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Neuroendocrinology and Pituitary

SUN-011 - Usp8 S680Del Knock-in Mice Recapitulate Endocrinological and Clinical Features of Cushing'S Disease

Sunday, June 14, 2026
9:00 AM - 4:00 PM CT
Location: ENDOExpo; Poster Floor
Disclosure(s):
Masaki Kobatake, MD: No financial relationships to disclose


Background: Cushing’s disease (CD), caused by ACTH-producing pituitary tumors, is associated with a poor prognosis. Somatic USP8 pathogenic variants (PVs) occur in approximately 30% and are confined to the 14-3-3 binding motif. The USP8 PVs are associated with smaller tumors, female predominance, and a high recurrence rate. These PVs enhance deubiquitinase activity and upregulate POMC expression; however, the underlying mechanisms remain unclear due to the lack of appropriate experimental models. Although germline USP8 PVs have been reported in pediatric patients with CD, it remains uncertain whether USP8 variants function as driver mutations. We aimed to generate USP8 PV knock-in (KI) mice as a model to elucidate CD and facilitate therapeutic development.

Methods: The murine orthologous variant S680del (the common human USP8 PV S718del) was introduced into the genome of B6C3F1 zygotes using the CRISPR–Cas9 system. Heterozygous KI mice were generated by crossing founders with wild-type (WT) mice. At 9–20 weeks of age, body weight, forelimb grip strength, blood glucose levels during an intraperitoneal glucose tolerance test, and plasma corticosterone (Cort) levels before and after dexamethasone (Dex) administration were measured. Primary pituitary cultures were established from these mice, and supernatant ACTH levels were quantified. Furthermore, FFPE pituitary sections were subjected to H&E, silver impregnation, and ACTH immunostaining. The pituitary tissues were independently evaluated by three genotype-blinded reviewers.

Results: Compared with WT, KI of both sexes exhibited lower body weight, reduced forelimb grip, and higher blood glucose levels at 30 min after glucose administration. In females, KI showed higher Cort before Dex (1,581 vs 1,112 pg/mL, p< 0.05) and after Dex (515 vs 154 pg/mL, p=0.02). In males, KI also demonstrated significantly higher Cort levels before Dex (1,381 ± 638 vs 873 ± 489 pg/mL, p=0.03) and tended to be higher after Dex (211 ± 325 vs 52 ± 69 pg/mL, p=0.11). ACTH concentrations in culture supernatants were higher in KI (69.7 vs 41.7 pg/mL, p< 0.01). No pituitary tumors were observed; however, ACTH-positive cells with reticulin disruption were detected in 75% of KI and 25% of WT (p=0.25).

Conclusions: USP8 PVs in mice induced pituitary ACTH hypersecretion and a biochemical phenotype consistent with CD, providing robust in vivo evidence supporting their role as disease drivers. The absence of overt tumors may reflect a feature of human CD, in which USP8 PVs lead to excessive ACTH secretion despite the presence of only small lesions. Furthermore, the phenotypes in both sexes suggest that the female predominance in human CD may stem from sex-dependent susceptibility to somatic tumorigenesis rather than the secretory effects. This genetically faithful model provides a valuable platform for mechanistic studies and preclinical drug screening.

*Unless otherwise noted, all abstracts presented at ENDO must not be released to the press or the public until the date and time of presentation. For oral presentations, the abstracts are embargoed until the session begins. The Endocrine Society reserves the right to lift the embargo on specific abstracts that are selected for promotion prior to or during ENDO.*