Genomic Analysis Identifies Renal Cell Carcinoma as a New Tumor Type Linked to Aristolochic Acid Exposure

Background: Dietary intake of nephrotoxic and carcinogenic aristolochic acid (AA) leads to endemic nephropathy (EN) marked by chronic tubulointerstitial nephropathy (CTN) and urinary tract transitional cell carcinomas (TCC). The role of AA in malignancies other than TCC is unexplored. We aimed to investigate a role of AA in the etiology of renal cell carcinomas (RCC), usually not related to EN, by conducting whole exome sequencing (WES) of tumor-enriched DNAs to detect the presence of the AA mutational signature. Methods: Four clear cell RCC patients from EN area linked to AA exposure and 1 non-EN RCC case were studied. Of the EN patients, 3 were farmers baking bread from grain from locally grown wheat ; 1 patient was diagnosed with CTN and 2 with TCC. All patients were in CKD stages≥3b.Tumor DNA was isolated by macrodissection from FFPE sections, processed for WES libraries, exome capture and sequencing on Illumina HiSeq2500 (multiplexed paired-end 50 bp run, >10x coverage/sample). Reads were aligned by BWA, variants called by GATK, annotated by ANNOVAR and genetic variants observed in normal population removed. The detection of the AA signature was performed by customized R functions. Results: High A:T>T:A transversion rates (0.9 to 1.8 /Mb) were found in all EN RCC samples. Three cases had A:T>T:A (48.1%, 38.2% and 21.6%) and 1 tumor was borderline positive (15.2% A:T>T:A). The non-EN RCC control was negative (6.4% A:T>T:A). In the positive samples the mutations occurred mainly in the CAG context and on the coding strand, meeting all definition criteria for the AA mutational signature. Conclusions: Our study finds AA mutational signature in 3 EN patients with RCC, suggesting a possible causal involvement of AA in the RCC etiology, with new implications for the worldwide RCC incidence due to widespread unregulated use of AA-containing herbs. Extension of this study is underway to validate the preliminary results in a larger cohort of cases. Supported by grant 04-38 Cro Nat Foundation