Treatment of endothelial dysfunction in diabetic patients
United States Patent: 7,956,035
Issued: June 7, 2011
Inventors: Stroes; Erik S.
G. (Amsterdam, NL)
Assignee: CSL Limited
(Parkville, Victoria, AU)
Appl. No.: 11/712,404
Filed: March 1, 2007
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A method for the treatment of endothelial
dysfunction in a diabetic patient, including both diabetes induced
macrovascular disorders and diabetes induced microvascular disorders,
comprises administration, preferably parenteral administration, to the
patient of an effective amount of high density lipoprotein (HDL).
Description of the
FIELD OF INVENTION
This invention relates to a method for the treatment of endothelial
dysfunction in diabetic patients. In particular, this invention relates to
a method for improving endothelial function in treatment of disorders
which are related to endothelial dysfunction, both macrovascular and
microvascular, in diabetic patients.
BACKGROUND OF THE INVENTION
In the chronic disease diabetes mellitus (diabetes), the body loses the
ability to properly produce or respond to the hormone insulin so that
cells of the peripheral tissues fail to actively take up glucose from the
blood for use or storage. In the diabetic individual, the level of glucose
in the peripheral blood can become elevated (hyperglycaemia) and typically
remains so unless some form of intervention is employed (e.g.,
administration of exogenous insulin) to return glucose in the blood to
normal levels. Left unchecked, the hyperglycaemia of diabetic individuals
can result in shock, organ degeneration or failure (e.g., kidney failure,
blindness, nerve disease, cardiovascular disease), tissue necrosis (e.g.,
requiring foot amputation), and even death.
Two major forms of diabetes are type 1 and type 2 diabetes. Type 1
diabetes, which was previously known as insulin-dependent diabetes
mellitus (IDDM) or juvenile onset diabetes, is an autoimmune disease in
which the body destroys the insulin-producing .beta. cells (islet cells)
of the pancreas resulting in an absolute requirement for daily
administration of exogenous insulin to maintain normal blood glucose
levels. Type 1 diabetes usually is diagnosed in children and young adults,
but can occur at any age. Type 1 diabetes accounts for 5-10% of diagnosed
cases of diabetes.
By far the more prevalent form of diabetes is type 2 diabetes, which was
previously known as non-insulin-dependent diabetes mellitus (NIDDM). Type
2 diabetes was also previously known as adult-onset diabetes, however,
this form of diabetes is becoming increasingly prevalent in the growing
population of overweight and clinically obese children and young adults.
Type 2 diabetes accounts for approximately 90-95% of all diagnosed cases
of diabetes. Type 2 diabetes typically begins with insulin resistance, a
disorder in which the body's cells do not respond to insulin properly,
followed by a gradual loss on part of the pancreas to produce and secrete
insulin. Type 2 diabetes is associated with a variety of factors including
older age, obesity, family history of diabetes, history of gestational
diabetes, impaired glucose metabolism, physical inactivity, and various
races or ethnicities. Individuals with type 2 diabetes must attempt to
control their blood glucose level with careful diet, exercise and weight
reduction, and additional medications.
Major factors contributing to the pro-atherogenic state in diabetes,
particularly type 2 diabetes mellitus (DM2), include dyslipidemia,
hyperglycemia, hypertension, visceral obesity and insulin resistance
(1,2). Observational studies have clearly demonstrated the importance of
diabetic dyslipidemia in contributing to atherogenesis in diabetes,
illustrated by the fact that the correlation between low density
lipoproteins (LDL) as well as high density lipoproteins (HDL) versus
cardiovascular events outweighs that of fasting plasma glucose (3). Statin
intervention studies have revealed a clear benefit of statin treatment on
reduction of cardiovascular events in DM2 (4, 5); however, in spite of
this impressive achievement, the majority of DM2 patients will still
suffer from cardiovascular events even when using statins (6).
During the last two decades, endothelial dysfunction has emerged as one of
the earliest stages of atherogenesis. Endothelial dysfunction, which is a
hallmark in all diabetic patients (ie both type 1 and 2) has been shown to
have predictive value for future cardiovascular events (7-9). In line with
the multifactorial pathogenesis of diabetes-induced vascular disease (10,
11), numerous therapeutic interventions have been evaluated for their
potential to improve endothelial function in DM2 patients (9, 12).
Surprisingly, whereas endothelial function could be fully restored by
statin therapy in dyslipidemic patients (13), several studies have
demonstrated that even intensive statin treatment cannot normalize
vascular dysfunction in DM2 (14, 15). The latter emphasizes possibilities
for additional therapeutic modalities in this high risk group.
