Pharmacogenomic analysis in adrenocortical carcinoma reveals genetic features associated with mitotane sensitivity and potential therapeutics.

Background: Adrenocortical carcinoma (ACC) is an aggressive endocrine malignancy with limited therapeutic options. Treating advanced ACC with mitotane, the cornerstone therapy, remains challenging, thus underscoring the significance to predict mitotane response prior to treatment and seek other effective therapeutic strategies. Objective: We aimed to determine the efficacy of mitotane via an in vitro assay using patient-derived ACC cells (PDCs), identify molecular biomarkers associated with mitotane response and preliminarily explore potential agents for ACC. Methods: In vitro mitotane sensitivity testing was performed in 17 PDCs and high-throughput screening against 40 compounds was conducted in 8 PDCs. Genetic features were evaluated in 9 samples using exomic and transcriptomic sequencing. Results: PDCs exhibited variable sensitivity to mitotane treatment. The median cell viability inhibition rate was 48.4% (IQR: 39.3-59.3%) and -1.2% (IQR: -26.4-22.1%) in responders (n=8) and non-responders (n=9), respectively. Median IC50 and AUC were remarkably lower in responders (IC50: 53.4 µM vs 74.7 µM, P<0.0001; AUC: 158.0 vs 213.5, P<0.0001). Genomic analysis revealed CTNNB1 somatic alterations were only found in responders (3/5) while ZNRF3 alterations only in non-responders (3/4). Transcriptomic profiling found pathways associated with lipid metabolism were upregulated in responder tumors whilst CYP27A1 and ABCA1 expression were positively correlated to in vitro mitotane sensitivity. Furthermore, pharmacologic analysis identified that compounds including disulfiram, niclosamide and bortezomib exhibited efficacy against PDCs. Conclusion: ACC PDCs could be useful for testing drug response, drug repurposing and guiding personalized therapies. Our results suggested response to mitotane might be associated with the dependency on lipid metabolism. CYP27A1 and ABCA1 expression could be predictive markers for mitotane response, and disulfiram, niclosamide and bortezomib could be potential therapeutics, both warranting further investigation.

The influence of cortisol co-secretion on clinical characteristics and postoperative outcomes in unilateral primary aldosteronism.

Context: The prevalence of unilateral primary aldosteronism (UPA) with cortisol co-secretion varies geographically. Objective: To investigate the prevalence and clinical characteristics of UPA with cortisol co-secretion in a Chinese population. Design: Retrospective cohort study. Methods: We recruited 580 patients with UPA who underwent cosyntropin stimulation test (CST) after the 1-mg dexamethasone suppression test (DST) and retrospectively analyzed the clinical characteristics and postoperative outcomes of UPA with and without cortisol co-secretion. Results: UPA with cortisol co-secretion (1 mg DST>1.8 ug/dL) was identified in 65 of 580 (11.2%) patients. These patients were characterized by older age, longer duration of hypertension, higher concentration of plasma aldosterone and midnight cortisol, lower adrenocorticotropic hormone (ACTH) and dehydroepiandrosterone sulfate (DHEAS), larger tumor diameter, and more history of diabetes mellitus. Cortisol and aldosterone levels were higher and DHEAS level was lower in UPA with cortisol co-secretion at 0-120 min after CST. Among 342 UPA patients with KCNJ5 gene sequencing and follow-up results, the complete clinical success rate was lower in UPA with cortisol co-secretion (33.3% vs. 56.4%, P<0.05); the complete biochemical success rate and KCNJ5 mutation did not differ between the two groups. Age, tumor size, and ACTH were independent predictors of UPA with cortisol co-secretion. Sex, BMI, duration of hypertension, KCNJ5 mutation, and cortisol co-secretion were independent predictors for complete clinical success in UPA after surgery. Conclusions: UPA with cortisol co-secretion is not uncommon in China, but the clinical features were distinctly different from those without co-secretion. Cortisol co-secretion is an independent risk factor for incomplete clinical success after surgery in UPA.

Cushing Syndrome Is Associated With Gut Microbial Dysbiosis and Cortisol-Degrading Bacteria.

