Low levels of raf kinase inhibitory protein in growth hormone-secreting pituitary adenomas correlate with poor response to octreotide treatment.
ABSTRACT Excessive GH production by pituitary tumors causes acromegaly. Medical treatment of acromegaly with somatostatin analogs (SMSs), like octreotide, is well established, but the clinical effect is variable. One mechanism for octreotide effect is inhibition of the MAPK signaling pathway after binding to the G protein-coupled somatostatin receptor. Nonphosphorylated Raf kinase inhibitory protein (RKIP) binds to and inhibits Raf1 kinase, and thereby attenuates MAPK signaling, whereas phosphorylated RKIP inhibits G protein receptor internalization and degradation due to inhibition of G protein receptor kinase 2. Objective: Our objective was to study RKIP levels in pituitary somatotroph adenomas, and relate them to clinical characteristics and response to octreotide treatment in patients with acromegaly.
RKIP level was analyzed by Western blot of proteins extracted from somatotroph tumors frozen a short time after surgery in 51 patients with active acromegaly. An acute somatostatin test was performed in 46 of the patients, and in 21 the IGF-I level before and 6 months after SMS treatment was available.
The adenoma RKIP level correlated significantly to both the acute and the long-term octreotide responses on serum levels of GH and IGF-I, respectively. In multiple regression analyses, the RKIP level was a significant determinant for both the GH reduction in the acute test and the IGF-I reduction after approximately 6 months.
The RKIP level in somatotroph adenomas seems to be important for the clinical effect of SMS treatment, in which low levels of RKIP correlate to poor clinical response to SMSs.
- SourceAvailable from: gla.ac.uk[Show abstract] [Hide abstract]
ABSTRACT: Raf Kinase Inhibitor Protein (RKIP) was originally described as an inhibitor of the Ras-Raf-MEK-ERK pathway, exerting its action by the physical inhibition of the interaction of Raf with MEK. It has subsequently been shown to play important roles in a number of other signalling pathways, including the NFκB pathway and in the stability of the mitotic spindle. Not surprisingly given that it impacts on many important signalling pathways RKIP levels have been shown to be important in the progression of a number of different cancers. RKIP expression is lost or decreased in a number of common human cancers and decreased still further in tumour metastases. One of the tumours in which RKIP is downregulated is colorectal cancer (CRC). Importantly it has been shown that not only is RKIP depleted in tumour tissue when compared with normal tissue but that the level of RKIP within a tumour is inversely correlated with the likelihood of metastatic relapse and with patient prognosis. Although we already have a number of very good prognostic indicators in CRC, one group of patients for whom new prognostic indicators would be useful are patients with Dukes B CRC. These are patients with locally advanced but non-metastatic disease and at present there is no firm consensus on their correct post-operative management. Therefore we set out to examine whether RKIP is a useful prognosticator in this particular group using a tissue microarray (TMA) with samples from over 200 patients with Dukes B CRC. The analysis revealed a strong inverse correlation between RKIP levels and disease specific survival. Moreover, in a multivariate analysis RKIP emerged as an independent prognostic indicator along with lympho-vascular invasion and peritoneal invasion, two well-known and powerful prognosticators. This allowed for the generation of a simple prognostic index, using information from the different independent indicators, allowing for improved patient risk stratification. This led us to examine whether RKIP could also function as a predictive marker in CRC. To do this we again used a TMA, this time consisting of a much larger cohort of patients across the whole range of tumour stages. The results confirmed the prognostic utility of RKIP and indicated that patients whose tumours have low levels of RKIP may derive a greater benefit from chemotherapy than those patients whose tumours have high levels, although this result did not reach statistical significance. In the second part of the thesis I have examined the effect of RKIP in previously characterised mouse models of CRC. To do this I have used a germline RKIP knockout mouse and in the first instance crossed it to the APC580S mouse. In this mouse APC is lost conditionally within the intestine and liver. RKIP knockout did not