Note : Les descriptions sont présentées dans la langue officielle dans laquelle elles ont été soumises.
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Treatment of Metabolic Syndrome
The invention relates to the treatment of metabolic syndrome.
S Metabolic syndrome is a cluster of health problems rather than a single
disease.
Multiple interrelated abnormalities in glucose and lipid metabolism result in
insulin-resistant hyperglycemia, hyperinsulinemia, a high triglyceride (TG)
level, a low
high-density lipoprotien cholesterol (HDL-C) level and abdominal or visceral
obesity.
The abnormalities seem to be connected by insulin resistance, that is a
reduced
sensitivity of the tissues of the body to the action of insulin, caused by
visceral obesity.
The body compensates for this state by secreting more insulin from the
pancreas, hence
causing hyperinsulinemia. Type II diabetes can develop when the pancreas is
unable to
sustain this level of insulin secretion. The high insulin levels in the blood
also
contribute to abnormalities in blood lipids, that is triglycerides and
cholesterol.
The consequences of metabolic syndrome are perhaps more important than its
causes.
The syndrome is characterised by glucose intolerance; abnormal cholesterol and
triglyceride levels, in particular there is usually a reduction in HDL-C; and
upper body
obesity. All of these features are independent high rislc factors for cardiac
disease.
Individually each substantially increases the risk of cardiac disease, but in
combination
the effect is dramatic. The result is a great increase in the risk of
hypertension,
atherosclerotic changes and myocardial infarction. As well as cardiac
diseases, other
resulting conditions include, as mentioned, Type II diabetes which can have
such
resultant problems as microangiopathy, neuropathy, retinopathy and
nephropathy; and
Polycystic Ovaryian Syndrome (PCOS) in women. PCOS is characterised by ovarian
cysts, high androgen levels, hirsuitism and infertility. Women with PCOS have
an
increased risk of endometrial cancer.
Striking similarities exist between the metabolic syndrome and untreated
growth
hormone (GH) deficiency due to pituitary or hypothalamic pathologies. The most
central findings in both these entities are abdominal obesity and insulin
resistance.
These similarities indicate that a reduced action of GH may be of importance
in the
pathogenesis of metabolic aberrations not only in GH deficiency but also in
metabolic
syndrome. In fact, there is a striking evidence for disturbances in the
neuroendocrine
3S regulation of GH secretion in obesity. With an increase in body weight, GH
secretion
becomes blunted with a decrease in the mass of GH secreted per burst without
any
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major impact on secretozy burst frequency. Furthermore, serum levels of IGF-I
are
inversely related to the percentage of body fat, in particular to the amount
of visceral
adipose tissue.
In patients with GH deficiency, the beneficial effects of replacement therapy
with
recombinant human GH (rhGH) on most of the clinical features of the adult GH
deficiency syndrome are well established. These effects include the reduction
of body
fat, the improvement in risk factors for cardiovascular disease, and the
normalization of
other metabolic alterations. Johannsson et al have recently provided first
evidence that
the beneficial effects of rhGH therapy might also be achieved in abdominally
obese
patients without evidence of pituitary disease. In this placebo-controlled
trial, treatment
of viscerally obese males with rhGH over a period of 9 months resulted in a
reduction
of abdominal fat mass, as well as in an improvement of glucose and lipid
metabolism.
The use of rhGH in patients with central obesity, however, may be limited by
an
increase in insulin resistance which is known to occur during the early course
of rhGH
therapy. This may be of particular importance in metabolic syndrome patients
who are
characterized by a highly increased risk to develop type 2 diabetes.
Present treatment for metabolic syndrome involves lifestyle changes and
pharmacological treatment with drugs such as metformin. Metformin is a
biguanide,
which improves glycaemic control by enhancing insulin sensitivity in the liver
and in
muscles. It does not stimulate insulin secretion.
It is clear that metabolic syndrome is an important disorder which can result
in serious
cardiovarcular disease. Whilst metabolic syndrome may be improved by lifestyle
changes, for example to diet and exercise levels, what is required is a
pharmacological
treatment which can be used to help combat this dangerous syndrome.
The inventors have surprisingly found that the combination of growth hormone
(GH)
and an insulin sensitising agent, for example, metformin provides a beneficial
treatment
for patients suffering from metabolic syndrome.
Although to date the studies have been performed with metformin, as the result
depends
on the insulin-sentising properties of that drug, similar results will be
obtainable with
other insulin-sensitising agents.
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The invention provides a pharmaceutical composition comprising growth hormone
and
an insulin sensitising agent, for use in the treatment of metabolic syndrome.
Preferably
the composition may be in combination with one or more pharmaceutically
acceptable
Garners.
Preferably the GH is recombinant human GH. By analogue we mean a substance
having
the same biological activity as described here and having at least 65%,
preferably 75%,
most preferably 85% homology with naturally occurring growth hormone.
The insulin sensitising agent is preferably a biguanide, most preferably
metformin.
Alternative insulin sensitising agents include PPAR gamma insulin sensitising
agents
and thiazolodeniones, for example the troglitazone and rosiglitazone families.
However, this list is not exclusive.
Insulin sensitising agents that are l~nown, or are under development include:
Troglitazone, Chemical Name:
5-[[4-[3,4-Dihydro-6-hydroxy-2,5,7,8-tetramethyl-2H-]-benzopyran-2-yl)
methoxy]phenyl]methyl]-2,4-thiazolidinedione
V411 (DIABII~, Glaucanin)
Pioglitazone (ACTOS, AD 4833, U 72107, U 72107A, U 72107E, ZACTOS~)
Chemical Name: 2,4-Thiazolidinedione, 5-[[4-[2-(5-ethyl-2-pyridinyl)
ethoxy]phenyl]methyl]-, monohydrochloride, (+/-) ;
Rosiglitazone (Avandia~, BRL 49653, BRL 49653C) Chemical Name: 2,4
Thiazolidinedione, 5-[[4-[2-(methyl-2-pyridinylamino)ethoxy]phenyl]methyl];
Bexaxotene-oral (LGD 1069 oral, Targretin oral, Targretin~, Targretyn oral
Targrexin
oral) Chemical Name: 4-[1-(3,5,5,8,8-Pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)
ethenyl]benzoic acid;
ZD 2079, (ICI D 2079) (Chemical Name: R)-N-[2-[4-(Caxboxymethyl)
phenoxy]ethyl)-N-(2-hydroxy-2-phenethyl) ammonium chloride: Netoglitazone,
(Isaglitazone, MCC 555, RWJ 241947) (Chemical Name:
5-[6(2-Fluorobenzyloxy)naphthalen-2-ylinethyl]thiazolidine-2,4-dione); INS 1
(D-chiro-inositol) (Chemical Name: D-1,2,3,4,5,6-Hexahydroxycyclohexane), NN
2344(DRF 2593); Dexlipotam, Chemical Narne: 5(R)-(1,2-Dithiolan-3-yl)
pentanoic
3 5 acid;
HQL 975, Chemical Name: 3-[4-[2-(5-Methyl-2-phenyloxazol-4-yl)
ethoxy]phenyl]-2(S)-(propylamino) propionic acid;
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YM 268, Chemical Name: 5,5'-Methylene-bis(1,4-phenylene)bismethylenebis
(thiazolidine-2,4-dione).
PPAR agonists under development include:
Reglitazar (JTT 501, PNU 182716, PNU 716) (Chemical Name:
Isoxazolidien-3,5-dione,
4-[[4-(2-phenyl-5-methyl)-1,3-oxazolyl]ethoxyphenyl-4]methyl-, (4RS));
I~RP 297, Chemical Name:
5-(2,4-Dioxothiazolidin-5-ylmethyl)-2-methoxy-N-[4-(trifluoromethyl)
benzyl]benzamide;
R 119702 (CI 1037, CS 011) Chemical Name:
(+/-)-5-[4-(5-Methoxy-1H benzimidazol-2-ylinethoxy)benzyl]thiazolin-2,4-dione
hydrochloride;
DRF 2189, Chemical Name: 5-[[4-[2-(1-Indolyl)ethoxy]phenyl]methyl]
thiazolidine-2,4-dione
A fuxther aspect of the invention provides the use of a growth hormone or
analogue
thereof and an insulin sensitising agent in the preparation of a
pharmaceutical
composition to reduce waist circumference. The invention also provides the use
of
growth hormone or analogue and an insulin sensitising agent in the preparation
of
pharmaceutical compositions for the treatment of metabolic syndrome.
Preferably the
insulin sensitising agent reduces the insulin antagonist action of the growth
hormone.
The invention further provides a method of treatment of metabolic syndrome
comprising the step of administering an insulin sensitising agent and growth
hormone
or analogue thereof to a patient suffering from metabolic syndrome.
The insulin sensitising agent and growth hormone may be administered
simultaneously,
or may be administered separately.
Also provided is the use of an insulin sensitising agent in the treatment of
metabolic
syndrome, in combination with treatment with growth hormone.
Further provided is the use of growth hormone in the treatment of metabolic
syndrome,
in combination with treatment with an insulin sensitising agent.
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Preferably metformin is administered orally in the form of metformin
hydrochloride
tablets. The preferred daily dose is in the range of 1000-2000 mg. The tablets
may be
given in two doses daily or in extended-release form in one dose.
5 The growth hormone is preferably administered in a subcutaneous inj ection,
the dose
being individually regulated according to body weight and IGF-levels.
The invention will now be described in detail by way of example with reference
to the
figures in which:
Figure 1 shows the baseline characteristics of 25 men with metabolic syndrome.
Figure 2 shows the lipid metabolism characteristics of the same 25 men with
metabolic
syndrome.
