U.S. patent application number 09/986509 was filed with the patent office on 2002-03-21 for treatment or prevention of menopausal symptoms and osteoporosis.
This patent application is currently assigned to Novogen, Inc.. Invention is credited to Kelly, Graham E..
Application Number | 20020035074 09/986509 |
Document ID | / |
Family ID | 25645414 |
Filed Date | 2002-03-21 |
United States Patent
Application |
20020035074 |
Kind Code |
A1 |
Kelly, Graham E. |
March 21, 2002 |
Treatment or prevention of menopausal symptoms and osteoporosis
Abstract
There is described a method for the treatment or prevention of
menopausal symptoms or osteoporosis wherein there is administered
to a subject in need of such treatment a therapeutically effective
amount of the isoflavone formononetin, or a method for the
treatment or prevention of menopausal symptoms wherein there is
administered to a subject in need of such treatment a
therapeutically effective amount of the isoflavone daidzein, the
isoflavone being optionally administered with one or more
pharmaceutically acceptable adjuvants, carriers and/or excipients.
Therapeutic uses and compositions/foods are also described,
comprising daidzein or formononetin optionally in association with
one or more pharmaceutically acceptable adjuvants, carriers, food
components and/or excipients.
Inventors: |
Kelly, Graham E.;
(Northbridge, AU) |
Correspondence
Address: |
FINNEGAN, HENDERSON, FARABOW, GARRETT &
DUNNER LLP
1300 I STREET, NW
WASHINGTON
DC
20005
US
|
Assignee: |
Novogen, Inc.
|
Family ID: |
25645414 |
Appl. No.: |
09/986509 |
Filed: |
November 9, 2001 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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09986509 |
Nov 9, 2001 |
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09077590 |
Jun 2, 1998 |
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09077590 |
Jun 2, 1998 |
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PCT/AU98/00313 |
May 1, 1998 |
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Current U.S.
Class: |
514/27 ;
514/456 |
Current CPC
Class: |
A61P 5/24 20180101; A61P
5/30 20180101; A61K 31/35 20130101; Y10S 514/899 20130101; A61K
31/35 20130101; A61P 15/12 20180101; A61K 31/353 20130101; A61K
2300/00 20130101; A61K 31/352 20130101; A61P 19/10 20180101 |
Class at
Publication: |
514/27 ;
514/456 |
International
Class: |
A61K 031/7048; A61K
031/353 |
Foreign Application Data
Date |
Code |
Application Number |
May 1, 1997 |
AU |
P06568 |
Dec 8, 1997 |
AU |
PP0814 |
Claims
1. A method for the treatment or prevention of menopausal symptoms
or osteoporosis wherein there is administered to a subject in need
of such treatment a therapeutically effective amount of the
isoflavone formononetin, or a method for the treatment or
prevention of menopausal symptoms wherein there is administered to
a subject in need of such treatment a therapeutically effective
amount of the isoflavone daidzein, the isoflavone being optionally
administered with one or more pharmaceutically acceptable
adjuvants, carriers and/or excipients.
2. A method according to claim 1 for the treatment or prevention of
osteoporosis.
3. A method according to claim 2 wherein formononetin is
administered to a subject.
4. A method according to claim 1 for the treatment or prevention of
menopausal symptoms.
5. A method according to claim 4 wherein formononetin or daidzein
is administered to a patient.
6. A method according to claim 1, wherein the isoflavone is present
in an amount of at least 90% (w/w) as against any other
isoflavones.
7. A method according to claim 1, wherein the isoflavone is
administered at least once per day.
8. A method according to claim 1, wherein the isoflavone is
administered in an amount from about 5 mg/day through to about 400
mg/day.
9. A method according to claim 1, wherein the isoflavone is
administered from 1 to 6 times in a 24 hour period so as to give a
daily dosage of from about 5 to about 400 mg.
10. A method according to claim 1 wherein maid isoflavone is
derived from a plant extract comprising daidzein and/or
formononetin substantially unaccompanied by other isoflavones.
11. A method according to claim 1 for the treatment or prevention
of osteoporosis wherein a combination of formononetin and daidzein
is administered to a patient.
12. A method according to claim 1 wherein the isoflavone is
substantially unaccompanied by other isoflavones.
13. A method according to claim 1 wherein daidzein is replaced by a
daidzein metabolite selected from equol, o-desmethylangolensin
(ODMA), dehydroequol, 2-dehydro ODMA, 6-hydroxy-ODMA,
dihydrodaidzein and tetra-hydrodaidzein.
14. A pharmaceutical composition for the treatment or prevention of
menopausal symptoms or osteoporosis where the said composition
comprises the isoflavone formononetin or a pharmaceutical
composition for the treatment or prevention of menopause wherein
said composition comprises the isoflavone daidzein, together with
one or more pharmaceutically acceptable adjuvants, carriers and/or
excipients.
15. A composition according to claim 14, which is a composition for
the treatment or prevention of osteoporosis.
16. A composition according to claim 15 which includes the
isoflavone formononetin.
17. A composition according to claim 14 which is a composition for
the treatment and/or prevention of acute menopausal symptoms.
18. A composition according to claim 17 wherein the isoflavone is
daidzein or formononetin.