have any effect on the rate of tumourigenesis or on the invasiveness of tumours in this model. However, in the setting of acute homozygous deletion of APC, RKIP knockout resulted in a decrease in apoptoses in the small intestine and an increase in aberrant mitotic activity in the liver. To follow this up I have examined the effect of RKIP knockout in a mouse model of superficially invasive CRC, specifically to see if RKIP knockout can promote invasive and metastatic behaviour. In this model the APC580S mouse is crossed to mice which conditionally express oncogenic KRas. Although RKIP knockout did not result in an increase in invasive tumours in this model there was a shift in tumour location from the small intestine to the colon. This shift appeared to be due, at least in part to an increase in chromosomal instability in the tumours. The final aim of the thesis was to develop a mouse model of CRC which more closely recapitulates the late stages of the human disease, specifically invasion and metastasis. To do this we have crossed the APC580S mouse with either a conditional p53 knockout or with a mouse that conditionally expresses a point mutation of p53 (p53R172H). In human tumours the majority of abnormalities of p53 are point mutations that result in the production of mutant protein that accumulates in tumour cells. There is evidence that this mutant protein may have oncogenic properties beyond the simple loss of normal p53 protein function. Therefore we have also used this model to study the differing effects of p53 loss and point mutation in CRC. We found that mice homozygous for p53 deletion (p53fl/fl) and those expressing a single copy of the mutant allele with loss of the second copy (p53R172H/fl) developed invasive tumours with nearly 100% penetrance and indeed metastasis was observed. Remarkably, although mice that were heterozygous for p53 deletion (p53fl/+) only rarely developed invasive tumours almost 100% of mice expressing a single copy of the mutant allele (p53R172H/+) developed invasive tumours. We went on to show that the increase in invasion seen in this model is related to an increase in Wnt signalling, which is associated with increased expression of pro-invasive Wnt targets such as fascin. We also showed a novel pro-invasive role for ARF in this process. This is also an excellent model of Dukes B CRC and therefore the ideal model to test the effect of RKIP deletion on invasion and metastasis. These studies led us to examine the differences in effect between knockout and mutant p53 in another tumour model. In this we used a novel model of the aggressive tumour pleomorphic rhabdomyosarcoma to demonstrate that mutant p53 can both promote both tumourigenesis and metastasis more potently than p53 knockout. These studies have demonstrated the value of RKIP in the clinically important Dukes B CRC population and shown its possible utility as a predictive marker in this group. Although we have not seen an effect of RKIP knockout in traditional mouse models of CRC we have developed a novel model which closely recapitulates Dukes B CRC and may be useful in elucidating the effect of RKIP knockout. We have also used this model to gain novel insights into the invasive process, in particular into the role played by mutant p53.
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ABSTRACT: Loss of E-cadherin is an important marker of epithelial tumour progression. The aims of this study were to explore whether E-cadherin expression and localization correlate to corticotroph tumour progression, relate the expression of the E-cadherin gene (CDH1) to immunohistochemical E-cadherin staining pattern, and study whether the E-cadherin levels were correlated to methylation status of the CDH1 promoter region. Immunohistochemical analyses of E-cadherin protein were performed, as was RT-qPCR of the CDH1 and the POMC genes. Methylation pattern of the promoter region of CDH1 was measured using pyrosequencing of bisulfite-treated DNA. Forty-five patients operated at a tertiary referral centre in Oslo, Norway. Adenoma tissue sections and RNA samples from patients with verified Cushing's disease or Nelson's syndrome were collected. Expression of E-cadherin mRNA and protein in pituitary corticotroph adenomas and average percentage of methylated cytosines in a cytosine-phosphate-guanosine island of the CDH1 promoter. Correlations were observed between tumour progression and both nuclear expression of E-cadherin and reduced CDH1 mRNA. The E-cadherin expression was not determined by the methylation pattern of the CDH1 promoter. Corticotroph tumour progression was associated with reduced expression of the epithelial marker E-cadherin.Clinical Endocrinology 05/2011; 75(6):811-8. · 3.40 Impact Factor
