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MaryO

~Chief Cushie~
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  1. MaryO'Note: I found this article very simplistic. What do you think? This article is for informational purposes only and is not a substitute for professional medical advice, diagnosis or treatment. Contact a qualified medical professional before engaging in any physical activity, or making any changes to your diet, medication or lifestyle. Imagine the heart-pounding rush of adrenaline you’d get while bungee jumping or zip lining — that’s what Angela Yawn felt all the time before receiving her diagnosis. In a span of six years, the 49-year-old gained 52 kg (115 lbs) and suffered from joint swelling, headaches, skin redness and a racing heart. “I would put my hand on my chest because it made me feel like that’s what I needed to do to hold my heart in,” Yawn, who lives in Griffin, U.S., told Today. “I noticed it during the day, but at night when I was trying to lie down and sleep, it was worse because I could do nothing but hear it beat, feel it thump.” Yawn recalled being the most frustrated with the weight gain, as she’d put on 1 kg (2 lbs) a day while only eating 600 calories. “I was going crazy,” she said. After dozens of doctors couldn’t piece together her seemingly unrelated symptoms, Yawn sought out the help of an endocrinologist in February 2021. Blood tests and an MRI confirmed that Yawn had a tumour in her pituitary gland — a small, pea-sized organ at the base of the brain — that caused the gland to release excess adrenocorticotropic hormones. As a result, she became inundated with cortisol, a steroid the body releases in response to danger or stress. This combination of factors led to her diagnosis — Cushing’s disease. Read on to learn more about Cushing’s disease, signs and symptoms as well as how it can be prevented. What is Cushing’s disease? “Cushing’s disease is a rare but serious condition that is caused by a pituitary tumour," a specialist from the University of California, Los Angeles (UCLA) pituitary team tells Yahoo Canada. "The gland releases excessive adrenocorticotropic hormones and cortisol into the blood over a long period of time. It’s a hormonal disorder that is sometimes called hypercortisolism, and you will need to see an endocrinologist or someone who specializes in hormonal-related diseases to confirm your diagnosis and to help you receive proper care.” Cushing’s disease is not the same as Cushing’s syndrome, which refers to elevated levels of cortisol in the blood and is much more common than Cushing’s disease. Unlike the disease, Cushing’s syndrome can be caused by taking medications that have cortisol such as prednisone, asthma inhalers and joint steroid injections. Who is at risk for Cushing’s disease? Cushing’s disease is incredibly rare, resulting in only 10 to 15 new cases per million people in the United States each year, according to UCLA Health. “It’s most commonly found in people between the ages of 20 and 50, and affects about three times more women than men,” the UCLA source, who asked not to be named, says. “However, you might be more at risk if you have high blood pressure, if you’re overweight or if you have type 2 diabetes.” What are the signs and symptoms of Cushing’s disease? Although each person may have a unique combination of symptoms, patients typically experience changes to their physical appearance, according to Mayo Clinic. “It’s very common to see rapid weight gain, red cheeks and bruising of the skin,” the UCLA source says. “I’ve also seen patients with generalized fatigue, depression, high blood pressure, a rapid heartbeat and loss of vision.” “The symptoms can seem random or unrelated, which is why it can be so hard to diagnose,” they add. To establish if you have the disease, your doctor will conduct a physical exam and ask you about your symptoms and medical history. Generally, the first step in diagnosing Cushing's disease is determining the state of excess cortisol in the blood. Afterwards, an MRI will determine if a pituitary tumour is visible. If you have symptoms of Cushing’s disease, you should make an appointment to see a doctor or endocrinologist. How is Cushing’s disease treated? In the last decade, treatment options have changed thanks to several breakthroughs in pituitary science. “Surgery to remove the tumour is normally the first treatment option. It’s minimally invasive, has a fairly high success rate and it’s the only long-term cure for Cushing’s disease at the moment,” explains the UCLA source. If surgery isn’t an option or doesn’t solve the problem, medication and radiation therapy are other ways to treat the disease. “No matter the stage of the disease at the time of diagnosis, treating it requires an experienced specialist or team of doctors familiar with pituitary tumours,” the UCLA source adds. How can I prevent Cushing’s disease? “There’s no tried and true method of preventing the condition,” the source explains. “But if you’re at risk or if you think you have the disease, I always recommend having a doctor monitor your cortisol levels on a regular basis.” The UCLA source also recommends implementing healthy lifestyle changes that can help prevent high blood pressure. Examples include reducing stress, getting adequate sleep, exercising regularly and eating a healthy diet that's rich in fruits, vegetables and whole grains. Adapted from https://ca.news.yahoo.com/what-is-cushings-disease-experts-warn-rare-serious-condition-120015725.html
  2. Abstract Cushing's disease causes numerous metabolic disorders, cognitive decline, and sarcopenia, leading to deterioration of the general health in older individuals. Cushing's disease can be treated with transsphenoidal surgery, but thus far, surgery has often been avoided in older patients. We herein report an older woman with Cushing's disease whose cognitive impairment and sarcopenia improved after transsphenoidal surgery. Although cognitive impairment and sarcopenia in most older patients show resistance to treatment, our case indicates that normalization of the cortisol level by transsphenoidal surgery can be effective in improving the cognitive impairment and muscle mass loss caused by Cushing's disease. References (27) 1. Lindholm J, Juul S, Jorgensen JO, et al. Incidence and late prognosis of Cushing's syndrome: a population-based study. J Clin Endocrinol Metab 86: 117-123, 2001. 2. Starkman MN. Neuropsychiatric findings in Cushing syndrome and exogenous glucocorticoid administration. Endocrinol Metab Clin N Am 42: 477-488, 2013. 3. Chen YF, Li YF, Chen X, Sun QF. Neuropsychiatric disorders and cognitive dysfunction in patients with Cushing's disease. CMJ 26: 3156-3160, 2013. 4. Priego T, Martín AI, González-Hedström D, Granado M, López-Calderón A. Role of hormones in sarcopenia. Vitam Horm 115: 535-570, 2021. 5. Grossman R, Mukherjee D, Chaichana KL, et al. Complications and death among elderly patients undergoing pituitary tumour surgery. Clin Endocrinol 73: 361-368, 2010. 6. Tiemensma J, Kokshoom NE, Biermasz NR, et al. Subtle cognitive impairments in patients with long-term cure of Cushing's disease. J Clin Endocrinol Metab 95: 2699-2714, 2010. 7. Brunetti A, Fulham MJ, Aloj L, et al. Decreased brain glucose utilization in patients with Cushing's disease. J Nucl Med 39: 786-790, 1998. 8. Liu S, Wang Y, Xu K, et al. Brain glucose metabolism is associated with hormone level in Cushing's disease: A voxel-based study using FDG-PET. Neuroimage Clin 12: 415-419, 2016. 9. Cheng H, Gao L, Hou B, et al. Reversibility of The cerebral blood flow in Patients with Cushing's Disease after Surgery Treatment. Metabolism 104: 154050, 2020. 10. Forget H, Lacroix A, Somma M, Cohen H. Cognitive decline in patients with Cushing's syndrome. J Int Neuropsychol Soc 6: 20-29, 2000. 11. Kim KJ, Filosa JA. Advanced in vitro approach to study neurovascular coupling mechanisms in the brain microcirculation. J Physiol 590: 1757-1770, 2012. 12. McEwen BS, Bowles NP, Gray JD, et al. Mechanisms of stress in the brain. Nat Neurosci 18: 1353-1363, 2015. 13. Rajkowska G, Miguel-Hidalgo JJ. Gliogenesis and glial pathology in depression. CNS Neurol Disord Drug Targets 6: 219-233, 2007. 14. Iuchi T, Akaike M, Mitsui T, et al. Glucocorticoid excess induces superoxide production in vascular endothelial cells and elicits vascular endothelial dysfunction. Circ Res 92: 81-87, 2003. 15. Cheng H, Gao L, Hou B, et al. Reversibility of the cerebral blood flow in patients with Cushing's disease after surgery treatment. Metabolism 104: 154050, 2020. 16. Frimodt-Møller KE, Møllegaard Jepsen JR, Feldt-Rasmussen U, et al. Hippocampal volume, cognitive functions, depression, anxiety, and quality of life in patients with Cushing syndrome. J Clin Endocrinol Metab 104: 4563-4577, 2019. 17. Siegel S, Kirstein CF, Grzywotz A, et al. Neuropsychological functioning in patients with cushing's disease and Cushing's syndrome. Exp Clin Endocrinol Diabetes 129: 194-202, 2021. 18. Chow Y, Masiak J, Mikołajewska E, et al. Limbic brain structures and burnout-A systematic review. Adv Med Sci 63: 192-198, 2018. 19. Starkman MN, Gebarski SS, Berent S, Schteingart DE. Hippocampal formation volume, memory dysfunction, and cortisol levels in patients with Cushing's syndrome. Biol Psychiatry 32: 756-765, 1992. 20. Goldberg AL, Tischler M, Demartina G, Griffin G. Hormonal regulation of protein degradation and synthesis in skeletal muscle. Fed Proc 39: 31-36, 1980. 21. Miller BS, Ignatoski KM, Daignault S, et al. A quantitative tool to assess degree of sarcopenia objectively in patients with hypercortisolism. Surgery 150: 1178-1185, 2011. 22. Delivanis D, Iñiguez-Ariza N, Zeb M, et al. Impact of hypercortisolism on skeletal muscle mass and adipose tissue mass in patients with adrenal adenomas. Clin Endocrinol 88: 209-216, 2018. 23. Kim JH, Kwak MK, Ahn SH, et al. Alteration in skeletal muscle mass in women with subclinical hypercortisolism. Endocrine 61: 134-143, 2018. 24. Gonzalez Rodriguez E, Marques-Vidal P, Aubry-Rozier B, et al. Diurnal salivary cortisol in sarcopenic postmenopausal women: the OsteoLaus Cohort. Calcif Tissue Int 109: 499-509, 2021. 25. Pivonello R, Fleseriu M, Newell-Price J, et al. Efficacy and safety of osilodrostat in patients with Cushing's disease (LINC 3): a multicentre phase III study with a double-blind, randomised withdrawal phase. Lancet Diabetes Endocrinol 8: 748-761, 2020. 26. Lau D, Rutledge C, Aghi MK. Cushing's disease: current medical therapies and molecular insights guiding future therapies. Neurosurg Focus 38: E11, 2015. 27. Villar-Taibo R, Díaz-Ortega C, Sifontes-Dubo M, et al. Pituitary surgery in elderly patients: a safe and effective procedure. Endocrine 2: 814-822, 2021. From https://www.jstage.jst.go.jp/article/internalmedicine/advpub/0/advpub_8326-21/_article
  3. RareCare® Since 1987, NORD has provided assistance programs to help patients obtain life-saving or life-sustaining medication they could not otherwise afford. These programs provide medication, financial assistance with insurance premiums and co-pays, diagnostic testing assistance, and travel assistance for clinical trials or consultation with disease specialists. Read NORD’s Coronavirus: Business Continuity Response for RareCare® patient assistance programs. Cushing’s Syndrome | Accepting Applications Medical Assistance Co-Pay Assistance Premium Assistance Contact: 1-855-864-4018 Email: cushings@rarediseases.org Fax: 1-203-517-0978 Other diseases:https://rarediseases.org/for-patients-and-families/help-access-medications/patient-assistance-programs-2
  4. Abstract Cushing syndrome is a rare disease that rarely presents as acute psychosis. In this case, the patient presented with acute psychosis and agitation as the first manifestations of the disease which led to the admission of the patient to a psychiatry hospital for one month, as it was difficult to restrain her sufficiently for performing appropriate diagnostic tests due to disturbing behavior. She responded well to treatment with olanzapine and lorazepam to treat the patient’s agitation, and successfully complete her evaluation. Thereafter, she was diagnosed with a pituitary tumor and underwent pituitary lesion resection via a microscopic transsphenoidal as needed. Two months after surgery, her cortisol levels returned to baseline, and she became calmer and decreased the tensity of her psychosis; however, it was only five months after surgery that her psychotic symptoms and disturbed behavior ceased. Introduction Cushing syndrome is comprised of a group of symptoms induced by prolonged exposure to high blood cortisol levels [1]. It is a rare disease, occurring in approximately 2.4 per million individuals per year [2]. Psychiatric and cognitive manifestations of Cushing syndrome occur in 70%-85% of patients, with irritability, emotional lability, and depression occurring most commonly. Rarer symptoms include mania, panic attacks, anxiety, suicidal ideation, and acute psychosis [3-5]. In this article, we describe a patient with Cushing syndrome who developed psychosis with agitation as the first manifestation of Cushing syndrome. The patient was difficult to manage since her agitation and refusal to undergo evaluation prevented her from receiving outpatient care. Case Presentation A 22-year-old woman with a three-month history of an increase in appetite, binge eating, and weight gain. After two weeks of her initial symptoms, she started to have grandiose and persecutory delusions, auditory hallucinations, decreased need for sleep, agitation, irritability, and aggression for which she went to a private psychiatry clinic and was given 10 mg olanzapine oral at night. After a month of starting oral olanzapine, she was not improving and was admitted to the psychiatry ward for evaluation. During her admission period, she started to have cognitive symptoms including worsened memory, attention, and orientation. After one month of admission with no improvement on medication, she was noted to have moon face and high blood pressure, and her laboratory investigation showed mild hypokalemia, high cortisol level, and adrenocorticotropic hormone (ACTH), elevated liver enzymes, and mild hypertriglyceridemia. A magnetic resonance imaging (MRI) scan of the brain revealed a 6 × 2-mm hyperintense lesion in the anterior pituitary on a T2-weighted image; therefore, she was transferred to our hospital for further work up and management as we have the endocrine facility. She had no past psychiatric history or family history of psychiatric illnesses, nor a history of substance abuse. She also had no past medical history and was not on any medication prior to this presentation. The patient was admitted to the endocrine department to evaluate the possibility of Cushing syndrome. Her blood pressure (150/98), heart rate (128 BPM), and respiratory rate (30 BPM) were elevated. She was treated with losartan, amlodipine, and spironolactone. Basic labs were done (Table 1). Therefore, insulin therapy was initiated. The evaluation of the patient’s condition was difficult as she was aggressive and uncooperative due to a lack of insight. Her primary team planned for sedation with anesthesia to facilitate a clinical evaluation; however, no intensive care unit bed was available. Lab test Patient result Reference values cortisol levels 1549 nmol/L 140 to 690 nmol/L ACTH (Adrenocorticotropic Hormone) 54 pg/mL 10 to 50 pg/mL ALT (Alanine transaminase) 305 U/L 7 to 56 U/L AST (Aspartate aminotransferase) 112 U/L 8 to 33 U/L Alkaline phosphatase 141 IU/L 44 to 147 IU/L Hemoglobin A1c 7.3% 5.7% to 6.4% Table 1: Lab results for the patient when she first came to our hospital Psychiatry was consulted to manage agitation. We started her on 5 mg olanzapine oral twice daily, and 2 mg lorazepam three times daily intravenous when oral was not possible. Maximum dosage of 5 mg olanzapine and 2 mg lorazepam every four hours were administered as required to manage agitation. Her ECG showed a QTC of 464. One-to-one nurse observation was initiated to detect risky behaviors. The patient slept well and became calmer and more cooperative throughout evaluations when receiving medication. One-to-one nurse observation was discontinued after five days, and lorazepam administration was reduced to two times daily. She remained easily provoked with grandiose and persecutory delusions, auditory hallucinations, and confusion. As the patient calmed, the primary team continued clinical evaluations. A contrast-enhanced MRI showed a focal non-deforming and hypo-enhancing lesion, measuring 7 mm (AP) x 6 mm (TV) x 6 mm (CC), in the anterior pituitary (Figures 1, 2). A minimal leftward deviated pituitary stalk with normal thickness was also identified. An 8 mg dexamethasone suppression test revealed cortisol levels had decreased from 1,500 to 900 nmol/L. The 24-hour cortisol level was not determined, as the patient was easily provoked. Inferior petrosal sinus sampling was performed under general anesthesia. These results are consistent with central Cushing disease. Figure 1: Coronal T1-weighted MRI of the pituitary gland with contrast showed a hypoenhancing nodular lesion at the midline of the anterior pituitary, with mild eccentric to the right Figure 2: Brain MRI sagittal view showing focal anterior pituitary hypoenhancing lesion at the midline and eccentric to the right Treatment with 250 mg metyrapone twice daily was initiated and the patient was scheduled for pituitary lesion resection via a microscopic transsphenoidal approach by neurosurgery. Her blood tests began normalizing post-surgery except for low cortisol (Table 2), and her vital signs were within normal range. Medications regulating blood pressure and glucose levels were decreased to monotherapy and discontinued thereafter. And 40 and 20 mg doses of hydrocortisone administered in the morning and night, respectively, were tapered to 5 mg twice daily over a period of two months after the surgery, and cortisol levels were regulated reaching 167 nmol/L. Agitation and irritability, grandiose and persecutory delusion and auditory hallucination tensity were reduced, with intact cognitive and memory function. Therefore, medication dosages were gradually reduced, starting with lorazepam. Lab Test Patient result Reference values cortisol levels 68 nmol/L 140 to 690 nmol/L ACTH (Adrenocorticotropic Hormone) 25 pg/ml 10 to 50 pg/mL ALT (Alanine transaminase) 17.2 U/L 7 to 56 U/L AST (Aspartate aminotransferase) 19.2 U/L 8 to 33 U/L Alkaline phosphatase 121 IU/L 44 to 147 IU/L TSH (Thyroid Stimulating Hormone) 1.8 mIU/L 0.5 to 5.0 mIU/L Table 2: Lab results after the surgery. Before discharge, the patient’s psychotropic medications were withheld by the primary team for two days due to oversedation. Upon discharge, due to the side effects of olanzapine, the patient was switched to oral risperidone 1 mg at night, with 0.5 mg oral clonazepam twice daily as needed for agitation and psychosis. Throughout follow-up, the patient experienced ongoing psychosis with disturbed behavior even though she is using received clonazepam twice daily. Therefore, her dosage of risperidone was increased to 2 mg orally at night, and oral clonazepam (0.5 to 1 mg) was administered three times daily as needed to manage agitation. After three months of discharge (five months from surgical intervention), her levels of agitation and irritability decreased, delusions and auditory hallucinations ceased, and she returned to baseline, and clonazepam was discontinued and risperidone dosage was tapered to 0.5 mg with observation and follow up in the clinic, and no symptom relapse was observed. The complete discontinuation of her medications is planned next visit while monitoring the patient for signs of relapse. Discussion Cushing syndrome may initially present as psychosis, which may be misdiagnosis as a primary psychotic disorder, delaying the proper diagnosis and management. Our patient presented to a psychiatry hospital before being referred to us because she resisted psychosis treatment, the resistance to treatment of primary illness due to psychiatric manifestation is not uncommon, as Fujii et al. [6] reported the management of a patient who resisted schizophrenia treatment for 10 years before being diagnosed with Cushing syndrome. Agitation with psychosis is likely the main obstacle for properly evaluating, diagnosing, and treating patients with Cushing syndrome. In our patient, we aimed to reduce her agitation to facilitate clinical evaluation. The organic cause of psychosis often responds poorly to antipsychotic medication and exhibits a challenge in managing agitation which necessitate the utilization of highly sedating medications, to facilitate further clinical evaluation. Shah et al. [7] reported similar difficulty treating a patient with agitation despite prescribing lorazepam and 1 mg haloperidol twice daily, agitation was poorly controlled. In our case, the patient responds to a high dose of Olanzapine with lorazepam in a better way than the case report that was managed with haloperidol with lorazepam. Psychiatric symptoms secondary to medical conditions usually occur transiently and they resolve after treatment of the primary cause, however, the duration for complete resolution of symptoms is unknown. In our case, the patient gradually improved for three months prior to achieving remission, whereas a patient reported by Wu et al. [8] went into complete remission one-month post-cortisol level correction. Conclusions Cushing syndrome, like many other endocrine diseases, can present as treatment-resistant psychiatric symptoms, which may be missed and treated as a primary psychiatric illness due to the lack of proper assessment and management. In this study, we tried to correlate the psychiatric symptoms with Cushing syndrome, the challenges we faced, and the response to the treatment. Our case report gives an insight into possible rare secondary causes of psychosis and advice a thorough evaluation of patients. References Your bibliography. (2021). Accessed: March 27, 2021: https://www.ncbi.nlm.nih.gov/books/NBK470218/. Etxabe J, Vazquez JA: Morbidity and mortality in Cushing's disease: an epidemiological approach. Clin Endocrinol (Oxf). 1994, 40:479-84. 10.1111/j.1365-2265.1994.tb02486.x Starkman MN, Schteingart DE: Neuropsychiatric manifestations of patients with Cushing’s syndrome. Relationship to cortisol and adrenocorticotropic hormone levels. Arch Intern Med. 1981, 215:9. 10.1001/archinte.1981.00340020077021 Dorn LD, Burgess ES, Dubbert B, et al.: Psychopathology in patients with endogenous Cushing's syndrome: 'atypical' or melancholic features. Clin Endocrinol (Oxf). 1995, 43:433-42. 10.1111/j.1365-2265.1995.tb02614.x Sharma ST, Nieman LK, Feelders RA: Cushing's syndrome: epidemiology and developments in disease management. Clin Epidemiol. 2015, 7:281-93. 10.2147/CLEP.S44336 Fujii Y, Mizoguchi Y, Masuoka J, et al.: Cushing’s syndrome and psychosis: a case report and literature review. Prim Care Companion CNS Disord. 2018, 20:18br02279. 10.4088/PCC.18br02279 Shah K, Mann I, Reddy K, John G: A case of severe psychosis due to Cushing’s syndrome secondary to primary bilateral Macronodular adrenal hyperplasia. Cureus. 2019, 11:e6162. 10.7759/cureus.6162 Wu Y, Chen J, Ma Y, Chen Z: Case report of Cushing’s syndrome with an acute psychotic presentation. Shanghai Arch Psychiatry. 2016, 28:169-72. 10.11919/j.issn.1002-0829.215126 From https://www.cureus.com/articles/98986-cushings-syndrome-with-acute-psychosis-a-case-report
  5. Dr. Friedman uses several medications to treat Cushing’s syndrome that are summarized in this table. Dr. Friedman especially recommends ketoconazole. An in-depth article on ketoconazole can be found on goodhormonehealth.com. Drug How it works Dosing Side effects Ketoconazole (Generic, not FDA approved in US) blocks several steps in cortisol biosynthesis Start 200 mg at 8 and 10 PM, can up titrate to 1200 mg/day • Transient increase in LFTs • Decreased testosterone levels • Adrenal insufficiency Levoketoconazole (Recorlev) L-isomer of Ketoconazole Start at 150 mg at 8 and 10 PM, can uptitrate up to 1200 mg nausea, vomiting, increased blood pressure, low potassium, fatigue, headache, abdominal pain, and unusual bleeding Isturisa (osilodrostat) blocks 11-hydroxylase 2 mg at bedtime, then go up to 2 mg at 8 and 10 pm, can go up to 30 mg Dr. Friedman often gives with spironolactone or ketoconazole. • high testosterone (extra facial hair, acne, hair loss, irregular periods) • low potassium • hypertension Cabergoline (generic, not FDA approved) D2-receptor agonist 0.5 to 7 mg • nausea, • headache • dizziness Korlym (Mifepristone) glucocorticoid receptor antagonist 300-1200 mg per day • cortisol insufficiency (fatigue, nausea, vomiting, arthralgias, and headache) • increased mineralocorticoid effects (hypertension, hypokalemia, and edema • antiprogesterone effects (endometrial thickening) Pasireotide (Signafor) somatostatin receptor ligand 600 μg or 900 μg twice a day Diabetes, hyperglycemia, gallbladder issues For more information or to schedule an appointment with Dr. Friedman, go to goodhormonehealth.com
