Note: Descriptions are shown in the official language in which they were submitted.
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1
INHIBITION OF THE ADHESION OF PATHOGENIC MICROORGANISMS BY A SUCROSE
STEA1RATE AND/OR A SORBITAN ESTER IN THE COSMETIC TREATMENT OF CUTANEOUS
ATOPY
SUMMARY OF THE INVENTION
The invention relates to a cosmetic treatment process for human atopic
dermatitis based on
coating the surface of the skin with a cosmetic composition that forms a film
on it.
According to the invention, this film¨by virtue of its constituents __
inhibits the adhesion
of pathogenic micro-organisms to the skin and reinforces the skin barrier.
More specifically, the cosmetic composition contains an ester of sucrose
and/or an ester of
sorbitan capable of blocking the adhesion of Staphylococcus aureus to the skin
and nasal
mucosa, thereby inhibiting the proliferation of pathogenic flora. The effect
of this inhibition
is to protect the cutaneous barrier by limiting the breakdown of lipids,
especially ceramides,
which is indirectly due to Staphylococcus aureus.
PRIOR ART
Atopic dermatitis or cutaneous atopy is an inflammatory, pruriginous skin
condition
associated with a hereditary immune predisposition and abnormalities of the
cutaneous
barrier. In clinical terms, this hereditary predisposition can also manifest
as allergic rhinitis
and asthma.
Atopic dermatitis leads to hypersensitivity to environmental allergens which
are tolerated
by healthy subjects.
In atopic dermatitis, eczema represents a delayed-type hypersensitivity
reaction mediated
by Th2 lymphocytes (which produce IL-4 and IL-5) and antigen-presenting cells.
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Like any immune reaction mediated by antigen-specific T lymphocytes, the
inflammatory
reaction of the eczema of atopic dermatitis proceeds through three phases.
The first phase is asymptomatic sensitisation. This phase is clinically
inapparent and leads
to the amplification of antigen-specific T lymphocytes. Typically,
sensitisation occurs in
young childhood through exposure to environmental allergens which are captured
by
dendritic cells.
In a second phase, eczema develops. After re-exposure to the allergen in
question,
Langerhans cells migrate and activate antigen-specific, cytokine-producing Th2
lymphocytes which, in turn, activate a variety of different cell-types in the
skin. This
mechanism helps recruit leukocytes into the dermis and epidermis where they
produce
inflammatory mediators. This phase of variable duration is characterised by
eczema lesions.
In most patients, it is accompanied by high levels of antigen-specific IgE
antibodies, some
of which are bound to the surface of Langerhans cells.
Finally, the lesions regress in the third phase. The eczema of atopic
dermatitis develops in
flare-ups separated by periods of spontaneous remission. The mechanisms
involved in
regulating the inflammation remain poorly understood.
Atopic dermatitis affects 10-30% of the population but its prevalence is
constantly on the
rise in developed countries, having doubled or tripled in the last twenty
years.
Atopic dermatitis is characterised by pruriginous flare-ups of acute eczema
separated by
periods of remission. It is most common in children. It onsets a matter of
months after birth
in the form of lesions on the cheeks and areas of skin subject to rubbing. It
then develops in
the form of attacks between one and two years of age. During this time, the
skin is dry and
has red, oozing patches, especially on the inner surfaces of skinfolds. After
five years of
age, attacks tend to disappear but the skin remains dry and sensitive. In some
people, atopic
dermatitis can persist into adulthood.
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Two phenomena follow on from atopic dermatitis. Firstly, the cutaneous barrier
is
weakened and secondly, the skin gets colonised by a flora of pathogenic micro-
organisms.
The main tissue of the cutaneous barrier is the stratum corneum, the outermost
layer of the
epidermis comprised of corneocytes and special lipids, including cholesterol,
free fatty
acids, cerebrosides and ceramides. These lipids act as intercellular cement
and make the
stratum corneum water-proof.
Analysis of the stratum corneum of patients with atopic dermatitis show marked
deficiencies in certain proteins and cutaneous lipids. There are deficiencies
in both
filaggrin, 'a protein involved in aggregating keratin filaments in
corneocytes, and
involucrin, a protein essential to constructing the protein skeleton of the
stratum corneum.
The lipids that are most notably deficient are ceramides 1 and 3.
Ceramides are particularly important in regulating skin barrier function by
limiting the
evaporation of water.