High-density lipoproteins (HDLs) represent a broad group of mostly
spheroidal plasma lipoproteins, which exhibit considerable diversity in
their size, apolipoprotein (apo) and lipid composition. HDL particles fall
into the density range of 1.063-1.21 g/ml (16) and as they are smaller
than other lipoproteins, HDLs can penetrate between endothelial cells more
readily allowing relatively high concentrations to accumulate in tissue
fluids (17). The major apolipoprotein of almost all plasma HDLs is apo
A-I, which in association with phospholipids and cholesterol, encloses a
core of cholesteryl esters (16). Nascent (i.e. newly synthesised) HDLs
secreted by the liver and intestine contain no cholesteryl esters and are
discoidal in shape (16). The negative association of plasma HDL
concentration with coronary artery disease has been well documented in
epidemiological studies (18). Although experiments in animals have
demonstrated an anti-atherogenic activity of HDLs (19), it is not yet
known whether this protective effect is related to the role of the
lipoprotein in reverse cholesterol transport or to a different mechanism.
The mechanism/mechanisms via which HDLs provide these cardioprotective
actions are not clearly understood, but may include a role for HDLs in
reverse transport of cholesterol from peripheral tissues to the liver,
inhibition of the oxidation of low-density lipoproteins, or modulation of
vasodilatation and platelet activation mediated by changes in the
production of prostacyclin (20). HDLs can also activate endothelial nitric
oxide (NO) synthase subsequent to its interaction with scavenger
In view of the emerging data on the NO promoting effects of HDL, compounds
with HDL-increasing capacity are of particular interest (21-24). Indeed,
in DM2 patients HDL is positively associated with endothelium-dependent
vasomotor responses (8). In work leading to the present invention, the
inventors have evaluated whether and to what extent HDL increase upon
infusion of exogenous reconstituted HDL (rHDL) would translate into an
improvement of vascular function. ApoA-I levels and endothelial function
were assessed both acutely (4 hours after infusion) as well as 7 days
after infusion of rHDL in DM2 and matched controls.
Bibliographic details of the publications referred to in this
specification are referenced at the end of the description. The reference
to any prior art document in the specification is not, and should not be
taken as, an acknowledgment or any form of suggestion that the document
forms part of the common general knowledge.
SUMMARY OF THE INVENTION
Throughout this specification and the claims which follow, unless the
context requires otherwise, the word "comprise", and or variations such as
"comprises" or "comprising", will be understood to imply the inclusion of
a stated integer or step or group of integers or steps but not the
exclusion of any other integer or step or group of integers or steps.
In one aspect, the present invention provides a method for the treatment
of endothelial dysfunction in a diabetic patient, which comprises
administration (preferably parenteral administration) to the patient of an
effective amount of high density lipoprotein (HDL).
In another aspect, the present invention provides the use of high density
lipoprotein (HDL) in the manufacture of a medicament for administration
(preferably parenteral administration) to a diabetic patient for the
treatment of endothelial dysfunction in the patient.
In yet another aspect, the present invention provides an agent for
administration (preferably parenteral administration) in the treatment of
endothelial dysfunction in a diabetic patient, which comprises high
density lipoprotein (HDL).
DETAILED DESCRIPTION OF THE INVENTION
Patients with diabetes, particularly type 2 diabetes mellitus (DM2), are
characterized by a marked increase in cardiovascular risk. Systemic
endothelial dysfunction, a hallmark in DM2, predicts future risk for
cardiovascular events. In view of the relation between HDL and the NO
pathway, the present inventors have evaluated the effect of rHDL infusion
on endothelial function in DM2. Specifically, in 7 DM2 patients and 7
normolipidemic controls, endothelial function was assessed using venous
occlusion plethysmography. Forearm blood flow (FBF) responses to
intra-arterial infusion of the endothelium-dependent and independent
vasodilators serotonin (5HT) and sodium nitroprusside, respectively, and
the inhibitor of nitric oxide synthase NG-monomethyl-l-arginine (L-NMMA)
were measured, both before, 4 hours after and 1 week after infusion of
rHDL (80 mg/kg based on protein).
At baseline HDL was similar in DM2 versus controls (1.1.+-.0.2 vs.
