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Low-Coverage Exome Sequencing Screen in Formalin-Fixed Paraffin-Embedded Tumors Reveals Evidence of Exposure to
Xavier Castells1, Sandra Karanović2, Maude Ardin1
1Molecular Mechanisms and Biomarkers Group, International Agency for Research on Cancer, Lyon, France.
Background:
Dietary exposure to cytotoxic and carcinogenic aristolochic acid (AA) causes severe nephropathy typically associated with urologic cancers. Monitoring of AA exposure uses biomarkers such as aristolactam-DNA adducts, detected by mass spectrometry in the kidney cortex, or the somatic A>T transversion pattern characteristic of exposure to AA, as revealed by previous DNA-sequencing studies using fresh-frozen tumors.
Methods:
Here, we report a low-coverage whole-exome sequencing method (LC-WES) optimized for multisample detection of the AA mutational signature, and demonstrate its utility in 17 formalin-fixed paraffin-embedded urothelial tumors obtained from 15 patients with endemic nephropathy, an environmental form of AA nephropathy.
Results:
LC-WES identified the AA signature, alongside signatures of age and APOBEC enzyme activity, in 15 samples sequenced at the average per-base coverage of approximately 10×. Analysis at 3 to 9× coverage revealed the signature in 91% of the positive samples. The exome-wide distribution of the predominant A>T transversions exhibited a stochastic pattern, whereas 83 cancer driver genes were enriched for recurrent nonsynonymous A>T mutations. In two patients, pairs of tumors from different parts of the urinary tract, including the bladder, harbored overlapping mutation patterns, suggesting tumor dissemination via cell seeding.
Conclusions:
LC-WES analysis of archived tumor tissues is a reliable method applicable to investigations of both the exposure to AA and its biologic effects in human carcinomas.
Impact:
By detecting cancers associated with AA exposure in high-risk populations, LC-WES can support future molecular epidemiology studies and provide evidence-base for relevant preventive measures.
Insights
Low-coverage whole-exome sequencing (LC-WES) detects aristolochic acid (AA) exposure signatures in archived urothelial tumors. This method aids in identifying AA-related cancers for epidemiological studies and preventive measures.
Area of Science:
- Genomics and Molecular Epidemiology
- Cancer Research
- Toxicology
Background:
- Aristolochic acid (AA) exposure causes nephropathy and urologic cancers.
- Biomarkers for AA exposure include DNA adducts and specific mutation patterns.
- Previous detection methods relied on fresh-frozen tumor samples.
Purpose of the Study:
- To develop and validate a low-coverage whole-exome sequencing (LC-WES) method for detecting the AA mutational signature.
- To assess the utility of LC-WES in formalin-fixed paraffin-embedded (FFPE) urothelial tumors from patients with endemic nephropathy.
- To investigate the potential for tumor dissemination in patients with overlapping mutation patterns.
Main Methods:
- Utilized a novel low-coverage whole-exome sequencing (LC-WES) approach.
- Applied LC-WES to 17 FFPE urothelial tumor samples from 15 patients.
- Optimized LC-WES for multisample detection of the AA mutational signature.
Main Results:
- LC-WES successfully identified the AA mutational signature in 91% of positive samples, even at low coverage (3-9×).
- The AA signature was detected alongside age and APOBEC signatures.
- Analysis revealed enrichment of recurrent A>T mutations in 83 cancer driver genes and suggested tumor dissemination in two patients.
Conclusions:
- LC-WES is a reliable method for analyzing archived tumor tissues to investigate AA exposure and its biological effects.
- This technique supports molecular epidemiology studies and informs preventive strategies for AA-related cancers in high-risk populations.
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