Note: Descriptions are shown in the official language in which they were submitted.
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Treatment of cytokine mediated conditions
Field of invention
The present invention refers to the use of certain sulfated glycosaminoglycans
for the
prevention and treatment of cytokine mediated conditions of the epithelial and
endothelial linings.
Background
Glycosaminoglycans (GAGs) constitute a highly heterogeneous class of large
macromolecules composed of repeating disaccharide units. In general, each of
the
repeating units comprises an aminosugar, either glucosamine or galactosamine,
and
an uronic acid or a hexose, either glucuronic acid, iduronic acid or
galactose. The
hydroxyl group at C2 of the uronic acid and the hydroxyl groups at C3, C4, C6
and the
amino group on C2 on the aminosugar may be substituted by sulfate groups. The
GAGs comprise the following polymers: heparin, heparan sulfate (HS), dermatan
sulfate (DS), chondroitin sulfate (CS), keratan sulfate and hyaluronan.
Heparin is a
GAG isolated on a commercial basis from animal tissues and used in the clinic
as an
antithrombotic drug. Naturally occuring heparin consists of unbranched,
negatively
charged polysaccharide chains of varying lengths, and molecular weights.
Chains of
molecular weight from 5 to over 40 kilodaltons (kDa), make up polydisperse
pharmaceutical-grade heparin.
Heparin derived from natural sources, mainly porcine intestine and bovine
lung, can
be used to prevent thrombosis. However, the effects of natural, or
unfractionated
heparin can be difficult to predict. After a standard dose of unfractionated
heparin,
coagulation parameters must be monitored very closely to prevent over- or
under-
anticoagulation.
Numerous brands of heparins and low molecular weight heparins (LMWH), such as
tinzaparin and dalteparin, are available for the treatments that rely on their
anti-
coagulant activity. LMWHs have been demonstrated to be at least as effective
and
safe as unfractionated heparin. Beside therapies relying on the anticoagulant
effect of
CA 02779838 2012-06-14
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heparin, other uses can be envisioned.
Heparin and LMWH have been suggested to be important agents in the treatment
of
cancer and the anti-tumor effect appears to be partially unrelated to their
anticoagulant effect. In Pathophysiology Haemost Thromb, 2007-08; 36; 195-203,
B
Casu et al. describe a modified non-anticoagulant heparin and explain its
dominant
anticancer mechanism as exerted through competition with functions of HS
chains in
HS proteoglycans associated with cell surfaces and extracellular matrices. In
this
context it is discussed that these heparins also effectively would inhibit
tumor cell
heparanases in experimental metastasis. Further they exert antimetastic action
by
disrupting selectin mediated tumor cell-vasular interactions and inhibition of
growth
factors. The authors also suggest that the heparin mediated release of tissue
factor
pathway inhibitor (TFPI) may explain their anti-angiogenetic and anti-
metastatic
effect, while attributing this effect to NS regions of heparin and suggesting
that a
minimum chain length of heparins is expected to accomplish the effect. In
Clinical
and Applied Thrombosis/Hemostasis, 2012; 000(00); 1-7, SA Mousa compares the
antiangiogenesis activity of the two LMWHs tinzaparin and dalteparin. It is
discussed
that the molecular weight profile is important for the release of TFPI and
concluded
that fragments within 2-8 kDa appear to primarily be responsible for increases
in
TFPI plasma levels. It is further concluded that tinzaprin induces higher
levels of TFPI
due to its relatively higher molecular weight fractions and higher
sulfate/carboxylate
ratio. It remains, however, obvious that heparin, LMWH and non-coagulant
modified
heparins exert its anti-tumor effects through different mechanisms and that
the field is
far from elucidated. US 8,071, 569 (Mousa) describes a supersulfated, oxidized
heparin fraction with angiogenesis inhibition characteristics that retains
certain
anticoagulant properties.