Figure 3 Shows the change over 18 months of IGF-I levels in two groups of
patients
treated either with Metformin+GH or with Metformin+Placebo. The figure gives
the
results from 14 of the 25 patients who completed the study (Mean~SEM).
Analysis of
the difference between the two groups by repeated measure analysis of variance
(RM
ANOVA) included the data from all 25 patients (p=0.001).
Figure 4. Shows the changes over 18 months in BMI, total body fat, and waist
circumference in the two groups of patients treated either with Metformin+GH
or with
Metformin+placebo. The Figure gives the results from the 14 patients who
completed
the study (mean~SEM). Analysis of the differences between the two groups by
repeated
measure analysis of variance (RM ANOVA) included the data from all 25 patients
and
gave significant results only for the change of waist circumference
(BMI:p=0.24; total
body fat: p=0.91; waist circumference: p=0.048).
Figure 5. Shows the change over 18 months in fasting plasma glucose and area
under
the curve (AUC) of glucose during oGTT in the two groups of patients treated
either
with Metformin+GH or with Metformin+placebo. The Figure gives the results from
the
14 patients who completed the study (mean~SEM). Analysis of the differences
between
the two groups by repeated measure analysis of variance (RM ANOVA) included
the
data from all 25 patients (FPG: p=0.09; AUC glucose: p=052).
Figure 6. Shows the percentage change in glucose disposal rate (GDR) in the
two
groups of patients treated either with Metformin+GH or with Metformin+Placebo.
The
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Figure gives the results from the 14 patients who completed the study
(mean~SEM)
(difference between the two groups: p=0.07, paired Wilcoxon/Mann/Whitney
test).
STUDY DESIGN
A randomized, double-blind, placebo-controlled trial was used to evaluate the
effect of
rhGH (recombinant human Growth Hormone) in addition to metformin in patients
suffering from metabolic syndrome. 25 non-smoking men with metabolic syndrome
were selected to complete the study. The patients had a body mass index (BMl'
between 30-40 kg/m2, fasting plasma glucose levels without antidiabetic
medication of
between 110 mg/dl (6.1 rilrilol/1) and 145 mg/dl (8.0 mmol/1), and HbAlc
<7.5%.
Hypertension, if present, was treated by angiotensin converting enzyme
inhibitors or
angiotensin II receptor antagonists. Antihypertensive drug therapy was kept
stable
during the study period. Lipid-lowering medication, the use of drugs to reduce
body
weight such as orlistat or sibutramine, and the use of glucocorticoids was not
allowed
during the study period. The same was true for any medication influencing
growth
hormone levels.
After providing informed consent, patients attended a pre-inclusion
assessment.
Growth hormone deficiency was excluded by a combined GHRH/arginine-test. After
considering inclusion and exclusion criteria, patients were randomized into
the placebo
arm or the rhGH arrn of the study.
In both treatment groups, patients received metformin 850 mg twice daily
(Glucophage~, Merck, Darmstadt, Germany) during the whole study period. In the
Met+GH group, rhGH (Genotropin~, Pharmacia GmbH, Erlangen, Germany) was
administered sc before bedtime at a daily does of 9.5 ~.g/kg (0.20 IU/leg
BW/week) after
an initial 4-week dose adjustment period. The rhGH chosen was equivalent to
the dose
used in the study from Johannsson et al. Patients in the Met+Placebo group
administered sc injections of placebo. The placebo vials contained the same
vehicle as
the rhGH vials, and both preparations were visually indistinguishable.
In both treatment groups, the daily dose was reduced by half in the event of
side effects.
Compliance was assessed by counting the returned empty vials.
Before start of the study, the patients were screened by two determinations of
FPG
levels. At baseline and after 18 months of treatment, all patients were
hospitalised.
During the study (after 6 weeks, 3 months, 6 months, and I2 months), they were
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monitored as outpatients. Measurements of blood pressure, ECG, as well as
physical
and laboratory examinations were performed during all visits. Body weight was
measured in the morning to the nearest 0.1 kg wearing indoor clothing, and
body height
was measured barefoot to the nearest 0.01 m. Waist circumference was measured
in the
standing position with a flexible plastic tape midway between the lower rib
margin and
the iliac crest. Systolic and diastolic blood pressure were measured after 10
min supine
rest and were repeated after 2 min to calculate mean value. Measurements of
body
composition were performed at baseline, and after 6, 12 and 18 months. A
combined
arginine-GHRH-test was performed at baseline, euglycemic hyperinsulinemic
clamp
tests were performed at baseline and after 18 months.
Inclusion Criteria
Patients had to fulfil all the following criteria before inclusion in the
study:
- Age >_ 35 years and _< 70 year;
- Male sex;
- Body mass index >_ 25 and <_ 40 kg/m2;
- Fasting plasma glucose measured at two different days >110 mg/dl (6.1
rmnol/1) and
<145 mg/dl (8.0 mmol/1) without antidiabetic medication;
- HbAlc <7.5 %;
- Increase of GH levels to >_ 5 ng/ml during the GHRH.arginine-test;
- The patient must give written informed consent before starting any study-
related
procedures;
- The patient must be willing and able to participate and comply with the
requirements of the study.
Exclusion Criteria
- S erum creatinin > 1.2 mg/dl;
- Previous cardiovascular or cerebrovascular events;
- Evidence of other major cardiac disease;
- Reasons for elevated blood glucose concentrations other than type 2
diabetes;
- Chronic obstructive pulmonary disease (COPD);
- Severe hyperlipidemia requiring treatment with lipid-lowering drugs;
- Use of drugs to reduce body weight such as orlistat or sibutramine during
the last 3
months before study entry;
- Use of glucocorticoids during the last 3 months before study entry;
- Present or past therapy with insulin (>1 week);
- Oral antidiabetic drugs during the last 6 months before study entry;
- Alcoholism or drug abuse;
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- Elevated liver enzymes (SGOT (AST) >100 U/1;
- Any suspicion of malignant disease;
- History of a malignant disease with a relapse-free interval of less than 10
years;
- State of kidney transplantation;
- Hypopituitarism;
- Treatment with glucocorticoids;
- Participation in any other study up to 30 days before study entry;
- Any disease influencing regular participation in the study;
- Active smoking or lustory of smoking during the last 12 months.
Criteria for Removal from the Study
- Cardiovascular or cerebrovascular events;
- Suspicion of malignant disease;
- Development of severe diabetes mellitus requiring insulin therapy;
- Evidence of impaired renal function (creatinine >_1.4 mg/dl.);
- Any other circumstance which prevents further therapy and regular
assessments;
- Decision of the patient to withdraw from the study.
RhGH was supplied in vials, each containing LE. of rhGH by Pharmacia & Upjohn.
In
accordance with the randomisation list, indistinguishable placebo vials were
given to
patients from the placebo group as a single subcutaneous injection before
bedtime.
RhGH and placebo were stored in a secure refrigerator at 2-8°C.
PATIENT ASSESSMENTS
Baseline Study Assessments (Pre-inclusion (Visit 1))
Before visit 1, patients were screened by two determinations of fasting plasma
glucose
levels. Baseline study assessments at visit 1 were only performed if both
values were
>110 mg/dl. (6.1 mmol/1) and <145 mg/dl. (8.0 mmol/1) without antidiabetic
medication. Prior to visit 1, the patients fasted overnight. The following
procedures
were carried out and recorded:
- Past medical history and history of concurrent diseases including any
allergies;
- Current medication;
- Family history;
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- Physical examination including determination of BMI, waist to hip ratio, 2
separate
measurements of systolic and diastolic blood pressure in the right arm with
the
subj ect seated and after a 10 min. rest, and assessment of heart rate;
- 12-lead electrocardiogram (ECG);
- Laboratory assessment (see below);
- GHRH/arginine-test.
Study Assessments (Visits 2, 3, 4, 5, 6, 7)
Before each visit, patients fasted for at least 8 hours. Study medication was
given to the
patient at each visit according to the randomisation list. Empty vials were
returned by
the patient and were counted to assess compliance with study medication. At
each visit
the patients had to provide samples of blood before receiving study
medication. Each
visit included:
- Medical history;
- Current medication;
- Adverse events;
- Physical examination including determination of BMI, waist to hip ratio, 2
separate
measurements of systolic and diastolic blood pressure in the right arm with
the
subject seated and after a 10 min, rest, and assessment of heart rate;
- 12-leas electrocardiogram (ECG);
- Laboratory assessments (see below);
- Oral glucose tolerance test;
- Assessment of body fat mass (DEXA) (visit 2 (begin), 5 (6 months), 6 (12
months)
and 7 (18 months;
- Euglycemic-hyperinsulinemic clamp test (visit 2, visit 7).
Blood Assessments
The following assessments were made:
- Haemoglobin;
- Haemotocrit;
- Platelets;
- Red blood cell count;
- White blood cell and differential count;
- Serum sodium;
- Serum potassium;
- Ser~un creatinine;
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- Serum alanine transaminase and asparetate transaminase;
- Total cholesterol, HDL-cholesterol, LDL-cholesterol, triglycerides;
- Lipoprotein (a), apolipoprotein Al, apolipoprotein B;
- HbAlc;
5 - Insulin-like growth factor 1, Insulin-like growth factor-binding protein
3;
- Growth hormone, growth hormone-linding protein;
- Fibrinogen, Plasminogen activator inhibitor (PAI) activity;
- Free fatty acids.
10 Pharmacodynamic Assessments
Oral Glucose Tolerance Test
After an overnight fast, subjects ingested a 75-g oral glucose load over a 3-
min. period.
