Is it best to collect the dexamethasone suppression test (DST) and adrenocorticotropic hormone (ACTH) level on different days when evaluating for hypercortisolism or Cushing's?

Timing of DST and ACTH Measurement in Cushing's Evaluation

ACTH levels should NOT be measured on the same day as the dexamethasone suppression test (DST)—these are separate diagnostic steps performed sequentially, not simultaneously. 1

The Correct Diagnostic Sequence

The diagnostic algorithm for Cushing's syndrome follows a clear two-stage approach:

Stage 1: Confirm Hypercortisolism

• The overnight 1mg DST is performed by administering 1mg dexamethasone between 11pm-midnight, followed by measuring only serum cortisol at 8am the next morning 1
• Normal cortisol suppression is defined as <1.8 μg/dL (50 nmol/L), which strongly predicts absence of Cushing's syndrome 2, 1
• ACTH should not be measured during the initial overnight DST, as this test is designed specifically to assess cortisol suppression only 1
• Alternative first-line screening tests include late-night salivary cortisol (LNSC) or 24-hour urinary free cortisol (UFC), but none of these involve ACTH measurement 2, 3

Stage 2: Determine ACTH Dependency (After Confirming Hypercortisolism)

• Once Cushing's syndrome is confirmed through abnormal DST and/or other screening tests, then proceed to measure ACTH levels 2, 1
• Morning (08:00-09:00h) plasma ACTH measurement is optimal for this assessment 4
• ACTH >5 ng/L indicates ACTH-dependent Cushing's syndrome, while low/undetectable ACTH indicates ACTH-independent disease 4, 3
• Using a cut-off of 29 ng/L provides 70% sensitivity and 100% specificity for diagnosing Cushing's disease 4, 3

Why This Sequential Approach Matters

Measuring ACTH during the overnight DST is not part of standard protocol and may lead to misinterpretation 1. The physiologic rationale is clear:

• The DST evaluates the hypothalamic-pituitary-adrenal axis's ability to suppress cortisol production in response to exogenous glucocorticoid 2
• ACTH levels during dexamethasone suppression do not provide diagnostically useful information and can confuse the clinical picture 5
• Research demonstrates that adding plasma ACTH levels to the DST does not improve diagnostic accuracy compared to measuring cortisol alone 5

Common Pitfalls to Avoid

• Do not order ACTH and DST simultaneously—this wastes resources and creates interpretive confusion 1
• False positive DST results can occur with rapid dexamethasone absorption/malabsorption, CYP3A4 inducers, or increased corticosteroid binding globulin from oral estrogens 2, 1
• In patients with cyclical Cushing's disease, hypercortisolemia must be confirmed immediately prior to any diagnostic procedures to ensure active disease phase 4, 3

Practical Implementation

The algorithmic approach is:

1. Day 1: Perform overnight 1mg DST (measure cortisol only at 8am) 1
2. If cortisol ≥1.8 μg/dL: Repeat screening test or use alternative screening (UFC, LNSC) to confirm hypercortisolism 2, 3
3. After confirming hypercortisolism: Obtain morning (08:00-09:00h) plasma ACTH on a separate day 4
4. Based on ACTH results: Proceed to imaging (pituitary MRI if ACTH-dependent, adrenal CT/MRI if ACTH-independent) 2, 4

This sequential approach maximizes diagnostic accuracy while avoiding the interpretive errors that occur when these tests are inappropriately combined 1, 5.