Growth Hormone Secretagogues in Clinical Practice

463 Pages·1998·20.077 MB·\463

by  Barry B. Bercu (Author), Richard F. Walker (Author)

rovvth
Hormone
Secretagogues
in Clinical Practice

rowth
Hormone
Secretagogues
in Clinical Practice
edited by
Barry B. Bercu
University of South Florida College of Medicine
Tampa, and
All Children's Hospital
St. Petersburg, Florida
Richard F. Walker
University of South Florida
Tampa, Florida
MARCEL
MARCEL DEKKER, INC. NEw YoRK· BAsEL· HoNG KoNG
DEKKER
The editors acknowledge the generous support of Pharmacia & Upjo/111, Inc.,
for an unrestricted educational grant.
ISBN: 0-8247-9832-5
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Preface
The symposium on which this book is based, held in Tampa, Florida, was the
second in a series devoted to achieving a better understanding of growth hor
mone (GH) secretagogues and the mechanisms by which they stimulate pitu
itary release of somatotropin. These objectives were achieved through the pre
sentation of recent findings and the discussion of contemporary issues in this
important area of endocrinology.
Historically, net GH release from the pituitary was thought to represent
the balance of single inhibitory and stimulatory influences. Although a stimu
latory agent, growth hormone-releasing factor (GRF) had been postulated in the
early 1960s but it was not until 1982 that growth hormone-releasing hormone
(GHRH) was isolated and characterized. That discovery served as the basis for
a regulatory system in which net GH secretion represented a balance of stimu
latory and inhibitory signals from two different hypothalamic peptides-GHRH
and somatostatin, respectively. Subsequent analysis of the GHRH molecule
revealed that the structural requirements for GH-stimulating activity resided in
the first 29 amino acids of the peptide.
Shortly after GHRH was characterized and identified, synthetic peptides
capable of stimulating GH secretion in vitro and in vivo were derived from met
enkephalin. It was known that other molecules such as arginine, morphine,
clinidine, and levodopa could also provoke GH secretion, but these increased
GH release by altering the secretion of endogenous GHRH or somatostatin.
Thus, they were not true GH secretagogues per se, and they were incapable of
directly altering somatotroph function in vitro when hypothalamic peptides were
absent. In contrast, the opioid-derived peptides were capable of directly stimu
lating GH release from cultured pituitary tissue and cells, even though none of
them shared structural homology with the active portion of the stimulatory
GHRH molecule (i.e., amino acids, 1-29). Furthermore, binding and functional
iii
iv Preface
studies demonstrated that GH-releasing activity and opioidergic character di
verged as GHRP efficacy and potency increased. The new molecules, which
were called GH-releasing peptides (GHRPs) to distinguish them from the natu
rally occurring hypothalamic peptide, GHRH, seemed to be xenobiotic analogs
of another, still-unidentified GH secretagogue.
However, the acceptance of another endogenous secretagogue for GH that
complemented GHRH function was not immediately forthcoming. The extremely
low activity of the early analogs, coupled with a poor understanding of their
relationship to endogenous GHRH and how it created differences in in vitro
activity, caused initial findings to be viewed skeptically and, occasionally,
dismissed as contamination artifacts of natural GHRH within the in vitro test
ing system.
Despite initial skepticism, Dr. Cyril Y. Bowers, who headed the team that
synthesized GHRP and discovered its potential as a GH secretatogue, persevered
in his efforts to have the scientific community recognize the importance of his
findings. After a decade of work demonstrating the efficacy of GHRP through
the publication of many important papers, his labors finally bore fruit.
One of us (A VS) has been fortunate to share a close friendship with Dr.
Bowers for more than 30 years and was privileged to collaborate with him during
the 1960s and early 1970s. This collaboration led to the isolation, elucidation
of structure, and synthesis of thyrotropin-releasing hormone (TRH), the first
hypothalamic hormone to be completely identified. It also placed the projects
involving other hypothalamic hormones on a solid foundation.
Through his vision and untiring devotion to the advancement of knowl
edge in the science of endocrinology, Dr. Bowers has earned the respect and
admiration of his colleagues. Accordingly, the symposium was conceived as a
tribute to Dr. Bowers and dedicated in his honor.
