Background : Diagnosis of pituitary-dependent hyperadrenocorticism (PDH) in cats is challenging because there is no specific diagnostic test. Hypothesis/Objective : The determination of plasma ACTH precursor (POMC and pro-ACTH) concentration might facilitate the diagnosis of PDH in cats. The aim of the study was to evaluate prospectively the plasma concentrations of ACTH precursors in a small cohort of cats with PDH and to estimate the value of this approach for diagnosis. Animals Four groups of cats were included: group 1 (cats with PDH), group 2 (cats with diabetes mellitus but not hyperadrenocorticism (HAC)), group 3 (cats with diabetes mellitus and confirmed acromegaly but not HAC), and group 4 (healthy cats). Methods : PDH diagnosis was based on clinical data, low-dose dexamethasone suppression test (LDDST), and adrenal and pituitary gland computed tomography (CT) scan. For groups 2, 3, and 4, hyperadrenocorticism was excluded by LDDST or urine cortisol:creatinine ratio (UCCR). An immunoluminometric assay was used to determine plasma concentrations of ACTH precursors in the 4 groups of cats. Results : Group 1 contained 9 cats (enlarged pituitary gland in 7/9). Plasma ACTH precursor concentrations ranged from <53 to >1010 similar to pmol/L with 8/9 concentrations =229 similar to pmol/L. Groups 2, 3, and 4 included 13, 7, and 13 cats, respectively. Plasma ACTH precursor concentrations ranged from <53 to 96 similar to pmol/L in group 2, <53 to 72 similar to pmol/L in group 3, and <53 to 99 similar to pmol/L in group 4. Conclusion and Clinical Importance : High plasma concentration of ACTH precursors in cats (>100 similar to pmol/L) is highly suggestive of PDH.