Introduction: There is no clinically established diagnostic biomarker for renal cell carcinoma (RCC) patients. This study aimed to identify a novel diagnostic biomarker using bacteria-derived DNA (b-DNA) in extracellular vesicles (EVs) of serum. Methods: A total of 88 clear cell RCC patients underwent radical or partial nephrectomy, and 10 healthy controls were enrolled in cohort A. EVs were collected from serum just before surgery and prior to antibiotic administration, and metagenomic sequencing was performed after amplification of 16S ribosomal RNA gene. Linear discriminant analysis effect size was used to compare b-DNA from both groups (Figure 1). Significantly abundant b-DNA in serum EVs of RCC patients were used to establish an index for RCC diagnosis. A total of 32 RCC patients and 16 controls were enrolled in cohort B for validation. Results: In cohort A, three kinds of b-DNA (Bacteroidia, TM7-1 and Sphingomonadales) were significantly abandant in serum EVs of RCC patients (Figure 2). In ROC analysis, the area under the curve (AUC) of each b-DNA was 0.66 (Bacteroidia), 0.68 (TM7-1), and 0.72 (Sphingomonadales), respectively. We established BTS index using three kinds of b-DNA to diagnose RCC patients, and it could distinguish RCC patients from healthy controls (AUC = 0.88, sensitivity 89%, specificity 40%). Logistic regression analysis showed that BTS index was an independent factor for RCC diagnosis both in all patients (Odds ratio [OR] 5.9, 95% Confidence interval [CI] 1.3-26.1, p = 0.02) and in early-stage (pT1) RCC patients (OR 6.0, 95% CI 1.2-29.7, p = 0.03). Finally, also in cohort B, BTS index could distinguish RCC patients from healthy controls (sensitivity 91%, specificity 40%), and it was an independent factor for RCC diagnosis both in all patients (OR 6.0, 95% CI 1.2-28.8, p = 0.03) and in pT1 RCC patients (OR 7.5, 95% CI 1.3-45.5, p = 0.03). Conclusions: B-DNA in serum EVs could become a novel diagnostic biomarker for RCC. SOURCE OF Funding: This work was supported by a KAKENHI grant(22K09469, 22H03213).