These protein and lipid deficiencies mean that corneocytes adhere to one
another less
strongly, compromising the cohesion and integrity of the stratum corneum. This
manifests
as a significant reduction in the thickness of the stratum corneum in these
patients.
Atopic dermatitis manifests as a significant loss of cutaneous proteins and
lipids which, as
mentioned above, leads to evaporation of moisture and dry skin. This is known
to make it
= easier for allergens to enter the skin.
Atopic dermatitis therefore triggers a vicious cycle: the evaporation of
intracellular water
leads to cutaneous dehydration and dry skin which in turn increases cutaneous
permeability. Greater cutaneous permeability makes it easier for allergens to
penetrate
through the tissue which sustains the skin's irritability.
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The skin's barrier function is also consolidated by the local ecosystem which
consists of a
flora of saprophytic bacteria.
A saprophytic bacterial flora is a natural, permanent feature of the surface
of the skin. The
density of bacteria on the surface of the skin is estimated at between 102 and
105/cm2. The
most common species are staphylococci, notably coagulase-negative
Staphylococcus
epidermidis and related species (Micrococcus) and coryneform bacteria, both
aerobic
(Corynebacterium, Brevibacterium) and anaerobic (Propionibacterium acnes).
These
bacteria are established on the stratum corneum where they adhere to
corneocytes to form a
protective biofilm.
This saprophytic bacterial flora therefore reinforces barrier function since
it occupies
potential adhesion sites for other micro-organisms __________________ which
might be pathogenic .. and
inhibit their proliferation.
However, if the skin is damaged as it is in atopic dermatitis, impaired
cutaneous barrier
function promotes water evaporation and creates a skin surface environment
that is
conducive to colonisation by bacteria, viruses or fungi. This permissive
environment is
exacerbated by the rise in skin temperature that occurs during an attack of
eczema.
Thus, a pathogenic bacterial flora can grow temporarily on the surface of the
skin. Most of
these species are found in the normal environment, including species belonging
to the
genera Pseudomonas and Acinetobacter; others derive from the digestive or
buccal flora
such as enterobacteria, streptococci or species of the genus Clostridium.
These bacteria are not usually capable of proliferating on the surface of
human skin but
they can cause infection if cutaneous barrier function is compromised as in
atopic
dermatitis in which they can form a pathogenic bacterial biofilm.
Among the bacteria of the transient pathogenic flora, Staphylococcus aureus
(S. aureus) is
the potentially most pathogenic species being the most common member of the
genus
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which is found in the nasal passages and throat of 15-30% of healthy people.
This Gram-
positive bacterium establishes itself on the skin by adhering to skin cells.
Recent studies have shown that the skin of 90% of patients with atopic
dermatitis is
5 colonised by S. aureus, compared with with only 5% of healthy subjects.
Remarkably, atopic dermatitis is also a proven risk factor for colonisation of
the nasal
mucosa by S. aureus which then provides a reservoir in the environment of
atopic patients
(Pascolini et al., 2011).
Some studies have also suggested that ceramides produced by the cutaneous
flora are
activated in the presence of S. aureus. Ceramidases hydrolyse ceramides in the
stratum
corneum which might suggest that colonisation of the skin by S. aureus is
partly
responsible for the deficiency in this type of lipid in the atopic epidermis
(Kita et al., 2002).
S. aureus is also a source of superantigens (proteinaceous toxins) which, in
subjects with
atopic dermatitis, interact with immune cells, amplifying the inflammatory
response and
thereby triggering flare-ups.
There is therefore a tangible link between the proliferation of S. aureus on
the epidermal
surface and impairment of cutaneous barrier function as a result of ceramide
deficiency,
especially in atopic subjects (remembering that impaired barrier function
promotes
sensitisation to environmental allergens and maintenance of inflammation of
the skin).
The two main viruses responsible for infections are herpes simplex virus (HSV)
and
vaccinia virus. Colonisation by the yeast Malassezia has also been observed in
patients
with atopic dermatitis.
Current treatment modalities for atopic dermatitis are based on managing the
inflammation
and controlling bacterial colonisation.
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Modalities aimed at managing the inflammation include the administration of
glucocorticosteroids and calcineurin inhibitors like pimecrolimus and
tacrolimus. These
products are usually combined with emollient products.