Blood samples for plasma glucose and serum insulin were obtained at baseline
and after
30, 60, 90 and 120 min. Glucose tolerance status of the subjects was defined
by WHQ
criteria.
GHRHlArginine-Test
Recently, application of GHRH and arginine has been reported as a potent,
reproducible
and age-independent test that is well correlated to the results of growth
horlnone during
insulin induced hypoglycemia and is able to reliably distinguish between
growth
hormone deficiency and normal adults (14).
The test was carried out according to the following protocol:
- 1 ~.g/lcg. i.v. Somatorelin (GHR_H_ Ferring~ at 0 min);
- 0.5 g/kg. Arginine i.v. over 30 min. from 0 to 30 min.);
- Blood samples for measurement of GH every 15 min. from -15 min. to +90 min.
In young, healthy, normal weight subj ects, a GH peak?9 ~,g/1 was regarded as
normal.
24 Houz~ Growth Hormone Profiles
The studies were started at 8 a.m. after a 12-h overnight fast. A polyethylene
catheter
was inserted into an antecubital vein and blood was collected at regular
intervals.
During the study, the patients received standardised meals.
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Measurement of Whole-Body Fat Mass and Fat Free Mass by Dual Energy X Ray
Absorptiometry (DEXA)
Whole-body fat mass and fat free mass (FFM) were assessed by dual energy X-ray
absorptiometry (DEXA) (model DPX-L, Lunar Corporation, Madison, WI, USA).
Analysis was performed using software version 1.31. A typical scan lasted
approximately 10 minutes, and the subject received 0.02 mrem of radiation.
Euglycemic-Hyperinsulinefnic Insuli~z-Clasnp
Euglycemic-hyperinsulinemic clamp test was performed based on the protocol of
De
Fronzo, et al. Prior to insulin clamp study, all subj ects consumed a diet
containing at
least 200 g. of carbohydrate per day for 3 days. The studies were performed at
8 a.m.
after a 12-h ovenught fast. A polyethylene catheter was inserted into an
antecubital
vein for infusion of insulin and 20% dextrose. A second catheter was inserted
into a
hand vein and the hand was heated during the study to ensure arterialization
of venous
blood. All blood samples for glucose and hormone analyses were drawn from the
heated hand catheter. Insulin-mediated glucose disposal was determined by the
euglycemic insulin clamp technique. Htunan insulin was achninistered as a
continuous
infusion (40 mU/m2~min) for 180 min. Plasma glucose concentration was measured
every 5 min. Infusion of 20% dextrose was not begun until 5 min. after the
initiation of
insulin infusion and was periodically adjusted to maintain the arterialized
plasma
glucose concentrations at 95 mg/dl. Blood samples for determination of serum
insulin
concentrations were collected before insulin infusion was started and after
120 and I80
min.
It has been previously demonstrated that hepatic glucose production is almost
completely suppressed in the hyperinsulinemic state. Under these conditions,
peripheral glucose utilisation is equal to the rate of glucose infusion to
maintain
euglycemia. In this study, the final 60 min. of the infusion period (between
120 and
180 min.) was used for determination of peripheral glucose utilisation. Data
are
expressed in milligrams per kilogram fat free mass per minute.
Assays
Serum GH levels were determined by a chemiluminescence immunometric assays
(Nichols Institute Diagnostics GmbH , Bad Nauheim, Germany). The assay was
calibrated against the WHO 1St international standard (80/505) fox human GH.
Intra-
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and interassay coefficients of variation (Cvs) for a low point of the standard
curve were
5.4% and 7.9%, respectively. Plasma IGF-I concentrations were measured by an
immunoradiometric assay (Nichols Institute Diagnostics GmbH, Bad Nauheim,
Germany). The assay was calibrated against the WHO 1St International Reference
Reagent 87/518. Intra- and interassay Cvs for low IGF-I concentrations were
2.4% and
5.2%, respectively. In our laboratory the normal IGF-I ranges were I I4-492
~,g/L for
adults aged 25-39 yr, 90-360 p,g/L for adults aged 40-54 yr, and 71-290 pg/L
for adults
aged >_ 55 yr.
Plasma glucose was determined by the glucose oxidase method (glucose
autoanalyzer,
EBIO 6666, Eppendorf, Germany). Plasma insulin levels Were assessed by
radioimmunoassay (Biochem Tmmunosystems, Freiburg, Germany). HbAlc was
determined by DCA 2000 (Bayer, Leverlusen, Germany, upper limit 6.3%). Total
testosterone was measured by an automated chemiluminescence immunoassay (ACS
180, Bayer diagnostics, Fernwald, Germany).
Red blood cell counts, white blood cell counts, serum levels of sodium,
potassium,
creatinine, liver enzymes, total cholesterol, HDL-cholesterol, LDL-
cholesterol,
triglycerides, lipoprotein (a) (Lp(a)), apolipoprotein A1, apolipoprotein B,
fibrinogen,
and prostate specific antigen were determined by routine methods.
Statistics
The data, if not marled otherwise, represent the mean~standard deviation (SD).
Comparisons between baseline values of the two groups were performed using the
unpaired Wilcoxon/Mann/Whitney test (WMW). Absolute differences between time
points were analyzed per group using the paired Wilcoxon signed rant test
(WSR). The
areas under the curve of glucose and insulin were determined per times. For
those AUC
measures as for BMI and total body fat time courses were compared by repeated
measure analysis of variance (RM ANOVA). Occurrence of side effects were
compared by chi-square test (CHIC. All p-values were given unadjusted and
therefore
interpreted explorative. All tests were conducted at a two-sided significance
level at
5%. Univariate analyses were performed with Graph.PadPrism (version 3.00 for
Windows, GraphPad Software, San Diego, CA, USA) and RM ANOVA was done in
SAS V6.12.
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For analysis of the time courses of parameters by RM ANOVA, data from all 25
patients were included. For analyzing the differences between time points,
only data
from the 14 patients who have completed the study were included.
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Schedule of Assessment
ScreeningBegin 6 weeks3 months6 months12 months18 months
Visit Visit Visit Visit Visit Visit Visit
1 2 3 4 5 6 7
Informed consentX
Physical
examination X X X x x X X
Medical historyX X X X X X X
Medication X x x X x x X
Adverse events X X X X X
Educational
programme x
Heart rate X X x X X X X
Blood pressure X X x X x x X
Body mass indexX X X X X X X
Waist to hip X X X X x X X
ratio
Haematology X X X X X X X
Clinical chemistry
x X x x x x x
ECG x x x x x x x
Oral glucose
tolerance test X X X X X X
Fasting blood
glucose X
HbAlc x x x x x x
Lipid metabolism X x X x X X
Lipoproteins X X X X X X
IGF-1, IGFBP-3,
GHBP X x x x x x
GHRH/arginine-
test
24-H GH profile X X
Insulin clamp X x
test
Body fat mass
(DEXA) x x x X
Asservation
of
whole blood x
samples (10m1)
Asservation
of
serum samples X X X x x X
(1 Oml)
Study medication X X X X X
Return of empty
vials X x X x x
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Results
Patients' assignment to the two treatment groups was stratified according to
BMI and
HbAlc. At baseline, the two groups did not differ significantly in terms of
all other
5 characteristics except the systolic blood pressure (Met+GH: 136~10 mmHg vs.
Met+Placebo: 126~17 mmGh; WMW p=0.033) (Tables 1,2).
Two patients in the Met+GH group and three patients in the Met+Placebo group
received treatment for hypertension (Met+GH group: I2.5 mg/d hydrochlothiazide
10 (12.5 mg/die) and 60 mg/d metoprolol, Met+placebo group: amlodipine 10
gm/d;
quinalapril 10 mg plus hydrochlorthiazide 12.5 mg/d and benazepril 10 mg/d).
These
medications were leept stable during the study period.
Side effects
Side effects were observed in 7/12 patients of the Met+GH group and in 8/13
patients
of the Met+placebo group (CHI p=0.81). The side effects reported in the Met+GH
group were arthralgia (5 patients) and peripheral edema (2 patients). These
side effects
appeared during the first 6 weeps of treatment and subsided spontaneously in 3
patients.
in 3 other patients side effects subsided in response to dose reduction. In
the
Met+Placebo group, side effects reported were headache (2 patients); transient
parasthesia (2 patients), diarrhoea (2 patients), muscle stiffiiess (1
patient) and
arthralgia (1 patient).
Dropout from the study occurred in S patients from the Met+GH group (4 due to
non-compliance, 1 due to the requirement for additional antidiabetic drugs).
In the
Met+placebo group, dropout occurred in 6 patients (5 due to non-compliance and
1
patient due to a newly diagnosed plasmocytoma).
Overall, 14 patients (7 from the Met+GH group, 7 from the Met+Placebo group)
completed the study.
Results are as shown in Tables 1-21
Those include data of the 14 patients who completed the study
Visits
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16
V 1 S creeping
V2 Baseline
V3 6 weeks
V4 3 months
VS 6 months
V6 12 months
V7 18 months
Treatment
Patient # 1-7
Treatment with metformin (1700 mg/day) + placebo
Patient # 8-14
Treatment with metformin (1700 mg/day) + growth hormone (daily dose 9.5
~,g/lcg
BW)
ARG-GHRH test and IGF-I levels
At baseline, mean GH peaks in the ART-GHRH test were reduced in both groups
(Met+GH: 6.8~3.7 ng/ml vs. Met+placebo: 3.8~2.9 ng/ml; differences between the
two
groups: WMW p=0.15). Overall, 22/25 patients (88%) showed a maximum GH
response <9 ~.g/1.
