Systematic assessment of histomorphologic and immunohistochemical features in metastatic castration-resistant prostate cancer biopsies

Background: Metastatic castration-resistant prostate cancer (mCRPC) represents a heterogeneous disease spectrum. While many tumors remain androgen receptor (AR)-driven, others undergo lineage plasticity toward neuroendocrine prostate cancer (NEPC). The significance of intermediate or mixed phenotypes-such as tumors with co-expression of AR and neuroendocrine (NE) markers or lacking all lineage markers-remains poorly defined. We sought to systematically characterize histomorphology and immunohistochemistry (IHC) in real-world mCRPC biopsies. Methods: 117 mCRPC biopsies from two institutions were retrospectively reviewed and classified by tumor morphology on H&E. IHC was performed for NKX3.1, AR, synaptophysin (SYP), insulinoma-associated protein 1 (INSM1), and Ki-67. Expression was semi-quantitatively scored using H-scores (0-300), and Ki-67 was reported as the percentage of tumor nuclei with positive staining. Results: Histologic distribution included 44 adenocarcinomas, 53 poorly differentiated carcinomas, 15 NEPC, and 5 mixed subtypes. Using H-score >40 as a cutoff, adenocarcinomas frequently expressed NKX3.1 (93%) and AR (100%), with limited NE marker expression (13% for INSM1 and 46% for SYP). Poorly differentiated carcinomas most often retained NKX3.1 (80%) and AR (84%), while NE marker expression was less common (SYP 27%, INSM1 26%). Mixed tumors demonstrated co-expression across lineages, (AR 100%, NKX3.1 100%, SYP 80%, INSM1 50%). NEPC showed diffuse NE differentiation (SYP 85%, INSM1 75%) with rare NKX3.1 or AR positivity (8% and 0%, respectively). High H-scores (>200) were enriched in NKX3.1 (83%) and AR (92%) positive adenocarcinomas, while majority NEPC showed high SYP expression and high Ki-67 proliferation index. Low H-score (<40) are seen across all four IHC markers in 1 poorly differentiated carcinoma and 2 NEPC. Conclusions: This study provides one of the first systematic, standardized assessments of mCRPC biopsy morphology and IHC marker expression. NKX3.1 and AR predominated in adenocarcinoma and poorly differentiated carcinomas, whereas SYP and INSM1 reliably identified neuroendocrine carcinomas. Mixed subtypes exhibited overlapping lineage features. Ki-67 indicated differences in tumor proliferation across subtypes. These findings highlight the morphologic and immunophenotypic heterogeneity of mCRPC and emphasize the need for integrated pathologic evaluation to refine diagnosis and support translational research.