Bacterial control has been extensively reported on, notably involving the
administration of
broad-spectrum antibiotics and the application of antiseptic solutions. First-
line treatment is
becoming difficult because of antibiotic-resistant bacteria. Moreover, both
types of
treatment not only kill the pathogens present on the skin but also the
beneficial saprophytic
bacteria.
The main disadvantage of both these modalities is that they destroy a
component that is
required for effective barrier function, namely the saprophytic bacterial
flora.
Recent studies have shown that the topical application of compositions
containing certain
sugars inhibit the adhesion of pathogenic bacteria to the skin.
Document WO 2006/106220 reports that certain sugars¨mono- and oligo-
saccharides such
as rhamnose, galactose, mannose and lauryl glucoside¨inhibit the adhesion of
bacteria to
corneocytes from atopic dogs, especially bacteria of the genus Staphylococcus
(S.
intermedius).
Document 'WO 96/23479 suggests that certain sugars can be used to inhibit the
adhesion of
certain micro-organisms. Singled out are monosaccharides like raffinose,
mannose and
rhamnose, disaccharides, oligosaccharides, aminated sugars and esters of
various sugars
including esters of glucose such as cetearylglucoside, caprylglucoside and
decylgincoside.
These various sugars can be blended into the compositions of cosmetic or
dermatological
preparations to inhibit bacterial adhesion, e.g. that of S. aureus, and/or for
therapeutic
purposes, notably the treatment of atopic dermatitis.
Document EP0875239A2 lists various esters of sugars including esters of
sucrose, for their
activity against certain Gram-positive species like Staphylococcus
epidermidis,
7
Staphylococcus aureus, Corynebacterium and Propionibacterium, involved in a
list of
pathologies on which atopic dermatitis appears. The esters listed are
palmitate/stearate
diester, stearate diester, laurate monoester, myristate monoester, and
stearate triester and
tetraester.
Document EP1340486A1 lists various sugar esters, including esters of sucrose,
fructose,
glucose and trehalose. Only the esters of fructose, glucose and trehalose are
tested for their
ability to inhibit the growth of Staphylococcus aureus. Sucrose esters are not
tested for this
activity. Only one representative of this family, namely sucrose stearate, is
tested and this
only for bleaching activity (see Table 7). Finally, this document does not
address atopic
dermatitis.
Document EP0815841A1 reports a composition to control skin redness due to
nappies
rubbing. This is not a treatment for atopic dermatitis as described above.
This document
describes the inhibitory effect on the growth of Staphylococcus aureus and
Staphylococcus
epidermidis of a mixture of monoesters of sucrose with palmitic acid and
stearic acid or a
mixture of monoesters with palmitic acid and lauric acid.
Document EP2210588A1 describes foaming compositions containing polysorbate 80,
i.e.
an ester of sorbitan. Polysorbate 80 is a very common emulsifying agent.
Document W04/037225A2 describes compositions free of either alcohol or
propylene
glycol as a vehicle for foaming aerosols. Atopic dermatitis is mentioned among
other
diseases that could be treated using the aerosol supplemented with some
specific active
substance. Sucrose stearate and polysorbate 80 mentioned in the examples are
commonly
used a surface-active agents.
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Document FR2798591A1 describes the use of special vegetable oil to enhance the
synthesis of cutaneous lipids, in particular for the treatment of atopic
dermatitis. Sucrose
distearate and sorbitan tristearate are commonly used as surface-active
agents.
Document EP1639989A1 describes the generation of micro-emulsions containing
esters of
sugar or sorbitan used as surface-active agents.
In Examples 5 and 8 of document US2005/158348A1, the compositions respectively
contain polysorbate 80 and a sucrose ester. These compositions are intended
for the
treatment of pain and inflammation. Esters of sorbitan and sucrose are used in
the
excipient.
Document US2010/080768A1 describes the presence of various ceramides and
sphingolipids in compositions intended for the treatment of various diseases,
including
atopic dermatitis.
Document US2011/101135 describes using sorbitan monocaprylate as a
preservative in a
cosmetic composition. Various bacterial strains are tested, including
Staphylococcus
aureus.
There is nevertheless a need for more effective alternatives to the various
modalities
currently available for managing atopic dermatitis.
It is in this context that the Applicant discovered, surprisingly and
unexpectedly, that
certain sugar esters like esters of sucrose and sorbitan, strongly inhibit the
adhesion of
pathogenic micro-organisms to the skin and mucous membranes.