1.2.+-.0.3 mmol/L, ns). 5HT-induced vasodilation (max 17.+-.10%) and L-NMMA
induced vasoconstriction (max -17.+-.15%) were reduced in DM2 versus
controls (5-HT 114.+-.22 and L-NMMA -48.+-.5%, both p<0.05). rHDL infusion
raised apoA-I levels (1.2.+-.0.2 to 2.8.+-.0.4 vs. 1.2.+-.0.2 to
2.7.+-.0.4 g/L, p<0.01) in DM2 and controls, respectively and restored FBF
responses to 5HT (86.+-.22%, p<0.05) and L-NMMA (-45.+-.9%, p<0.01) in
DM2. This effect persisted 7 days after infusion (5HT; 80.+-.25%, p<0.05
and L-NMMA -37.+-.7%, p<0.01 compared to baseline). rHDL infusion had no
effect in controls. Accordingly, this work demonstrates that acute HDL
increase improves endothelial function in DM2 and that the improvement
persists for at least 7 days in spite of return-to-baseline of HDL
In one aspect, the present invention provides a method for the treatment
of endothelial dysfunction in a diabetic patient, which comprises
administration to the patient of an effective amount of high density
Preferably, the administration is parenteral administration.
Reference herein to "treatment" is to be considered in its broadest
context. The term "treatment" does not necessarily imply that a subject is
treated until total recovery. Accordingly, treatment includes amelioration
of the symptoms of a particular condition or disorder as well as reducing
the severity of, or eliminating a particular condition or disorder.
As used herein, references to "treatment of endothelial dysfunction" are
to be considered as references to improvement of endothelial function in
treatment of disorders which are related to endothelial dysfunction. Such
disorders include both macrovascular disorders (relating to the large
blood vessels) such as transient ischaemic attack, stroke, angina,
myocardial infarction, cardiac failure, and peripheral vascular disease,
as well as microvascular disorders (relating to the small blood vessels)
such as diabetic retinopathy (non-proliferative, proliferative, macular
oedema), microalbuminuria, macroalbuminuria, end stage renal disease,
erectile dysfunction, autonomic neuropathy, peripheral neuropathy,
osteomyelitis and lower limb ischaemia.
References herein to a "diabetic" patient are to be understood as a
reference to a patient suffering from either type 1 diabetes (DM1) or type
2 diabetes (DM2).
In accordance with the present invention, HDL is administered to a
diabetic patient. The term "HDL" as used herein relates to all forms of
high density lipoproteins and includes mature HDL, nascent HDL or
reconstituted HDL (rHDL) or any mixture thereof, as well as rHDL produced
from recombinant apolipoprotein or an analogue thereof with functional
relationship to nascent or reconstituted HDL. Such analogues include
functional peptides derived from the apolipoprotein (Apo) structure such
as those described in International Patent Publications Nos. WO 99/16459
and WO 99/16408, the contents of which are incorporated herein by
The high density lipoproteins comprise a protein component, and lipid. The
proteins are preferably apolipoproteins, e.g. human apolipoproteins such
as apolipoprotein A-I (apoA-I), apolipoprotein A-II (apoA-II) or
apolipoprotein A-IV (apoA-IV) or recombinant apolipoproteins, or
functionally homologous peptides with similar properties. Suitable lipids
are phospholipids, preferably phosphatidyl choline, optionally mixed with
other lipids (cholesterol, cholesterol esters, triglycerides,
sphingolipids, or other lipids). The lipids may be synthetic lipids,
naturally occurring lipids or combinations thereof.
Preferably, the HDL is reconstituted HDL.
Production of reconstituted HDL is described, by way of example, in U.S.
Pat. No. 5,652,339 and by Matz and Jonas (25) and Lerch et al. (26).
Production of rHDL with recombinant apolipoproteins is described, by way
of example, in European Patent No. EP 469017 (in yeast), U.S. Pat. No.
6,559,284 (in E. coli), and International Patent Publications Nos. WO
87/02062 (in E. coli, yeast and Cho cells) and WO 88/03166 (in E. coli).
The contents of each of these documents are incorporated herein by
The HDL is administered in an effective amount. An "effective amount"
means an amount necessary at least partly to attain the desired response,
or to delay the onset or inhibit progression or halt altogether, the onset
or progression of the particular condition or disorder being treated. The
amount varies depending upon the health and physical condition of the
individual to be treated, the racial background of the individual to be
treated, the degree of protection desired, the formulation of the
composition, the assessment of the medical situation, and other relevant
factors. It is expected that the amount will fall in a relatively broad
range that can be determined through routine trials.
