|
Weight Loss Patent Abstract
The present invention relates to a method of treating or preventing
weight loss of patients with inflammatory bowel diseases comprising
administering to said patients an effective amount of Saccharomyces
boulardii.
Weight Loss Patent Claims
1. A method of treating or preventing weight loss of patients with
inflammatory bowel diseases comprising administering to said patients
an effective amount of Saccharomyces boulardii (S. boulardii).
2. A method according to claim 1, wherein the inflammatory bowel
diseases mentioned above are selected from Crohn's disease, ulcerative
colitis, pouchitis, or indeterminate colitis.
3. A method according to claim 1, of treating or preventing weight
loss of patients with Crohn's disease comprising administering to
said patients an effective amount of S. boulardii.
4. A method according to claim 1, wherein S. boulardii is in the
form of a lyophilizate.
5. A method according to claim 1, wherein S. boulardii is in the
form of a lyophilizate such as obtained according to the following
method: culturing S. boulardii cells (ATCC 74012 or CNCM I-745)
in a nutritive medium until the cells reach a stationary phase,
concentrating the cultured S. boulardii cells and freezing the concentrate,
vacuum drying the frozen concentrate.
6. A method according to claim 1, wherein S. boulardii is administered
orally.
7. A method according to claim 1, wherein S. boulardii is administered
in the form of tablets, capsules, dragees, sachets or suspensions.
8. A method according to claim 1, wherein the daily dosage of S.
boulardii is from about 1 mg/kg to about 100 mg/kg and in particular
of about 10 mg/kg.
Weight Loss Patent Description
BACKGROUND OF THE INVENTION
[0001] The present invention relates to a therapeutic method for
treating weight loss linked to inflammatory bowel diseases (IBD),
such as Crohn's disease and ulcerative colitis.
[0002] Inflammatory bowel diseases are chronic relapsing conditions
of the intestinal tract for which essentially no therapy is available.
The major known forms of IBD are Crohn's disease and ulcerative
colitis (see Carter et al. (2004) Gut 53(supplement V):v1-v16 for
a review).
[0003] Crohn's disease was first described in 1932 and its prevalence
is as high as 75/100,000 in Britain and Scandinavia. It mainly affects
the ileon and/or the colon, with 60% of patients suffering from
an ileocolonic form of the disease. Its clinical manifestations
include abdominal pain, nausea, diarrhea, anorexia, abdominal tenderness,
and weight loss.
[0004] Ulcerative colitis (UC) was first completely described in
1909, its prevalence is approximately 80/100,000 in the Northern
world. UC affects the colonic and rectal mucosa. Among others, it
is manifested by pain, diarrhea, rectal bleeding, anorexia, and
weight loss.
[0005] Current treatments of IBD aim at inducing and preserving
remission and mainly rely on corticosteroids, such as prednisone,
aminosalicylates, such as mesalazine, immunosuppressants, and anti-TNF
monoclonal antibodies, such as infliximab. If a patient fails to
respond to drug therapy and relapses, a surgical intervention is
usually deemed mandatory to remove the dysfunctional part of the
intestinal tract.
[0006] Saccharomyces boulardii (Sacharomyces cerevisiae var. boulardii,
Mallie et al. (2001) J. Mycol. Med. 11:19-25) is a thermophilic
non pathogenic yeast. Clinically, S. boulardii is essentially used
as a probiotic for short course prevention or treatment of diarrhea.
It is used in particular for the management of antibiotic-associated
diarrhea (McFarland et al. (1998) Am. J. Gastroenterol. 90:439-448)
and for the prevention of Clostridium difficile-associated diarrhea
(Surawicz et al. (2000) Clin. Infect. Dis. 31:1012-1017). S. boulardii
has also been used in association with conventional drug treatment
to reduce Crohn's disease-associated diarrhea (Plein and Hotz (1993)
Z. Gastroenterol. 31:129-134).
[0007] Weight loss in inflammatory bowel disease is multifactorial.
Proinflammatory cytokines (IL-1, IL-6 and TNF.alpha.) from the inflamed
intestinal mucosa are known to have several effects leading to disruption
of normal metabolism (Jahnsen et al. (2003) Am. J. Gastroenterol.
98:1556-1562). However, there are currently no therapeutic methods
effective at specifically treating weight loss in IBD patients.
[0008] Thus, an object of the present invention is to provide a
therapeutic method for treating weight loss in patients suffering
from inflammatory bowel diseases.