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ABSTRACT: Recently, correlations between corticotroph tumor dedifferentiation and both E-cadherin immunostaining and reduced mRNA expression of the E-cadherin gene (CDH1) have been demonstrated. The purpose of this study was to explore whether tumor dedifferentiation correlated with glucocorticoid resistance and whether the resistance was associated with both positively and negatively regulated genes. Tumor material from 20 patients with verified Cushing's disease or Nelson's syndrome operated on at Rikshospitalet, Oslo. Reverse transcription polymerase chain reaction analysis of genes such as E-cadherin (CDH1), proopiomelanocortin (POMC), glucocorticoid-induced leucine zipper (GILZ), and thioredoxin-interacting protein (TXNIP) was performed. The correlations between the expression of the GILZ, TXNIP, and POMC genes in different stages of corticotroph adenomas, the E-cadherin mRNA expression and staining pattern, and the preoperative 24-h cortisol excretion were examined. The GILZ and TXNIP expression levels were positively correlated to the CDH1 expression and were highest in microadenomas and in tumors with a high membranous E-cadherin reactivity. In contrast, the POMC expression was not significantly different between the groups. This divergence between the genes that were positively and negatively regulated by glucocorticoids could not be supported by other gene expression analyses. No correlations to urinary cortisol were found. The expression of the glucocorticoid-responsive genes POMC, GILZ, and TXNIP in corticotroph adenomas showed a remarkable variation. The pattern and variability of glucocorticoid resistance in corticotroph adenomas seem to correlate with a loss of the epithelial phenotype associated with corticotroph tumor dedifferentiation.Endocrine 01/2013; · 1.42 Impact Factor
Low Levels of Raf Kinase Inhibitory Protein in Growth
Hormone-Secreting Pituitary Adenomas Correlate
with Poor Response to Octreotide Treatment
Stine Lyngvi Fougner, Jens Bollerslev, Fahim Latif, John K. Hald, Terje Lund, Jon Ramm-Pettersen,
and Jens Petter Berg
Research Institute for Internal Medicine (S.L.F.), University of Oslo, N-0027 Oslo, Norway; Hormone Laboratory, Aker University Hospital
(F.L., T.L., J.P.B.), University of Oslo, N-0027 Oslo, Norway; and Section of Endocrinology (S.L.F., J.B.), and Departments of Radiology
(J.K.H.) and Neurosurgery (J.R.-P.), Rikshospitalet Medical Centre, N-0027 Oslo, Norway
Context: Excessive GH production by pituitary tumors causes acromegaly. Medical treatment of
acromegaly with somatostatin analogs (SMSs), like octreotide, is well established, but the clinical
after binding to the G protein-coupled somatostatin receptor. Nonphosphorylated Raf kinase
whereas phosphorylated RKIP inhibits G protein receptor internalization and degradation due to
inhibition of G protein receptor kinase 2.
Objective: Our objective was to study RKIP levels in pituitary somatotroph adenomas, and relate
them to clinical characteristics and response to octreotide treatment in patients with acromegaly.
Patients and Methods: RKIP level was analyzed by Western blot of proteins extracted from so-
after SMS treatment was available.
Results: The adenoma RKIP level correlated significantly to both the acute and the long-term
octreotide responses on serum levels of GH and IGF-I, respectively. In multiple regression analyses,
the RKIP level was a significant determinant for both the GH reduction in the acute test and the
IGF-I reduction after approximately 6 months.
Conclusion: The RKIP level in somatotroph adenomas seems to be important for the clinical effect
of SMS treatment, in which low levels of RKIP correlate to poor clinical response to SMSs. (J Clin
Endocrinol Metab 93: 1211–1216, 2008)
GH is secreted from a somatotroph pituitary adenoma. The pri-
mary treatment is transsphenoidal surgery, which in a recent
large study, has been reported to give a total cure rate of 28–
39%, depending on cure definition (2). Therefore, a relatively
large proportion of patients are in need of additional treatment.
cromegaly is a clinical syndrome caused by excessive GH
able for clinical use, octreotide and lanreotide. Both bind with
high affinity to the somatostatin receptor (SSTR) subtype 2
(SSTR2), less affinity to subtype 5, and with low affinity to re-
ceptor subtype 3. Reduced tumor expression of SSTR2 is sug-
gested to explain the lack of response to SMSs in acromegaly.
