Full text of DOI:10.2165/00003088-200241070-00003

Clin Pharmacokinet 2002; 41 (7): 485-504
0312-5963/02/0007-0485/$25.00/0
DRUG DISPOSITION
© Adis International Limited. All rights reserved.
Clinical Pharmacokinetics of
Depot Leuprorelin
Piero Periti, Teresita Mazzei and Enrico Mini
Department of Preclinical and Clinical Pharmacology, Università di Firenze, Florence, Italy
Contents
Abstract
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 485
1. General Pharmacology of Leuprorelin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 487
2. Pharmaceutical Formulation Issues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 488
3. Pharmacokinetics and Pharmacodynamics in Animals . . . . . . . . . . . . . . . . . . . . . . . . . 490
4. Clinical Pharmacokinetics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 495
4.1 Short-Acting Formulations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 490
4.2 Long-Acting Formulations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 497
5. Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 501
Leuprorelin acetate is a synthetic agonist analogue of gonadotropin-releasing
hormone. Continued leuprorelin administration results in suppression of gonadal
steroid synthesis, resulting in pharmacological castration.
Abstract
Since leuprorelin is a peptide, it is orally inactive and generally given subcu-
taneously or intramuscularly. Sustained release parenteral depot formulations, in
which the hydrophilic leuprorelin is entrapped in biodegradable highly lipophilic
synthetic polymer microspheres, have been developed to avoid daily injections.
The peptide drug is released from these depot formulations at a functionally
constant daily rate for 1, 3 or 4 months, depending on the polymer type [poly-
lactic/glycolic acid (PLGA) for a 1-month depot and polylactic acid (PLA) for
depot of >2 months], with doses ranging between 3.75 and 30mg.
Mean peak plasma leuprorelin concentrations (Cmax) of 13.1, 20.8 to 21.8,
47.4, 54.5 and 53 μg/L occur within 1 to 3 hours of depot subcutaneous admini-
stration of 3.75, 7.5, 11.25, 15 and 30 mg, respectively, compared with 32 to 35
μg/L at 36 to 60 min after a subcutaneous injection of 1mg of a non-depot for-
mulation. Sustained drug release from the PLGA microspheres maintains plasma
concentrations between 0.4 and 1.4 μg/L over 28 days after single 3.75, 7.5 or
15mg depot injections.
Mean areas under the concentration-time curve (AUCs) are similar for subcu-
taneous or intravenous injection of short-acting leuprorelin 1mg; a significant
dose-related increase in the AUC from 0 to 35 days is noted after depot injection
of leuprorelin 3.75, 7.5 and 15mg. Mean volume of distribution of leuprorelin is
37L after a single subcutaneous injection of 1mg, and 36, 33 and 27L after depot
administration of 3.75, 7.5 and 15mg, respectively. Total body clearance is 9.1
L/h and elimination half-life 3.6 hours after a subcutaneous 1mg injection; cor-
responding values after intravenous injection are 8.3 L/h and 2.9 hours.

486
Periti et al.
A 3-month depot PLA formulation of leuprorelin acetate 11.25mg ensures a
Cmax of around 20 μg/L at 3 hours after subcutaneous injection, and continuous
drug concentrations of 0.43 to 0.19 μg/L from day 7 until before the next injection.
Recently, an implant that delivers leuprorelin for 1 year has been evaluated.
Serum leuprorelin concentrations remained at a steady mean of 0.93 μg/L until
week 52, suggesting zero-order drug release from the implant.
In general, regular or depot leuprorelin treatment is well tolerated. Local re-
actions are more common after application of the 3- or 4-month depot in com-
parison with the 1-month depot.
Within the past two decades, a number of pep-
nadotropes respond to an acute need for gonado-
tropins.[2]
tide drugs have been approved for clinical use
through advances in molecular biochemistry; some
of these are natural or synthetic hormones. A recent
development of considerable interest is the synthe-
sis of various analogues of gonadotropin-releasing
hormone (GnRH) that constitute a new class of
pharmacological agents.
The natural human GnRH (or LHRH) is respon-
sible for regulating the synthesis and secretion of
the gonadotropins luteinising hormone (LH) and
follicle-stimulating hormone (FSH). The secretion
of GnRH is governed by the GnRH pulse genera-
tor which resides in the arcuate nucleus of the
mediobasal hypothalamus and preoptic area. The
mechanisms of this governance are not yet well
understood. GnRH is released from some hypothal-
amic neurons into the hypophysioportal circulation
and comes into contact with specific receptors on
pituitary gonadotropes. The keystone of GnRH se-
cretion is its pulsatile release, with the frequency
or amplitude of GnRH pulses determining whether
FSH and/or LH are secreted and the relative
amounts of each.[1]
Pituitary gonadotropes are remarkable cells in
that they have the ability to regulate the number of
their own GnRH receptors. During the pulsatile
GnRH signal and after short-term stimulation by
the endogenous peptide or its synthetic superactive
agonists, an increase in the number of GnRH re-
ceptors amplifies the synthesis and release of go-
nadotropins. This receptor upregulation has been
shown to occur after a single administration of a
GnRH agonist and is the mechanism by which go-
Conversely, when target pituitary cells are ex-
posed to the GnRH stimulus for a prolonged pe-
riod, they lose the ability to respond to the same
stimulation with their original sensitivity. This
process of desensitisation is rapid and reversible
and is part of a broader biological phenomenon of
adaptation, by which cells adjust their response to
the level of the stimulus.[3] An important action
mechanism of GnRH agonist analogues is the loss
of receptors and their inability to react with endo-
genous GnRH during prolonged exposure to the
synthetic agonist peptide. The ensuing ligand-
receptor complex eventually becomes internalised
by pinocytosis and degraded by hydrolytic lyso-
somal enzymes, thus decreasing the number of
receptors in the plasma membrane of gonado-
tropes.[4] A prolonged occupancy of these recep-
tors by a GnRH agonist also contributes to down-
regulation, since as long as receptors are occupied
by the analogue they cannot be stimulated by en-
dogenous GnRH. During this desensitisation of go-
nadotropes, an impairment of calcium channels oc-
curs and consequently the ability of GnRH to
evoke Ca2+ entry through these channels is reduced
and the pituitary gonadotropin secretory response
to Ca2+ is attenuated.[5]
Mature endogenous GnRH is a 10-amino-acid
peptide with blocked amino and carboxyl termini
arising from cleavage of a 92-amino-acid polypep-
tide precursor. Since GnRH also can be readily
synthesised through solid phase peptide synthesis,
it is possible to introduce specific amino acid sub-
stitutions into the synthetic products and thus de-
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

Depot Leuprorelin
487
sign potentially useful derivatives of the natural
chemical model. Efforts to synthesise longer-acting
GnRH agonists have been directed toward stabilis-
ing the molecule against enzymatic attack, increas-
ing binding to circulating proteins and membranes,
and enhancing the affinity of the agonist peptide
for the GnRH receptor. This pharmaceutical re-
search has resulted in thousands of GnRH ana-
logues, some of which are potent agonists.[6]
tracellular, GnRH analogues can accumulate in the
extracellular space with an apparent volume of
distribution of about 30L, thus facilitating de-
sensitisation regardless of the role of GnRH pro-
teases in physiological signal termination.[10]
Whereas pulsatile delivery of natural GnRH
from the hypothalamus, with the correct period-
icity, stimulates gonadotropin synthesis and secre-
tion, a continuous infusion of GnRH actually
suppresses gonadotropin secretion by desensitis-
ing the gonadotropes. Short-term use of GnRH ag-
onists stimulates pituitary gonadotropin release,
while long-term administration leads to inhibition
of the pituitary-gonadal axis due to downregula-
tion of the GnRH pituitary receptors, thus causing
so-called ‘chemical castration’ in men and pre-
menopausal women.[8] As a result, secretion of LH
and FSH is inhibited, and the serum concentration
of testosterone falls to castration levels in men (50
ng/dl) and estrogen concentrations decrease to
postmenopausal values in women. These proper-
ties of GnRH or synthetic long-acting GnRH ago-
nists are used successfully as therapy for a variety
of endocrine disorders that are responsive to reduc-
tions in gonadal steroids, the clearest indications
being children with gonadotropin-dependent pre-
cocious puberty, assisted fertilisation techniques,
prostate cancer, estrogen-positive breast cancers,
endometriosis, uterine leiomyomas, benign pros-
1. General Pharmacology
of Leuprorelin
Most of the GnRH agonists contain one or two
substitutions in the peptide chain, where a hydro-
phobic D-amino acid residue replaces glycine in
position 6 and N-ethylamide replaces glycinamide
in position 10.[7] Leuprorelin acetate, des-Gly[8]-
(D-Leu[6])-GnRH ethylamide acetate (figure 1), is
the first superactive nonapeptide agonistic ana-
logue of GnRH to be synthesised.[9] It is defined as
‘superactive’ because leuprorelin has been shown
to be 31- and 90-fold more active than GnRH in
inducing ovulation in rabbits and rats, respec-
tively.[9] Because of their non-natural amino acid
substituents, synthetic peptides can be less suscep-
tible to proteolysis and bind with greater affinity
to GnRH receptors than does natural GnRH, result-
ing in an increase in their agonist biological activ-
ity. Furthermore, if deactivating enzymes are in-
1
2
3
4
5
6
7
8
9
5-oxo Pro
-
His
-
Trp
-
Ser
-
Tyr
-
D-Leu
-
Leu
-
Arg
-
ProNHC2H5• acetate
O
C
H
N
O
C
H
N
O
C
H
N
O
C
H
N
O
C
H
N
O
C
H
N
O
C
H
N
O
C
O
C
H
N
CH
CH
CH
CH
CH
CH
CH
N
CH2CH3•CH3COOH
O
N
H
CH2
CH2
CH2
OH
CH2
CH2
CH2
CH2
CH3
CH
CH3
CH
CH2
CH2
N
N
H
CH3
CH3
H
H
OH
C
NH
NH2
N
H
Fig. 1. Chemical structure of leuprorelin acetate, des-Gly10-(D-Leu6)-GnRH ethylamide acetate, molecular weight 1209.41 as free
base. GnRH = gonadotropin releasing hormone.
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

