herein elsewhere in conjunction with one or more conventional excipients needed to (e.g.) facilitate processing of said combination into a tablet core that remains intact during processing, storing and distribution, that is convenient for a patient to self-administer and swallow and that disintegrates in the patient’s gastrointestinal tract upon swallowing at the appropriate rate. Typically, the tablet core comprises, besides the combination of pharmacologically and/or physiologically active ingredients, one or more of a bulking agent, such as sorbitol or microcrystalline cellulose, anti-caking agents, such as colloidal sillicon, lubricants, such as glyceryl behenate, binders, fillers, disintegrants, preservatives, emollients, humectants, sweetening-agents, plasticizers, flim-forming agents, hardness-increasing agents, flavours, etc.

The tablet core can suitably be produced by direct compression of a (free-flowing) mixture or blend of the combination of pharmacologically and/or physiologically active ingredients and the excipients or by the formation of a granulate comprising the combination of pharmacologically and/or physiologically active ingredients and the excipients, optionally using a granulation liquid, which granulate is subsequently compressed into a tablet (core).

In preferred embodiments of the invention, the tablets comprise a coating surrounding the core as defined here above. It is conventional in the art to apply a coating on a tablet formulation, in particular in order to improve the visual appearance of the tablet, in order to facilitate swallowing of the tablet, in order to mask unpleasant taste of tablet components, in order to confer a distinctive and recognizable appearance and/or in order to control disintegration of the tablet and release of the active ingredients from the tablet. In accordance with the present invention, a coating is applied using conventional coating compositions and techniques, e.g. using coatings comprising film-forming agents, thickener, anticaking agent, lubricant and colorants.

In particularly preferred embodiments of the invention, the coating comprises a conventional colorant or pigment, such as titanium dioxide, indigo carmine, iron black oxide, carminic acid, etc. The presence of a colorant in the coating not only renders the unit dosage form more visually appealing and/or attractive, but also serves the purpose of making the unit dosage form more easily recognizable. More in particular, it is contemplated to provide day-time unit dosage forms and night time unit dosage-forms, in accordance with what is described herein elsewhere, wherein the coating of the day-time unit dosage form has a color distinct from that of the coating of the night time unit dosage form. For instance, day-time unit dosage forms are provided with a coating having a colour that is generally associated with day-time, e.g. a bright colour, such as a skye-blue colour, and night-time unit dosage forms are provided with a coating having a colour that is generally associated with night-time, e.g. a dark colour, such as a navy-blue colour.

Kit of parts

A further aspect of the invention is directed to a kit of parts comprising a plurality of first unit dosage forms as a day-time component, having the composition as defined herein elsewhere, as well as a plurality of second unit dosage forms as the night-time component, having the composition as defined herein elsewhere. In accordance with the invention, such a kit may comprise a plurality of unit dosage forms, such as 5, 10, 15, 20, 25, 30, 40, 50, 60, 70, 75, 80, 90 or 100 of each of the first unit dosage forms and the second unit dosage forms. In one particularly preferred embodiment, a kit as

described herein comprises said first unit dosage forms in a total number of 30 and said second unit dosage forms in a total number of 30 so that the kit can be used by a subject to cover a treatment period of 30 days, during which period said subject takes one of said first unit dosage forms per day and one of said second unit dosage forms per day.

In preferred embodiments, said kit of parts further comprises printed instructions for using the product in the treatment of a perimenopausal, menopausal or postmenopausal female subject or a female subject that suffers from and/or is at risk of suffering from menopausal syndrome, by administering one of said first unit dosage forms in the morning, preferably the early morning and/or within a certain time interval after the subject wakes up as defined/explained herein elsewhere, and by administering one of said second unit dosage forms in the evening and/or within a certain time interval before the subject in need of the treatment goes to sleep or intends to go to sleep, as defined/explained herein elsewhere.