  6. Abstract Introduction Stress-related brain disorders can be associated with glucocorticoid disturbance and hippocampal alteration. However, it remains largely unknown how cortisol affects the structure and function of hippocampus. Cushing's disease (CD) provides a unique “hyperexpression model” to explore the effects of excessive cortisol on hippocampus as well as the relation between these effects and neuropsychological deficits. Methods We acquired high-resolution T1-weighted and resting-state functional magnetic resonance imaging in 47 CD patients and 53 healthy controls. We obtained the volume and functional connectivity of the hippocampal rostral and caudal subregions in both groups. Relationships between hippocampal alterations, neuroendocrine, and neuropsychological assessments were identified. Results Relative to control subjects, the CD patients had smaller volumes of all four hippocampal subregions. Furthermore, whole brain resting-state functional connectivity analyses with these four different hippocampal regions as seeds revealed altered hippocampal functional connectivity with high-order networks, involving the DMN, frontoparietal, and limbic networks in CD patients. The intrinsic hippocampal functional connectivity was associated with the quality of life of the CD patients. Conclusions Our findings elucidate the cumulative effect of excess cortisol on the morphology and function of hippocampus and reinforce the need for effective interventions in stress-related brain disease to halt potential hippocampal damage. 1 INTRODUCTION Converging evidence has pointed to a strong linkage between the cortisol and human brain and stress-related neuropsychiatry disorders, such as major depression disorder and posttraumatic stress disorder (de Kloet et al., 2005). However, it remains to be established how this stress hormone influences specific brain structures and functions, particularly in humans, which is of particular importance for both treatment of stress-related disorders and research on cortisol effects in the brain. Cushing's disease (CD) is caused by an adrenocorticotropic hormone pituitary adenoma and characterized by chronic hypercortisolism. This condition is therefore a unique and natural “hyperexpression model” to investigate the chronic effects of cortisol on brain physiology and cognition (Zhang et al., 2021). By applying multimodal neuroimaging techniques to CD patients, previous studies have observed that chronic hypercortisolism could cause a number of abnormalities in various brain phenotypes. Among these neural changes of CD patients, hippocampal anomalies are the most replicated findings. Studies on CD patients report hippocampal changes that converge with morphologic alterations such as reduction in volume (Burkhardt et al., 2015; Toffanin et al., 2011). Moreover, abnormal cerebral blood flow and glucose metabolism in hippocampus have also been found in CD patients. Both structural and functional alterations in the hippocampus might contribute to the psychotic symptoms in CD patients (Frimodt-Møller et al., 2019). However, it is well established that psychosis is better described as a brain connectional diaschisis rather than isolated regional dysfunctions (Matthews & Hampshire, 2016). These current hippocampus-related findings were mainly obtained by voxel-based or regional analyses of brain volume or metabolism properties, and researchers have not determined whether the organizational patterns of hippocampal functional connectivity are disrupted in CD patients. The hippocampus is easily targeted by long-term hypercortisolism because this area is a part of the stress response system and is abundant in mineralocorticoid receptors and glucocorticoid receptors (McEwen et al., 2016). Also recently, studies on macaques and humans have observed that hippocampus is an anatomically and functionally heterogeneous region along the rostral/caudal-dorsal/ventral axis (Schultz & Engelhardt, 2014). Specifically, the rostral hippocampus has connections with prefrontal regions and relates to stress, emotion, and affect. In contrast, the caudal hippocampus mainly connects to sensory cortical areas and performs primarily cognitive functions (Fanselow & Dong, 2010). Therefore, the hippocampus should be studied in a set of separate structures with rostral and caudal hippocampus. Whether the hippocampal subregions exhibit differentially altered connectivity patterns responding to chronic hypercortisolism remains largely unknown. The present study further extends this work by examining the relationship between hippocampal subregions and resting-state functional connectivity in large-scale brain networks, as measured by resting-state fMRI (rs-fMRI) (Park & Friston, 2013). We focus on default mode network (DMN), frontoparietal, and limbic networks, given their involvement in stress related psychiatric illnesses. The first is the DMN, which supports self-related cognitive functions. Complementing the DMN is the frontoparietal network, which supports the cognitive regulation of behavior and emotion. Finally, the limbic networks play a key role in emotion regulation. In this study, first, to explore the structural changes of hippocampal subregions in CD patients, we performed a volumetric MRI analysis of the four subregions (left rostral hippocampus, left caudal hippocampus, right rostral hippocampus, and right caudal hippocampus). Given the known direct neurotoxic effects of cortisol on hippocampus, we predicted that chronic hypercortisolism caused smaller hippocampal volumes in CD patients. Second, we used these four subregions as seed regions separately and mapped whole-brain functional connectivity patterns associated with each subregion to examine alterations in hippocampal functional connectivity in CD patients. Considering the psychiatric symptoms in CD patients, it is reasonable to expect the presence of altered hippocampal functional connectivity with high-order networks. 2 MATERIAL AND METHODS 2.1 Participants A total of 47 participants with a diagnosis of CD and 53 healthy control (HC) subjects were included in this study. The CD patients underwent transsphenoidal surgery at the Department of Neurosurgery, The First Medical Center of Chinese People's Liberation Army (PLA) General Hospital between May 2017 and November 2019. According to the clinical practice guideline (Nieman et al., 2015), CD was diagnosed by experienced endocrinologists and confirmed by postsurgical pathology. The detailed preoperative assessments of diagnostic criteria have been reported in our previous study. HCs were recruited from the local community and were controlled for any history of psychopathology abnormalities. All participants were right-handed and had normal vision and auditory sensation. The study was approved by the local ethics committee of the Chinese PLA General Hospital and written informed consent was obtained from each participant. The data of these 47 CD and 53 HC subjects have been partially used in our previous studies (Wang et al., 2019; Zhang et al., 2021). 2.2 Neuroendocrine and neuropsychological assessment All participants underwent biochemical evaluation to assess their cortisol level. We quantified the levels of 24-h urinary free cortisol (24hUFC, nmol/24h); serum cortisol (nmol/L) at 0:00, 8:00, and 16:00. Cortisol was detected with an ADVIA Centaur Analyzer (Siemens Healthcare Diagnostics, Tarrytown, NY, USA). Cortisol levels at 8:00 as well as 24hUFC were also measured in 51 HC subjects. All participants underwent a comprehensive neuropsychological assessment with an expert psychiatrist, including Self-Rating Depression Scale (SDS), Self-Rating Anxiety Scale (SAS), Mini-mental State Examination (MMSE), and Montreal Cognitive Assessment (MoCA). Moreover, health-related quality of life and neuropsychiatric symptoms of CD patients were evaluated with the Cushing's Quality-of-Life (CushingQoL) questionnaire (Nelson et al., 2013) and Chinese version of the neuropsychiatric inventory (CNPI) (Leung et al., 2001), respectively. 2.3 Image acquisition Structural and functional MRI data were acquired on a 3.0-Tesla MR system (Discovery MR750, General Electric) with an 8-channel head coil. High-resolution structural 3D T1-weighted images were conducted using a sagittal Fast Spoiled Gradient-Echo (FSPGR) sequence with the following parameters: repetition time = 6.7 ms, echo time = 2.9 ms, flip angle = 7°, field of view = 250 × 250 mm2, number of slices = 192, voxel size = 1 × 1 × 1 mm3 with no gap. The functional images were acquired using an echo-planar imaging (EPI) sequence with repetition time = 2000 ms, echo time = 30 ms, flip angle = 90°, thickness/gap = 3.5 mm/0.5 mm, slices = 36, field of view = 224 × 224 mm2, voxel size = 3.5 × 3.5 × 3.5 mm3, number of total volumes = 240. Soft earplugs were used to attenuate scanner noise and head motion was restrained with foam padding. During functional scanning, all participants were requested to keep their eyes closed and stay awake. 2.4 rs-fMRI data preprocessing Preprocessing of the rs-fMRI images was conducted using SPM12 and Data Processing Assistant for Resting-State fMRI (DPABI, http://www.restfmri.net/forum/DPARSF). The first 10 volume of the functional images were removed to avoid initial steady-state problems. Then functional images were spatially realigned to the first image for motion correction, and reslicing for acquisition temporal delay. The head motion of all participants in this study had no more than 2-mm translation or 2° rotation in any direction. Next, functional images were coregistered to each participant's segmented gray matter T1 image, and then spatially normalized to the MNI space, resampled to 3-mm isotropic voxels. Subsequently, the global signal, white matter signal, cerebrospinal fluid signal and 24-motion vectors were regressed from the data. Finally, linear detrending and bandpass filter (0.01−0.08 Hz) were carried out to reduce the effects of low-frequency drift and high-frequency physiological noise. 2.5 Hippocampal functional connectivity The hippocampus has been functionally parcellated into four subregions (left rostral hippocampus, left caudal hippocampus, right rostral hippocampus, and right caudal hippocampus) based on Human Brainnetome Atlas (Fan et al., 2016). On each hippocampal subregion, we performed seed-based functional connectivity analysis. Briefly, hippocampal functional connectivity maps were obtained by computing the Pearson correlation coefficient for each voxel's time course with the average time course inside the region of interest. Notably, the computation was constrained within a gray-matter mask which was generated by thresholding (a threshold of 0.2) a prior gray-matter probability map in SPM12. The resulting correlation coefficients were further converted to z scores using Fisher's r-to-z transform to improve normality. For each subject, we obtained 4 z-score maps indicative of the intrinsic functional connectivity patterns of the four hippocampal subregions. To exclude the possible confounding effect of hippocampal volume in CD patients, we performed a voxel-based morphometry analysis on structural MRI images and took the volume of hippocampal subregions as a covariate in the functional connectivity statistical analyses. 2.6 Statistical analysis All demographic and clinical variables including neuroendocrine and neuropsychological scores were compared by two-sample t-tests. Sex composition of the two groups was compared using a Pearson's chi-square test (two-tailed). To explore differences in hippocampal functional connectivity between CD patients and HCs, general linear models were performed in a voxel-wise fashion. To exclude the possible confounding effects of age, gender, education level, and volume of hippocampal subregions, we used these measures as covariates in the general linear models. Multiple comparison correction was performed using a FDR of 0.05 within the grey matter mask. In CD patients group, a linear regression analysis was further performed to explore the relationship between functional connectivity of the clusters showing significant group differences and neuropsychological scores as well as the endocrinological indicators (cortisol and 24hUFC). Multiple comparisons were also corrected using the FDR method with a corrected threshold of q < 0.05. 3 RESULTS 3.1 Demographic, endocrinological, and neuropsychological results Table 1 shows the demographic characteristics of the CD patients and the HCs. There were no significant differences in terms of age, sex distribution, and years of education between groups. Compared with HCs, CD patients had significantly lower MoCA scores and higher SDS and SAS scores (Table 1). As expected, the CD patients had significantly higher levels of serum cortisol and 24hUFC (p < .001). Moreover, we calculated the volumes of the four hippocampal subregions and found smaller volumes of all four hippocampal subregions in the CD patients. TABLE 1. Participant characteristics CDs (n = 47) HCs (n = 53) p Value Age (years) 37.38 ± 10.61 (20–59) 34.79 ± 10.72 (21–63) .113 Gender (male/female) 4/43 4/49 .859 Education (years) 11.00 ± 4.11 11.74 ± 3.10 .311 Illness duration (months) 41.62 ± 53.71 — — Neuropsychological tests MoCA 22.47 ± 3.98 (n = 45) 27.72 ± 2.00 <.001 SDS 40.18 ± 9.96 (n = 45) 27.13 ± 4.42 <.001 SAS 38.27 ± 7.90 (n = 45) 26.98 ± 4.47 <.001 CNPI 11.93 ± 9.68 (n = 45) — — Cushing QOL 37.76 ± 8.29 (n = 45) — — Endocrinological tests Serum cortisol (nmol/L) 0:00 am 633.81 ± 237.59 (n = 46) — — 8:00 am 735.34 ± 279.44 (n = 47) 358.51 ± 107.43 (n = 51) <.001 16:00 pm 671.05 ± 273.56 (n = 47) — — 24hUFC (nmol/24 h) 2381.59 ± 1653.16 (n = 41) 252.03 ± 119.47 (n = 47) <.001 Volume of hippocampal subregions (mm3) Left rostral hippocampus 343.75 ± 39.15 (257.18–423.27) 365.69 ± 27.19 (313.21–442.06) .001 Left caudal hippocampus 272.69 ± 32.74 (206.63–339.04) 296.39 ± 23.13 (249.62–347.61) <.001 Right rostral hippocampus 305.10 ± 33.71 (229.67–396.89) 336.76 ± 25.98 (274.95–415.16) <.001 Right caudal hippocampus 320.42 ± 32.60 (238.16–396.58) 347.87 ± 27.16 (294.00–415.80) <.001 Abbreviations: 24hUFC, 24-h urinary free cortisol.; CDs, Cushing's disease patients; CNPI, Chinese version of neuropsychiatric inventory; Cushing QOL, Cushing Quality of Life Scale; HCs, healthy controls; MoCA, Montreal Cognitive Assessment; SAS, Self-Rating Anxiety Scale; SDS, Self-Rating Depression Scale. Note: All values are expressed as mean ± SD. Group differences in sex between CDs and HCs were examined using chi-square test. Group differences in the other demographic and clinical characteristics between CDs and HCs were examined using two-sample t-tests (two-tailed). 3.2 Spatial distribution of hippocampal functional connectivity The hippocampal functional connectivity maps of both CD patients and HCs are presented in Figure 1. Visually, the spatial distributions of hippocampal functional connectivity were highly similar between groups, in spite of some differences in strength. We observed that the brain regions significantly positively connecting to hippocampus were primarily distributed in several limbic network regions (the orbital frontal cortex, bilateral medial temporal regions, and temporal pole) and DMN regions (bilateral medial frontal cortex, posterior cingulate gyrus/precuneus, and anterior cingulate cortex). Brain regions with negative connectivity to hippocampus were chiefly distributed in the frontoparietal network regions (dorsolateral prefrontal cortex, supramarginal gyrus, and angular gyrus). FIGURE 1 Open in figure viewerPowerPoint Between-group differences in functional connectivity of the hippocampal subregions. The first column shows the hippocampal functional connectivity subregions. The second and third columns show the hippocampal functional connectivity maps within CD and HC groups, respectively. Further between-group comparisons showed that CD patients had significantly altered hippocampal functional connectivities relative to HCs, with a corrected statistical threshold of p < .05. ROI1, left rostral hippocampus; ROI2, left caudal hippocampus; ROI3, right rostral hippocampus; ROI4, right caudal hippocampus; ROI, region of interest; CD, Cushing's disease; HC, healthy control 3.3 Altered hippocampal functional connectivity in CD patients The significant differences in functional connectivity with each hippocampal subregion between the CD patients and HCs groups are illustrated in third column of Figure 1. Both the right and left rostral hippocampus exhibited significantly decreased functional connectivity with the superior parietal lobe (SPL), a component of the frontoparietal network. Moreover, right rostral hippocampus exhibited additional increased functional connectivity with right inferior frontal gyrus, a component of DMN. For the left caudal hippocampus, significantly altered functional connectivity was found to the DMN regions, including (bilateral medial frontal cortex, angular gyrus, anterior, and posterior cingulate cortex). We also observed decreased functional connectivity between the right caudal hippocampus and anterior cingulate cortex. Additionally, the right caudal hippocampus exhibited increased functional connectivity with some limbic regions including the right orbital frontal cortex and temporal pole (Table 2). TABLE 2. Brain regions showing changed RSFC between CDs and HCs groups Peak MNI coordinate Brain regions BA Cluster size (voxels) x y z Peak T ROI-based RSFC ROI1 R IFG 48 219 57 21 —3 4.598 L angular 39 423 −27 −72 51 −5.530 RIO2 R thalamus − 114 9 −6 3 −5.905 L angular 39 195 −27 −72 54 −4.830 R angular 39 384 36 −66 48 −5.607 ROI3 R MTG 20 633 39 6 −21 4.410 L angular 39 195 −27 −72 54 −4.830 R angular 39 384 36 −66 48 −5.607 MFG/ACC 10/32 572 −3 42 −3 −4.033 PCC/PreCUN 26/23 709 12 −45 27 −4.502 ROI4 MFG/ACC 32 465 3 48 6 −4.670 R MTG/OFC 48/21 747 30 3 −21 4.208 Note: Statistical threshold was set at p < .05, corrected. Abbreviations: CDs, Cushing's disease patients; HCs, healthy controls; ROI, regions of interest; BA, Brodmann areas; MNI, Montreal Neurological Institute; RSFC, resting-state functional connectivity; SFG, superior frontal gyrus; MFG, middle frontal gyrus; dMFG, dorsal medial frontal gyrus; IPL, inferior parietal lobule; AG, angular gyrus; ROL, rolandic operculum; Ins, insular; PrCG, precentral gyrus; L, left; R, right; ROI1, left rostral hippocampus; ROI2, left caudal hippocampus; ROI3, right rostral hippocampus; ROI4, right caudal hippocampus. 3.4 Brain–behavior relationships in the CD patients In the correlation analyses of CD patients, the mean values of the functional connectivity between the left caudal hippocampus and anterior cingulate cortex correlated positively with the Cushing's QoL scores (r = .327, p < .05) (Figure 2). No other correlations were found for volumes and functional connectivity of the four hippocampal subregions with neuroendocrine and neuropsychological assessment in the CD patients. FIGURE 2 Open in figure viewerPowerPoint Significant correlations between left hippocampal functional connectivity and the quality of life in CD patients. CD, Cushing's disease; Hip, hippocampus; ACC, anterior cingulate cortex 4 DISCUSSION Using a cohort of CD patients and HCs, the present study performed a comprehensive investigation to reveal how the chronic hypercortisolism affects the morphology and connectivity of hippocampal subregions and their relationships with neuroendocrine and neuropsychological assessment. Compared with the HCs, the CD patients had smaller volumes of all four hippocampal subregions. Furthermore, CD patients exhibited differential patterns of altered hippocampal functional connectivity with high-order networks, involving the DMN, frontoparietal, and limbic networks. The intrinsic hippocampal functional connectivity was associated with the quality of life of the CD patients. Together, these findings elucidate the cumulative effect of cortisol on the morphology and function of hippocampus and provide important information to further understand the role of hippocampus in stress-related brain disease. Cortisol, the end product of the hypothalamic–pituitary–adrenal axis, plays a critical role in the body's response to stress and maintenance of homeostasis (Sapolsky et al., 2000); however, chronic hypercortisolism is known to impair neurons in the hippocampus. CD patients naturally demonstrate chronic excessive amounts of cortisol; therefore these patients serve as a natural “hyperexpression model” to investigate the chronic effects of cortisol on human hippocampus. Importantly, we showed the CD patients are associated with smaller hippocampal volumes in all four subregions. In line with our study, previous structural imaging studies have shown hippocampal volume decreases in CD patients (Frimodt-Møller et al., 2019; Toffanin et al., 2011). Furthermore, Brown et al. found that healthy volunteers were associated with a significant reduction in hippocampal volume following only 3-day stress doses of corticosteroid administration, strongly suggesting the effects of cortisol on hippocampal size. It is important to note that chronic hypercortisolism can affect the hippocampus in at least two ways: by direct neurotoxic effects on the hippocampus (Lupien et al., 2018; Uno et al., 1994) and by reduction in hippocampal neurogenesis (Saaltink & Vreugdenhil, 2014). Moreover, cortisol stimulates the release of excitatory amino acids glutamate on hippocampal cells (de Kloet et al., 2005). On the other hand, chronic elevations of cortisol also reduce neurotrophic factors that includes nerve growth factor and brain-derived neurotrophic factor (McEwen et al., 2015). The different patterns of functional connectivity in rostral hippocampus versus caudal hippocampus might be associated to the specific cytoarchitecture along the rostral/caudal hippocampus. Accumulated evidence from both animal and human studies suggests that different parts of the hippocampus display distinctive gene expression and anatomical projections patterns (Fanselow & Dong, 2010). In detail, gene expression in the rostral hippocampus correlates with regions involved in emotion and stress (amygdala and hypothalamus). Moreover, the rostral hippocampus has connections with prefrontal regions, exerts strong regulatory control of the hypothalamic–pituitary–adrenal axis with a negative feedback (Toffanin et al., 2011). Accordingly, as demonstrated in this study, chronic hypercortisolism predominantly disrupted the functional connectivity in rostral hippocampus. Another major finding in this study was altered hippocampal functional connectivity with DMN, frontoparietal, and limbic networks in CD individuals relative to that in HCs. Emerging evidence proposes that interactions within and between these large-scale brain networks play important roles on brain functions and may be affected in multiple psychiatric disorders (Menon, 2011; Sha et al., 2019). Among these brain networks, the DMN is anchored in the medial prefrontal cortex and posterior cingulate cortex and is implicated in internally directed attention and self-referential processing (Raichle, 2015), while the frontoparietal and limbic networks support the cognitive regulation of emotion, attention, and behavior (Buhle et al., 2014; Kohn et al., 2014). The engagement of these high-level functional networks may suggest the linkage of abnormal stress hormone cortisol to cognitive deficits in CD patients. In line with our study, previous studies have shown stress-induced cortisol increase was associated with altered connectivity within the major brain networks (Zhang et al., 2019, 2020, 2020). Meanwhile, structural and functional alterations in these brain systems are also found in CD patients. For example, many functional imaging studies have consistently demonstrated altered brain activities and functional connectivity involving in DMN, frontoparietal, and limbic networks (Jiang et al., 2017; Wang et al., 2019; Zhang et al., 2021), even in the patients with long-term remission of CD (van der Werff et al., 2015). Importantly, previous studies have shown that the CD patients had widespread reductions of white matter integrity, which provide further evidence for the structural substrate for the persistence of these functional deficits (Pires et al., 2015; van der Werff et al., 2014). Here, we propose that by altering hippocampal processes via the abundant glucocorticoid and mineralocorticoid receptors, exposure to hypercortisolism disrupts the interactions with DMN, frontoparietal, and limbic networks in CD patients, thus engender vulnerability for emotional and cognitive problems. In line with this view is evidence that altered hippocampal functional connectivity is associated with the quality of life in CD patients. Because impaired quality of life is a persistent complaint from CD patients (Webb et al., 2018), it is important to accurately assess which aspects of QoL are affected in order to better understand the severity of hypercortisolism on patients and the potential efficacy of treatment. CushingQoL questionnaire has proven to be a valuable resource for assessing health-related QoL in CD patients, based on the combination of psychosocial issues and physical problems (Nelson et al., 2013). A better understanding of the neuroplasticity and continuing quality of life change may in turn facilitate advances in management and intervention. Several issues need to be addressed further. First, although the sample size of this study was relatively large, the findings still need to be further replicated in an independent sample. Second, the cross-sectional, observational nature of our study design precludes any causal conclusions. Therefore, studies tracking dynamic changes in hippocampal functional connectivity following the remission of hypercortisolism are needed. We are currently following up participants as part of a longitudinal study. Finally, a combined analysis of multimodal imaging including structural and metabolic data would provide integrated information on the effect of cortisol excess on human brain. In short, we demonstrate that CD patients present atypical morphology and functional connectivity of hippocampus. Here we observed the chronic hypercortisolism caused smaller volumes of all hippocampal subregions. This volume change was in line with the preclinical research that excess cortisol cause dendritic shrinkage and loss of spines in the hippocampus. Functionally, CD patients demonstrated altered hippocampal connectivity whose nodes include key components of the DMN, frontoparietal, and limbic networks. These multimodal results reinforce the need for effective therapeutic interventions in stress-related brain disease to halt possible hippocampal damage. ACKNOWLEDGMENTS This study was supported by the National Natural Science Foundation of China (No. 82001798 and No. 81871087), Military Young Scholar Medical Research Fund of Chinese PLA General Hospital (No. QNF19071), and Medical Big Data and Artificial Intelligence Development Fund of Chinese PLA general Hospital (No. 2019MBD-039). CONFLICT OF INTEREST The authors report no biomedical financial interests or potential conflicts of interest. Read more, including references, at https://onlinelibrary.wiley.com/doi/10.1002/brb3.2507
  7. Let’s keep the ball rolling: the more members in the community, the smarter the insights. We're about 1,468 contributors away from being able to tell which treatments were reported to work better for different subgroups with common aspects. You can help get there by simply inviting others! Every invite posted adds about 10 new members to this community. Together, we have the power to build the largest real-world knowledge-base for Cushing's syndrome Take the survey: https://stuff.health/s/uyZ91Lzz
  8. Clinical trial analyses focus on the human body’s homeostatic response to potent HSD-1 inhibition by SPI-62 Results highlight that urinary free cortisol is distinct from intracellular cortisol that causes symptoms in patients with Cushing’s syndrome or autonomous cortisol secretion May 24, 2022 07:20 AM Eastern Daylight Time PORTLAND, Ore.--(BUSINESS WIRE)--Sparrow Pharmaceuticals, an emerging, clinical-stage biopharmaceutical company developing novel, targeted therapies for disorders of glucocorticoid excess, today presented new pharmacological data during a poster session and a Rapid Communications session titled, “HPA axis modulation by a potent inhibitor indicates 11β-hydroxysteroid dehydrogenase type 1 (HSD-1) is a main source of cortisol that can bind intracellular receptors” at the 24th European Congress of Endocrinology (ECE 2022). Sparrow scientists examined the steroid hormone changes after administration of its lead therapeutic candidate, SPI-62, an HSD-1 inhibitor, to healthy adults. “Normalized urinary free cortisol, or UFC, is a standard therapeutic target for patients with Cushing’s syndrome,” said David A. Katz, Ph.D., CSO at Sparrow Pharmaceuticals, “But that biomarker doesn’t measure the cortisol that can access intracellular receptors and cause symptoms. UFC normalization has been shown not to correlate with clinical endpoints in patients with Cushing’s syndrome. Many patients with autonomous cortisol secretion have normal UFC, yet substantial cortisol morbidity. As we conduct clinical trials for patients with those diseases, we’re in search of better ways to measure the cortisol that makes patients ill.” The study analyzed historical clinical trial data to better characterize how SPI-62 impacts cortisol levels and the body’s homeostatic response to those changes. Conclusions of the study include: Half of hepatocellular cortisol with access to intracellular receptors is generated in healthy adults by HSD-1. ACTH increase compensates for the effect of HSD-1 inhibition on systemic cortisol levels. Secondary increases of androgen levels have not been associated to date with clinical consequences. Large changes of the amount of cortisol that can bind intracellular receptors, and thus cause cortisol-related morbidity, can occur independently of urinary free cortisol levels. HSD-1 converts cortisone to cortisol in tissues in which cortisol excess is associated with morbidity including liver, adipose, bone, and brain. SPI-62 is a potent HSD-1 inhibitor in clinical development for treatment of Cushing’s syndrome and autonomous cortisol secretion, and as adjunctive therapy to prednisolone in polymyalgia rheumatica. In Phase 1 clinical trials SPI-62 was generally well tolerated and associated with maximal liver and brain HSD-1 inhibition. To register and view the abstracts, visit ECE’s website here. From https://www.businesswire.com/news/home/20220524005465/en/Sparrow-Pharmaceuticals-Presents-New-Clinical-Trial-Data-Analyses-on-HSD-1-Inhibitor-SPI-62-at-the-24th-European-Congress-of-Endocrinology