The mean IGF-I levels at baseline were in the lower age-adjusted normal range
in both
groups (Met+GH: 173~58 ng/ml vs. Met+placebo: 144~40 ng/ml; differences
between
the two groups: WMW p=0.31). In the Met+GH group, IGF-I increased from
173~58~g/1 to 4.34~61 ~,g/1 (WSR p<0.001) after 3 months and remained stable
thereafter (differences between the two groups, RM ANOVA p<0.001) (Fig. 1)
Body weight and body composition (Tables 1 and 2)
Body weight (Met+GH: -2.2~5.7 kg vs. Met+placebo: -3.8~4.2 kg; differences
between
the two groups: WMW p=0.54) and BMI (Met+GH: -1.16~1.61 lcg/m2 vs.
Met+Placebo: -1.21~1.37 lcg/m2; differences between the two groups: M ANOVA
CA 02483005 2004-10-19
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17
p=0.24) slightly decreased in both groups without reaching statistical
significance.
Mean total body fat significantly decreased by 4.6~S.8 kg (range: 2.2-14.5 kg)
in the
Met+GH group (WSR p=0.04) and by 3.8~2.1 kg (range: 0.4-6.7 kg) in the
Met+Placebo group (WSR p=0.07). The difference between the two groups,
however,
S was not significant (RM ANOVA p=0.91) (Fig 2). Fat free mass did not
significantly
change in both groups (Met+GH group: O.S~3.8 kg; WSR p=0.99; Met+placebo:
-1.7~2.7 kg; WSR p=0.69; differences between the two groups: WMW p=0.26).
However, waist circumference significantly decreased during the study period
only in
the Met+GH group (Met+GH: 120~9 cm vs. 114~10 cm; Met+placebo: 112~9 cm vs.
110~9 cm; differences between the two groups: WMW p=0.048). This is an
important
result.
Glucose metabolism
1S
According to the inclusion criteria, baseline FPG levels were slightly
elevated in both
groups (Met+GH: range 6.2-8.0 mmol/l; Met+placebo : range 6.2-7.6 mmol/1;
differences between the two groups: WMW p=0.32). During the first 6 months of
treatment, FPG and AUC of glucose levels during the oGTT significantly
increased in
the Met+GH group (FPG: baseline S.9~0.7, 6 months 6.7~0.4 mmol/1/ WSR p-O.OOS;
AUC glucose: baseline 1005~2S7 mmol/1~ 120 min, 6 months 1186 ~203 mmol/1~120
min, WSR p=O.OOS) (Fig 3). These increases in FPG and AUC of glucose levels at
6
months returned to baseline levels during further treatment (after 18 months:
FPG S.8
~0.2 mmol/l; AUC glucose 1028 ~202 mmol/1~120 min). W the Met+Placebo group,
2S FPG remained stable until 12 months and decreased thereafter (baseline: 6.2
~0.3, 18
months: S.S ~0.6 mmol/1, WSR p=0.02). No significant differences were seen
between
both groups with respect to the overall changes of FPG (WSR p=0.09), the AUC
of
glucose (RM ANOVA p=0.52), the AUC of insulin (RM ANOVA p-O.S1), and HbAlc
(Met+GH: S.6 ~0.4% at baseline to S.6 ~0.3% after 18 months, WSR p=1.0;
Met+placebo: 6.0 ~0.7 to S.6 ~0.4%, WSR p=0.22; differences between the two
groups: WMW p-0.20).
At baseline, GDR did not differ significantly between the Met+GH and the
Met+placebo group (Met+GH: 3.9 ~1.8 mg/kg~min; Met+placebo: S.7 ~2.1
3S mg/kg~min; differences between the two groups: WMW p=0.12). After 18 months
of
treatment, mean GDR increased in the Met+GH group (4.6 ~2.4 mg/kg~min) and
slightly decreased in the Met+Placebo group (4.8 ~1.4 mg/kg~min). The
difference
CA 02483005 2004-10-19
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18
between the two groups, however, did not reach statistical significance (WMW
p=0.07)
(Fig.6.)
Lipid metabolism and fibrinogen
At baseline, levels of triglycerides, total cholesterol, Lp (a),
apolipoprotein A1 and B
were not statistically different between the two groups (Table 2). In both
groups,
triglycerides, total cholesterol, LDL-cholesterol and fibrinogen levels did
not change
significantly during the 18 months period. HDL-cholesterol significantly
increased in
both groups (Table 6). In the Met+placebo group the apolipoprotein A1 and B
increased
significantly, whereas these parameters were unaffected in response to the
additional
administration of rhGH. A statistically significant increase in Lp (a) was
observed in
both groups without a siguficant difference between the two groups.
Blood Pressure
A slight decrease of systolic and diastolic blood pressure was seen in both
treatment
groups without statistical difference between the groups (Met+GH: 136 ~11/88
~10 at
baseline to 127 ~13 (WSR p=0.25) / 81 ~4 (WSR p=0.38) rmnHg after 18 months;
Met+Placebo: 126 ~16 / 83 ~11 to 118 ~9 (WSR p=0.009)/75 ~8 (WSR p=0.11)
mmHg).
Total testosterone levels
Serum total testosterone levels significantly increased in both groups after
18 months
(Met+GH: 8.57 ~1.7 to 10.9 ~3.6 nmol/l, WSR p=0.03; Met+placebo: 8.9 ~1.6 to
14.2
~3.5 nmol/1, WSR p=0.02). Despite a higher increase in the Met-Placebo group,
the
difference between the two groups did not reach statistical significance (WMW
p=0.06).
CA 02483005 2004-10-19
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19
Table 1
Body Mass Index (kg/m2)
v2 v~ = u.~ _t v~. . ua
- _ = ~ ~,. .
;,, ~ a , ~
;:
1 31,6J 30,6 30,4 31,1 31,2 30,5
2 34,131,8 33,0 35,1 35,0 34,7
3 31,230,1 29,9 28,8 28,9 28,4
4 33,732,2 31,6 31,7 33,8 33,3
30,529,4 29,5 28,7 27,0 27,8
6 34,633,8 34,3 32,8 32,8 32,4
7 29,429,4 29,3 29,7 29,2 29,5
~ ~ ~~ j
~ I'
~ ,.~~
I
~~
..." ,.., ...,. k" ,... .... , w. _
Min :. _ .. ,... . ", _ ... . ,
29,4......... ...- " . -_...,......., ,... ,. ......
29,4 . .. , " a 27,8
.. .-., 28,7 27,0
29,3
Max 34,633,8 34,3 35,1 35,0 34,7
mean 32,231,0 31,1 31,1 31,1 30,9
SD 2,01,6 1,9 2,3 2,9 2,6
_. 3 . ._.., ., .". .. 30'3 ..29 .. 30 6
7 _", 31 $ 31.'Ov s . 6. : ..
.,. . . _. .....,. . .. .u,.._ a_::_.a
. . ,..,
: . .
9 41,241,2 39,0 38,2 39,7 38,3
33,032,8 32,6 32,4 31,8 32,1
11 30,729,6 29,4 28,9 30,0 30,2
12 33,032,8 31,5 30,9 32,2 30,3
13 35,535,7 35,6 33,9 35,2 34,7
14 29,630,7 31,0 31,4 30,3 31,4
FE ~~'~ ,1
::Y
HI~
3.
~'
~.=3'3
(
'~..,
~,
~
~
~
I
F . ~, , ( . ~ E .....~~~~s ~v
i ~. ~~ - = r ~L
_ ~ - = 3~s
~ "
Min ..."."w,..,.. ~e. ~.~ ,;~._."" , .
29,6~..",";,.,.,., _ ~. .~,.,..,....r .
...,.,,~_ 29,4 _ .....,..30,2
_ _ 28,9 .
29,6 ~.~..
29,6
Max 41,241,2 39,0 38,2 39,7 38,3
mean 33,733,5 32,9 32,3 32,7 32,5
~D 3,83,9 3,3 3,0 3,6 3,0
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Table 2
Waist Circumference (cm)
S
1 109,5109,0 109,5 116,0 112,0 110,0
2 120,0109,5 107,0 114,5 116,0 116,0
3 106,0107,0 107,0 104,0 n.d. n.d.
4 119,5113,0 113,5 113,0 117,0 116,0
5 105,5103,5 103,0 104,0 94,0 97,0
6 121,0119,0 118,5 116,5 115,0 117,0
7 100,0101,0 100,5 104,0 97,0 101,0
I ~ .,.. ~ x. .. ~ > . ,.",.
Min 100,0.. ,. -.- . ~,... , ,.94 ,
"':... .,. . .. 00,5 104,p',.~~ ou.:.,. - 97 O
....101,p'~ . ...... ,
.'
Max 121,0119,0 118,5 116,5 117,0 117,0
mean111,6108,9 108,4 110,3 108,5 109,5
SD 8,5 6,0 6,1 6,0 10,3 8,6
~ ~ ' ~; ( 1
1
118,0._..,~.. __.- .~.... ..9EO.~ 104'0-....._.__._~... ,..
- ....109,5-....103,5. .._. . ...~07,p-1
_.
9 140,0133,5 126,5 126,5 133,0 128,0
10 117,0117,0 113,0 113,0 113,0 108,0
11 115,0104,0 102,0 102,0 97,0 n.d.
12 119,0112,0 114,0 109,5 112,5 n.d.
13 117,5121,5 116,0 120,5 127,0 n.d.
14 111,0115,0 114,0 117,0 111,0 111,0
' ~
Min .1_11;~~~~w_ w, __. .w_.102,0y g7,0 107,0
~.' 104'0._ -_ ~~ jv j5;0
:~._.