Another type of modified heparins with an anti-tumour effect is described by
JW Park
et al in Journal of Controlled Release, 2010; 148; 317-326. These heparin
derivatives
are prepared by conjugating a LMWH (Fraxiparin 4,5 kDa) with deoxycholic acid
(DOCA) in order to reduce their anticoagulant activity and establish an orally
available preparation. The derivatives are suggested to retain the capacity of
heparin
and HS bind to factors important for the angiogenic process and specifically
to
reduce the angiogenic activity of FGF2 and VEGF. It is further generally
discussed
CA 02779838 2012-06-14
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herein that there are structural requirements necessary for obtaining the
interactions
between heparins and the growth factors related to the capability of
electrostatic and
hydrophobic interactions.
EP 710483 describes low molecular weight heparins and other sulfated
glycosaminoglycans for prevention of structural and functional alterations of
the
peritoneal membrane in patients suffering from renal insufficiency who thereby
risk
peritoneal loss of proteins and compromised transport of urea and creatinine.
Heparin and LMWH also have well documented anti-inflammatory properties and
efficacy against sepsis and shock which suggest direct anti-inflammatory
activity
rather than anticoagulant properties, see AD Cornet et al in Thromb Heaemost
2007;
98; 579-586. This article refers to studies that indicate that a non-
anticoagulant
heparin prevents from vascular endothelial dysfunction during hyperdynannic
sepsis.
The non-anticoagulant heparins are further described in US 5,583,121 (Chaundry
et
al) who describe 0-desulfated heparins or heparin fragments in the treatment
of
hypovolemic shock.
Heparin is reported to protect against protein-losing enteropathy (PLE), a
life-
threatening complication developed from different unrelated diseases, such as
Crohn's diseases and complications following the Fontan procedure, see L Bode
et al
The Journal of Biological Chemistry, 2006; 281; 7809-7815. It is reported that
loss of
HS from the basolateral surface of intestinal epithelial cells has an
important role in
the development of PLE, while an increased availability of cytokines appears
to
further contribute to aggravate the complication. Heparin therapy is concluded
to
compensate for the HS loss and also to inactivate cytokines like IFNy and TNFa
and
restore the intestinal epithelium from protein leakage. KM Bhagirath et al in
Ann
Thorac Surg, 2007; 84; 2110-2 compares LMWH with unfractionated heparin for
the
treatment in PLE in the context of an effective treatment with LMWH
(dalteparin). It is
discussed that LMWH is less ionic than unfractionated heparin and has less
propensity to incorporate itself into epithelial cells which may question the
need of
heparin sulfate binding to epithelial cells of the intestine.
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Furthermore, N Brodszki et at in Acta Paediatrica, 2011; 100; 1448-1453,
report a
successful treatment of Gorham-Stout syndrome with the low anticoagulant
heparin
tafoxiparin. In the report comparisons are drawn with earlier therapies with
heparin
for treating PLE.
It is apparent that the role of heparin and LMWH in treating or preventing
from protein
leakage in the different discussed complications is not fully understood and
it is not
clarified to what extent they exert similar mechanisms as in the context of
inhibiting
angiogenesis in tumor treatment or when counteracting inflammation. It is also
evident that there is no clear picture of the requirements of a heparin
derivative that
effectively would replace heparin, being an even more suitable therapy with
less of
the side-effects normally associated with heparin. For the treatment or
prevention of
protein leakage from the endothelium or the epithelium as precipitated by the
underlying discussed diseases, it would be highly advantageous if an effective
LMWH agent was available with low or negligible anticoagulant effect. Such an
agent
should also exhibit a suitable bioavailability, at least comparable to that of
tinzaparin
or dalteparin, and retain other usual advantages compared to unfractionated
heparin.
It would also be highly desirable to obtain such a heparin derivative that is
perfected
to both treat protein leakage and inhibit endothelial proliferation (such as
angiogenesis) with consideration to the multiple outlined mechanisms involved
in the
activity of heparin or LMWH. Accordingly, there is demand for a low
anticoagulant
heparin with a highly favorable molecular weight distribution of the
polysaccharide
chains and with a favorable content of charged groups on the chains for the
purpose
of treating both protein leakage and treat abnormal endothelial proliferation
associated with tumors and also other degenerative diseases wherein inhibition
of
endothelial proliferation is required, such as diabetic retinopathy and
macular
degeneration.