  9. Data presented at AACE 2022 detail levoketoconazole-specific effects observed among patients with endogenous Cushing's syndrome from the phase 3 LOGICS trial. New research presented at the American Academy of Clinical Endocrinology (AACE) annual meeting provides insight into the effects of treatment with levoketoconazole (Osilodrostat) among patients with endogenous Cushing’s syndrome. An analysis of data from a double-blind, placebo-controlled, randomized withdrawal study, results of the study demonstrate levoketoconazole provided benefits across a range of etiologies and provide evidence of levoketoconazole-specific effects through the withdrawal and reintroduction of therapy during the trial. “This LOGICS study showed that treatment with levoketoconazole benefitted patients with Cushing’s syndrome of different etiologies and a wide range in UFC elevations at baseline by frequent normalization of mUFC and concurrent improvements in serum cholesterol,” said Maria Fleseriu, MD, professor of medicine and neurological surgery and director of the Northwest Pituitary Center at Oregon Health and Science University, during her presentation. “The benefits observed were established as levoketoconazole-specific via the loss of therapeutic effect upon withdrawal to placebo and restoration upon reintroduction of levoketoconazole.” An orally administered cortisol synthesis inhibitor approved by the US FDA for treatment of endogenous hypercortisolemia in adult patients with Cushing’s syndrome considered ineligible for surgery, levoketoconazole received approval based on results of the phase 3 open-label SONICS trial, which demonstrated . Launched on the heels of SONICS, the current trial, LOGICS, was designed as phase 3, double-blind, placebo-controlled, randomized withdrawal study aimed at assessing the drug-specificity of cortisol normalization in adult patients with Cushing’s syndrome through a comparison of the effects of withdrawing levoketoconazole to placebo against continuing treatment. The trial began with an open-label titration maintenance phase followed by a double-blind randomized withdrawal phase and a subsequent restoration phase, with the randomized withdrawal and restoration phase both lasting 8 weeks. A total of 89 patients with Cushing’s syndrome received levoketoconazole to normalize mUFC. Of these, 39 patients on a stable dose for 4 weeks or more were included in the randomized withdrawal stage of the study. These 39, along with 5 completers of the SONICS trial, were randomized in a 1:1 ratio to continue therapy with levoketoconazole or placebo therapy, with 22 patients randomized to each arm. The primary outcome of interest in the study was the proportion of patients with loss of mean urinary free cortisol response during the randomized withdrawal phase of the study, which was defined as an mUFC 1.5 times the upper limit of normal or greater or an mUFC 40% or more above baseline. Secondary outcomes of interest included mUFC normalization at the end of the randomized withdrawal phase of the study and changes in comorbidity biomarkers. Overall, 21 of the 22 patients randomized to placebo during the withdrawal stage met the primary endpoint of loss of mUFC compared to just 9 of 22 among the levoketoconazole arm of the trial (treatment difference: -54.5% [95% CI, -75.7 to -27.4]; P=.0002). Additionally, at the conclusion of the randomization phase, mUFC normalization was observed among 11 patients in the levoketoconazole arm of the trial compared to 1 patient receiving placebo (treatment difference: 45.5% [95% CI, 19.2 to 67.9]; P=.0015). Further analysis indicated the restoration of levoketoconazole therapy was associated with a. Reversal of loss of contrail control in most patients who had been randomized to placebo. Investigators pointed out the mean change from randomized withdrawal baseline to the end of the randomized withdrawal period in total cholesterol was -0.04 mmol/L for levoketoconazole and 0.9 mmol/L for placebo (P=.0004) and the mean change in LDL-C was -0.006 mmol/L and 0.6 mmol/L, respectively (P=0.0056), with the mean increases in cholesterol observed among the placebo arm reversed during the restoration phase. In safety analyses, results suggest the most commonly reported adverse events seen with levoketoconazole treatment, during all study phases combined were nausea and hypokalemia, which occurred among 29% and 26% of patients, respectively. Investigators also pointed out liver-related events, QT interval prolongation, and adrenal insufficiency, which were respecified adverse events of special interest occurred among 10.7%, 10.7%, and 9.5% of patients receiving levoketoconazole, respectively. This study, “Levoketoconazole in the Treatment of Endogenous Cushing’s Syndrome: A Double-Blind, Placebo-Controlled, Randomized Withdrawal Study,” was presented at AACE 2022. Related Content: American Academy of Clinical EndocrinologyClinicalCushing's Syndrome From https://www.endocrinologynetwork.com/view/no-increased-risk-of-fracture-in-dkd-with-sglt2-inhibitors-vs-dpp-4-inhibitors
  10. Osilodrostat is associated with improvements in physical manifestations of hypercortisolism and reductions in mean body weight and BMI in adults with Cushing’s syndrome, according to a speaker. As Healio previously reported, in findings from the LINC 4 phase 3 trial, osilodrostat (Isturisa, Recordati) normalized mean urinary free cortisol level at 12 weeks in more than 75% of adults with Cushing’s disease. In new findings presented at the AACE Annual Scientific and Clinical Conference, most adults with Cushing’s syndrome participating in the LINC 3 phase 3 trial had improvements in physical manifestations of hypercortisolism 72 weeks after initiating osilodrostat, with more than 50% having no dorsal fat pad, supraclavicular fat pad, facial rubor, proximal muscle atrophy, striae, ecchymoses and hirsutism for women at 72 weeks. Source: Adobe Stock “Many patients with Cushing’s syndrome suffer from clinical manifestations related to hypercortisolism,” Albert M. Pedroncelli, MD, PhD, head of clinical development and medical affairs for Recordati AG in Basel, Switzerland, told Healio. “The treatment with osilodrostat induced a rapid normalization of cortisol secretion, and improvements in physical manifestations associated with hypercortisolism were observed soon after initiation of osilodrostat and were sustained throughout the study.” Albert M. Pedroncelli Pedroncelli and colleagues analyzed changes in the physical manifestations of hypercortisolism in 137 adults with Cushing’s syndrome (median age, 40 years; 77.4% women) assigned osilodrostat. Dose titration took place from baseline to 12 weeks, and therapeutic doses were administered from 12 to 48 weeks, with some participants randomly assigned to withdrawal between 26 and 34 weeks. An extension phase of the trial took place from 48 to 72 weeks. Investigators subjectively rated physical manifestations of hypercortisolism in participants as none, mild, moderate or severe. Participants were evaluated at baseline and 12, 24, 34, 48 and 72 weeks. At baseline, the majority of the study cohort had mild, moderate or severe physical manifestations of hypercortisolism in most individual categories, including dorsal fat pad, central obesity, supraclavicular fat pad, facial rubor, hirsutism in women and striae. Central obesity was the most frequent physical manifestation rated as severe. The percentage of participants with improvements in physical manifestations of hypercortisolism increased from week 12 on for all individual manifestations evaluated in the study, and improvements were maintained through week 72. At 72 weeks, the percentage of participants who had no individual physical manifestations was higher than 50% for each category except central obesity, where 30.6% of participants had no physical manifestations. In addition to improvement in physical manifestations, the study cohort had decreases in body weight, BMI and waist circumference at weeks 48 and 72 compared with baseline. “The main goal of treating patients with Cushing’s syndrome is to normalize cortisol secretion,” Pedroncelli said. “The rapid reduction and normalization of cortisol levels is accompanied by improvement in the associated clinical manifestations. This represents an important objective for patients.” From https://www.healio.com/news/endocrinology/20220512/osilodrostat-improves-physical-manifestations-of-hypercortisolism-for-most-adults
  11. Crinetics Pharmaceuticals, Inc. (Nasdaq: CRNX) today announced positive results from the multiple-ascending dose (MAD) portion of a first-in-human Phase 1 clinical study of CRN04894, the company's first-in-class, investigational, oral, nonpeptide adrenocorticotropic hormone (ACTH) antagonist that is being developed for the treatment of Cushing’s disease, congenital adrenal hyperplasia (CAH) and other conditions of excess ACTH. Following administration of CRN04894, results showed serum cortisol below normal levels and a marked reduction in 24-hour urine free cortisol excretion in the presence of sustained, disease-like ACTH concentrations. “The design of our Phase 1 healthy volunteer study allowed us to demonstrate CRN04894’s potent pharmacologic activity in the presence of ACTH levels that were in similar range to those seen in CAH and Cushing’s disease patients,” said Alan Krasner, M.D., Crinetics’ chief medical officer. “The observation of dose-dependent reductions in serum cortisol levels to below the normal range even in the presence of high ACTH indicates that CRN04894 was effective in blocking the key receptor responsible for regulating cortisol secretion. We believe this is an important finding that may be predictive of CRN04894’s efficacy in patients.” ACTH is the key regulator of the hypothalamic-pituitary adrenal (HPA) axis controlling adrenal activation. It is regulated by cortisol via a negative feedback loop that acts to inhibit ACTH secretion. This feedback loop is dysregulated in diseases of excess ACTH. In Cushing’s disease, a benign pituitary tumor drives excess ACTH secretion even in the presence of excess cortisol. While in CAH, an enzyme deficiency results in excess androgen synthesis without normal cortisol synthesis, allowing unchecked ACTH production and requiring lifelong glucocorticoid use. In both diseases, excess ACTH drives over-stimulation of the adrenal gland and leads to a host of symptoms including infertility, adrenal rest tumors, and metabolic complications in CAH and, in Cushing’s disease, symptoms include hypertension, central obesity, neuropsychiatric disorders and metabolic complications. To our knowledge, no other ACTH antagonists are currently in clinical development for diseases of ACTH excess such as Cushing’s disease or CAH. The 49 healthy adults evaluated in the multiple ascending dose portion of the Phase 1 study were administered 40, 60 or 80 mg doses of CRN04894, or placebo, daily for 10 days. After 10 days of dosing was complete, evaluable participants were administered an ACTH challenge to stimulate adrenal activation to disease relevant levels. Safety and pharmacokinetic data were consistent with expectations from the single-ascending dose cohorts in the Phase 1 study. There were no discontinuations due to treatment-related adverse events and no serious adverse events reported. Glucocorticoid deficiency was the most common treatment-related adverse event in the MAD cohorts. This was an expected extension of pharmacology given the mechanism of action of CRN04894. CRN04894 showed consistent oral bioavailability in the MAD cohorts with a half-life of approximately 24 hours, which is anticipated to support once-daily dosing. Participants in the MAD cohorts who were administered once nightly CRN04894 experienced a dose-dependent suppression of adrenal function as measured by suppression of serum cortisol production of 17%, 29% and 37% on average from baseline over 24 hours for the 40, 60 or 80 mg dosing groups respectively, (despite requirement for glucocorticoid supplementation in some of these subjects to prevent clinical adrenal insufficiency), compared to an average 2% increase in serum cortisol for individuals receiving placebo. The strong, dose-dependent suppression of serum and urine free cortisol was achieved despite ACTH levels in subjects in the 60 and 80 mg cohorts similar to those typically seen in patients with CAH and Cushing’s disease. Even when an additional exogenous ACTH challenge was administered on top of the already increased ACTH levels, cortisol levels remained below the normal range in subjects receiving CRN04894, indicating clinically significant suppression of adrenal activity. “Due to its central position in HPA axis, ACTH is the obvious target for inhibiting excessive stimulation of the adrenal in diseases of ACTH excess. Even though the field of endocrinology has known about its clinical significance for more than 100 years, we are not aware of any other ACTH antagonist that has entered clinical development. This is an important milestone for endocrinology and for our company.” said Scott Struthers, Ph.D., founder and chief executive officer of Crinetics. “We are very excited to initiate patient studies in Cushing’s disease and CAH with CRN04894, which will be our third home-grown NCE to demonstrate pharmacologic proof-of-concept and enter patient trials.” Crinetics plans to present additional details of safety, efficacy, and biomarker results from the CRN04894 Phase 1 study at an endocrinology-focused medical meeting in 2022. Data Review Conference Call Crinetics will hold a conference call and live audio webcast today, May 25, 2022, at 8:00 a.m. Eastern Time to discuss results from the MAD cohorts of the Phase 1 study of CRN04894. To participate, please dial 1-877-407-0789 (domestic) or 1-201-689-8562 (international) and refer to conference ID 13730000. To access the webcast, click here. Following the live event, a replay will be available on the Events page of the Company’s website. About the CRN04894 Phase 1 Study Crinetics has completed enrollment of the 88 healthy volunteers in this double-blind, randomized, placebo-controlled Phase 1 study. Participants were divided into multiple cohorts in the single ascending dose (n=39) and multiple ascending dose (n=49) portions of the study. In both the SAD and MAD portions of the study, safety and pharmacokinetics were assessed. In addition, pharmacodynamic responses were evaluated before and after challenges with injected synthetic ACTH to assess pharmacologic effects resulting from exposure to CRN04894. From https://www.streetinsider.com/Corporate+News/Crinetics+Pharmaceuticals+(CRNX)+Reports+Positive+Top-line+Results+Including+Strong+Adrenal+Suppression+from+CRN04894+Phase+1+Study/20126484.html
  12. I'm not around much right now - I have the other dred C-disease :(

  13. Published: May 15, 2022 (see history) DOI: 10.7759/cureus.25015 Cite this article as: Iturregui J, Shi G (May 15, 2022) Recurrent Metatarsal Fractures in a Patient With Cushing Disease: A Case Report. Cureus 14(5): e25015. doi:10.7759/cureus.25015 Abstract Cushing syndrome (CS) can result from excess exposure to exogenous or endogenous glucocorticoids. The most common endogenous cause of CS is an adrenocorticotropic hormone (ACTH)-secreting pituitary adenoma, known as Cushing disease (CD). Patients typically present with characteristics including truncal obesity, moon facies, facial plethora, proximal muscle weakness, easy bruising, and striae. Insufficiency fractures of the metatarsals are a rare presentation for CS. A 39-year-old premenopausal woman presented to the orthopedic outpatient clinic with recurrent metatarsal fractures and no history of trauma. A metabolic bone disease was suspected, and after further evaluation by endocrinology services, the CD was diagnosed. Surgical resection was performed, and pathology confirmed the presence of a pituitary adenoma. Multiple, recurrent, non-traumatic metatarsal fractures can be the initial presentation of CD in a premenopausal woman. Introduction Cushing syndrome (CS) is a rare clinical and metabolic disorder caused by excessive exposure to glucocorticoids. In the United States, an estimated 10 to 15 people per million population are affected by CS each year, while studies in Europe report an incidence of 0.7 to 2.4 per million people affected annually [1,2]. Furthermore, CS more commonly affects women [2]. Common characteristics of CS include truncal obesity, moon facies, proximal muscle weakness, fatigue, facial plethora, supraclavicular fullness, peripheral edema, weight gain, striae, easy bruising, acne, hirsutism, amenorrhea, dorsocervical "buffalo" hump, depression, hypertension, impaired glucose tolerance, and osteoporosis [1,3,4]. The most common cause of CS is exogenous glucocorticoid therapy. Meanwhile, endogenous cortisol hypersecretion commonly results from either an adrenocorticotropic hormone (ACTH)-secreting pituitary adenoma or a cortisol-secreting adrenal tumor. When CS is caused by a pituitary adenoma, this is referred to as Cushing disease (CD). CD is the most common endogenous cause of CS, accounting for 80-85% of cases [1,5]. Whether a patient’s CS is caused by exogenous or endogenous sources, excessive exposure to steroids can have deleterious effects on the bones, resulting in secondary osteoporosis. The decrease in bone mass and microarchitectural changes increase the risk of fragility fractures, with reported rates as high as 30-67% [6]. The most commonly reported fracture site in CS patients is the vertebrae; however, other reported fracture sites include the ribs, sternum, wrist, elbow, shoulder, pelvis, hip, femoral condyles, tibia, fibula, calcaneus, metatarsals, and phalanges [4,6-16]. There are reports of metatarsal fractures occurring in patients diagnosed with endogenous CS [3,6,7,16-19]. However, to the best of our knowledge, there are no reports of multiple, recurrent, bilateral metatarsal fractures as the initial presentation in a pre-menopausal woman with CD. Here, we present a case of a premenopausal woman with recurrent metatarsal stress fractures who was diagnosed with CD after further evaluation. Case Presentation A 39-year-old premenopausal woman was evaluated by her primary care physician due to right foot pain after feeling a pop while walking. She reported swelling and some bruising along the lateral aspect of her foot. Her exercise regimen consisted of walking twice a week for 30 minutes at each session. She did not report any traumatic injuries to her foot. Imaging revealed a fifth metatarsal fracture (Figure 1). The patient was placed in a cast walker boot and referred to orthopedics for further evaluation. Orthopedic management included no weight bearing on her right foot and continuing using the cast walker boot or a postop shoe, with reevaluation in four weeks. Figure 1: Oblique radiograph of the right foot demonstrating a mildly displaced transverse fracture of the proximal fifth metatarsal (arrow). At the time of evaluation, the patient was 161.5 cm tall, weighed 101 kg, and had a BMI of 38.86 kg/m2. Her medical history included hypertension, hyperglycemia, hyperlipidemia, hypothyroidism, obesity, anxiety, obstructive sleep apnea, and colon polyps. The patient reported a history of metatarsal fractures in her left foot in 2008, which healed slowly and without surgical intervention. She also underwent bunion and bunionette surgery on her left foot. Her medications included alprazolam, levothyroxine, lisinopril, bimatoprost, ergocalciferol, meloxicam, and ondansetron. She was a former smoker (2007-2010), a daily wine drinker, and had an active job working as a nurse. Her family history included lung cancer and alcohol abuse in her father; hypertension, hypothyroidism, and alcohol abuse in her mother; and osteoporosis and end-stage renal disease secondary to polycystic kidney disease in her sister. At the three-month follow-up visit, the fracture line remained clearly visible, and minimal callus had formed at the fracture site. Surgical fixation was recommended and performed four months after the fracture occurred. Six months after her right foot's fifth metatarsal fracture, she developed new-onset swelling and tenderness over the middle metatarsals dorsally in her right foot with no history of trauma. Radiographs demonstrated new second and third metatarsal neck fractures (Figure 2). Conservative management with a postop shoe for six weeks and re-evaluation was recommended. In the interim between her initial right foot fifth metatarsal fracture and the new right foot second and third metatarsal fractures, the patient was diagnosed with diabetes mellitus type II, treated with a plant-based diet, hospitalized for urolithiasis, and diagnosed with depression. She was started on bupropion. Figure 2: Anteroposterior radiograph of bilateral feet demonstrating second and third metatarsal neck fractures of the right foot (arrows). Due to the recurrent metatarsal stress fractures with no associated trauma, the patient was referred to endocrinology for workup of metabolic bone disease. Her physical exam revealed no abnormalities, and her overall workup was negative. Bone mineral density results demonstrated osteopenia in the lumbar spine (T-score: -1.8) and left femoral neck (T-score: -1.0), and normal bone density in the left total hip (T-score: -0.80). Six months following her right foot's second and third metatarsal fractures, the patient developed right great toe and second toe swelling and bruising. Two months later, after trying supportive tennis shoes and reducing weightbearing on her right foot, she did not notice any improvement and sought orthopedic care. Radiographs revealed a new subacute fracture of the right second proximal phalanx (Figure 3). A magnetic resonance imaging (MRI) scan was ordered, which revealed a first metatarsal shaft stress fracture as well (Figure 4). She underwent conservative management with a Cam walker boot and was referred to endocrinology for re-evaluation for suspected endocrinopathy. Figure 3: AP radiograph of bilateral feet demonstrating a subacute fracture of the second proximal phalanx of the right foot (arrow). Figure 4: T1-weighted sagittal MRI of the right foot demonstrating a first metatarsal shaft stress fracture (arrow). At her endocrinology visit, a physical exam revealed some facial hair, frontal hair loss, and a significant dorsocervical and anterior cervical fat pad. A Cushingoid face shape, facial redness, acne, oligomenorrhea, incremental weight gain over the last decade, centripetal adiposity, easy bruising, and lower leg swelling were also reported. Bone mineral density results reported spine and hip Z-scores within the expected range for age, indicating no osteoporosis. Since she had features of hypercortisolism, labs to evaluate for Cushing syndrome were ordered. The 11:00 pm salivary cortisol levels were elevated to 173 ng/dL and 168 ng/dL in two samples. The 1 mg dexamethasone suppression test failed to suppress her cortisol levels, with an elevated cortisol value of 29 mcg/dL. The 24-hour urine-free cortisol level was elevated at 135 mcg/24 hours. These lab results confirmed a diagnosis of Cushing syndrome. Her ACTH was elevated at 86 pg/mL, which indicated an ACTH-dependent CS. Pituitary MRI demonstrated a 1.1 cm × 1.5 cm × 1.1 cm pituitary lesion, representing a pituitary macroadenoma (Figure 5). The patient underwent endoscopic endonasal transsphenoidal pituitary tumor resection with the goal of treating her Cushing disease and preventing further fragility fractures. Pathology evaluation confirmed a pituitary adenoma. Figure 5: T1-weighted coronal MRI of the pituitary demonstrating a 1.1 cm × 1.5 cm × 1.1 cm cystic sellar mass which represents a pituitary macroadenoma (arrow). Discussion This is a case of a 39-year-old woman who presented with recurrent metatarsal fractures with no history of trauma, raising suspicion of a metabolic bone disease. The patient also developed centripetal weight gain, glucose intolerance, kidney stones, depression/anxiety, and Cushingoid features. A laboratory workup performed by endocrinology services confirmed a diagnosis of ACTH-dependent CS. An MRI revealed a pituitary lesion which represented a pituitary macroadenoma, for which surgical resection was performed. Pathology confirmed a pituitary adenoma. The association of multiple, non-traumatic metatarsal fractures occurring in premenopausal women with endogenous CS has been reported in the literature [3,7,19]. However, to the best of our knowledge, this is the first report presenting a premenopausal woman with multiple, recurrent metatarsal fractures as the initial manifestation of CD. Several mechanisms play a role in glucocorticoid-induced bone loss, which is more prominent in trabecular bone compared to cortical bone [3,4,6,8]. Normally, trabecular bone has a greater bone turnover rate than cortical bone. In the presence of excess glucocorticoids, trabecular bone has greater sensitivity to glucocorticoids and undergoes slower bone turnover. The most significant effects of excess glucocorticoids on bones are decreased osteoblast function and quantity, which explain the reduced trabecular bone turnover rate [4,10]. The proposed mechanisms for this are glucocorticoid-induced inhibition of osteoblast proliferation and genesis, as well as induction of osteoblast and osteocyte apoptosis [4,10,11]. Furthermore, glucocorticoids decrease bone protein synthesis (e.g., osteocalcin), type I collagen formation, and alkaline phosphatase activity [4]. Additional effects include greater bone resorption, inhibition of intestinal calcium absorption, inhibition of renal calcium reabsorption, and decreased secretion of gonadal steroids and growth hormones [8]. Glucocorticoids also induce protein catabolism, which can result in muscle weakness, decreased bone stimulation from weakened muscle contraction, and further bone loss and debility [4]. Multiple fragility fractures in the foot with no history of trauma or overuse are uncommon. When evaluating a patient with this presentation, secondary causes for these fractures need to be investigated. Differential diagnoses include osteoporosis, Charcot foot, multiple myeloma, celiac disease, avascular necrosis, and endocrine disorders such as hyperthyroidism, primary hyperparathyroidism, or CS, among others [3,6,7]. There is a high rate of fragility fractures due to secondary osteoporosis in CS patients, with the vertebrae being most commonly affected [6]. LiYeung and Lui [7] and Albon et al. [19] each reported a case of a pre-menopausal woman who initially presented with multiple metatarsal fractures secondary to an adrenal adenoma causing CS. In each case, the patient’s densitometry indicated osteoporosis. However, in our case and the case reported by Molnar et al. [3] of a pre-menopausal woman with multiple fractures due to CD (recurrent fractures were not reported), the bone densitometries performed did not indicate osteoporosis. The patients reported by LiYeung and Lui [7], Albon et al. [19], and Molnar et al. [3] did not demonstrate marked clinical characteristics of CS. In comparison to our patient, she did have multiple Cushingoid features upon her second evaluation by endocrinology. Furthermore, in all our cases, the patients were first evaluated for metatarsal fractures as the initial presentation, which resulted in a diagnosis of endogenous CS after further evaluation. Finally, early recognition and treatment of CS are important, as there is an increased risk of morbidity and mortality as the condition progresses [20]. In addition, the treatment of CS can reverse the bone loss that occurs with excess glucocorticoid exposure [4,10]. This case also highlights the importance of collaboration between physicians in the different branches of medicine. Conclusions Excess glucocorticoid exposure can have deleterious effects on the bones, increasing the risk for secondary osteoporosis and fragility fractures. There needs to be an index of suspicion for metabolic bone disease, including endogenous CS caused by CD, as the underlying etiology of multiple, recurrent, atraumatic metatarsal fractures in pre-menopausal women. Early diagnosis and management of CD can lower the risk of morbidity and mortality as well as reverse bone loss. References Guaraldi F, Salvatori R: Cushing syndrome: maybe not so uncommon of an endocrine disease. J Am Board Fam Med. 2012, 25:199-208. 10.3122/jabfm.2012.02.110227 Valassi E, Santos A, Yaneva M, et al.: The European Registry on Cushing's syndrome: 2-year experience. Baseline demographic and clinical characteristics. Eur J Endocrinol. 2011, 165:383-92. 10.1530/EJE-11-0272 Molnar V, Zekan P, Dušek T, Ivković A: Multiple metatarsal fractures: the first manifestation of Cushing’s disease—a case report. J Am Podiatr Med Assoc. 2021, 111:10.7547/19-024 Han JY, Lee J, Kim GE, et al.: A case of cushing syndrome diagnosed by recurrent pathologic fractures in a young woman. J Bone Metab. 2012, 19:153-8. 10.11005/jbm.2012.19.2.153 Barahona MJ, Sucunza N, Resmini E, et al.: Deleterious effects of glucocorticoid replacement on bone in women after long-term remission of Cushing's syndrome. J Bone Miner Res. 2009, 24:1841-6. 10.1359/jbmr.090505 Papadakis G, Uebelhart B, Goumaz M, Zawadynski S, Rizzoli R: An unusual case of hypercortisolism with multiple weight-bearing bone fractures. Clin Cases Miner Bone Metab. 2014, 10:213-7. LiYeung LL, Lui TH: Bilateral adrenal adenoma presented as multiple metatarsal and phalangeal fractures. J Orthop Case Rep. 2015, 5:77-8. 10.13107/jocr.2250-0685.353 Trementino L, Appolloni G, Ceccoli L, Marcelli G, Concettoni C, Boscaro M, Arnaldi G: Bone complications in patients with Cushing's syndrome: looking for clinical, biochemical, and genetic determinants. Osteoporos Int. 2014, 25:913-21. 10.1007/s00198-013-2520-5 Abdel-Kader N, Cardiel MH, Navarro Compan V, Piedra Priego J, González A: Cushing's disease as a cause of severe osteoporosis: a clinical challenge. Reumatol Clin. 2012, 8:278-9. 10.1016/j.reuma.2011.11.004 Lee HJ, Je JH, Seo JH, Na YJ, Yoo HJ: Multiple spontaneous rib fractures in patient with Cushing’s syndrome. J Bone Metab. 2014, 21:277-82. 10.11005/jbm.2014.21.4.277 Poonuru S, Findling JW, Shaker JL: Lower extremity insufficiency fractures: an underappreciated manifestation of endogenous Cushing's syndrome. Osteoporos Int. 2016, 27:3645-9. 10.1007/s00198-016-3712-6 Belaya ZE, Hans D, Rozhinskaya LY, et al.: The risk factors for fractures and trabecular bone-score value in patients with endogenous Cushing's syndrome. Arch Osteoporos. 2015, 10:44. 10.1007/s11657-015-0244-1 Tajika T, Shinozaki T, Watanabe H, Yangawa T, Takagishi K: Case report of a Cushing's syndrome patient with multiple pathologic fractures during pregnancy. J Orthop Sci. 2002, 7:498-500. 10.1007/s007760200087 Baron E, Sheinfeld M, Migdal EA, Hardoff R: Multiple pathologic fractures mimicking bone metastases in a patient with Cushing's syndrome. Clin Nucl Med. 1996, 21:506-8. 10.1097/00003072-199606000-00027 Bosch S, Bogaerts S: Pituitary adenoma presenting with bilateral calcaneal stress fracture: a case report. JOSPT Cases. 2021, 1:109-111. Kostoglou-Athanassiou I, Spiliotis G, Athanassiou L, Myriokefalitakis I: Cushing’s syndrome in a patient with systemic lupus erythematosus. Endocrine Abstracts. 2018, 56:106. 10.1530/endoabs.56.P106 Kaur K, Findling JW: Cushing’s disease. A Case-Based Guide to Clinical Endocrinology. Davies TF (ed): Humana Press, Totowa; 2008. 27-33. 10.1007/978-1-60327-103-5_3 Ontell FK, Shelton DK: Multiple stress fractures. An unusual presentation of Cushing's disease. West J Med. 1995, 162:364-6. Albon L, Rippin J, Franklyn J: “My feet are killing me!” An unusual presentation of Cushing’s syndrome. Endocrine Abstracts. 2003, 5:26. Nieman LK: Recent updates on the diagnosis and management of Cushing’s syndrome. Endocrinol Metab (Seoul). 2018, 33:139-46. 10.3803/EnM.2018.33.2.139 From https://www.cureus.com/articles/91295-recurrent-metatarsal-fractures-in-a-patient-with-cushing-disease-a-case-report
