| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
ARTICLES |
Endocrine, Polypeptide and Cancer Institute, Veterans Affairs Medical Center (Z.R., J.L.V., A.V.S., G.H., P.A., K.G., T.C.), and Department of Medicine, Tulane University School of Medicine (Z.R., J.L.V., A.V.S., G.H., T.C.), New Orleans, Louisiana 70112
Address all correspondence and requests for reprints to: Dr. Andrew V. Schally (151), Veterans Affairs Medical Center, 1601 Perdido Street, New Orleans, Louisiana 70112-1262.
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Receptors for VIP (VPAC-R), with two subtypes, VPAC1-R and VPAC2-R, are abundant in many types of cancers (20, 21, 22, 23, 24). It was also reported that VIP (22, 25) and its antagonists (26, 27) can inhibit the growth of various human carcinomas. Because of the structural similarity between VIP and GHRH as well as considerable homology between their receptor proteins, GHRH analogs may interact with VPAC-R (28, 29). This might contribute to their antiproliferative effect on human tumors. This prompted us to study the possible interactions of GHRH antagonists with VPAC-R on cancer cells.
We recently synthesized three new GHRH analogs, JV-1–50, JV-1–51, and JV-1–52, which are derivatives of the potent GHRH antagonists JV-1–36, JV-1–38, and JV-1–42 (8), but contain D-Phe2 instead of D-Arg2 (30). Analogs JV-1–50 and JV-1–51 are structurally related to JV-1–42 and JV-1–36, whereas JV-1–52 is derived from JV-1–38. Analogs JV-1–50, JV-1–51, and JV-1–52 have reduced antagonistic activities on GHRH-R and increased antagonistic properties on VPAC1-R and VPAC2-R compared with their parent compounds. We also synthesized a potent VPAC1-R and VPAC2-R antagonist, JV-1–53, that is devoid of detectable antagonistic activity on GHRH-R (30). The inhibitory activities of these peptides on GHRH-R, VPAC1-R, and VPAC2-R were evaluated on rat pituitary and pineal cells (30) and compared with that of a selective VPAC1-R antagonist, PG 97–269 (31).
According to earlier reports, human prostate has relatively rich VIP-like immunopositive innervation (32, 33), and VPAC-R have been identified in both normal and malignant human prostate cells and tissue (20, 34). VIP stimulates prostatic secretion and increases the proliferation of prostatic epithelial cells in culture (35, 36). VIP is also known to potentiate the invasive capacity of the androgen-responsive human carcinoma cell line LNCaP (37). In addition, VIP increases the secretion of human prostate-specific antigen (PSA) and the production of cAMP in LNCaP cancer cells (38). PSA is produced primarily by prostatic epithelial cells (39) and is established as a marker for the early detection of prostate cancer (40).
In this study we compared the inhibitory effects of GHRH antagonists and VPAC-R antagonists on the proliferation and PSA secretion of VPAC-R-positive LNCaP human prostatic cancer cells in static culture. In addition, the inhibitory effects of the analogs on the growth of the MiaPaCa-2 human pancreatic cancer cell line, which served as a VPAC-R- negative (41) control, were evaluated. The expression of VPAC1-R, VPAC2-R, and GHRH-R on LNCaP and MiaPaCa-2 cancer cells was studied by RT-PCR and radioligand competition assay. We also measured, in a superfusion system, the inhibitory activity of the antagonists on the rate of inducible PSA release from LNCaP cells and evaluated their VPAC-R binding affinity. The overall aim of this project was comparison of the specificity and antitumor activity of GHRH antagonists and VIP antagonists.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Tissue cultures
Human prostatic (LNCaP) and pancreatic (MiaPaCa-2) cancer cell
lines were obtained from American Type Culture Collection
(Manassas, VA). The cell culture media and reagents were purchased from
Life Technologies, Inc. (Grand Island, NY). The LNCaP cell
line was grown in RPMI 1640 medium supplemented with 10% FBS and a
mixture of antibiotics and antimycotics (100 U/ml penicillin G, 100
µg/ml streptomycin, and 0.25 µg/ml amphotericin). The
MiaPaCa-2 cell line was routinely maintained in DMEM containing 1
mM pyruvate, 10% FBS, 2.5% horse serum, and a mixture of
antibiotics and antimycotics as described above. The cultures were
maintained in a humidified atmosphere containing 95% air-5%
CO2 at 37 C. The cells were passaged weekly and
were routinely monitored for the presence of mycoplasma using a test
kit from Roche Molecular Biochemicals (Indianapolis,
IN).
