Krasic Jure, Soic Dinko, Abramovic Lucija Skara, Kolosnjaj Ivona, Tomic Miroslav, Vrtaric Alen, Oguic Sasa Kralik, Gelo Nina, Bojanac Ana Katusic, Jezek Davor, Mitrecic Dinko, Ulamec Monika, Kulis Tomislav, Gornik Olga, Sincic Nino
Department of Histology and Embryology, School of Medicine, University of Zagreb, Šalata 3, 10000, Zagreb, Croatia.
Laboratory for Stem Cells, Croatian Institute for Brain Research, School of Medicine, University of Zagreb, 10000, Zagreb, Croatia.
Cancer Cell Int. 2025 Jul 11;25(1):257. doi: 10.1186/s12935-025-03887-8.
Nonseminomatous testicular germ cell tumors (NSE) present significant diagnostic challenges, especially for the early detection of serum tumor marker (STM) negative cases. Current diagnostic tools are limited, highlighting the need for innovative approaches. This study investigates a novel multi-analyte approach combining circulating cell-free DNA (cfDNA) and N-glycan profiling in both blood and seminal plasma to improve NSE diagnostics.
The study included 41 NSE patients and 114 healthy controls. Diagnostic potential of cfDNA parameters (quality and fragmentation), cfDNA methylation (RASSF1A, PRSS21, and LINE-1) and N-glycan alterations in blood plasma and seminal plasma samples was investigated using logistic regression models. Pre- and post-radical orchidectomy longitudinal samples from NSE patients were analyzed to assess surgical treatment response and disease monitoring utility.
Blood plasma analysis of combined cfDNA and N-glycan profiling demonstrated high diagnostic precision, with an AUC of 0.96, identifying 85% of STM-negative patients and all pure-form teratomas. Post-operative blood plasma analysis showed that LINE-1 cfDNA methylation levels returned to those of healthy controls. Seminal plasma analysis revealed an increased cfDNA fragmentation index (CFI) and cfDNA methylation changes in LINE-1 and PRSS21, with an AUC of 0.83 and identifying 85% of STM-negative patients.
The proposed multi-analyte approach significantly improves early diagnostics of NSE, particularly for STM-negative cases and teratomas. LINE-1 cfDNA methylation is a promising biomarker for NSE detection and treatment monitoring. These findings could transform diagnostic strategies and patient management in testicular germ cell tumors, with potential applications in reducing overtreatment and improving outcomes .
非精原性睾丸生殖细胞肿瘤（NSE）带来了重大的诊断挑战，尤其是对于血清肿瘤标志物（STM）阴性病例的早期检测。当前的诊断工具有限，凸显了创新方法的必要性。本研究调查了一种新颖的多分析物方法，该方法结合循环游离DNA（cfDNA）以及血液和精浆中的N-聚糖谱分析，以改善NSE的诊断。
该研究纳入了41例NSE患者和114名健康对照。使用逻辑回归模型研究了cfDNA参数（质量和片段化）、cfDNA甲基化（RASSF1A、PRSS21和LINE-1）以及血浆和精浆样本中N-聚糖改变的诊断潜力。对NSE患者根治性睾丸切除术前和术后的纵向样本进行分析，以评估手术治疗反应和疾病监测效用。
联合cfDNA和N-聚糖谱分析的血浆分析显示出高诊断精度，曲线下面积（AUC）为0.96，识别出85%的STM阴性患者和所有纯形式畸胎瘤。术后血浆分析表明，LINE-1 cfDNA甲基化水平恢复到健康对照的水平。精浆分析显示cfDNA片段化指数（CFI）增加，以及LINE-1和PRSS21中的cfDNA甲基化变化，AUC为0.83，识别出85%的STM阴性患者。
所提出的多分析物方法显著改善了NSE的早期诊断，特别是对于STM阴性病例和畸胎瘤。LINE-1 cfDNA甲基化是NSE检测和治疗监测的一个有前景的生物标志物。这些发现可能会改变睾丸生殖细胞肿瘤的诊断策略和患者管理，在减少过度治疗和改善治疗结果方面具有潜在应用价值。
