Histopathology revealed three distinct tumor components (Fig. 1C). The first component was characterized by scant infiltrates of deeply basophilic cells with barely discernible cytoplasm (Fig. 1D). These cells showed nuclear PIT1-immunopositivity (Fig. 1E) and immunonegativity for growth hormone (GH, Fig. 1F) and alpha-subunit (Fig. 1G). CAM5.2-immunostaining revealed fibrous bodies in roughly 80% of tumor cells (Fig. 1H). The Ki-67 proliferative index was less than 1% (Fig. 1I).

The second component consisted of densely packed, mildly pleomorphic cells with basophilic cytoplasm and round nuclei with dense chromatin (Fig. 1J). Strong nuclear TPIT expression (Fig. 1K), diffuse expression of ACTH (Fig. 1L), and strong PAS positivity (Fig. 1M) were present. CAM5.2 showed a diffuse staining pattern (Fig. 1N). The Ki-67 proliferative index was calculated at 3–5% (Fig. 1O).

The third component exhibited nests of monomorphic cells with chromophobic cytoplasm and round, chromatin-dense nuclei (Fig. 1P). These cells showed nuclear SF1 expression (Fig. 1Q), partially strong FSH expression (Fig. 1R), and no definite LH expression (Fig. 1S). CAM5.2 staining pattern was diffuse (Fig. 1T). The Ki-67 proliferative index was less than 1% (Fig. 1U).

Representative punch biopsies from each component were subjected to global DNA methylation profiling. Dimensionality reduction including previously published pituitary reference tumors [1], revealed epigenomic affiliation of the first, second and third components with sparsely granulated somatotroph, corticotroph and gonadotroph PitNETs, respectively (Fig. 1 V). For the first, second, and third component, the brain tumor methylation classifier (v12.8) calculated scores of 0.99, 0.99, and 0.79 for the “pituitary adenoma” methylation classes “subtype STH producing, subclass sparsely granulated”, “subtype ACTH producing”, and “subtype gonadotrophin producing”, respectively. All three tumors exhibited quiet copy number profiles (Fig. 1W–Y).

Three synchronous pituitary neuroendocrine tumors (PitNETs) of distinct lineages, comprising a sparsely granulated somatotroph, a densely granulated corticotroph, and a gonadotroph PitNET.