Max 140,0133,5 126,5 126,5 133,0 128,0
mean119,6116,1 112,7 112,1 113,9 113,5
~D 9,4 9,5 8,2 10,6 12,5 9,8
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21
Table 3
HbAlc (%)
vzm v~ m v~ , ... .. ~r~
1 6,26,1 5,7 5,9
2 7,45,3 5,7 5,7
3 5,55,2 5,5 5,4
4 5,75,8 5,0 5,0
5 5,45,7 6,0 6,0
.
6 5,95,4 5,0 5,4
7 5,65,1 5,3 5,6
_... ... _ .:: _.,: .._
Mm 5 5,1 . ,. _..:_ !
4 5 0 _.~
5,0
Max 7,46,1 6,0 6,0
mean 6,05,5 5,5 5,6
SD 0,70,4 0,4 0,3
f
,. ~ . " .".. ", ....
...".5,4. . . . .., ~ . 5 4.,
8 5,2 .. ,5 6. ,- -,~~~~~~~z"~
., .....
9 5,36,1 5,7 4,9
1 5,46,1 5,6 6,3
0
11 6,35,9 5,1 5,4
1 5,45,4 5,7 5,9
2
1 5,45,6 5,6 5,6
3
1 5,85,1 5,6 5,4
4
3
~ ._ _ ~.~_. ., ~~. _.~.~z
~ u..u-... ~
' ~
___ 5 5,1 5,1 4,9
Min 3
Max 6,36,1 5,7 6,3
mean 5,65,6 5,6 5,6
~D 0,40,4 0,2 0,4
1~
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22
Table 4
Total Cholesterol (mmol/1)
y
1 6,346,26
2 4,324,91
3 5,565,64
4 5,645,51
5 3,905,53
6 4,555,09
7 4,655,87
Minv~.'. ._..___.....
.3'904'91 ,_ .
...
Max 6,346,26
mean4,995,55
SD 0,860,45
~
~
.........,.
8 6.88~
.......~~~.
5,77
9 6,215,82
5,285,20
11 4,244,89
12 5,724,19
13 4,294,47
14 4,99n.d.
~
..__'~. lE.i.. _. ~..
_ 4,24.: :.'1
Min . ~
4,19
Max 6,885,82
mean5,375,06
~D 0,970,67
Storage samples from V3-V6 are available for analysis of lipid parameters
CA 02483005 2004-10-19
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23
Table 5
LDL Cholesterol (mmol/1)
._ v~ v~ - ..a,'
. -
..~
,
1 4,533,36
2 2,842,61
3 3,653,62
4 4,093,59
5 3,903,52
6 3,032,95
7 3,083,80
'
3Min , a ......,_..:_..,~
., 2'$~.....,... 2;~1
Max 4,533,80
mean 3,593,35
SD 0,630,42
: 33 ..~
_ .~I
8 ; ..3
......3 ~ ..3'70. ..
75 . .... ..".~
9 4,653,47
3,833,41
11 2,842,97
12 n.d.2,25
13 2,072,53
14 3,57n.d.
_. _,n_
Min . ,.... ....~,..
-.. 2 , 2 25~ .-.
~7 N
Max 4,653,70
mean 3,453,06
~D 0,890,57
10 Storage samples from V3-V6 are available for analysis of lipid parameters
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24
Table 6
HDL Cholesterol (mmol/I)
v~.. ~.r = ~,
_~
1 1,08 1,42
2 0,88 1,32
3 1,03 1,53
4 0,96 1,50
5 0,96 1,01
6 0,95 1,29
7 0,89 1,45
_,_ . . .~J
Min O $$~._. l O~
rvJ .~..v.
Max 1,08 1,53
mean0,96 1,36
SD 0,07 0,18
i
8 0 78 ~.,: l;o~
!, :~.. ..
9 1,01 1,06
1,08 1,42
11 0,91 1,32
12 0,88 1,16
13 0,80 0,93
14 0,80 n.d.
Min-~0,78
..J ~....~._
~ __ _O 93~
Max 1,08 1,42
mean0,89 1,15
~D 0,12 0,19
Storage samples from V3-V6 are available for analysis of lipid parameters
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WO 03/090784 PCT/EP03/04357
Table 7
Triglycerides (mmol/1)
5
~r-
1 2,75 2,46
2 1,35 1,76
3 1,10 1,00
4 1,57 1,15
5 1,72 2,04
6 2,24 1,41
7 2,07 1,74
-. ~'
.
Min ~ ~....-. ..
1~~0 1,0~ - _.
~' .a...
Max 2,75 2,46
mean1,83 1,E5
SD 0,57 0,51
. . ~..,.a ":'...>
8 2,36 >"
. . 2,28 .
9 2,21 3,40
10 0,67 0,82
11 1,33 1,25
12 3,59 2,44
13 1,66 2,92
14 1,58 n.d.
Min .. ' 0 _ .
0,67
Max 3,59 3,40
mean1,91 2,19
~D 0,93 0,98
Storage samples from V3-V6 are available for analysis of lipid parameters
CA 02483005 2004-10-19
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26
Table 8
Maximal GH-response in GHRH-arginine stimulation test (mmol/1)
l~z
1 2,75 2,46
2 1,35 1,76
3 1,10 1,00
4 1,57 1,15
5 1,72 2,04
6 2,24 1,41
7 2,07 1,74
., ~ ..... ,.~.w
Min_.~~.,.~ ...'
. 1,00
1,10
Max2,75 2,46
mean1,83 1,65
SD 0,57 0,51
,
-
3
1
~ ~ s
9 2,21 3,40
0,67 0,82
11 1,33 1,25
12 3,59 2,44
13 1,66 2,92
14 1,58 n.d.
i r~~
," ~ 3
Min0,67 ~.
0,82
Max3,59 3,40
mean1,91 2,19
~D 0,93 0,98
V7b are data collected 4 weeks after withdrawal of GH / placebo
CA 02483005 2004-10-19
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27
Table 9
IGF-I (~,g/1)
__... _ .. ,._ _ .. . ~ . ~ ~ _.__,~ _._ v._ ,
_. _ vs . ~_ ~ __._.' ; ... v7 _ _ _~ ._..
. ._,_ v~. _ vs u7rz*
vz '~~
1 190 136 127 176 151 162 182
2 97 120 111 134 121 127 162
3 185 153 173 202 202 223 212
4 109 105 102 135 113 113 127
5 159 178 234 244 202 208 212
6 105 90 101 99 124 93 94
7 164 262 210 216 199 192 222
_ - 1. r ' _. _.3. _ ,
_" . __ .__ _ . . ., "~ ~._ ..._ <. ....,...
. ._._._.. . - ~_ ~_... ._... . '
' > '. ,.. .. ~~
_" . ..
Min 97 90 101 99 113 93 94
Max 190 262 234 244 202 223 222
mean144 149 151 172 159 160 173
SD 40 58 55 52 41 50 48
,. _ .. ~~, -. _
,,: , ~ , :. E _- ;
,:: _ . . 3 r;
.- ~" ~
'~:. ~c
- ~
,:
,
,~3
:
~k
-.
, ... . ., J
z. ~ _~ ,i
3 '
~. , . _ .. ... .m.F.489~._~ ,............... 500_....508 --~.,......
175..,.485 __ _. _____._~_ ~46._s .,n....w"_,.
--_.~ 478
9 71 305 377 458 306 folgtfolgt
157 349 273 368 416 390 148
11 142 352 352 335 239 190 166
12 210 358 420 431 489 376 155
13 202 372 484 466 419 525 folgt
14 251 433 490 501 430 563 244
a"3 ~ - ,~-, r _,
- . M~~:. ..-M.. ....~...:.... , s
._ .. ... .. , ~
.~... .~._..,... . a _ ..-.
..
Min 7~ 305 273 . . " ..~._~- w
335 . ~ ,146
.. 90 ~ ~_ .._:
239 ._
Max 251 485 490 501 500 563 244
mean173 379 412 434 400 425 172
~SD 58 60 83 61 95 138 41
V7b are data collected 4 weeps after withdrawal of GH / placebo
CA 02483005 2004-10-19
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2~
Table 10
Glucose tolerance test: Glucose 0 min (mmol/1)
# V~ = V3:' V~ , ~~~~ , .;!_ m~ SIT.
. VG'.,' , I _,
~ .:
1 6,33 5,94 7,33 6,22 6,27 5,61
2 6,27 5,38 5,33 5,83 6,00 4,61
3 6,05 7,05 6,33 6,16 5,83 5,61
4 6,49 5,50 5,88 6,22 6,44 5,88
5,77 5,83 6,27 5,38 5,50 4,94
6 6,49 6,38 7,55 6,77 6,22 5,66
7 6,05 6,05 6,77 6,83 6,99 6,33
Min 5,77 5,38 5,33 5,38 5,50 4,61
Max 6,49 7,05 7,55 6,83 6,99 6,33
mean6,21 6,02 6,49 6,20 6,18 5,52
SD 0,27 0,57 0,78 0,50 0,48 0,58
i ~
~ ' _ i~~
" ' _.. ~ " " ' 3 t.
. __."~ ~ ~',,"".3, """", ..,..:..~.3
~....z= z ~ . ~~........n..d.:
... . N i
-
T
8 5,94 fi,11 6,27 6,99 n.d. _.
.
...