Description of the invention
Before the present invention is described, it is to be understood that the
terminology
employed herein is used for the purpose of describing particular embodiments
only
and is not intended to be limiting, since the scope of the present invention
will be
limited only by the appended claims and equivalents thereof.
CA 02779838 2012-06-14
It must be noted that, as used in this specification and the appended claims,
the
singular forms "a," "an," and "the" include plural referents unless the
context clearly
dictates otherwise.
Also, the term "about" is used to indicate a deviation of +1- 2 % of the given
value,
5 preferably +1- 5 %, and most preferably +1- 10 % of the numeric values,
where
applicable.
In the context of the present invention the term "cytokine mediated condition
of the
epithelial or endothelial linings" relates to a complication or condition
wherein the
integrity or functionality of the epithelium or endothelium is impaired and
the activity
of cytokines place an important role in the deteriorative process which
becomes
manifested for example as leakage of proteins. Alternatively, the cytokine
mediated
complication relates to abnormalities in the endothelium generation including
abnormal proliferation, expressed for example as angiogenesis associated with
tumor
growth and also other diseases such as diabetic retinopathy and macular
degeneration, especially the wet form of age-related macular degeneration.
In the context of the present invention the term "protein leakage" relates to
extravasation of fluid and proteins over the endothelial cell layer, as for
example is
seen in sepsis, or transepithelial leakage of proteins and fluid, as for
example is seen
in PLE.
The invention generally relates to the usefulness of least one sulfated GAG
selected
from the group consisting of HS, depolymerised HS, DS, depolymerised DS, LMWH,
and depolymerised heparin having an anti-factor Xa activity of 200 Ill/mg or
less for
treating of or preventing at least one cytokine mediated condition of the
endothelial or
epithelial linings.
The present invention further relates to a method for treating or preventing
at least
one cytokine mediated condition of the endothelial or epithelial linings
comprising
administering a sulfated GAG selected from the group consisting of HS,
depolymerised HS, DS, depolymerised DS, LMWH, and depolymerised heparin
having an anti-factor Xa activity of 200 IU/mg or less.
CA 02779838 2012-06-14
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The cytokine-mediated condition of the endothelial or epithelial linings in
particular
relates to abnormalities in such issues, as expressed by loss of integrity and
leakage.
Thereby, present invention aims at preventing or treating such complications
which
have severe consequences for surrounding organs or tissues. In one aspect,
protein
leakage is linked to vascular dysfunctions related to endothelial
proliferation (such as
angiogenesis).
Protein leakage from endothelial and epithelial linings occurs in many
different
disorders. Non-limiting examples of disorders wherein protein leakage is a
complication are Gorham Stout Syndrome, lymphangioma, sepsis, PLE, and
nephrosis.
Likewise endothelial proliferation (e.g. angiogenesis) occurs in several
different
disorders. Non-limiting examples of disorders wherein endothelial
proliferation
leading to dysfunctional neovascularization are cancer, diabetic retinopathy,
Gorham
Stout syndrome and lymphangioma and macular degeneration.
Accordingly the presently invented treatments with sulfated GAGs is in an
aspect
directed to treatment of complications wherein proliferating vessels are
dysfunctional
as manifested by defective retaining capacity
The present invention also relates to the prevention and treatment of protein
leakage
as a part of a treatment of Gorham stout syndrome, lymphangioma, sepsis, and
PLE.
The present invention further relates to the prevention and treatment
ofcomplications
of cancer arriving from neovascularization and dysfunctional vessels,
treatment of
diabetic retinopathy, Gorham Stout syndrome and lymphangioma.
HS, DS and CS, are composed of alternating hexosamine and uronic acid
residues.