Cell proliferation assay
For all experiments, cells were grown to 80–90% confluence,
harvested by use of trypsin-EDTA solution, and seeded at low
concentration in 96-well plates. After 24 h, the culture medium
was removed and replaced with test medium (for LNCaP: RPMI 1640 and 2%
heat-inactivated dextran-coated charcoal-treated FBS; for MiaPaCa-2:
DMEM, 1 mM pyruvate, 1.6% FBS, and 0.4% horse serum)
containing the test compounds (in octuplicate wells each) at 3- and
10-µM concentrations or the vehicle (0.1%
dimethylsulfoxide) as a control for 72 h. Cell growth was
determined by a microculture tetrazolium assay based on a method
described by Plumb et al. (42). This assay measures the
reduction of substrate
[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide]
(MTT) to a dark blue formazan product by mitochondrial
dehydrogenases in living cells. Briefly, after 72-h treatment with
various compounds, 40 µl 2 mg/ml MTT (Sigma, St. Louis,
MO) were added to each well, and the cells were incubated for 4 h
at 37 C in darkness. The medium was replaced by 200 µl DMSO and then
with 25 µl Sorensen’s glycine buffer (0.1 M
glycine plus 0.1 M NaCl, pH 10.5). After shaking
to dissolve the formazan, absorbance was read at 540 nm in the plate
reader (Beckman Coulter, Inc., Palo Alto, CA). Results
were calculated as a percentage of the control, where the
OD540nm of treated cultures (medium plus
compounds) was related to the OD540nm of the
control culture (medium plus vehicle) x 100. Aliquots of medium
were kept frozen for PSA immunoradiometric assay (IRMA) at -20 C.
Superfusion
Superfusion analysis of cultured LNCaP cells was performed in a
system similar to that described previously for studying dispersed
anterior pituitary cells (43, 44). Tissue culture medium and reagents
for the superfusion technique were obtained from Sigma. In
brief, cultured tumor cells were harvested and resuspended in 1.5 ml
medium, and the cells were transferred onto superfusion columns (4.5–5
million cells each) and allowed to sediment simultaneously with 0.8 ml
Sephadex G. The dead volume of the system was set at 1 ml. Medium 199
containing BSA (1 g/liter), NaHCO3 (2.2 g/liter),
penicillin G (50 mg/liter), and gentamicin sulfate (87 mg/liter) was
equilibrated with a mixture of 95% air-5% CO2
and used as the culture medium. The medium was pumped at a flow rate of
1 ml/3 min. After an overnight recovery period, in which the baseline
stabilized and the cells regained their full responsiveness, 2-ml
fractions of the effluent medium were collected every 6 min. The cells
were exposed periodically to VIP (1 nM) or GHRH (10
nM), which was dissolved in fresh medium immediately before
application, for 12 min at 60-min intervals. The antagonists were
infused at 100 or 300 nM for 12 min (two fractions),
immediately followed by the mixture of an antagonist and 1
nM VIP or 10 nM GHRH for an additional 12 min.
Fractions of effluent medium were kept frozen for PSA IRMA at -20 C.
Each experiment was performed in three superfusion columns
simultaneously.