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Inhibition of the adhesion of S. a=treus to the surface of the skin prevents
the proliferation
of pathogenic micro-organisms, even in the absence of antibiotic or topical
antiseptic.
One of the advantages of inhibiting the adhesion of pathogenic micro-organisms
to the
skin, in particular S. aureus, is to maintain the saprophytic bacterial flora
and reinforce
cutaneous barrier function by cutting down breakdown of its component lipids.
Sugar esters can be advantageously combined with lipids that occur naturally
in the
epidermis to form a film that acts still more effectively against breakdown of
the epidermal
barrier and colonisation of the skin by pathogens.
Other aims and aspects of the invention will emerge on reading the following
detailed
description which is given for the purposes of illustration and is in no way
limiting.
DETAILED DESCRIPTION OF THE INVENTION
A first aspect of the invention relates to a cosmetic composition for topical
application
containing at least one sucrose ester and/or sorbitan ester as an agent to
inhibit the adhesion
of Staphylococcus aureus to human skin and nasal mucosa.
More specifically, the invention concerns a composition for topical
application containing
at least one sucrose ester and/or sorbitan ester for use in the treatment of
atopic dermatitis,
as an agent to inhibit the adhesion of Staphylococcus aureus to human skin and
nasal
mucosa.
A sugar ester is a sugar in which at least one free alcohol group has been
esterified with a
fatty acid Chain.
A series of different families of sugar ester have been described. Notably,
there are esters of
glucose, sucrose and sorbitan (Piccicuto et at., 2001). They have the
advantage of being
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non-toxic and non-irritant and are therefore in widespread use in the food
processing,
pharmaceutical and cosmetic industries, often for their surface-active
properties.
Out of these three distinct families, this Application only concerns esters of
sucrose and
.. sorbitan.
These are non-ionic surfactants with a hydrophilic group consisting of sucrose
or sorbitan
joined through an ester linkage to a hydrophobic group consisting of a fatty
acid. The
surface-active properties of these esters also make them easily to combine
with lipids in
cosmetic compositions, especially emulsions.
=
a-D-glucopyranosyl-P-D-fructofuranoside or sucrose carries eight free alcohol
groups and
can therefore be esterified with up to eight fatty acids.
Sorbitan is generated by the dehydration of sorbitol, a polyhydroxylated
compound which
is itself obtained by reducing the aldehyde group of glucose to create an
alcohol group.
Sorbitan carries four free alcohol groups, each of which can be esterified by
a fatty acid.
In the context if the present invention, sorbitan esters means esters of the
"Span" type, e.g.
with the following developed structural formula:
OH
0
HO OH
in which R is a fatty acid, in this case a monoesterified span
The present invention also concerns "Tween"-type polyethoxylated sorbitan
esters, e.g. the
following formula:
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o(cH2cH2o>1H
op-1204201w ____________________
R
H(OCH2CH2ixo 0(CH2CH20)1FI
in which R is a fatty acid, in this case a monoesterified tween.
The carbon chains of the fatty acids may be short or long, e.g. mention could
be made of
lauric acid, myristic acid, palmitic acid and stcaric acid with respective
chains of C12, C14,
C16 and C18. The fatty acid may also be chosen from among saturated and
unsaturated fatty
acids, e.g. oleic acid (a C18 carbon chain).
Thus, according to another characteristic of the invention, the cosmetic
composition
contains sucrose and/or sorbitan with at east one alcohol group esterified
with a fatty acid
from the group with carbon chains of between C12 and C22, advantageously
between C12 and
C18.
Depending on the ratio of fatty acids to sucrose, esterification can yield the
monoester,
diester, triester or polyester (up to the octa-ester), or a mixture of all
these forms. The
esterification of sorbitan can yield the monoester, diester, triester, tetra-
ester or a mixture of
these various forms.
According to another embodiment, the cosmetic composition of the invention
contains
sucrose and/or sorbitan in which some or all of the alcohol groups have been
esterified with
the same fatty acid.
In this case and advantageously, the sucrose ester will be selected from the
group
containing sucrolaurate, sucromyristate, sucropalmitate and sucrostearate, and
the sorbitan
ester will be selected from the group containing sorbitan laurate (Span 20),
sorbitan
monostearate (Span 60), sorbitan oleate (Span 80) and sesquioleate sorbitan
(Montane 83).