19. A composition according to claim 17 which includes a
combination of daidzein and formononetin.
20. A composition according to claim 14 which comprises from 5 to
400 mg of isoflavone.
21. A composition according to claim 14 wherein daidzein is
replaced by a daidzein metabolite selected from equol,
o-desmethylangolensin (ODMA), dehydroequol, 2-dehydro-ODMA,
6-hydroxy-ODMA, dihydrodaidzein and tetra-hydrodaidzein.
22. Use of the isoflavone formononetin in the treatment or
prevention of menopausal symptoms or osteoporosis, or use of the
isoflavone daidzein in the treatment or prevention of menopausal
symptoms, the isoflavone being optionally administered with one or
more pharmaceutically acceptable adjuvants, carriers, and/or
excipients.
23. Use according to claim 22, wherein the isoflavone is
administered in an amount from about 5 mg to about 400 mg.
24. Use according to claim 22 for the treatment or prevention of
osteoporosis wherein the isoflavone is formononetin.
25. Use according to claim 22 for the treatment or menopausal
symptoms wherein the isoflavone is formononetin or daidzein.
26. Use according to claim 25 wherein a combination of daidzein and
formononetin are administered to a subject.
27. Use of formononetin, for the manufacture of a medicament for
the treatment or prevention of menopausal symptoms or osteoporosis,
or use of daidzein or formononetin for the manufacture of a
medicament for the treatment or prevention of osteoporosis.
28. Use according to claim 22 wherein the daidzein or formononetin
is administered from 1 to 6 times in a 24 hour period so as to give
a daily dosage of about 5 mg to about 400 mg.
29. Use according to claim 27 wherein the isoflavone daidzein is
combined with formononetin.
30. An agent for the treatment or prevention of menopausal symptoms
or osteoporosis which comprises formononetin, or an agent for the
treatment or prevention of menopausal symptoms which comprises
daidzein, optionally in association with one or more
pharmaceutically acceptable adjuvants, carriers and/or
excipients.
31. A method or composition according to claim 1 or 14, wherein the
isoflavone daidzein is in the aglycone, glycoside, malonyl or
acetyl form.
Description
[0001] This invention relates to compositions, therapeutic uses and
methods of treatment or prevention of menopausal symptoms and
osteoporosis.
[0002] Menopausal symptoms and osteoporosis are significant
scourges in the female population, generally affecting many women
in later life.
[0003] Menopausal symptoms are very well known and are described,
for example, by Greene, J. G. and Cooke, D. J. (1980) British
Journal of Psychiatry Volume 136, 486-491 (incorporated herein by
reference). Hot flushes are one of the principal menopausal
symptoms which are uncomfortable and irritating. Greene and Cooke
have developed a score in order to measure menopausal symptoms in
women. This score is approved by the US Department of Health and
widely used in the medical community. The indicators of menopausal
symptoms according to Greene and Cooke comprise hot flushes,
sweating at night, heart beating quickly or strongly, feelings of
tension or nervousness, difficulty in sleeping, excitability,
attacks of panic, difficulties in concentrating, feelings of
tiredness or lack of energy, unhappiness or depression, crying
spells, irritability, feelings of dizziness or faintness, pressure
or tightness in head or body, parts of the body feeling numb or
tingling, dry vagina and/or dry mouth, headaches, muscle and joint
pains, loss of feeling in hands or feet, breathing difficulties,
and loss of interest in sex.
[0004] The peri-menopausal stage of life in women is associated
with a fall in blood levels of the three major
estrogens--estradiol, estrone and estriol--which occurs naturally
in women usually between 45-55 years of age. The primary or acute
menopause symptoms which effect menopausal women include hot
flushes and night sweats. These are associated with often
dramatically increased blood perfusion of the skin producing
discomfort and sweating.
[0005] The precise mechanism of these symptoms is unknown but
generally is thought to represent disturbance to normal homeostatic
mechanisms controlling vasomotor activity and thermoregulation.
[0006] The fact that treatment and/or prevention with replacement
estrogens usually relieves the symptoms (so-called estrogen
replacement therapies) establishes the link between these symptoms
and an estrogen deficiency. The menopausal stage of life is
associated with a wide range of other acute symptoms as described
above and these symptoms are generally estrogen-responsive.
[0007] Osteoporosis is believed to affect one third to one half of
all post-menopausal women. In the United States it is been reported
flat annually 500,000 bone fractures occur as a result of
osteoporosis. It is further reported that nearly one third of women
over 65 will suffer at least one bone fracture resulting from
osteoporotic bone weakening. Increased calcium intake and other
approaches are suggested to have some effect. However, the
widespread effects of osteoporosis indicates effective approaches
for prevention/treatment have not yet arisen.
[0008] It has previously been thought that reduction in endogenous
estrogen levels which occurs prior to menopause causes or
contributes to the symptoms of menopause, as well as
post-menopausal osteoporosis.
[0009] Isoflavones, being plant chemicals which occur largely in
members of Leguminosae, display a range of biological functions
which have suggested they may be useful in treating a host of
medical conditions.