488
Periti et al.
tatic hyperplasia, hirsutism and polycystic ovarian
syndrome in women.[2,11,12]
therapeutic value in several serious and wide-
spread diseases in which suppression of gonadal
activity is desired. Only when leuprorelin is admin-
istered continuously is pituitary-gonadal function
suppressed and this paradoxical effect, attributed
to down-regulation of the receptors, is temporary
and reversible. The effects of continuous exposure
to GnRH or an analogue such as leuprorelin on
serum levels of gonadotropins and sex hormones
in humans is independent of drug dosage.[11] On
the other hand, long-term treatment with a GnRH
analogue has the practical disadvantage of requir-
ing prolonged daily injections, since these ana-
logues are water-soluble peptides with a relatively
short biological half-life and are poorly absorbed
from the gastrointestinal tract. To overcome this
disadvantage, parenteral long-acting depot forms
of GnRH agonist, such as leuprorelin microsphere
drug delivery system, have been developed and
successfully used in the last decade.[12,15-20] The
oral bioavailability of leuprorelin is low and cur-
rently this drug is administered by parenteral
routes as 3.75 and 7.5mg monthly depot intramus-
cular injections, or 1mg daily subcutaneous injec-
tions of the short-term formulation.[11] Recent
multicentre studies have reported the use of
11.25mg or 22.5mg 3-month and 30mg 16-week
leuprorelin depots in patients with stage D2 adeno-
carcinoma of the prostate.[21-24] Other researchers
have evaluated an implant that delivers leuprorelin
for 12 months for the palliative treatment of pros-
tate cancer. [25,26]
Native GnRH has a short half-life of less than
15 minutes due to lack of plasma protein binding
and because proteolytic enzymes split the mole-
cule between the fifth and sixth amino acid into
two biologically inactive residues. Replacement of
glycine in position 6 by a D-isomer of leucine ren-
ders the GnRH analogue molecule resistant to en-
zymatic cleavage and greatly increases its protein
binding (46%) and circulating half-life (around 3.5
hours). Obviously protein binding and half-life do
not necessarily correlate, leuprorelin clearance be-
ing about 9 L/h. The first three amino acids are sites
of receptor activation. Replacement of pyrogluta-
mic acid by 5-oxo-L-proline confers the attribute
of GnRH antagonist on such a compound because
the derivative still binds with the receptor but fails
to trigger the signal-transduction pathway from re-
ceptor to effectors within the cell.[2]
Splitting off the glycine molecule in position 10
results in the loss of 90% of the biological activity
of native GnRH. Nevertheless, the attachment of
NH-ethylamide to proline at position 9 restores
most of the agonistic power of GnRH and, in asso-
ciation with changes at position 6, creates a potent
compound: the leuprorelin molecule.
The leuprorelin base has three ionisable sites,
two cationic and one anionic: the imidazolyl nitro-
gen of histidine, the phenol hydroxyl of tyrosine
and the guanidine nitrogen of arginine. As a result
of the synthetic procedure, the peptide is asso-
ciated with at least 1 mol of acetic acid, therefore
existing as an acetate salt.[13] It appears that the
amino acids at both ends of the GnRH peptide
chain are associated with expression of hormonal
activity, whereas the amino acid in the sixth posi-
tion affords both metabolic stability and an in-
crease in hydrophobicity leading to the increased
potency of the synthetic derivative (figure 2).
Leuprorelin acetate is approximately 80 to 100
times more potent than natural GnRH.[14]
2. Pharmaceutical Formulation Issues
Like polypeptides in general, the GnRH ana-
logues are poorly absorbed due to their high mo-
lecular weight and because they undergo enzy-
matic degradation in the digestive tract that
appears to be mediated by a combination of Tyr[5]-
Gly[6] endopeptidase activity as well as by en-
zymes that hydrolyse the pyro-Gly[1] or cleave the
carboxyl side of Pro9.[6] GnRH is also readily ac-
cessible in blood to these proteases, since it is not
bound to human serum proteins. Leuprorelin and
similar analogues have enhanced resistance to en-
GnRH and its agonist analogue leuprorelin can
be used in conditions that require stimulation of the
pituitary-gonadal axis, but they have much more
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

Depot Leuprorelin
489
pyroGlu
1
His
2
Trp
3
Ser
4
Tyr
5
1+2+3 = receptor activation
5+6 = metabolic stability, affinity to receptors
9+10 = biological activity
A
10
9
8
7
6
Gly
Pro
Arg
Leu
Gly
1 = still bound to receptor but lends an
antagonist property
5-oxoPro
1
His
2
Trp
3
Ser
4
Tyr
5
6 = potentiation of metabolic stability,
increase of hydrophobicity
B
10 = attachment of ethylamide to carboxyl
group of proline at position 9 restores
the biological activity lost due to splitting
off Gly10
10
9
Pro
8
Arg
7
Leu
6
H3CH2C-HN
D-Leu
Fig. 2. Schematic peptide structure of native GnRH (A) and its synthetic agonist analogue leuprorelin (B). The modification of GnRH
that increases its hydrophobicity also increases its biological potency. GnRH = gonadotropin releasing hormone.
zymatic degradation and relatively high protein
binding capacity due to their non-natural amino
acid substituents,[10] resulting in increased biolog-
ical activity, increased accumulation in extracellu-
lar space[10] and prolonged serum half-life.[8,27]
Leuprorelin is eliminated by enzymatic break-
down and renal excretion.[28]
Various nonparenteral routes have been consid-
ered as alternative methods for administration of
leuprorelin acetate. However, sublingual, intra-
nasal, intravaginal or percutaneous administration
of leuprorelin in rats and healthy humans has vary-
ing and unpredictable absorption, whereas subcu-
taneous or intramuscular administration assures
complete absorption.[11,27,29-31]
During the past 30 years, drug delivery systems
using various kinds of biodegradable synthetic
polymers have been studied extensively. Around
the middle of the 1970s, rapid and innovative prog-
ress in analytical or biological technology made
possible the discovery and production of therapeu-
tic hormone peptides, including GnRH agonist an-
alogues. In the 1980s, injectable and biodegrada-
ble microspheres (20μm diameter) of the synthetic
aliphatic polyesters of hydroxyl acids, polylactic
acid (PLA) and polylactic/glycolic acid (PLGA),
appeared to be a particularly ideal drug delivery
system due to avoidance of insertion of large im-
plants and no need to remove the device after the
drug supply is exhausted.[32] Moreover, the PLGA
and PLA drug carriers completely disappear from
the injection site in about 6 or 12 weeks, respec-
tively,[33] and a PLGA with molecular weight of
:
14 000 and lactic/glycolic acid ratio of 75 25
most effectively meets the requirements for a
once-monthly depot.[34]
The small, spherical microspheres with a
smooth surface, containing a high leuprorelin con-
centration, are easily dispersed in the vehicle, and
are easily injectable,[33] with a small initial burst
and persistent serum drug concentrations after in-
tramuscular or subcutaneous injection into ani-
mals[34-42] and humans.[20,23,24,43-45]
The biocompatibilities of PLGA and PLA were
originally demonstrated using absorbable surgical
sutures. The tissue response to PLA or PLGA
microspheres does not play a major role in control-
ling the degradation rate of these polymers.[33]
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

490
Periti et al.
These synthetic, biodegradable polymers are
highly lipophilic and most water-soluble com-
pounds, such as GnRH peptide analogues, are eas-
ily released from microspheres prepared using
them. The polymers slowly decompose in the
body, yielding lactic acid and glycolic acid, which
are normally found in blood and tissues.[46] There-
fore, in terms of safety, these polymers constitute
an ideal drug delivery system. This system has a
high entrapment efficiency and long-lasting re-
lease, predominantly due to polymer erosion. The
time courses of release of leuprorelin acetate and
the change in the average molecular weight of the
polymer were determined in in vitro and in vivo
tests using microspheres prepared with PLGA (77
crease in the molecular weight of the polymer
when the copolymer ratio of the microsphere wall
is around 75 25.[34] The release profile of the drug
:
from microspheres prepared with PLGA was ideal
for 1 month but was effected by different and in-
dependent mechanisms up to and after day 14. Dur-
ing the first 2 weeks, drug release occurred by a
diffusion-dependent mechanism from the PLGA
matrix and subsequently by a copolymer-degrada-
tion-dependent release. The release rate of the pep-
tide after day 14 was related to the spontaneous
hydrolysis rate of the PLGA.[47]
Leuprorelin and sodium oleate, chosen as an
acidic amphipathic surface-active agent, were
combined to increase the lipophilicity of the
leuprorelin via hydrophobic ion pairing:[48] the
hydrophobically modified leuprorelin-oleate com-
plex was lyophilised and directly encapsulated
within biodegradable PLGA microspheres via a
single oil-in-water emulsion method. The PLGA
microspheres encapsulated with the complex
leuprorelin exhibited higher loading amount and
greater encapsulation efficiency than those with
free leuprorelin, reducing an initial burst effect and
showing similar drug release behaviour and stabil-
ity.
Recent results of screening biodegradable PLA
and PLGA for the 3- or 4-month depot formula-
tion[33,34] indicate that microspheres prepared us-
ing PLA with a molecular weight of 12 000 to
18 000 give linear sustained drug release for over
13 weeks; a similar release pattern was confirmed
by serum leuprorelin concentration assays follow-
ing subcutaneous injection of these microspheres.
Microspheres prepared using PLA with a molecu-
lar weight of about 15 000 containing less than
0.1% water-soluble oligomers and loaded with
12% leuprorelin represent the most desirable long-
acting depot formulation.[21,22]
:
23) -12 000 and, as shown in figure 3, the results
were comparable, suggesting that PLGA is hydro-
lysed by a nonenzymatic process. Release of
leuprorelin acetate is generally correlated with de-
Leuprorelin acetate remaining in vitro
In vivo*
Molecular weight in vitro
In vivo*
100
90
12 000
10 000
8 000
80
70
60
50
40
30
20
10
0
6 000
4 000
2 000
0
5
10 15 20 25 30 35
Time (days)
Fig. 3. Comparison of in vitro and in vivo release of leuprorelin
acetate from drug delivery systems and degradation of poly-
lactic/glycolic acid (PLGA) microspheres (based on data of
Ogawa et al.[34]). Microspheres containing 0.9mg of the drug
dispersed in 0.3ml of diluent were injected subcutaneously into
the nucal region of 6-week-old rats and the amount remaining
at the site of injection was quantified by HPLC: the polymer was
extracted from the microspheres with methylene chloride and
the average molecular weight was measured by gel permeation
chromatography.
3. Pharmacokinetics and
Pharmacodynamics in Animals
Animal model studies have been conducted to
evaluate alternative routes to subcutaneous and in-
tramuscular leuprorelin injections.
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