SUMMARY OF THE INVENTION
[0009] The present invention follows from the unexpected finding
that Saccharomyces boulardii could be used to treat weight loss
in an animal model of inflammatory bowel disease. Furthermore, the
Inventors have shown that this beneficial effect on the prevention
of weight loss is parallel to a previously unrecognized specific
Saccharomyces boulardii-induced lowering of the inflammatory lesions
of the intestinal tract, which is fundamentally different from the
known anti-diarrheal effects of Saccharomyces boulardii.
[0010] Thus, the present invention relates to a method of treating
or preventing weight loss of patients with inflammatory bowel diseases
comprising administrating to said patient an effective amount of
Saccharomyces boulardii.
BRIEF DESCRIPTION OF THE FIGURES
[0011] FIG. 1A, FIG. 1B and FIG. 1C
[0012] FIGS. 1A, 1B and 1C represent the evolution of the weight
(vertical axis, in grams) of IBD mice administered daily with 200
.mu.l of Saccharomyces boulardii solutions, as a function of time
(horizontal axis, in weeks).
[0013] FIG. 1A: diamond shapes represent IBD mice not administered
with S. boulardii, black circles IBD represent mice having received
a 0.1 mg/ml S. boulardii solution, black triangles represent mice
having received a 1 mg/ml S. boulardii solution, crosses represent
mice having received a 10 mg/ml S. boulardii solution, and white
squares represent normal mice.
[0014] FIG. 1B: diamond shapes represent IBD mice not administered
with S. boulardii, black circles IBD represent mice having received
a 0.001 mg/ml S. boulardii solution, black triangles represent mice
having received a 0.01 mg/ml S. boulardii solution, and crosses
represent mice having received a 0.1 mg/ml S. boulardii solution.
[0015] FIG. 1C: diamond shapes represent IBD mice not administered
with S. boulardii, black circles IBD represent mice having received
a 0.001 mg/ml S. boulardii solution, black triangles represent mice
having received a 0.1 mg/ml S. boulardii solution, and crosses represent
mice having received a 1 mg/ml S. boulardii solution.
DETAILED DESCRIPTION OF THE INVENTION
[0016] The yeast Saccharomyces boulardii is a variant of Saccharomyces
cerevisiae. It is notably characterized in Mallie et al. (2001)
J. Mycol. Med. 11:19-25. In particular, isolates of S. boulardii
are deposited under the Budapest treaty at the Centre National de
culture de Microorganismes (Institut Pasteur, Paris, France) under
reference number 1-745 (Mar. 28, 1988) and at the American type
Culture Collection (Rockville, USA) under reference number 74012
(Jul. 18, 1990). Saccharomyces boulardii is also commercially available
as Ultra-Levure.RTM. (Laboratoires Biocodex, Gentilly, France).
[0017] As intended herein, inflammatory bowel diseases (IBD) are
conditions characterized by chronic inflammatory lesions of the
intestinal tract consecutive to an activation of the intestinal
immune system. Manifestations of IBD notably comprise abdominal
pain, diarrhea, anorexia and weight loss.
[0018] The present invention relates to a method of treating or
preventing weight loss of patients with inflammatory bowel diseases
comprising administering to said patients an effective amount of
Saccharomyces boulardii (S. boulardii).
[0019] In a particular embodiment of the above defined method,
the above mentioned inflammatory bowel diseases are selected from
Crohn's disease, ulcerative colitis, pouchitis or indeterminate
colitis.
[0020] In a more particular embodiment, the invention relates to
a method as defined above of treating or preventing weight loss
of patients with Crohn's disease, which comprises administering
to said patients an effective amount of S. boulardii.
[0021] According to a preferred embodiment of the above defined
method of treating or preventing weight loss of patients with inflammatory
bowel diseases, S. boulardii is in the form of a lyophilizate.
[0022] Lyophilization, or freeze drying, is a conservation method
in which yeast cells are freezed and then submitted to a sublimation
of their iced water content to yield a dry yeast powder containing
advantageously less than 2% water, preferably less than 1%. Lyophilizates
are usually obtained by vacuum drying S. boulardii frozen concentrates.
Any yeast lyophilization method known to the man pertaining to the
art can be used. However, a preferred method is as follows.
[0023] Saccharomyces boulardii cells are cultured in a liquid nutritive
medium (from 4700 to 9500 liters) until they reach a stationary
phase, which corresponds to a cell concentration of 10.+-.1% (w/v).