  14. — More than half of patients saw physical manifestations fully resolve by week 72 by Kristen Monaco, Staff Writer, MedPage Today May 16, 2022 SAN DIEGO -- Osilodrostat (Isturisa) improved many physical features associated with Cushing's disease, according to additional findings from the phase III LINC-3 study. Among 137 adults with Cushing's disease, a 39.5% improvement in central obesity scores was observed from baseline to week 72 with osilodrostat, reported Alberto Pedroncelli, MD, PhD, of Recordati AG in Basel, Switzerland. Not only was central obesity the most common physical manifestation associated with hypercortisolism among these Cushing's disease patients, but it was also more frequently rated as severe at baseline, Pedroncelli explained during the American Association of Clinical Endocrinology (AACE) annual meeting. Osilodrostat treatment also led to a 34.9% improvement in proximal muscle atrophy at week 72, along with a 34.4% improvement in hirsutism scores. By week 72, nearly all physical manifestations of hypercortisolism saw significant improvement -- marked by more than 50% of patients scoring these physical traits as nonexistent: Dorsal fat pat: 50.6% Central obesity: 30.6% Supraclavicular fat pad: 51.8% Facial rubor: 64.7% Hirsutism in women: 53.1% Proximal muscle atrophy: 61.2% Striae: 63.5% Ecchymoses: 87.1% Most of these physical manifestation improvements were notable soon after treatment initiation with osilodrostat, Pedroncelli pointed out. When stratified according to testosterone levels, hirsutism scores remained either stable or improved in the majority of patients who had normal or above normal testosterone levels. More women with normal testosterone levels over time experienced improvements in hirsutism versus those with levels above the upper limit of normal, who mostly remained stable. Osilodrostat is an oral agent that was first FDA approved in March 2020 for adults with Cushing's disease who either cannot undergo pituitary gland surgery or have undergone the surgery but still have the disease. Available in 1 mg, 5 mg, and 10 mg film-coated tablets, the drug acts as a potent oral 11-beta-hydroxylase inhibitor -- the enzyme involved in the last step of cortisol synthesis. Osilodrostat is taken orally twice daily, once in the morning and once in the evening. Approval was based upon findings from the LINC-3 and LINC-4 trials, which found osilodrostat was able to normalize cortisol levels in 53% of patients, based on mean 24-hour urinary free cortisol (UFC) concentrations. During an initial 10-week randomization phase, 86% of patients maintained their complete cortisol response if they remained on osilodrostat versus only 29% of those who were switched to placebo. As expected, 77.4% of the 137 adults included in the trial were women. The median participant age was 40 and about 47 months had passed since their initial diagnosis. A total of 87.6% underwent previous pituitary surgery and 16.1% underwent previous pituitary irradiation. At baseline, median and mean 24-hour UFC levels were 3.5 nmol and 7.3 nmol, respectively, based on two or three urine samples. Participants had an average body weight of 176.4 lb, body mass index (BMI) of 30, and 41 in waist circumference at baseline. Throughout the trial, all measures dropped, reaching the nadir at week 72: body weight of 165 lb, BMI of 27, and 37.8 in waist circumference. The most common side effects reported with the agent include adrenal insufficiency, fatigue, nausea, headache, and edema. Kristen Monaco is a staff writer, focusing on endocrinology, psychiatry, and nephrology news. Based out of the New York City office, she’s worked at the company since 2015. Disclosures The study was supported by Recordati AG. Pedroncelli reported employment with Recordati. Primary Source American Association of Clinical Endocrinology Source Reference: Pedroncelli AM, et al "Osilodrostat therapy improves physical features associated with hypercortisolism in patients with Cushing's disease: findings from the phase III LINC 3 study" AACE 2022. From https://www.medpagetoday.com/meetingcoverage/aace/98745
  15. Published: May 15, 2022 (see history) DOI: 10.7759/cureus.25017 Cite this article as: Fernandez C, Bhatia S, Rucker A, et al. (May 15, 2022) Intermittent Blurry Vision: An Unexpected Presentation of Cushing’s Syndrome Due to Primary Bilateral Macronodular Adrenal Hyperplasia (PBMAH). Cureus 14(5): e25017. doi:10.7759/cureus.25017 Abstract Cushing’s syndrome (CS) is an uncommon endocrine disorder resulting from prolonged exposure to elevated glucocorticoids, with 10-15 million annual cases per the American Association of Neurological Surgeons. Exogenous and endogenous causes can further be divided into adrenocorticotropic hormone (ACTH) dependent (i.e Cushing’s Disease) or ACTH independent. ACTH-independent CS can be caused by primary bilateral macronodular adrenal hyperplasia (PBMAH) representing less than 1% cases of CS. We report a case of a woman presenting with chronic resistant hypertension, episodic blurry vision, weight gain and wasting of extremities. She was diagnosed with Cushing’s syndrome due to PBMAH. Our patient’s presentation was unusual as she presented at 40 years old, 10 years earlier than expected for PBMAH; and primarily with complaints of episodic blurry vision. Her symptoms also progressed rapidly as signs and symptoms largely presented over the course of 12 months, however responded well to surgical resection. Introduction Cushing’s syndrome (CS) is an uncommon endocrine disorder caused by prolonged exposure to elevated glucocorticoids [1]. There are exogenous or endogenous causes. The National Institute of Health’s (NIH) Genetic and Rare Diseases Information Center (GARD) estimated the prevalence of endogenous CS to be 1 in 26,000 [2]. According to a large study, the annual incidence of CS in individuals less than 65 years old was nearly 49 cases per million [3]. Cushing’s disease (CD), which is defined as Cushing’s syndrome caused by an adrenocorticotropic hormone (ACTH)-secreting pituitary tumor, accounts for approximately 80% of patients with CS; whereas ACTH-independent CS accounts for the remaining 20% [4]. Among the causes of pituitary ACTH-independent CS is bilateral macronodular adrenal hyperplasia which is rare, comprising less than 1% of patients with CS [5]. Herein is a case of rapid onset Cushing’s syndrome due to PBMAH initially presenting as episodes of bilateral blurry vision. Case Presentation The patient is a 40-year-old female with a past medical history of resistant hypertension (on four agents), and recently diagnosed type 2 diabetes mellitus (started on insulin regimen). Patient was recently seen by her primary care provider, with complaints of intermittent episodes of blurry vision going on for months. As part of evaluation in December 2020, the patient underwent a renal ultrasound as part of evaluation by the primary physician for uncontrolled hypertension. The doppler incidentally showed an indeterminate hypoechoic mass on the right kidney and presumably located within the right adrenal gland, measuring 3.4 x 5.4 cm, without sonographic evidence of renal artery stenosis. The left kidney appeared normal. She was recommended to have further evaluation with contrast enhanced MR or CT with adrenal protocol. In January 2021, the patient was sent from her PCP’s office to the ED as the patient was having blurred vision. She had a plain CT scan of the brain that was unremarkable. The patient's systolic blood pressure was in the 160s-170s mm Hg upon arrival to ED compliance with home medications of 5mg of amlodipine daily, 25mg of metoprolol succinate daily, 100mg of losartan daily, and 25mg of hydrochlorothiazide daily. Physical exam reported obesity without evidence of abdominal striae. Blood work in the ED showed elevated blood glucose level over 600 (mg/dL) despite being on a regimen of lantus 60 units, metformin 1000mg twice a day, and semaglutide SQ weekly. Hemoglobin A1c was greater than 15.5%, and vitamin D was low (15.6 ng/mL). The morning ACTH was low (<5pg/mL) (nAM levels: 7.2 - 63.3 pg/mL), AM cortisol was high at 26.1 ug/ml (normal: 5.0 - 23.0 ug/mL), plasma aldosterone was normal at 4.2 ng/dL with a normal plasma renin at 1.96 (0.25 - 5.82 ng/mL/h). 24-hour urine free cortisol (UFC) was high at 1299.5 (4.0-50.0 mcg/24h). CT of the abdomen/pelvis with and without contrast showed low-attenuation masses (less than 5 Hounsfield units) present in both adrenal glands measuring 6.9 x 5.3 cm on the right and 4.5 x 3.9 cm on the left, and did not demonstrate significant arterial enhancement (Figure 1). MR imaging of the abdomen without and with contrast was also obtained and showed the same masses of the bilateral adrenal glands, with largest on the left measured 3.6 cm and largest on the right measured 3.7 cm, as well as mild fatty infiltration of the liver. General surgery and hematology/oncology were consulted and recommendations were made for outpatient follow-up with PCP and endocrinology. Figure 1: CT of the abdomen/pelvis with contrast showing low-attenuation masses present in both adrenal glands measuring 6.9 x 5.3 cm on the right (dark gray arrow) and 4.5 x 3.9 cm on the left (light gray arrow) In early February 2021, the patient again presented to the ED complaining of recurrent episodes of bilateral blurry vision. Examination was unremarkable, including an ophthalmological exam with slit lamp exam. Blurred vision was suspected to be due to osmotic swelling in the setting of severe hyperglycemia as the patient had persistently uncontrolled blood sugars. Recommendations were for tighter control of blood glucose, and follow-up with primary care and ophthalmology. Patient followed up with the endocrinologist in mid-February to which the patient reported first noticing a difference in her energy and changes to her weight around one year prior. She communicated a weight gain of 30 to 40 lbs over the past year. Patient had a reported history of gestational hypertension diagnosed five years ago when she gave birth to her daughter, which was steadily worsening over the past year. She reported intermittent myalgias and easy bruising. Patient had no family history or any apparent features to suggest multiple endocrine neoplasia (MEN) syndrome. Blood work revealed ACTH less than 1.5 pg/mL, AM cortisol was high at 24.5 mcg/dL, and normal aldosterone at 3.6 ng/dL, with normal renin and metanephrine levels. Physical examination revealed truncal obesity as well as a round face, cushingoid in appearance, and relatively thin extremities and abdominal striae. She was then referred to a surgical specialist, and it was decided that she would undergo laparoscopic bilateral adrenalectomy due to severe Cushing’s syndrome. The surgical pathology report revealed macro-nodular cortical hyperplasia of both left and right adrenal gland masses with random endocrine atypia. The largest nodule on the left measured 4.5 cm and the largest nodule on the right measured 6.6 cm. Post-operatively she was started on hydrocortisone 20 mg every morning and 10 mg every evening, and fludrocortisone 0.1 mg twice a day as part of her steroid replacement regimen. Eventually she changed to hydrocortisone 10 mg three times a day and fludrocortisone 0.1 mg once a day. For her diabetes, her insulin glargine decreased from 60 units to 20 units. Amlodipine and hydrochlorothiazide were discontinued from her antihypertensive medications; she continued losartan and metoprolol. Follow up blood work showed stable electrolytes with potassium 4.2 mmol/L (3.5-5.2 mmol/L), sodium 137 mmol/L (134-144mmol/L), chloride 100 mmol/L (96-106 mmol/L), and carbon dioxide 23 mmol/L (20-29mmol/L). Discussion ACTH-independent Cushing’s syndrome due to bilateral cortisol-secreting nodules is rare, accounting for 2% of CS cases. The majority of causes include primary bilateral macronodular adrenal hyperplasia (PBMAH), primary pigmented nodular adrenocortical disease (PPNAD), and bilateral adrenocortical adenomas (BAA). In PBMAH, typically patients are diagnosed within the fifth or sixth decade of life [4]. The usual age of onset for PPNAD is within the first to third decade of life, with median age in the pediatric population at age 15 years [6]. BAA is such a rare entity that there exists little epidemiological data with less than 40 reported cases until 2019 [7]. A small subset of patients present with overt clinical symptoms of CS, as hypercortisolism often follows an insidious course that can delay diagnosis from years to decades, with one series reporting a diagnostic delay of approximately eight years [8]. Serum and urine hormone screening in the right clinical setting can provide clues to these endocrine disorders, however diagnosis of ACTH-independent CS often occurs incidentally wherein a radiographic study was done for reasons other than to identify adrenal disease [9]. CT or MRI alone are not able to differentiate these disease entities, requiring pathological examination for final determination [7]. Adrenal venous sampling (AVS) and I-6B-iodomethyl-19-norcholesterol (I-NP-59) can aid in identifying hormone-secreting status of each adrenal lesion, however usefulness is debated among experts [10-12]. In all cases the end goal is to normalize adrenocortical hormones, and PBMAH primarily involves surgical resection with exogenous hormone replacement. Bilateral adrenalectomy is generally the treatment of choice with overt Cushing syndrome regardless of cortisol level. These patients require lifelong steroid administration [9,13]. Another approach is unilateral adrenalectomy of the larger or more metabolically active gland, which can be identified after AVS or I-NP-59 testing. This has been proposed in order to preserve some autonomous hormonal production and prevent adrenal crisis, however remission rates of Cushing syndrome as high as 84% have been reported with eventual need for bilateral adrenalectomy [7,8,14]. Steroid enzyme inhibition to control cortisol secretion has been used as an adjunct before surgery. In some patients with identified aberrant adrenal hormone receptors, targeted pharmacological inhibition remains an alternative medical approach [8]. Despite these alternatives to surgery, surgical resection remains the optimal approach [1]. Conclusions ACTH-independent Cushing’s syndrome due to PBMAH usually presents as an indolent course, with typical diagnosis in the fifth to sixth decade. As the use of imaging for other non-endocrine related investigations becomes more utilized, PBMAH being less of a rare entity. Clinical presentation usually dictates the timing of and type of surgical intervention. Although there are some reports of unilateral resection resulting in a cure, many of these cases eventually proceed to staged bilateral resection. Our patient’s presentation as her primary complaint was recurrent episodes of blurry vision that were suspected to be due to osmotic swelling because of her uncontrolled hyperglycemia. Her case was also unusual as she presented at 40 years old, an average of 10 years earlier than is typically diagnosed for PBMAH. Her symptoms also progressed rapidly over the course of 12 months with development of resistant hypertension and insulin-dependent diabetes requiring high basal insulin. Following surgical resection, her antihypertensive regimen was de-escalated and had significant reduction in insulin requirements, and was maintained on adrenocorticoid therapy. References Nieman LK: Recent updates on the diagnosis and management of Cushing's syndrome. Endocrinol Metab (Seoul). 2018, 33:139-46. 10.3803/EnM.2018.33.2.139 Rare Disease Database: Cushing Syndrome. (2021). Accessed: 12/17/2021: https://rarediseases.org/rare-diseases/cushing-syndrome/. Broder MS, Neary MP, Chang E, Cherepanov D, Ludlam WH: Incidence of Cushing's syndrome and Cushing's disease in commercially-insured patients <65 years old in the United States. Pituitary. 2015, 18:283-9. 10.1007/s11102-014-0569-6 Lacroix A, Feelders RA, Stratakis CA, Nieman LK: Cushing’s syndrome. Lancet. 2015, 386:913-27. 10.1016/S0140-6736(14)61375-1 Tokumoto M, Onoda N, Tauchi Y, et al.: A case of adrenocoricotrophic hormone -independent bilateral adrenocortical macronodular hyperplasia concomitant with primary aldosteronism. BMC Surg. 2017, 17:97. 10.1186/s12893-017-0293-z Stratakis CA: Cushing syndrome caused by adrenocortical tumors and hyperplasias (corticotropin- independent Cushing syndrome). Endocr Dev. 2008, 13:117-32. 10.1159/000134829 Gu YL, Gu WJ, Dou JT, et al.: Bilateral adrenocortical adenomas causing adrenocorticotropic hormone-independent Cushing's syndrome: a case report and review of the literature. World J Clin Cases. 2019, 7:961-71. 10.12998/wjcc.v7.i8.961 Lacroix A: ACTH-independent macronodular adrenal hyperplasia. Best Pract Res Clin Endocrinol Metab. 2009, 23:245-59. 10.1016/j.beem.2008.10.011 Sweeney AT, Srivoleti P, Blake MA: Management of the patient with incidental bilateral adrenal nodules. J Clin Transl Endocrinol Case Rep. 2021, 20:100082. 10.1016/j.jecr.2021.100082 Lumachi F, Zucchetta P, Marzola MC, Bui F, Casarrubea G, Angelini F, Favia G: Usefulness of CT scan, MRI and radiocholesterol scintigraphy for adrenal imaging in Cushing's syndrome. Nucl Med Commun. 2002, 23:469-73. 10.1097/00006231-200205000-00007 Builes-Montaño CE, Villa-Franco CA, Román-Gonzalez A, Velez-Hoyos A, Echeverri-Isaza S: Adrenal venous sampling in a patient with adrenal Cushing syndrome. Colomb Med (Cali). 2015, 46:84-7. Guo YW, Hwu CM, Won JG, Chu CH, Lin LY: A case of adrenal Cushing's syndrome with bilateral adrenal masses. Endocrinol Diabetes Metab Case Rep. 2016, 2016:150118. 10.1530/EDM-15-0118 Wei J, Li S, Liu Q, et al.: ACTH-independent Cushing's syndrome with bilateral cortisol-secreting adrenal adenomas: a case report and review of literatures. BMC Endocr Disord. 2018, 18:22. 10.1186/s12902-018-0250-6 Osswald A, Quinkler M, Di Dalmazi G, et al.: Long-term outcome of primary bilateral macronodular adrenocortical hyperplasia after unilateral adrenalectomy. J Clin Endocrinol Metab. 2019, 104:2985-93. 10.1210/jc.2018-02204 From https://www.cureus.com/articles/90069-intermittent-blurry-vision-an-unexpected-presentation-of-cushings-syndrome-due-to-primary-bilateral-macronodular-adrenal-hyperplasia-pbmah
  16. Abstract Corticotroph cells give rise to aggressive and rare pituitary neoplasms comprising ACTH-producing adenomas resulting in Cushing disease (CD), clinically silent ACTH adenomas (SCA), Crooke cell adenomas (CCA) and ACTH-producing carcinomas (CA). The molecular pathogenesis of these tumors is still poorly understood. To better understand the genomic landscape of all the lesions of the corticotroph lineage, we sequenced the whole exome of three SCA, one CCA, four ACTH-secreting PA causing CD, one corticotrophinoma occurring in a CD patient who developed Nelson syndrome after adrenalectomy and one patient with an ACTH-producing CA. The ACTH-producing CA was the lesion with the highest number of single nucleotide variants (SNV) in genes such as USP8, TP53, AURKA, EGFR, HSD3B1 and CDKN1A. The USP8 variant was found only in the ACTH-CA and in the corticotrophinoma occurring in a patient with Nelson syndrome. In CCA, SNV in TP53, EGFR, HSD3B1 and CDKN1A SNV were present. HSD3B1 and CDKN1A SNVs were present in all three SCA, whereas in two of these tumors SNV in TP53, AURKA and EGFR were found. None of the analyzed tumors showed SNV in USP48, BRAF, BRG1 or CABLES1. The amplification of 17q12 was found in all tumors, except for the ACTH-producing carcinoma. The four clinically functioning ACTH adenomas and the ACTH-CA shared the amplification of 10q11.22 and showed more copy-number variation (CNV) gains and single-nucleotide variations than the nonfunctioning tumors. Keywords: corticotroph; Cushing disease; ACTH-secreting carcinoma; single nucleotide variation; copy number variation; exome 1. Introduction The pathological spectrum of the corticotroph includes ACTH (adrenocorticotropic hormone)-secreting pituitary adenomas (PA), causing Cushing disease (CD), silent corticotroph adenomas (SCA), Crooke cell adenomas (CCA) and the rare ACTH-secreting carcinoma (ACTH-CA). Pituitary carcinomas account for 0.1 to 0.2% of all pituitary tumors and are defined by the presence of craniospinal or distant metastasis [1,2,3]. Most pituitary carcinomas are of corticotroph or lactotrope differentiation [3]. Although a few cases present initially as CA, the majority develop over the course of several months or years from apparently benign lesions [3,4]. CCA are characterized by the presence of hyaline material in more than 50% of the cells of the lesion, and most of them arise from silent corticotroph adenomas (SCA) or CD-provoking ACTH-secreting adenomas [5]. SCA are pituitary tumors with positive immunostaining for ACTH but are not associated with clinical or biochemical evidence of cortisol excess; they are frequently invasive lesions and represent up to 19% of clinically non-functioning pituitary adenomas (NFPA) [6]. ACTH-secreting PA represents up to 6% of all pituitary tumors and causes eloquent Cushing disease (CD), which is characterized by symptoms and signs of cortisol hypersecretion, including a two- to fivefold increase in mortality [7,8]. The 2017 World Health Organization (WHO) classification of PA considers not only the hormones these tumors synthesize but also the transcription factors that determine their cell lineage [9]. TBX19 is the transcription factor responsible for the terminal differentiation of corticotrophs [9]. All tumor lesions of corticotroph differentiation are positive for both ACTH and TBX19. ACTH-secreting PA causing CD are among the best genetically characterized pituitary tumors, with USP8 somatic variants occurring in up to 25–35% of sporadic cases [9]. Yet, information regarding the molecular pathogenesis of the lesions conforming to the whole pathological spectrum of the corticotroph is scarce. The aim of the present study is to characterize the genomic landscape of pituitary tumors of corticotroph lineage. For this purpose, we performed whole exome sequencing to uncover the mutational burden (single-nucleotide variants, SNV) and copy-number variations (CNVs) of these lesions. 2. Results 2.1. Clinical and Demographic Characteristics of the Patients A total of 10 tumor samples from 10 patients were evaluated: 4 ACTH-secreting adenomas causing clinically evident CD, three non-functioning adenomas that proved to be SCA upon immunohistochemistry (IHC), one ACTH-secreting CA with a prepontine metastasis, one rapidly growing ACTH-secreting adenoma after bilateral adrenalectomy (Nelson syndrome) in a patient with CD and one non-functioning, ACTH-producing CCA (Table 1). All except one patient were female; the mean age was 38.8 ± 16.5 years (range 17–61) (Table 1). They all harbored macroadenomas with a mean maximum diameter of 31.9 ± 13 mm (range 18–51). Cavernous sinus invasion was evident on MRI in all but one of the patients (Table 1). Homonymous hemianopia was present in seven patients, whereas right optic nerve atrophy and amaurosis were evident in patient with the ACTH-CA, and in patient with CD and pituitary apoplexy (Table 1). Detailed clinical data are included in Supplementary Table S1. Death was documented in only the patient with pituitary apoplexy, and one patient was lost during follow-up, as of October 2018. Table 1. Clinical features of the tumors analyzed and SNV present in each tumor. 