Depot Leuprorelin
491
A comparison of the ovulation-inducing activ-
ity of leuprorelin acetate following intravenous,
subcutaneous, oral, rectal and vaginal administra-
tion in diestrous rats clearly indicated that injec-
tion is the most reliable route of administration of
this GnRH analogue: 100% of ovulation was
reached with a dose inferior to 0.5 μg/kg by injec-
tion, whereas the same result required more than
1.0 μg/kg by vaginal or rectal administration and
more than 50 or 100 μg/kg by the nasal or oral
routes.[46]
Intravenous
Subcutaneous
Vaginal
100
10
1
Figure 4 shows leuprorelin serum concentra-
tions after parenteral or vaginal administration to
diestrous female rats according to radioimmuno-
assay. After intravenous administration, leu-
prorelin rapidly disappeared from serum with an
0
2
4
6
8
10
12
Time (h)
Fig. 4. Serum concentrations of leuprorelin in rats during the
diestrous as evaluated by radioimmunoassay. 100 μg/kg in sa-
line was administered intravenously and subcutaneously, and
500 μg/kg in 5% citricacid solution (pH 3.5)vaginally. Eachpoint
represents the mean standard error. The pharmacokinetic
parameters after intravenous administration were obtained by
simulating a two-compartment open model. The serum concen-
tration (Ct) at time t was expressed by: Ct = Ae – dt + Be – βt,
with A = 241.5 99.6 μg/L, B = 122.4 43.1 μg/L, α = 8.37
4.12 h–1, and β = 1.35 0.23 h–1. The transfer rate constants
between central and tissue compartments were 2.99 2.32 h–1
and 3.79 1.35 h–1. The elimination rate constant from the
central compartment was 2.94 0.92 h–1 and the volumes were
0.32 0.07 L/kg for the central compartment and 1.31 0.71
L/kg for the tissue compartment (reproduced from Okada et
al.,[49] with permission from John Wiley & Sons, Inc).
1
α phase half-life (t ⁄ α) of 8.4 4 min and a terminal
2
1
phase half-life (t ⁄ β) of 33.2 6.8 min, correspond-
2
ing to a two-compartment open model with vol-
umes of distribution (Vd) of 0.32 0.07 L/kg (cen-
tral compartment) and 1.31 0.71 L/kg (tissue
compartment). Subcutaneous administration re-
sulted in a sustained serum concentration three
times higher at 4 hours than that after the intrave-
nous route. In the case of vaginal administration,
leuprorelin was absorbed rapidly and a long-last-
ing serum concentration was observed.[49]
In another study, leuprorelin was administered
by subcutaneous, vaginal, rectal and nasal routes
to pregnant rats. Almost all the drug was rapidly
absorbed and eliminated after subcutaneous ad-
ministration of a 10μg dose, with 94% reaching the
systemic circulation, a peak concentration of 110
1
To develop a new dosage form for self-admin-
istration, a nasal formulation of leuprorelin was
studied in rats and dogs.[51] After the peptide was
administered with a bioavailability promoter (5%
α-cyclodextrin), the percentage of dose absorbed
was about three times that obtained with no pro-
moter. After nasal administration to pregnant rats,
the fraction of the leuprorelin dose that reached the
systemic circulation was very low, but the rate of
absorption was enormously high and there was a
rapid decrease in serum concentration.[52] The
intranasal bioavailability in rats was significantly
increased by a surfactant,[52] α-cyclodextrin, EDTA,
and solution volume.[29] Prolonged release through
the intranasal route makes this form of administra-
μg/L and t of 32 min. After vaginal administra-
⁄2β
tion of the same dose, the rate of absorption was
very low with a consequent low peak concentra-
tion, but with longer persistence of drug concen-
trations than after parenteral route. Organic acids
enhance vaginal absorption of leuprorelin in dies-
trous rats by lowering the pH of the solution. The
absolute peptide bioavailability after vaginal admin-
istration of the 5% citric acid solution was 16.7%
at pH 3.5 and 38.4% at pH 1.8, and the enhanced
absorption correlated well with the chelating abil-
ity of the organic acids.[50]
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

492
Periti et al.
tion useful when lengthy GnRH receptor occupa-
tion is desired.
into rats using a suspending vehicle and a conven-
tional syringe with a 23-gauge needle. Leuprorelin
acetate was found to be released at a constant rate
over 30 days and its kinetics were as expected by
comparison with zero-order kinetics.[36] Neither
subcutaneous nor intramuscular injection of the
depot resulted in local irritation at the injection
site. Intramuscular injection of microspheres con-
taining leuprorelin acetate 13.5μg to 1.35mg,
equivalent to 1 to 100 μg/kg/day, suppressed serum
testosterone levels in male rats, and the effect was
maintained for 6 weeks. When the PLGA micro-
spheres were administered to female rats at a
leuprorelin acetate dose of 0.9mg per rat, equiva-
lent to 100 μg/kg/day, effective serum concentra-
tions of leuprorelin acetate (approximately 5 μg/L)
and suppressed levels of LH, FSH and estradiol
were maintained for 4 or more weeks, as ex-
pected.[54]
Leuprorelin in serum was measured by radioim-
munoassay (RIA)[55] after a single parenteral injec-
tion of PLGA depot formulation, consisting of
9.8% analogue content, at a dose of 0.9mg per rat
(corresponding to about 100 μg/kg/day for 30
days). Serum concentrations showed an initial
burst of more than 20 μg/L, followed by a steady
state with a pseudo-zero-order release, and a re-
markably constant concentration of around 5 μg/L
for 4 weeks. There were no differences between the
drug release profiles after subcutaneous and intra-
muscular injection.[51,54]
The bioavailability of leuprorelin (isotonic so-
lution diluent) after an oral dose in male adult bea-
gle dogs and rats was less than 1%, while the re-
lease patterns of the drug from the microspheres of
the subcutaneous injectable depot formulation
showed a slight initial burst and subsequent effec-
tive steady-state serum drug concentrations over 1
month.[51]
In another study from the same research group
at Takeda Chemical Industries,[54] serum concen-
trations of leuprorelin were assayed in rats after
PLGA microspheres were administered subcutane-
ously, as shown in figure 5. After a single injection
there was an immediate sharp increase, resulting
In addition to the various nonparenteral routes
considered as alternative methods for administra-
tion of leuprorelin acetate, percutaneous absorp-
tion in vitro was investigated in skin from nude
mouse, snake and human cadavers, focusing on
passive transdermal delivery by evaluating per-
meabilities with a high-performance liquid chro-
matography (HPLC) assay.[13] The low membrane
permeability of leuprorelin is due to its large mo-
lecular size and the highly hydrophilic cationic na-
ture of the acetate salt (partition coefficients octa-
nol/water: k = 0.022 for leuprorelin acetate and k
= 0.119 for leuprorelin base). Chemical enhancers
strongly increase skin penetration of the peptide,
particularly when in the form of leuprorelin base.
The in vitro permeability in mouse skin was 10-
or 100-fold higher than that obtained in human ca-
daver skin, depending on the type of enhancer that
was used in the leuprorelin formulation. Snake skin
was at least 10-fold less permeable than cadaver
skin. Skin permeability of the peptide in the base
form was 10-fold higher than in the acetate form
with the enhancers.[13] In healthy male humans, a
low level of electrical current was shown to induce
changes in cutaneous leuprorelin permeability suf-
ficient to promote the transdermal absorption of
therapeutically relevant amounts of the polypep-
tide.[53]
In a recent study, sublingual gel formulations of
leuprorelin were administered to dogs, monkeys
and humans;[31] the absolute bioavailability of
leuprorelin following sublingual administration
was 46.7% in dogs compared with 2.7% in mon-
keys at an equivalent dose of 0.45 mg/kg. Absolute
bioavailabilities in humans were 2.0, 2.3 and 2.4 at
doses of 1.125, 2.25 and 4.5mg, respectively. The
sublingual formulations used for the bioavailabil-
ity study were homogeneous gel solutions contain-
ing leuprorelin acetate 45 mg/L.
An injectable slow-release PLGA formulation
of leuprorelin was studied in rats and dogs.[51]
Microspheres of 20μm in diameter were easily in-
jected either subcutaneously or intramuscularly
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