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In an advantageous embodiment, the sucrose ester is a sucrose stearate
containing at least
70%, advantageously at least 75% monoesters of sucrose and stearic acid by
weight, the
rest being di, tri and polyesters of sucrose and stearic acid.
Advantageously, the cosmetic composition according to the invention contains a
polyethoxylated sorbitan ester. Mention could be made of the ethoxylated
sorbitan
monolaurate (Polysorbate 20), polyoxyethylene sorbitan monostearate 20
(Polysorbate 60)
and polyoxyethylene sorbitan mono-oleate (Polysorbate 80). In a preferred
embodiment,
the sorbitan ester is polysorbate 20 or polysorbate 80.
According to an alternative embodiment, the cosmetic composition can be made
up with
sucrose and/or sorbitan in which some or all of the alcohol groups have been
esterified with
at least two different fatty acids.
In this case and advantageously, the sucrose ester will be selected from the
group
containing sucropalmitate stearate and sucrotetrastearate triacetate, and the
sorbitan ester
will be selected from the group containing the oleate/stearic acid ester of
sorbitan
(Montane 481) and sorbitan esterified with fatty acids from olive oil (olivate
sorbitans).
According to a preferred embodiment, the sucrose ester in the cosmetic
composition
according to the invention is the sucrostearate, i.e. sucrose esterified with
at least one
molecule of stearic acid (a C18 fatty acid).
Those skilled in the art are aware that, for a given fatty acid carbon chain
length, the
proportion of monoesters in the mix will affect the
hydrophobicity/hydrophilicity of the
sugar ester.
These properties are characterised by means of the Hydrophile/Lipophile
Balance (HLB)
index. A value of between 1 and 10 indicates a lipophilic sugar ester,
corresponding to
relatively few monoesters and therefore a relatively high proportion of
polyesters. But a
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value of between 11 and 20 indicates a hydrophilic sugar ester, corresponding
to a high
proportion of monoesters and therefore a relatively low proportion of
polyesters.
In a particular embodiment of, the invention, the sucrostearate has a HLB of
16,
corresponding to a relative proportion of sucrose monoesters and stearic acid
of between
75% and 80% in weight. Hereafter, this product is referred to as sucrose
monostearate.
Preferably, the sucrose ester and/or the sorbitan ester represents 0.1-5% of
the composition
by weight,,advantageously between 1% and 3%.
Advantageously, the cosmetic composition also contains lipids that are capable
of restoring
the cutaneous barrier, respectively:
- at least one lipid not found naturally in the skin, advantageously an
oil; and/or
- a mixture of constituents that occur naturally in the skin, including
ceramides 1,
3, 6, cholesterol, free fatty acids and phytosphingosine.
In practice, supplementation with certain specific lipids can help restore the
lipid layer
between skin cells, i.e. the extracellular cement, thereby helping to restore
the cutaneous
barrier. As a result, the evaporation of water from the skin is cut down
together with the
risk of S. aureus adhering on the surface of the skin and proliferating there.
This action
combines with the effects of the esters of the invention which inhibit the
adhesion of S.
aureus and prevent the hydrolysis of ceramidcs in the stratum corneum.
Lipids that are not naturally found in the skin, like oils, mediate short-term
moisturisation
of the tissue. They are'advantageously selected from the group including
sunflower oil and
rapeseed oil (canola) because of the high concentrations of essential omega-6
and omega-3
fatty acids in these oils.
In a particular embodiment of the invention, the cosmetic composition contains
lipids not
found naturally in skin, respectively sunflower oil representing 3-15% of the
composition
by weight and rapeseed oil (canola) representing 0.05-10% of the composition
by weight.
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Supplementation with lipids that occur naturally in the skin can correct the
deficiency in
certain stratum corneum lipids seen in patients suffering from atopic
dermatitis. This can
help restore a functional cutaneous barrier in the long term, especially when
combined with
the effects of the esters described above.
In a particular embodiment, the mixture of naturally-occurring constituents
corresponds to
a lipid composition with an INCI designation, namely Water, Ceramide 3,
Ceramide 611,
Ceramide 1, Phytosphingosine, Cholesterol, Sodium Lauroyl Lactylate, Carbomer,
Xanthan
Gum. This lipid composition is advantageously supplied as a commercial
product, SK
Influx VTM (Evonik Industries).
Preferably, the lipid composition represents 0.01-5% of the overall
composition by weight.