The presence of D-glucuronic acid (GIcA) and its C-5 epimer L-iduronic acid
(IdoA)
and the specific sulfation of hexosamines and uronosyl residues endow the
polymer
an extreme structural variation. The structure is built on repeating
disaccharides
containing from none or very few to nearly 100% iduronic acid-containing
disaccharides. The organization of GIcA-and IdoA-N-hexosamine containing
CA 02779838 2012-06-14
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disaccharides can vary from long blocks to an alternating disaccharide
pattern. The
variation of sulfation and the degree of IdoA sulfate generates a wide variety
of
biological activity. There are different well-defined polysaccharides of DS,
CS, HS
and depolymerised heparin.
The glucosamine of the repeating building blocks of HS is either N-acetylated
or N-
sulfated, and their presence is arranged mainly in a segregated manner. HS has
ubiquitous distribution on cell surfaces and in the extracellular matrix. It
is generally
less sulfated and has a lower IdoA content than heparin and has a more varied
structure. Interactions between HS and proteins are implicated in a variety of
physiological processes, such as cell adhesion, cell proliferation, enzyme
regulation,
cytokine action, virus entry and anticoagulant properties. HS possesses
anticoagulant activity depending on the presence of a specific anticoagulant
pentasaccharide, however considerably less than heparin. Heparan sulfate is a
linear
polysaccharide which can be prepared from porcine intestinal mucosa or from
bovine
lung, from heparin side fractions using cetylpyridinium chloride fractions and
sequential salt extraction as described by Fransson et at., Structural studies
on
heparan sulphates, Eur. J. Biochem. 106, 59-69 (1980).
CS is a linear polysaccharide consisting of alternating glucuronic acid and N-
acetyl-
galactosamine residue, the latter being sulfated at either C4 or C6 or both.
They can
be prepared from bovine trachea or nasal cartilage. CS is of importance for
the
organization of extracellular matrix, generating an interstitial swelling
pressure and
participating in recruitment of neutrophils.
DS is a linear polysaccharide consisting of alternating uronic acid and N-
acetyl-
galactosamine residue. The uronic acids are either GIcA or IdoA and the
disaccharide can be sulfated at C4 and/or C6 on the galactosamine and at C2 of
the
IdoA. DS can be prepared from porcine skin and intestinal mucosa and possesses
biological activities such as organization of extracellular matrix,
interactions with
cytokines, anti-coagulant activities and recruitment of neutrophils.
LMWH or depolymerized heparins are linear oligosaccharides mainly consisting
of
alternating N-sulfated glucosamine and IdoA residue and often containing the
CA 02779838 2012-06-14
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anticoagulant pentasaccharide. They can be prepared from heparin by specific
chemical cleavage. Their main clinical function is to inhibit factor Xa,
resulting in an
antithrombotic effect. It is proposed to have antimetastatic properties.
Fragmin
(Pfizer, USA) is an example of a LMWH obtained by controlled depolymerisation
of
heparin and having an antithrombotic effect owing to inhibition of factor Xa.
Heparin
fragments having selective anticoagulant activity, as well as methods for the
preparation thereof, are described in US patent number 4,303,651. According to
the
European Pharmacopoeia (PharmEur) a heparin, in order to be called a LMWH (low
molecular mass heparin), should have an antifactor Xa activity not less than
70
IU(International Unit)/mg and an Mw of less than 8 kDa.
Accordingly, the GAGs administered in methods and in compositions of the
present
invention preferably have a weight average molecular weight (Mw) of 30 kDa or
less,
preferably less than 20 kDa, more preferably 10 kDa or less, even more
preferably
not higher than 8 kDa and most preferable not higher than 7 kDa.
Preferably, the GAG belongs to the group consisting of depolymerized heparins
having an average molecular weight below 20 kDa, preferably below 10 kDa, more
preferably not higher than 8 kDa and most preferably not higher than 7 kDa.
