Impact of SLC20A1 on the Wnt/ß­catenin signaling pathway in somatotroph adenomas.

Studies have revealed that genetic and functional aberrations of oncogenes, tumor­suppressor genes, signaling pathways and receptors are among the most prominent events in pituitary tumorigenesis, and a potent biomarker would be helpful for early diagnosis, subsequent treatment and disease control. The present study investigated the expression signatures of solute carrier family 20 member 1, also known as phosphate transporter 1 (SLC20A1) and the Wnt/ß­catenin signaling pathway in 52 patients with somatotroph adenomas. According to immunohistochemistry analysis, the H­score of SLC20A1 was 222.6±15.2 in invasive tumor samples and 144.5±30.4 in non­invasive tumor samples (P<0.01), while the H­scores of ß­catenin were 210.1±21.4 and 134.9±32.7, respectively (P<0.05). The H­scores of Wnt inhibitory factor 1 (Wif1) exhibited the opposite trend, with scores of 134.5±22.7 and 253.6±14.8, respectively (P<0.01). The H­scores of SLC20A1 were negatively associated with those of Wif1 in somatotroph adenomas (correlation coefficient r=­0.367). The mean progression­free survival in the low SLC20A1 group was longer than that in the group with high SLC20A1 H­scores (P=0.024). Reverse transcription­quantitative PCR (RT­qPCR) and western blotting confirmed the interference efficiency of the segments short hairpin (Sh)­B­SLC20A1 and Sh­C­SLC20A1. Cell proliferation experiments revealed that the cell viability of the Sh­B­SLC20A1 group was 76.3±4.5, 65.7±3.7 and 53.1±3.2% of that of control GH3 cells after 24, 48 and 72 h of transfection, respectively, while the cell viability of the Sh­C­SLC20A1 group was 86.4±5.7, 75.6±4.4 and 67.5±3.8%, respectively (P<0.05). ELISA analysis demonstrated the growth hormone (GH) levels in the Sh­B­SLC20A1 and Sh­C­SLC20A1 groups to be 34.7±10.4 and 54.6±14.4%, respectively, of that of control GH3 cells (P<0.05). The transmembrane invasion assay revealed that knocking down SLC20A1 significantly suppressed cell invasion in the Sh­B­SLC20A1 and Sh­C­SLC20A1 groups. RT­qPCR and western blotting demonstrated that Sh­B­SLC20A1 and Sh­C­SLC20A1 evidently increased the levels of Wif1 and secreted frizzled­related protein 4. The present data suggested that SLC20A1 levels are positively associated with tumor size, invasive behavior and tumor recurrence in somatotroph adenomas. Furthermore, SLC20A1 may be associated with the activation of the Wnt/ß­catenin signaling pathway.

Expression of Cyclin E/Cdk2/p27Kip1 in Growth Hormone Adenomas.

OBJECTIVE: The present study was conducted to evaluate the levels of Cdk2, cyclin E, p21, and p27 in growth hormone adenomas (GHPAs) and analyze their association with clinicopathologic features. METHODS: We collected 46 GHPA specimens and clinical materials from March 2012 to December 2015 at Beijing Tiantan Hospital. We analyzed the expression of Cdk2, cyclin E, p21, and p27 using immunohistochemistry, quantitative real-time polymerase chain reaction, Western blot, and methylation-specific polymerase chain reaction and sequencing. RESULTS: The levels of cyclin E and Cdk2 were much greater in the invasive group than in the noninvasive group (P < 0.05). Significant differences were found between cyclin E and p21 and tumor size (P < 0.05) and between cyclin E expression and invasion (P < 0.05). Tumors were more likely to require whole resection in patients with low cyclin E expression (P < 0.05). The high-level p27 group had better progression-free survival than did the low-level p27 group (P < 0.01). Quantitative real-time polymerase chain reaction and Western blot analysis revealed a similar trend for Cdk2, cyclin E, p21, and p27 protein levels in growth hormone specimens (P < 0.01). An average of 33 CpG sites per sample were analyzed using matrix-assisted laser desorption ionization time-of-flight mass array, and in 7 of the 33 CpG sites, the methylation levels of p27 were >50%. Statistically significant differences were found in 4 CpG sites between the invasive and noninvasive specimens (P < 0.01). CONCLUSIONS: Overexpression of cyclin E/Cdk2 and loss of p27 appears to be associated with a poor prognosis and might play a role in the treatment of GHPAs in the future.

Stereotactic radiation therapy for the treatment of functional pituitary adenomas associated with feline acromegaly.