2.2. General Genomic Characteristics of Neoplasms of Corticotrophic Lineage Overall, approximately 18,000 variants were found, including missense, nonsense and splice-site variants as well as frameshift insertions and deletions. Of these alterations, the majority corresponded to single-nucleotide variants, followed by insertions and deletions. The three most common base changes were transitions C > T, T > C and C > G; most of the genetic changes were base transitions rather than transversions (Figure 1). There were several genes across the whole genome affected in more than one way, meaning that the same gene presented missense and nonsense variants, insertions, deletions and splice-site variants (Figure 2). Many of these variants are of unknown pathogenicity and require further investigation. Gains in genetic material were found in 44 cytogenetic regions, whereas 72 cytogenetic regions showed loss of genetic material in all corticotroph tumors. Figure 1. Panel (A) shows the gadolinium-enhanced magnetic resonance imaging of the patient with ACTH-CA, highlighting in red the metastatic lesion in the prepontine area. Panel (B) shows the hematoxylin and eosin staining displaying the hyaline structures in the perinuclear areas denoting a Crooke cell adenoma. Panel (C,D) depict a representative corticotroph tumor with positive ACTH and TBX19 immunohistochemistry, respectively. Panel (E) shows four graphics: variant classification, variant type, SNV class and transition (ti) or transversion (tv) describing the general results of exome sequencing of the corticotroph tumors. Figure 2. Representative rainfall plots showing the SNV alterations throughout the whole genome of corticotroph tumors (A) CCA, (B) SCA, (C) CD and (D) ACTH-CA, displaying all base changes, including transversions and transitions. No kataegis events were found. Alterations across the genome were seen in all corticotroph tumors. 2.3. ACTH-Secreting Carcinoma (Tumor 1) SNV missense variants were found in the genes encoding TP53 (c.215G > C [rs1042522], p.Pro72Arg); AURKA (c.91T > A [rs2273535], p.Phe31Ile); EGFR (epidermal growth factor receptor, c.1562G > A [rs2227983], p.Arg521Lys); HSD3B1 (3-ß-hydroxisteroid dehydrogenase, c.1100C > A [rs1047303], p.Thr367Asn); CDKN1A (cyclin-dependent kinase inhibitor 1A or p21, c.93C > A [rs1801270], p.Ser31Arg); and USP8 (c.2159C > G [rs672601311], p.Pro720Arg). Interestingly, the previously reported USP48, BRAF, BRG1 and CABLES1 variants in pituitary CA cases were not found in this patient’s tumor (Figure 3). All SNV detected in WES experiments were validated by Sanger sequencing. The variants described were selected due to their potential pathogenic participation in other tumors and the allelic-risk association with tumorigenesis. Hereafter, all the mentioned variants in other corticotroph tumors are referred to by these aforementioned variants. Even though these same genes presented other variants, currently the significance of those variants is unknown. Figure 3. Panel (A) shows the oncoplot from the missense variants of the selected genes and their clinical–pathological features. Panels (B–G) depict USP8, EGFR, TP53, AURKA, CDKN1A and HSD3B1 proteins, respectively, with the changes found in DNA impacting aminoacidic changes. In general, the pituitary CA presented more CNV alterations than the benign tumors, with 27 and 32 cytogenetic regions showing gains and losses of genetic material, respectively. The cytogenetic regions showing gains were 10q11.22, 15q11.2, 16p12.3, 1p13.2 and 20p, where genes SYT15, POTEB, ARL6IP1, HIPK1 and CJD6 are coded, respectively. By contrast, 8p21.2 was the cytogenetic region showing loss of genetic material. The previously reported amplification of 1p13.2 was also detected in this tumor (Figure 4) [10]. Figure 4. Hierarchical clustering of corticotroph tumors according to their gains and losses across the whole genome (somatic chromosomes only). High contrast was used to enhance potential CNV alterations; nevertheless, there were only 44 unique cytogenetic regions that showed gains in genetic material with statistical significance, whereas only 72 unique cytogenetic regions showed loss of genetic material with statistical significance. 2.4. Crooke Cell Adenoma (Tumor 2) The CCA showed SNV in the genes encoding TP53, EGFR, HSD3B1 and CDKN1A. However, neither the genes encoding AURKA and USP8 nor those encoding USP48, BRAF, BRG1 and CABLES were affected in this tumor. In CCA, only two and fifteen gains and losses were observed in copy-number variation, respectively. CNVs only showed gains in cytogenetic regions 17q12 and 10q11.22, harboring genes CCL3L1 and NPY4R, respectively, whereas losses were found in cytogenetic regions 18q21.1, 15q12 and 2q11.2, harboring genes KATNAL2, TUBGCP5 and ANKRD36. 2.5. Silent Corticotroph Adenomas (Tumors 3–5) The three SCA shared SNVs in the genes encoding HSD3B1 and CDKN1A. SCA 4 and 5 showed SNV in the genes encoding EGFR, whereas SNV in the genes encoding AURKA and TP53 were present in SCA 3 and 5. None of the SCA were found to have SNV in the genes encoding USP8, USP48, BRAF, BRG1 or CABLES1. The SCA presented only two and eighteen gains and losses (CNV), respectively. In regard to CNV, the these clinically silent tumors presented gains of genetic material in cytogenetic regions 17q22 and 10q11.22, which harbor genes encoding CCL3L1 and NPY4R. Eighteen losses were found distributed in cytogenetic regions 18q21.1, 15q12 and 2q11.2, encompassing the genes encoding KATNAL2, TUBGCP5 and ANKRD36. This CNV pattern closely resembles the one found in the CCA, which is somewhat expected if we consider that both neoplasms are clinically non-functioning 2.6. ACTH-Secreting Adenomas (Cushing Disease) (Tumors 6–9) SNV of the genes encoding TP53 and HSD3B1 were present in tumor samples from all four CD patients, whereas none of these patients harbored adenomas with SNV in the genes encoding USP8 or CDKN1A. An SNV in the gene encoding AURKA was identified in only one of these tumors (tumor 8). EGFR SNV were found in tumors 7 and 9. None of the CD-causing ACTH-secreting adenomas showed the previously reported SNV in the genes encoding USP48, BRAF, BRG1 and CABLES1. CNV analysis in this group of eloquent-area corticotroph tumors revealed 25 gains and 55 losses of genetic material. The gains occurred in cytogenetic regions 17q12, 2p12, 9p24 and 10q11.22, where genes CCL3L1, CTNNA2, FOXD4 and NPY4R are coded, respectively. The losses were localized in cytogenetic regions 21p12, 15q11.2, and 8p23, harboring genes USP16, KLF13 and DEF130A, respectively. We also detected the previously reported 20p13 amplification [10]. 2.7. ACTH-Secreting Adenoma Causing Nelson Syndrome (Tumor 10) This patient’s tumor showed SNV in the genes encoding USP8, TP53, HSD3B1 and CDKN1A but no alterations were found in the genes encoding EGFR and AURKA. This tumor and the ACTH-CA were the only two neoplasms that harbored a USP8 variant. No SNV were identified in the genes encoding USP48, BRAF, BRG1 and CABLES1. Interestingly, CNV analysis revealed the same gains and losses of genetic material found in tumors from other patients with CD. 2.8. Tumor Phylogenic Analysis We performed a phylogenetic inference analysis to unravel a hypothetical sequential step transformation from an SCA to a functioning ACTH-secreting adenoma and finally to an ACTH-CA. The theoretical evolutive development of the ACTH CA, departing from the SCA, shows two main clades, with the smallest one comprising two of the three SCA and two of the five ACTH-adenomas causing CD. Since these four tumors have the same SNV profile, we can assume that they harbor the genes that must be altered to make possible the transition from a silent to a clinically eloquent adenoma; the gene encoding ATF7IP (c.1589A > G [rs3213764], p.K529R) characterizes this clade. The second and largest clade includes the CCA, the ACTH-CA, one of the three SCA and three of the five most aggressive ACTH adenomas causing CD, including the adenoma of the patient with Nelson syndrome. This clade represents the molecular alterations required to evolve from a CD-causing ACTH-adenoma to a more aggressive tumor, or even to a CA and is characterized by the gene encoding MSH3 (c.235A > G [rs1650697], p.I79V) (Figure 5). Figure 5. Phylogenetic analysis of the corticotroph tumors. The theoretical evolutive development of the ACTH-CA, departing from the SCA shows two main clades. The first clade, characterized by ATF7IP gene, comprises 2 of the 3 SCA and 2 of the 5 ACTH-adenomas causing CD. The second clade is characterized by the gene encoding MSH3 and includes the CCA, the ACTH-CA, one of the 3 SCA and 3 of the 5 most aggressive ACTH adenomas causing CD, including the adenoma of the patient with Nelson syndrome. Red dots represent the Cushing Disease provoking adenomas, green dots represent the silent corticotroph tumors, brown dot represent the Crooke cell adenoma and the blue dot represent the corticotroph carcinoma. 2.9. Correlation between Gene Variants and Clinicopathological Features The USP8 variant positively correlated with increased tumor mass (p = 0.019). The CDKN1A variant was significantly associated with silent tumors (p = 0.036). The rest of the genetic variants did not correlate with any of the clinicopathological features tested. The presence of the EGFR variant was not distinctly associated with any of the clinical parameters and was equally present in functional as well as non-functional tumors (p = 0.392). AURKA SNV did not correlate with any of the features, including recurrence (p = 0.524). Detailed statistical results are presented in Supplementary Table S2. 3. Discussion Corticotrophs are highly specialized cells of the anterior pituitary that synthesize and secrete hormones that are essential for the maintenance of homeostasis. In this study, we sequenced the exome of 10 corticotroph tumors, including three SCA, four ACTH adenomas causing CD, an ACTH adenoma in a patient with Nelson syndrome, a CCA and an ACTH-CA in total, representing the broad pathological spectrum of this cell. Our results portray the genomic landscape of all the neoplasms that are known to affect the corticotroph. The neoplasm with the highest number of genomic abnormalities, including SNV and CNV, was the ACTH-CA, followed by the CCA and the CD tissues. Of all the genes harboring SNVs, six were found to be present in at least two of our tumor samples: HSD3B1, TP53, CDKN1A, EGFR, AURKA and USP8. The HSD3B1 gene encodes a rate-limiting enzyme required for all pathways of dihydrotestosterone synthesis and is abundantly expressed in adrenal tumors. Gain of function of this HSD3B1 variant, which has a global allelic prevalence of 0.69678 [11], results in resistance to proteasomal degradation with the consequent accumulation of the enzyme and has been associated with a poor prognosis in patients with prostate cancer [12]. Nine of the ten corticotroph tumors in our cohort harbored an SNV of the tumor suppressor gene TP53. The TP53 variant described in our cohort has been reported to be present in 80% of non-functioning pituitary adenomas and is apparently associated with a younger age at presentation and with cavernous sinus invasion [13]. Furthermore, this TP53 variant results in a reduced expression of CDKN1A and an increased expression of vascular endothelial growth factor (VEGF) as well as an increased cellular proliferation rate [13]. CDKN1A (also known as p21) is a cyclin-dependent kinase inhibitor regulating cell cycle progression. The SNV described in our study was reported to alter DNA binding ability and expression and has a global allelic frequency of 0.086945 [14]. This cyclin-dependent kinase inhibitor SNV was found to be associated with breast carcinoma [15] and lung cancer [16]. The presence of this SNV has not been previously explored in pituitary adenomas, although CDKN1A is downregulated in clinically non-functioning pituitary adenomas of gonadotrophic lineage but not in hormone-secreting tumors [17]. EGFR encodes a transmembrane tyrosine kinase receptor, activation of which leads to mitogenic signaling [18]. This gene is upregulated in several cancers and represents a target for molecular therapies [19]. The EGFR SNV described in our corticotroph tumor series was found to be associated with the response to neoadjuvant chemotherapy in patients with breast and lung cancer [18]. EGFR is normally expressed in corticotrophs, where it participates in the regulation of POMC (proopiomelanocortin) gene transcription and cellular proliferation [20]. The EGFR rs2227983 has a 0.264334 global allelic frequency [21]. AURKA is a cell-cycle regulatory serine/threonine kinase that promotes cell cycle progression by the establishment of the mitotic spindle and centrosome separation [22]. Alterations of these gene are related to centrosomal amplification, dysfunction of cytokinesis and aneuploidy [22]; it has a global allelic frequency of 0.18078 [23]. This same SNV has been associated with overall cancer risk, particularly breast, gastric, colorectal, liver and endometrial carcinomas, but it has never been formally studied in pituitary tumors [22]. Activating somatic variants of the gene encoding USP8 were recently found in 25–40% of ACTH-secreting adenomas causing CD [24,25]. Patients harboring these variants are usually younger, more frequently females and were found to have higher long-term recurrence rates in some but not all studies [26,27]. USP8 mediates the deubiquitination of EGFR by inhibiting its interaction with protein 14-3-3, which in turn prevents its proteosomal degradation. Signaling through the recycled deubiquitinated EGFR is increased, leading to increased POMC transcription and cellular proliferation. Most activating USP8 variants are located within its 14-3-3 binding motif [24,25]. Recently, USP8 and TP53 SNV were described in corticotroph tumors as drivers of aggressive lesions [28]. To our knowledge, USP8 variants have not been evaluated in patients with pituitary carcinomas, and none of the previously mentioned studies have included patients with Nelson syndrome. In our cohort, neither the CCA nor the SCA showed variants in USP8, in concordance with previously published studies [25,29], or in the genes USP48, BRAF, BRG1 and CABLES1 [9], and none of them were present in our cohort. Genetic structural variations in the human genome can be present in many forms, from SNV to large chromosomal aberrance [30]. CNV are structurally variant regions, including unbalanced deletions, duplications and amplifications of DNA segments ranging from a dozen to several hundred base pairs, in which copy-number differences have been observed between two or more genomes [31,32]. CNV are involved in the development and progression of many tumors and occur frequently in PA [30,33]. Hormone-secreting pituitary tumors show more CNV than non-functioning tumors [34]. Accordingly, our non-functioning SCA and CCA had considerably fewer chromosomal gains and losses than the CD-causing adenomas and the ACTH-CA. Expectedly, the ACTH-CA had significantly more cytogenetic abnormalities than any other tumor in our series. Interestingly, the ACTH-adenomas causing CD, the SCA and the CCA shared the gain of genetic material in 17q12, highlighting their benign nature. The 17q12 amplification has been described in gastric neoplasms [35]. The only cytogenetic abnormality shared by all types of corticotroph tumors was the gain of genetic material in 10q11.22. Amplification of 10q11.22 was previously described in Li–Fraumeni cancer predisposition syndrome [36]. The ACTH-CA, the CCA and one SCA clustered together showing a related CNV pattern; this CNV profile could be reflective of the aggressive nature of these neoplasms, since both CCA and SCA can follow a clinically aggressive course [5,6]. Our results show that all lesions conforming to the pathological spectrum of the corticotroph share some of the SNV and CNV profiles. These genomic changes are consistent with the potential existence of a continuum, whereby silent tumors can transform into a clinically eloquent tumor and finally to carcinoma, or at least a more aggressive tumor. It can also be interpreted as the common SNV shared by aggressive tumors. It is known that silent corticotroph adenomas may switch into a hormone-secreting tumor [37] and are considered a marker for aggressiveness and a risk factor for malignancy since most of the carcinomas are derived from functioning hormone-secreting adenomas. Our phylogenetic inference analysis showed that the genes ATF7IP and MSH3 could participate in a tumor transition ending in aggressive entities or even carcinomas. ATF7IP is a multifunctional nuclear protein mediating heterochromatin formation and gene regulation in several contexts [38], while MSH3 is a mismatch-repair gene [39]. Events related to heterochromatin remodeling and maintenance have been related to aggressive pituitary adenomas and carcinomas [40]. Additionally, alterations in mismatch-repair genes are related to pituitary tumor aggressiveness and resistance to pharmacologic treatment [41,42]. The variants described in ATF7IP and MSH3 are related to prostate and colorectal cancer, respectively [43,44]. There is evidence suggesting that the ATF7IP variant could be deleterious because it leads to a negative regulation of transcription [45]. Thus, these events could be biologically relevant to corticotroph tumorigenesis, although more research is needed. 4. Conclusions We have shown genomic evidence that within the tumoral spectrum of the corticotroph, functioning ACTH-secreting lesions harbor more SNV and CNV than non-functioning ACTH adenomas. The ACTH-secreting CA shows more genomic abnormalities than the other lesions, underscoring its more aggressive biological behavior. Phylogenetic inference analysis of our data reveals that silent corticotroph lesions may transform into functioning tumors, or at least potentially, into more aggressive lesions. Alterations in genes ATF7IP and MSH3, related to heterochromatin formation and mismatch repair, could be important in corticotroph tumorigenesis. The main drawback of our study is the limited sample size. We are currently increasing the number of samples to corroborate our findings and to be able to perform a more comprehensive complementary phylogenetic analysis of our data. Finally, further research is needed to uncover the roles of these variants in corticotroph tumorigenesis. 5. Materials and Methods 5.1. Patients and Tumor Tissue Samples Ten pituitary tissues were collected: one ACTH-CA, one CCA, three SCA, and five ACTH-secreting PA causing CD, including the tumor of a patient who developed Nelson syndrome after bilateral adrenalectomy. All tumors included in the study were sporadic and were collected from patients diagnosed, treated and followed at the Endocrinology Service and the Neurosurgical department of Hospital de Especialidades, Centro Médico Nacional Siglo XXI of the Instituto Mexicano del Seguro Social, Hospital General de Mexico “Dr. Eduardo Liceaga” and Instituto Nacional de Neurologia y Neurocirugia “Manuel Velazquez”. All participating patients were recruited with signed informed consent and ethical approval from the Comisión Nacional de Ética e Investigación Científica of the Instituto Mexicano del Seguro Social, in accordance with the Helsinki declaration. CD was diagnosed according to our standard protocol. Briefly, the presence of hypercortisolism was documented based on two screening tests, namely a 24 h urinary free-cortisol level above 130 µg and the lack of suppression of morning (7:00–8:00) cortisol after administration of 1 mg dexamethasone the night before (23:00) to less than 1.8 µg/dL, followed by a normal or elevated plasma ACTH to ascertain ACTH-dependence. Finally, an overnight, high-dose (8 mg) dexamethasone test, considered indicative of a pituitary source, and a cortisol suppression > 69%, provided that a pituitary adenoma was clearly present on magnetic resonance imaging (MRI) of the sellar region. In none of the 10 patients included in the study was inferior petrosal venous sampling necessary to confirm the pituitary origin of the ACTH excess. Invasiveness was defined by the presence of tumor within the cavernous sinuses (CS). DNA was extracted from paraffin-embedded tumor tissues using the QIAamp DNA FFPE tissue kit. From frozen tumors, DNA was obtained using the Proteinase K-ammonium acetate protocol. 5.2. Construction and Sequencing of Whole Exome Libraries Exome libraries were prepared according to the Agilent SureSelect XT HS Human All exon v7 instructions. Briefly, 200 ng of DNA was enzymatically fragmented with Agilent SureSelect Enzymatic Fragmentation Kit. Fragmented DNA was end-repaired and dA-tail was added at DNA ends; then, molecular barcode adaptors were added, followed by AMPure XP bead purification. The adaptor-ligated library was amplified by PCR and purified by AMPure XP beads. DNA libraries were hybridized with targeting exon probes and purified with streptavidin-coated magnetic beads. The retrieved libraries were amplified by PCR and purified by AMPure XP beads and pooled for sequencing in NextSeq 500 using Illumina flow cell High Output 300 cycles chemistry. All quality controls of the libraries were carried out using Screen tape assays and quantified by Qubit fluorometer. Quality parameters included a DNA integrity number above 8 and a 100X sequencing depth aimed with at least 85% of coverage. 5.3. Bioinformatics Analysis The fastq files were subjected to quality control using FastQC v0.11.9, the adapters were removed using Cutadapt v3.4, the alignment was carried out with Burrows–Wheeler Alignment Tool v0.7.17 with the -M option to ensure compatibility with Picard and GRCh38 as a reference genome. The marking of duplicates as well as the sorting was carried out with Picard v2.26.4 with the AddOrReplaceReadGroups programs with the option SORT_ORDER = coordinate and MarkDuplicates, respectively. Variant calling was carried out using Genomic Analysis Toolkit (GATK) v4.2.2.0 following the Best Practices guide (available at https://gatk.broadinstitute.org/) [46] and with the parameters used by Genomic Data Commons (GDC), available at https://docs.gdc.cancer.gov/ [47]. The GATK tools used were CollectSequencingArtifactMetrics, GetPileupSummaries, CalculateContamination and Mutect2. Mutect2 was run with the latest filtering recommendations, including a Panel of Normal and a Germline Reference from the GATK database. Filtering was performed with the CalculateContamination, LearnReadOrientationModel and FilterMutectCalls tools with the default parameters. For the calculation of CNV GISTIC v2.0.23 was used with the parameters used by GDC. Catalog of Somatic Mutation in Cancer (COSMIC) was used to uncover pathogenic variants. For the analysis of variants and CNV, the maftool v2.10.0 and ComplexHeatmap 2.10.0 packages were used. All analyses were carried out on the GNU/Linux operating system under Ubuntu v20.01.3 or using the R v4.0.2 language in Rstudio v2021.09.0+351. A second bioinformatics pipeline was also used, SureCall software (Agilent) with the default parameters used for SNV variant calling. The variants found by both algorithms were taken as reliable SNV. Data were deposited in Sequence Read Archive hosted by National Center for Biotechnology Information under accession number PRJNA806516. Phylogenetic tree inference (PTI) was run by means of the default parameters using matrices for each sample. These matrices contain an identifier for each variant, mutant read counts, counts of reference reads and the gene associated with the variant. The only PTI parameter was Allele Frequency of Mutation and was used to improve the speed of the algorithm. Briefly, PTI uses an iterative process on the variants shared between the samples. First, it builds the base of the tree using the variants shared by all the samples; second, it eliminates these variants and establishes a split node; and third, it eliminates the variants of the sample that produced the division (split). PTI iteratively performs these three steps for all division possibilities. Each tree is given a score based on an aggregated variant count, and the tree with the highest score is chosen as the optimal tree. 5.4. Sanger Sequencing forConfirmation of Exome Findings Exome variant findings in exome sequencing were validated by Sanger sequencing using BigDye Terminator v3.1 Cycle Sequencing kit (ThermoFischer) in a 3500 Genetic Analyzer. Primers used for USP8 [48], TP53 [49], EGFR [50], AURKA [51], CDKN1A [52,53] and HSD3B1 sequencing have been previously reported. 5.5. Hormone and Transcription Factor Immunohistochemistry Paraffin-embedded, formalin-fixed tissue blocks were stained with hematoxylin–eosin and reviewed by a pathologist. Tumors were represented with a 2-fold redundancy. Sections (3 μm) were cut and placed onto coated slides. Immunostaining was performed by means of the HiDef detection HRP polymer system (Cell Marque, CA, USA), using specific antibodies against each pituitary hormone (TSH, GH, PRL, FSH, LH and ACTH) and the lineage-specific transcription factors TBX19, POU1F1 and NR5A1, as previously described [54]. Two independent observers performed assessment of hormones and transcription factors expression at different times. 5.6. Statistical Analysis Two-tailed Fisher exact tests and Student’s t tests were used to evaluate the relationship between the identified gene variants and clinicopathological features. A p value of <0.05 was considered statistically significant. Statistical software consisted of SPSS v28.0.1 Supplementary Materials The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/ijms23094861/s1. Author Contributions D.M.-R., K.T.-P. and M.M. conceived, designed and coordinated the project, performed experiments, analyzed, discussed data and prepared the manuscript. S.A.-E., G.S.-R., E.P.-M., S.V.-P., R.S., L.B.-A., C.G.-T., J.G.-C. and J.T.A.-S. performed DNA purification, library preparation, sequencing experiments, bioinformatics analysis and wrote the manuscript. A.-L.E.-d.-l.-M., I.R.-S., E.G.-A., L.A.P.-O., G.G., S.M.-J., L.C.-M., B.L.-F. and A.B.-L. provided biological samples and detailed patient information. All authors have read and agreed to the published version of the manuscript. Funding This work was partially supported by grants 289499 from Fondos Sectoriales Consejo Nacional de Ciencia y Tecnologia, Mexico, and R-2015-785-015 from Instituto Mexicano del Seguro Social (MM). Institutional Review Board Statement Protocol approved by the Comisión Nacional de Ética e Investigación Científica of the Instituto Mexicano del Seguro Social, in accordance with the Helsinki declaration (R-2019-785-052). Informed Consent Statement Informed consent was obtained from all subjects involved in the study. Data Availability Statement Data were deposited in Sequence Read Archive hosted by National Center for Biotechnology Information under accession number PRJNA806516. Acknowledgments Sergio Andonegui-Elguera is a doctoral student from Programa de Doctorado en Ciencias Biomédicas, Universidad Nacional Autónoma de México (UNAM) and received fellowship 921084 from CONACYT. KTP is a recipient of Consejo Nacional de Ciencia y Tecnología (CONACyT) fellowship “Estáncias posdoctorales por Mexico 2021” program. DMR is a recipient of the National Council for Science and Technology Fellowship “Catedra CONACyT” program. Conflicts of Interest The authors declare no conflict of interest. References Heaney, A. Clinical review: Pituitary carcinoma: Difficult diagnosis and treatment. J. Clin. Endocrinol. Metab. 2011, 96, 3649–3660. [Google Scholar] [CrossRef] [PubMed] Raverot, G.; Burman, P.; McCormack, A.; Heaney, A.; Petersenn, S.; Popovic, V.; Trouillas, J.; Dekkers, O.M.; The European Society of Endocrinology. European Society of Endocrinology Clinical Practice Guidelines for the management of aggressive pituitary tumours and carcinomas. Eur. J. Endocrinol. 2018, 178, G1–G24. [Google Scholar] [CrossRef] Todeschini, A.B.; Beer-Furlan, A.; Montaser, A.S.; Jamshidi, A.O.; Ghalib, L.G.; Chavez, J.A.; Lehman, N.L.; Prevedello, D.M. 