Depot Leuprorelin
493
from the initial burst of the microspheres. Sub-
sequent serum drug concentrations were sustained
at effective steady-state concentrations over 1
month. The regression line of the amount of the
analogue remaining at the incised injection site re-
vealed that the rate of release of leuprorelin was
2.8% of the dose/day and the initial burst was
17.8%.[54]
Suppression of the serum levels of gonadotro-
pin and sex hormones, change in weight of genital
organs, and regression of experimental endometri-
osis after injection of the 1-month depot leu-
prorelin formulation were seen in studies in both
rats and dogs. Striking suppression of serum estra-
diol, testosterone, LH and FSH for 6 weeks follow-
ing an initial elevation (flare-up) of the levels of
these hormones were induced in rats or dogs by a
single injection.[36,37,54,56,57] The initial flare-up
was due to agonistic activity and could not be elim-
inated by this therapy. The flare-up of serum tes-
tosterone disappeared completely after 3 days in
rats[38] and after 2 weeks in dogs.[36] During re-
peated depot leuprorelin injection, the second and
succeeding parenteral administrations caused very
little flare-up as long as chemical castration was
sufficiently maintained.[57]
vesicles and prostate), as a marker of prostate tu-
mour suppression, in rats within 2 weeks of, and
lasting for over 6 weeks after, a single injection at
dosages of more than 100 μg/kg/day.[36,38] The
adrenal gland was not affected by the treatment. A
periodic challenge test using a solution of the pep-
tide (100 μg/kg) also revealed that a single injec-
tion of the microspheres caused dramatic suppres-
sion of the ability of the pituitary-gonadal system
to secrete gonadotropin and testosterone for over
5 weeks. Complete recovery of these functions was
observed 10 weeks after the injection; thus, it was
proven to be only a temporary chemical castration.
Cytological examination of vaginal smears follow-
ing leuprorelin depot injection also revealed re-
versible and sustained suppression; the oestrus cy-
cle was arrested in diestrus for 6 weeks after
treatment and recurred 8 to 10 weeks after.[57]
In these pharmacokinetic animal studies,
:
leuprorelin microspheres with PLGA (75 25)
14 000 were proven to maintain persistent serum
drug concentrations for over 4 weeks after injec-
tion and achieved reliable pharmacological effects
with the expectation of sufficient medical effi-
cacy.[33,34,38] It is possible to estimate the optimal
dose for clinical study in humans from these results
in animals. The effective serum concentration of
the peptide for suppression of serum testosterone
and genital organ weight was 0.4 to 1 μg/L in rats
after constant infusion using an osmotic mini-
pump.[38]
Additionally, 2-, 3- and 4-month depot formu-
lations of leuprorelin acetate in microspheres pre-
pared using various polymers were evaluated for
their release profiles and pharmacological ef-
fects.[39-45] The microspheres were fairly spherical
particles with a mean diameter of 20 to 30μm and
had many small pores on their surface, similar to
those of the 1-month depot, with a trap efficiency
of over 95% for a peptide load of between 5 and
18%.[33] The microspheres prepared using PLA
with a molecular weight of about 15 000, contain-
ing less than 0.1% water-soluble oligomers and
loaded with 12% leuprorelin, were the most desir-
able formulation for a 3-month depot injection,
The leuprorelin microspheres produced marked
regression of the genital organs (testis, seminal
25
20
15
10
5
Initial microcapsules burst
Steady state
0
0
2
4
6
8
Time (wk)
Fig. 5. Serum concentrations of leuprorelin acetate in rats after
a single subcutaneous injection of polylactic/glycolic acid
(PLGA) microcapsules (9.8% of leuprorelin acetate and a dose
of 0.9mg per rat); each point represents the mean standard
error for five rats (reproduced from Okada et al.,[54] with permis-
sion).
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

494
Periti et al.
giving linear sustained drug release for over 13
weeks in rats after a subcutaneous injection of 4.5
mg, corresponding to a dose of around 100
μg/kg/day for 90 days.[44] This linear sustained re-
lease pattern was confirmed by serum leuprorelin
concentrations determined by a double-antibody
RIA system[55] and depended on the rate of poly-
mer degradation, with a steady-state serum con-
centration of about 2 μg/L following a large initial
peak during the first week. Serum concentrations
of leuprorelin acetate in rats and dogs after paren-
teral injection were elevated just after administra-
tion because of the initial burst from the PLA
microspheres, and the subsequent concentrations
in rats were lower than those in dogs (figure 6). It
was assumed that clearance of the microspheres,
due to uptake by macrophages, in rats might be
more rapid than in dogs.[45] The serum steady-state
concentrations obtained with this 3-month depot
formulation were slightly lower than those after
injection of the 1-month depot formulation, prob-
ably due to the more rapid clearance of PLA micro-
spheres than PLGA microspheres.[45]
Subcutaneous
Intramuscular
Rat
C3 = 154 15 (subcutaneous)
102 10 (intramuscular)
4
2
0
Dog
C3 = 55 8 (subcutaneous)
64 8 (intramuscular)
4
2
0
0
4
8
12
16
Time (wk)
Fig. 6. Serum leuprorelin concentrations in rats and dogs
(double-antibody radioimmunoassay system) after subcutane-
ous or intramuscular injection of the 3-month polylactic acid
(PLA) depot. Dose was 100 (rat) and 25.6 (dog) μg/kg (repro-
duced from Okada et al.,[45] with permission). C3 = concentration
at 3 hours after administration.
Leuprorelin was incorporated into a variety of
PLGA matrices using a solvent extraction-evapo-
ration method, demonstrating that microsphere
combinations as two- or three-part systems made
:
from a 75 25 acid and ester-terminated PLGA, or
strongly suppressed for 21 weeks, indicating a per-
sistent suppression of the pituitary over a period of
3 or 4 months by the depot preparation.[58]
The objective of other preclinical studies was to
develop a leuprorelin acetate depot based on an in
situ-formed delivery system which suppressed and
maintained serum testosterone concentrations in
animals in the range of 0.5 μg/L for over 90 days.
The formulation evaluated contained 45% (w/w)
:
:
an ester-terminated 75 25 and 50 50 PLGA, pro-
vided a composite profile with onset of 10 days and
duration of approximately 100 days. In addition, a
single-polymer system composed of high molecu-
:
lar weight ester-terminated 75 25 PLGA was em-
ployed to produce leuprorelin release over the de-
sired 90-day release period. These formulations
were administered to an animal model at 3 or 9
mg/kg doses, and elicited chemical castration (tes-
tosterone RIA method) from 10 to 100 days.[39]
The leuprorelin serum concentration profile in
female rats after subcutaneous injection of the 3-
month depot microspheres prepared with PLA was
similar to that observed in male rats. Serum gonad-
otropin concentrations transiently flared up within
the first 6 hours because of the agonistic stimula-
tory action of the peptide, but thereafter were
:
75 25 PLGA having an intrinsic viscosity of 0.20
dl/g, dissolved in 55% (w/w) N-methyl-2-pyrro-
lidone with 3% (w/w) leuprorelin acetate, which
was dispersed within the polymer solution imme-
diately prior to use. With this formulation, testos-
terone concentrations in rat and dog models dimin-
ished to the target concentrations of 0.5 μg/L by
day 14 and remained suppressed up to day 91.[40,41]
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

Depot Leuprorelin
495
The new formulation of leuprorelin depot with its
simple manufacturing process is expected to pro-
vide an economic benefit to the user compared
with products currently available.[41]
of this neurohormone are present only in this sys-
tem.[6]
GnRH analogues are used clinically to induce a
selective and reversible suppression of the pituitary-
gonadal axis. This clinical application requires the
continuous administration of GnRH or the use of
a long-acting GnRH agonist, such as the leu-
prorelin acetate depot formulation, to induce bio-
chemical castration.
Single daily injections of leuprorelin or its re-
tard formula administered every 4 or more weeks
have been introduced for several disorders where
treatment relies on the complete extinction of
gonadotropin secretion and the subsequent sup-
pression of gonadal steroids to castration lev-
els.[6,11,21,22,30]
The development of lower-molecular-weight
GnRH analogues or of novel potent and orally ac-
tive nonpeptide GnRH antagonists[60] may, in the
years to come, improve bioavailability through the
oral route.
The clinical pharmacokinetic properties of
leuprorelin acetate after subcutaneous intramuscu-
lar administration of regular or depot formulations
have been studied in healthy volunteers and in pa-
tients, predominantly those with prostatic cancer.
The studies investigating direct or indirect leu-
prorelin pharmacokinetics in humans have been
previously reviewed by Chrisp and Sorkin,[30]
Plosker and Brogden[11] and Okada and Tog-
uchi.[33]
Another recent study evaluated formulation
variables by altering carrier characteristics, in-
creasing the drug load and adding calcium chloride
to organic and aqueous phases of the emulsion dur-
ing processing, yielding particles with increased
porosity, lower bulk density, higher specific sur-
face area and higher initial release. In an animal
model, these formulations showed a faster onset of
testosterone suppression compared with micro-
spheres, without higher drug load or calcium chlor-
ide release.[42]
Recent clinical studies in patients with ad-
vanced prostatic cancer demonstrate a therapeutic
equivalence of the 3-month high-dose (11.25, 22.5
and 30mg) and 1-month 3.75 or 7.50mg depot
leuprorelin formulations.[21,22,59] An implant that
delivers leuprorelin for 1 year in the palliative treat-
ment of prostate cancer was recently found to be
efficacious.[25,26]
4. Clinical Pharmacokinetics
Physiological production of gonadotropins re-
quires intermittent GnRH secretion while the pitu-
itary is continuously stimulated by either natural
GnRH or long-acting GnRH synthetic agonists
which desensitise gonadotropin secretion, a pro-
cess that has important clinical applications. The
frequency of the hypothalamic secretion of GnRH
is approximately circahoral.[6] Most of the infor-
mation regarding GnRH secretion in humans has
been inferred from an indirect approach in animal
experiments in which the pulsatile release of LH
in the peripheral circulation is measured, due to the
short plasma half-life of GnRH as well as the ana-
tomical inaccessibility of the hypophysioportal
circulation. GnRH is not bound to circulating pro-
teins but is readily accessible to plasma enzymes,
liable to rapid degradation and to a substantial di-
lution as it leaves the hypothalamic-pituitary por-
tal system. Thus, biologically important amounts
Table I summarises the studies evaluating the
pharmacokinetic properties of leuprorelin acetate
given as a single subcutaneous or intravenous in-
jection in healthy volunteers and patients with
prostatic cancer; in these studies, serum drug con-
centrations were measured with the RIA procedure
described by Yamazaki and Okada.[55]
4.1 Short-Acting Formulations
The study of Sennello et al.[27] was conducted
in a group of six healthy adult males and was of a
randomised, complete crossover design, each volun-
teer receiving bolus intravenous and subcutaneous
1mg doses of leuprorelin acetate administered as
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