BACKGROUND: Conventional fractionated radiotherapy has been shown to be partially effective for management of pituitary tumors in cats that cause acromegaly and diabetes mellitus (DM), but, the efficacy and safety of stereotactic radiation therapy (SRT) as a treatment for acromegalic cats has not been described. HYPOTHESIS: Stereotactic radiation therapy is an effective and safe treatment for controlling acromegaly associated with pituitary adenomas in cats. Additionally, SRT-treated acromegalic cats with DM will experience a decrease in insulin requirements after radiation therapy. ANIMALS: Fifty-three client-owned cats referred to Colorado State University for SRT to treat pituitary tumors causing poorly controlled DM secondary to acromegaly. METHODS: Retrospective study of cats treated for acromegaly with SRT between 2008 and 2016 at Colorado State University. Diagnosis of acromegaly was based on history, physical examination, laboratory results, and cross-sectional imaging of the pituitary. Signalment, radiation protocol, insulin requirements over time, adverse effects, and survival were recorded. RESULTS: Median survival time was 1072 days. Of the 41 cats for which insulin dosage information was available, 95% (39/41) experienced a decrease in required insulin dose, with 32% (13/41) achieving diabetic remission. Remission was permanent in 62% (8/13) and temporary in 38% (5/13) cats. Median duration to lowest insulin dose was 9.5 months. Of the treated cats, 14% developed hypothyroidism and required supplementation after SRT. CONCLUSIONS: Stereotactic radiation therapy is safe and effective for treating cats with acromegaly. Cats treated with SRT have improved survival time and control of their DM when compared to previously reported patients treated with non-SRT.

Decreasing mortality and changes in treatment patterns in patients with acromegaly from a nationwide study.

CONTEXT: New therapeutic strategies have developed for the management of acromegaly over recent decades. Whether this has improved mortality has not been fully elucidated. OBJECTIVE: The primary aim was to investigate mortality in a nationwide unselected cohort of patients with acromegaly. Secondary analyses included time trends in mortality and treatment patterns. DESIGN: A total of 1089 patients with acromegaly were identified in Swedish National Health Registries between 1987 and 2013. To analyse time trends, the cohort was divided into three periods (1987-1995, 1996-2004 and 2005-2013) based on the year of diagnosis. MAIN OUTCOME MEASURES: Using the Swedish population as reference, standardized mortality ratios (SMRs) were calculated with 95% confidence intervals (CIs). RESULTS: Overall SMR was 2.79 (95% CI: 2.43-3.15) with 232 observed and 83 expected deaths. Mortality was mainly related to circulatory diseases (SMR: 2.95, 95% CI: 2.35-3.55), including ischemic heart disease (2.00, 1.35-2.66) and cerebrovascular disease (3.99, 2.42-5.55) and malignancy (1.76, 1.27-2.26). Mortality decreased over time, with an SMR of 3.45 (2.87-4.02) and 1.86 (1.04-2.67) during the first and last time period, respectively (P = .015). During the same time periods, the frequency of pituitary surgery increased from 58% to 72% (P < 0.001) and the prevalence of hypopituitarism decreased from 41% to 23% (P < 0.001). CONCLUSIONS: Excess mortality was found in this nationwide cohort of patients with acromegaly, mainly related to circulatory and malignant diseases. Although still high, mortality significantly declined over time. This could be explained by the more frequent use of pituitary surgery, decreased prevalence of hypopituitarism and the availability of new medical treatment options.

Assessment of sFRP4 as a bio-marker for predicting aggressiveness and recurrence of growth hormone-secreting pituitary adenomas.

The association of sFRP4 expression with aggressiveness and recurrence of growth hormone (GH)-secreting pituitary adenomas was investigated. Ten normal pituitary and 52 GH-secreting pituitary adenoma specimens were classified into three groups: normal pituitary (control) group, non-aggressive group, and aggressive group, according to preoperative evaluation by magnetic resonance imaging (MRI)/computed tomography (CT). Expression of sFRP4 was determined by quantitative real-time polymerase chain reaction (qRT-PCR), western blot analysis, and tissue microarrays, to assess the association between sFRP4 and aggressiveness. Follow-up information of all 52 patients was collected to evaluate the impact of sFRP4 expression on the recurrence/progression of GH-secreting pituitary adenomas. qRT-PCR results showed a lower level of sFRP4 mRNA in the aggressive group, as compared to that in the non-aggressive group (P=0.001). A similar trend was observed on western blot analysis for sFRP4 protein expression (P=0.004). On analysis by tissue microarrays, weak sFRP4 expression was detected in the aggressive group (10/15, 66.7%). Univariate analysis showed a significant relationship between low sFRP4 expression and aggressiveness (P=0.024). On multivariate analysis weak sFRP4 expression was found to be an independent factor of recurrence/progression (odds ratio: 0.063, P=0.026). Methylation of the sFRP4 promoter was increased in low sFRP4 staining group compared to that in the high sFRP4 staining group (P<0.001). In this study, weak sFRP4 expression appeared to predict aggressive behavior, and was associated with recurrence/progression of GH-secreting pituitary adenomas. Methylation of the sFRP4 promoter may account for the low sFRP4 expression.

Preoperative somatostatin analogues versus direct transsphenoidal surgery for newly-diagnosed acromegaly patients: a systematic review and meta-analysis using the GRADE system.