CONTEXT: Cushing syndrome (CS) is a severe endocrine disease characterized by excessive secretion of cortisol with multiple metabolic disorders. While gut microbial dysbiosis plays a vital role in metabolic disorders, the role of gut microbiota in CS remains unclear. OBJECTIVE: The objective of this work is to examine the alteration of gut microbiota in patients with CS. METHODS: We performed shotgun metagenomic sequencing of fecal samples from 78 patients with CS and 78 healthy controls matched for age and body mass index. Furthermore, we verify the cortisol degradation capacity of Ruminococcus gnavus in vitro and identify the potential metabolite by LC-MC/MS. RESULTS: We observed significant differences in microbial composition between CS and controls in both sexes, with CS showing reduced Bacteroidetes (Bacteroides vulgatus) and elevated Firmicutes (Erysipelotrichaceae_bacterium_6_1_45) and Proteobacteria (Enterobacter cloacae). Despite distinct causes of hypercortisolism in ACTH-dependent and ACTH-independent CS, we found no significant differences in metabolic profiles or gut microbiota between the 2 subgroups. Furthermore, we identified a group of gut species, including R. gnavus, that were positively correlated with cortisol levels in CS. These bacteria were found to harbor cortisol-degrading desAB genes and were consistently enriched in CS. Moreover, we demonstrated the efficient capacity of R. gnavus to degrade cortisol to 11-oxygenated androgens in vitro. CONCLUSION: This study provides evidence of gut microbial dysbiosis in patients with CS and identifies a group of CS-enriched bacteria capable of degrading cortisol. These findings highlight the potential role of gut microbiota in regulating host steroid hormone levels, and consequently host health.

Association between contralateral adrenal and hypothalamus-pituitary-adrenal axis in benign adrenocortical tumors.

Context: Adrenal incidentaloma (AI) is commonly discovered on cross-sectional imaging. Mild autonomous cortisol secretion is the most common functional disorder detected in AI. Objective: To delineate the association between radiological characteristics of benign adrenocortical tumors and hypothalamus-pituitary-adrenal (HPA) axis. Methods: In the study, 494 patients diagnosed with benign unilateral adrenocortical tumors were included. Mild autonomous cortisol secretion (MACS) was diagnosed when cortisol after 1mg-dexamethasone suppression test (1-mg DST) was in the range of 1.8-5ug/dl. Non-functional adrenocortical tumor (NFAT) was diagnosed as cortisol following 1-mg DST less than 1.8ug/dL. We performed Logistics regression and causal mediation analyses, looking for associations between radiological characteristics and the HPA axis. Results: Of 494 patients, 352 (71.3%) with NFAT and 142 (28.7%) with MACS were included. Patients with MACS had a higher tumor diameter, thinner contralateral adrenal gland, and lower plasma ACTH and serum DHEAS than those with NFAT. ACTH (OR 0.978, 0.962-0.993) and tumor diameter (OR 1.857, 95%CI, 1.357-2.540) were independent factors associated with decreased serum DHEAS (all P<0.05). ACTH was also associated with decreased contralateral adrenal diameter significantly (OR 0.973, 95%CI, 0.957-0.988, P=0.001). Causal mediation analysis showed ACTH mediated the effect significantly for the association between 1-mg DST results and DHEAS level (Pmediation<0.001, proportion=22.3%). Meanwhile, we found ACTH mediated 39.7% of the effects of 1-mg DST on contralateral adrenal diameter (Pmediation=0.012). Conclusions: Patients with MACS had thinner contralateral adrenal glands and disturbed HPA axes compared with NFAT. ACTH may partially be involved in mediating the mild autonomous cortisol secretion to DHEAS and the contralateral adrenal gland.

Effects of a low-sodium diet in patients with idiopathic hyperaldosteronism: a randomized controlled trial.