For the purpose of treatment or prevention of at least one cytokine mediated
condition of the endothelial or epithelial linings, the GAG of the present
invention has
an anti-factor Xa activity of 200 KU/mg or less, preferably less than 150
IU/mg, more
preferably less than 100 IU/mg, more preferably less than 70 Ili/mg, more
preferably
less than 30 IU/mg, and most preferable less than 10 Ili/mg.
The anticoagulant activity of heparin, LMWH and other heparin derivatives is
often
measured as their ability to potentiate the inhibition of coagulation factor
Xa and
factor Ila by antithrombin. Methods for measuring anti-factor Xa- and anti-
factor Ila
activity are well known to the skilled person and are also described in
pharmacopoeias such as the European pharmacopoeia (Pharm Eur) and the United
States pharmacopoeia (USP).
CA 02779838 2012-06-14
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The anticoagulant activity can be abrogated by for example selective periodate
oxidation (see e.g. Fransson LA, and Lewis W, Relationship between
anticoagulant
activity of heparin and susceptibility, to periodate oxidation, FEBS Lett.
1979, 97:
119-23; Lindahl et al.and Proc Natl Acad Sci USA, 1980; 77(11):6551-6555).
In one embodiment the disaccharide structure of the GAG is essentially devoid
of
non-sulfated monosaccharide glucuronic and iduronic units and having an
antifactor
Xa activity of 10 IU/mg or less.
In another embodiment the GAG is a LMWH with an antifactor Xa activity of 10
IU/mg
or less and an average molecular weight not higher than 8 kDa, and preferably
not
higher than 7 kDa. A suitable such LMWH has an anticoagulant activity (< 10
IU/mg
by pharmacopoeia! anti-factor Xa- and anti-factor ha assays) and an weight
average
Mw of 5-7 kD, while having a distribution of polysaccharides and their
corresponding
molecular mass in cumulative % according to the table below.
Molecular mass, Cumulative weight,
kDa %
>15 0
>10 6-15
>9 7-20
>8 13-23
>7 18-32
>6 26-42
>5 40-53
>4 52-68
>3 >70
>2 >85
It is of desirable that the GAGs to be used in the present invention comprise
a
suitable distribution of chains with molecular weight in the range of 4-7 kDa
while
retaining chains differently sized outside the range. This can preferably be
obtained
by purposeful depolymerisation and fractionation.
CA 02779838 2012-06-14
It is also important to retain the distribution of clusters of negatively
charged groups
of the chains as have been found important in the mode of action in native
unfractionated heparin. This is preferably accomplished by a selective process
to
eliminate the pentasaccaharide sites of the chain associated with the
anticoagulant
5 effect.
Without being bound to theory, it has been hypothesized that the GAGs as
defined
above and administered according to the present invention will bind to the
cell
surface and decrease the ability of proteins to pass through the cellular
barrier and
thereby treat or prevent protein leakage. It is further hypothesized that the
GAGs as
10 defined above bind factors such as cytokines and growth factors (such as
VEGF) and
thereby modulate the activity of these factors. Another such factor, heparin-
binding
protein (HBP, azurocidin) is involved in endothelial leakage and was recently
suggested to be the prime marker of early sepsis. Given the fact that HBPs
seem to
be involved in both the pathological angiogenic process in for example Gorham
stout
syndrome and in conditions with leaky vessels (HBP), and that loss of heparan
sulfate can lead to leakage of proteins over the intestinal epithelium, the
present
inventors postulate that giving heparin in conjunction with more conventional
therapies, would be beneficial and that the GAG will, for example, bind to the
proliferating factors (eg VEGF), and thereby slow the process down.
One non-limiting way of obtaining a sulfated GAG to be used according to the
invention is to treat a sulfated GAG, such as heparin (e.g. pig intestinal
mucosa
heparin) by periodate oxidation followed by alkaline 8-elimination of the
product. This
process leads elimination of the anticoagulant activity and depolymerization.