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  17. Abstract Purpose: Literature regarding endogenous Cushing syndrome (CS) largely focuses on the challenges of diagnosis, subtyping, and treatment. The enigmatic phenomenon of glucocorticoid withdrawal syndrome (GWS), due to rapid reduction in cortisol exposure following treatment of CS, is less commonly discussed but also difficult to manage. We highlight the clinical approach to navigating patients from GWS and adrenal insufficiency to full hypothalamic-pituitary-adrenal (HPA) axis recovery. Methods: We review the literature on the pathogenesis of GWS and its clinical presentation. We provide strategies for glucocorticoid dosing and tapering, HPA axis testing, as well as pharmacotherapy and ancillary treatments for GWS symptom management. Results: GWS can be difficult to differentiate from adrenal insufficiency and CS recurrence, which complicates glucocorticoid dosing and tapering regimens. Monitoring for HPA axis recovery requires both clinical and biochemical assessments. The most important intervention is reassurance to patients that GWS symptoms portend a favorable prognosis of sustained remission from CS, and GWS typically resolves as the HPA axis recovers. GWS also occurs during medical management of CS, and gradual dose titration based primarily on symptoms is essential to maintain adherence and to eventually achieve disease control. Myopathy and neurocognitive dysfunction can be chronic complications of CS that do not completely recover. Conclusions: Due to limited data, no guidelines have been developed for management of GWS. Nevertheless, this article provides overarching themes derived from published literature plus expert opinion and experience. Future studies are needed to better understand the pathophysiology of GWS to guide more targeted and optimal treatments. Introduction Endogenous neoplastic hypercortisolism - Cushing syndrome (CS) - is one of the most challenging diagnostic and management problems in clinical endocrinology. CS may be due to either a pituitary tumor (Cushing disease, CD), or a non-pituitary (ectopic) tumor secreting ACTH. ACTH-independent hypercortisolism due to unilateral or bilateral adrenal nodular disease has been increasingly recognized as an important cause of CS. Regardless of the cause of CS, the clinical manifestations are protean and include a myriad of clinical, biochemical, neurocognitive, and neuropsychiatric abnormalities. The catabolic state of hypercortisolism causes signs and symptoms including skin fragility, bruising, delayed healing, violaceous striae, muscle weakness, and low bone mass with fragility fractures. Other clinical features include weight gain, fatigue, depression, difficulty concentrating, insomnia, facial plethora, and fat redistribution to the head and neck with resultant supraclavicular and dorsocervical fullness[1]. Metabolic consequences of hypercortisolism including hypertension, diabetes, and dyslipidemia are common. In addition, women often experience hirsutism and menstrual irregularity, while men may have hypogonadism. Management options of CS include surgery, medications, and radiation. The preferred first line treatment, regardless of source, is surgery, which offers the potential for remission[2,3,4]. The primary literature, reviews, and clinical practice guidelines for CS have traditionally focused on the diagnosis, subtyping, and surgical approach to CS. This bias derives first from the profound diagnostic challenge posed in the evaluation of cortisol production and dynamics, given that circulating cortisol follows a circadian rhythm, exhibits extensive protein binding and metabolism, and rises acutely with stress. CD and ectopic ACTH syndrome may be difficult to distinguish clinically and biochemically, and inferior petrosal sinus sampling is required in many patients to resolve this differential diagnosis. Ectopic ACTH-producing tumors can also be small, and these tumors can escape localization despite the best current methods. Although diagnosis and initial surgical remission can be achieved in the majority of patient with CS at experienced centers, up to 50% of patients with CD will require additional therapies after unsuccessful primary surgeries or recurrence up to many years later[5]. For patients who do not achieve surgical cure or who are not surgical candidates, several medical treatment options are now available. Pharmacotherapies directed at the pituitary include pasireotide[6, 7] (FDA approved) and cabergoline[8]. Adrenal steroidogenesis inhibitors such as osilodrostat[9] (FDA approved), metyrapone[10], levoketoconazole[11] (FDA approved) and ketoconazole[12], as well as the glucocorticoid antagonist, mifepristone[13] (FDA approved), are now widely used to treat CS. Pituitary radiotherapy is an additional treatment option for CD but can take months to years to lower cortisol production. Bilateral adrenalectomy (BLA) provides immediate, reliable correction of hypercortisolism but mandates life-long corticosteroid replacement therapy, and, in patients with CD, may be complicated by corticotroph tumor progression syndrome in 25–40% of patients[14]. After successful surgery for CS, the rapid onset of adrenal insufficiency (AI) is anticipated and usually portends a favorable prognosis [15,16,17,18]; however, despite the use of post-operative corticosteroid replacement, the rapid reduction in cortisol exposure often results in an enigmatic phenomenon referred to as the glucocorticoid withdrawal syndrome (GWS). This article addresses the clinical presentation and the pathogenesis of GWS, as well as its distinction from AI. When available, appropriate references are provided. Statements and guidance provided without references are derived from expert opinion and experience. Clinical Presentation and Pathogenesis of GWS GWS occurs following withdrawal of supraphysiologic exposure to either exogenous or endogenous glucocorticoids of at least several months duration[19]. After surgical cure of endogenous CS, GWS is usually characterized by biochemical evidence of hypothalamic-pituitary-adrenal (HPA) axis suppression with many signs and symptoms consistent with cortisol deficiency despite the use of supraphysiologic glucocorticoid replacement therapy. The degree of physical or psychologic glucocorticoid dependence experienced by patients may not correlate with the degree of HPA axis suppression[20, 21]. GWS symptom onset is typically 3–10 days postoperatively, often after the patient has been discharged from the hospital. The first symptoms of GWS vary but usually consist of myalgias, muscle weakness, fatigue, and hypersomnolence. Anorexia, nausea, and abdominal discomfort are common, but vomiting should raise concern for hyponatremia, cerebrospinal fluid leak, hydrocephalus, or other perioperative complications. Mood changes develop more gradually and range from mood swings to depression, and the fatigue with myalgias can exacerbate mood changes. An atypical depressive disorder has been described in many patients after CD surgery[22]. Weight loss should ensue in most patients but gradually and proportionate to the reduction in glucocorticoid exposure. It is important to complete a thorough symptom review and physical exam at postoperative visits, as the differentiation between GWS and bona fide AI – and even between GWS and recurrence of CS – can be challenging (Fig. 1). All three conditions are associated with symptoms of myalgias, weakness, and fatigue; however, rapid weight loss, hypoglycemia, and hypotension are suggestive of AI and the need for an increase in the glucocorticoid dose. In parallel, hypersomnia is more suggestive of GWS, while insomnia is more associated with recurrence of CS. Given the anticipation of GWS onset shortly after discharge and the potential for hyponatremia during this time, a widely employed strategy is a generous glucocorticoid dose for the first 2–3 weeks, at least until the first postoperative outpatient visit (Table 1). Fig. 1 Overlapping clinical features of Cushing syndrome (CS), glucocorticoid withdrawal syndrome (GWS), and adrenal insufficiency (AI) Full size image Table 1 Glucocorticoid Therapy Options After Surgery for CS Full size table The mechanisms responsible for the precipitation of the GWS after surgery for CS and the variability in its manifestations are not completely understood, yet alterations in the regulation of cortisol and cortisol-responsive genes appear to contribute. Down-regulation of corticotropin-releasing hormone (CRH) and proopiomelanocortin (POMC) expression, combined with up-regulation of cytokines and prostaglandins are likely to be important components of GWS. Low CRH has been associated with atypical depression[23], and CRH levels in cerebrospinal fluid of patients with CD are significantly lower compared to healthy subjects[24]. CRH suppression gradually resolves after surgical cure over 12 months during glucocorticoid replacement[25], illustrative of the slow recovery process. The expression of POMC, the ACTH precursor molecule, is also suppressed with chronic glucocorticoid exposure[26], and the normalization of POMC-associated peptides mirrors HPA axis recovery[19]. In the acute phase of glucocorticoid withdrawal, interleukins IL-6 and IL-1β, as well as tumor-necrosis factor alpha (TNFα) have been observed to rise[27], suggesting that glucocorticoid-mediated suppression of cytokines and prostaglandins is then released in GWS, and these cytokines induce the associated flu-like symptoms. Glucocorticoid replacement with dexamethasone 0.5 mg/d reduced but did not normalize IL-6 after 4–5 days[27], consistent with resistance to suppression during GWS. Acute Care: Perioperative Planning, Coaching, and Management For patients with CD, transsphenoidal surgery performed by an experienced surgeon achieves remission in about 80% of pituitary microadenomas and 60% of macroadenomas[28,29,30,31]. Post-operative AI and GWS are some of the most challenging phases of management for endocrinologists and one of the most disheartening for CS patients. Many patients report feeling unprepared for the postsurgical recovery process[32]. For these reasons, it is important to prepare the patient prior to surgery for the difficult months ahead, and the same considerations apply to the commencement of medical therapies, as will be discussed later. On the one hand, more potent glucocorticoids and higher doses reliably mitigate symptoms, but on the other hand, substitution of exogenous for endogenous CS delays recovery of the HPA axis and perpetuates CS-related co-morbidities. Limited data that compare management strategies preclude evidence-based decisions, yet some themes can be derived from expert opinion and extensive experience from CS centers. In centers dedicated to the management of CS, surgeons and endocrinologists work closely together through all phases of the process. Although the goal of primary surgery for CD is adenoma resection, the tumor might not be found and/or removed completely after initial exploration. To prepare for this possibility, the surgeon should determine in advance with the patient and endocrinologist what to do next in this situation – dissect further, perform a hypophysectomy or hemi-hypophysectomy, or stop the operation. The plan for perioperative testing and glucocorticoid treatment varies widely among centers. The conundrum faced in the immediate perioperative period is that withholding glucocorticoids allows for rapid testing and demonstration of remission; however, complete resection of the causative tumor causes AI from prolonged suppression of the HPA axis and concerns for acute decompensation. Abundant evidence has shown that post-pituitary adenomectomy patients are not at risk for an adrenal crisis when monitored closely in an intensive care unit or equivalent setting[33]. Many studies have confirmed that post-operative AI almost always suggests a remission of CD[15,16,17,18, 34]. A standard protocol includes securing serum electrolytes and cortisol, plasma ACTH, capillary blood glucose, blood pressure, and urine specific gravity every 6 h for 24–48 h while withholding all glucocorticoids. Consecutive serum cortisol values less than 2–5 µg/dL (we use < 3 µg/dL) are sufficient to document successful tumor resection and to begin glucocorticoid therapy[35]. Post-operative signs and symptoms of AI including vomiting, hyponatremia, hypoglycemia, and hypotension should also mandate immediate glucocorticoid support. Although not clinically useful in the immediate post-operative period, some investigators have shown that low ACTH and DHEAS levels may be better predictors of long-term remission than serum cortisol[36]. A similar strategy for the management of possible post-operative AI/GWS following unilateral adrenalectomy for nodular adrenal disease has recently been reported. A post-operative day 1 basal cortisol and its response to cosyntropin stimulation can reliably segregate those patients with HPA axis suppression requiring cortisol replacement from those with an intact HPA axis who do not need to be discharged with glucocorticoid therapy[37]. Once remission is achieved, exogenous glucocorticoid replacement should be initiated and maintained during the months required for HPA axis recovery. Several glucocorticoids and dosing options are available (Table 1), and the initial dose is generally 3- to 4-fold higher than the physiologic range and graded based on age, comorbidities, and severity of disease. Fludrocortisone acetate should also be initiated following BLA for patients who receive glucocorticoids other than hydrocortisone, the only glucocorticoid with mineralocorticoid activity. By comparison, post-BLA patients receiving supraphysiologic hydrocortisone doses usually do not need mineralocorticoid support until their dose is tapered to near physiologic replacement. In the acute postoperative period, several medical comorbidities accompanying CS may reverse rapidly and require medication adjustments[35]. In particular, insulin and oral hypoglycemic drugs, potassium-sparing diuretics such as spironolactone, and other cardiovascular drugs are typically tapered or discontinued as glucose counter-regulation and electrolyte balance change rapidly upon cortisol reduction. Due to the high risk of postoperative venous thromboembolism[38,39,40], prophylaxis is frequently recommended and continued for several weeks after discharge. Posterior pituitary manipulation can disturb water balance and result in serum sodium alterations, including transient or permanent central diabetes insipidus, and in rare cases the triphasic response of diabetes insipidus, followed by syndrome of inappropriate secretion of antidiuretic hormone (SIADH), and finally permanent diabetes insipidus[41, 42]. In the first week or two after discharge, the most common cause for readmission is hyponatremia[43, 44], although the mechanisms responsible for this transient SIADH state are not known. For this reason, patients should be instructed to drink only when thirsty and not as an alternative to solid foods or for social reasons for 7–10 days after the surgery. Both diabetes insipidus and SIADH may not manifest for weeks after surgery; consequently, serum sodium should be monitored after hospital discharge as well [42]. Subacute Care: The GWS and HPA Axis Recovery When managing GWS symptoms, it is important to repeatedly emphasize to the patient that not only are GWS symptoms to be expected, but in fact these manifestations portend a favorable prognosis of sustained remission from CS. The most important treatment intervention is frequent reassurance to the patient that GWS typically resolves as the HPA axis recovers. Family members must be included in the conversation to help provide as much support as possible, as patients report that support from family and friends is the most helpful coping mechanism during the recovery process[32]. When appropriate, it may be necessary to provide the patient with temporary disability documentation, since GWS symptoms may be so severe to preclude gainful employment. The patient must know that the myalgias reflect the body’s attempts to repair the muscle damage, similar to the soreness experienced the day after resistance weight training, and these aches will eventually subside. Due to the challenges of differentiating between GWS and AI, a higher glucocorticoid dose can be briefly trialed to assess if this increased glucocorticoid exposure improves symptoms, but late-day dosing should be avoided to support recovery of the circadian rhythm. In parallel, the patient should be encouraged to adequately rest, particularly going to sleep early but limiting daytime sleep to short naps. Several other classes of medications can be trialed to target specific patient symptoms (Table 2). Antidepressants such as fluoxetine, sertraline, and trazodone might help to improve mood, sleep and appetite. A non-steroidal anti-inflammatory medication to address the musculoskeletal discomfort might be used early in the GWS, with the cyclooxygenase type 2 (COX-2) inhibitor celecoxib (100–200 mg once or twice daily) preferred when several weeks of daily treatment is needed, generally not more than 3 months. With anorexia and reduced food intake, adequate protein intake is necessary to allow muscle recovery. Egg whites, nuts, and lean meats are nutritionally dense and generally easy to tolerate despite poor appetite. Table 2 Pharmacotherapy and Ancillary Treatment Options for GWS Symptoms Full size table Following surgical remission, the duration of glucocorticoid taper can vary from 6 to 12 months or more, depending on age, severity of disease, and duration of disease [45, 46]. Monitoring for HPA axis recovery involves both clinical and biochemical assessments. Since the HPA axis is likely to remain suppressed with prolonged supraphysiologic glucocorticoid replacement, the first goal is to shift from all-day dosing to a circadian schedule as soon as possible, such as hydrocortisone 20 mg on rising and 10 mg in the early afternoon by 2–6 weeks after surgery. The advantages of hydrocortisone include rapid absorption for symptom mitigation, the ability to measure serum cortisol as a measure of drug exposure when helpful, and the relatively short half-life [47], which ensures a glucocorticoid-free period in the early morning when it is most critical to avoid prolonged HPA axis suppression and to enhance recovery. The second goal, which should not be attempted until GWS symptoms – particularly the anorexia and myalgias – are considerably improved, is to limit replacement to a single morning dose. Biochemical assessment should begin once patients are taking a physiologic dose of glucocorticoid replacement (total daily dose of hydrocortisone 15 to 20 mg per day) and clinically feel well enough to begin the final stage to discontinuation of glucocorticoid replacement (Fig. 2). Biochemical evaluation begins with basal testing, and dynamic assessment of adrenal function might be necessary to confirm completion of recovery. For basal testing, patients should not take their afternoon hydrocortisone dose (if prescribed) the day before testing and then have a blood draw by 0830 prior to the morning hydrocortisone dose on the day of testing. While a serum cortisol alone is adequate to taper hydrocortisone, a simultaneous plasma ACTH assists in gauging the state of HPA axis recovery. Often the ACTH and cortisol rise gradually in parallel, but sometimes the ACTH rises above the normal range despite a low cortisol, which indicates recovery of the hypothalamus (CRH neuron) and pituitary corticotrophs in advance of adrenal function. Serum DHEAS can remain suppressed for months to years after cortisol normalization, and a low DHEAS does not indicate continued HPA axis suppression. A rapid rise in DHEAS, in contrast, is concerning for disease recurrence, but a slow drift to a measurable amount in parallel with the cortisol rise is consistent with HPA axis recovery. Periodic assessment of electrolytes is prudent to screen for hyponatremia and hypo- or hyperkalemia as medications are changed, particularly diuretics. Hypercalcemia that is parathyroid-hormone independent might be observed during the recovery phase, probably related to the rise in cytokines that accompany resolution of hypercortisolemia[48, 49]. Fig. 2 Glucocorticoid withdrawal algorithm. TDD, total daily dose Full size image Basal testing is performed at 4- to 6-week intervals during glucocorticoid replacement. A rule of thumb is that the AM cortisol in µg/dL plus the morning dose of hydrocortisone in milligrams should sum to 15–20. Thus, once endogenous cortisol production is measurable, the hydrocortisone dose should be not more than 20 mg on arising. Once the AM cortisol rises to near 5 and then 10 µg/dL, the AM hydrocortisone dose is dropped to 15 and then 10 mg, respectively. Once the AM cortisol is 12–14 µg/dL, recovery is essentially complete, and the morning hydrocortisone dose is dropped to 5 mg for 4–6 weeks and then stopped or held for dynamic testing (Fig. 2). A clinical pearl related to HPA axis recovery is that patients who state that they are finally feeling better and getting over the GWS usually have started to make some endogenous cortisol, yet not enough to stop glucocorticoid tapering. Nevertheless, a smidgeon of endogenous cortisol production with the waning of GWS symptoms is a harbinger that HPA axis recovery is imminent. If basal testing is equivocal, dynamic testing might be necessary. The gold standard testing for central AI is the insulin tolerance test, which is rarely used, and metyrapone testing might be employed once the basal cortisol is > 10 µg/dL. Although designed to test for primary adrenal insufficiency, the cosyntropin stimulation test is often employed in this setting due to greater availability, simplicity, and safety than insulin or metyrapone testing. The duration of full HPA axis recovery can be highly variable depending on the individual and postoperative glucocorticoid dosing[50]. GWS During Medical Management of CS Patients who are not surgical candidates or do not have successful remission of CS following surgery may be offered medical treatment or BLA. After BLA, the GWS will ensue without eventual recovery of the HPA axis, so glucocorticoids are tapered until a chronic physiologic replacement dose is reached as described previously. With medical management, patients might also experience GWS, particularly at the onset of treatment. Therefore, patients must be counseled that the typical symptoms of fatigue, myalgias, and anorexia are not only possible but indeed expected, rather than “side effects” of the medication, with two caveats. First, as described for glucocorticoid replacement following surgical remission, the endocrinologist must distinguish GWS from AI due to over-treatment of CS. The same parameters of vomiting, hypotension, and hypoglycemia favor inadequate cortisol exposure and the need for dose reduction or treatment pause and/or supplementation with a potent glucocorticoid such as dexamethasone to reverse an acute event. Second, known adverse effects of the specific drug in use should be considered and excluded. The quandary of distinguishing GWS from over-treatment raises an important principle of medical management: under-dose initially and gauge primarily the severity of GWS symptoms in the first several days. The initial goal of medical therapy is not to rapidly achieve normal cortisol milieu, but rather to “dial in” just enough inhibition of cortisol production or receptor antagonism to precipitate mild to moderate GWS symptoms. Once GWS symptoms appear and/or a typical dose of the medication is achieved, further assessments, including glucose, serum cortisol and/or UFC (except when treated with mifepristone), clinical appearance, and body weight are conducted while the dose is maintained constant until GWS symptoms begin to dissipate. If the patient is not experiencing adequate clinical and/or biochemical benefit from the medication in the absence of GWS symptoms, the dose is gradually raised incrementally. This iterative process might require periodic dose reduction or perhaps even temporarily discontinuing the medication if the patient’s daily living activities are affected at any point in the process. For several medications, a block-and-replacement strategy is an option[3], particularly for very compliant patients for whom a priority is placed on avoidance of over-treatment. This strategy resembles thionamide-plus-levothyroxine therapy for the treatment of Graves disease. The patient is given both a generous dose of medication to completely block endogenous glucocorticoid production, plus simultaneous exogenous glucocorticoid therapy, titrated to replacement dose or greater. This approach allows for greater control over glucocorticoid exposure and low risk of AI, as long as the patient always takes both medications each day. Long-acting pasireotide, for example, would not be an appropriate drug for the block-and-replace strategy. Based on the drug mechanism of action, this block-and-replace strategy is feasible with ketoconazole or levoketoconazole, the 11β-hydroxylase inhibitors osilodrostat and metyrapone, and the adrenolytic agent mitotane (the latter three are off-label uses). Alternatively, the patient might be given a double replacement dose of glucocorticoid to take only if symptoms concerning for over-treatment occur, and the medical therapy for hypercortisolemia is then interrupted until the patient communicates with the endocrinologist. Treatment monitoring with medical management includes biochemical and symptom assessment. For all medications other than mifepristone, normalization of 24-hour UFC is the minimal goal [2]. Basal morning cortisol and late-night salivary cortisol may be more challenging to interpret in the setting of diurnal rhythm loss characteristic of CS. Because mifepristone blocks glucocorticoid receptors, ACTH and cortisol increase with treatment for most forms of CS; dose titration therefore relies on assessment of clinical features, glycemia, body weight, and other metabolic parameters [2]. For occult tumors, periodic imaging to screen for a surgical target and/or tumor regrowth is prudent, and a pause in treatment for repeat surgery might be indicated. The End Game: Comprehensive Recovery for the Patient with CS Besides navigating the GWS and shepherding recovery of the HPA axis, recovery from co-morbidities of CS must be addressed to the extent possible. Hypertension, hyperglycemia, hypokalemia, and dyslipidemia often improve substantially but do not always resolve. Insomnia, skin thinning and bruising, and risk of thrombosis also generally resolve, and associated treatments might be discontinued. Although there is usually an improvement in bone density and decreased fracture risk following correction of CS, anabolic and/or anti-resorptive therapies may be warranted in some patients. The deformities of vertebral compression fractures may be permanent, and some authors have recommended the use of vertebroplasty for symptom relief[51]. Violaceous striae and chronic skin tears might heal with hyperpigmentation, leaving “the scars of Cushing’s,” which can persist for a lifetime. These milestones or minor victories can be used as evidence of healing and encouragement for the patient during the dark days of the GWS, and these changes herald further improvements. Fat redistribution and significant weight loss take some weeks to manifest and usually follow next. The myopathy from CS is an example of a co-morbidity that rarely improves without targeted treatment, and the German Cushing’s Registry has provided evidence for chronic muscle dysfunction following cure of CS[52]. Recent data indicate that a low IGF-1 after curative surgery is associated with long-term myopathy [53]. This persistent myopathy is a common source of chronic fatigue following HPA axis recovery, which is unresponsive to glucocorticoids. For these reasons, an important ancillary modality is physical therapy, and an ideal time to initiate this treatment is at the first signs of HPA axis recovery when the GWS symptoms have subsided. A complete evaluation from an experienced physical therapist should focus on core and proximal muscle strength, balance, and other factors that limit function. Exercises targeting these factors (stand on one foot, sit-to-stand, straight-arm raises with 1- to 5-pound weights) rather than traditional gym exercises (arm curls, bench press, treadmill) are