Adrenal Hyperfunction - Cushing Syndrome

Adrenal hyperfunction (Cushing syndrome) causes adrenal tissue to secrete excess cortisol and is manifested by a constellation of symptoms, including central obesity, hypertension, type 2 diabetes, easy bruisability, abdominal striae, and myopathy. The signs and symptoms of high cortisol are common and nonspecific, making the diagnosis challenging.

Quick Answers for Clinicians

Which testing algorithms are related to this topic?


Indications for Testing

  • Resistant diabetes
  • Refractory hypertension
  • Multiple signs/symptoms of Cushing syndrome after ruling out metabolic syndrome or polycystic ovarian syndrome (PCOS)
  • Do not test patients who have hypercortisolism secondary to administration of glucocorticoids (oral, inhaled, injections, topical)

Laboratory Testing

  • Initial testing (Nieman, Endocrine Society, 2008; National Comprehensive Cancer Network [NCCN], 2018)
    • Salivary cortisol (collected between 2300 hours and 0000 hours) – at least two measurements
      • Reflects circadian nadir so elevations are more relevant
      • Samples are stable over time
      • Cutoff values are test and age specific
      • Counsel individual to collect on quiet, restful evening
    • 24-hour urine free cortisol (UFC) – at least two measurements
      • 24-hour urine collection – random measurements not recommended
      • Urinary cortisol levels >3 x upper limit of normal are unequivocal and confirm diagnosis
      • Results not affected by taking oral estrogen
    • Dexamethasone suppression testing (DST) – overnight and 2-day tests are available
      • Does not interfere with cortisol tests
      • Other glucocorticoids will influence cortisol levels and must be discontinued prior to testing
        • Other hormones that may interfere with cortisol testing – exogenous testosterone, Nature-Throid, estrogens (eg, birth control), megestrol acetate
        • Estrogen-containing drugs should be withdrawn for 6 weeks prior to testing
      • Low-dose (1 mg) DST
        • Patient should take 1 mg dexamethasone between 2300 hours and 0000 hours – should suppress adrenocorticotropic hormone (ACTH) production if hypothalamic-pituitary-adrenal axis (HPA) is normal
        • Serum or urine should be collected between 0800 and 0900 hours and tested for free cortisol
          • ACTH collected at same time may be helpful in case of positive test
        • Dexamethasone is measured to confirm administration and normal metabolism
      • 2-day, 2 mg test – may be used to confirm that ACTH can be suppressed if overnight, low-dose test is equivocal
        • Administer 0.5 mg every 6 hours starting at 0800 for 2 days for a total of eight doses (2 mg)
        • Collect serum for cortisol 2-6 hours after last dose administered
        • Collecting ACTH at same time will assist with etiology in case of positive test
        • Collect dexamethasone to confirm administration and normal metabolism
        • Urinary corticosteroid excretion on day two of dexamethasone administration has been used (compare to baseline day urinary corticosteroid excretion)
          • Not currently recommended, but literature often cites concentrations
      • To confirm elevated results – repeat either of the DST tests or the cortisol tests
        • If high suspicion remains – use high-dose dexamethasone (8 mg) overnight test
          • Similar protocol to low-dose overnight dexamethasone suppression test
          • Negative suppression testing – suggests etiology other than Cushing syndrome
  • Follow-up testing (Nieman, Endocrine Society, 2008; NCCN, 2018)
    • Endocrinology referral
    • Serum ACTH in conjunction with suppression tests – aid in determining whether disease is adrenal, pituitary, or ectopic ACTH based
      • Suppressed ACTH (<10 pg/mL) – suggests etiology is ACTH independent and pathology is adrenal (adenoma, carcinoma)
      • High ACTH (>20 pg/mL) – suggests ectopic ACTH
      • Intermediate ACTH (10 pg/mL – 20 pg/mL) – likely Cushing syndrome and magnetic resonance imaging (MRI) should be done to visualize tumor
        • ACTH-producing tumors are often not visualized (microadenoma)
        • Bilateral inferior petrosal sinus sampling is indicated  – testing is technically difficult and should only be performed in a high-volume center (Pappachan, 2017)
          • Petrosal ACTH to serum ACTH ratio is measured before and after corticotropin stimulation
          • Absence of change in gradient – suggestive of ectopic ACTH-producing tumor
          • Significant increase in pre- and poststimulation – expected in Cushing syndrome
  • Mechanisms of potential interference with test evaluation when diagnosing Cushing syndrome and selected drugs that may cause these effects (Nieman, Endocrine Society, 2008)
    • CYP3A4 and dexamethasone metabolism
      • Accelerate dexamethasone metabolism by induction of CYP3A4
        • Phenobarbital
        • Phenytoin
        • Carbamazepine
        • Primidone
        • Rifampin
        • Rifapentine
        • Ethosuximide
        • Pioglitazone
      • Impair dexamethasone metabolism by inhibition of CYP3A4
        • Aprepitant/fosaprepitant
        • Itraconazole
        • Ritonavir
        • Fluoxetine
        • Diltiazem
        • Cimetidine
    • Increase cortisol-binding globulin, therefore falsely elevate cortisol results
      • Estrogens
      • Mitotane
    • Increase urine free cortisol results
      • Carbamazepine – increase
      • Fenofibrate – increase if measured by high-performance liquid chromatography (HPLC)
      • Some synthetic glucocorticoids – immunoassays
    • Drugs that inhibit 11β-HSD2 – licorice, carbenoxolone​