496
Periti et al.
Table I. Summary of studies evaluating pharmacokinetic properties of leuprorelin given as a single subcutaneous dose in healthy volunteers
and patients with prostatic cancer. Serum leuprorelin was measured using RIA as described by Yamazaki and Okada[55]
Pharmacokinetic
parameter
Mean value
SD
Dosage Formulationa Study details
regimen
(mg)
References
Cmax (μg/L)
32.0 11.1
1
A
9 volunteers (6 healthy and 3 with prostatic cancer); Cmax
achieved within 30-60 min
27,61
13.09 6.16b 3.75
B
B
B
B
C
11 prostatic cancer patients; Cmax within 1h
5 prostatic cancer patients; Cmax within 1h
6 prostatic cancer patients; Cmax within 3h
1 prostatic cancer patient; Cmax within 3h
62,63
47.4 18.6b
54.5 26.5b
53.05b
7.5
15
30
20.8
11.25
25 prostatic cancer patients; Cmax within 3h; median Cmin at
week 12, 0.19 μg/L
23
10.7
3.75
11.25
1
B
C
A
15 prostatic cancer patients; Cmax within 1h; median Cmin at
week 4, 0.18 μg/L
23 prostatic cancer patients; Cmax within 3h; Cmin at week 13, 64
0.17 0.07 μg/L
6 healthy normal adult males; AUC∞ after intravenous
administration = 125.8 32.9 μg • h/L
21.8
AUC∞ (μg • h/L)
118.6 34.7
27
AUC35 (μg • h/L) 541.6 160.1 3.75
990.5 275.7 7.5
B
B
B
A
11 prostatic cancer patients
5 prostatic cancer patients
6 prostatic cancer patients
63
1323.8 565.1 15
F
0.96 0.26
1
6 healthy volunteers: AUC 118.6 34.7 μg • h/L
subcutaneous vs 125.8 32.9 μg • h/L intravenous,
assumed as 1.00
27
Vss (L)
37.1 16.8
1
A
6 healthy volunteers: Vss after intravenous administration
27
26.5 10.1L
36 5.1
3.75
7.50
15
B
B
B
11 prostatic cancer patients
5 prostatic cancer patients
63
33.5 3.5
27.3
2
6 prostatic cancer patients: inverse relationship of Vss to
dose (p < 0.01)
1
t
⁄ β (h)
2
3.6 1.2
1
A
6 healthy volunteers: 2.9 0.5h after intravenous
administration (harmonic mean half-life, calculated from
corresponding mean value of β-phase)
27
MRT (h)
4.2 0.8
1
1
A
A
6 healthy volunteers: 3.1 0.6h after intravenous
administration
CL [L/h (ml/min)]
9.1 2.6 (151
43)
6 healthy volunteers: 8.3 1.8 L/h (139 30 ml/min) after
intravenous administration
a
A = isotonic solution diluent (1 mg/ml); B = suspension of lyophilised 1-month depot powder in 2ml distilled water diluent; C = suspen-
sion of lyophilised 3-month depot powder in 2ml of distilled water diluent.
b
Mean leuprorelin concentrations declined after Cmax, reaching at 24 hours 1.6 0.8 (3.75mg dose), 2.71 0.8 (7.5mg dose), 3.3 1.1
(15mg dose) and 4.17 (30mg dose, one patient) μg/L.
AUC∞ = area under the concentration-time curve from zero to infinity; AUC35 = AUC from zero to 35 days; CL = plasma clearance; Cmax
=
1
peak plasma concentration; Cmin = trough plasma concentration; F = bioavailable fraction; MRT = mean residence time; t ⁄ β = terminal half-life;
2
Vss = volume of distribution at steady state.
1ml of a 1 mg/ml isotonic solution. Following sub-
cutaneous injection, leuprorelin was rapidly ab-
sorbed and the calculated circulating drug concen-
trations reached a mean peak concentration of 32.3
μg/L at 0.6 hours after subcutaneous injection. The
mean zero to infinity areas under the curve (AUC∞)
were similar (94.3%) between the subcutaneous
and intravenous groups, and plasma drug clearance
appeared to be independent of the administration
route; the difference between the two mean resi-
dence times (4.25 minus 3.11 hours = 1.14 hours)
represents the residence at the injection site of the
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

Depot Leuprorelin
497
subcutaneous doses. The mean drug fraction
reaching the general circulation (F = 0.96 0.26;
range 0.68 to 1.34) was not significantly different
from 1.0 for the subcutaneous doses. The mean
1
t ⁄ β following intravenous and subcutaneous doses
2
were 2.9 + 0.5 and 3.6 + 1.2 hours, respectively
30
20
10
(harmonic mean half-life, calculated from corre-
sponding mean value of β); the difference was not
significant.
Imai et al.[61] reported the serum concentrations
of leuprorelin acetate after a single 1mg subcuta-
neous injection (isotonic solution diluent) in three
patients with prostate cancer (figure 7), and judg-
ing from their clinical response estimated that this
peptide offers considerable therapeutic potential
for the treatment of prostatic carcinoma[65] when
the serum concentration is maintained at higher
than 1 μg/L.
0
0
2
4
6
8
10 12 14 16 18 20 22 24
Time (h)
Fig. 7. Serum leuprorelin concentration (radioimmunoassay
sensitivity 50pg per assay tube) after a 1mg subcutaneous in-
jection in three patients with prostatic carcinoma. Each point
represents the mean standard deviation (reproduced from
Imai et al.,[61] with permission from John Wiley & Sons, Inc.).
4.2 Long-Acting Formulations
Imai et al.[61] also studied a first model for con-
trolled slow release of leuprorelin from a drug-
vinyl polymer composite (VPC) made by radia-
tion-induced polymerisation of glass-forming
comonomers. Five patients with prostate cancer
received 150 or 100mg leuprorelin-polymer com-
posite subcutaneous implants (14mm diameter ×
4mm). Serum peptide concentrations higher than
1 μg/L were maintained over a period of several
months (figure 8). Unfortunately, the size of the
composite makes it impossible to inject and they
concluded that incision for implantation and re-
moval was not clinically convenient.
Some of the new types of small biodegradable
leuprorelin polymer composites have succeeded in
maintaining effective drug concentrations for more
than 4 weeks. Successful commercialisation has oc-
curred with the development of depot-injectable
PLGA or PLA microspheres of the GnRH ana-
logue agonist leuprorelin acetate as once monthly
(PLGA) or once every 3 or 4 months (PLA) injec-
tions that are easily performed using a fine 23-
gauge needle intramuscularly or subcutaneously.
These depots maintain persistent blood drug con-
centrations for 1, 3 or 4 months due to zero-order
release from drug doses of 3.75, 7.5 or
15mg,[11,23,30,32,62,63] 11.25mg[59,64] or 22.5[21] or
30mg,[22] respectively.
Mazzei et al.[62,63] evaluated the pharmaco-
kinetics, endocrine and antitumour effects of
leuprorelin PLGA depot in 22 patients with ad-
vanced prostatic cancer and 1 patient with benign
prostatic hypertrophy, giving the drug by subcuta-
neous injection at doses of 3.75, 7.5, 15 or 30mg.
Drug concentrations were measured by a double-
antibody RIA[55] over an observation period of 5
weeks. Serum concentrations of leuprorelin
showed a rapid increase immediately after injec-
tion, reaching a peak proportional to dose within 3
hours (range of mean values 13.1 to 54.5 μg/L).
Subsequently, mean drug concentrations declined
to a plateau proportional to dose (0.49 to 0.85 μg/L
at 5 weeks). A significant (p < 0.01) dose-dependent
increase from 541.7 to 1323.8 μg • h/L in the area
under the serum concentration-time curve from
zero to 35 days (AUC35) was also noted, as re-
ported in table I. The mean values of the zero-order
rate of leuprorelin release into serum increased
from 583.42 to 2658.76 μg/h in relation to dose (p
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

