Repurposing public sarcoma multi-omics for neoantigen discovery

Background : Soft tissue sarcomas, particularly Complex Karyotype Sarcomas (CKS), are characterized as "immunologically cold" malignancies driven by structural instability rather than a high tumor mutational burden (TMB). Public “legacy” cohorts are a useful resource to uncover immunotherapy biomarkers. This study used the Whole Exome Sequencing (WES) and RNA-sequencing of CKS patients, to overcome technical limitations and to identify and prioritize neoantigens. Methods : The systematic immunogenomic reanalysis was performed on a landmark cohort of CKS patients (Kim et al., 2018) with a custom bioinformatics workflow which was developed to uncover interpretable immunogenomic signals. This approach consisted of: (1) defining a quality-controlled "callable territory" and normalizing TMB metrics respectively; (2) utilizing RNA-seq not only for expression filtering but as an orthogonal validation check for variant transcription and to distinguish functional amplifications from technical depth artifacts; and (3) applying a multi-modal epitope prediction pipeline to identify and prioritize high-affinity neoantigens derived from both somatic SNVs indels and expressed gene fusions. Results : The reanalysis shows that standard genome-wide metrics frequently underestimated the immunogenic potential. Normalizing the TMB refined the identification of hypermutated and microsatellite instability-like phenotypes. Furthermore, integration of transcriptomic data facilitated the recovery of actionable targets in "low-TMB" tumors. A subset of fusion-derived peptides demonstrated predicted binding affinities competitive with SNV-derived candidates. Conclusion : This study illustrates that technically constrained multi-omic datasets can be systematically re-analyzed to identify potential therapeutic targets. These data argue for looking beyond aggregate biomarkers; patient-specific, expressed neoepitopes may exist even in sarcomas typically described as immunologically “cold”.