  • Definitive solid tumor diagnosis requires biopsy and pathologist examination
  • Useful immunohistochemical stains may include ACTH by immunohistochemistry
  • For detailed descriptions, refer to ARUP’s Immunohistochemistry Stain Offerings

Imaging Studies

  • MRI pituitary with inferior bilateral petrosal sampling of ACTH – differentiate between Cushing syndrome and ectopic source (if necessary)
    • If ACTH is low – MRI of adrenal glands
    • If ACTH >20 pg/mL – search for ectopic ACTH-secreting tumor
  • Computed tomography (CT) adrenal protocol or MRI adrenals – identify unilateral/bilateral tumor or hyperplasia


  • Untreated Cushing syndrome – associated with excess morbidity/mortality secondary to cardiovascular disease
  • If tumor is benign and removed – mortality associated with comorbid diseases developed in association with tumor may normalize
  • Malignant tumors producing ACTH have a poor prognosis

Differential Diagnosis


  • Following definitive treatment (surgery of adrenals or pituitary, medical adrenal suppression, radiation therapy), consider monitoring (Nieman, Endocrine Society, 2015)
    • Cortisol – for hypocortisolism or hypercortisolism
    • Pituitary hormone (refer to Hypopituitarism)
    • Serum sodium – 5-14 days following transsphenoidal surgery



  • Incidence – 2-3/100,000 (Nieman, Endocrine Society, 2008)
  • Age – uncommon in children; peaks in 20s-50s
  • Sex – M<F, 1:4-6


  • Endogenous
    • Pituitary (Cushing syndrome)
      • Hyperplasia
      • Adenoma, often microadenoma
    • Adrenal
      • Adenoma
      • Carcinoma
        • Most are sporadic tumors
        • Hereditary syndromes (eg, Li-Fraumeni) may present as carcinoma
      • Ectopic production of ACTH – tumors (carcinoid, small cell lung cancer)
    • Exogenous – glucocorticoid administration
  • Genetics (Lacroix, 2015)
    • Pituitary adenoma – USP8, MEN1, CDKNB/p27Kip1, AIP, DICER1
      • Ectopic ACTH secretion – RET, MEN1
      • Bilateral macronodular adrenal hyperplasia – ARMC5, MEN1, FH, GNAS1, PDE11A, PDE88, MC2R, PRKACA
      • Adrenal adenoma – PRKACA, CTNNB1, GNAS1, PRKAR1A
      • Primary pigmented nodular adrenocortical disease – PRKAR1A, PDE11A, PDE8B, PRKACA


  • Corticotropin-releasing hormone (CRH) in the hypothalamus stimulates release of ACTH from the pituitary gland
  • ACTH acts on the adrenal glands to produce cortisol
  • Most endogenous cases are caused by hypersecretion of pituitary ACTH or ectopic production of ACTH from nonpituitary source

Clinical Presentation

  • Centripetal obesity, moon facies, buffalo hump, hirsutism, reddish-purple striae
  • Hypertension
  • Menstrual abnormalities (eg, amenorrhea)
  • Irritability, impaired memory
  • Osteoporosis
  • Fatigue, weakness
  • Proximal myopathy
  • Impaired glucose tolerance

ARUP Laboratory Tests

Primary Tests

Rule out Cushing syndrome

Do not use for patients taking glucocorticoids

Rule out Cushing syndrome

Do not use for patients taking glucocorticoids

Differential diagnosis of Cushing syndrome

Screen and diagnose primary and secondary adrenal insufficiency

Related Tests

Compliance assessment of dexamethasone suppression testing

Screen, diagnose, and monitor diseases associated with excess or deficient cortisol production

Aid in diagnosing adrenal insufficiency and determining the presence of anterior pituitary tumors

Screen, diagnose, and monitor diseases associated with excess or deficient cortisol production

Screen, diagnose, and monitor diseases associated with excess or deficient cortisol production

Screen, diagnose, and monitor diseases associated with excess or deficient cortisol production

Aid in histologic diagnosis of adrenal hyperfunction

Stained and returned to client pathologist for interpretation; consultation available if needed

Medical Experts



Brittany A. Young, MD, PhD
Assistant Professor of Pathology (Clinical), University of Utah
Co-Director of Clinical Laboratories at University Hospital, ARUP Laboratories


Additional Resources