Therapeutic Indication

A further aspect of the invention is directed to, stated generally, a method of treating a subject in need thereof, using the compositions as defined herein. Based on the information provided for the first time herein, it is contemplated that the compositions of the invention are useful in methods of treating female subjects in perimenopause, menopause or post-menopause, so as to prevent, alleviate and/or suppress one or more symptoms associated with menopause and/or to improve the well-being of said subjects in general.

As used herein, the term ‘method of treating a subject’ generally refers to an intervention, in particular to the act of administering the composition, carried out with the aim of affecting one or more physiological and/or pathological processes in the subject in a way that is beneficial to the subject’s health and/or well-being. Improving health and/or well-being can, generally speaking, be accomplished by preventing or curing a disease or condition, but also by preventing, suppressing or alleviating one or more symptoms of a disease or condition (i.e. even if it does not remove, or lessen the impact of, the underlying cause) or by affecting (inducing or suppressing) any physiological effect that is not perse associated with the disease or condition but that aids to the overall feeling of well-being of the subject nonetheless.

As explained in the introductory part of the present document, menopause is a process every female goes through as a consequence of the normal aging process and is characterized by a sharp decrease in endogenous estrogen production following loss of ovarian follicular activity, as a consequence of which menses becomes irregular and eventually ceases. The present invention relates to treatment of perimenopausal subjects, menopausal subjects as well as postmenopausal subjects. In the context of the present invention, the term ‘perimenopausal subject’ is used to denote subjects undergoing the transition process, starting at the point the first physical changes and the first irregularities in menstrual patterns become apparent and up to the point that the subject has not experienced menses for more than 12 months, at which point ‘menopause’ is conventionally diagnosed. The term ‘postmenopausal subject’ is used to denote subjects going through the subsequent stage, during which symptoms associated with menopause may, and often do, persist. It may take years, typically up to 5 years after the subject’s last menses, until the subject does not experience any menopause symptoms anymore.

The terms ‘menopause’ and ‘menopausal’ are used herein to generally refer to anything related to the entire transition process, unless otherwise indicated. Hence, the term ‘symptoms associated with menopause’ refers to symptoms caused by the entire transition process, irrespective of the precise period of their occurrence.

It will be understood by those skilled in the art, based on the present teachings, that compositions of the invention can suitably prevent, suppress and/or treat symptoms in each of the respective stages and groups of subjects.

Similar transitions can occur at an earlier age, typically as a consequence of (idiopathic) pathology (‘pre-mature ovarian failure’), surgical intervention (oophorectomy), medical treatments (radiotherapy, chemotherapy) and life-style (smoking). For these physiological processes, the term ‘menopause’ is also used.

In one embodiment, the invention provides compositions as defined herein, for use in a method of treating a subject in need thereof, especially a perimenopausal, menopausal or postmenopausal female subject. In one embodiment, the invention provides compositions as defined herein, for use in a method of preventing, suppressing and/or alleviating one or more symptoms associated with menopause in a subject in need thereof. In one embodiment, the invention provides compositions as defined herein, for use in a method of preventing and/or treating menopausal syndrome in a subject in need thereof.

In one embodiment, the invention provides a method of treating a subject in need thereof, especially a perimenopausal, menopausal or postmenopausal female subject, said method comprising the administration of a composition as defined herein. In one embodiment, the invention provides methods for preventing, suppressing and/or alleviating one or more symptoms associated with menopause in a subject in need thereof. In one embodiment, the invention provides methods for preventing and/or treating menopausal syndrome in a subject in need thereof, comprising the administration of a composition as defined herein.

In one embodiment, the invention provides the use of a composition as defined herein in the manufacture of a pharmaceutical or nutraceutical formulation for use in a method of treating a subject in need thereof, especially a perimenopausal, menopausal or postmenopausal female subject. In one embodiment, the invention provides the use of a composition as defined herein in the manufacture of a pharmaceutical or nutraceutical formulation for use in a method of preventing, suppressing and/or alleviating one or more symptoms associated with menopause. In one embodiment, the invention provides the use of a composition as defined herein in the manufacture of a pharmaceutical or nutraceutical formulation for use in a method of preventing and/or treating menopausal syndrome.