Preferred HDL dosage ranges are from 0.1-200 mg, more preferably 10-80 mg,
HDL (weight based on apolipoprotein) per kg body weight per treatment. For
example, the dosage of HDL which is administered may be about 0.2-100 mg
HDL per kg body weight (weight based on apolipoprotein) given as an
intravenous injection and/or as an infusion for a clinically necessary
period of time, e.g. for a period ranging from a few minutes to several
hours, e.g. up to 24 hours. If necessary, the HDL administration may be
repeated one or several times. The actual amount administered will be
determined both by the nature of the condition or disorder which is being
treated and by the rate at which the HDL is being administered.
Preferably, the patient is a human, however the present invention extends
to treatment and/or prophylaxis of other mammalian patients including
primates, livestock animals (e.g. sheep, pigs, cattle, horses, donkeys),
laboratory test animals (e.g. mice, rabbits, rats, guinea pigs), companion
animals (e.g. dogs, cats) and captive wild animals.
In accordance with the present invention, the HDL is preferably
administered to a patient by a parenteral route of administration.
Parenteral administration includes any route of administration that is not
through the alimentary canal (that is, not enteral), including
administration by injection, infusion and the like. Administration by
injection includes, by way of example, into a vein (intravenous), an
artery (intraarterial), a muscle (intramuscular) and under the skin
(subcutaneous). The HDL may also be administered in a depot or slow
release formulation, for example, subcutaneously, intradermally or
intramuscularly, in a dosage which is sufficient to obtain the desired
Compositions suitable for parenteral administration conveniently comprise
a sterile aqueous preparation of the active component which is preferably
isotonic with the blood of the recipient. This aqueous preparation may be
formulated according to known methods using suitable dispersing or wetting
agents and suspending agents. The sterile injectable preparation may also
be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable
diluent or solvent, for example as a solution in a polyethylene glycol and
lactic acid. Among the acceptable vehicles and solvents that may be
employed are water, Ringer's solution, suitable carbohydrates (e.g.
sucrose, maltose, trehalose, glucose) and isotonic sodium chloride
solution. In addition, sterile, fixed oils are conveniently employed as a
solvent or suspending medium. For this purpose, any bland fixed oil may be
employed including synthetic mono- or di-glycerides. In addition, fatty
acids such as oleic acid find use in the preparation of injectables.
The formulation of such therapeutic compositions is well known to persons
skilled in this field. Suitable pharmaceutically acceptable carriers
and/or diluents include any and all conventional solvents, dispersion
media, fillers, solid carriers, aqueous solutions, coatings, antibacterial
and antifungal agents, isotonic and absorption delaying agents, and the
like. The use of such media and agents for pharmaceutically active
substances is well known in the art, and it is described, by way of
example, in Remington's Pharmaceutical Sciences, 18th Edition, Mack
Publishing Company, Pennsylvania, USA. Except insofar as any conventional
media or agent is incompatible with the active ingredient, use thereof in
the pharmaceutical compositions of the present invention is contemplated.
Supplementary active ingredients can also be incorporated into the
Other delivery systems can include sustained release delivery systems.
Preferred sustained release delivery systems are those which can provide
for release of the active component of the invention in sustained release
pellets or capsules. Many types of sustained release delivery systems are
available. These include, but are not limited to: (a) erosional systems in
which the active component is contained within a matrix, and (b)
diffusional systems in which the active component permeates at a
controlled rate through a polymer.
The present invention also provides the use of high density lipoprotein (HDL)
in the manufacture of a medicament for administration, preferably
parenteral administration, to a diabetic patient for the treatment of
endothelial dysfunction in the patient.
In yet another aspect, the invention provides an agent for administration,
preferably parenteral administration, in the treatment of endothelial
dysfunction in a diabetic patient, which comprises high density
Claim 1 of 11 Claims
1. A method for the treatment of
endothelial dysfunction in a Type 2 diabetic patient, which comprises
administering to a Type 2 diabetic patient suffering from endothelial
dysfunction associated with a diabetes induced microvascular disorder
selected from the group consisting of diabetic retinopathy (non-proliferative,
proliferative, macular oedema), microalbuminuria, macroalbuminuria, end
stage renal disease, erectile dysfunction, autonomic neuropathy,
peripheral neuropathy, osteomyelitis and lower limb ischaemia, an amount
of high density lipoprotein (HDL) effective to treat said endothelial
dysfunction and improve endothelial function.
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