USP8 Mutations in Pituitary Cushing Adenomas—Targeted Analysis by Next-Generation Sequencing

Abstract

Gain-of-function somatic mutations in the ubiquitin specific protease 8 (<i>USP8</i>) gene have recently been reported as a cause of pituitary adenomas in Cushing disease. Molecular diagnostic testing of tumor tissue may aid in the diagnosis of specimens obtained through therapeutic transsphenoidal surgery; however, for small tumors, availability of fresh tissue is limited, and contamination with normal tissue is frequent. We performed molecular testing of DNA isolated from single formalin-fixed and paraffin-embedded (FFPE) tissue sections of 42 pituitary adenomas from patients with Cushing disease (27 female patients and 15 male patients; mean age at surgery, 42.5 years; mean tumor size, 12.2 mm). By Sanger sequencing, we identified previously reported <i>USP8</i> missense mutations in six tumors. Targeted next-generation sequencing (NGS) revealed known or previously undescribed missense mutations in three additional tumors (two with two different mutations each), with mutant allele frequencies as low as 3%. Of the nine tumors with <i>USP8</i> mutations (mutation frequency, 21.4%), seven were from female patients (mutation frequency, 25.9%), and two were from male patients (mutation frequency, 13.3%). Mutant tumors were on average 11.4 mm in size, and patients with mutations were on average 43.9 years of age. The overall <i>USP8</i> mutation frequency in our cohort was lower than in previously described cohorts, and we did not observe <i>USP8</i> deletions that were frequent in other cohorts. We demonstrate that testing for <i>USP8</i> variants can be performed from small amounts of FFPE tissue. NGS showed higher sensitivity for <i>USP8</i> mutation detection than did Sanger sequencing. Assessment for <i>USP8</i> mutations may complement histopathological diagnosis.