Studies correlating SSTR2 mRNA levels to octreotide efficacy
have had contradictory results, but most have shown a correla-
tion between receptor mRNA level and octreotide response (3–
Printed in U.S.A.
Copyright © 2008 by The Endocrine Society
doi: 10.1210/jc.2007-2272 Received October 9, 2007. Accepted January 18, 2008.
First Published Online January 29, 2008
Abbreviations: GRK2, G protein-coupled receptor kinase 2; MRI, magnetic resonance im-
and posterior; SMS, somatostatin analog; SSTR2, somatostatin receptor subtype 2.
O R I G I N A LA R T I C L E
E n d o c r i n eC a r e
J Clin Endocrinol Metab, April 2008, 93(4):1211–1216jcem.endojournals.org
8). Recently, we have demonstrated a better octreotide response
in patients with adenomas containing a large proportion of im-
patients exhibited poor response despite a high proportion of
SSTR2a-positive adenoma cells and vice versa.
served and widely expressed protein. It inhibits the MAPK sig-
naling pathway Ras/Raf-1/MEK/ERK by inhibition of Raf-1
phosphorylation and activation (10–12). This intracellular sig-
C phosphorylates RKIP, which then dissociates from Raf-1, and
binds to and inhibits G protein-coupled receptor kinase 2
(GRK2). This inhibition leads to decreased internalization and
degradation of G protein-coupled receptors, and, therefore, a
the MAPK pathway (14). Internalization of the G protein-cou-
pled receptor SSTR2a is shown to be dependent of GRK2-me-
diated phosphorylation of the receptor (15). Moreover, RKIP
way (16). Low levels of RKIP have been associated with an in-
creased propensity of tumors to metastasize and were the stron-
gest predictor for reduced survival in colorectal cancer (17–19).
Loss of RKIP has also been linked to resistance to chemothera-
peutic drugs in prostate and breast cancer cell lines (20).
The purpose of this study was to examine the RKIP levels in
somatotroph pituitary adenomas, and relate this to tumor size
and invasiveness, hormone levels, and to the clinical response to
Subjects and Methods
A total of 51 patients with active acromegaly were included in the
tomy at Rikshospitalet Medical Centre in the period 1996–2006. The
by elevated serum IGF-I compared with the normal reference range of
their age and failure to suppress GH less than 2.5 mU/liter after a 75-g
oral glucose load. In all patients, a pituitary tumor was visualized on a
magnetic resonance imaging (MRI) scan.
One patient had previously undergone transsphenoidal surgery.
There were 15 patients who received treatment with octreotide before
octreotide treatment in ongoing clinical trials. One patient was treated
preoperatively with pegvisomant. None had received radiation therapy
before surgery. Table 1 provides an overview of the study population.
The study was approved by the local ethical committee and con-
ducted according to the Declaration of Helsinki II. Written informed
consent was obtained from all patients.
In 26 patients additional serum was stored at ?80 C until analyzed.
Serum IGF-I was measured by RIA (Nichols Institute, Nijmegen, The
Netherlands) except in the four last included patients (Immulite; Diag-
nostic Products Corp., Los Angeles, CA). The same method was used
when IGF-I levels were compared before and after SMS treatment. Over
methods) were used. At every change of method, cross-calibration was
performed and, if necessary, factorial adjustments introduced. In most
patients, an additional three to seven samples for measurement of GH
were drawn during daytime (0800–1500 h) for calculation of mean GH
level. The acute somatostatin test was performed in 46 of the patients
before medical treatment with a SMS. During the test, one to three mea-
surements of basal GH levels were performed, and two to three analyses
between 2 and 4 h after sc injection of octreotide. The percent reduction
of GH was calculated and nadir GH recorded for each patient.
In addition, serum IGF-I was measured in the stored samples in one
run to minimize between-assay variability and was compared with pre-
used for statistical analysis.
