RUNX2 Rearrangement as a Recurrent Molecular Event and Diagnostic Marker in Salivary Gland Keratocystoma

Keratocystoma is considered a rare, benign salivary gland tumor predominantly occurring in the parotid gland. Owing to its rarity, insufficiently recognized clinicopathological features, immunohistochemical (IHC) phenotype, and molecular characteristics, keratocystoma remains diagnostically challenging. This study aimed to investigate the clinical, histological, and molecular features of keratocystoma to refine the histological diagnostic boundaries. Nineteen cases of keratocystoma were analyzed using fluorescence in situ hybridization, targeted RNA sequencing, and IHC. Histological mimics, including 20 cases of squamous cell carcinoma, 21 cases of metaplastic Warthin tumor, 20 cases of pleomorphic adenoma, 19 cases of mucoepidermoid carcinoma, 19 cases of necrotizing sialometaplasia, 21 cases of branchial cleft cyst, and 21 cases of epidermoid cyst, were incorporated for IHC comparative analysis. All keratocystoma cases arose in the parotid gland. Of the 19 cases, 13 were female and 6 were male patients. Patient age ranged from 5 to 74 years, with a median age of 29 years. Histologically, the multiple cystic spaces were lined by parakeratinized and/or orthokeratinized stratified squamous epithelium. Interestingly, a distinct granular layer was observed in 2 cases, although only in focal areas. Multinucleated giant cell infiltration, cholesterol clefts, and calcification were noted in 13 cases, 11 cases, and 5 cases, respectively. Involvement of salivary gland parenchyma by solid squamous cell nests was observed in 9 cases. Two cases showed areas in which ductal epithelium appeared to undergo transition to keratocystoma components. Two cases presented with few mitoses in the solid squamous cell nests. One case demonstrated perineural invasion. All keratocystoma cases showed RUNX2 rearrangements. RNA sequencing identified a recurrent IRF2BP2::RUNX2 fusion in all successfully tested cases. It revealed significantly higher rate of RUNX2 protein expression in keratocystoma compared with squamous cell carcinoma, epidermoid cyst, and branchial cleft cyst. Two cases experienced recurrence. These findings confirmed that RUNX2 rearrangements and IRF2BP2::RUNX2 fusions were specific molecular features of keratocystoma. Although molecular testing for RUNX2 gene rearrangement or fusion is diagnostically helpful, RUNX2 IHC is not significantly helpful. Although keratocystoma is benign, recurrence may occur occasionally, warranting close postoperative follow-up.