Whole-proteome phage immunoprecipitation sequencing reveals germ cell tumor–specific immunosignature

Germ cell tumors (GCTs) pose significant diagnostic challenges because of the limited performance of existing tumor markers. Here, we used phage immunoprecipitation sequencing (PhIP-Seq) to develop a unique immunosignature panel to improve diagnosing and differentiating GCT. Using 427 serum samples (150 GCT, 277 controls), we developed and validated an immunosignature panel (GCT-iSIGN) comprising 24 peptides from 16 unique proteins. This panel achieved 93% sensitivity, 99% specificity, and an area under the curve (AUC) of 0.98, identifying 23/24 biomarker-negative GCT cases. A secondary model (Sem-iSIGN), consisting of 17 peptides from five proteins, differentiated seminoma from nonseminoma with 96% specificity, 65% sensitivity, and AUC of 0.77. RNA sequencing data from The Cancer Genome Atlas confirmed differential overexpression of target antigens in testicular cancer. ELISA validation of ERVK7 and LUZP4 and immunohistochemical detection of ERVK7, MUC4, ZNF91, and LUZP4 in tumor tissues supported target expression. This study highlights PhIP-Seq immunoprofiling to identify serum-based immunosignature panels that can serve as biomarkers for GCTs. This approach addresses the shortcomings of conventional markers and offers a scalable, cost-effective tool for improving cancer diagnosis and management.