Simultaneous expression of CD5 and INSM1 may distinguish parotid CASTLE from other primary tumors with a squamous phenotype.

Parotid carcinoma showing thymus-like element (CASTLE) is positive for the immunohistochemical markers CD5 and CD117. However, the specificity of the markers has not been evaluated in parotid tumors with a squamous phenotype. This study aims to validate the efficiency of CD5, CD117, and INSM1 in distinguishing CASTLE from other parotid tumors with a squamous phenotype, including squamous cell carcinoma (SCC, n = 4), lymphoepithelial carcinoma (LEC, n = 5), and pleomorphic adenoma with squamous differentiation (PA-SD, n = 3). The parotid CASTLE exhibited diffusely CD5 membranous positivity (Immunoreactive Score, IRS = 12) and moderate INSM1 nuclear expression (IRS = 6), while SCC, PA-SD, and LEC showed negligible CD5 (IRS ≤ 2) and weak/focal INSM1 (IRS ≤ 3) expression. Although CD117 was diffusely positive in CASTLE, it was moderately to strongly expressed in the LECs (IRS = 9), limiting its diagnostic utility in differential diagnosis. EBER in situ hybridization (ISH) confirmed EBV association in all LECs but not in the CASTLE. In summary, simultaneous CD5 and INSM1 expression may serve as a distinct feature for parotid CASTLE, facilitating differential diagnosis in salivary gland pathology. In addition, EBER ISH remains essential to exclude LEC.