The process disclosed in US Patent 4,990,502 (Lormeau et al) demonstrates one
way of treating native heparin to selectively cleave residues of the
pentasaccharide
residues responsible for the anticoagulant effect and a following
depolymerization
that results in a low anticoagulant, LMWH with a an average molecular weight
5,8 to
7,0 kDa.
The invention further relates to a parenteral composition of least one
sulfated GAG
as described above for treating of or preventing at least one cytokine
mediated
condition of the endothelial or epithelial linings.
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In the described parenteral compositions the active compounds can be
incorporated
into a solution or suspension, which also contain one or more adjuvants such
as
sterile diluents such as water for injection, saline, fixed oils, polyethylene
glycol,
glycerol, propylene glycol or other synthetic solvents, antibacterial agents,
antioxidants, chelating agents, buffers and agents for adjusting the
osmolarity. The
compositions can be delivered in ampoules, vials, disposable syringes or as
infusion
arrangements, also for self administration.
Encompassed by the present invention is any combination with the disclosed
embodiments.
The invention will be further disclosed in the following non-limiting examples
Examples
Tafoxiparin is a depolymerised heparin that is essentially deprived of its
anticoagulant
activity (< 10 Ill/mg by pharmacopoeia! anti-factor Xa- and anti-factor Ila
assays).
The weight average Mw is 5-7 kDa.
Table I. Distribution of polysaccharides and their corresponding molecular
mass in
cumulative % of weight for several batches of Tafoxiparin
Molecular mass, Cumulative weight,
kDa
>15 0
>10 6-15
>9 7-20
>8 13-23
>7 18-32
>6 26-42
>5 40-53
>4 52-68
>3 >70
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>2 >85
The pharmaceutical preparation of DF01 is a solution for subcutaneous
injection, 8
mL dispensed in glass vials sealed with a rubber stopper and covered with a
tear-off
aluminum cap.
CASE DESCRIPTIONS OF TREATMENT OF GORHAM STOUT SYNDROME
P1, a 2.5-year-old boy, with an unremarkable family history and previously
healthy
except for nonallergic asthma, was referred to the clinic for the chest tube
drainage of
an extensive and progressive pleural effusion detected when the patient was
hospitalized for pneumonia. Computed tomography (CT) had shown extensive
amounts of pleural effusion, atelectasis and emphysematous areas in the right
lung.
A tube thoracostomy was performed with direct exchange of 700 mL of whitish
fluid,
with triglyceride levels at 6 mM (reference value for plasma 1.8 mM),
indicative of
chylothorax. A new chest CT was performed the next day showing regression of
the
pleural effusion but, in addition, it detected multiple skeletal, possibly
lytic,
abnormalities in several vertebrae, the pelvis, and in the right clavicle as
well as
rounded changes within the spleen. During the following days, extensive
pleural
drainage, infection, malignancy and immunodeficiency were ruled out. A bone
marrow biopsy and a fine needle biopsy of one of the lytic skeletal changes
were
inconclusive. The patient was started on total parenteral nutrition (TPN).