In a preferred embodiment of the compositions for use according to the invention, the one or more symptom(s) associated with menopause are selected from the group consisting of vasomotor symptoms, including hot flushes, palpitations and night sweating; symptoms associated with vulvar and vaginal atrophy, such as vaginitis, vaginal dryness, loss of libido, vaginal pain associated with sexual activity; bleeding associated with sexual activity; dyspareunia, dysuria, vaginal infections, pruritus; dryness, irritation, itching, and/or burning in and around the vaginal area; symptoms associated with nervous system changes, such as nervousness, sleeping problems, insomnia, fatigue, depression, hypomnesis, loss of concentration and dizziness; symptoms resulting from muscoskeletal changes, such as myalgia, muscle pain, joint pain, arthralgia and back pain; symptoms caused by urogenital changes such as urinary frequency or urinary incontinence; and emotional disturbances, such as irritability, depressed mood, nervousness, feelings of sadness, lack of motivation, anxiety, aggressiveness, difficulty concentrating, fatigue, mood changes and tension.

In particularly preferred embodiments of the invention, the method of treating a subject in need thereof, especially a perimenopausal, menopausal or postmenopausal female subject, results in at least one effect selected from:

a decrease in the severity of vasomotor symptoms,

a decrease in the frequency of vasomotor symptoms,

a decrease in the severity of vulvar atrophy, and

a decrease in the severity of vaginal atrophy.

In an embodiment of the invention, the method results in a decrease in the severity of vasomotor symptoms. In an embodiment of the invention, the method results in a decrease in the frequency of vasomotor symptoms. In an embodiment of the invention, the method results in a decrease in the severity of moderate to severe hot flushes. In an embodiment of the invention, the method results in a decrease in the frequency of moderate to severe hot flushes. In an embodiment of the invention, the method results in a decrease in the severity of vulvar and/or vaginal atrophy.

In other embodiments of the invention, the method results in an improvement of the subjects’ overall perception of physiological and/or mental well-being.

In certain embodiments of the invention, the method results in an improvement of the subjects’ perception of one or more menopause-related symptoms, as determined using a menopausal specific questionnaire, such as a questionnaire selected from the group consisting of the Women’s health questionnaire (WHQ), Validated Kupperman Index (Kl), Menopause Rating Scale (MRS), Menopause-Specific Quality of Life Questionnaire (MNQOL), Green Climateric Scale, MenQol, Utian Quality of Life Scale (UQOL). The WHQ, MRS, MNQOL or the Kl are preferably used to measure therapy effectiveness, in particular to monitor a decrease in the severity or in the frequency of one or more symptoms. These instruments measure the intensity or severity as assessed by the individual patient. They provide a measure for the quality of life. The person skilled in the art is familiar with these questionnaires and knows how to use them (Jenabi et al. , Oman medical Journal, Vol. 30, No. 3:151-156, 2015).

In certain embodiments of the invention, the method results in one or more of the effects defined here above within at least 12, 10, 8, 6, 4 or 2 weeks of treatment, based on the compositions and treatment regimens as defined herein.

The subject

As the invention is directed specifically to the treatment of menopause-associated symptoms and/or menopausal syndrome, it will be understood that the subject is a female. The subject can be of any age, e.g. the human subject can be a juvenile, an adolescent, an adult or an elderly subject.

In preferred embodiments of the invention the female human subject suffers from and/or is at risk of suffering from menopause-associated symptoms or from menopausal syndrome, due to the normal aging process. Hence, in preferred embodiments the female human subject is at least 40 years of age, at least 45 years of age, at least 48 years of age, at least 49 years of age, at least 50 years of age, at least 51 years of age, at least 52 years of age, at least 53 years of age, at least 54 years of age or at least 55 years of age. Human subjects treated in accordance with the invention will typically be less than 65 years of age, less than 60 years of age or less than 55 years of age.