Background: Idiopathic hyperaldosteronism (IHA) is one of the most common types of primary aldosteronism (PA), an important cause of hypertension. Although high dietary sodium is a major risk factor for hypertension, there is no consensus on the recommended dietary sodium intake for IHA. Objective: This study investigated the effect of a low-sodium diet on hemodynamic variables and relevant disease biomarkers in IHA patients, with the aim of providing a useful reference for clinical treatment. Methods: Fifty IHA patients were evenly randomized into two groups and provided, after a 7-day run-in period (100 mmol/d sodium), either a low-sodium diet (50 mmol/d sodium) or a normal sodium diet (100 mmol/d sodium) for an additional 7 days. After the 14-day intervention (conducted without potassium supplementation), changes in blood pressure (BP) and serum potassium were evaluated in both groups. Results: After the dietary intervention, the low sodium group exhibited, compared to the normal sodium group, decreased BP (SBP: 121.8 ± 12.8 vs. 129.9 ± 12.1 mmHg, p < 0.05; DBP: 82.6 ± 7.6 vs. 86.4 ± 8.2 mmHg, p < 0.05; MAP: 95.7 ± 8.8 vs. 100.9 ± 8.4 mmHg, p < 0.05) and increased serum potassium levels (3.38 ± 0.33 vs. 3.07 ± 0.27 mmol/L, p < 0.001). The low sodium group showed also better control of both BP and serum potassium: BP <140/90 mmHg in 70.0% of total patients (76.0% vs. 64.0%, in the low and normal sodium groups, respectively; p > 0.05), BP <130/85 mmHg in 38.0% of total patients (56.0% vs. 20.0%, p < 0.05), BP <120/80 mmHg in 28.0% of total patients (44.0% vs. 12.0%, p < 0.05); serum potassium ≥3.5 mmol/L in 22.0% of total patients (32.0% vs. 12.0% in the low and normal sodium groups, respectively; p = 0.088). There were differences between the controlled BP group (<120/80 mmHg) and the non-controlled BP group (≥120/80 mmHg) in gender, BP at baseline, and type of diet (low vs. normal sodium). Female gender and low-sodium diet were protective factors for BP control. Conclusions: A low-sodium diet is effective in lowering BP and elevating serum potassium in IHA patients. Female patients on a low-sodium diet are more likely to achieve BP control (<120/80 mmHg). We advocate a dietary sodium intake of 50 mmol/d for IHA patients. Clinical trial registration: https://clinicaltrials.gov, Identifier NCT05649631.

SDHB immunohistochemistry for prognosis of pheochromocytoma and paraganglioma: A retrospective and prospective analysis.

Introduction: Pheochromocytomas and paragangliomas (PCC/PGL) are rare neuroendocrine tumors and can secrete catecholamine. Previous studies have found that SDHB immunohistochemistry (IHC) can predict SDHB germline gene mutation, and SDHB mutation is closely associated with tumor progression and metastasis. This study aimed to clarify the potential effect of SDHB IHC as a predictive marker for tumor progression in PCC/PGL patients. Methods: We included PCC/PGL patients diagnosed in Ruijin Hospital, Shanghai Jiao Tong University School of Medicine from 2002 to 2014 for retrospective analysis and discovered that SDHB (-) staining patients had poorer prognoses. Then we examined SDHB protein expression by IHC on all tumors in the prospective series, which was composed of patients from 2015 to 2020 in our center. Results: In the retrospective series, the median follow-up was 167 months, and during follow-up, 14.4% (38/264) patients developed metastasis or recurrence, and 8.0% (22/274) patients died. Retrospective analysis revealed that 66.7% (6/9) of participants in the SDHB (-) group and 15.7% (40/255) of those in the SDHB (+) group developed progressive tumors (OR: 10.75, 95% CI: 2.72-52.60, P=0.001), and SDHB (-) was independently associated with poor outcomes after adjusting by other clinicopathological parameters (OR: 11.68, 95% CI: 2.58-64.45, P=0.002). SDHB (-) patients had shorter disease-free survival (DFS) and overall survival (OS) (P<0.001) and SDHB (-) was significantly associated with shorter median DFS (HR: 6.89, 95% CI: 2.41-19.70, P<0.001) in multivariate cox proportional hazard analysis. In the prospective series, the median follow-up was 28 months, 4.7% (10/213) patients developed metastasis or recurrence, and 0.5% (1/217) patient died. For the prospective analysis, 18.8% (3/16) of participants in the SDHB (-) group had progressive tumors compared with 3.6% (7/197) in the SDHB (+) group (RR: 5.28, 95% CI: 1.51-18.47, P=0.009), statistical significance remained (RR: 3.35, 95% CI: 1.20-9.38, P=0.021) after adjusting for other clinicopathological factors. Conclusions: Our findings demonstrated patients with SDHB (-) tumors had a higher possibility of poor outcomes, and SDHB IHC can be regarded as an independent biomarker of prognosis in PCC/PGL.