[0010] A small subgroup of isoflavones (comprising daidzein,
genistein, biochanin, and formononetin and) is distinguished by
their ability to bind to estrogen receptors on animal (including
human) cells. This is due to the close similarity of the steric
structure of the diphenolic rings of isoflavones with the steroidal
ring structure of estrogens such as estradiol, estrone and estriol.
Although having substantially lower binding affinity to the
receptor compared to steroidal estrogens, estrogenic isoflavones
are weakly estrogenic. This group of five isoflavones have the most
basic diphenolic structure possible in contrast to the relatively
more complex structures of other isoflavonoid compounds. This
simplicity of structure and its close proximity in shape to the
steroidal ring structure of estrogenic hormones is believed to
grant these compounds their estrogenicity. This group also exhibits
a range of biological functions in animal cells which appear to be
independent of the estrogen receptor and these include antioxidant,
diuretic, anti-spasmolytic and anti cancer effects. These
interesting functions with their potential therapeutic benefits has
brought this particular group of isoflavones to the attention of
medical researchers in recent years.
[0011] In the plant, the isoflavones can occur in a variety of
forms--(i) in the basic form, (ii) in a malonyl form, and (iii) in
an acetyl form; the isoflavones are biologically active in each of
these forms. The naturally-occurring state for each of these forms
is as a glycoside, being bound to a sugar moiety such as glucose to
produce a water-soluble form. In this form, the isoflavone has
enhanced stability to degradative factors such as heat, oxidation
and ultraviolet irradiation. This water-soluble form also permits
transport of the isoflavone both around the plant and
intra-cellularly. At the intra-cellular site of biochemical
function of the isoflavone, an intra-cellular glucoside enzyme
cleaves the sugar moiety, leaving the more biologically active, but
water-insoluble, aglucone form.
[0012] When ingested in the diet, the isoflavones undergo varying
degrees of metabolism within the gut, within the gut wall, and
within the liver before entering the parenteral bloodstream to
exert their biological effects. The first metabolic process is the
hydrolysis of the glucosidic form to the aglucone form. This occurs
as a result both of low pH from gastric acid and of the action of
.beta.-glycosidase enzyme activity within bowel bacteria.
[0013] Some of the aglucone isoflavones are absorbed intact and in
passing through the gut wall are believed to be glucoronated or
sulphonated as per steroidal compounds. The bulk of isoflavones are
fermented within colonic bacteria. One of the fermentation
processes is to demethylate isoflavones (e.g. formononetin gives
daidzein and biochanin gives genistein on demethylation). In
another series of fermentation steps, daidzein and genistein are
converted to a range of end-products including equol, dehydroequol,
O-desmethylangolensin (ODMA), 6-hydroxy-ODMA, 2-dehydro-ODMA,
dihydrodaidzein, tetra-hydrodaidzein and dihydrogenistein. The
liver is capable of further demethylation of isoflavones such as
formononetin and biochanin to the more basic daidzein and genistein
structures. The isoflavones and their metabolites and derivatives
circulate freely within the body and are excreted primarily in the
urine with smaller amounts in the faeces.
[0014] The possibility that dietary estrogenic isoflavones may have
some therapeutic benefit in acute menopausal symptoms was suggested
by the observation that Japanese women who typically have much
higher dietary levels of isoflavones (mostly derived from soya)
compared to women in Western countries have a reportedly lower
incidence of acute menopausal syndrome symptoms such as hot
flushes. This has led to somewhat speculative claims of therapeutic
benefit of the isoflavones from the group daidzein, formononetin,
biochanin and genistein in the treatment and/or prevention of acute
menopausal syndrome symptoms (U.S. Pat. No. 5,498,631--Gorbach et
al).
[0015] Gorbach analysed urinary isoflavone excretion in Japanese
subjects who consumed a traditional Japanese low-fat diet. The
presence of estrogenic isoflavones in the urine of the women, men
and children studied suggested to Gorbach that the isoflavones
produced a therapeutic effect. The obvious flaws in this study,
namely that in a diet with high isoflavone intake significant
urinary out-put of isoflavones would be expected, and the huge
number of biochemically active species in any diet make it
impossible to ascribe biological effects to any particular
component or components indicate that Gorbach's claims do not stand
scrutiny. The fact that a community with a particular health
profile happens to have a high dietary level (and high body levels)
of a certain plant component in no way establishes cause and
effect. This is only achieved through appropriately conducted
clinical studies where well accepted scientific principles can be
applied.
[0016] Clinical and other studies done to date in this area are
highly equivocal, with no consistent effect reported. Reported
studies have involved the challenge of peri-menopausal women either
with whole foodstuffs (such as soyflour) containing isoflavones or
with extracts of soya or other legumes, often together with other
agents such as vitamins, or isoflavones together with estrogen
and/or vitamins and various minerals. It is to be noted that soy
does not contain the isoflavones formononetin and biochanin. Even
when a positive clinical effect has been obtained, it has been with
a mixture of a plurality of isoflavones, as well as a wide range of
other unidentified dietary components and other biologically active
components--it is known for example that other compounds present in
legumes such as flavonoids (e.g. quercetin, luteolin, kaempferol
and lignans) also are estrogenic and it is also likely that among
the other 700 or so isoflavonoids present in the Leguminosae family
there are as yet unidentified isoflavonoids with estrogenic
activity.