Receptor binding
Preparation of membrane fractions of human prostatic (LNCaP) and
pancreatic (MiaPaCa-2) cancer cells and receptor binding of VIP and
GHRH were performed as previously reported (45, 46). Radioiodinated VIP
was purchased from Amersham Pharmacia Biotech (Arlington
Heights, IL), and 125I-labeled
[His1,Nle27]hGHRH-(1–32)NH2
was prepared as previously described (46). The protein concentration
was determined using a Bio-Rad Laboratories, Inc., protein
assay kit. The LIGAND PC computerized curve-fitting program
developed by Munson and Rodbard (47) was used to determine the type of
receptor binding, the dissociation constant (Kd),
and the maximal binding capacity of receptors. The receptor binding
affinity of GHRH-related peptides to tumor membranes of LNCaP cells was
measured in displacement experiments based on competitive inhibition of
radiolabeled VIP binding using various concentrations
(10-12-10-6
M) of GHRH analogs. IC50, defined as
the dose causing 50% inhibition of specific binding of
[125I]VIP, was calculated by a computerized
curve-fitting program (48). Relative affinities of GHRH analogs for
VPAC-R compared with VIP were calculated as the ratio of the
IC50 of the tested compounds to the
IC50 of VIP (1.07 nM).
RT-PCR analysis
Total RNA of cultured LNCaP and MiaPaCa-2 cancer cells was
extracted according to the Tri-Reagent protocol (Sigma).
The concentration of RNA was determined by spectrophotometric analysis
at A260/280nm. One microgram of total RNA was
reverse transcribed and then amplified using the reagents and protocol
of the GeneAmp RNA PCR Core kit (Perkin-Elmer Corp.,
Norwalk, CT). The RT reaction was performed in a final volume of 20
µl containing 2.5 µM random hexamers, 1 mM
each of deoxynucleoside triphosphate, 1 x PCR buffer, 5
mM MgCl2, 1 U/µl ribonuclease
inhibitor, and 2.5 U/µl Moloney leukemia virus reverse transcriptase.
One fourth (5 µl) of the RT reaction was used for each PCR
amplification with a primer set that would amplify complementary DNA
(cDNAs) for human VPAC1-R and
VPAC2-R, GHRH-R, or ß-actin (Table 1
). The PCR reaction included 1 x
PCR buffer, 2 mM MgCl2, 0.15
µM (for VPAC1-R and ß-actin), or
1.0 µM (for VPAC2-R and GHRH-R) of
each primer and 2.5 U/100 µl AmpliTaq DNA polymerase in a 25-µl
volume. The PCR amplification was performed in a GeneAmp PCR System
2400 (Perkin-Elmer Corp.) with the following cycle
profile: 95 C for 180 sec, followed by 45 (receptors) or 25 (ß-actin)
cycles of 95 C for 30 sec, 60 C for 30 sec, and 72 C for 45 sec. After
the last cycle, there was a final extension for 7 min at 72 C. PCR
products were electrophoresed on a 1.5% agarose gel stained with 0.5
µg/ml ethidium bromide and visualized under UV light, followed by
scanning and quantification of gel (GDS 7500 Gel Documentation System,
UVP, Upland, CA; and GS-700 Imaging Densitometer, Bio-Rad Laboratories, Inc., Hercules, CA). The quality of RNA extracted
was tested by PCR amplification of human ß-actin cDNA (49) from the
same RT reaction as that used for various receptor cDNA
amplifications.
|
Statistical analysis
IRMA results from superfusion system were further analyzed with
computer software developed in our institute (43). In brief, the
program first separates the fractions containing a basal level of the
secreted PSA from the response fraction by using several iterations.