In accordance with other embodiments of the invention, the female human subject suffers from and/or is at risk of suffering from menopause-associated symptoms due to a medical condition or pathology. Hence, in preferred embodiments, the female human subjects is a subject suffering from premature ovarian failure.

In accordance with other embodiments of the invention, the female human subject suffers from and/or is at risk of suffering from menopause-associated symptoms as a consequence of surgical intervention. Hence, in preferred embodiments, the female human subjects is a subject that has undergone oophorectomy.

In accordance with other embodiments of the invention, the female human subject suffers from and/or is at risk of suffering from menopause-associated symptoms as a consequence of other medical treatments. Hence, in preferred embodiments the female human subjects is a subject that has undergone and/or is undergoing therapeutic treatments resulting in suppression of circulating estrogen levels, such as a subject that has undergone and/or is undergoing chemotherapy and/or radiotherapy.

In embodiments of the invention, the methods as described herein comprise the step of identifying a subject that is in need of receiving treatment with the compositions of the invention and/or identifying a subject suffering from or at risk of suffering from any of the conditions as described here above and/or a subject meeting any of the above-described physiological characteristics.

In one embodiment such method of treatment is carried out for non-medical or non-therapeutic reasons. Hence, in an embodiment of the invention, the subject is a healthy subject.

Treatment

Typically, the method of the invention entails the repeated administration, to the subject in need thereof, of the compositions in the form of a ‘unit dosage form’, most preferably a coated tablet, as defined herein before, via the oral route. The compositions or unit dosage forms as described herein are administered in an amount and at a frequency effective to achieve one or more of the therapeutic objectives as defined herein before.

In accordance with a preferred embodiment of the invention, the method comprises the administration of hyaluronic acid or a pharmaceutically acceptable salt thereof in a daily dose of 2 mg or more, preferably 20 mg or more, preferably 36 mg or more and/or in an amount of 400 mg or less; preferably 100 mg or less, preferably 40 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of proanthocyanidins in a daily dose of 0.2 mg or more, as determined by gel-permeation chromatography, preferably 2 mg or more, preferably 16 mg or more and/or 400 mg or less, preferably 100 mg or less, preferably 30 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of Nw-methylserotonin in a daily dose of 0.02 mg or more, preferably 0.2 mg or more, preferably 0.1 mg or more and/or 100 mg or less, preferably 30 mg or less, preferably 4 mg or less

In accordance with a preferred embodiment of the invention, the method comprises the administration of vitamin C in a daily dose of 1 mg or more, preferably 50 mg or more, preferably 200 mg or more and/or in an amount of 1500 mg or less, preferably 700 mg or less, preferably 250 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of ginkgolides in a daily dose of 0.001 mg or more,

preferably 0.01 mg or more, preferably 0.1 mg or more and/or in an amount of 100 mg or less, preferably 10 mg or less, preferably 1 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of valerenic acid in a daily dose of 0.001 mg or more, preferably 0.01 mg or more, preferably 0.1 mg or more and/or in an amount of 100 mg or less, preferably 10 mg or less, preferably 1 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of apigenins in a daily dose of 0.1 mg or more, preferably 1 mg or more, preferably 5 mg or more and/or in an amount of 100 mg or less, preferably 50 mg or less, preferably 20 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration a magnesium compound, preferably trimagnesium citrate in a daily dose of 1 mg or more, preferably 50 mg or more, preferably 350 mg or more and/or in an amount of 2000 mg or less, preferably 1000 mg or less, preferably 400 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of Pinus pinaster bark extract in a daily dose of 0.2 mg or more, preferably 2 mg or more, preferably 20 mg or more and/or in an amount of 400 mg or less, preferably 200 mg or less, preferably 40 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of Cimicifuga racemosa root extract in a daily dose of 0.2 mg or more, preferably 2 mg or more, preferably 30 mg or more and/or in an amount of 400 mg or less, preferably 200 mg or less, preferably 50 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of powdered Ginkgo biloba leaf in a daily dose of 1 mg or more, preferably 10 mg or more, preferably 80 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 160 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of Valeriana officinalis root extract in a daily dose of 1 mg or more, preferably 10 mg or more, preferably 70 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 130 mg or less.

