Xinqing Zhu,1,* Abdullah Al-danakh,1,* Yuli Jian,2,* Mohammed Safi,3,* Sijie Luo,1,* Qiwei Chen,1 Shujing Wang,2 Deyong Yang1,4 

1Department of Urology, First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning, People’s Republic of China; 2Department of Biochemistry and Molecular Biology, Institute of Glycobiology, Dalian Medical University, Dalian, Liaoning People’s Republic of China; 3Thoracic/Head and Neck Medical Oncology Department, MD Anderson Cancer Center, Houston, TX, USA; 4Department of Surgery, Healinghands Clinic, Dalian, Liaoning, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Deyong Yang; Shujing Wang, Email yangdeyong@dmu.edu.cn; wangshujing@dmu.edu.cn

Background: Clear cell renal cell carcinoma (ccRCC), the predominant subtype of RCC, is distinguished by unique biological characteristics and heterogeneity, including eosinophilic and clear subtypes. Notwithstanding progress in therapy, immune checkpoint inhibitors (ICIs), and tyrosine kinase inhibitors (TKIs), the prognosis for individuals with metastatic ccRCC remains poor, presumably owing to metabolic alterations leading to mitochondrial dysfunction, which affects treatment response variability.
Methods: We analyzed histological and immunohistochemical data from a cohort at Dalian Medical University’s First Affiliated Hospital alongside RNA-sequencing transcriptome data from the TCGA database. Histologically, eosinophilic and clear ccRCC subtypes were evaluated using Kaplan-Meier and Cox proportional hazards models for survival analysis and prognosis. Differential gene expression (DEG) analysis and Gene Set Enrichment Analysis were performed to explore transcriptomic differences and relevant pathways.
Results: The study discovered substantial histological and molecular differences between the eosinophilic and clear cell subtypes of ccRCC. The eosinophilic subtype linked with frequent high-grade tumors (69.05% eosinophil vs 35.35% clear) and a poorer prognosis (HR=2.659, 95% CI:1.437– 4.919, P=0.002). DEG analysis revealed distinct expression patterns among subtypes and identified a risk score signature that remained significant even after adjusting for clinical variables (HR=3.967, 95% CI: 1.665– 9.449, P=0.002), showing less favorable survival in the high-risk group (P < 0.0001). RRM2 emerged as the most prognostic gene from this risk score, particularly in the eosinophilic subtype, alongside other clinical variables. By IHC, RRM2 shows high IHC score in eosinophilic compared to clear subtype (P=0.019). In addition, highly expressed RRM2 correlates with poor outcomes and is linked to mitochondrial genes, immunological pathways, and ICIs treatment.
Conclusion: These findings show significant differences in prognosis between subtypes. RRM2 was the most prognostic gene from the discovered novel risk score signature associated with subtypes. Future research is essential to validate these insights and their therapeutic implications for ccRCC management.

Keywords: clear cell renal cell carcinoma subtypes, metabolic reprogramming. mitochondria, tumor microenvironment, RRM2, immune checkpoint inhibitors