Appropriately, the meeting began with Dr. Bower's historical perspective
on the discovery and development of GHRPs as therapeutic agents. He explained
that the prototype molecule of the GHRP-type GH secretagogues, growth hor
mone-releasing hexapeptide (GHRP-6, His-D-Trp-Ala-Trp-D-Phe-Lys-NH2), was
used for pharmacological analysis of this new family of potential drug sub
stances. Although much was learned about their activity, there was little inter
est in clinical application of GHRPs until oral bioavailability was later demon
strated in rats, dogs, monkeys, and finally, humans, leading to a concerted effort
by several pharmaceutical companies to develop better orally bioavailable
molecules. Those efforts culminated in the discovery of orally active, non
peptidyl, and truncated peptide GH secretagogues. Despite the apparent struc
tural diversity of the xenobiotic molecules, they all shared the pharmacodynamic
and mechanistic characteristics of GHRP-6, which were distinct from those for
GHRH. Furthermore, the GH secretagogue characteristics of GHRPs were not
shared by the other molecules that were capable of indirectly stimulating GH
Preface v
secretion, such as TRH, arginine, clonidine, morphine, and others. Although
GHRP had certain constraints on its ability to stimulate GH secretion, it was a
reliable secretagogue that produced unique patterns of GH secretion that were
distinctly different from those produced by GHRH. Subsequent comparative
analysis of GHRH and GHRP efficacies as GH secretagogues revealed that they
acted through clearly dissimilar signal transduction mechanisms.
Following Dr. Bower's interesting and thorough review of GHRP evolu
tion, contemporary issues related to the broad scope of GH secretagogue ac
tion were presented for discussion. The topics discussed during this first ses
sion included an analysis of structural requirements for GH secretagogue
efficacy, the use of mathematical models for predicting the differential values
of individual stimuli for GH secretion from the pituitary, and the presentation
of a unifying mechanism by which peptidyl and nonpeptidyl GH secretagogues
of the GHRP class elicited their action. Structural analyses of the xenobiotic
GHRPs indicated that small-molecule mimetics of the prototypic GHRP-6 could
be useful for overcoming many of the potential disadvantages associated with
the peptides, which are poorly absorbed and rapidly metabolized, and thus suffer
from low bioavailability. In an attempt to circumvent these problems, a new
series of nonpeptidyl GHRPs was derived from a molecule-modeling analysis
of reported small-molecule mimetics. These included the benzolactam L-692,429
and the acyclic dipeptides L-162, 752 and MK-0677. From a satisfactory con
formational overlay, a series of new compounds containing substituted tetra
hydroisoquinoline and isoindoline rings was synthesized. These nonpeptidyl
mimetics of GHRP were proposed as potentially useful tools to investigate the
utility and mechanisms of action of non-GHRH GH secretagogues.
Data were presented within this context to show that all of the non-GHRH
type secretagogues used a common receptor type that had binding and disso
ciation characteristics different from those identified for GHRH-related secre
tagogues. Furthermore, phosphotidyl inositide second messengers were shown
to mediate intracellular transduction of the GHRP message, compared with cyclic
nucleotides that were used by GHRH. As an extension of the concept that unique
binding sites subserve GHRP activity, the results of a study providing the mo
lecular characterization of the new receptor were presented. By utilizing Xeno
pus oocyte expression cloning, a swine pituitary eDNA was isolated that en
coded a protein with a high-affinity binding site for MK-0677, a nonpeptidyl
GHRP. From its pharmacology for GHRP binding, the investigators concluded
that the eDNA encoded a swine GHRP receptor. Similar receptors were also
identified for human and rat GHRP eDNA clones. The GHRP receptor repre
sents a G-protein-coupled receptor that is most closely related to the neuro
tension receptor (35% amino acid sequence identity) and is encoded by a single
highly conserved gene of diverse vertebrate species. This receptor is expressed
in pituitary tissues, as well as in areas of the brain that include the hypothala-
vi Preface
mus, hippocampus, and other sites. The central message of this study was that
the presence of a GHRP receptor implies the existence of a naturally occurring
ligand that may be part of the GH secretion regulatory complex.
The initial session was followed by one in experimental pharmacology and
involved topics such as agonists of growth hormone-releasing hormone, inter
actions of the exogenous GH secretagogues, evidence for extrahypophyseal sites
of action for the GHRP-like molecules, and an analysis of the physiological state
as it affects GHRH and GHRP activity. The presentations were wide and var
ied. It was recognized that such efforts, as described in the foregoing, had been
devoted to the design, synthesis, and evaluation of the different families of GH
secretagogues. However, suitable screening systems for the rapid evaluation of
numerous novel compounds representing potential candidates were lacking.