Platelet-Lymphocyte and Neutrophil-Lymphocyte Ratios Are Prognostic Markers for Pheochromocytomas and Paragangliomas.

CONTEXT: Preoperative inflammatory markers, such as the neutrophil-lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR), and lymphocyte-monocyte ratio (LMR), have recently been proposed as prognostic markers in different tumors. However, their predictive values in patients with pheochromocytomas and paragangliomas (PPGLs) are uncertain. OBJECTIVE: This study aimed to investigate the prognostic significance of inflammatory biomarkers in PPGL patients. METHODS: Data from 1247 consecutive PPGL patients between 2002 and 2020 were evaluated. The preoperative inflammatory markers were evaluated. The prognostic roles were identified by X-tile software, Kaplan-Meier curves, and Cox regression models. RESULTS: A total of 728 patients were included in the analysis, with a median follow-up of 63 months (IQR, 31-111 months); 31 individuals died, 28 patients developed metastases, and 12 patients developed recurrence. Our study showed that deaths were observed significantly more frequently in patients with high NLR(≥3.5) and high PLR (≥217.4) than those with low NLR (<3.5) (P = .003) and low PLR (<217.4) (P = .005). Elevated NLR (≥3.5) and elevated PLR (≥217.4) was significantly associated with decreased overall survival (OS) (P = .005), and elevated PLR (≥238.3) was significantly associated with decreased metastasis-free survival (MFS) (P = .021). Cox models illustrated that NLR and PLR were independent prognostic factors for OS, and PLR was an independent prognostic factor for MFS. CONCLUSION: Both elevated NLR and PLR are associated with poor prognosis in PPGLs. They are convenient predictive markers that could be used in daily clinical practice.

Efficacy and safety of adjuvant radiation therapy in localized adrenocortical carcinoma.

Context: Adrenocortical carcinoma (ACC) is rare and have high rates of recurrence and mortality. The role of adjuvant radiation therapy (RT) in localized ACC was controversial. Methods: We conducted a retrospective study in our center between 2015 and 2021 to evaluate the efficacy and safety of adjuvant RT in localized ACC. Overall survival (OS) and disease-free survival (DFS) were estimated using the Kaplan-Meier method. Cox proportional hazards regression models were used to estimate the independent risk factors. Adverse events associated with RT were documented according to the toxicity criteria of the radiation therapy oncology group (RTOG) and the common terminology criteria for adverse events (CTCAE v5.0). Results: Of 105 patients with localized ACC, 46 (43.8%) received adjuvant RT after surgery. The median radiation dose was 45.0Gy (range:30.0-50.4) and median follow up time was 36.5 (IQR: 19.7-51.8) months. In comparison to the no adjuvant RT group, patients with adjuvant RT had better 3-year OS (87.9% vs 79.5%, P=0.039), especially for patients with ENSAT I/II stage (P=0.004). Adjuvant RT also improved the median DFS time from 16.5months (95%CI, 12.0-20.9) to 34.6months (95%CI, 16.1-53.0). Toxicity of RT was generally mild and moderate with six grade 3 events. Conclusions: Postoperative adjuvant RT significantly improved OS and DFS compared with the use of surgery alone in resected ACC patients. Although this retrospective study on RT in localized ACC indicates that RT is effective in ACC, its findings need to be prospectively confirmed.

Reoperation for Recurrent and Persistent Cushing's Disease without Visible MRI Findings.