The statistical parameters of the baseline are determined from the
values of the basal fractions. A fraction is considered as a response
if it is larger or smaller than the 95% confidence limits of the
baseline. Further statistical analysis is based on the net integral
values (area of the response curve above the baseline) of the responses
(NET INT). Results of IRMA and MTT test from static cell culture were
subjected to one-way ANOVA, and differences between groups were
determined by Tukey’s test. P < 0.05 was considered
significant. Data are expressed as a percentage of the control values
(set at 100%) and represent the mean ± SEM
PSA content or the formazan level of octuplicates of wells with the
same treatment.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
). Both peptides caused a significant
increase in the rate of PSA secretion (total PSA secretion per cell
number) in cell culture [3 µM GHRH, 1.9-fold; 10
µM GHRH, 2.2-fold (P < 0.001); 3
µM VIP, 1.7-fold (P < 0.05);
10 µM VIP, 2.0-fold (P <
0.001); Fig. 1, B and C]. GHRH antagonists JV-1–38, JV-1–36, and
JV-1–42 at 10-µM concentrations significantly
inhibited cell proliferation by 73%, 54%, and 44%, respectively
(P < 0.001); JV-1–38 was the most potent inhibitor
of the peptides tested (Fig. 1A). Their structurally related
analogs JV-1–52, JV-1–51, and JV-1–50, with decreased GHRH-R
antagonistic potency and increased VPAC-R antagonistic activity,
produced a weaker inhibition of cell proliferation than the parent
peptides (Fig. 1A). JV-1–52 was the most potent inhibitor among these
nonselective antagonistic analogs of VPAC-R/GHRH-R (Fig. 1A). However,
its antiproliferative effect was weaker than that of its parent
compound, JV-1–38 (53% vs. 73%). JV-1–53, the most
potent antagonist of VPAC1-R and
VPAC2-R and devoid of measurable antagonistic
activity on GHRH-R, caused only a marginal, statistically insignificant
inhibition (P > 0.05). The selective
VPAC1-R antagonist PG 97–269 also had a weak
antiproliferative effect. The specific GHRH antagonists JV-1–38,
JV-1–36, and JV-1–42 caused the strongest inhibition of total PSA
levels (69%, 45%, and 46%, respectively; P < 0.001;
Fig. 1B). The nonspecific VPAC-R/GHRH-R antagonist JV-1–52 also
markedly reduced total PSA secretion. These changes in total PSA
secretion from LNCaP cells after exposure to various antagonists
correlated well with the alteration in cell number, indicating that the
rate of PSA secretion was not reduced (Fig. 1C).
|
). Among their derivatives with
decreased GHRH-R antagonistic potency and increased VPAC-R antagonistic
activity, only JV-1–52 inhibited the proliferation of this
VPAC-R-negative cell line (55% inhibition; P <
0.001). VPAC1-R and VPAC2-R
antagonist JV-1–53, devoid of GHRH-R antagonistic activity, caused
only a marginal, statistically insignificant inhibition
(P > 0.05), and the selective
VPAC1-R antagonist PG 97–269 proved ineffective
(Fig. 2). VIP and GHRH also did not influence the proliferation of
MiaPaCa-2 pancreatic cancer cells.
|
). Using primers for human
GHRH-R mRNA, we could not find the expression of GHRH-R (387 bp) in
either LNCaP or MiaPaCa-2 cells. In contrast, the amplification with
specific primers for human VPAC1-R produced a
large amount of a single product of 324 bp in LNCaP cells. Using
specific primers for human VPAC2-R, only a low
expression of VPAC2-R mRNA (586 bp) could be
detected in LNCaP cells. MiaPaCa-2 pancreatic cancer cells proved to be
negative for both VPAC1-R and
VPAC2-R mRNA.
|
). We could not identify VPAC-R on
membranes of MiaPaCa-2 pancreatic cancer cells. Similarly, using
[125I]His1,Nle27-hGHRH-(1–32)NH2
as radioligand, no binding sites for GHRH were detected on either the
LNCaP prostatic or MiaPaCa-2 pancreatic cancer cells examined.
|
). The binding of radioligand was
completely displaced by increasing concentrations
(10-12–10-6
M) of unlabeled VIP (IC50 = 1.07
nM; Table 2). The selective
VPAC-R antagonist JV-1–53 had the highest binding affinity to VPAC-R
on prostate cancer cells, even slightly higher than that of VIP itself
(Fig. 5). Other peptides (JV-1–50, JV-1–51, and JV-1–52), also more
selective for VPAC-R, displayed lower receptor binding affinity than
VIP, but this affinity was still 1 order of magnitude higher than that
of GHRH antagonists JV-1–36, JV-1–38, and JV-1–42 (Fig. 5). The
selective VPAC1-R antagonist (PG 97–269) also
showed a very strong binding affinity to VPAC-R (Table 2). GHRH
antagonists containing D-Arg in position 2 (JV-1–36,
JV-1–38, and JV-1–42) displayed the lowest binding affinity to VPAC-R
on LNCaP cell membranes (Fig. 5). In our study
hGHRH-(1–29)NH2 had an almost negligible binding
affinity for VPAC-R.
|
|
). PSA release reached the maximum value
in 3–9 min and then rapidly returned to basal values (337.2 ±
17.8 pg/ml) The areas of the peaks of PSA responses caused by VIP
stimulation gradually declined during the experiment (after first VIP,
2356.5 ± 49.5 pg; after third VIP, 1985.1 ± 54.6 pg). GHRH
infused at 10 nM for 12 min at 60-min intervals was also
able to evoke PSA release from LNCaP prostatic cancer cells, with
similar kinetics, but its effects were approximately 25-fold smaller.
