Urine test improves reliability of Cushing’s syndrome diagnosis: Study
It provides additional biomarkers for more accurate interpretation of results
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A urine test that measures cortisol, cortisone, and dexamethasone simultaneously can improve the reliability of 24-hour urinary free cortisol (UFC) testing for diagnosing Cushing’s syndrome, including Cushing’s disease, according to a study
It also helps detect interfering medications and provides doctors with additional biomarkers for more accurate interpretation of results, according to a study by Spanish researchers.
This test, which uses a technique known as liquid chromatography with two stages of mass spectrometry (LC-MS/MS) to separate and measure compounds in a sample, including those present in very small amounts, “would help to avoid delays in the diagnosis/follow-up of [Cushing’s syndrome] or the need for additional tests when UFC results are not entirely conclusive,” researchers wrote.
The study, “Urinary free cortisol analyses: Enhancing their clinical performance in Cushing’s syndrome management by means of LC-MS/MS,” was published in the Clinica Chimica Acta.
Standard testing methods can sometimes produce inaccurate results
Cushing’s syndrome occurs when cortisol levels in the body are elevated. Cortisol is a hormone produced by the adrenal glands above the kidneys. In Cushing’s disease, excess cortisol results from a pituitary tumor that produces and releases adrenocorticotropic hormone, which signals the production of cortisol by the adrenal glands.
One of the main tests used to investigate Cushing’s syndrome is the 24-hour UFC, which measures the amount of cortisol excreted in urine over a full day and provides an estimate of the amount produced by the body. Clinical guidelines recommend it as a first-line test. However, standard testing methods can sometimes produce inaccurate results, making diagnosis more difficult.
Immunoassays are commonly used to measure the 24-hour UFC. These tests use antibodies that bind to cortisol and are popular because they are fast and automated. However, they are not always specific and may also detect similar compounds. This cross-reactivity can lead to misleading or falsely high results, especially in patients taking corticosteroids, which mimic cortisol.
To address these limitations, the researchers developed a new LC-MS/MS-based method. LC separates compounds in a liquid sample based on their chemical properties as they move through a column. After separation, MS identifies and measures compounds according to their mass-to-charge ratio, a molecular fingerprint. Tandem MS, which involves two stages of MS, allows for more detailed results.
The herein presented LC-MS/MS approach not only offers the possibility of discontinuing the use of [immunoassays], but also provides additional biomarkers which are significantly relevant in [Cushing’s syndrome].
The researchers also found that adding a small amount of a compound called ammonium fluoride greatly improved the signal-to-noise ratio, which measures how clearly the target compound can be detected compared with background noise. The signal increased by about 250% to 350% for cortisol and 200% to 300% for cortisone, which is produced from cortisol.
Dexamethasone, a lab-made corticosteroid used in diagnostic tests, was detected in 7.7% of samples from patients with Cushing’s syndrome, including Cushing’s disease. Those samples were excluded because dexamethasone can affect the accuracy of cortisol measurement. Cortisone increased as UFC increased, but the increase slowed once UFC exceeded 60 micrograms/day.
The reference range, which defines the interval of values expected in healthy individuals, is essential for interpreting results. To determine this range, the investigators analyzed urine samples from individuals without known cortisol diseases. By calculating the distribution of measured values, they defined normal limits for urinary cortisol and cortisone, as well as the ratio between cortisol and cortisone.
This ratio differed significantly between healthy individuals and patients with either hypercortisolism (excess cortisol) or hypocortisolism (not enough cortisol). Therefore, “the herein presented LC-MS/MS approach not only offers the possibility of discontinuing the use of [immunoassays], but also provides additional biomarkers which are significantly relevant in [Cushing’s syndrome],” the researchers wrote.