PURPOSE: Transsphenoidal surgery is the first-line treatment for Cushing's disease (CD), even with negative preoperative magnetic resonance imaging (MRI) results. Some patients with persistent or recurring hypercortisolism have negative MRI findings after the initial surgery. We aimed to analyze the efficacy of repeat surgery in two groups of patients and determine if there is an association between positive MRI findings and early remission. PATIENTS AND METHODS: Clinical, imaging, and biochemical information of 42 patients who underwent repeat surgery by a single neurosurgeon between 2002 and 2021 was retrospectively analyzed. We compared the endocrinological, histopathological, and surgical outcomes before and after repeat surgery among 14 CD patients with negative MRI findings and 28 patients with positive MRI findings. RESULTS: Immediate remission was achieved in 29 patients (69.0%) who underwent repeat surgery. Among all patients, 28 (66.7%) had MRI findings consistent with solid lesions. There was no significant difference in remission rates between the recurrence and persistence groups (77.8% vs. 57.1%, odds ratio = 2.625, 95% confidence interval = 0.651 to 10.586). Patients in remission after repeat surgery were not associated with positive MRI findings (odds ratio = 3.667, 95% confidence interval = 0.920 to 14.622). CONCLUSIONS: In terms of recurrence, repeat surgery in patients with either positive or negative MRI findings showed reasonable remission rates. For persistent disease with positive MRI findings, repeat surgery is still an option; however, more solid evidence is needed to determine if negative MRI findings are predictors for failed reoperations for persistent hypercortisolism.

Deep multi-scale resemblance network for the sub-class differentiation of adrenal masses on computed tomography images.

OBJECTIVE: The accurate classification of mass lesions in the adrenal glands ('adrenal masses'), detected with computed tomography (CT), is important for diagnosis and patient management. Adrenal masses can be benign or malignant and benign masses have varying prevalence. Classification methods based on convolutional neural networks (CNNs) are the state-of-the-art in maximizing inter-class differences in large medical imaging training datasets. The application of CNNs, to adrenal masses is challenging due to large intra-class variations, large inter-class similarities and imbalanced training data due to the size of the mass lesions. METHODS: We developed a deep multi-scale resemblance network (DMRN) to overcome these limitations and leveraged paired CNNs to evaluate the intra-class similarities. We used multi-scale feature embedding to improve the inter-class separability by iteratively combining complementary information produced at different scales of the input to create structured feature descriptors. We augmented the training data with randomly sampled paired adrenal masses to reduce the influence of imbalanced training data. RESULTS: We used 229 CT scans of patients with adrenal masses for evaluation. In a five-fold cross-validation, our method had the best results (89.52 % in accuracy) when compared to the state-of-the-art methods (p < 0.05). We conducted a generalizability analysis of our method on the ImageCLEF 2016 competition dataset for medical subfigure classification, which consists of a training set of 6776 images and a test set of 4166 images across 30 classes. Our method achieved better classification performance (85.90 % in accuracy) when compared to the existing methods and was competitive when compared with methods that require additional training data (1.47 % lower in accuracy). CONCLUSION: Our DMRN sub-classified adrenal masses on CT and was superior to state-of-the-art approaches.

Newly diagnosed diabetes mellitus is a risk factor for cardiocerebrovascular events in primary aldosteronism.

OBJECTIVE: To explore the prevalence and clinical significance of newly diagnosed diabetes mellitus (DM) in patients with primary aldosteronism (PA). Investigating the risk factors for cardiocerebrovascular disease (CCVD) will guide strategies for reducing CCVD in patients with PA. METHODS: We retrospectively included 729 PA patients without DM and conducted oral glucose tolerance tests. RESULTS: We found that 15.0% of PA patients had newly diagnosed DM. The DM prevalence increased with elevated aldosterone levels [OR = 3.20 (1.77, 5.78), P value < 0.001]. The rate of CCVD in newly diagnosed diabetic PA patients was higher than that in nondiabetic PA patients at diagnosis (11.9% vs. 5.0%, P = 0.005). Furthermore, multivariate logistic analysis revealed that HT duration [1.055 (1.002,1.111), P = 0.041] and newly diagnosed DM [2.600 (1.072,6.303), P = 0.034] were significantly associated with CCVD in PA patients. CONCLUSION: The prevalence of newly diagnosed DM in PA patients was higher than that in the general population. Aldosterone level was an independent risk factor for DM not for CCVD. CCVD was correlated with longer HT duration and newly diagnosed DM. Therefore, it is crucial to screen DM at the diagnosis in PA patients.

Association of Famine Exposure on the Changing Clinical Phenotypes of Primary Hyperparathyroidism in 20 years.