6,00
9 6,77 n.d. 6,33 7,05 6,77 6,11
5,44 6,00 6,16 6,44 5,77 5,94
11 6,72 6,77 6,66 7,16 6,38 5,77
12 5,77 6,00 6,44 6,33 6,27 5,77
13 5,88 5,83 5,77 6,27 6,44 5,50
14 4,94 n.d. 6,11 6,49 5,55 5,83
Min 4~j4.. ..- 5.83 5 77 ..- . _ . ",5 5 50<-
=,~ . -..._.... F~~. _-6 27.; 55 ...,.,._...,
~ .~~ .~, ~.~~
Max 6,77 6,77 6,66 7,16 6,77 6,11
mean5,92 6,14 6,25 6,68 6,20 5,84
~D 0,66 0,37 0,28 0,38 0,45 0,20
1~
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29
Table 11
Glucose tolerance test: Glucose 60 min (mmol/1)
v~ , ,
_.._m r _.._ ~_. v~ ors
. _. u~ v4 -
v~, ~' , : ~ ~
.
.
1 11,99n.d. 10,10 16,7112,93 14,93
2 8,33 10,05 n.d. 11,9912,05 10,55
3 9,66 9,71 11,32 10,998,55 12,93
4 8,49 9,44 9,88 9,339,38 10,88
5 10,279,38 10,32 8,107,16 5,11
6 12,1614,49 16,82 12,779,71 12,82
7 13,497,27 14,99 10,5513,77 13,38
. ,... 3
.
Min L . ~ . . 9 8$! _,. _.8'10_.H... ~.,- 7 16
' $,33-...~ ! v
Max 13,4914,49 16,82 16,7113,77 14,93
mean10,6310,06 12,24 11,4910,51 11,51
SD 1,97 2,38 2,94 2,782,44 3,20
f ~ ~i
-., ~
~
~
~
~
~
t
3
~
I~
'. 4 ~~ . v.,...."
$,. 9'49 ~..., .. ~~ , ...
~ ,. .. " ~ 9,38 . 9'05. _. ".n.d. .~
.., p 21 .. '." . .
... _
9 14,9913,99 10,05 12,1616,04 n.d.
9,38 9,49 11,21 10,8811,82 12,27
11 15,7112,49 13,88 16,389,77 15,10
12 12,3217,37 16,54 14,4912,16 7,38
13 9,33 9,27 9,88 10,278,49 9,05
14 6,61 12,38 11,49 11,388,72 12,60
~i ~ ~ ~ 1
~ ,
-
Min-... ... -> . . 9 3$ , ..... .. ...,. _. ..
_6'~1... .9'27 ". ~.,. .9;058'49:.. _....._T 38
., . ... ..
Max 15,7117,37 16,54 16,3816,04 15,10
mean11,1212,17 11,78 12,0911,17 10,94
~D 3,34 2,89 2,58 2,542,84 2,86
1~
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Table 12
Glucose tolerance test: Glucose 120 min (mmol/1)
5
T ~t4 E V~ v~ rr~:.=
..
1 7,27 10,27 7,16 12,108,10 12,49
2 13,218,60 n.d. 7,99 8,27 8,44
3 6,94 5,33 7,05 6,55 5,50 7,05
4 5,05 5,38 5,38 6,61 7,22 7,60
5 4,83 5,00 8,27 3,33 2,78 3,66
6 6,11 10,66 11,38 7,49 4,11 8,33
7 4,83 13,77 7,22 7,94 6,22 8,49
i.
. .. , .. ;~
Min 483- ..... ...,.. - 5 3 _~ _ -_ >_..
' . _._r 38, ,... 33 _ ~~ . ~~ , r
.~.5 . .... '. 78 y 3,6.6 '
0ov.. .
Max 13,2113,77 11,38 12,108,27 12,49
mean89 43 7,74 7,43 6,03 8,01
6 8
SD , , 2,01 2,60 2,06 2 60.....
2,96 3,36
._.4.'66.__.5 83 5,16 n.a. ~+,u~
g g~g4 13,21 11,38 6,16 8,44 8,10
10 5,44 5,88 7,60 10,05 4,94 8,66
11 9,10 6,44 9,94 10,27 7,16 4,88
12 5,22 9,77 6,00 11,55 3,83 3,77
13 10,82 10,88 9,71 10,32 8,66 9,77
14 4,50 7,99 9,33 7,27 7,94 8,44
v
Min 4,44 ... 4 ... , .... 5 31.83 ,. -..
~~ ._ 6fi.. , 1 ~ -... '_. ,.........
A-,.. ~ .... .5 _~ ' 3,77
" 83 >
.~
Max 10,82 13,21 11,38 11,55 8,66 9,77
mean6,92 8,41 8,54 8,68 6,83 6,81
~D 2,62 3,04 2,11 2,45 1,99 2,48
CA 02483005 2004-10-19
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31
Table 13
Glucose tolerance test: Glucose AUC (mmol/1/120 min)
v~. = ~ v~ ~~. .' , x~ , b s~
-
1 856 1041 1552 1207 1439
2 934 1022 1134 1151 1024
3 1107 954 1081 1041 853 1156
4 1136 893 931 944 973 1057
5 881 888 1056 748 678 565
6 1371 13811577 1194 893 1189
7 ggg 10311319 1076 1222 1247
._ , - -~ i
' ~ _ . _ _ _,...
.
Min 856 -......g88,..._...931 .,.,... _ ".,., ,.,, .,.._
. , .. _ --;~48, g78 -565.
_.
Max 1371 13811577 1552 1222 1439
mean1025 10281167 1098 997 1097
SD 189 183 238 247 205 271
'
~ y
' s - .,.... .. .
.
8 ' 1417 _.936- .... , . ..... .~.. . ~.
.. ' . .._. ... 926.'v ., "...,..........r 854
. . ~.. , 908
. . _,.,. "
9 1069 1134 1126 1419
1061 926 1086 1147 1031 1174
11 679 11461331 1505 993 1226
12 1069 15151366 1406 1032 729
13 1061 10571057 1114 963 1001
14 679 1152 1096 928 1184
' I 1 3 3
f ~ ~~ ~3 ~...
~ ,.w u. .
"
. F 679 g26 .. .,... ~.,.,. .., .,
Mm . ,..H. ' g26~.., . 'g08 o,na ". _ .. 729
.~ M " .. . 'g28
.
Max 1417 15151366 1505 1419 1226
mean1005 11161150 1186 1061 1028
~D 257 241 154 203 180 202
1~
CA 02483005 2004-10-19
WO 03/090784 PCT/EP03/04357
32
Table 14
Glucose tolerance test: Insulin 0 min (pmol/1)
V~ '. A '~~ V~ ~ " m
_. . . ....~~ . "V~
. V2 ' V4
~.. m =
1 114,1 179,4 514,4 200,2 124,8 246,1
2 96,1 127,7 169,3 116,2 370,9 175,8
3 70,3 230,3 48,1 65,3 65,3 83,9
4 115,5 76,8 104,8 112,6 148,5 149,2
5 35,9 78,9 86,8 94,7 66,7 87,5
6 66,0 65,3 79,6 66,7 92,6 86,1
7 61,0 62,4 76,1 75,3 40,2 153,5
~.., '
x
Min 35,9% . ~2~4~'3 _._. ., __.. _._ 83;9..::.
'-~ ___...~ _ 48'1 653 - - .40 _. .
-_..h._... ~. ~.,
.._
Max 115,5 230,3 514,4 200,2 370,9 246,1
mean79,8 117,3 154,2 104,4 129,9 140,3
SD 29,7 65,3 163,3 47,0 112,6 60,0
y ,.,_ ..,
.:.... ..=. ... -...z.. __. a>=i
..... . ~." ".~_ .~ ...
.. >... .
..
8.....61.;0 9~ 9 76;1 100,5 45 ~ 83
._ __. 9
,
9 223,1 495,1 278,4 231,8 414,7 478,6
172,2 264,8 196,6 172,2 132,7 279,1
11 122,0 125,6 108,3 92,6 97,6 99,0
12 295,6 160,0 265,5 171,5 176,5 199,5
13 283,4 260,5 261,2 238,9 252,6 298,5
14 147,1 184,4 142,8 199,5 203,1 290,6
~
~
L . , _ o..
' ~ ... , .., __..
. .. .. ,
-
. .g1 -~ ..... ....... - 45 Z 83'9
Min .... . 7~,1 _
~ . . 98'9 9216
~_
Max 295,6 495,1 278,4 238,9 414,7 478,6
mean186,3 226,7 189,8 172,4 188,9 247,0
~D 85,9 134,1 82,2 58,1 120,8 135,4
CA 02483005 2004-10-19
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33
Table 15
Glucose tolerance test: Insulin 60 min (pmol/1)
V2 Y~ - -. ~ Y"~' ! '~, . .,
, :::, . .
1 1021,7 1643,1 1404,9 1626,6 1580,7 1987,5
2 965,8 1239,8 3055,8 2011,2 1054,0 975,8
3 880,4 851,0 1323,1 1435,0 1435,0 1553,4
4 860,3 999,5 910,5 723,2 533,1 1169,5
5 1808,1 922,0 749,1 840,2 1240,6 1043,2
6 695,3 507,3 584,8 355,9 437,7 560,4
7 952,1 462,8 1238,4 481,4 827,3 932,0
. . _ ...._ , ,. _..
Min 695,3"'-_..462,g-...'~_ ..584,8,- 355'9..-437,7 ...r~
= .. .~. .. ..... ~ 560,4
...,
Max 1808,1 1643,1 3055,8 2011,2 1580,7 1987,5
mean 2 5 1323,8 1067,6 1015,5 1174,5
1026 946
, ,
SD 360,3 410,4 822,7 626,9 437,7 465,0
, '~.: ~ ~F '; ~~ ..