498
Periti et al.
and dihydrotestosterone were maintained in all pa-
tients for up to 5 weeks.
25
20
15
10
5
100mg
150mg
In another study, plasma leuprorelin concentra-
tions were monitored to determine if there was an
accumulation with repeated monthly single injec-
tions of 7.5mg as depot formulation.[20] Figure 10
shows the mean plasma leuprorelin acetate con-
centrations for 56 patients with prostatic cancer.
The initial rapid increase in leuprorelin concentra-
tions immediately after injection is the charac-
teristic pharmacokinetic pattern of depot formula-
tions containing a biodegradable copolymer of
lactic and glycolic acids such as PLGA. Plasma
leuprorelin concentrations decreased during the
initial 2 to 4 weeks and stabilised after week 8,
without accumulation.
In two recent pharmacokinetic studies in a total
of 48 patients with advanced prostate carcinoma
who received the 3-month PLA formulation con-
taining 11.25mg of leuprorelin, the depot subcuta-
neous injection provided a median serum Cmax
higher than 20 μg/L after 3 hours (tmax) and a steady
state concentration of around 0.2 μg/L starting on
day 7 and persisting at 0.17 to 0.19 μg/L until the
next scheduled administration.[23,64] Figure 11 il-
lustrates the release pattern of leuprorelin as me-
0
5
10
15
20
25
Time (wk)
Fig. 8. Serum leuprorelin concentration (radioimmunoassay
sensitivity 50pg per assay tube) after subcutaneous implanta-
tion of vinyl polymer composite containing 100 or 150mg of the
peptide in four patients with prostatic cancer (reproduced from
Imai et al.,[61] with permission from John Wiley & Sons, Inc.).
< 0.01) during the initial phase and lasting, as a
mean, 1.38 to 1.98 hours, while the estimated mean
duration of the slower release phase was 1077 to
1193 hours.[62] In all patients, serum leuprorelin
concentrations after injections of 3.75, 7.5 or 15mg
leuprorelin acetate depot reached a peak propor-
tional to the dose within 3 hours (figure 9). Mean
serum leuprorelin concentrations then fell rapidly
and by 24 hours were in the range 1.6 to 3.3 μg/L
for all three doses. After the first day, the decline
in serum concentrations of the nonapeptide oc-
curred more gradually and reached steady-state
concentrations of 0.49 to 0.85 μg/L at 5 weeks,
again proportional to dose.[63] The mean value of
the apparent volume of distribution (Vd/F) varied
between 27.3 and 36L, in inverse relationship (p <
0.01) to the leuprorelin dose. The values of AUC35
varied between 541.6 160 and 1323.8 565 μg
• h/L and this dose-related increase was statisti-
cally significantly different between patients re-
ceiving 3.75 and 7.5mg or those receiving 3.75 and
15mg, but not between those receiving 7.5 and
15mg.[63] Castration levels of serum testosterone
3.75mg leuprorelin acetate depot (n=11)
7.5mg (n=5)
15mg (n=6)
60
50
40
30
20
10
0
0
0.5
(h)
1
3
1
2
7
14 21 28 35
Time (days)
Fig. 9. Mean serum leuprorelin concentration (double-antibody
radioimmunoassay) as a function of time after subcutaneous
administration of 3.75, 7.5 or 15mg of the depot formulation
(reproduced from Mazzei et al.,[63] with permission from Cam-
bridge Medical Publications).
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

Depot Leuprorelin
499
2.0
1.0
0.0
seven patients with prostatic cancer after intramus-
cular administration of 3.75mg as depot formula-
tion.[66] The serum concentration of the pentapep-
tide 5-9 was 150 ng/L at 1 hour, reaching a peak
concentration of 860 ng/L at 3 hours. In a further
patient receiving the same leuprorelin dosage,
peak urine concentration of this metabolite was
4.97 μg/L within 2 days and remained detectable
at 1.74 μg/L after 29 days. Since other GnRH ago-
nists are predominantly excreted in urine, this
would also be expected to occur with leu-
prorelin.[11]
pre 0
2
4
6
8 10 12 14 16 18 20 22 24
Time (wk)
Clinical trials using leuprorelin depot have
demonstrated that a 1-month release formulation
is potentially useful for the therapy of prostate can-
cer, endometriosis and other endocrine-dependent
diseases without producing any serious adverse ef-
fects.[15] The serum testosterone suppression pat-
tern obtained with the leuprorelin 1mg daily injec-
tion (30 mg/month) and that with once-a-month
3.75 or 7.5mg depot overlapped almost perfectly,
effectively reducing the dose to one-eighth or one-
quarter of that required with the daily injec-
tion.[20,33] The microspheres with PLGA contain-
ing about 10% leuprorelin (3.75 or 7.5mg) were
demonstrated to provide a linear sustained release
for 4 weeks after injection: the serum drug concen-
trations were evaluated using a modified RIA sys-
tem.[29,54,55,62,63,67] Increasing the dose of leu-
prorelin acetate depot to more than 3.75mg once
every 4 weeks did not significantly increase objec-
tive response rate in patients with stage D2 pros-
tatic cancer.[15]
In a non-blind randomised phase II pharmaco-
kinetic study conducted in Germany and Italy, 42
patients with advanced or metastatic prostate can-
cer were treated for 9 months with subcutaneous
leuprorelin acetate depot as two different formula-
tions. Fifteen patients received the 1-month depot
and 27 patients received the newly developed 3-
month depot, containing 3.75 and 11.25mg of
leuprorelin acetate, respectively. Whether injected
monthly or at 3-month intervals, leuprorelin ace-
tate produced a complete downregulation of the
pituitary and led to persistent suppression of tes-
Fig. 10. Mean plasma leuprorelin concentrations at each week
for 56 patients with prostatic cancer. Drug concentrations were
measured by a double antibody radioimmunoassay; values are
means standard error. The arrows indicate the scheduled
injection day of 7.5mg of the peptide as a copolymer depot
formulation (reproduced from Sharifi et al.,[20] with permission).
dian serum concentrations for both the 1-month
depot (PLGA) and the 3-month depot (PLA),
which were comparably therapeutic and provided
a constant concentration during 9 months of treat-
ment in 40 patients with advanced prostatic can-
cer.[23] With the 1-month depot, the Cmax reached
10.7 μg/L (tmax 1 hour), a lower concentration than
with the 3-month depot, which reached a Cmax of
20.8 μg/L slightly later (tmax 3 hours). The median
leuprorelin troughs after 1 and 3 months of treat-
ment were 0.18 and 0.19 μg/L, respectively. Com-
paring the median leuprorelin concentrations at
steady state (from day 3 or 7 until before the next
injection) showed ranges of 0.18 to 0.38 μg/L for
the 1-month depot and 0.19 to 0.43 μg/L for the
3-month depot formulation.
Metabolism and excretion of leuprorelin have
been studied in animals but not extensively in hu-
mans.[11] In animals, leuprorelin is metabolised to
the pentapeptide 5-9, tripeptides 1-3 and 5-7, and
dipeptide 1-2.[66] The pentapeptide is expected to
be the major metabolite formed by peptidase
cleavage of the Ser4-Tyr5 bond, and further pepti-
dase activity could then result in smaller peptide
fragments.[30] The metabolism and excretion of
leuprorelin were determined by HPLC in only
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

500
Periti et al.
100
10
1
1-month depot (n = 1-15)
3-month depot (n = 1-25)
0.1
0.01
0
1
2
3
4
5
6
9
Time (mo)
Fig. 11. Release pattern of leuprorelin as median serum concentrations for two depot formulations injected subcutaneously monthly
(3.75mg) or at 3-month intervals (11.25mg) [reproduced from Tunn et al.,[23] with permission].
tosterone and dihydrotestosterone to the castration
range (≤50 ng/dl for testosterone) within the first
month of treatment, which thereafter could be
maintained over the entire observation period of 9
months.[23] The 3-month depot showed a higher
median Cmax of leuprorelin at 20.8 μg/L than the
1-month depot at 10.7 μg/L but, conversely, this
did not influence the flare-up rise in serum testos-
terone levels. Cmax occurred at 3 hours for the 3-
month and at 1 hour for the 1-month depot formu-
lations. During the steady state, constant release
was detected, starting on day 3 for the 1-month and
day 7 for the 3-month depots. A marked decrease
in median prostate-specific antigen (PSA) levels of
97.8% (1-month depot) and 96.6% (3-month de-
pot) compared with baseline was observed, indi-
cating an objective clinical response for more than
80% of all patients in both arms. Based on Eu-
ropean Organisation for Research and Treatment
of Cancer criteria, the best responses in terms of
complete/partial remissions and stabilisation were
comparable in the two arms at 86.7% (1-month de-
pot) and 85.2% (3-month depot); 6.7% of patients
in the 1-month group and 3% in the 3-month group
had disease progression. The most common ad-
verse effects in both treatment groups were related
to hormone deprivation.
A second prospective randomised multicentre
phase II study was conducted on 62 patients with
advanced prostate cancer in Germany. They re-
ceived long-term treatment with subcutaneous in-
jections of leuprorelin acetate 3-month depot as
part of a European trial.[24] The median survival
time was 3.1 years, time to tumour progression was
2.8 years and median period of progression-free
survival was 1.9 years, confirming an adequate
suppression of testosterone serum levels to the cas-
tration range.
An evaluation of a new leuprorelin acetate im-
plant that delivers the drug during 1 year for pal-
liative treatment of prostate cancer was recently
published.[25,26] The subcutaneous implant (4mm
× 45mm) uses osmotic pressure to deliver leu-
prorelin continuously at a controlled rate for 12
months and effectively suppressed testosterone in
99% of 80 patients within 2 to 4 weeks. Leu-
prorelin was rapidly absorbed, resulting in mean
serum concentrations of 16.8 μg/L at 24 hours;
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