In the inhibitory tests, the cells were preincubated with 100 or 300
nM antagonistic peptides for 12 min and then immediately
exposed to a mixture of the analogs and 1 nM VIP or 10
nM GHRH for an additional 12 min (Fig. 6, A and B). The
results obtained from superfusion system indicate that the order of
inhibitory potencies of these analogs on PSA release was similar to
their VPAC-R binding affinities: JV-1–53
PG
97–269>JV-1–51>JV-1–52JV-1–50>>JV-1–36JV-1–38JV-1–42
(Table 2
). The analogs with D-Phe2
substitution, which had been designed to possess enhanced VIP
antagonistic characteristics (JV-1–50, JV-1–51, JV-1–52, and
JV-1–53), were able to block more powerfully the VIP- or GHRH-induced
PSA release than D-Arg2-containing
peptides, JV-1–36, JV-1–38, and JV-1–42. JV-1–53 (Fig. 6B),
JV-1–51, and the selective VPAC1-R antagonist
(PG 97–269) at 300 nM concentrations completely blocked
VIP- or GHRH-stimulated PSA response, whereas JV-1–50 and JV-1–52
inhibited VIP- or GHRH-evoked PSA release to a lesser extent. In
contrast, GHRH antagonists JV-1–36, JV-1–42, and JV-1–38 (Fig. 6A)
proved to be practically ineffective at concentration tested. None of
the GHRH-R and VPAC-R antagonists tested was able to influence basal
PSA secretion.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Consequently, in this study we also characterized VPAC1-R, VPAC2-R, and GHRH-R on LNCaP human prostatic cancer cells, which could be involved in the antiproliferative effect of the antagonists. Our findings, based on RT-PCR, radioligand competition assay, and superfusion system, clearly indicate that LNCaP cells have functional VPAC-R and are devoid of classic pituitary-type GHRH-R. The expression of mRNA for both types of VPAC-R was detected in LNCaP prostatic cancer cells, with VPAC1-R predominating in accordance with recent observations of rat prostate membranes (50). We also demonstrated in a superfusion system that a pulsatile stimulation of LNCaP cells with GHRH leads to a transient increase in the rate of PSA secretion, however, to a 25-fold lesser extent than with VIP. Because the release of this marker is not related to cell proliferation in this dynamic system, the superfusion method allowed characterization of the receptor-mediated, short-term endocrine effects of GHRH analogs. Antagonistic analogs of GHRH, JV-1–36, JV-1–38, and JV-1–42, inhibited VIP- or GHRH-evoked PSA release much less effectively than the selective and potent VPAC-R antagonists JV-1–53 and PG 97–269. These GHRH antagonists were also able to bind to VPAC-R on LNCaP cells, however, with a lesser affinity than VPAC-R antagonists. These findings suggest that GHRH and its antagonists may influence inducible PSA release through tumoral VPAC-R by acting as pharmacological analogs of VIP with lower affinity. A similar rank order of potency for these GHRH analogs was recently observed by us in tests on pinealocytes (blockade of VIP-evoked cAMP efflux) (30), where VPAC1-R were found to be predominant (51). These results indicate that the same receptor subtype may play a role in the induction of PSA secretion from LNCaP cells by both VIP and GHRH. A joint use of superfusion method and ligand binding assay proved to be suitable for screening GHRH analogs for their VPAC-R inhibitory activity on LNCaP cells.
In contrast, the antiproliferative effects of analogs were not correlated with their VPAC-R antagonistic potencies. Instead, a clear association was observed between tumor inhibition and antagonistic activity of analogs on GHRH-R, suggesting that VPAC-R are not the main receptors that mediate the antiproliferative effect of GHRH antagonists.