Yeast cells are then concentrated by centrifugation to yield cream-like
concentrates weighing from 500 to 1300 kg depending on the initial
culture volume. From 65 kg to 170 kg of a cryoprotecting 25% (w/w)
lactose solution are added to the concentrates, depending on the
initial culture volume. The concentrates are then freezed at -30.degree.
C. by 2.4 kg aliquots during about 18.5 hours. The frozen concentrates
are lyophilizated using CS 1500 SERAIL (Argenteuil, France) automated
lyophilizers or SMH 1000, SMH 1500, and SMH 3600 USIFROID (Maurepas,
France) automated lyophilizers. The solid cake-like lyophilizates
obtained are then grinded to yield a powder.
[0024] Advantageously, the viability and the vitality of yeast
cells recovered from lyophilizates is higher than what can be obtained
by other conventional conservation methods.
[0025] Thus, the present invention relates to a method of treating
or preventing weight loss of patients with inflammatory bowel diseases
as defined above, wherein S. boulardii is in the form of a lyophilizate
such as obtained according to the following method:
culturing S. boulardii cells (ATCC 74012 or CNCM I-745) in a nutritive
medium until the cells reach a stationary phase,
concentrating the cultured S. boulardii cells and freezing the
concentrate,
vacuum drying the frozen concentrate.
[0026] In a preferred embodiment of the above defined method, S.
boulardii is administered orally.
[0027] In another preferred embodiment of the above defined method
S. boulardii is administered in the form of tablets, capsules, dragees,
sachets or suspensions.
[0028] In yet another preferred embodiment of the above defined
method, the daily dosage of S. boulardii, in particular lyophilizated
S. boulardii, is from about 1 mg/kg to about 100 mg/kg and in particular
of about 10 mg/kg.
[0029] In particular lyophilizated S. boulardii is administered
to patients at dosages ranging from about 100 mg/day to about 1000
mg/day, and in particular at a dosage of about 750 mg/day.
[0030] In an advantageous embodiment of the above defined method,
S. boulardii is administered in association with at least one other
medicine intended for the prevention or the treatment of inflammatory
bowel diseases, such as a medicine selected from the group constituted
of corticosteroids, such as prednisolone, hydrocortisone, or budesonide,
aminosalicylates, such as sulfasalazine or mesalazine, immunosuppressants,
such as azathioprine or 6-mercaptopurine, methotrexate, ciclosporin,
thalidomide, mycophenolate mofetil, and anti-TNF.alpha. monoclonal
antibodies, such as infliximab.
[0031] The present invention also relates to the use of Saccharomyces
boulardii for the manufacture of a medicament intended for the prevention
or the treatment of weight loss in patients with inflammatory bowel
diseases.
EXAMPLE 1
Prevention of Weight Loss by Saccharomyces boulardii in an Animal
Model of Inflammatory Bowel Disease (IBD)
[0032] The animal model which was used in the following experiments
was initially described by Powrie et al. (1993) Int. Immunol. 5:1461-1471
and has been widely in use since then (Powrie et al. (1994) Immunity
1:553-562; Groux et al. (1997) Nature 389:737-742) for studies focusing
on inflammatory bowel diseases, such as Crohn's disease.
[0033] Briefly, immunodeficient mice of the C.B-17 scid type are
intraperitoneously injected with approximately 2.5 10.sup.5 CD4+
T cells which highly express the CD45RB molecule (CD4+CD45RB.sup.hi
T cells), the CD4+CD45RB.sup.hi T cells being isolated by using
a FACS from BALB/C mice.
[0034] These mice, hereafter designated "IBD mice", present
bowel lesions similar to those induced by Crohn's disease in association
to a substantial weight loss.
[0035] The study of the effects of the oral administration of lyophilized
Saccharomyces boulardii on the weight of IBD mice was undertaken
by the Inventors. The evolution of weight variation parallels IBD
evolution.
[0036] Groups of 5 IBD mice were respectively force fed daily with
200 .mu.l of PBS solutions (Phosphate Buffer Saline) containing
lyophilized Saccharomyces boulardii (Ultra Levure.RTM., Laboratoires
Biocodex, France) at concentrations ranging from 0 to 10 mg/ml for
4 weeks. A non-pathogenic control group (i.e. not injected with
CD4+CD45RB.sup.hi T cells) was also included in the study.