necessary to restore functional status and avoid frustration and injury when the patient is not yet prepared for the latter stages of recovery. Professional supervision of this initial phase is a critical component of the recovery process, and failure to attend to musculoskeletal rehabilitation – as would be routine following survival of a critical illness – risks long-term morbidities from a curable disease. Patients with CS often complain of cognitive defects, which usually improve but may not completely recover following treatment[54, 55]. Glucocorticoids are toxic to the hippocampus, and both rats treated with high-dose corticosterone and patients with CD experience reductions in hippocampal volume, which does not completely return to normal even with correction of hypercortisolemia[56, 57]. Because the hippocampus is an important brain region for memory, the main complaint is impaired formation of new memories and recall of recent events. When significant cognitive dysfunction persists, a formal neuropsychologic testing session is prudent, both to screen for additional sources of memory loss (degenerative brain diseases) and to identify aspects that might be amenable to functional management approaches. Cognitive therapy can be effective for mental health and overall disease coping strategies as well. Finally, for patients undergoing transsphenoidal surgery for CD, complications associated with pituitary surgeries in general should also be considered. Anterior pituitary hormone axes should be assessed biochemically and symptomatically for hypothyroidism and hypogonadism, as hypopituitarism is an independent predictor of decreased quality of life after surgical cure [58]. Hypopituitarism can not only complicate the assessment of GWS with overlapping symptoms such as fatigue, but treatment of hypopituitarism can also be important for GWS recovery. Prior to initiating physical therapy, testosterone replacement in male patients with hypogonadism should be optimized. Hypothyroidism can contribute to hyponatremia and can also slow the metabolism of glucocorticoids. Therefore, optimizing the treatment of hypothyroidism and hypogonadism prior to completing glucocorticoid taper is prudent. Growth hormone deficiency may also be evaluated in symptomatic patients in the setting of other anterior pituitary hormone deficiencies, although formal evaluation is best delayed for at least 6–12 months when HPA axis recovery has occurred or at least the glucocorticoid dose is reduced to a physiologic range [2]. Summary and Final Thoughts After a diagnosis of CS has been well established, a multidisciplinary team of endocrinologists and surgeons must design the best treatment strategy for the patient. Expectations and possible adverse side effects of surgery or pharmacotherapy should be reviewed with the patient. The GWS is a very difficult concept for patients to understand. It seems inconceivable to them that they could possibly feel worse (and that this is a good omen) six weeks after resolution of their hypercortisolism than they do pre-operatively; however, there are no studies that address whether comprehensive pre-operative patient education regarding GWS has any impact on the patient’s post-operative perception and outcome after successful surgery. An addiction metaphor is sometimes helpful: the patient’s body and brain has become addicted to steroids (cortisol) and after steroids are abruptly reduced, their body and brain are dysphoric — much like removal of any other addictive substance (e.g., opioids, alcohol, nicotine). The patient and their care team need to know that this treatment odyssey will be a marathon, not a sprint. It may take as long as 12–18 months for patients to have full HPA axis recovery, regression of GWS, and, most importantly, resolution of the devastating effects of chronic excessive glucocorticoid exposure. Conclusions GWS following surgery or during medical treatment of CS can be challenging to manage. There are currently no standard guidelines for management of GWS, but various available medical and ancillary therapies are discussed here. Studies are needed to better understand the pathophysiology of GWS to guide more targeted treatments. There may be yet unrecognized steroids produced by the adrenal glands, the withdrawal of which contributes to GWS symptoms[59]. Future observational and interventional studies would be beneficial for identifying optimal management options. References Carroll TB, Findling JW (2010) The diagnosis of Cushing’s syndrome. 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Funding XH is supported by grant T32DK07245 from the National Institutes of Diabetes and Digestive and Kidney Diseases. Author information Affiliations Department of Internal Medicine, Division of Metabolism, Endocrinology and Diabetes, University of Michigan Medical School, Ann Arbor, MI, USA Xin He & Richard J. Auchus Department of Medicine, Division of Endocrinology and Molecular Medicine, Medical College of Wisconsin, Milwaukee, WI, USA James W. Findling Endocrinology Center and Clinics, Medical College of Wisconsin, Milwaukee, WI, USA James W. Findling Department of Pharmacology, University of Michigan Medical School, Ann Arbor, MI, USA Richard J. Auchus Lieutenant Colonel Charles S. Kettles Veterans Affairs Ann Arbor Healthcare System, Ann Arbor, MI, USA Richard J. Auchus Contributions All authors contributed to the manuscript conception, design, and content. All authors read, edited, and approved the final manuscript. Corresponding author Correspondence to Richard J. Auchus. Ethics declarations Financial Interests Dr. Auchus has received research support from Novartis Pharmaceuticals, Corcept Therapeutics, Spruce Biosciences, and Neurocrine Biosciences and has served as a consultant for Corcept Therapeutics, Janssen Pharmaceuticals, Novartis Pharmaceuticals, Quest Diagnostics, Adrenas Therapeutics, Crinetics Pharmaceuticals, PhaseBio Pharmaceuticals, OMass Therapeutics, Recordati Rare Diseases, Strongbridge Biopharma, and H Lundbeck A/S. Dr. Findling has received research support from Novartis Pharmaceuticals and has served as a consultant for Corcept Therapeutics and Recordati Rare Diseases. Human Subjects and Animals No human subjects or animals were used to collect data for this manuscript. Additional information Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Electronic Supplementary Material Below is the link to the electronic supplementary material. Supplementary Material 1 Rights and permissions Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. Reprints and Permissions Cite this article He, X., Findling, J.W. & Auchus, R.J. Glucocorticoid Withdrawal Syndrome following treatment of endogenous Cushing Syndrome. Pituitary (2022). https://doi.org/10.1007/s11102-022-01218-y Download citation Accepted15 March 2022 Published26 April 2022 DOIhttps://doi.org/10.1007/s11102-022-01218-y From https://link.springer.com/article/10.1007/s11102-022-01218-y
  18. Compared with placebo, levoketoconazole improved cortisol control and serum cholesterol levels for adults with endogenous Cushing’s syndrome, according to results from the LOGICS study presented here. Safety and efficacy of levoketoconazole (Recorlev, Xeris Biopharma) for treatment of Cushing’s syndrome were established in the pivotal phase 3, open-label SONICS study. The phase 3, double-blind LOGICS study sought to demonstrate the drug specificity of levoketoconazole in normalizing mean urinary free cortisol (mUFC) level. “Treatment with levoketoconazole benefited patients with Cushing’s syndrome of different etiologies and a wide range in UFC elevations at baseline by frequent normalization of UFC,” Ilan Shimon, MD, professor at the Sackler Faculty of Medicine at Tel Aviv University and associate dean of the Faculty of Medicine at Rabin Medical Center and director of the Institute of Endocrinology in Israel, told Healio. “This is a valuable Cushing’s study as it includes a placebo-controlled randomized withdrawal phase.” LOGICS participants were drawn from a cohort of 79 adults with Cushing’s syndrome with a baseline mUFC at least 1.5 times the upper limit of normal who participated in a single-arm, open-label titration and maintenance phase of approximately 14 to 19 weeks. Researchers randomly assigned 39 of those participants plus five from SONICS who had normalized mUFC levels on stable doses of levoketoconazole for at least 4 weeks to continue to receive the medication (n = 22) or to receive placebo with withdrawal of the medication (n = 22) for 8 weeks. At the end of the withdrawal period, all participants received levoketoconazole for 8 more weeks. Primary endpoint was proportion of participants who lost mUFC normalization during the randomized withdrawal period, and secondary endpoints included proportion with normalized mUFC and changes in total and LDL cholesterol at the end of the restoration period. During the withdrawal period, 95.5% of participants receiving placebo vs. 40.9% of those receiving levoketoconazole experienced loss of mUFC response, for a treatment difference of –54.5% (95% CI, –75.7 to –27.4; P = .0002). At the end of the withdrawal period, 4.5% of participants receiving placebo vs. 50% of those receiving levoketoconazole maintained normalized mUFC, for a treatment difference of 45.5% (95% CI, 19.2-67.9; P = .0015). Among participants who had received placebo and lost mUFC response, 60% regained normalized mUFC at the end of the restoration period. During the withdrawal period, participants in the placebo group had increases of 0.9 mmol/L in total cholesterol and 0.6 mmol/L in LDL cholesterol vs. decreases of 0.04 mmol/L (P = .0004) and 0.006 mmol/L (P = .0056), respectively, for the levoketoconazole group. The increases seen in the placebo group were reversed when participants restarted the medication. The most common adverse events with levoketoconazole were nausea (29%) and hypokalemia (26%). Prespecified adverse events of special interest were liver-related (10.7%), QT interval prolongation (10.7%) and adrenal insufficiency (9.5%). “This study has led to the FDA decision to approve levoketoconazole for the treatment of Cushing’s syndrome after surgical failure or if surgery is not possible,” Shimon said. From https://www.healio.com/news/endocrinology/20220512/logics-levoketoconazole-improves-cortisol-control-in-endogenous-cushings-syndrome
  19. A worldwide, observational study of adults and adolescents with growth hormone deficiency (GHD) found long-term GH replacement was safe. These findings were published in the Journal of Clinical Endocrinology & Metabolism. Data for this long-term follow-up study were sourced from the KIMS Pfizer International Metabolic Database cohort. Patients (N=15,809) with confirmed GHD were prescribed GH by their primary care physician. Adverse events were evaluated at up to 18 years (mean, 5.3 years). The median age of study participants was 44.8 (range, 5.6-91.2) years, 50.5% were girls or women, 94.4% were White, 57.6% were true-naive to treatment at baseline, 59.7% had pituitary or hypothalamic tumor, 21.6% had idiopathic or congenital GHD, and 67.8% had at least 2 pituitary deficiencies. Patients were administered a mean GH dosage of 0.30±0.30 mg/d. At year 15, patients (n=593) had a 1.7-kg/m2 increase in body mass index (BMI), a 4.3-kg increase in weight, a 0.4-cm decrease in height, a 6.2-cm increase in waist circumference, a 0.03 increase in waist to hip ratio, a 6.3-mm Hg increase in systolic blood pressure, a 1.0-mm Hg increase in diastolic blood pressure, and a 0.5-bpm decrease in heart rate. Approximately one-half of the patients (51.2%) experienced at least 1 adverse event, but few patients (18.8%) reported treatment-related adverse events. The most common all-cause adverse events included arthralgia (4.6%), peripheral edema (3.9%), headache (3.6%), influenza (2.8%), depression (2.8%), and recurrence of pituitary tumor (2.7%). The most common treatment-related adverse events were peripheral edema (3.1%) and arthralgia (2.6%). The rate of all-cause (P =.0141) and related (P =.0313) adverse events was significantly related with age at enrollment, with older patients (aged ³45 years) having higher rates than younger patients. The rate of all-cause and related adverse events was higher among patients with pituitary or hypothalamic tumor, adult-onset GHD, and insulin-like growth factor 1 standard deviation score greater than 0; those who had prior pituitary radiation treatment; and those who took a GH dosage of no more than 0.30 mg/d (all P £..014). A total of 1934 patients discontinued treatment, and 869 patients reduced their dose due to adverse events. Study discontinuation was highest among patients with idiopathic or congenital GHD (45.0%). At least 1 serious adverse event occurred among 4.3% of patients. The most common serious events included recurrence of pituitary tumor (n=154; 1.0%) and death (n=21; 0.1%). The highest mortality rate was observed among patients who enrolled at 45 years of age and older (4.7%). In total, 418 patients who had no history of cancer at baseline were diagnosed with cancer after starting GH treatment, which equated to a standardized incidence ratio of 0.92 (95% CI, 0.83-1.01). This study was limited as data were collected during routine clinical practice and no predefined windows or reporting were set. This study found that GH replacement therapy was safe at up to an 18-year follow-up among adolescents and adults. Disclosure: Multiple authors declared affiliations with industry. Please refer to the original article for a full list of disclosures. Reference Johannsson G, Touraine P, Feldt-Rasmussen U, et al. Long-term safety of growth hormone in adults with growth hormone deficiency: overview of 15,809 GH-treated patients. J Clin Endocrinol Metab. Published online April 3, 2022. doi:10.1210/clinem/dgac199 From https://www.endocrinologyadvisor.com/home/topics/general-endocrinology/safety-of-long-term-growth-hormone-treatment-assessed/
  20. Abstract Context Arginine-vasopressin and CRH act synergistically to stimulate secretion of ACTH. There is evidence that glucocorticoids act via negative feedback to suppress arginine-vasopressin secretion. Objective Our hypothesis was that a postoperative increase in plasma copeptin may serve as a marker of remission of Cushing disease (CD). Design Plasma copeptin was obtained in patients with CD before and daily on postoperative days 1 through 8 after transsphenoidal surgery. Peak postoperative copeptin levels and Δcopeptin values were compared among those in remission vs no remission. Results Forty-four patients (64% female, aged 7-55 years) were included, and 19 developed neither diabetes insipidus (DI) or syndrome of inappropriate anti-diuresis (SIADH). Thirty-three had follow-up at least 3 months postoperatively. There was no difference in peak postoperative copeptin in remission (6.1 pmol/L [4.3-12.1]) vs no remission (7.3 pmol/L [5.4-8.4], P = 0.88). Excluding those who developed DI or SIADH, there was no difference in peak postoperative copeptin in remission (10.2 pmol/L [6.9-21.0]) vs no remission (5.4 pmol/L [4.6-7.3], P = 0.20). However, a higher peak postoperative copeptin level was found in those in remission (14.6 pmol/L [±10.9] vs 5.8 (±1.4), P = 0.03]) with parametric testing. There was no difference in the Δcopeptin by remission status. Conclusions A difference in peak postoperative plasma copeptin as an early marker to predict remission of CD was not consistently present, although the data point to the need for a larger sample size to further evaluate this. However, the utility of this test may be limited to those who develop neither DI nor SIADH postoperatively. Cushing disease, copeptin, cortisol, remission Issue Section: Clinical Research Article Arginine vasopressin (AVP) and CRH act synergistically as the primary stimuli for secretion of ACTH, leading to release of cortisol [1, 2]. The role of AVP in the hypothalamic-pituitary-adrenal (HPA) axis is via release from the parvocellular neurons of the paraventricular nuclei (and possibly also from the magnocellular neurons of the paraventricular and supraoptic nuclei), the secretion of which is stimulated by stress [3-6]. AVP release results in both independent stimulation of ACTH release and potentiation of the effects of CRH [3, 7-9]. Additionally, there is evidence that glucocorticoids act by way of negative feedback to suppress AVP secretion [10, 11-20]. Further, parvocellular neurons of the hypothalamic paraventricular nuclei have been shown to increase AVP production and neurosecretory granule size after adrenalectomy, and inappropriately elevated plasma AVP has been reported in the setting of adrenal insufficiency with normalization of plasma AVP after glucocorticoid administration [21-24]. This relationship of AVP and its effect on the HPA axis has been used in the diagnostic evaluation of Cushing syndrome (CS) [14] and evaluation of remission after transsphenoidal surgery (TSS) in Cushing disease (CD) by administration of desmopressin [25]. Copeptin makes up the C-terminal portion of the AVP precursor pre-pro-AVP. Copeptin is released from the posterior pituitary in stoichiometric amounts with AVP, and because of its longer half-life in circulation, it is a stable surrogate marker of AVP secretion [26-28]. Plasma copeptin has been studied in various conditions of the anterior pituitary. In a study by Lewandowski et al, plasma copeptin was measured after administration of CRH in assessment of HPA-axis function in patients with a variety of pituitary diseases. An increase in plasma copeptin was observed only in healthy subjects but not in those with pituitary disease who had an appropriately stimulated serum cortisol, and the authors concluded that copeptin may be a sensitive marker to reveal subtle alterations in the regulation of pituitary function [7]. Although in this study and others, plasma copeptin was assessed after pituitary surgery, it has not, to the best of our knowledge, been studied as a marker of remission of CD before and after pituitary surgery [7, 29]. In this study, plasma copeptin levels were assessed as a surrogate of AVP secretion before and after TSS for treatment of CD. Because there is evidence that glucocorticoids exert negative feedback on AVP, we hypothesized that there would be a greater postoperative increase in plasma copeptin in those with CD in remission after TSS resulting from resolution of hypercortisolemia and resultant hypocortisolemia compared with those not in remission with persistent hypercortisolemia and continued negative feedback. In other words, we hypothesized that an increase in copeptin could be an early marker of remission of CD after TSS. We aimed to complete this assessment by comparison of the peak postoperative copeptin and change in copeptin from preoperative to peak postoperative copeptin for those in remission vs not in remission postoperatively. Subjects and Methods Subjects Adult and pediatric patients with CD who presented at the Eunice Kennedy Shriver National Institute of Child Health and Human Development under protocol 97-CH-0076 and underwent TSS between March 2016 and July 2019 were included in the study. Exclusion criteria included a prior TSS within 6 weeks of the preoperative plasma copeptin sample or a preoperative diagnosis of diabetes insipidus, renal disease, or cardiac failure. Written informed consent was provided by patients aged 18 years and older and by legal guardians for patients aged < 18 years to participate in this study. Written informed assent was provided by patients aged 7 years to < 18 years. The 97-CH-0076 study (Investigation of Pituitary Tumors and Related Hypothalamic Disorders) has been approved by the Eunice Kennedy Shriver National Institute of Child Health and Human Development institutional review board. Clinical and Biochemical Data Clinical data were extracted from electronic medical records. Age, sex, body weight, body mass index (BMI), pubertal stage (in pediatric patients only), and history of prior TSS were obtained preoperatively during the admission for TSS. Clinical data obtained postoperatively included TSS date, histology, development of central diabetes insipidus (DI) or (SIADH), time from TSS to most recent follow-up, and clinical remission status at postoperative follow-up. Preoperatively, serum sodium, 24-hour urinary free cortisol (UFC), UFC times the upper limit of normal (UFC × ULN), midnight (MN) serum cortisol, MN plasma ACTH, and 8 AM plasma ACTH were collected. Postoperatively, serum sodium, serum and urine osmolality, urine specific gravity, serum cortisol, and plasma ACTH were collected. For serum cortisol values < 1 mcg/dL, a value of 0.5 mcg/dL was assigned for the analyses; for plasma ACTH levels < 5 pg/mL, a value of 2.5 pg/mL was assigned. Additionally, plasma copeptin levels were obtained preoperatively and on postoperative days (PODs) 1 through 8 after TSS at 8:00 AM. Peak postoperative copeptin was the highest plasma copeptin on PODs 1 through 8. The delta copeptin (Δcopeptin) was determined by subtracting the preoperative copeptin from the peak postoperative copeptin; hence, a positive change indicated a postoperative increase in plasma copeptin. Plasma copeptin was measured using an automated immunofluorescent sandwich assay on the BRAHMS Kryptor Compact PLUS Copeptin-proAVP. The limit of detection for the assay was 1.58 pmol/L, 5.7% intra-assay coefficient of variation, and 11.2% inter-assay coefficient of variation, with a lower limit of analytical measurement of 2.8 pmol/L. For those with multiple preoperative plasma copeptin values within days before surgery, an average of preoperative copeptin levels was used for analyses. Diagnosis of CD was based on guidelines published by the Endocrine Society and as previously described for the adult and pediatric populations [30, 31]; diagnosis was further confirmed by either histologic identification of an ACTH-secreting pituitary adenoma in the resected tumor specimen, decrease in cortisol and ACTH levels postoperatively, and/or clinical remission after TSS at follow-up evaluation. All patients were treated with TSS at the National Institutes of Health Clinical Center by the same neurosurgeon. Remission after surgical therapy was based on serum cortisol of < 5 μg/dL during the immediate postoperative period, improvement of clinical signs and symptoms of cortisol excess at postoperative follow up, nonelevated 24-hour UFC at postoperative follow-up, nonelevated midnight serum cortisol at postoperative follow up when available, and continued requirement for glucocorticoid replacement at 3 to 6 months’ postoperative follow-up. Diagnosis of SIADH was based on development of hyponatremia (serum sodium < 135 mmol/L) and oliguria (urine output < 0.5 mL/kg/h). Diagnosis of DI was determined by development of hypernatremia (serum sodium > 145 mmol/L), dilute polyuria (urine output > 4 mL/kg/h), elevated serum osmolality, and low urine osmolality. Statistical Analyses Results are presented as median (interquartile range [IQR], calculated as 25th percentile-75th percentile) or mean ± SD, as appropriate, and frequency (percentage). Where appropriate, we compared results using parametric or nonparametric testing; however, the median (IQR) and the mean ± SD were both reported to allow for comparisons with the appropriate testing noted. Subgroup analyses were completed comparing those who developed water balance disorders included patients who developed DI only (but not SIADH), those who developed SIADH only (but not DI), and those with no water balance disorder; hence, for these subgroup analyses, those who developed both DI and SIADH postoperatively (n = 4) were excluded. Preoperative copeptin, peak postoperative copeptin, and Δcopeptin were compared between those with and without remission at follow-up, using either t test or Wilcoxon rank-sum test, depending on the distribution of data. These were done in all patients combined, as well as within each subgroup. The same tests were used for comparing other continuous variables (eg, age, BMI SD score [SDS], cortisol excess measures) between those with and without remission. Categorical data (eg, sex, Tanner stage) were analyzed using the Fisher exact test. Comparisons of copeptin levels among the subgroups (DI, SIADH, neither) were carried out using mixed models and the Kruskal-Wallis test, as appropriate. Post hoc pairwise comparisons were adjusted for multiplicity using the Bonferroni correction, and as applicable, only corrected P values are reported. Mixed models for repeated measures also analyzed copeptin, serum sodium, and cortisol data for PODs 1 through 8. In addition, maximum likelihood estimation (GENMOD) procedures analyzed the effects of copeptin and serum sodium on the remission at follow-up. Correlation analyses were done with Spearman ρ. All analyses were tested for the potential confounding effects of age, sex, BMI SDS, and pubertal status, and were adjusted accordingly. For plasma copeptin reported as < 2.8 pmol/L, a value of 1.4 pmol/L (midpoint of 0 and 2.8 pmol/L) was used; sensitivity analyses repeated all relevant comparisons using the threshold limit of 2.8 pmol/L instead of 1.4 pmol/L. Odds ratios (OR) and 95% CIs, other magnitudes of the effect, data variability, and 2-sided P values provided the statistical evidence for the conclusions. Statistical analyses were performed in SAS version 9.4 software (SAS Institute, Inc, Cary, NC). Results Patient Characteristics Forty-four adult and pediatric patients, aged 7 to 55 years (77.2% were < 18 years old), with CD were included in the study. The cohort included 28 female patients (64%), and the median BMI SDS was 2.2 (1.1-2.5). Thirty-four percent (15/44) had prior pituitary surgery (none within the prior 6 weeks). Seventy-five percent (33/44) had postoperative follow-up evaluations available, with median follow-up of 13.5 months (11.3-16.0). Of those 33 patients, 85% were determined to be in remission at follow-up. Comparing those in remission vs no remission, there was no difference in age, sex, BMI SDS, pubertal status (in pediatric ages only), preoperative measures of cortisol excess (UFC × ULN, PM serum cortisol, MN plasma ACTH, AM plasma ACTH), duration of follow-up, or development of DI or SIADH. There was a lower postoperative serum cortisol nadir in those in remission at follow-up compared with those not in remission at follow-up, as expected, because a postoperative serum cortisol < 5 μg/dL was included in defining remission status. Postoperatively, 8/44 (18%) developed DI, 13/44 (30%) developed SIADH, 4/44 (9%) developed both DI and SIADH, and 19/44 (43%) developed no water balance disorder (Table 1). There were no differences by remission status when assessing these subgroups (ie, DI, SIADH, and no water balance disorder) separately. Table 1. Demographic and clinical characteristics of subjects All subjects, n = 44 All subjects by remission status, n = 33 All subjects by remission status, excluding those with DI or SIADH, n = 13 Remission, n = 28 No remission, n = 5 P Remission, n = 10 No remission, n = 3 P Age, median (range), y 14.5 (7-55) 17.4 ± 10.7 14.5 (12.5-17.5) 15.6 ± 13.2 11.0 (9.0-12.0) 0.11 13.7 ± 3.1 14.0 (13.0-15.0) 19.7 ± 16.8 11.0 (9.0-39.0) 0.60a Sex  Female 28 (64%) 22 (78.6%) 3 (60.0%) 0.57 9 (90.0%) 2 (66.7%) 0.42 BMI SDS 2.2 (1.1-2.5) 1.7 ± 1.0 2.0 (0.9-2.5) 2.2 ± 0.4 2.2 (2.1-2.3) 0.70 1.7 ± 1.1 2.0 (0.7-2.5) 2.0 ± 0.4 2.1 (1.5-2.3) 0.65a Pubertal status Female (n = 19) (n = 15) (n = 2) 0.51 (n = 8) (n = 1) 0.44   Tanner 1-2 6 4 (26.7%) 1 (50.0%) 3 (37.5%) 1 (25.0%)   Tanner 3-5 13 11 (73.3%) 1 (50.0%) 5 (62.5%) 0 Male (n = 14) (n = 5) (n = 2) (n = 1) (n = 1) --- Testicular volume < 12, mL 10 4 (80.0%) 2 (10.00%) 1 (100.0%) 1 (100.0%) Testicular volume ≥ 12, mL 4 1 (20.0%) 0 1.0 0 0 Preoperative UFC ULN 3.3 (1.2-6.1) 4.9 ± 6.1 2.6 (1.0-7.6) 3.2 ± 1.3 3.7 (3.0-3.9) 0.70 7.2 ± 8.4 3.9 (1.8-9.1) 3.8 ± 0.7 3.9 (3.0-4.4) 0.93 Preoperative PM cortisol 11.9 (9.2-14.8) 13.3 ± 