...,..~ .. .u_ .
.. ,
~
8 _..a.~8,. F........$39,53". ~ - 639,3 g01
. ....~. , . ,2
.1063 ~ X85
683, , ,
9 2903,7 2874,3 1466,6 2128,1 2227,1 2226,4
2196,3 1879,9 1705,5 819,4 2340,5 2062,8
11 1045,4 645,8 602,0 548,9 615,6 1183,2
12 1884,9 2247,9 1624,4 1408,5 1719,8 1153,7
13 2884,4 2382,1 1908,6 1443,6 1262,8 1664,6
14 1991,8 1533,3 1483,1 1435,0 1788,7 2337,6
_ , ~ :
i
8 ..~ ~ ; , .
a ... 602,0 .. ~ 548,9815,6 .... ~~
g~. _.. _ ..5 ..... - 801,2'
...--ra= ,.. .
645 .
Min 683, ,
Max 2903,7 2874,3 1908,6 2128,1 2340,5 2337,6
mean 1941,5 1803,8 1375,7 1281,3 1513,4 1647,1
~D 843,6 779,5 476,2 507,0 700,6 577,4
1~
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34
Table 16
Glucose tolerance test: Insulin 120 min (pmol/1)
~_. . ; F V3 . 1f~# ,,- ~5 ~ , VC V7 , ,-' ' .
. : ,' . .. :~;
', ~VZ' '
1 791,4 1435,0 726,8 2031,21380,5 1881,3
2 1018,1 797,1 1972,4 855,3 863,2 770,6
3 602,7 134,9 277,0 352,3 351,6 657,2
4 284,8 491,5 431,9 647,2 645,8 697,4
5 265,5 413,3 1152,3 122,0 88,3 236,8
6 264,8 495,8 406,8 352,3 94,0 348,0
7 301,4 1139,4 434,8 551,0 191,6 856,0
_.. , .. " -.. ~ ,.i
_... . _..l
Min'264,8 . x.34 277,0 . 122,p___. .. _. ...236,g~~
~ . 9 ___._ ~."~. _ $8 _...
~ ... .
Max 1018,1 1435,0 1972,4 2031,21380,5 1881,3
mean504,1 701,0 771,7 701,6 516,4 778,2
SD 305,5 452,7 604,9 632,1 479,2 535,8
' 3~'3
""" ,~_~_~ia..3._... ~ "......-,
i_ . ,.".,~... . 336,5 210,2 378,8 218,8
,_.._ 190,1
., 115,5
8
9 4734,8 3135,5 3935,5 1188,92674,8 1927,2
653,6 795,0 1912,9 1686,8524,5 1826,8
11 731,1 222,4 636,4 375,3 422,6 299,2
12 347,3 852,4 446,3 2019,0170,0 1772,2
13 3497,1 5237,8 2225,7 1949,42446,7 3511,4
14 1693,3 3312,7 3842,9 1435,02888,7 2392,1
Min ' ._. ~_._... . -_ 336,5~~_~.._ - .. 1.70'0...____ ._
X15,5_ ~90,1~__...... 210,2~~.: 218,8
___..~ " .-...._~I
Max 4734,8 5237,8 3935,5 2019,0 2888,7 3511,4
mean 1681,81963,7 1905,2 1266,4 1358,0 1706,8
~D 1770,7 1944,7 1537,2 725,0 1238,4 1153,9
CA 02483005 2004-10-19
WO 03/090784 PCT/EP03/04357
Table 17
Glucose tolerance test: Insulin AUC (pmol/I/120 min)
5
Y.G. ; ,.;', ~,; . ....Ya YV ~'', ~~'; ..:y
, Xs7;::,.,Y~ty . :. ~: ~~ I
. :~
1 88468147016 121530 164537 139999 183070
2 91374102136 247602 149814 100263 86939
3 7301362014 89135 98628 98606 115439
4 6362877016 70731 66189 55814 95571
5 11752770085 82118 56912 79083 72324
6 5163847269 49680 33923 31857 46645
7 6799763822 89630 47678 56589 86208
.. ~ ~
, u. -. . _..:......r , ":-: i . , ,...:
Min . _ . _...::. ~> ....... .. .,..>..
..... , , ;: 31857 46645
51638 .. ~.~.~.-.,.
47269 49680 33923
Max 117527147016 247602 164537 139999 183070
mean7909281337 107204 88240 80316 98028
SD 2186733497 65601 51290 36073 43021
,
_ ~~ ~ ~
__ . ... ,.: ~ ....__..~...F.... ..~. .. .,....'
.~. . . ...". ..- ~., ..,..,..r. .a.r - . .....;
.~ -.. '. . .a,....~~~ .. ...,.,...1
. ..
.
8 4632272410 62745 80439 51079 63154
9 322961281375 214411 170306 226314 205757
10 156551144583 165613 104934 160146 186945
11 8831749185 58462 46968 52543 82936
12 132379165247 118818 150223 113587 128375
13 286476307872 189119 152268 156745 214174
14 174718196911 208556 135134 200075 220739
Min 46322 49185 58462 46968 51079 63154
Max 322961 307872 214411 170306 226314 220739
mean172532 173940 145389 120039 137213 157440
~D 100421 97312 66040 44422 68245 65468
CA 02483005 2004-10-19
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36
Table 18
C-peptide (ng/ml)
.,
1 3,6 4,7 9,0 4,6 3,7 4,2
2 5,0 3,4 3,3 3,6 5,7 3,1
3 2,2 7,0 1,8 2,3 2,3 2,6
4 3,5 2,6 3,1 3,1 3,8 3,5
5 3,3 3,0 2,8 3,8 2,7 2,5
6 2,9 2,5 2,9 2,6 2,3 2,3
7 2,4 2,0 2,4 2,1 1,7 4,9
... , . ....,
Min .. ,.. 1,;$ 2'1.. _..1'7_ ... 2'3
2 2.. 2'O ._ .. ....~..~..,.. . . -. .,.-
.'. .. . ... ...., ~..., --x
.. ... . .
Max 5,0 7,0 9,0 4,6 5,7 4,9
mean 3,3 3,6 3,6 3,2 3,2 3,3
SD 0,9 1,7 2,4 0,9 1,4 1,0
~ I '
,' ~ '
1 _
"
a
__.
8 . 1 9_. ,~,2 _2'5..->,., .......
..... _.._ 4 . .._>,~ _ ._ 1.9 .....
... . ... ..2'7, . ...
.
9 4,6 7,0 4,5 4,6 5,3 8,5
1 3,6 5,0 4,1 4,3 3,3 4,8
0
11 3,1 3,0 3,3 2,8 2,9 2,7
1 5,4 7,4 6,5 7,4 6,0 7,4
2
13 4,5 5,9 5,4 6,1 6,5 7,5
1 5,3 8,5 6,3 7,3 5,9 6,7
4
I
$ t i
~
F ~ ~ ......,~..,x.._
I F~ ,
e a.~. _-
Min~ ...~'9x _ v ~.~~w~ ~...~- 1 9 26
.~._ 2,4___..__ .a u.~27.~,~~...
_25
Max 5,4 8,5 6,5 7,4 6,5 8,5
mean 4,1 5,6 4,7 5,0 4,5 5,7
~D 1,3 2,3 1,5 2,0 1,8 2,4
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37
Table 19
Glucose disposal (mg/kg/min) (euglycemic hyperinsulinemic clamp)
.~ y,~
~
..
1 3,24 2,70
2 4,31 3,27
3 5,71 6,20
4 4,91 5,84
5 7,27 5,81
6 4,58 5,37
7 9,64 4,61
l
Min 3,24 ~ ~~ 2,70
~
Max 9,64 6,20
mean5,67 4,83
SD 2,15 1,36
v~ ~ .,
a., I .. ....;
..-' r ~- '
8 5,78 7,56
9 1,67 1,96
3,14 6,38
11 5,03 6,14
12 4,74 5,55
13 1,36 1,95
14 5,35 2,47
.-. ,
Min 1,3:6_.._....,.
- 1'95 _._
...._
Max 5,78 7,56
mean3,87 4,57
~D 1,81 2,37
CA 02483005 2004-10-19
WO 03/090784 PCT/EP03/04357
3~
Table 20
Truncal fat (DEXA) (kg)
vz ~~ ~- ~~ , F . ~~ : ~m ~ I
'..
,
1 17,86 16,86 16,32 14,59
2 16,42 13,63 n.d. 13,55
3 13,11 12,72 12,72 n.d.
4 16,49 15,90 15,88 16,84
5 15,43 13,28 17,61 11,28
6 17,25 18,44 17,52 16,85
7 11,29 10,53 10,50 11,79
; ; t~ r
_ 3 r ..a.
A
Min 11,29 10,53 10,50 11,28
Max 17,86 18,44 17,61 16,85
mean15,40 14,48 15,09 14,15
SD 2,37 2,71 2,87 2,40
I
1.3'28.."10,63 .....,.~.r 8'59'.,~ 12;10..> ~., . ~....-
: ........
..
9 25,42 17,56 15,01 16,01
17,37 16,13 16,48 16,44
11 16,91 13,56 21,64 12,94
12 15,12 12,18 13,49 12,31
13 16,82 16,99 16,89 16,66
14 15,55 16,25 15,21 16,24
v " ~ ~~~n~
3 .'
? _ ", _ . .. ' ,.~..,_ .,...~,..