Background & Aims: Primary hyperparathyroidism(PHPT) has been evolving into a milder asymptomatic disease. No study has assessed the association between famine exposure and such a shift. We aim to explore the effects of China's Great Famine exposure on the changing pattern of PHPT phenotypes. Methods: 750 PHPT patients diagnosed from 2000 to 2019 were studied. The clinical presentations were compared between them in recent 10 years (2010-2019) and previous 10 years (2000-2009). Participants were then categorized into fetal, childhood, adolescent, adult exposure, and unexposed groups. Logistic regression was used to estimate the odds ratios (ORs) and confidence intervals (CIs) of famine exposure as factors contributing to the changes in the clinical presentations of PHPT. Results: Serum levels of PTH, albumin-corrected Ca, tumor size, eGFR, BMDs (all P<0.001), and clinical symptoms became milder in recent 10 years. Famine exposure (72.6% vs 58.4%, P<0.001), especially the adult exposure (18.8% vs 4.1%, P<0.001)was significant less in recent 10 years. The ORs (95%CIs) of having upper 3rd tertile PTH were 2.79(1.34,5.8), 2.07(1.04,4.11), 3.10(1.15,8.38) and 8.85(2.56,30.56) for patients with fetal, childhood, adolescent and adult famine exposure, respectively. The ORs (95%CIs) of upper 3rd tertile albumin-corrected Ca and upper 3rd tertile of tumor size was 4.78(1.39, 16.38) and 4.07(1.12,14.84) for participants with adult famine exposure, respectively. All these associations were independent of age, sex, disease duration and other confounders. Conclusions: The clinical manifestations of PHPT in China continue to be milder. Exposure to famine is associated with PHPT. Less famine exposure might be responsible for the mile form of PHPT in recent years.

Mutational landscape of non-functional adrenocortical adenomas.

Adrenal incidentalomas are the most frequent human neoplasms. Recent genomic investigations on functional adrenocortical tumors have demonstrated that somatic mutations in PRKACA and KCNJ5 responsible for the development of adrenocortical adenomas (ACAs) are associated with hypercortisolism and aldosteronism, respectively. Several studies have identified CTNNB1 mutations in ACAs and have been mostly involved in the tumorigenesis of non-functional ACA (NFACA). However, integrated genomic characterization of NFACAs is lacking. In the current study, we utilized pan-genomic methods to comprehensively analyze 60 NFACA samples. A total of 1264 somatic mutations in coding regions among the 60 samples were identified, with a median of 15 non-silent mutations per tumor. Twenty-two NFACAs (36.67%) had genetic alterations in CTNNB1. We also identified several somatic mutations in genes of the cAMP/PKA pathway and KCNJ5. Histone modification genes (KMT2A, KMT2C, and KMT2D) were altered in 10% of cases. Germline mutations of MEN1 and RET were also found. Finally, by comparison of our transcriptome data with those available in the TCGA, we illustrated the molecular characterization of NFACA. We revealed the genetic profiling and molecular landscape of NFACA. Wnt/ß-catenin pathway activation as shown ssby nuclear and/or cytoplasmic ß-catenin accumulation is frequent, occurring in about one-third of ACA cases. cytochrome P450 enzymes could be markers to reveal the functional status of adrenocortical tumors. These observations strongly suggest the involvement of the Wnt/ß-catenin pathway in benign adrenal tumorigenesis and possibly in the regulation of steroid secretion.

Immunohistochemical Analysis of CYP11B2, CYP11B1 and ß-catenin Helps Subtyping and Relates With Clinical Characteristics of Unilateral Primary Aldosteronism.