The MRI scan was available for renewed analysis in 46 patients, and
be reevaluated. The measurements were performed by two investigators
the coronal view before and after contrast injection was used for the
used for the measurement of maximal vertical and anteroposterior di-
ameter. For each tumor, the suprasellar, infrasellar, parasellar, anterior,
and posterior (SIPAP) grading score was determined (21). The formula
width ? height ? length ? 0.5 was used to estimate tumor volume (22).
used: noninvasive, maximum SIPAP 1 in the suprasellar extension; bor-
imum 1 in the parasellar extension; and invasive, all adenomas that did
not classify under the specifications for noninvasive or borderline
Shortly after pituitary surgery, adenoma tissue was frozen at ?70 C.
The tissue was homogenized in TRIZOL Reagent (Invitrogen Corp.,
Carlsbad, CA), and protein was extracted following the manufacturer’s
instructions, then precipitated and washed as described in detail previ-
binding procedure using Ultrospec 3300 pro spectrophotometer (Am-
ersham Biosciences, Buckinghamshire, UK).
4–12% BisTris gel (Invitrogen) and blotted onto a polyvinylidene fluo-
ride membrane. Nonfat dry milk in Tris-buffered saline with Tween 20
was used for blocking (5%) and antibody incubation (2.5%). The mem-
was incubated with rabbit polyclonal anti-RKIP antibody (1:2,000; Ab-
cam plc, Cambridge, UK) at 4 C overnight, washed three times, and
Baseline characteristics of the patients
No. of women/men
Tumor volume (cm3)
Serum GH (mU/liter)
Serum IGF-I (nmol/liter)
1.21 (0.59–2.48)1.18 (0.72–4.37)
1.35 (0.91–2.31)1.66 (0.68–2.60)
Unless stated, values are expressed as median (interquartile range).
Fougner et al.Adenoma RKIP Level and Octreotide EfficacyJ Clin Endocrinol Metab, April 2008, 93(4):1211–1216
incubated with the secondary antibody antigoat IgG (1:10,000; Calbio-
chem, San Diego, CA) for 1 h at room temperature. The same protocol
was used for the membrane part containing the largest proteins but with
Louis, MO) and as secondary antibody an antimouse IgG (1:10,000;
incubated with a mouse anti-?-tubulin antibody (1:1000; Sigma-
Aldrich) and with the antimouse IgG as secondary antibody. The
enhanced chemiluminescence Western Blotting Detection Reagents
(Amersham Biosciences) were used for visualization. Protein levels were
adenoma the mean of RKIP/?-actin and RKIP/?-tubulin was designated
the RKIP ratio and used in the analyses.
Most adenomas have previously been examined for SSTR2a protein
Laboratories, Schwabhausen, Germany), standardized against the ?-ac-
tin signal. This procedure has been described in detail previously (9).
Semiquantitative scores of SSTR2a immunohistochemical expres-
multiple regression analysis with RKIP in this study. The SSTR2a scores
were based on less than 75% (grade 1 or 2) or more than 75% (grade 3)
of the cells positively stained by immunohistochemistry (9).
Differences between groups were analyzed using the Mann-Whitney
rank sum test and Kruskal-Wallis test when more than two groups were
compared. Relationships between variables were tested by Spearman
correlation analysis. Multiple linear regression analysis was performed
with stepwise addition of the variables that had P values less than 0.1 in
the univariate analyses. P ? 0.05 was considered significant for all tests.
The statistical analyses were performed using SPSS software version 13
(SPSS, Inc., Chicago, IL).
Staining with anti-RKIP antibody gave a single band at ap-
proximately 22 kDa. As described in the Subjects and Methods
section, the mean of RKIP/?-actin and RKIP/?-tubulin was used
RKIP level and acute octreotide response
The RKIP level in the somatotroph adenomas correlated to
both the percent reduction of serum GH concentration during
the acute octreotide test (R ? 0.342; P ? 0.020) and the nadir
GH level during the test (R ? ?0.457; P ? 0.001) (Fig. 2). In a
multiple regression model, both SSTR2a immunohistochemical
expression scores of the adenoma and RKIP levels were signifi-
cant determinants of the acute octreotide response, defined as
The good responders, defined as patients with more than
75% GH reduction in the acute test, had a significantly higher
(P ? 0.004).