In accordance with a preferred embodiment of the invention, the method comprises the administration of Matricaria recutita ray floret extract in a daily dose of 1 mg or more, preferably 10 mg or more, preferably 80 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 140 mg or less.

In preferred embodiments of the invention, the method comprises the daily administration of one or more unit dosage forms according to the present invention.

In particularly preferred embodiments of the invention, the method comprises the daily administration of one day-time unit dosage form as well as the daily administration of one night-time unit dosage form, as defined herein.

As will be apparent from the present teachings, the day-time unit dosage form is specifically adapted to be efficacious in suppressing and/or alleviating menopause associated symptoms and supporting typical day-time activity levels. Hence, in particularly preferred embodiments of the invention, the day-time unit dosage form is administered early during the day for optimal support. In preferred embodiments of the invention the day-time unit dosage form is administered in the morning, preferably the early morning, e.g. during the time interval of 5:00 h am to 11 :00 h am, more preferably during the time interval of 6:00 am to 10:30 am, most preferably during the time interval of 7:00 am to 10:00 am. In preferred embodiments of the invention the day-time unit dosage form is administered within a certain time interval after the subject in need of the treatment wakes up, typically in the morning, after a full night’s sleep, e.g. within 5 hours after the subject wakes up, within 4 hours after the subject wakes up, within 3 hours after the subject wakes up, within 2 hours after the subject wakes up, within 1 hour after the subject wakes up or within 0.5 hour after the subject wakes up.

As will be apparent from the present teachings, the night-time unit dosage form is specifically adapted to be efficacious in suppressing and/or alleviating menopause associated symptoms and supporting night-time rest. Hence, in particularly preferred embodiments of the invention, the night-time unit dosage form is administered by the end of the day for optimal support. In preferred embodiments of the invention the night time unit dosage form is administered in the evening, e.g. during the time interval of 19:00 h pm to 00:00 h pm, more preferably during the time interval of 20:00 pm to 23:30 pm, most preferably during the time interval of 21 :00 pm to 23:00 pm. In preferred embodiments of the invention the night-time unit dosage form is administered within a certain time interval before the subject in need of the treatment goes to sleep or intends to go to sleep, typically in the evening, for a full night’s sleep, e.g. within 5 hours before the subject goes to sleep, within 4 hours before the subject goes to sleep, within 3 hours before the subject goes to sleep, within 2 hours before the subject goes to sleep, within 1 hour before the subject goes to sleep or within 0.5 hour before the subject goes to sleep.

Hence, in a particularly preferred embodiment of the invention the method comprises the oral administration of:

• hyaluronic acid or a pharmaceutically acceptable salt thereof, preferably sodium hyaluronate in an amount of 1 mg or more, preferably 10 mg or more, preferably 18 mg or more and/or in an amount of 200 mg or less; preferably 50 mg or less, preferably 20 mg or less;

• Pinus pinaster bark extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 10 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 20 mg or less, such as 12 mg, 14 mg, 15 mg, 16 mg or 18 mg;

• Cimicifuga racemosa root extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 15 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 25 mg or less, such as 17 mg, 19 mg, 20 mg, 21 mg or 23 mg;

• vitamin C in an amount of 1 mg or more, preferably 50 mg or more, preferably 200 mg or more and/or in an amount of 1500 mg or less, preferably 700 mg or less, preferably 250 mg or less; and

• powdered Ginkgo biloba leaf in an amount of 1 mg or more, preferably 10 mg or more, preferably 80 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 160 mg or less, such as 100 mg, 115 mg, 120 mg, 125 mg or 140 mg;

preferably in a unit dosage form as defined herein, to the female subject in need thereof, in the morning, preferably the early morning and/or within a certain time interval after the subject wakes up as defined in the foregoing.