[0017] Gorbach suggests that isoflavanoids bind to estrogen
receptors, exerting an estrogenic effect in menopausal women.
However, it is acknowledged generally that no direct evidence
exists to link the presence of isoflavones with a therapeutic
effect in treatment and/or prevention of acute menopausal syndrome
symptoms. But even if such a link could be inferred from the
current epidemiological and clinical studies, which it can not, the
question remains what, if any, therapeutic effect is the result of
a collective effect of the estrogenic isoflavones--daidzein,
formononetin, biochanin, genistein. All four isoflavones are
estrogenic, but they have quite different estrogenic potencies. The
relative estrogenicity of genistein, daidzein, formononetin,
biochanin is 1.3, 0.09, 0.01, 0.07 respectively (relative to
17.beta.-estradiol 100). Hence formononetin and biochanin have
negligible estrogenic activity. On this basis, and given the
relative proportions of daidzein and genistein in the blood of
Japanese maintaining a typical Japanese diet, it might be inferred
that genistein potentially is the most potent isoflavone as far as
acute menopause syndrome symptoms are concerned.
[0018] Estrogenic isoflavones have also been identified as possible
therapeutic compounds in the treatment and/or prevention of
osteoporosis. Asian populations consuming large amounts of
phytoestrogen-rich soybeans and vegetables appear to be protected
to a greater extent than western populations from the problems
associated with osteoporosis. These observations are by no means
clear, and are contradicted in a number of studies.
[0019] Fujita and Fukase (Proc. Soc. Exp. Biol. Med. 200(2) 149-5,
1992) indicates that in osteoporosis analysis between Japanese and
US populations diet is not of particular significance, with bone
mass being very similar in both populations. Instead they suggest
the outcomes of osteoporosis are more likely associated with
lifestyle affecting muscle development and motor control. Arjanandi
et al (American Institute of Nutrition, p161-167, 1995) indicates
that any protective effects of soy on bone is associated with soy
protein. Hunt et al (Am. J. Clin. Nutr., p517-523, 1989) indicate
that there is no appreciable difference in bone density between
elderly menopausal omnivores, and elderly menopausal vegetarians
whose diet included isoflavone rich plant materials.
[0020] Published European Patent Application No. 0135172 (Takeda,
published May 27, 1985) discloses methods for the treatment of
osteoporosis by administration of 7,4-dihydroxy isoflavone
(daidzein) relying on its estrogenic activity. This finding is
inconsistent with the biological studies reported above. Moreover,
Tobe et at (Biosci. Biotech. Biochem. 61(2) 370-371, 1997) show
that daidzein stimulates bone resorption, that is bone breakdown,
and would be contra-indicated for treating osteoporosis, as it
would worsen an existing condition, and possibly pre-dispose a non
affected person to osteoporosis.
[0021] Against the foregoing background, the present invention is
predicated upon our surprising finding that, within the framework
of what was conjectured regarding the treatment/prevention of
menopausal symptoms and osteoporosis, formononetin may be used to
treat both menopausal symptoms and osteoporosis, and daidzein may
be used in the treatment of menopausal symptoms. Our findings
indicate that formononetin has pronounced clinical activity in the
treatment and/or prevention of menopausal symptoms and in the
treatment and/or prevention of osteoporosis as does daidzein in the
treatment/prevention of menopausal symptoms. This is highly
unexpected given the negligible estrogenic effect of formononetin,
the bone resorption activity of daidzein and the established view
that formononetin was very rapidly metabolised to daidzein in the
gut.
SUMMARY OF THE INVENTION
[0022] There is provided in a first aspect of this invention a
method for the treatment or prevention of menopausal symptoms or
osteoporosis wherein there is administered to a subject in need of
such treatment a therapeutically effective amount of the isoflavone
formononetin, or a method for the treatment or prevention of
menopausal symptoms wherein there is administered to a subject in
need of such treatment a therapeutically effective amount of the
isoflavone daidzein, the isoflavone being optionally administered
with one or more pharmaceutically acceptable adjuvants, carriers
and/or excipients.
[0023] In another aspect of the invention there is provided a
pharmaceutical composition for the treatment or prevention of
menopausal symptoms or osteoporosis wherein the said composition
comprises the isoflavone formononetin or a pharmaceutical
composition for the treatment or prevention of menopause wherein
said composition comprises the isoflavone daidzein together with
one or more pharmaceutically acceptable adjuvants, carriers and/or
excipients.
[0024] In another aspect of the invention there is provided use of
the isoflavone formononetin in the treatment or prevention of
menopausal symptoms or osteoporosis, or use of the isoflavone
daidzein in the treatment or prevention of menopausal symptoms, the
isoflavone being optionally administered with one or more
pharmaceutically acceptable adjuvants, carriers, and/or
excipients.
[0025] In a further aspect of the invention there is provided an
agent for the treatment or prevention of menopausal symptoms or
osteoporosis, or an agent for the treatment or prevention of
menopausal symptoms which comprises daidzein optionally in
association with one or more pharmaceutically acceptable adjuvants,
carriers and/or excipients.