[0037] The results obtained from 3 independent experiments are
presented in the following Tables 1, 2 and 3 and in the corresponding
FIGS. 1A, 1B and 1C. TABLE-US-00001 TABLE 1 effect on weight variation
of S. boulardii administration to IBD mice (experiment 1) Concentration
of S. boulardii solution (quantity Weight variation Group administered)
after 4 weeks Control -- +1 g +5% IBD mice 0 mg/ml (0 mg/kg) -9.5
g -47.5% IBD mice 0.1 mg/ml (1 mg/kg) -5 g -25% IBD mice 1 mg/ml
(10 mg/kg) +0.5 g -2.5% IBD mice 10 mg/ml (100 mg/kg) +1 g +5%
[0038] TABLE-US-00002 TABLE 2 effect on weight variation of S.
boulardii administration to IBD mice (experiment 2) Concentration
of S. boulardii solution (quantity Weight variation Group administered)
after 4 weeks IBD mice 0 mg/ml (0 mg/kg) -6.5 g -32.5% IBD mice
0.001 mg/ml (0.01 mg/kg) -5.2 g -26% IBD mice 0.01 mg/ml (0.1 mg/kg)
-4.8 g -24% IBD mice 0.1 mg/ml (1 mg/kg) -3.5 g -17.5%
[0039] TABLE-US-00003 TABLE 3 effect on weight variation of S.
boulardii administration to IBD mice (experiment 3) Concentration
of S. boulardii solution (quantity Weight variation Group administered)
after 4 weeks IBD mice 0 mg/ml (0 mg/kg) -6.5 g -32.5% IBD mice
0.001 mg/ml (0.01 mg/kg) -5.2 g -26% IBD mice 0.1 mg/ml (1 mg/kg)
-3.5 g -17.5% IBD mice 1 mg/ml (10 mg/kg) -2.2 g -11%
[0040] The results obtained by the Inventors indicate that S. boulardii
administration to IBD mice results in a potent reduction in weight
loss. Thus, patients suffering from inflammatory bowel diseases,
such as Crohn's disease could benefit from S. boulardii administration,
at doses ranging from 1 mg/kg to 100 mg/kg, to reduce weight loss.
[0041] Besides, it is to be noted that parallel studies carried
on normal mice administered with the above S. boulardii solutions
did not show any adverse effect of S. boulardii administration,
even with high quantities
EXAMPLE 2
Prevention of Bowel Inflammation by Saccharomyces boulardii in
Parallel with Prevention of Weight Loss in an Animal Model of Inflammatory
Bowel Disease
[0042] Since it was shown by the Inventors that lyophilized S.
boulardii administration to IBD mice has a beneficial effect on
the limitation of weight loss, they further investigated to determine
if S. boulardii had a favorable impact on the inflammatory status
of the colon.
[0043] Colonic sections taken from IBD mice treated as described
in Example 1 were graded from 0 to 5, in a blind study, relatively
to their inflammatory status.
[0044] The following grades were affected to each section analyzed:
[0045] Grade 0: normal; [0046] Grade 1: slight scattered mucosal
leukocyte infiltration optionally associated to a slightly hyperplastic
epithelium; [0047] Grade 2: modest scattered or grouped leukocyte
infiltration reaching sub-mucosa associated to an eroded and a slightly
hyperplastic epithelium; [0048] Grade 3: slight transmural inflammatory
infiltrations, usually associated to ulcerations and a slight hyperplasia;
[0049] Grade 4: strong infiltrations, usually transmural, associated
to ulcerations and a strong hyperplasia; [0050] Grade 5: strong
transmural inflammation associated to severe ulcerations and an
intestinal gland loss.
[0051] The results obtained from 3 independent experiments are
presented in the following
[0052] Tables 4, 5 and 6: TABLE-US-00004 TABLE 4 effect on bowel
inflammation of S. boulardii administration to IBD mice (experiment
1) Concentration Clinical grade of S. boulardii solution of colonic
section Group (quantity administered) after 4 weeks IBD mice 0 mg/ml
(0 mg/kg) 4 .+-. 0.5 IBD mice 0.1 mg/ml (1 mg/kg) 3.5 .+-. 0.2 IBD
mice 1 mg/ml (10 mg/kg) 2.5 .+-. 0.5 IBD mice 10 mg/ml (100 mg/kg)
2.2 .+-. 0.3
[0053] TABLE-US-00005 TABLE 5 effect on bowel inflammation of S.