Advantageously, the cosmetic composition also contains at least one additional
polyhydroxylated compound selected from the group containing rhamnose, xylitol
and
rnannitol.
These additional polyhydroxylated compounds like rhamnose and xylitol help
inhibit the
adhesion of pathogenic bacteria such as S. aureus to human skin and nasal
mucosa.
Mannitol also has free radical-quenching activity.
In a particular embodiment of the invention, the cosmetic composition contains
a mixture
of three of the additional polyhydroxylated compounds mentioned above.
Advantageously, rhamnose represents 0.01-1% of the composition by weight,
xylitol
represents 0.05-2% of the composition by weight and mannitol represents 0.005-
1% of the
composition by weight.
Advantageously, the composition also contains at least one anti-pruriginous
agent selected
from the group containing in particular:
- palmitoyl ethanolamide, CAS Number 544-31-0;
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- hydroxy-a-sanshool, CAS Number 83883-10-7, part of the composition of
zanthaleneTM
- a lipo-dipeptide based on tyrosyl-arginine, part of the composition of
'calmosensineTm ;
5 - ichtyol or ammonium ichthosulphonate, CAS Number 8029-68-3.
Advantageously, the cosmetic composition according to the invention can also
include at
least one anti-inflammatory agent, preferably selected from the group
containing:
- beta-sitosterol, CAS Number 83-46-5;
10 - ,enoxolone or glycyrrhetinic acid, CAS Number 471-53-4.
Advantageously, the cosmetic composition also includes Vitamin PP which is
known to
stimulate lipid synthesis in the stratum corneum, including that of ceramides,
free fatty
acids and cholesterol. Vitamin PP acts by stimulating the activity of serine
palmitoyl
15 transferase, a key enzyme in the synthesis of sphingosine which is the
precursor for
ceramide production.
Another aspect of the invention concerns a cosmetic composition for the
cosmetic
treatment of atopic dermatitis.
The invention also concerns a cosmetic treatment process for atopic dermatitis
based on
forming a film on the skin to protect the epidermis by means of the
composition described
hereafter.
=
In an advantageous embodiment, the treatment process involves applying to the
skin a film-
forming composition, notably containing a sucrose and/or sorbitan ester to
inhibit the
adhesion of S. aureus to the surface of the skin, together with lipids, e.g.
fatty acids,
ceramides and cholesterol, to reinforce the skin's defences against insults
from micro-
organisms, and notably against enzymes secreted by such micro-organisms that
hydrolyse
constituents of the stratum corneum.
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This film can be applied to the skin according to the stage of the disease, in
order to:
- arrest its progress;
- act during flare-ups of the disease;
- prevent recurrence.
Advantageously, the composition used to stop progression contains (in
percentage weight
of the composition):
between 1% and 5% sucrose monostearate and/or a sorbitan ester;
between 3% and 25% of a mixture of oils;
between 0.01% and 5% SK Influx VTM;
between 0.06% and 4% of a mixture of rhamnose, xylitol and mannitol.
Advantageously, the composition used to treat flare-ups contains (in
percentage weight of
the composition):
between 1% and 5% sucrose monostearate and/or a sorbitan ester;
between 3% and 25% of a mixture of oils;
between 0.01% and 5% SK Influx VTM;
between 0.06% and 4% of a mixture of rhamnose, xylitol and mannitol;
between 0.01% and 1% of an anti-pruriginous agent;
- = between 0.5% and 1% of an anti-inflammatory agent;
Advantageously, the composition used notably to prevent recurrence contains
(in
percentage weight of the composition):
between 1% and 5% sucrose monostearate and/or a sorbitan ester;
between 3% and 25% of a mixture of oils;
between 0.01% and 5% SK Influx VTM;
between 0.06% and 3% of a mixture of rhamnose, xylitol and mannitol;
between 0.01% and 1% of an anti-pruriginous agent;
between 0.01% and 2% Vitamin PP.
=
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These various compositions are film-forming, i.e. they can create a film over
the surface of
the skin to protect it against insults from pathogenic micro-organisms.