_ _ ~~ 10,63 ~ ' ~
Min 13,28 ~ 12,10
8,59
Max 25,42 17,56 21,64 16,66
mean17,21 14,76 15,33 14,67
~D 3,88 2,65 3,93 2,10
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Table 21
Total fat (DEXA) (kg)
vz u~ ~~ ~, ~. ~~
~
1 34,6533,45 33,96 29,16
2 28,99n.d. n.d. 25,06
3 24,0222,34 n.d. n.d.
4 41,5336,73 38,57 41,09
5 26,5124,49 17,61 19,84
6 40,9439,70 30,18 37,65
7 21,1739,70 20,82 18,09
.. - ... . . -
."_ ...._.,.... ._._ . .._... ..
_ .,....' .., ....3r W ....tilJ~..,.,:~.,.,
_ 21,17:.,. - ..>...-...a.
Min ...:... ._ - H
.- 17,61 18,09
22,34
Max 41,5339,70 38,57 41,09
mean 31,1131,34 28,23 28,48
SD 8,097,60 8,82 9,36
t
I 25,66'~. . "."., ~...u~ .~,~".22,88
8 '~ ~... ....16'77..,w~~_ .
.. 19 __..~~.
89 .~
9 50,0942,49 35,58 38,60
32,9131,04 32,57 30,58
11 33,0826,49 n.d. 26,37
12 32,1725,18 28,18 23,40
13 34,1835,29 34,92 34,67
14 30,3133,48 31,45 32,72
' '
, '
I
__~. _ y - ,...,~ .. ,_...__.
_ ' ~
.~..~ V 4~
~
Min 25,6619,89 16,77 22,88
Max 50,0942,49 35,58 38,60
mean 34,0630,55 29,91 29,80
~5D 7,617,45 6,96 5,47
15
CA 02483005 2004-10-19
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Discussion
This randomised, double-blind, placebo-controlled trial has provided first
evidence, that
rhGH in combination with an insulin sensitising agent such as metformin might
be a
5 promising therapeutic approach in high risk patients with metabolic syndrome
and that
treatment with an insulin-sensitising agent such as metformin might be
effective in
reducing the insulin antagonistic effects of rhGH. Since the insulin
antagonist action of
GH has to be regarded as the principal factor for GH induced insulin
resistance and
since rises in FPG levels during rhGH treatment would primarily be expected in
10 patients with pre-existing insulin resistance, such an effect of metformin
is of special
interest when treating patients with metabolic syndrome who are characterized
by an
insulin resistant state and a high risk of developing type 2 diabetes.
The study included highly selected male patients presenting with obesity and
impaired
15 glucose metabolism at baseline. Consistent with earlier findings of
disturbances in the
neuroendocrine regulation of GH secretion in obesity, the majority of the
patients
showed a reduced GH peak in the ARG-GHRH test at baseline.
Despite the fact that only patients with impaired FPG levels were included in
this study,
20 no sustaining negative effects or rhGH on glucose metabolism were seen and
the
overall course of FPG and insulin levels did not significantly differ between
those
patients receiving only metformin and those receiving metformin and rhGH.
Consistent
to previous studies, we have seen a slight but significant increase in FPG
levels and
glucose levels during oGTT after 6 months of treatment in the group receiving
25 metformin and rhGH. One patient with a baseline FPG level of 9.0 mmol/1
dropped out
of the study during this period due to the need for additional antidiabetic
drug
treatment.
The overall increase in FPG and AUC glucose levels, however, was transient
with both
30 parameters returning to baseline levels during further rhGH and metformin
treatment.
Interestingly, after 18 months of treatment, GDR as a measure of insulin
sensitivity
increased in those patients receiving metformin and rhGH and slightly
decreased in the
metformin group. This effect, however, did not reach statistical significance
due to the
small sample size included. Similar effects of rhGH on insulin sensitivity
were seen in
35 the study of Johannsson et al., who investigated obese male patients, who,
in contrast to
the present study, had normal FPG levels and did not receive metformin. The
long-term
course of FPG levels seen in our study, together with the GDR results after 18
months
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41
underlines the need for long-term studies when investigating the effects of
rhGH on
parameters of insulin resistance.
The study also showed a significant decrease of total body fat in both
treatment groups
without a significant difference between the groups. These data are in good
accordance
with previous studies showing that the administration of rhGH can reduce total
body fat
and visceral fat in obese patients due to the lipolytic effect of GH. Albeit
visceral fat
was not directly measured in the present study, waist circumference as a
parameter of
visceral fat accumulation decreased in those patients receiving a combination
of
metformin a~.zd rhGH, but not in those patients treated only with metformin.
Tlus
strongly suggests a rhGH specific effect on visceral fat and is of major
importance since
most of the clinical features of the metabolic syndrome are closely related to
the
accumulation of visceral fat, which is by itself one of the major determinants
of insulin
resista~lce in this syndrome. In consequence, the slight increase in GDR seen
in patients
treated with metformin and rhGH is most probably due to changes of body
composition
with a preferential reduction in visceral adipose depots associated with the
reduction in
visceral adipose depots associated with the reduction of waist circumference.
W
addition, GDR might have been influenced by the slight increase of skeletal
muscle
mass seen in the rhGH treated group.
Despite the additional effects of rhGH, patients treated with metformin alone
also
experienced positive effects regarding the course of body composition, which
is well
lmown from previous studies. In the metformin treated group, FPG levels
significantly
decreased consistent to previous studies demonstrating a significant
improvement of
glucose metabolism. The slight decrease of GDR after 18 months may be due to
the
natural course of disease and due to the fact that metformin has no direct
effect on
visceral fat mass and muscle mass.
Dyslipoproteinemia is one of the major characteristics of the metabolic
syndrome and a
central finding in patients with growth hormone deficiency.
Previous investigations on the importance of GH as a regulating factor for
serum lipids
have produced conflicting results in terms of the effects on serum
cholesterol,
LDL-cholesterol, HDL-cholesterol, and apoliprotein B. We have seen no
significant
changes of total cholesterol, triglycerides and LDL-cholesterol, neither in
the
metformin-treated group nor in response to the additional administration of
rhGH. This
may be due to the small sample size as well as the fact that patients with
severe
hyperlipidemia requiring lipid lowering drug treatment were excluded from the
study.
CA 02483005 2004-10-19
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42
The increase of HDL-cholesterol in response to metformin alone is most
probably due
to the reduction of total body fat and not due to a direct effect of
metformin. In the
group treated with metformin and rhGH, the increase of HDL-cholesterol may be
influenced by the additional administration of rhGH. As shown for the changes
of
serum lipids, the increase of lipoproteins were also similar in both groups.
This
observation might indicate an rhGH independent effect, even though rhGH may
increase Lp(a). The increase of lipoproteins in the group treated with
metformin alone
remains speculative, even though some previous trials could not demonstrate an
effect
of metformin on lipoprotein levels.
Low total testosterone levels were observed in both groups and are most
probably the
consequence of increased body weight, accumulation of visceral fat, and
patients' age.
The increase of total testosterone levels in both groups may be related to the
effects of a
reduced total body fat on testosterone and SHBG levels and most probably are
due to
direct effects of metformin or rhGH.
One shortcoming of the present study is the high number of dropouts. The
dropout rate
was similar in the two treatment groups and most of the dropouts were due to
non-compliance. Since most of the dropouts due to non-compliance occurred
after the
first 12 months of treatment, the high dropout rate is partly due to the study
design
including double blind sc injections over a period of 18 months.
The dose of rhGH used in his study was higher than the dose currently used in
adult
patients with GH deficiency. This resulted in supraphysiological IFG-I levels
in the
rhGH treated patients and also is the obvious cause for the high rate of GH
related side
effects during the early phase of treatment. In consequence, lower doses of
rhGH
titrated according to individual IGF-I levels are suggested for future
studies.
In summary, the present study demonstrates that in patients with metabolic
syndrome
and impaired FPG levels, and 18-month treatment with metformin in combination
with
rhGH was not associated with sustaining negative effects on glucose metabolism
and
was even more effective in reducing waist circumference than treatment with
metformin alone. Thus, the results provide first evidence that rhGH in
combination with
an insulin sensitising agent, for example metformin, might be an effective
therapeutic
approach in high risk patients with central obesity and that treatment with
metformin
might be effective in reducing the insulin antagonistic effects of rhGH. The
transient
increase in glucose levels despite the use of metformin as well as the case of
one patient
with an elevated FPG level at baseline who dropped out due to the need for
further
CA 02483005 2004-10-19
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43
antidiabetic drug treatment indicate, that such an approach would be most
applicable in
insulin resistant patients who show still normal or only slightly elevated FPG
levels.
Further studies including higher numbers of patients are necessary to
investigate the
effects of this therapeutic approach with regard to central findings in
metabolic
syndrome and, most importantly, with regard to its effects on cardiovascular
risk.
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44
REFERENCES
1. Johannsson G, Bengtsson BA 1999 Growth hormone and the metabolic
syndrome. J Endocrinol Invest 22:41-6.
2. De Fronzo RA, Gunnarsson R, Bjorkman O, Olsson M, Wahren J 1985
Effects of insulin on peripheral and splanchnic glucose metabolism in
noninsulin-dependent (type II) diabetes mellitus. J Clin Invest 76:149-55.
3. Reaven GM 1988 Banting lecture 1988. Role of insulin resistance in human
disease. Diabetes 37:1595-607.
4. Rosen T, Eden S, Larson G, Wilhelmsen L, Bengtsson BA 1993
Cardiovascular risk Factors in adult patients with growth hormone deficiency.
Acta Endocrinol (Copenh) 129:195-200.
5. Franco C, Bengtsson BA, Johannsson G 2001 Visceral obesity and the role of
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