boulardii administration to IBD mice (experiment 2) Concentration
of Clinical grade S. boulardii solution of colonic section Group
(quantity administered) after 4 weeks IBD mice 0 mg/ml (0 mg/kg)
4.2 .+-. 0.4 IBD mice 0.001 mg/ml (0.01 mg/kg) 4.5 .+-. 0.4 IBD
mice 0.01 mg/ml (0.1 mg/kg) 4.1 .+-. 0.8 IBD mice 0.1 mg/ml (1 mg/kg)
3.3 .+-. 0.7
[0054] TABLE-US-00006 TABLE 6 effect on bowel inflammation of S.
boulardii administration to IBD mice (experiment 3) Concentration
of Clinical grade S. boulardii solution of colonic section Group
(quantity administered) after 4 weeks IBD mice 0 mg/ml (0 mg/kg)
4.2 .+-. 0.6 IBD mice 0.01 mg/ml (0.1 mg/kg) 4 .+-. 0.5 IBD mice
0.1 mg/ml (1 mg/kg) 2.4 .+-. 0.4 IBD mice 1 mg/ml (10 mg/kg) 2.1
.+-. 0.7
[0055] The results obtained by the Inventors indicate that S. boulardii
administration to IBD mice results in a significant reduction of
colon inflammation, which correlates with the reduction in weight
loss observed in Example 1. Furthermore, the dose of lyophilized
S. boulardii effective at reducing inflammation is determined as
ranging from 1 mg/kg to 100 mg/kg.
EXAMPLE 3
Prevention of IFN-.gamma. Secretion by Saccharomyces boulardii
in an Animal Model of Inflammatory Bowel Disease
[0056] Following the finding that S. boulardii had a favorable
impact on the prevention of weight loss and on the inflammatory
status of the colon, the Inventors further investigated the effect
of S. boulardii on the production of the Th1-secreted inflammatory
cytokine IFN-.gamma. by cells of the colon.
[0057] Briefly, colonic T cells were purified from inflammatory
colonic sections taken from IBD mice treated as described in Example
1 and were cultured in the presence of mitogens. IFN-.gamma. concentration
in the cultures supernatant of the purified T cells was then measured
by ELISA.
[0058] The results obtained from 3 independent experiments are
presented in the following
[0059] Tables 7, 8 and 9: TABLE-US-00007 TABLE 4 effect on IFN-.gamma.
secretion of S. boulardii administration to IBD mice (experiment
1) Concentration of S. boulardii solution IFN-.gamma. Group (quantity
administered) concentration (ng/ml) IBD mice 0 mg/ml (0 mg/kg) 33
IBD mice 0.1 mg/ml (1 mg/kg) 0 IBD mice 1 mg/ml (10 mg/kg) 0 IBD
mice 10 mg/ml (100 mg/kg) 0
[0060] TABLE-US-00008 TABLE 5 effect on IFN-.gamma. secretion of
S. boulardii administration to IBD mice (experiment 2) Concentration
of S. boulardii solution IFN-.gamma. Group (quantity administered)
concentration (ng/ml) IBD mice 0 mg/ml (0 mg/kg) 29 IBD mice 0.001
mg/ml (0.01 mg/kg) 32 IBD mice 0.01 mg/ml (0.1 mg/kg) 28 IBD mice
0.1 mg/ml (1 mg/kg) 9
[0061] TABLE-US-00009 TABLE 6 effect on IFN-.gamma. secretion of
S. boulardii administration to IBD mice (experiment 3) Concentration
of S. boulardii solution IFN-.gamma. Group (quantity administered)
concentration (ng/ml) IBD mice 0 mg/ml (0 mg/kg) 18 IBD mice 0.01
mg/ml (0.1 mg/kg) 16 IBD mice 0.1 mg/ml (1 mg/kg) 1 IBD mice 1 mg/ml
(10 mg/kg) 3
[0062] The results obtained by the Inventors indicate that S. boulardii
administration to IBD mice results in a significant reduction of
IFN-.gamma. secretion by colonic T cells of the Th1 subtype, which
correlated with the observed reduction in inflammation and prevention
of weight loss. As seen in Examples 1 and 2, the effective dose
of lyophilized S. boulardii ranges from 1 mg/kg to 100 mg/kg.
[0063] All the cited publications are incorporated herein by reference. |