RKIP level and the long-term octreotide response
The RKIP level in the tumor correlated to both the percent
reduction in serum IGF-I concentration (R ? 0.674; P ? 0.001)
and the absolute IGF-I reduction (R ? 0.619; P ? 0.003) after
3). Both the RKIP level and the adenoma SSTR2a immunohis-
tochemical grading were significant determinants for the long-
term percent serum IGF-I reduction in the multiple regression
model (R20.722) (Table 2). The good responders, defined as
higher RKIP level (median 1.85) than the bad responders (me-
dian 0.73) (P ? 0.015).
The effect of octreotide on tumor shrinkage did not signifi-
cantly correlate to the level of RKIP in the adenoma (R ? 0.41;
P ? 0.116).
RKIP level and baseline patient characteristics
The RKIP level did not correlate to patient age or baseline
serum GH or IGF-I levels (P ? 0.41, P ? 0.70). No correlation
to preoperative tumor size or invasiveness could be demon-
strated (P ? 0.45, P ? 0.84).
There was no difference in RKIP levels among the patients
preoperatively treated with octreotide compared with those not
pretreated (P ? 0.98).
A Gs? mutation analysis was performed in 47 of the patients
positive. There was no difference in RKIP level between the gsp-
positive and gsp-negative adenomas (P ? 0.344).
RKIP level and adenoma SSTR2a status
There was no difference in RKIP level between tumors with
different immunohistochemical scores for SSTR2a (P ? 0.60),
and no correlation between the RKIP level and the SSTR2a level
measured by Western blot in a previous study (P ? 0.17) (9).
FIG. 1. A representative Western blot analysis of RKIP (lower panel; the band at
?22 kDa) and ?-actin (upper panel; the band at ?42 kDa) from tumors in six
patients. MW, Molecular weight.
J Clin Endocrinol Metab, April 2008, 93(4):1211–1216jcem.endojournals.org
The present study demonstrates a significant correlation be-
tween the adenoma RKIP level and the effect of octreotide in
tumors with low RKIP levels.
ment in acromegaly has been the tumor expression of SSTRs,
mainly the subtype 2 receptor. Generally, a low receptor level
correlates to a poor octreotide response with some exceptions
level, which may not necessarily correlate to the protein level of
the receptor (23, 24). In our recent study of adenoma SSTR2a
protein level, we demonstrated a correlation between octreotide
response and adenoma SSTR2a expression immunohistochemi-
cally, in which adenomas with a high proportion of SSTR2a-
positive cells responded significantly better to octreotide treat-
ern blot analysis of protein extracts from the tumors was found
(9). Nevertheless, exceptions were observed, suggesting that
other mechanisms downstream of the SSTRs may also be in-
volved in determining the clinical effect of SMSs in the soma-
totroph adenomas. In the present study, we found a strong cor-
level in tumor extracts or expressed immunohistochemically. In
the multiple regression models, both SSTR2a immunohisto-
chemical grading and the RKIP level were significant determi-
nants of the octreotide response. Patients with poor octreotide
response despite a high proportion of SSTR2a-positive cells
seemed to have low RKIP levels, and patients with few SSTR2a-
positive cells, but relatively good octreotide response, had high
RKIP levels. This suggests a model in which SSTR2a expression
is a permissive factor for octreotide response in acromegaly but
that RKIP is an independent modulator of the efficacy of SMSs.
Multiple regression analyses
Univariate analysesMultivariate analyses
For % GH reduction in acute octreotide test
SSTR2a to actin ratio
Log RKIP ratio
For % IGF-I reduction after approximately 6 months
SSTR2a to actin ratio
Log RKIP ratio
aSSTR2a grading represents the immunohistochemical grading of the adenomas in two categories: less than 75% of the cells positively stained (grade 1 and 2), and
more than 75% of the cells positively stained by immunohistochemistry (grade 3).
bThe Mann-Whitney rank sum test P value is presented. For the other variables, the Spearman correlations are presented in the univariate analyses.
FIG. 2. Correlation between tumor RKIP level and short-term octreotide response in patients with somatotrophinomas. Each symbol represents the logarithm of the
mean RKIP/?-actin and RKIP/?-tubulin ratios in the tumor, and the individual short-term octreotide response on the percent serum GH reduction (left panel) and the
nadir GH level (right panel) during the acute octreotide test (n ? 46). Correlation between variables was tested with Spearman correlation analysis.