DETAILED DESCRIPTION
[0026] Throughout this specification and the appended claims,
unless the context requires otherwise, the word "comprise", or
variations such as "comprises" or "comprising" or "include" or
"including", will be understood to imply the inclusion of a sated
element or integer or group of elements or integers but not the
exclusion of any other element or integer or group of elements or
integers.
[0027] The present invention provides in a first aspect a method
for the treatment or prevention of menopausal symptoms or
osteoporosis wherein there is administered to a subject in need of
such treatment a therapeutically effective amount of the isoflavone
formononetin, or a method for the treatment or prevention of
menopausal symptoms wherein there is administered to a subject in
need of such treatment a therapeutically effective amount of the
isoflavone daidzein, the isoflavone being optionally administered
with one or more pharmaceutically acceptable adjuvants, carriers
and/or excipients.
[0028] It is believed that the invention described represents a
substantial breakthrough in the field of treatment and/or
prevention of menopausal symptoms and osteoporosis. The
administration to subjects of plant extracts containing a range of
isoflavones may be unpleasant in the sense that the extract may not
be particularly palatable, and/or may contain a host of ill-defined
compounds which may affect disadvantageous biological activity.
Wilcox et al (British Medical Journal (1990) 301: 905) have
reported increases in vaginal cell proliferation amongst
post-menopausal women consuming soybean phytoestrogens for six
weeks. In addition Markiewicz et al (J. Steroid Biochem. (1993) 45:
399) have shown experimentally that the soy isoflavone genistein
exhibited an estrogen effect on endometrial cancer cells, that is,
potentiated cancer cell growth in this cell type. Such reports
raise questions about the safely of comparatively high doses of
genistein. The present invention provides the treatment or
prevention of menopausal symptoms and osteoporosis without any side
effects caused by uncharacterised biologically active plant
materials (such as coumesterols), or other disadvantageous
effects.
[0029] The menopausal symptoms which may be treated according to
the method of this invention are those described by Greene, J. G.
and Cooke, D. J. (1980) British Journal of Psychiatry Volume 136,
486-491. Preferably the menopausal symptoms treated or prevented
according to the present invention are hot sweats and night time
sweats, more particularly hot sweats. Having said this, the method
of the invention is applicable to the treatment and/or prevention
of other symptoms of menopause as previously described. The
isoflavone formononetin is of the formula (I): 1
[0030] Although it was previously thought that formononetin was
almost immediately metabolised demethylated) to daidzein upon
administration to a subject, the present inventors have found that
formononetin persists in the blood stream for a considerable the
(having a half life of generally about 20 hours).
[0031] The isoflavone daidzein is of the formula (II): 2
[0032] Formononetin or daidzein arm preferably administered to a
subject substantially unaccompanied by other isoflavones. By this
is meant that any composition or preparations may contain minor
amounts of other isoflavones, in the order of 10% (w/w) or less.
Preferably the formononetin or daidzein represents at least 90% of
isoflavone content, more preferably 95%, even more preferably 98%
or more. Genistein, if presents is in amounts of about 5% or less,
more preferably less than 1% (w/w) with regard to isoflavone
content. It is recognised by regulatory agencies that an isoflavone
content in the order of 95 % of total isoflavones represents
effective purity.
[0033] In the treatment of menopausal symptoms formononetin may be
administered in combination with daidzein, for example from a ratio
of 1:10 to 10:1.
[0034] Daidzein metabolites may be used in place of daidzein in the
various embodiments of this invention. These metabolites include
equol, o-desmethylangolensin (ODMA), dehydroequol, 2-dehydro-ODMA,
6-hydroxy-ODMA, dihydrodaidzein and tetra-hydrodaidzein (which
collectively may be referred to herein as daidzein metabolites).
Accordingly, in a further aspect this invention extends to a method
for the treatment or prevention of menopausal symptoms or
osteoporosis wherein, there is administered, to a subject in need
of such treatment and/or prevention, formononetin or a daidzein
metabolite, optionally administered with one or more
pharmaceutically acceptable adjuvants, carriers and/or excipients.
The formononetin or daidzein metabolites may be administered in a
form substantially unaccompanied by other isoflavones.
[0035] Daidzein and/or formononetin compositions or preparations
are administered in an amount, and under a dosage regime which
gives relief to menopausal symptoms or osteoporosis. With regard to
menopausal symptoms this can be readily determined by the subject
who is being treated, or by their physician. Generally, it is found
that prevention or therapy of menopausal symptoms and osteoporosis
results from daily administration of formononetin such as from one
to six times in a 24 hour period, as does the treatment or
prevention of menopausal symptoms with daidzein, so as to give a
daily dose of the isoflavone in an amount from about 5 mg to about
400 mg per day (this dosage range may be referred to as the
"effective amount").
[0036] Formononetin and daidzein may be prepared by synthesising
the compounds by conventional chemical synthetic techniques as are
well known in the art, or by purification from extracts of plants
of the genus Leguminosae, particularly from soy (such as from soy
flour, soy hypocotyls) and clover (such as red clover, and
subterranean clover) such as to form a formononetin or daidzein
composition or preparation.