Thus, one laboratory reported the results of its efforts to develop a simple
screening system based on polyclonal antibodies. For this purpose, a pentapep
tide (Gly-Ala-Asn-Ala-Gly) was bound to the COOH or NH-terminus of GHRP-
6, and the resulting undecapeptides, termed P1 and P2, respectively, were
further conjugated with bovine serum albumin to serve as antigens for the
immunization of rabbits and preparation of antisera.
Another presentation focusing on GHRH reported that many agonistic
analogs intended for potential clinical and veterinary applications had been
synthesized by various laboratories. Analogs with enhanced biological activi
ties and, thus, greater potential therapeutic usefulness were shown to result from
derivatization of functional groups or exchanges of amino acids. Many differ
ent analogs were identified, and their potential role as therapeutic agents sug
gested. Similarly, a clinical need for antagonistic analogs of GHRH was re
ported. Among proposed applications for these molecules were acromegaly,
diabetic retinopathy, diabetic nephropathy (glomerulosclerosis), and in cancer
treatment GHRH antagonists could be given alone or in combination with
superactive somatostatin analogs to suppress GH and insulin-like growth factor
I (IGF-1) levels in patients with neoplasms potentially dependent on IGF-1.
GHRH antagonists could also be used for inhibiting growth of tumors that do
not express somatostatin receptors, such as osteosarcomas or pancreatic can
cers. The main point of this presentation was to summarize the work of one of
us (A VS) in synthesis of these antagonists and also to review oncological in
vestigations performed in various cancer models with the new antitumor agents.
Potential clinical applications of antagonists in treating IGF-1-dependent can
cers were discussed in light of increasing evidence of the involvement of growth
factors IGF-I and IGF-II involvement in the progression and metastases of
various malignancies.
Consistent with the discovery of GHRP receptors in the hypothalamus,
the more pronounced effect of GHRP in vivo compared with in vitro, and
GHRH and GHRP synergy in vivo, was the direct demonstration of GHRP
Preface vii
activity in brain structures through enhancement of cjos expression and elec
trical activity in hypothalamic neurons. GHRP was reported to behave like a
hypothalamic neurohormone that released GH in two phases characterized by
early hypersensitivity and late hyposensitivity. These responses were typical of
receptors activated by neurohormones, and the investigator provided evidence
for extrahypophyseal site of action of non-GHRH GH secretagogues. Further
direct evidence was provided by the use of electrophysiological tests. Neuro
secretory neurons in the arcuate nucleus were identified antidromically, and their
electrical characteristics were recorded. Physiological evidence for the effects
of GHRP on behavior was provided in a series of tests involving voluntary and
forced exercise. The data showed that GHRP has an effect on the ability to
perform a behavioral task; however, it was unclear whether this improvement
resulted from enhanced physical capacity, attitude, or a combination of these
neurophysiological factors. Finally, action of GH secretagogues in the brain were
discussed by determination of the activity of these substances on hypothalamic
hormones in hypothalamic incubation systems. GHRP stimulated arginine va
sopressin release, but had no stimulatory effect on corticotropin-releasing hor
mone. In addition, release of posterior pituitary hormones was inhibited by small
synthetic molecules capable of stimulating GH release from the anterior pitu
itary.
After the basic aspects of GH secretagogue endocrinology had been dis
cussed, the meeting turned to topics of clinical application for GH secretagogues.
Within this context, investigators presented a novel diagnostic test for evaluat
ing pituitary function in slowly growing children and aging adults. The objec
tive was to show that differential secretory responses to GHRH and GHRP could
be used to allow appropriate selection of therapeutic entities, ranging from
GHRH and GHRP alone, the combination of GHRH, or recombinant GH, when
patients lack a pituitary mechanism for GH production or secretion. In another
study, the presence of a significant GH response to GHRP in at least a subset
of patients suggested the possibility of anabolic benefits from multiple doses
given over time. Another study reported different effects of GHRH and GHRP
on sleep, indicating that the two secretagogues have different activities here.
GHRH was effective in producing slow-wave sleep, whereas GHRP was asso
ciated with rapid eye movement (REM) sleep. Finally, reports were presented
on interactions of GH secretagogues with other hormones and endogenous
substances, ranging from sex steroids to interferons.
In conclusion, this book provides the reader with information based on
presentations from the Second International Symposium on GH Secretagogues.
Through those details provided by pioneers in the field, one can view the ori
gins of a new area of GH research involving the xenobiotic GH secretagogues.
Dr. Bowers is recognized for his significant contributions to this research and,
accordingly, we thank him, the scientific committee, session chairs, speakers,