Background: Primary aldosteronism is caused by aldosterone overproduction. While conventional hematoxylin-eosin staining can demonstrate morphological abnormality, it cannot provide any functional histopathological information. We aimed to identify the diagnostic, functional and prognostic value of CYP11B2, CYP11B1, and ß-catenin immunostaining in unilateral hyperaldosteronism. Method: A total of 134 patients with unilateral hyperaldosteronism were recruited in our study. The expression of CYP11B2, CYP11B1, and ß-catenin was evaluated semiquantitatively on 134 patients' sections using immunohistochemistry technology and the relationship with clinical data was assessed. Results: Patients were classified into four subtypes based on CYP11B2 staining as below: (1)118 patients with unilateral single aldosterone-producing adenoma (APA), (2)11 with unilateral multiple APA, (3)four with aldosterone-producing cell cluster (APCC), and (4)one with an undefined source. Adjusted CYP11B2 H-score was correlated with serum aldosterone, aldosterone to renin ratio (ARR), and serum potassium. In the abnormal ß-catenin staining group, hypertension duration, aldosterone, ARR, cortisol, tumor diameter, tumor area, and CYP11B2 H-score were significantly higher than those of the wild-type group. Serum potassium level was significantly lower in the abnormal ß-catenin staining group. Age, gender, BMI, family history of hypertension, adjusted CYP11B2 and CYP11B1 H-scores differed significantly between complete clinical success and incomplete clinical success groups. Age, gender and family history of hypertension were independently associated with complete clinical success based on multivariate logistic regression analysis. Conclusion: CYP11B2 immunostaining could improve the differential diagnosis of unilateral hyperaldosteronism. Adjusted CYP11B2 H-score could be used as a histopathological marker to reflect the severity of unilateral APA. Dysregulation of Wnt/ß-catenin signaling and impaired ß-catenin degradation may provoke the proliferation and enhance the steroidogenic ability of APA tumor cells, indicating that the Wnt pathway might be a potential, actionable, therapeutic target in the treatment of hyperaldosteronism. Age, sex and family history of hypertension were independent predictors of clinical outcome after adrenalectomy for unilateral hyperaldosteronism.

KCNJ5 Mutation Contributes to Complete Clinical Success in Aldosterone-Producing Adenoma: A Study From a Single Center.

OBJECTIVE: The KCNJ5 mutation is the most frequent mutation in aldosterone-producing adenoma (APA). We aimed to illustrate the relationship between KCNJ5 and prognosis after adrenalectomy as a guide for further treatment. METHODS: Our study included 458 patients with APA. Tumor tissues were screened for somatic mutations in KCNJ5 hot-spot regions. We performed a retrospective analysis to identify correlations between KCNJ5 and clinical outcomes in 334 patients with adrenal venous sampling lateralization. RESULTS: Somatic KCNJ5 mutations were identified in 324 of 458 patients with APA (70.7%). Compared with the KCNJ5-wild type patients, patients with KCNJ5 mutations were younger, had a higher proportion of women, and had shorter durations of hypertension, lower body mass indexes (BMIs), and lower systolic blood pressure values (P < .05). During follow-up, among the 334 patients with APA with adrenal venous sampling lateralization, 320 (95.8%) presented complete biochemical success and 187 (56.0%) presented complete clinical success. One hundred eighty-seven patients with primary aldosteronism who achieved complete clinical success presented the following characteristics: age <40 years (78.7%), BMI <24 kg/m2 (71.0%), hypertension duration <5 years (78.4%), females (66.9%), and KCNJ5 mutation (65.5%). A multivariate logistic regression analysis identified BMI, hypertension duration, and KCNJ5 mutation as independent predictors of complete clinical success. CONCLUSION: The prevalence of KCNJ5 mutations was 70.7%. KCNJ5 mutation is a protective factor of complete clinical success, while BMI and hypertension duration were risk factors of incomplete clinical success.

The Clinical Features and Molecular Mechanisms of ACTH-secreting Pancreatic Neuroendocrine Tumors.