Furthermore, in a particularly preferred embodiment of the invention the method comprises the oral administration of:

• hyaluronic acid or a pharmaceutically acceptable salt thereof, in an amount of 1 mg or more, preferably 10 mg or more, preferably 18 mg or more and/or in an amount of 200 mg or less; preferably 50 mg or less, preferably 20 mg or less;

• Pinus pinaster bark extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 10 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 20 mg or less, such as 12 mg, 14 mg, 15 mg, 16 mg or 18 mg; • Cimicifuga racemosa root extract in an amount of 0.1 mg or more, preferably 1 mg or more, preferably 15 mg or more and/or in an amount of 200 mg or less, preferably 100 mg or less, preferably 25 mg or less, such as 17 mg, 19 mg, 20 mg, 21 mg or 23 mg;

• Valeriana officinalis root extract in an amount of 1 mg or more, preferably 10 mg or more, preferably 70 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 130 mg or less, such as 80 mg, 95 mg, 100 mg, 105 or 120 mg;

• Matricaria recutita ray floret extract in an amount of 1 mg or more, preferably 10 mg or more, preferably 80 mg or more and/or in an amount of 1000 mg or less, preferably 500 mg or less, preferably 140 mg or less, such as 90 mg, 105 mg, 110 mg, 115 mg or 130 mg; and

• a magnesium compound, preferably trimagnesium citrate in an amount of 1 mg or more, preferably 50 mg or more, preferably 350 mg or more and/or in an amount of 2000 mg or less, preferably 1000 mg or less, preferably 400 mg or less; preferably in a unit dosage form as defined herein, to the female subject in need thereof, in the evening and/or within a certain time interval before the subject in need of the treatment goes to sleep or intends to go to sleep, as defined in the foregoing.

Methods comprising the administration of unit dosage forms in the frequency and at the points in time as described herein are continued for a certain period. In preferred embodiments of the invention, the methods according to the regimens described herein are continued for a period of at least 1 week, at least 2 weeks, at least 3 weeks, at least 4 weeks, at least 1 month, at least 5 weeks, at least 6 weeks, at least 7 weeks, at least 8 weeks, at least 2 months, at least 3 months, at least 4 months, at least 5 months or at least 6 months. In principle this period can extend for as long as the subject experiences (therapeutic) benefits from the method, such as a noticeable reduction of one or more menopause associated symptoms, perceptions of improved energy-levels during the day-time and/or improved night-rest and/or an improvement in overall feelings of well-being. Typically, methods according to the regimens described herein may be continued for periods of up to 5 years, up to 4 years, up to 3 years, or up to 2 years.

MISCELLANEOUS

Unless otherwise defined, all terms used in disclosing the invention, including technical and scientific terms, have the meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. By means of further guidance, term definitions are included to better appreciate the teaching of the present invention.

"A", "an", and "the" as used herein refer to both singular and plural forms unless the context clearly dictates otherwise. By way of example, "a compartment" refers to one or more than one compartment.

"About" as used herein referring to a measurable value such as a parameter, an amount, a temporal duration, and the like, is meant to encompass variations of +/-10% or less, more preferably +/-5% or less, even more preferably +/-1% or less of and from the specified value, in so far such variations are appropriate to perform in the disclosed invention. However, it is to be understood that the value to which the modifier "about" refers is itself also specifically disclosed.

"Comprise," "comprising," "comprises" and "comprised of" as used herein are synonymous with "include", "including", "includes" or "contain", "containing", "contains" and are inclusive or open-ended terms that specifies the presence of what follows e.g. component and do not exclude or preclude the presence of additional, non-recited components, features, element, members, steps, known in the art or disclosed therein.

The recitation of numerical ranges by endpoints includes all numbers and fractions subsumed within that range, as well as the recited endpoints. The skilled person will appreciate that the present invention can incorporate any number of the specific features described above.

Throughout this text, the use of terms in brackets, especially if directly preceding a noun, a verb or adjective, usually means that the term within brackets specifies a possible option or a possible meaning and should thus not be considered limiting. By way of example, the term “(peri)menopause” should be read as “perimenopause and menopause”.