[0037] Compositions/preparations administered to subjects for the
treating and/or prevention of, or for reducing the predispositon
to, menopausal symptoms or osteoporosis may comprise in addition to
the specific isoflavones previously mentioned formononetin
optionally administered with one or more pharmaceutically
acceptable adjuvants, carriers and/or excipients, so as to form a
composition or preparation. Pharmaceutically acceptable adjuvants,
carriers and/or excipients, and the like, are well known in the
art, for example as described in the Handbook of Pharmaceutical
Excipients, second edition, American Pharmaceutical Association,
1994 (incorporated herein by reference). Daidzein or formononetin
may be administered in the form of tablets, capsules, powders for
reconstitution, syrups, foods (such as food bars, biscuits, snack
foods and other standard food forms well known in the art) or in
drink formulations. Drinks may contain flavouring, buffers and the
like.
[0038] In the method of this invention calcium may be
co-administered (that is before at the same time or after the
isoflavones previously mentioned), for example as a separate
tablet, or as part of a suitable dosage form.
[0039] In a further aspect of this invention there is provided a
pharmaceutical composition for the treatment or prevention of
menopausal symptoms or osteoporosis wherein the said composition
comprises the isoflavone formononetin or a pharmaceutical
composition for the treatment or prevention of menopause wherein
said composition comprises the isoflavone daidzein, together with
one or more pharmaceutically acceptable adjuvants, carriers and/or
excipients. As mentioned above, pharmaceutically acceptable
adjuvants, carriers and/or excipients are well known in the art.
Examples include compositions according to the present invention
may include one or more pharmaceutically acceptable carriers. The
carriers are selected so as to be acceptable in the sense of being
ingredients in the composition and must not be deleterious to the
patient. The carriers may be solid or a liquid, or both, and may be
formulated with the extract as a unit-dose, for example a tablet,
which may contain from 0.5% to 59% by weight of the active compound
or up to 100% by weight to the active compound. Compositions may be
prepared by any of the well known techniques of pharmacy, for
example admixing the components, optionally including excipients,
diluents (for example water) and auxiliaries as are well known in
the pharmaceutical field.
[0040] The compositions of the invention include those suitable for
oral, rectal, optical, buccal (for example sublingual), parental
(for example subcutaneous, intramuscular, intradermal and
intravenous) and transdermal administration. The most suitable
route in any given case will depend on the nature and severity of
the condition being treated and the state of the patient.
[0041] Compositions suitable for oral administration may be
presented in discrete units, such as capsules, cachets, lozenges,
or tablets, each containing a predetermined amount of the extract;
as a powder or granules; as a solution or a suspension in an
aqueous or non-aqueous liquid; or as an oil-in-water or
water-in-oil emulsion. Such compositions may be prepared by any
suitable method of pharmacy which includes the step of bringing
into association the active isoflavone and one or more suitable
carriers (which may contain one or more accessory ingredients as
noted above). In general the compositions of the invention are
prepared by uniformly and intimately admixing the isoflavone with a
liquid or finely divided solid carrier, or both, and then, if
necessary, shaping the resulting mixture. For example, a tablet may
be prepared by comprising or moulding a powder or granules
containing the extract, optionally with one or more accessory
ingredients. Compressed tables may be prepared by compressing in a
suitable machine, the extras in the form of a powder or granules
optionally mixed with a binder, lubricant, inert diluents, and/or
surface active/dispersing agent(s). Moulded tablets may be made by
moulding, in a suitable machine, the powdered compound moistened
with an inert liquid binder.
[0042] Suitable carriers may be fillers, such as sugars, for
example lactose, saccharose, mannitol or sorbitol, cellulose
preparations and/or calcium phosphates, for example tricalcium
phosphate or calcium hydrogen phosphate, and also binders, such as
starch pastes using, for example, corn, wheat, rice or potato
starch, gelatin, tragacanth, methylceullose and/or
polyvinylpyrrolidone, and, if desired, disintegrators, such as the
above-mentioned starches, also carboxymethyl starch, cross linked
polyvinyl pyrrolidone, agar or alginic acid or a salt thereof, such
as sodium alginate. Excipients may be flow conditioners and
lubricants, for example silicic acid, talc, stearic acid or salts
thereof, such as magnesium or calcium stearate, and/or polyethylene
glycol. Drage cores are provided with suitable, optionally enteric,
coatings, there being used, inter alia, concentrated sugar
solutions which may comprise gum arabic, talc,
polyvinylpyrrolidone, polyethylene glycol and/or titanium dioxide,
or coating solutions in suitable organic solvents or solvent
mixtures, or, for the preparation of enteric coatings, solutions of
suitable cellulose preparations, such as acetylcellulose phthalate
or hydroxypropylmethylcellulose phthalate. Dyes or pigments may be
added to the tablets or drage coatings, for example for
identification purposes or to indicate different doses of active
ingredients.
[0043] Other orally administrable pharmaceutical compositions are
dry-filled capsules made, for example, of gelatin, and soft, sealed
capsules made of gelatin and a plasticiser, such as glycerol or
sorbitol. The dry-filled capsules may comprise the extracts in the
form of granules, for example in admixture with fillers, such as
lactose, binders, such as starches, and/or glicants, such as talc
or magnesium stearate, and, where appropriate, stabilisers. In soft
capsules, the extract is preferably dissolved or suspended in
suitable liquids, such as fatty oils, paraffin oil or liquid
polyethylene glycols, to which stabilisers may also be added.