OBJECTIVE: Pancreatic neuroendocrine tumors (pNETs) causing ectopic adrenal corticotropic hormone (ACTH) syndrome (EAS) are rare and aggressive with little known information. We aimed to elucidate the clinical features and molecular mechanisms of pNETs with EAS by methylation analysis. METHODS: Seven patients with ectopic ACTH-secreting pNETs who were diagnosed at Shanghai Clinical Endocrine and Metabolic Diseases Center and Pancreatic Disease Center in Ruijin Hospital between 2001 and 2019 were enrolled. Twenty patients with ectopic ACTH-secreting thymic neuroendocrine tumors (TNETs) and 7 with nonfunctional pNETs (nf-pNETs) were also enrolled as controls. We collected clinical data and measured POMC promoter CpG methylation. RESULTS: All 7 patients had elevated ACTH and urinary free cortisol (UFC) levels with positive ACTH staining in the pancreas and were diagnosed with ectopic ACTH-secreting pNET. Of the 7 patients, 6 underwent surgery and 1 underwent transarterial embolization (TAE). Two patients were free of disease after surgery; 2 died within 90 days after surgery; and 3 had metastases and died within 1 year. Compared with ACTH-secreting TNETs, ACTH-secreting pNETs had similar clinical and biochemical features but a significantly poorer prognosis. POMC promoter CpG methylation was significantly lower in ACTH-secreting pNETs than in nf-pNETs and normal pancreas. CONCLUSIONS: ACTH-secreting pNETs are aggressive and fatal. Surgery is definitively curative for patients with resectable primary tumors without metastasis. Pro-opiomelanocortin (POMC) promoter hypomethylation caused pNETs to produce ACTH. This study further supplements the genetic features of ACTH-secreting NETs.

Association between thyroid function and serum cortisol in cortisol-producing adenoma patients.

PURPOSE: Thyroid dysfunction has been reported in hypercortisolism. Previous findings regarding changes in thyroid function due to cortisol-producing adenoma (CPA) have been inconsistent. The study aimed to investigate the association between thyroid function and excessive cortisol secretion in patients with CPA and to explore the changes in pituitary function after adrenalectomy. METHODS: We conducted a retrospective study; thyroid function was evaluated in 94 patients with CPA and 94 healthy controls (HC) matched for age and sex. A total of 94 patients with nonfunctioning adrenal incidentalomas (NFAIs) were recruited as a second control group. RESULTS: Serum thyroid stimulating hormone (TSH) and free thyroxine (T4) levels were significantly lower in the CPA group than in the HC and NFAIs groups (P < 0.001). The prevalence of central hypothyroidism was 12.8% in the CPA group and increased according to serum cortisol quartiles (P for trend = 0.025). According to the stepwise multiple linear regression analysis, serum cortisol was negatively associated with TSH and free T4 levels in the CPA group after adjustment for body mass index and age. Furthermore, decreased TSH levels were corrected by adrenalectomy [0.75 (0.50, 1.14) vs. 1.91 (1.36, 2.71) µIU/ml, P < 0.001], in parallel with a recovery in free T4 levels [11.20 (10.00, 12.43) vs. 12.04 (11.24, 13.01), P < 0.001]. Postoperative growth hormone and prolactin levels did not change compared with baseline. CONCLUSION: Serum TSH and free T4 levels were decreased in patients with CPA, and dysfunction of the hypothalamic-pituitary-thyroid axis might be reversible after surgery.

From clinic to mechanism: Proteomics-based assessment of angiogenesis in adrenal pheochromocytoma.

Adrenal pheochromocytoma (PCC) is a very rare tumor that stems from chromaffin cells, which can develop into malignant tumor. During the operation, abundant blood vessels were often observed in PCC than other adrenal tumors, which increases the difficulty and risk of the surgery. Therefore, it is important to investigate the mechanism of PCC angiogenesis. Twelve surgical specimens of PCC from Ruijin Hospital, Shanghai Jiaotong University were grouped into high and low microvessel density (MVD) group. They were also divided into rich blood supply and nonenriched blood supply group, according to computed tomography (CT) manifestation. Comparative proteomic analysis based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) and bioinformatics analysis revealed that 206 proteins differentially regulated in the high MVD group compared with low MVD group (p < 0.05). Besides, 61 proteins were discovered to be significantly changed when the 12 samples were grouped according to CT manifestation. By intersecting the differentially changed protein from MVD and CT grouping, 25 proteins were filtered out, with pathological function. COX4I2 was verified to be increased gradually with angiogenesis with increasing severity, and PLAT was shown to be decreased with angiogenesis in PCC, by quantitative reverse-transcription polymerase chain reaction and immunohistochemistry. The quantitative proteomics result indicated that the tumor angiogenesis in PCC is associated with hypoxia. COX4I2 and PLAT were highly correlated with blood supply in PCC which contribute to angiogenesis in PCC, which could be used as biomarkers to better indicate tumor angiogenesis, or targets to regress tumor angiogenesis as treatment.