Although the potent GHRH antagonists JV-1–36, JV-1–38, and JV-1–42 have more than 1000-fold higher affinity for GHRH-R than for VPAC1-R and VPAC2-R on rat pituitary and pineal cells (8, 30), their antagonistic activity on VPAC-R is not negligible. These GHRH antagonists displaced radiolabeled VIP from VPAC-R on LNCaP cell membranes with IC50 values of 27–82 nM. This affinity is higher than that of a previously reported VPAC-R antagonist, VIPhyb, (IC50 = 500–700 nM), which inhibited the growth of nonsmall cell lung cancers and breast cancers in vitro and in vivo (26, 27). Despite this relatively high VPAC-R binding affinity of analogs JV-1–36, JV-1–38, and JV-1–42, their antagonistic properties on VPAC-R appear to be less involved in the antiproliferative mechanism than their GHRH-R inhibitory potency. This is supported by the findings that D-Phe2-containing analogs JV-1–50, JV-1–51, and JV-1–52, with decreased GHRH-R antagonistic potency and increased VPAC-R antagonistic activity, caused less inhibition of cell proliferation than their parent compounds. A possible nonspecific inhibitory effect of these analogs due to their high concentration (10 µM) can be also excluded, because GHRH, VIP, and a potent GHRH agonistic analog (JI-38) (52), structurally similar to the antagonists, either did not inhibit or slightly stimulated the proliferation of LNCaP and MiaPaCa-2 cells (data not shown).
These results provide, for the first time, evidence that the antiproliferative effects of GHRH antagonists in vitro are mainly exerted through a VPAC-R-independent mechanism. This conclusion is supported by the ability of GHRH antagonists to inhibit proliferation of VPAC-R-negative human pancreatic cancer cell line (MiaPaCa-2).
Therefore, it is likely that unknown receptors, which are different from the well known VPAC-R and the classic pituitary GHRH-R, might be involved in the antiproliferative mechanism of our GHRH antagonists. Using primers for human GHRH-R mRNA, a single band of a 387-bp product was amplified in the human pituitary cells, but no GHRH-R mRNA expression was found in LNCaP and MiaPaCa-2 human cancer cells, in accordance with an earlier report on ovarian tumors (53). The presence of splicing variants of GHRH-R was detected in human pituitary adenomas (53, 54), and thus, in extrapituitary tumor cells other GHRH-R variants or structurally related receptors may also exist, the cDNA of which was not recognized by our primers. The radiolabeled [His1,Nle27]human GHRH-(1–32)NH2, previously used for the characterization of pituitary GHRH-R (46, 55) might have very low affinity to these variant forms of receptors. Consequently, we could not find specific binding sites for this ligand on cancer cells. In contrast, some of the GHRH analogs with structures such as JV-1–36, JV-1–38, and JV-1–42 might bind to these variant forms of receptors with higher affinity and thus inhibit tumor proliferation. The identification of these unknown receptors involved in the antiproliferative action of our GHRH antagonists is in progress. In conclusion, although much additional work is required, the studies in vitro described herein and other findings suggest that potent antitumor effects of GHRH antagonists (5) are exerted by mechanisms independent of receptors for VIP.
|
Acknowledgments |
|---|
|
Footnotes |
|---|
2 On leave from Department of Human Anatomy, University
Medical School, H-7643 Pécs, Hungary.