[0044] Formulations suitable for buccal (sublingual) administration
include lozenges comprising the extracts in a flavoured base,
usually sucrose and acacia or tragacanth; and pastilles comprising
the compound in an inert base such as gelatin and glycerin or
sucrose and acacia.
[0045] Formulations suitable for rectal administration are
preferably presented as unit dose suppositories. These may be
prepared by admixing the isoflavones with one or more conventional
solid carriers, for example cocoa butter, and then shaping the
resulting mixture.
[0046] Compositions may include calcium or other active agents
suggested to provide some amelioration of osteoporosis or symptoms
of menopause.
[0047] Pharmaceutical compositions generally comprise from about 5
mg to about 400 mg of the isoflavone/s and may be administered one
or more times per day.
[0048] In a further aspect of this invention there is provided use
of the isoflavone formononetin in the treatment or prevention of
menopausal symptoms or osteoporosis, or use of the isoflavone
daidzein in the treatment or prevention of menopausal symptoms, the
isoflavone being optionally administered with one or more
pharmaceutically acceptable adjuvants, carriers, and/or
excipients.
[0049] In another aspect of the invention there is provided use, of
formononetin, for the manufacture of a medicament for the treatment
or prevention of menopausal symptoms or osteoporosis, or use of
daidzein or formononetin for the manufacture of a medicament for
the treatment or prevention of osteoporosis. Generally, the
isoflavone is provided in the form of a medicament in association
with one or more pharmaceutically adjuvants, carriers and/or
excipients. Daidzein and/or formononetin may be conveniently
blended as a dry powder with other components and formed into
appropriate dosage forms. Such medicaments generally comprise from
about 5 mg to about 400 mg of isoflavone.
[0050] According to a still further aspect of the invention there
is provided an agent for the treatment or prevention of menopausal
symptoms or osteoporosis, or an agent for the treatment or
prevention of menopausal symptoms which comprises daidzein
optionally in association with one or more pharmaceutically
acceptable adjuvants, carriers and/or excipients.
[0051] Daidzein and/or formononetin may be administered in the form
of a dietary product, as mentioned above, for example in a
palatable food carrier such as a confectionary bar, biscuit, cereal
or beverage.
[0052] The methods and compositions of the present invention do not
include the use of estrogens, or components such as licorice,
cholecalciferol and vitamin E. Estrogen administration has many
side effects such as genital bleeding and hepatic disorders.
Licorice has vasoactive activity and may exacerbate menopause
symptoms or osteoporosis- Cholecalciferol and vitamin E are also
disadvantageous.
[0053] The foregoing description in relation to daidzein, applies
to the daidzein metabolites equol, o-desmethylangolensin (ODMA),
dehydroequol, 2-dehydro-ODMA, 6-hydroxy-ODMA, dihydrodaidzein and
tetra-hydrodaidzein, as well as combinations of the above
species.
[0054] Hence daidzein may be replaced by one or more of these
metabolites. Daidzein or formononetin may also take the form of
aglycones, glycosides, malonyl or acetyl derivatives.
[0055] This invention will now be described with reference to the
following non-limiting examples.
EXAMPLE 1
[0056] Peri-menopausal women with symptoms of acute menopausal
syndrome including at least an average of 3 hot flush episodes per
day were treated with a preparation containing a concentrated
amount of daidzein, formononetin, genistein and biochanin prepared
according to the method described in published Australian patent
application 40523/93 (incorporated herein by reference). The
preparation contained 100 mg isoflavones containing biochanin and
formononetin (in the ratio of 1.8:1). Urine samples were collected
from each woman at the start of the test and then again at the end
of the trial and the levels of particular isoflavones in the urine
(as a marker of total body isoflavone ratios) then correlated with
the degree of reduction in the incidence of hot flushes per day
over the course of the study. Daidzein and genistein were detected
in the urine of all test subjects but the levels were highly
variable, ranging from barely detectable to relatively high.
Formononetin and biochanin levels ranged between undetectable and
significant. Genistein levels showed no correlation with
therapeutic response; daidzein levels correlated well with
therapeutic response indicating potential therapeutic utility of
daidzein and formononetin.
EXAMPLE 2
[0057] Daidzein substantially free of other isoflavones was
prepared from soy by the following method. 1 kg of defatted
soyflour (readily available from many commercial sources) was added
to 10 L of water containing 50 g of glucan hydrolase enzyme
(BioFeed Beta CT, Novo Nordisk, Denmark). To this suspension 5 L of
ethyl acetate is then added. This mixture is mixed vigorously using
a high pressure pump for 3 hours so that it forms an emulsion. This
mixing ensures effective contact between the aqueous phase and the
miscelles of ethyl acetate so that the enzymatically hydrolysed
aglucone forms of the isoflavones move from the aqueous to the
organic solvent phase. The three phases (soyflour, aqueous, ethyl
acetate) are separated by centrifugation in a swing bucket
centrifuge at 2000 g for 30 minutes. The upper phase comprising
ethyl acetate is aspirated, 200 ml of water added, and then placed
into a rotary evaporator at 75.degree. C. under a weak vacuum. Upon
removal of the ethyl acetate, the residual aqueous phase containing
the isoflavones is extracted once with 500 ml of hexane to remove
oils and fats, and then twice with 500 ml of octanol which
selectively removes genistein. The aqueous phase then was taken to
dryness overnight in an oven at 80.degree. C. This material was
shown by high pressure liquid chromatographic analysis to comprise
65% isoflavones comprising daidzein (95%): genistein(5%):
formononetin(0%): biochanin(0%). This material was mixed with
standard carriers/excipients and tableted to give 500 mg tablets
containing 25 mg daidzein. In this particular example, the daidzein
was tableted with equivalent (w/w) amounts of microcrystalline
cellulose, calcium hydrogen phosphate, magnesium stearate and
anhydrous colloidal silica.