Open biopsy
of one of the lytic bone changes revealed lymphatic vessels dissecting the
bone,
resulting in the zones of bone resorption and marrow hypoplasia. The
pathological
changes were consistent with Gorham¨Stout disease, or disseminated
lymphangiomatosis. Treatment with interferon a-2b (IntronA, Schering-Plough
Sweden, Stockholm, Sweden) was started at a dose of 2 million unitsAn2 body
surface, every second day. A prompt response with reduced pleural fluid
drainage
could be seen, within a few days. A repeat chest CT showed clearing of the
chylothorax, but a pericardial effusion appeared and was confirmed by
echocardiography. The pericardial effusion was successfully managed
conservatively
with diuretic therapy. The patient was discharged but returned after 15 days
owing to
increasing pericardial and pleural fluid, despite continued interferon a-2b
and diuretic
therapy. His clinical condition quickly deteriorated, despite continuous
positive airway
CA 02779838 2012-06-14
13
pressure treatment. The patient was sedated and intubated. The lymph leakage
continued, some days exceeding 6 V24 h or 545 mVkg/24 h. TPN was restarted
and the interferon a-2b therapy was intensified with doses daily instead of
every
second day. Explorative thoracoscopy was performed revealing large amounts of
gelatinous fluid and thick mesh-like tissue in the pleural compartments, and a
pleural
biopsy was taken. The large drainage volumes continued and tafoxiparin (low
anticoagulant, LMWH) was started. The patient received a total of 3 weeks
treatment
with tafoxiparin. Because no changes in the drainage volumes were observed,
chest
radiation therapy (RT) was performed with 15 Gy given in 10 fractions over 2
weeks,
as suggested by the current literature. Tafoxiparin and interferon a-2b were
paused
during the RT. In spite of all the aforementioned efforts, the chest tube
drainage
volumes remained unaltered and ultimately surgical intervention was attempted
after
thoracoscopy revealed, surprisingly, that the mesh-like tissue had completely
resolved by this time. A resection of the right middle lobe with concomitant
pleurodesis and thoracic duct ligation was performed; the lung specimen showed
lymphangiogenesis. This operation finally stemmed the fluid leakage, and the
patient
could subsequently return to his home clinic for continued interferon a-2b
therapy.
Four years after the treatment, he is clinically stable with no breathing
problems and
without any new symptoms.
P2, a 4-year-old girl with an unremarkable family history and in good prior
health,
started complaining about nonspecific back pain approximately 8 months before
her
first admission. She was admitted to the local hospital after mild trauma to
the thorax,
when an X-ray of her chest showed substantial bilateral pleural effusions with
the
compression of the inferior vena cava and several fractured ribs. Routine
blood tests
and analysis of the pleural exudates (shown to be chylous) revealed no
infection or
malignant cells. A CT scan confirmed, besides massive chylothorax, several rib
fractures, fracture of the sternum and diffuse osteolytic changes in the
humerus,
femur, pelvis, sacrum and multiple vertebrae. Also, poorly defined changes
were
noted in the spleen. A chest tube was inserted and the amount of drained
pleural
fluid averaged between 1400 and 2000 mV24 h. The patient was started on TPN.
The clinical picture suggested lymphangiomatosis, and treatment with pegylated
interferon a-2b (Peglntron, Schering-Plough Sweden, Stockholm, Sweden) was
initiated (weekly injections of 1 Ig, later 0.5 Ig/kg bodyweight). She
received a 7-day
CA 02779838 2012-06-14
14
course of continuous intravenous infusion of octreotide, 1 Ig/kg bodyweight/h.
Despite the treatment, no remission of the chylothorax was observed during 2
months of treatment and P2 was referred to our unit. Biopsies from pleurae and
ribs
confirmed the diagnosis of lymphangiomatosis. A cytokine profile of the fluid
was
determined. Peglntron was switched to subcutaneously administered IntronA at a
dose of 2 million units/ft-12 body surface, every second day. Furthermore,
tafoxiparin
was included in the treatment. The patient underwent RT directed towards all
affected areas; 15 Gy was given in 10 fractions over 2 weeks in two sessions,
4
weeks apart to allow bone marrow recovery. Interferon treatment was paused
during
RT. During the 4 weeks between the RT blocks, she was treated with filgrastim
(Neupogen, Amgen AB, Solna, Sweden) for the purpose of collecting CD34+ cells,
should an autologous rescue stem cell transplant be necessary. She responded
adequately to this treatment regimen. After the second round of RT, interferon
was
administered at an intensified dosage, with daily injections instead of every
other day.
Her clinical status improved steadily. After approximately 2.5 months, the
pleural
drain could be removed. Tafoxiparin was stopped after a total of 3 months'
treatment.
She continues with daily injections of IntronA. An MR control at 3 months
after
discharge showed clear improvement on the previously noted changes in the
lungs
and sen, and moreover, a reduction in the lytic changes in the bones was seen.