A number of advantages emerge from reading about the invention:
reducing S. aureus adhesion to atopic skin prevents the growth of a
pathogenic bacterial flora at the same time as maintaining the saprophytic
bacterial flora;
it is known that a bacterial biofilm grows on the surface of the skin in a
series of different phases from bacterial adhesion through growth,
maturation and expansion of the film. Therefore, inhibiting the adhesion of
S. aureus to atopic skin leads to inhibition of the formation of a pathogenic
biofilm;
using a non-toxic, non-irritant sucrose and/or sorbitan ester cuts down the
need for antimicrobial agents such as antibiotics and topical antiseptic
products;
the presence of lipids that do not occur naturally in the skin restores
barrier function and helps cut down the evaporation of water from the
epidermis;
complementary action by the sucrose and/or sorbitan ester and the lipids
reduces the burden of S. aureus as a result of the inhibition of adhesion
and then restores the stratum corneum's barrier function;
the sucrose and/or sorbitan ester and the lipids therefore form a film on the
epidermis that protects it against insults from pathogenic micro-organisms
= and prevents breakdown of the barrier function.
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EXAMPLES
1/ EFFECT OF SUCROSTEARATE ON IN VITRO ADHESION OF STAPHYLOCOCCUS AUREUS
ONTO A SOLID MATRIX.
1) Method
The following protocol was used:
- S. aureus was grown in the Clinical Bacteriology Laboratory at La Timone
Hospital
(CIP 65-8) in 'Nutrient Broth N 2' (Oxoid), for 12 hours at 37 C;
- The culture was centrifuged at 9,000 g for 10 min and then the bacterial
pellet was
resuspended in phosphate-buffered saline (PBS);
- This suspension was diluted 50-fold in an aqueous solution or in an
aqueous
solution containing the test polyhydroxylated compound. This was either
xylitol or
sucrostearate HLB 16 (SURFHOPETM C1816);
- The bacteria were pre-incubated with the test polyhydroxylated compound. for
45
mm at 37 C;
- The mixture (100 I) was then spotted into a well on a glass cytology
slide, of fixed
surface area (0.8 cm2) with walls, i.e. a special slide for cell culture
(Chamber Slide
System, Labtek);
- The bacterial cultures were then incubated at 37 C for 60 mm after which
they were
rinsed with UltraPure water to wash off any bacteria that had not stuck to the
surface of the well;
- Adherent bacteria were stained with Crystal Violet for 10 seconds, rinsed
and dried
at 25 C for 12 hours;
- Finally, the bacteria were examined by microscope at a magnification of X
1,000
(oil immersion) with perfectly homogeneous fields;
- Finally, the number of bacteria in each well was counted automatically
using
appropriate software.
CA 02881267 2015-02-04
19
2) Results
In this protocol, the effect of the polyhydroxylated compounds was analysed
vis-a-vis
inhibition of the adhesion of S. aureus onto the solid matrix. The tests were
carried out with
xylitol at a final concentration of 0.5% and the HLB16 sucrostearate (sucrose
monostearate,
SURFHOliE C18-C16) at a final concentration of 1%.
Table 1: Effects of xylitol (0.5%) and HLB 16 sucrostearate (1%) on the
adhesion of S.
aureus to the surface of glass slides. Results are presented as the mean count
of three fields
.. together with the corresponding standard deviation.
Condition Number of cells per Inhibition of adhesion (% with
respect
field to the water control)
Water (control) 99 11 0%
Xylitol (0.5%) 91 6 8%
HLB 16 12 1 88%
sucrostearate (1%)
3) Conclusion
Strong inhibition of the adhesion of S. aureus to the surface of glass slides
was observed in
the presence of 1% HLB 16 sucrostearate.
2/ EFFECT OF ESTERS OF THE INVENTION ON EX VIVO ADHESION OF S. AUREUS ON HUMAN
CORNEOCYTES.
To corroborate the in vitro findings, an ex vivo adhesion test on human
corneocytes was
adapted from a method developed on dogs (McEwan et al., 2005).
CA 02881267 2015-02-04
1) Bacterial strain
A clinical* strain of S. aureus (CIP 65-8) was isolated from a patient in the
Clinical
Bacteriology Laboratory at La Timone Hospital in Marseille.
5
2) Protocol
S. aureus was inoculated into 10 ml of 'Nutrient Broth N 2' (Oxoid) and
incubated with
stirring at 37 C for 12 hours. The culture was centrifuged and the bacterial
pellet was
10 resuspended in PBS. Bacterial density was adjusted to about 106
CFU/ml with osmotically
purified water.
