The genomic landscape of cancer is characterized by dozens of mutations. Currently, massive parallel sequencing allows sequencing of hundreds of genetic hotspots in tens of genes that may provide prognostic and predictive information.
As a validation step we examined 20 samples of melanoma, lung and colon cancer with previously identified BRAF, EGFR or KRASmutations. Formalin fixed paraffin embedded (FFPE) tissue was examined and cancer tissue identified by a pathologist. DNA extracted and the ion ampliseq™ cancer panel applied. Four samples were loaded on a 316 chip 100 Mb and run on Ion PGM.
Ninety five percent of the samples were amplified successfully. All 316 chips were run successfully. The average base call was 229 Mb per chip; on average each base was read 3503 times. All known mutations and deletions were detected providing support for the system's robustness. In addition, mutations in the APC, ATM, JAK3, KDR, KIT, MET, PTEN, STK11and TP53 genes were found.
As a clinical service we examined 5 samples of melanoma, gastric, appendix, parotid and sinonasal undifferentiated carcinoma. All of the samples were amplified successfully. All chips were run successfully; on average each base was read 1073 times. All known mutations were detected. In addition, mutations in ATM, GNAS, KRAS, ABL1 and ERBB2 genes were found. A multidisciplinary team including biologists, genetic counselors, oncologists, pathologists and surgeons discussed cases and provided instructions for further treatment.
We conclude that analysis of FFPE samples using the ampliseq™ cancer panel with the Ion PGM™ is doable and can identify known and new mutations in clinical samples. Routine use of this tool in treating oncological patients is feasible. Many methodological, ethical and medical issues need to be addressed in the wide spread clinical use of this test.