Received December 20, 1999.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
This article has been cited by other articles:
 |
|
 |
|
  F. F. Casanueva, J. P. Camina, M. C. Carreira, Y. Pazos, J. L. Varga, and A. V. Schally Growth hormone-releasing hormone as an agonist of the ghrelin receptor GHS-R1a PNAS, December 23, 2008; 105(51): 20452 - 20457. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Xie, D. W. Wolff, M.-F. Lin, and Y. Tu Vasoactive Intestinal Peptide Transactivates the Androgen Receptor through a Protein Kinase A-Dependent Extracellular Signal-Regulated Kinase Pathway in Prostate Cancer LNCaP Cells Mol. Pharmacol., July 1, 2007; 72(1): 73 - 85. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Buchholz, A. V. Schally, J. B. Engel, F. Hohla, E. Heinrich, F. Koester, J. L. Varga, and G. Halmos Potentiation of mammary cancer inhibition by combination of antagonists of growth hormone-releasing hormone with docetaxel PNAS, February 6, 2007; 104(6): 1943 - 1946. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Havt, A. V. Schally, G. Halmos, J. L. Varga, G. L. Toller, J. E. Horvath, K. Szepeshazi, F. Koster, K. Kovitz, K. Groot, et al. The expression of the pituitary growth hormone-releasing hormone receptor and its splice variants in normal and neoplastic human tissues PNAS, November 29, 2005; 102(48): 17424 - 17429. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Keller, A. V. Schally, K. Groot, G. L. Toller, A. Havt, F. Koster, P. Armatis, G. Halmos, M. Zarandi, J. L. Varga, et al. Effective treatment of experimental human non-Hodgkin's lymphomas with antagonists of growth hormone-releasing hormone PNAS, July 26, 2005; 102(30): 10628 - 10633. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. B. Engel, G. Keller, A. V. Schally, G. L. Toller, K. Groot, A. Havt, P. Armatis, M. Zarandi, J. L. Varga, and G. Halmos Inhibition of Growth of Experimental Human Endometrial Cancer by an Antagonist of Growth Hormone-Releasing Hormone J. Clin. Endocrinol. Metab., June 1, 2005; 90(6): 3614 - 3621. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Z. Rekasi, T. Czompoly, A. V. Schally, F. Boldizsar, J. L. Varga, M. Zarandi, T. Berki, R. A. Horvath, and P. Nemeth Antagonist of growth hormone-releasing hormone induces apoptosis in LNCaP human prostate cancer cells through a Ca2+-dependent pathway PNAS, March 1, 2005; 102(9): 3435 - 3440. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. L. Toller, J. E. Horvath, A. V. Schally, G. Halmos, J. L. Varga, K. Groot, D. Chism, and M. Zarandi Development of a polyclonal antiserum for the detection of the isoforms of the receptors for human growth hormone-releasing hormone on tumors PNAS, October 19, 2004; 101(42): 15160 - 15165. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Z. Szereday, A. V. Schally, J. L. Varga, C. A. Kanashiro, F. Hebert, P. Armatis, K. Groot, K. Szepeshazi, G. Halmos, and R. Busto Antagonists of Growth Hormone-Releasing Hormone Inhibit the Proliferation of Experimental Non-Small Cell Lung Carcinoma Cancer Res., November 15, 2003; 63(22): 7913 - 7919. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Halmos, A. V. Schally, T. Czompoly, M. Krupa, J. L. Varga, and Z. Rekasi Expression of Growth Hormone-Releasing Hormone and Its Receptor Splice Variants in Human Prostate Cancer J. Clin. Endocrinol. Metab., October 1, 2002; 87(10): 4707 - 4714. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Busto, A. V. Schally, J. L. Varga, M. O. Garcia-Fernandez, K. Groot, P. Armatis, and K. Szepeshazi The expression of growth hormone-releasing hormone (GHRH) and splice variants of its receptor in human gastroenteropancreatic carcinomas PNAS, September 3, 2002; 99(18): 11866 - 11871. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Halmos, A. V. Schally, J. L. Varga, A. Plonowski, Z. Rekasi, and T. Czompoly Human renal cell carcinoma expresses distinct binding sites for growth hormone-releasing hormone PNAS, September 12, 2000; 97(19): 10555 - 10560. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Z. Rekasi, T. Czompoly, A. V. Schally, and G. Halmos Isolation and sequencing of cDNAs for splice variants of growth hormone-releasing hormone receptors from human cancers PNAS, September 12, 2000; 97(19): 10561 - 10566. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Endocrinology | Endocrine Reviews | J. Clin. End. & Metab. |
| Molecular Endocrinology | Recent Prog. Horm. Res. | All Endocrine Journals |
), human
GHRH-(1–29)NH2 (
), JV-1–36 (
), JV-1–38 (•),
JV-1–42 (
), JV-1–50 (
), JV-1–51 (
), JV-1–52 (
),
JV-1–53 (
), and PG 97–269 (
) were calculated using a
computerized curve-fitting program (