Advantages of the invention will become apparent from the following examples, which are given below as mere illustrations, and are non-limitative.

EXAMPLES

Example 1: day- and night-formulations according to the invention

An example of product according to the most preferred embodiment of the invention comprises a combination of (skye-blue) day-time coated tablets and (navy-blue) night-time coated tablets. The exact composition of the two types of tablets is given in Table 1 below.

Tablets are prepared by dry-blending the respective tablet components to obtain free-flowing dry powder blends. These dry-powder blends are processed into tablet cores by direct compression. A coating liquid is produced by combining the coating components listed in Table 1 , together with an adequate quantity of water that enables spraying of the liquid (in a conventional pan-coating process).

Skye-blue day-time tablets and navy-blue night-time tablets are packaged into separate blister strips. Carton boxes are filled with equal quantities of day-time tablets and night-time tablets (in separate blister strips) together with a package insert with instructions to self-administer one day-time tablet each day (in the morning) and one night-time tablet each day (in the evening).

Table 1 : day-time tablet formulation according to an embodiment of the invention



Table 2: night-time tablet formulation according to an embodiment of the invention


Example 2: Therapeutic use of the day- and night-formulations according to the invention

To confirm efficacy of the treatments according to the present invention a group of 200 perimenopausal women is recruited for a 6-month clinical study. Perimenopausal women between 45-55 years old participate in the study. These

women have experienced disappearance of menses for 3-11 months, after which normal cycles appear again. Hormone levels are used as a further inclusion criterion (serum levels of FSH>30 lU/ml and estrogen E2 levels<20 pg/L). Participants are interviewed for socio-economical status, smoking and dietary habits and examined at first visit for BMI, heart rate, blood pressure, mammography, vaginal sonography and Pap test. Participants visit the study center for screening, enrollment and 1 , 3 and 6 months following start of treatment.

Study participants receiving the treatment of the invention were supplied with sufficient quantities of the product according to example 1 , i.e. the sky-blue day-time tablets and the navy-blue night-time tablets, with instructions to take one day-time tablet every day, shortly after waking up in the morning, and one night-time tablet every day, shortly before going to bed in the evening. Participants in the control group received day-time and night-time placebo tablets (identical in shape, color and appearance as the treatment day-time and night-time tablets), and were instructed to follow the same administration protocol as the treatment group.

Women's Health Questionnaire, designed by Hunter (1992), was used by the subjects to describe level of discomfort and menopause associated symptoms (Hunter M. The women's health questionnaire: a measure of physical and emotional well-being of mid-aged women. Psychology & Health 1992; 7(1 ):45-54). Patients deliver the filled forms at each visit to the study center. Women's Health Questionnaire (WHQ) contains a total of 36 questions related to perimenopausal symptoms: 1 . Somatic symptoms (7 items), 2. depressed mood (8 items), 3. vasomotor symptoms (2 items), 4. memory/concentration (3 items), 5 attractiveness (2 items), 6 anxiety (4 items), 7 sexual behavior (3 items), 8 sleep problems (3 items), 9 menstrual symptoms (4 items). Each symptom is evaluated according to its frequency of occurrence and discomfort level. Points given for frequency: 4: never occurring; 3: sometimes occur; 2: frequently; 1 point: always occur. Points given for discomfort level: 4: no discomfort; 3: little discomfort; 2: clear discomfort; 1 : heavy discomfort. Thus, a higher the score is indicative of a lower level of distress and dysfunction.

Statistical analysis is done using SPSS (Statistical Package for the Social Science).

The perimenopausal symptom scores obtained during the study period reveal that the treatment of the invention results in a lowering of the levels of distress and dysfunction in the treatment group compared to the placebo group. These results

support earlier conclusions concerning treatment efficacy, based on reports of noticeable and meaningful improvements in well-being by perimenopausal women that took the product of the invention for a number of weeks.