Fougner et al. Adenoma RKIP Level and Octreotide EfficacyJ Clin Endocrinol Metab, April 2008, 93(4):1211–1216
No correlation between RKIP level and tumor shrinkage was
demonstrated, but MRI scan for this analysis was limited to 16
patients. Previous studies have not shown any correlation be-
tween biochemical effect and tumor size reduction during long-
term treatment of acromegaly (25, 26).
to medical treatment. Some prostate and breast cancer cell lines
low levels of RKIP. Increasing the levels by transfecting the cells
with an RKIP expression vector re-sensitized the cells to drug-
induced apoptosis. In drug-sensitive cell lines, down-regulation
of RKIP led to the resistance to the chemotherapeutic drug (20).
non-Hodgkin’s lymphoma cell lines to chemotherapeutic drug-
induced apoptosis (27). Both inhibition of the MEK-ERK path-
way and inhibition of the nuclear factor-?B pathway were sug-
gested as possible mechanisms for the drug-sensitizing effects.
Studies have indicated RKIP as a metastasis suppressor gene
in cancer. Loss of RKIP was associated with metastasis devel-
opment, and increased angiogenesis and vascular invasion were
suggested as possible mechanisms (18). Somatotroph pituitary
tumors rarely metastasize, but growth can be locally invasive
with erosion of surrounding bone. The tumor often expands
outside of the sella turcica at diagnosis, making radical surgery
difficult. In our study adenoma RKIP level did not correlate to
tumor size and invasiveness. This could be due to the general
benign nature of these adenomas.
29). Therefore, a synergistic inhibitory effect on the MEK-ERK
pathway is a possible mechanism for the association between
adenoma RKIP levels and the clinical efficacy of octreotide.
Phosphorylation of RKIP by protein kinase C activates RKIP
binding to and inhibition of GRK2. Therefore, internalization
correlation between total RKIP level and the level of SSTR2a in
We found a significant correlation between the adenoma
RKIP level and the clinical effect of octreotide, both in the acute
test situation and after long-term treatment. The RKIP level was
also a significant determinant of octreotide efficacy in the mul-
tiple regression models, in addition to the SSTR2a protein ex-
pression. However, potentially RKIP by itself might not be cru-
cial but could be related to an associating factor of an unknown
mechanism. Yet, there are several potential mechanisms for
RKIP to be of importance for the SMS effect, e.g. the effect of
RKIP on drug sensitivity of other cell types.
In conclusion, RKIP levels in somatotroph tumors correlated
ment, even in patients who expressed high levels of the main
octreotide receptor SSTR2a. The study suggests that RKIP is an
important modulator of the therapeutic effect of octreotide of
GH producing tumors. Further studies are needed to reveal the
mechanism for this and to evaluate the role of RKIP on SMS
response in other tumors.
We thank Dr. Olivera Casar Borota for the immunohistochemical anal-
yses of somatostatin receptor subtype 2a, and Mrs. Aase-Brith Jensen
and Mrs. Vigdis Enge for assistance with the protein analyses.
Fougner, Research Institute for Internal Medicine, Section of Endocri-
nology, Rikshospitalet Medical Centre, N-0027 Oslo, Norway. E-mail:
Disclosure Summary: S.L.F. has received lecture fees from Novartis
fees from Novartis and Pfizer, and J.P.B. from Pfizer. The other authors
have nothing to disclose.
kanen H, Loyttyniemi E, Ebeling T, Jaatinen P, Laine H, Nuutila P, Salmela P,
FIG. 3. Correlation between tumor RKIP level and the long-term octreotide response in patients with somatotrophinomas. Each symbol represents the logarithm of the
mean RKIP/?-actin and RKIP/?-tubulin ratios in the tumor and the individual change in serum-IGF-I levels presented as percent (left panel) or absolute (right panel)
reduction after approximately 6-month treatment with octreotide (n ? 21). Correlation between variables was tested with Spearman correlation analysis.
J Clin Endocrinol Metab, April 2008, 93(4):1211–1216jcem.endojournals.org
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