Depot Leuprorelin
501
steady-state mean serum leuprorelin concentra-
tions were then maintained throughout the year, at
approximately 0.9 μg/L (figure 12). Most inser-
tion-site reactions were mild and transient, and
systemic adverse events were predictable effects
of testosterone suppression.
Depot formulations of leuprorelin 15 or 30mg,
composed of PLA with a molecular weight of
12 400 to 17 300, release the drug in an almost
zero-order manner without a large initial burst or
time lag. The microspheres sustained serum drug
concentrations and pharmacological effect for
over 3 or 4 months after injection.[23,64]
The response rate and incidence of adverse ef-
fects in these leuprorelin depot studies did not dif-
fer significantly from those occurring with the
daily formulation. Adverse effects, those to be ex-
pected as a result of chemical castration, were mild
and never troublesome, including hot flushes
(50%), sweating (10.7%), peripheral oedema
(7%), pain, constipation, dyspnoea, chest pain, im-
potence and increased urinary frequency.[15,21-
23,33,62,64,65,68] Local reactions (localised pain, ery-
thema and hardness) were more common after
application of the 3- or 4-month high-dose leu-
prorelin depots than with the 1-month de-
pot.[59,64,69]
Hormonal therapy remains the mainstay of
treatment of advanced prostate cancer, and the
widespread use of PSA testing has led to the iden-
tification of a large group of patients early in the
course of disease in whom the only evidence of
progression is increasing PSA. However, essen-
tially all patients treated with androgen depriva-
tion will eventually develop disease progression as
indicated by rising PSA levels, new lesions on ra-
diological studies and worsening symptoms. Thus,
new strategies for long-term hormone-delivering
therapy are needed.[70]
5. Conclusions
The unique feature of GnRH-producing cells is
that they release the hormone rhythmically in short
bursts. Only when the hypothalamic sexual centre
sends GnRH messages to the gonadotropes in a
pulsatile fashion do they respond with the synthe-
sis and rhythmic release of FSH and LH in the
amounts and ratios required for adequate gonadal
function in sexually mature women and men.
The pharmacokinetic characteristics of leu-
prorelin determine its mode of administration, fre-
quency of administration and clinical utility. The
most important clinical applications of this GnRH
agonist derive from its capacity, during prolonged
administration, to cause rapid desensitisation of
the pituitary gonadotropes with inhibiting effects
on the reproductive system. Preclinical study of
the pharmacokinetic properties of leuprorelin after
subcutaneous or intramuscular administration has
indicated that continuous administration or release
of this drug is necessary for pituitary desensitisa-
tion due to receptor downregulation. The effect on
serum levels of gonadotropins or sex hormones is
independent of drug dosage as part of a broader
biological phenomenon of adaptation by which
cells adjust their sensitivity to the level of the stim-
ulus. This phenomenon is termed ‘paradoxical’ be-
cause it is achieved with an inherently stimulating
drug. The biological effect is strictly time-dependent
and not concentration-dependent if the serum con-
centration is maintained higher than 1 μg/L, which
determines the administration interval.
25
20
15
Insertion of
new implant
4
2
0
0
2
4
8 12
20
28
36
44
52
60
Study week
Fig. 12. Serum leuprorelin concentrations measured in 80 pa-
tients on days 3 and 7, and in a subset of 21 patients also at 4,
24 and 48 hours and on day 5, after implantation; afterwards
samples were obtained only every month. Values are means
standard deviation (reproduced from Fowler et al.,[26] with per-
mission from Elsevier Science).
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

502
Periti et al.
6. Conn PM, Crowley WF. Gonadotropin-releasing hormone and
its analogues. N Engl J Med 1991; 324 (2): 93-103
7. Karten MJ, Rivier JE. Gonadotropin-releasing hormone analog
design. Structure-function studies toward the development of
agonists and antagonists: rationale and perspective. Endocr
Rev 1986; 7: 44-66
8. Lønning PE, Lien EA. Pharmacokinetics of anti-endocrine
agents. In: Workman P and Graham MA, guest editors. Can-
cer surveys, vol 17: pharmacokinetics and cancer chemother-
apy 1993. New York: CSHL Press, 1993
9. Fujino M, Fukuda T, Shinagawa S, et al. Synthetic analogs of
luteinizing hormone-releasing hormone (LH-RH) substituted
in position 6 and 10. Biochem Biophys Res Commun 1974;
60: 406-13
10. Handelsman DJ, Swerdloff RS. Pharmacokinetics of gonado-
tropin-releasing hormone and its analogs. Endocr Rev 1986;
7 (1): 95-101
11. Plosker GL, Brogden RN. Leuprorelin. A review of its pharma-
cological and therapeutic use in prostatic cancer, endometri-
osis and other sex hormone-related disorders. Drugs 1994; 48
(6): 930-67
12. Ascoli M, Segaloff DL. Adenohypophyseal hormones and their
hypothalamic releasing factors. In: Hardman JG, Goodman
Gilman A, Limbird LE, editors. The pharmacological basis of
therapeutics. 9th ed. New York: McGraw-Hill, 1996: 1363-80
13. Lu MF, Lee D, Rao GS. Percutaneous absorption enhancement
of leuprolide. Pharm Res 1992; 9 (12): 1575-9
14. Perrin MH, Rivier JE, Vale WW. Radioligand assay for gonado-
tropin-releasing hormone: relative potencies of agonists and
antagonists. Endocrinology 1980; 106: 1289-93
15. Rizzo M, Mazzei T, Mini E, et al. Leuprorelin acetate depot in
advanced prostatic cancer: a phase II multicenter trial. J Int
Med Res 1990; 18 Suppl. 1: 114-25
16. Covens A, Thomas G, Shaw P, et al. A phase II study of leupro-
lide in advanced/recurrent endometrial cancer. Gynecol
Oncol 1997; 64 (1): 126-9
17. Kuhn JM, Abourachid H, Brucher P, et al. A randomized com-
parison of the clinical and hormonal effects of two GnRH
agonists in patients with prostate cancer. Eur Urol 1997; 32
(4): 397-403
18. Horwich A, Huddart RA, Gadd J, et al. A pilot study of inter-
mittent androgen deprivation in advanced prostate cancer. Br
J Urol 1998; 81 (1): 96-9
19. Hornstein MD, Surrey ES, Weisberg GW, et al. Leuprorelide
acetate depot and hormonal add-back in endometriosis: a 12-
month study. Lupron Add-Back Study Group. Obstet
Gynecol 1998; 91 (1): 16-24
20. Sharifi R, Soloway M, and the Leuprolide Study Group. Clini-
cal study of leuprolide depot formulation in the treatment of
advanced prostate cancer. J Urol 1990; 143: 68-71
21. Sharifi R, Bruskewitz RC, Gittleman MC, et al. Leuprolide ac-
etate 22.5 mg 12-week depot formulation in the treatment of
patients with advanced prostate cancer. Clin Ther 1996; 18
(4): 647-57
Therefore, depot formulations that release leu-
prorelin at an almost constant rate for 1 month or
more improve convenience, compliance and effi-
cacy compared with daily injections of a regular
short-acting formulation. The serum sex hormone
suppression pattern obtained with the depot formu-
lation overlaps almost perfectly with that of the
daily dose, thus reducing the total dose by 8- to
4-fold.
The most recent clinical studies of the leu-
prorelin depot formulation composed of PLA show
that doses higher than 7.5mg sustain the pharma-
cological effect for over 3 or 4 months after a single
injection, with biologically equivalent serum pep-
tide concentrations, as demonstrated in the phar-
macokinetic analyses reported in the present re-
view. On the other hand, results from animal
experiments with high-dose leuprorelin depots can
be extrapolated to the clinic, where intramuscular
or subcutaneous single injections of depot formu-
lations containing 11.25 to 30mg are given every
1, 3 or 4 months.
The local and systemic tolerability of regular or
depot leuprorelin formulations is excellent, since
most adverse effects associated with the drug result
from changes in levels of circulating sex hor-
mones, reflecting the goal of the treatment.
Acknowledgements
No sources of funding were used to assist in the prepara-
tion of this article. There are no potential conflicts of interest
directly relevant to the contents of this manuscript.
References
1. Belchetz PE, Plant TM, Nakai Y, et al. Hypophysiological re-
sponses to continuous and intermittent delivery of hypotha-
lamic gonadotropin-releasing hormone. Science 1978; 202:
631-41
2. Henzl MR. Gonadotropin-releasing hormone and its analogues:
from laboratory to bedside. Clin Obstet Gynecol 1993; 36 (3):
617-35
22. Sharifi R, Knoll LD, Smith J, et al. Leuprolide acetate (30-mg
depot every four months) in the treatment of advanced pros-
tate cancer. Urology 1998; 51 (2): 271-6
23. Tunn UW, Bargelloni U, Cosciani S, et al. Comparison of LH-
RH analogue 1-month depot and 3-month depot by their hor-
mone levels and pharmacokinetic profile in patients with
advanced prostate cancer. Urol Int 1998; 60 Suppl. 1: 9-16
24. Jocham D. Leuprorelin three-month depot in the treatment of
advanced and metastatic prostate cancer: long-term follow-up
results. Urol Int 1998; 60 Suppl. 2: 18-24
3. Alberts B, Bray D, Lewis J, et al. The cell. 2nd ed. New York:
Garland Publishing, 1989
4. Conn PM, Hazem E. Luteinizing hormone release and gonado-
tropin-releasing hormone (GnRH) receptor internalization in-
dependent actions of GnRH [letter]. Endocrinology 1982;
109: 2040
5. Stojilkovic SS, Rojas E, Stutzin A, et al. Desensitization of
pituitary gonadotropin secretion by agonist-induced inactiva-
tion of voltage-sensitive calcium channels. J Biol Chem 1989;
264: 10939-42
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