UNLABELLED: Whether the preoperative use of somatostatin analogues (SA) improves surgical outcomes in acromegaly is still a matter of debate. OBJECTIVE: We conducted a systematic review of randomized, controlled trials that compared the short-term outcomes of preoperative use of SA (Pre-SA) with direct TSS (No-SA) for the treatment of newly diagnosed acromegaly. METHODS: Embase, Pubmed, Lilacs, and Central Cochrane were used as our data sources. The primary outcomes were no need for any adjuvant treatment 3 months after surgery, based on biochemical results (GH nadir after OGTT <1 µg/L and normal IGF-1 for age and gender), quality of life and mortality. The included trials were analyzed using the Grading of Recommendations Assessment, Development, and Evaluation approach. RESULTS: A total of 2.099 references were identified and two reviewers independently screened the titles and abstracts. From the 14 potentially eligible studies, four were included and ten were excluded due to lack of randomization or different outcomes. A pool of 261 patients was randomly assigned to Pre-SA or No-SA. Meta-analysis of IGF1 normalization showed a significant difference in favor of Pre-SA (RR 2.47; 95% CI 1.66, 3.77). Adding a GH nadir on OGTT ≤1 µg/L, we found a RR of 2.15 (95% CI 1.39, 3.33). Quality of evidence for no need of adjuvant postoperative treatment was moderate, but for improving quality of life was very low and for mortality was absent. CONCLUSION: Pre-SA increases the chance of biochemical control of acromegaly 3 months after TSS in patients harboring GH-secreting pituitary macroadenomas.

Glucocorticoid replacement and mortality in patients with nonfunctioning pituitary adenoma.

CONTEXT: Current treatment guidelines generally suggest using lower and weight-adjusted glucocorticoid replacement doses in patients with insufficiency of the hypothalamic-pituitary-adrenal (HPA) axis. Although data in patients with acromegaly revealed a positive association between glucocorticoid dose and mortality, no comparable results exist in patients with nonfunctioning pituitary adenomas (NFPA). OBJECTIVE: Our objective was to assess whether higher glucocorticoid replacement doses are associated with increased mortality in patients with NFPA and HPA axis insufficiency. DESIGN, PARTICIPANTS, AND INTERVENTION: We included 105 patients receiving glucocorticoid replacement after pituitary surgery due to NFPA and concomitant HPA axis insufficiency. Patients were grouped according weight-adapted and absolute hydrocortisone (HC) replacement doses. Mortality was assessed using Kaplan-Meier methodology as well as multivariable Cox regression models. SETTING: This was a retrospective analysis conducted at a tertiary referral center. MAIN OUTCOME: We evaluated overall mortality based on HC replacement doses. RESULTS: Average age at inclusion was 58.9±14.8 yr, and mean follow-up was 12.7±9.4 yr. The groups did not differ according to age, follow-up time, pattern of hypopituitarism, and comorbidities. Kaplan-Meier survival probabilities differed significantly when comparing individuals with differing weight-adjusted HC dose (P=0.001) as well as absolute HC dose (5-19, 20-29, and ≥30 mg, P=0.009). Hazard ratios for mortality increased from 1 (0.05-0.24 mg/kg) to 2.62 (0.25-0.34 mg/kg) to 4.56 (≥0.35 mg/kg, P for trend=0.006) and from 1 (5-19 mg) to 2.03 (20-29 mg) to 4 (≥30 mg, P for trend=0.029), respectively. CONCLUSION: Higher glucocorticoid replacement doses are associated with increased overall mortality in patients with NFPA and insufficiency of HPA axis. This further substantiates the importance of a balanced and adjusted glucocorticoid replacement therapy in these patients.

Prognostic significance of the Ki-67 labeling index in growth hormone-secreting pituitary adenomas.

CONTEXT: Ki-67 is a marker of proliferation activity associated with invasiveness and prognosis in human tumors. OBJECTIVE: The aim of the study was to evaluate the Ki-67 index prognostic relevance in a group of acromegalic patients who underwent transsphenoidal surgery for a GH-secreting pituitary adenoma. MATERIAL AND METHODS: We selected 68 consecutive acromegalic patients referred to our hospital during a 5-yr period. The Ki-67 index was determined by immunohistochemistry on tissue samples obtained from each adenoma after surgery. Those patients who were not completely cured after surgery began medical therapy with somatostatin analogs (SSAs). Periodical pituitary magnetic resonance imaging and hormonal evaluation were performed during the follow-up. RESULTS: Twenty-eight of 68 patients were cured after surgery (41%). Among the 40 patients treated with SSAs, 13 were considered uncontrolled. Pituitary magnetic resonance imaging showed residual/recurrent disease in 25 of 68 patients after 6 months. No correlation was found between Ki-67 index and age, tumor size, GH, or IGF-I plasma levels. Tumors described as having cavernous sinus invasion had a higher mean Ki-67 index as compared with noninvasive tumors (P < 0.01). The Ki-67 index was significantly lower in tumors in patients cured after surgery as compared with patients considered not cured (P < 0.01) and in tumors in patients controlled by SSA therapy as compared with patients considered as uncontrolled (P < 0.05). CONCLUSION: The Ki-67 labeling index may predict clinical outcome in postsurgical management of acromegalic patients. We suggest routine Ki-67 evaluation in GH-secreting pituitary adenomas.