4.7 12.2 (9.2-16.8) 10.8 ± 2.1 11.5 (9.0-11.6) 0.30 13.3 ± 6.0 11.2 (8.4-16.5) 11.1 ± 2.6 11.6 (8.3-13.6) 0.57a Preoperative MN ACTH 43.4 (29.3-51.6) 44.2 ± 25.5 46.1 (27.6-50.5) 40.9 ± 15.3 11.5 (9.0-11.6) 0.74 36.6 ± 16.6 37.4 (29.1-48.8) 34.0 ± 9.4 39.3 (23.1-39.5) 0.67 Preoperative AM ACTH 44.6 (31.4-60.5) 46.9 ± 28.9 44.0 (29.8-56.2) 48.6 ± 28.8 58.7 (21.7-60.5) 0.84 35.2 ± 16.2 40.3 (28.0-44.0) 45.4 ± 24.6 58.7 (17.0-60.5) 0.41a Postoperative cortisol nadir 0.5 (0.5-0.5) 0.7 ± 0.7 0.5 (0.5-0.5) 7.8 ± 6.6 5.2 (2.2-12.3) <0.001 0.6 ± 0.3 0.5 (0.5-0.5) 8.1 ± 7.9 5.2 (2.1-17.0) 0.003 Duration of follow-up 13.5 (11.3-16.0) 15.3 ± 7.9 14.0 (12.0-16.5) 14.0 ± 13.0 11.0 (6.0-14.0) 0.30 18.6 ± 11.2 15.5 (12.0-27.0) 16.7 ± 17.2 11.0 (3.0-36.0) 0.82a DI only 8 (18%) 7/8 (87.5%) 1/8 (12.5%) 0.91 --- --- --- SIADH only 13 (30%) 8/9 (88.9%) 1/9 (11.1%) Neither DI/SIADH 19 (43%) 10/13 (76.9%) 3/13 (23.1%) Both DI and SIADH 4 (9%) 3/3 (100%) 0/3 Demographic and clinical characteristics of all subjects (n = 44) with Cushing disease. Data are also presented by remission status for all subjects with postoperative follow-up (n = 33) and by remission status after excluding those who developed DI or SIADH postoperatively with postoperative follow-up (n = 13). Both median (IQR) and mean ± SD reported to allow for comparisons, with P value provided using appropriate testing depending on distribution of data sets. Data are mean ± SD, median (25th-75th IQR), or frequency (percentage) are reported, except for age, which is presented as median (range). Abbreviations: AM, 7:30-8 PM; BMI, body mass index; DI, diabetes insipidus; IQR, interquartile range; MN, midnight; N/A, not applicable; SDS, SD score; SIADH, syndrome of inappropriate antidiuresis; UFC, urinary free cortisol; ULN, upper limit of normal. p-values below the threshold of 0.05 are in bold. aP value indicates comparison using parametric testing, as appropriate for normally distributed data. Open in new tab Preoperative copeptin levels were higher in males (7.0 pmol/L [5.1-9.6]) than in females (4.0 pmol/L [1.4-5.8], P = 0.004) (Fig. 1). Age was inversely correlated with preoperative copeptin (rs = -0.35, P = 0.030) and BMI SDS was positively correlated with preoperative copeptin (rs = 0.54, P < 0.001) (Fig. 2). Figure 1. Open in new tabDownload slide Preoperative plasma copeptin and sex. Preoperative plasma copeptin in all patients, comparing by sex. A higher preoperative plasma copeptin was found in males (7.0 pmol/L [5.1-9.6]) than in females (4.0 pmol/L [1.4-5.8], P = 0.004). Horizontal lines = median. Whiskers = 25th and 75th interquartile ranges. Figure 2. Open in new tabDownload slide Preoperative plasma copeptin and BMI SDS. Association of preoperative plasma copeptin and BMI SDS in all patients. A BMI SDS was positively associated with a preoperative plasma copeptin (rs = 0.54, P < 0.001). Shaded area = 95% confidence interval. Copeptin Before and After Transsphenoidal Surgery for CD Among the 33 patients with postoperative follow-up, there was no difference in peak postoperative copeptin for patients in remission vs those not in remission (6.1 pmol/L [4.3-12.1] vs 7.3 pmol/L [5.4-8.4], P = 0.88). There was also no difference in the Δcopeptin for those in remission vs not in remission (2.3 pmol/L [-0.5 to 8.2] vs 0.1 pmol/L [-0.1 to 2.2], P = 0.46) (Fig. 3). Including all subjects, the mean preoperative copeptin was 5.6 pmol/L (±3.4). For patients with follow-up, there was no difference in preoperative copeptin for those in remission (4.8 pmol/L [±2.9]) vs no remission (6.0 pmol/L [±2.0], P = 0.47). POD 1 plasma copeptin ranged from < 2.8 to 11.3 pmol/L. Figure 3. Open in new tabDownload slide (A) Peak postoperative plasma copeptin in all patients, comparing those in remission with no remission (6.1 pmol/L [4.3-12.1] vs 7.3 pmol/L [5.4-8.4], P = 0.88). (B) ΔCopeptin (preoperative plasma copeptin subtracted from postoperative peak plasma copeptin) in all patients, comparing those in remission with no remission (2.3 pmol/L [-0.5 to 8.2] vs 0.1 pmol/L [-0.1 to 2.2], P = 0.46). Horizontal lines = median. Whiskers = 25th and 75th interquartile ranges. When those who developed DI or SIADH were excluded, there was no difference in peak postoperative copeptin in those in remission vs no remission (10.2 pmol/L [6.9-21.0] vs 5.4 pmol/L [4.6-7.3], P = 0.20). However, because the distribution of the peak postoperative copeptins was borderline normally distributed, parametric testing was also completed for this analysis, which showed a higher peak postoperative copeptin in remission (14.6 pmol/L [±10.9]) vs no remission (5.8 [±1.4], P = 0.03). There was no difference in the Δcopeptin for those in remission vs not in remission (5.1 pmol/L [0.3-19.5] vs 1.1 pmol/L [-0.1 to 2.2], P = 0.39) (Fig. 4). Preoperative copeptin was not different for those in remission (4.7 pmol/L [±2.4]) vs no remission (4.9 pmol/L [±20.3], P = 0.91). There was no association between serum cortisol and plasma copeptin over time postoperatively (Fig. 5). Figure 4. Open in new tabDownload slide (A) Peak postoperative plasma copeptin excluding those who developed DI or SIADH, comparing those in remission with no remission (10.2 pmol/L [6.9-21.0] vs 5.4 pmol/L [4.6-7.3], P = 0.20). (B) ΔCopeptin (preoperative plasma copeptin subtracted from postoperative peak plasma copeptin) excluding those who developed DI or SIADH, comparing those in remission with no remission (5.1 pmol/L [0.3-19.5] vs 1.1 pmol/L [-0.1 to 2.2], P = 0.39). Horizontal lines = median. Whiskers = 25th and 75th interquartile ranges. Figure 5. Open in new tabDownload slide Plasma copeptin and serum cortisol vs postoperative day for patients who did not develop DI or SIADH. Plasma copeptin (indicated by closed circle) and serum cortisol (indicated by “x”). Results shown as (median, 95% CI). All analyses here were repeated adjusting for serum sodium, and there were no differences by remission status for preoperative, peak postoperative, or Δcopeptin for all subjects or after excluding those who developed a water balance disorder (data not shown). Copeptin and Water Balance Disorders As expected, peak postoperative copeptin appeared to be different among patients who developed DI, SIADH, and those without any fluid balance disorder (P = 0.029), whereas patients with DI had lower median peak postoperative copeptin (4.4 pmol/L [2.4-6.9]) than those who developed no fluid abnormality (10.0 pmol/L [5.4-16.5], P = 0.04), the statistical difference was not present after correction for multiple comparisons (P = 0.13). Peak postoperative copeptin of patients with SIADH was 9.4 pmol/L (6.5-10.4) and did not differ from patients with DI (P = 0.32) or those with no fluid abnormality (P = 1.0). There was a difference in Δcopeptin levels among these subgroups (overall P = 0.043), which appeared to be driven by the lower Δcopeptin in those who developed DI (-1.2 pmol/L [-2.6 to 0.1]) vs in those with neither DI or SIADH (3.1 pmol/L [0-9.6], P = 0.05). However, this pairwise comparison did not reach statistical significance, even before correction for multiple comparisons (P = 0.16) (Fig. 6). Preoperative copeptin levels were also not different among the subgroups (P = 0.54). Figure 6. Open in new tabDownload slide (A) Peak postoperative plasma copeptin, comparing those who developed DI, SIADH, or neither (P = 0.029 for comparison of all 3 groups). (B) ∆ Copeptin (preoperative plasma copeptin subtracted from postoperative peak plasma copeptin), comparing those who developed DI, SIADH, or neither (P = 0.043 for comparison of all 3 groups). Horizontal lines = median. Whiskers = 25th and 75th interquartile ranges. Top brackets = pairwise comparisons. P values presented are after Bonferroni correction for multiple comparisons. Association of Sodium and Copeptin Longitudinal data, adjusting for subgroups (ie, DI, SIADH, neither), were analyzed. As expected, there was a group difference (P = 0.003) in serum sodium over time (all DI was missing preoperative serum sodium), with the difference being driven by DI vs SIADH (P = 0.007), and SIADH vs neither (P = 0.012). There was no group difference in plasma copeptin over POD by water balance status (P = 0.16) over time (Fig. 7). There was also no effect by remission status at 3 to 6 months for either serum sodium or plasma copeptin. Figure 7. Open in new tabDownload slide (A) Serum sodium and (B) plasma copeptin by POD and water balance status longitudinal data, adjusting for subgroups (ie, DI, SIADH, neither). Data points at point 0 on the x-axis indicate preoperative values. As expected, there was a group difference (P = 0.003) in serum sodium over time (all with DI were missing preoperative serum sodium), with the difference being driven by DI vs SIADH (P = 0.007), and SIADH vs neither (P = 0.012). There was no group difference in plasma copeptin over POD by water balance status (P = 0.16) over time. Higher serum sodium levels from PODs 1 through 8 itself decreased the odds of remission (OR, 0.56; 95% CI, 0.42-0.73; P < 0.001) in all CD patients. Copeptin levels from these repeated measures adjusting for serum sodium did not correlate with remission status at 3 to 6 months’ follow-up (P = 0.38). There were no differences in preoperative, peak postoperative, or delta sodium levels by remission vs no remission in all patients and in those with no water balance disorders. Discussion AVP and CRH act synergistically to stimulate the secretion of ACTH and ultimately cortisol [1, 2], and there is evidence that glucocorticoids act by way of negative feedback to suppress AVP secretion [10, 11-20]. Therefore, we hypothesized that a greater postoperative increase in plasma copeptin in those with CD in remission after TSS because of resolution of hypercortisolemia and resultant hypocortisolemia, compared with those not in remission with persistent hypercortisolemia and continued negative feedback, would be observed. Although a clear difference in peak postoperative and Δcopeptin was not observed in this study, a higher peak postoperative copeptin was found in those in remission after excluding those who developed DI/SIADH when analyzing this comparison with parametric testing, and it is possible that we did not have the power to detect a difference by nonparametric testing, given our small sample size. Therefore, postoperative plasma copeptin may be a useful early marker to predict remission of CD after TSS. The utility of this test may be limited to those who do not develop water balance disorders postoperatively. If a true increase in copeptin occurs for those in remission after treatment of CD, it is possible that this could be due to the removal of negative feedback from cortisol excess on pre-pro-AVP secretion, as hypothesized in this study. However, it is also possible that other factors may contribute to an increase in copeptin postoperatively, including from the stress response of surgery and postoperative hypocortisolism and resultant stimulation of pre-pro-AVP secretion from these physical stressors and/or from unrecognized SIADH. It was anticipated that more severe hypercortisolism to be negatively correlated with preoperative plasma copeptin because of greater negative feedback on AVP. However, no association was found between preoperative plasma copeptin and markers of severity of hypercortisolism (MN cortisol, AM ACTH, UFC × ULN) in this study. Similarly, we would expect that the preoperative plasma copeptin would be lower compared with healthy individuals. However, comparisons of healthy individuals may be difficult because the fluid and osmolality status at the time of the sample could influence the plasma copeptin, and depending on those factors, copeptin could be appropriately low. A healthy control group with whom to compare the preoperative values was not available for this study, and the thirsted state was not standardized for the preoperative copeptin measurements. Future studies could be considered to determine if preoperative plasma copeptin is lower in patients with CD, or other forms of CS, compared with healthy subjects, with all subjects thirsted for an equivalent period. Further, if preoperative plasma copeptin is found to be lower in thirsted subjects with CS than a thirsted healthy control group, the plasma copeptin could potentially be a diagnostic test to lend support for or against the diagnosis of endogenous CS. In the comparisons of those who developed DI, SIADH, or neither, no difference was found in the Δcopeptin. Peak copeptin was lower in DI compared with those without DI or SIADH (but not different from SIADH). Again, it is possible that there is a lower peak postoperative copeptin and change in copeptin in those with DI, but we may not have had the power to detect this in all of our analyses. These comparisons of copeptin among those with or without water balance disorders postoperatively are somewhat consistent with a prior study showing postoperative copeptin as a good predictor of development of DI, in which a plasma copeptin < 2.5 pmol/L measured on POD 0 accurately identified those who developed DI, and plasma copeptin > 30 pmol/L ruled out the development of DI postoperatively [29]. In the current study, 3 of 6 subjects with DI had a POD 1 plasma copeptin < 2.5 pmol/L, and none had a POD 1 plasma copeptin > 30 pmol/L. However, the study by Winzeler et al found that copeptin measured on POD 0 (within 12 hours after surgery) had the greatest predictive value, and POD 0 plasma copeptin was not available in our study. Further, we used the preoperative, peak, and delta plasma copeptin for analyses, so the early low copeptin levels may not have been captured in our data and analyses. Additionally, this study revealed that increasing levels of serum sodium have lower odds of remission. Those who have an ACTH-producing adenoma that is not identified by magnetic resonance imaging and visual inspection intraoperatively have lower rates of remission and are more likely to have greater manipulation of the pituitary gland intraoperatively [32-36], and the latter may result in greater damage to the pituitary stalk or posterior pituitary, increasing the risk for development of DI and resultant hypernatremia. A higher preoperative copeptin was associated with male sex and increasing BMI SDS. Increasing preoperative copeptin was also found in pubertal boys compared with pubertal girls, with no difference in copeptin between prepubertal boys and girls. It is particularly interesting to note that these associations were only in the preoperative plasma copeptin levels, but not the postoperative peak copeptin or Δcopeptin. Because the association of higher plasma in adult males and pubertal males in comparison to adult females and pubertal females, respectively, have been reported by others [26, 37-40], it raises the question of a change in the association of sex and BMI with plasma copeptin in the postoperative state. An effect of BMI or sex was not found by remission status, so it does not seem that the postoperative hypocortisolemic state for those in remission could explain this loss of association. However, this study may not have been powered to detect this. Strengths of this study include the prospective nature of the study. Further, this is the first study assessing the utility of copeptin to predict remission after treatment of CD. Limitations of this study include the small sample size because of the rarity of the condition, difficulty in clinically diagnosing DI and SIADH, potential effect of post-TSS fluid balance disorders (particularly for those who may have developed transient partial DI or transient SIADH), lack of long-term follow-up, lack of any postoperative follow-up in 11 of the 44 total subjects, as well the observational nature of the study. Further, it is possible that pubertal status, sex, and BMI may have affected copeptin levels, which may have not been consistently detected because of lack of power. Lack of data on the timing of hydrocortisone replacement is an additional limitation of this study because postoperative glucocorticoid replacement could affect AVP secretion via negative feedback. Additional studies are needed to assess to further assess the role of vasopressin and measurement of copeptin in patients before and after treatment of CD. A clear difference in peak postoperative plasma copeptin as an early marker to predict remission of CD after TSS was not found. Further studies with larger sample sizes are needed to further evaluate postoperative plasma copeptin as an early marker to predict remission of CD, though the utility of this test may be limited to those who do not develop water balance disorders postoperatively. Future studies comparing copeptin levels before and after treatment of adrenal CS would be of particular interest because this would minimize the risk of postoperative DI or SIADH which also influence copeptin levels. Additionally, comparison of thirsted preoperative plasma copeptin in those with endogenous CS and thirsted plasma copeptin in healthy controls could potentially provide evidence of whether or not preoperative plasma copeptin is lower in patients with CD, or other forms of CS, compared with healthy subjects. Further, if this is found to be true, it could potentially be a diagnostic test to lend support for or against endogenous CS. Abbreviations AVP arginine vasopressin BMI body mass index CD Cushing disease CS Cushing syndrome DI diabetes insipidus HPA hypothalamic-pituitary-adrenal IQR interquartile range MN midnight OR odds ratio POD postoperative day SDS SD score SIADH syndrome of inappropriate antidiuresis TSS transsphenoidal surgery UFC urinary free cortisol ULN upper limit of normal Acknowledgments The authors thank the patients and their families for participating in this study. Funding This work was supported by the Intramural Research Program, Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD), National Institutes of Health. Disclosures C.A.S. holds patents on technologies involving PRKAR1A, PDE11A, GPR101, and related genes, and his laboratory has received research funding support by Pfizer Inc. for investigations unrelated to this project. C.A.S. is associated with the following pharmaceutical companies: ELPEN, Inc., H. Lunbeck A/S, and Sync. Inc. Clinical Trial Information ClinicalTrials.gov registration no. NCT00001595 (registered November 4, 1999). Data Availability Some or all datasets generated during and/or analyzed during the current study are not publicly available but are available from the corresponding author on reasonable request. Published by Oxford University Press on behalf of the Endocrine Society 2022. This work is written by (a) US Government employee(s) and is in the public domain in the US. From https://academic.oup.com/jes/article/6/6/bvac053/6564309?login=false
  21. Research opportunity for Human Growth Hormone Deficiency caregivers of adolescent patients. This is a 75 min web-assisted phone interview, and the compensation is $125. Please sign up at the link below to receive an email invite to the survey. https://rarepatientvoice.com/CushingsHelp/
  22. Abstract Objective We aimed to perform a systematic review and meta-analysis of all-cause and cause-specific mortality of patients with benign endogenous Cushing's syndrome (CS). Methods The protocol was registered in PROSPERO (CRD42017067530). PubMed, EMBASE, CINHAL, Web of Science and Cochrane Central searches were undertaken from inception to January 2021. Outcomes were the standardized mortality ratio (SMR), proportion and cause of deaths. The I 2 test, subgroup analysis and meta-regression were used to assess heterogeneity across studies. Results SMR was reported in 14 articles including 3,691 patients (13 Cushing's disease (CD) and 7 adrenal CS (ACS) cohorts). Overall SMR was 3.0 (95%CI 2.3-3.9; I 2=80.5%) for all CS, 2.8 (95%CI 2.1-3.7 I 2=81.2%) for CD and 3.3 (95%CI 0.5-6.6; I 2=77.9%) for ACS. Proportion of deaths, reported in 87 articles including 19,181 CS patients (53 CD, 24 ACS, and 20 combined CS cohorts) was 0.05 (95%CI 0.03, 0.06) for all CS subtypes with meta-regression analysis revealing no differences between CS subtypes (P=0.052). The proportion of deaths was 0.1 (10%) in articles published before 2000 and 0.03 (3%) in 2000 until the last search for CS (P<0.001), CD (p<0.001), and ACS (P=0.01). The causes of death were atherosclerotic diseases and thromboembolism (43.4%), infection (12.7%), malignancy (10.6%), active disease (3.5%), adrenal insufficiency (3.0%), and suicide (2.2%). Despite improved outcomes in recent years, increased mortality from CS persists. The causes of death highlight the need to prevent and manage co-morbidities in addition to treating hypercortisolism. Cushing's syndrome, mortality, meta-analysis, causes of death, meta-regression analysis Issue Section: META-ANALYSIS Accepted manuscripts Accepted manuscripts are PDF versions of the author’s final manuscript, as accepted for publication by the journal but prior to copyediting or typesetting. They can be cited using the author(s), article title, journal title, year of online publication, and DOI. They will be replaced by the final typeset articles, which may therefore contain changes. The DOI will remain the same throughout. PDF This content is only available as a PDF. © The Author(s) 2022. Published by Oxford University Press on behalf of the Endocrine Society. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
  23. Osilodrostat is associated with rapid normalization of mean urinary free cortisol (mUFC) excretion in patients with Cushing disease and has a favorable safety profile, according to the results of a study published in the Journal of Clinical Endocrinology & Metabolism. The phase 3 LINC-4 study (ClinicalTrials.gov Identifier: NCT02697734) evaluated the safety and efficacy of osilodrostat, a potent, orally available 11β­-hydroxylase inhibitor, compared with placebo in patients with Cushing disease. The trial, which was conducted at 40 centers in 14 countries, included a 12-week, randomized, double-blind, placebo-controlled period that was followed by a 36-week, open-label osilodrostat treatment period with an optional extension. Eligible patients were aged 18 to 75 years with a confirmed diagnosis of persistent or recurrent Cushing disease after pituitary surgery and/or irradiation or de novo disease, as well as an mUFC level greater than 1.3 times the upper limit of normal (ULN). The patients were randomly assigned 2:1 to osilodrostat 2 mg twice daily or matching placebo, stratified by prior pituitary irradiation. The primary endpoint was the proportion of patients who achieved mUFC ≤ULN at week 12. The key secondary endpoint was the proportion of patients who achieved mUFC ≤ULN at week 36. A total of 73 patients (median age, 39.0 years; 83.6% women) were randomly assigned to either osilodrostat (n=48) or placebo (n=25) and received at least 1 study drug dose from November 2016 to March 2019. The participants had a median (interquartile range [IQR]) time since diagnosis of Cushing disease of 67.4 (26.4-93.8) months. The median treatment duration in the randomized, placebo-controlled period was 12.0 weeks in both the osilodrostat group (IQR, 2.0-13.0 weeks) and the placebo group (IQR, 11.7-13.7 weeks). The proportion of patients who achieved mUFC ≤ULN (≤138 nmol/24 h) at week 12 was significantly increased in those who received osilodrostat (n=37, 77.1%) vs those who received placebo (n=2, 8.0%), with an estimated odds ratio of 43.4 (95% CI, 7.1-343.2) in favor of osilodrostat (P <.0001). A total of 59 patients (80.8%; 95% CI, 69.9-89.1) also achieved the key secondary endpoint of mUFC ≤ULN at week 36, after 24 weeks of open-label osilodrostat. The most frequently occurring adverse events in the placebo-controlled period in the osilodrostat and placebo groups, respectively, were decreased appetite (37.5% vs 16.0%), arthralgia (35.4% vs 8.0%), nausea (31.3% vs 12.0%), and fatigue (25.0% vs 16.0%). A potential study limitation is that although osilodrostat exposure was greater than 1 year among the participants, some adverse effects may take longer to be observed. “This randomized, placebo-controlled trial demonstrates that osilodrostat is a highly effective treatment for Cushing disease, normalizing UFC excretion in 77% of patients after 12 weeks’ treatment,” stated the investigators. “Cortisol reductions were maintained throughout 48 weeks of treatment and were accompanied by improvements in clinical signs of hypercortisolism and quality of life. The safety profile was favorable.” Disclosure: This study was funded by Novartis Pharma AG. Some of the study authors declared affiliations with biotech, pharmaceutical, and/or device companies. Please see the original reference for a full list of disclosures. Reference Gadelha M, Bex M, Feelders RA, et al. Randomized trial of osilodrostat for the treatment of Cushing’s disease. J Clin Endocrinol Metab. Published online March 23, 2022. doi:10.1210/clinem/dgac178 From https://www.endocrinologyadvisor.com/home/topics/general-endocrinology/osilodrostat-effective-for-cushing-disease/
  24. The popular website "How Stuff Work"s is doing a survey of all kinds of diseases and Cushing's is one of them! Share your information and help get the word out to the world in general. (I'm MaryO there, too and I shared about my pituitary surgery and its aftermath. I hope this info helps someone else like these boards and related websites have) The questionnaire is here: https://stuff.health/s/u0A9djA5 Together, we’ll figure out which treatments work best for Cushing's syndrome.
  25. Abstract Background Neuroendocrine tumors can cause ectopic Cushing syndrome, and most patients have metastatic disease at diagnosis. We identified risk factors for outcome, evaluated ectopic Cushing syndrome management, and explored the role of bilateral adrenalectomy in this population. Methods This was a retrospective study including patients with diagnosis of ectopic Cushing Syndrome secondary to neuroendocrine tumors with adrenocorticotropic hormone secretion treated at our quaternary referral center over a 40-year period (1980–2020). Results Seventy-six patients were included. Mean age at diagnosis was 46.3 ± 15.8 years. Most patients (N = 61, 80%) had metastases at ectopic Cushing syndrome diagnosis. Average follow-up was 2.9 ± 3.7 years (range, 4 months–17.2 years). Patients with neuroendocrine tumors before ectopic Cushing syndrome had more frequent metastatic disease and resistant ectopic Cushing syndrome. Patients with de novo hyperglycemia, poor neuroendocrine tumor differentiation, and metastatic disease had worse survival. Of those with nonmetastatic disease, 8 (53%) had ectopic Cushing syndrome resolution after neuroendocrine tumor resection, 3 (20%) were medically controlled, and 4 (27%) underwent bilateral adrenalectomy. In patients with metastatic neuroendocrine tumors, hypercortisolism was initially medically managed in 92%, 3% underwent immediate bilateral adrenalectomy, 2% had control after primary neuroendocrine tumor debulking, and 2% were lost to follow-up. Medical treatment resulted in hormonal control in 7 (13%) patients. Of the 49 patients with metastatic disease and medically resistant ectopic Cushing syndrome, 23 ultimately had bilateral adrenalectomy with ectopic Cushing syndrome cure in all. Conclusion Patients with neuroendocrine tumors before ectopic Cushing syndrome development were more likely metastatic and had worse survival. De novo hyperglycemia and poor neuroendocrine tumor differentiation were predictive of worse prognosis. Medical control of hypercortisolism is difficult to achieve in patients with neuroendocrine tumors–ectopic Cushing syndrome. Well-selected patients may benefit from bilateral adrenalectomy early in the treatment algorithm, and multidisciplinary management is essential in this complex disease. Graphical abstract More information at https://www.surgjournal.com/article/S0039-6060(22)00158-1/fulltext
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