Target areas were marked out on the arms of a human volunteer. On each area,
cutaneous
debris was removed with 5 successive "pre-stripping" procedures with discs of
15 SellotapeTmOriginal (diameter 22 mm). Corneocytes were then
collected from each target
area using D-SquameTM sticky discs (diameter 22 mm) with a D-Squame disc
applicator set
to a pressure of 150 g/cm2.
The discs were then placed in Petri dishes (diameter 35 mm) and covered with
0.5 ml water
20 or 0.5 ml of a suspension of S. aureus at a density of 106 CFU/ml,
containing the test
polyhydroxylated compound (triplicate areas for each).
The polyhydroxylated compounds tested in this way were:
xylitol, at a final concentration of 0.5%;
HLB 16 sucrose stearate (sucrose monostearate, SURFHOPE C1816,) at a
final concentration of 1%,
sucrose laurate monoester (SURFHOPE C1216, 80% monoester), at a
final concentration of 0.1%,
polysorbate 60, :t a final concentration of 0.1%;
polysorbate 60, at a final concentration of 1%;
polysorbate 20, at a final concentration of 0.1%;
CA 02881267 2015-02-04
21
All solutions were made up in water and put in contact with S. aureus at room
temperature
45 mm before the start of the adhesion experiment. The dishes were then
incubated at 37 C
for 60 min. Then the discs were rinsed in osmotically purified water to remove
any bacteria
that had not stuck to the corneocytes. Adherent bacteria were stained with
oxalated Crystal
Violet for 10 seconds. Finally, the discs were rinsed off with osmotically
purified water,
placed on a microscope slide and air-dried for 24 hours.
3) Image acquisition and bacterial counts
Adherent bacteria were photographed using an Olympus BX 53 microscope fitted
with a
digital camera. The images were then analysed using UTHSCSA Image Tool
software
(Reindeer Graphics Inc.). The mean number of bacteria per disc was calculated
from 10-12
images, corresponding to the surfaces of about 100 corneocytes. Means were
compared
using Statgraphics Plus software (Manugistics Inc.).
4) Results and conclusion
These test ,results show that xylitol inhibits the adhesion of S. aureus to
corneocytes by a
_____________________________________________________________ factor of about
20% (Table 2). Very strong inhibitory activity 50% or over was
observed with the HLB 16 sucrostearate and polysorbate 60 at final
concentrations of 1%
(Table 2). Polysorbate 20 and 60 at 0.1% gave very promising inhibitory
activity. In
contrast, sucrose laurate monoester induced a significant increase in
adhesion.
CA 02881267 2015-02-04
22
Table 2: Effect of xylitol and sugars of the invention on the adhesion of S.
aureus on
human corneocytes.
Inhibition of S. aureus adhesion (%)
PolyhYdroxylated compound (final
Test 1 Test 2
concentration)
Xylitol (0.5%) 21,0 20,2
HLB 16 sucrostearate (1%) 57Ø 49.6.
Sucrose laurate (0.1%) -22.8.
Polysorbate 60 (0.1%) 39.8.
Polysorbate 60 (1%) 54.
Polysorbate 20 (0.1%) 57.
Each sugar was tested in two independent experiments for xylitol and sucrose
stearate
(1%).
These ex vivo results therefore confirm the in vitro results, i.e. that the
presence of HLB 16
sucrostearate in its surroundings inhibits the adhesion of S. aureus to both a
solid matrix
like a cytology slide and to human corneocytes.
3/ EXAMPLES OF TOPICAL COMPOSITIONS
ATODERMTm STOP EVOLUTIVE
Product % by weight in the composition
HLB 16 sucrostearate (SURFHOPE 2Ø
C1816)
Lipid phase 11Ø
Other polyhydroxylated compounds 0.61.
Water Q.s. 100
=
CA 02881267 2015-02-04
23
ATODERMTm STOP CRISE
Product % by weight in
the composition
HLB 16 sucrostearate (SURFHOPE 2Ø
C1816)
Lipid phase 11Ø
Other polyhydroxylated compounds 0.61.
Anti-pruriginous agent 0.3.
Anti-inflammatory agent 0.5.
Water Q.s. 100
ATODERMTm STOP RECIDIVE
, Product % by weight in
the composition
HLB 16 sucrostearate (SURFHOPE 2Ø
C1816)
Lipid phase 11Ø
Other polyhydroxylated compounds 1.1.
Anti-pruriginous agent 0.3.
Vitamin PP 0.3.
Water Q.s. 100
CA 02881267 2015-02-04
24
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