Six
months postdischarge, the patient is doing remarkably well. Measurements of
selected cytokine levels are shown in Table 2.
CA 02779838 2012-06-14
. 15
,
. ,
,
Table 2 Measurement of cytokines, using specific ELISA kits, from serum or
pleural
effusion (PE) taken from Pi and P2 at afferent states of disease activity
values outside
the reference intervals are indicated in bold
State of disease
Source Cytokine
Maximal activity Remission Reference interval
P1 ¨ serum VEGF-A (ng/L) 730 570 62-707
VEGF-C (ng/L) 4050 3910 2459-6651
P2¨ serum VEGF-A (ng/L) 1200 370 62-707
VEGF-C (ng/L) 6930 2260 2459-6651
IL-5 (ng/L) <8 16
<8
IL-12 (ng/L) <7,8 <8
<16
IL-18 (ng/L) 140 630
140-500
IL-10 (ng/L) <5 <5
<9
IL-1fl (ng/L) <5 <5
<5
IL-6 (ng/L) 7 13
<8
IL-8 (ng/L) <5 12
<60
sIL-2R (kU/L) 479 1030
<700
TNF-a (ng/L) 11 21
<15
IL-4 (ng/L) <16 ¨
<100
P2¨ PE VEGF-A (ng/L) 220
¨ nk
VEGF-C (ng/L) 200 ¨ nk
IL-5 (ng/L) <8 ¨
nk
IL-12 (ng/L) <7,8 ¨
nk
IL-10 (ng/L) <5 ¨
nk
IL-1fl (ng/L) <5 ¨
nk
IL-6 (ng/L) 2828 ¨
nk
IL-8 (ng/L) 559 ¨
nk
sIL-2R (kU/L) 530 ¨
nk
TNF-a (ng/L) 23 ¨
nk
IL-4 (ng/L) <16 ¨
nk
Discussion
CA 02779838 2012-06-14
16
=
Given the fact that heparin binding proteins seem to be involved in both the
pathological lymphangiogenetic process in GSS (e.g. IL- 8, VEGF) and in
conditions
with leaky vessels (HBP), and that the loss of HS or HSPG ??? can lead to the
leakage of proteins over the intestinal epithelium, we postulated that giving
the
patients with GSS heparin in conjunction with more conventional therapies
(discussed later on) would be beneficial. However, our initial patient lost up
to 5 L of
lymph daily, corresponding to approximately 50% of his body weight, which had
to be
replenished by the massive infusions of fresh frozen plasma (FFP), blood and
albumin. We thus chose not to give any drug that could possibly further worsen
his
coagulation status. Heparin and LMWH can, however, be enzymatically or
chemically
modified to selectively almost completely lose its anticoagulant properties
while
keeping most of its charge and hence ability to bind cationic proteins. One
such
preparation is tafoxiparin, a periodate-oxidized, LMWH currently used in a
clinical
obstetric study The Swedish Medical Products Agency (Lakemedelsverket)
approved
its usage in our patients owing to the extreme situation, despite no available
data on
its use in children. The anticipated risks were HIT (heparin-mediated
thrombocytopenia), an allergic reaction or coagulation disturbances. We first
administered a test dose and then choose to give a dose corresponding to 2500
EATI2 twice daily, subcutaneously, in accordance with the doses of heparin
used in
PLE.
The mortality of patients afflicted with Gorham Stout syndrome and the
condition
chyclothorax is high, about 70 % (although based on a small number of
patients).
Thus, there is of great importance to improve or find alternative treatments
for these
patients. The case reports exemplified herein present an interesting
alternative or
supplemental therapy to the currently applied treatment regimen.
Although particular embodiments have been disclosed herein in detail, this has
been
done by way of example for purposes of illustration only, and is not intended
to be
limiting with respect to the scope of the appended claims that follow. In
particular, it is
contemplated by the inventor that various substitutions, alterations, and
modifications
may be made to the invention without departing from the spirit and scope of
the
invention as defined by the claims.