Depot Leuprorelin
503
25. Fowler JE Jr, Gottesman JE, Reid CF, et al. Safety and efficacy
of an implantable leuprolide delivery system in patients with
advanced prostate cancer. J Urol 2000; 164: 730-4
44. Okada H, Doken Y, Ogawa Y, et al. Preparation of three-month
depot injectable microspheres of leuprorelin acetate using bi-
odegradable polymers. Pharm Res 1994; 11: 1143-7
45. Okada H, Doken Y, Ogawa Y, et al. Sustained suppression of
the pituitary-gonadal axis by leuprorelin three-month depot
microspheres in rats and dogs. Pharm Res 1994; 11: 1199-203
46. Toguchi H. Formulation study of leuprorelin acetate to improve
clinical performance. Clin Ther 1992; 14 Suppl. A: 121-30
47. Ogawa Y, Yamamoto M, Takada S, et al. Controlled-release of
leuprolide acetate from polyactic acid or copoly(lactic/gly-
colic) acid microcapsules: influence of molecular weight and
copolymer ratio of polymer. Chem Pharm Bull (Tokyo) 1988;
36 (4): 1502-7
48. Choi SH, Park TG. Hydrophobic ion pair formation between
leuprolide and sodium oleate for sustained release from bio-
degradable polymeric microspheres. Int J Pharm 2000; 203:
193-202
49. Okada H, Yamazaki I, Takatsura Y, et al. Vaginal absorption
of a potent luteinizing hormone-releasing hormone analogue
(Leuprolide) in rats. IV: evaluation of the vaginal absorption
and gonadotropin responses by radioimmunoassay. J Pharm
Sci 1984; 73 (3): 298-302
50. Okada H, Yamazaki I, Yashiki T, et al. Vaginal absorption of
a potent luteinizing hormone-releasing hormone analogue
(leuprolide) in rats. II: mechanism of absorption enhancement
with organic acids. J Pharm Sci 1983; 72 (1): 75-8
51. Shimamoto T. Pharmaceutical aspects: nasal and depot formu-
lations of leuprolide. J Androl 1987; 8 (1): S14-6
52. Yamazaki I. Serum concentration patterns of an LHRH agonist,
gonadotrophins and sex steroids after subcutaneous, vaginal,
rectal and nasal administration of the agonist to pregnant rats.
J Reprod Fert 1984; 72: 129-36
53. Meyer BR, Kreis W, Eschbach J, et al. Successful transdermal
administration of therapeutic doses of polypeptide to normal
human volunteers. Clin Pharmacol Ther 1988; 44 (6): 607-12
54. Okada H, Heya T, Ogawa Y, et al. One-month release injectable
microcapsules of a luteinizing hormone-releasing hormone
agonist (leuprolide acetate) for treating experimental endo-
metriosis in rats. J Pharmacol Exp Ther 1988; 244: 744-50
55. Yamazaki I, Okada H. A radioimmunoassay for a highly active
luteinizing hormone-releasing hormone analogue and rela-
tion between the serum level of the analogue and that of go-
nadotropin. Endocrinol Jpn 1980; 27: 593-605
56. Okada H, Inoue Y, Heya T, et al. Pharmacokinetics of once-a-
month injectable microspheres of leuprolide acetate. Pharm
Res 1991; 8: 787-91
57. Okada H, Heya T, Ogawa Y, et al. Sustained pharmacological
activities in rats following single and repeated administration
of once-a-month injectable microspheres of leuprolide ace-
tate. Pharm Res 1991; 8: 584-7
58. Okada H, Doken Y, Ogawa Y. Persistent suppression of the
pituitary-gonadal system in female rats by three-month depot
injectable microspheres of leuprorelin acetate. J Pharm Sci
1996; 85 (10): 1044-8
59. Fornara P, Jocham D. Clinical study results of the new formu-
lation leuprorelin acetate three-month depot for the treatment
of advanced prostate carcinoma. Urol Int 1996; 56 Suppl. 1:
18-22
60. Cho NB, Harada M, Imaeda T, et al. Discovery of a novel,
potent and orally active nonapeptide antagonist of the human
luteinizing hormone-releasing hormone (LHRH) receptor. J
Med Chem 1998; 41 (22): 4190-5
26. Fowler JE Jr, Flanagan M, Gleason DM, et al. Evaluation of an
implant that delivers leuprolide for 1 year for the palliative
treatment of prostate cancer. Urology 2000; 55: 639-42
27. Sennello LT, Finley RA, Chu SY, et al. Single-dose pharmacoki-
netics of leuprolide in humans following intravenous and sub-
cutaneous administration. J Pharm Sci 1986; 75 (2): 158-60
28. Stetler-Stevenson MA, Fluoret G, Nakamura S, et al. In vivo
metabolism of tritiated LHRH by the whole kidney and indi-
vidual tubules of rats. Am J Physiol 1983; 244: F628-32
29. Adjei A, Sundberg D, Miller J, et al. Bioavailability of leuprol-
ide acetate following nasal inhalation delivery to rats and
healthy humans. Pharm Res 1992; 9: 244-9
30. Chrisp P, Sorkin EM. Leuprorelin: a review of its pharmaco-
logy and therapeutic use in prostatic disorders. Drugs Aging
1991; 1 (6): 487-509
31. Qiu Y, Johnson HW, Reiland TL, et al. Sublingual absorption
of leuprolide: comparison between human and animal mod-
els. Int J Pharm 1999; 179: 27-36
32. Okada U, Yamamoto M, Heya T, et al. Drug delivery using
biodegradable microspheres. J Control Release 1994; 28:
121-9
33. Okada H, Toguchi H. Biodegradable microspheres in drug de-
livery. Crit Rev Ther Drug Carrier Syst 1995; 12 (1): 1-99
34. Ogawa Y, Okada H, Yamamoto M, et al. In vivo release profiles
of leuprolide acetate from microcapsules prepared with poly-
lactic acids or copoly(lactic/glycolic) acids and in vivo deg-
radation of these polymers. Chem Pharm Bull (Tokyo) 1988;
36: 2576-81
35. Ogawa Y, Yamamoto M, Okada H, et al. A new technique to
efficiently entrap leuprolide acetate into microcapsules of
polyactic acid or copoly(lactic/glycolic) acid. Chem Pharm
Bull (Tokyo) 1988; 36: 1095-103
36. Ogawa Y, Okada H, Heya T, et al. Controlled release of LHRH
agonist, leuprolide acetate, from microcapsules: serum drug
level profiles and pharmacological effects in animals. J
Pharm Pharmacol 1989; 41: 439-44
37. Okada H. Once-month release injectable microspheres of
leuprolide acetate, a superactive agonist of LH-RH. Proc Int
Symp Control Rel Bioact Mater 1989; 16: 12-3
38. Okada H, Heya T, Igari Y, et al. One-month release injectable
microspheres of leuprolide acetate inhibit steroidogenesis
and genital organ growth in rats. Int J Pharm 1989: 231-9
39. Burton KW, Shameem M, Thanoo BC, et al. Extended release
peptide delivery systems through the use of PLGA micro-
sphere combinations. J Biomater Sci Polym Ed 2000; 11 (7):
715-29
40. Ravivarapu HB, Moyer KL, Dunn RL. Parameters affecting the
efficacy of a sustained release polymeric implant of leupro-
lide. Int J Pharm 2000; 194 (2): 181-91
41. Ravivarapu HB, Moyer KL, Dunn RL. Sustained suppression
of pituitary-gonadal axis with an injectable, in situ forming
implant of leuprolide acetate. J Pharm Sci 2000; 89 (6): 732-41
42. Ravivarapu HB, Lee H, DeLuca PP. Enhancing initial release
of peptide from poly(d,l-lactide-co-glycolide) (PLGA)
microspheres by addition of a porosigen and increasing drug
load. Pharm Dev Technol 2000; 5 (2): 287-96
43. Okada H, Inoue Y, Ogawa Y, et al. Three-month release inject-
able microspheres of leuprorelin acetate. Proc Int Symp Con-
trol Rel Bioact Mater 1992; 19: 52-3
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)

504
Periti et al.
61. Imai K, Yamanaka H, Yuasa H, et al. The sustained release of
LH-RH agonist from LH-RH agonist-polymer composite in
patients with prostatic cancer. Prostate 1986; 8: 325-32
62. Mazzei T, Mini E, Eandi M, et al. Pharmacokinetics, endocrine
and antitumour effects of leuprolide depot (TAP-144-SR) in
advanced prostatic cancer: a dose-response evaluation. Drugs
Exp Clin Res 1989; XV (8): 373-87
63. Mazzei T, Mini E, Rizzo M, et al. Human pharmacokinetic and
pharmacodynamic profiles of leuprorelin acetate depot in
prostatic cancer patients. J Int Med Res 1990; 18 Suppl. 1:
42-56
teinizing hormone-releasing hormone analogue, leuprorelin,
and its metabolite. J Chromatogr 1991; 566: 57-66
67. Marshall JC, Odell WD. Preparation of biologically active
[
125I]LH-RH suitable for membrane binding studies. Proc Soc
Exp Biol Med 1975; 149 (2): 351-5
68. Giraud B. Interim report of a large French multicentre study of
efficacy and safety of 3.75 mg leuprorelin depot in metastatic
prostatic cancer. J Int Med Res 1990; 18 Suppl. 1: 84-9
69. Kienle E, Lübben G, The German Leuprorelin Study Group.
Efficacy and safety of leuprorelin acetate depot for prostate
cancer. Urol Int 1996; 56 Suppl. 1: 23-30
70. Harris KA, Small EJ. Hormonal treatment for prostate cancer.
64. Khan MS, O’Brien A. An evaluation of pharmacokinetics and
pharmacodynamics of leuprorelin acetate 3M-depot in pa-
tients with advanced and metastatic carcinoma of the prostate.
Urol Int 1998; 60 (1): 33-40
Expert Opin Investig Drugs 2001; 10 (3): 493-510
65. Leuprolide Study Group. Leuprolide versus diethylstilbestrol for
metastatic prostate cancer. N Engl J Med 1984; 311: 1281-6
66. Ueno H, Matsuo S. High-performance liquid chromatography
followed by radioimmunoassay for the determination of a lu-
Correspondence and offprints: Dr Piero Periti, Via XX
Settembre 104, 50129 Firenze, Italy.
© Adis International Limited. All rights reserved.
Clin Pharmacokinet 2002; 41 (7)