[0058] Formononetin substantially free of other isoflavones was
prepared from clover by enzyme/solvent as above. Formononetin was
recovered as a purity level greater than 95% by chromatographic
analysis or by HPLC.
EXAMPLE 3
[0059] The dried end product of the first part of Example 2 above
can be used to further concentrate genistein or daidzein
with/without. 3 kg of this material is mixed with 1000 L of an
organic solvent such as acetonc, chloroform or octanone, but
preferably acetone for reasons of safety and cost. Each of these 3
solvents has been shown by the inventors to have high affinity for
genistein but not daidzein. The mixture is stirred continuously at
room temperature for between 1-24 hours but preferably 2 hours
during which time a large amount (approximately 75%) of the
genistein transfers into the solvent phase. The mixture is allowed
to settle for about 2 hours, the solvent is separated from the
residue (Sample 2) and transferred to a still for evaporation.
Sample 2 material preferably is extracted with acetone a further
1-5 times (preferably 4 times). This material typically contains
76% daidzein, 1% genistein, 0.5% glycetein, with the remainder
comprising residual lipid soluble material such as short chain
fatty acids. On a w/w basis compared with the other isoflavones,
this preparation contains 98% (w/w) daidzein. Daidzein may be
purified to 95% (w/w) of total material or more purity by
preparative purification regimens such as preparative HPLC. A
daidzein preparation according to this example is tableted with
conventional inert excipients according to Example 3 to give a 200
mg tablet containing 50 mg daidzein.
EXAMPLE 4
[0060] A group of 36 post-menopausal women experiencing menopausal
symptoms were treated with a composition containing either 15 mg
daidzein or 60 mg daidzein administered on a daily basis for 3
months. Compared to a placebo control group there was a significant
decrease in the Greene Score for menopausal symptoms which
corresponds to treatment/amelioration of menopausal symptoms (such
as hot flushes). The urinary profile of these subjects demonstrated
the therapeutic effectiveness of daidzein as against other
isoflavones.
[0061] In a similar study the same dosage levels of formononetin
again gave a significant decrease in Greene Score for menopausal
symptoms as for daidzein.
EXAMPLE 5
[0062] A pharmacokinetic study involving 16 human patients aged
between 18 and 40 was conducted, to determine the pharmacokinetics
of formononetin following oral administration. Each patient was
orally administered 9.3 mg of formononetin with 200 ml of purified
water and maintained a low isoflavone diet for one week prior to,
and during the study. Blood samples were taken at 0, 0.25, 0.5,
0.75, 1.0. 1.5, 2.0, 2.5, 3, 4, 5, 6. 8, 10 and 12 and 24 hours
post administration, and analysed for formononetin
concentration.
[0063] An analysis of formononetin concentrations (ng/ml) against
time demonstrated that contrary to previous opinion, unmetabolised
formononetin persists in the blood stream for considerable time
following administration (having a half life of about 20
hours).
EXAMPLE 6
[0064] 15 post-menopausal women who are experiencing menopausal
systems are each administered 60 mg formononetin daily for three
months. Relative to a control group the treatment group show a
significant decrease in Greene Score for menopausal symptoms.
[0065] A similar study involving 10 post-menopausal women in a high
risk group for osteoporosis were also administered 60 mg
formononetin daily for three months. Preliminary results indicate
protection against osteoporosis in the treatment group when
measured by bone density and bone turnover markers were
measured.
EXAMPLE 7
[0066] The second study (double-blind, placebo-controlled) involved
a six month study of the effects of phytoestrogen formononetin on
bone resorption markers in twenty post-menopausal women. The aim of
the study was to evaluate the efficacy of a defined daily quantity
of isoflavone at 25 mg, 50 mg or 75 mg on bone resorption makers in
post menopausal women and compare these to normal controls The
effects of isoflavones on endometrial thickness, circulatory lipids
and coagulation factors were also evaluated. Subjects were also
given supplemental calcium in a dose of 1200 mg per day.
[0067] Bone density measurements using a bone densitometer were
made at three sites of the forearm at 0.3 and 6 months and showed
significant improvement in bone density. Bone markers for
osteocalcin, deoxy pyridinoline crosslinks, N-terminal collagen
crosslinks, calcium and other markers showed a similar improvement
in bone resorption and turnover.
[0068] It is to be recognized that the present invention has been
described by way of example only, and that various modifications
and/or alterations which would be obvious to a person skilled in
the art, on the basis of the teaching herein, can be made thereto
without departing from the intended scope or spirit of the
invention.
* * * * *