Whole-Exome Sequencing of Muscle-Invasive Bladder Cancer Identifies Recurrent Mutations of UNC5C and Prognostic Importance of DNA Repair Gene Mutations on Survival

Abstract

Purpose Due to suboptimal outcomes in muscle-invasive bladder cancer even with multimodality therapy, determination of potential genetic drivers offers the possibility of improving therapeutic approaches and discovering novel prognostic indicators. Experimental Design Using pTN staging, we case-matched 81 patients with resected ≥pT2 bladder cancers for whom perioperative chemotherapy use and disease recurrence status were known. Whole exome sequencing was conducted in 43 cases to identify recurrent somatic mutations and targeted sequencing of 10 genes selected from the initial screening in an additional 38 cases was completed. Mutational profiles along with clinicopathologic information were correlated with recurrence-free survival (RFS) in the patients. Results We identified recurrent novel somatic mutations in the gene UNC5C (9.9%), in addition to TP53 (40.7%), KDM6A (21.0%), and TSC1 (12.3%). Patients who were carriers of somatic mutations in DNA repair genes (one or more of ATM, ERCC2, FANCD2, PALB2, BRCA1 or BRCA2) had a higher overall number of somatic mutations (p=0.011). Importantly, after a median follow-up of 40.4 months, carriers of somatic mutations (n=25) in any of these six DNA repair genes had significantly enhanced RFS compared to non-carriers (median 32.4 vs. 14.8 months; hazard ratio of 0.46, 95% CI 0.22 to 0.98; p=0.0435), after adjustment for pathologic pTN staging and independent of adjuvant chemotherapy usage. Conclusion Better prognostic outcomes of individuals carrying somatic mutations in DNA repair genes suggest these mutations as favorable prognostic events in muscle-invasive bladder cancer. Additional mechanistic investigation into the previously undiscovered role of UNC5C in bladder cancer is warranted.

Molecular analysis of urothelial cancer cell lines for modeling tumor biology and drug response

“…Eleven genes encoding proteins involved in phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)-mammalian target of rapamycin (mTOR) signaling were altered in 60% of the lines, most frequently by altered PI3K regulatory subunit (PIK3CA) and the tyrosine kinase gene, ERBB2 . In agreement with previous observations (2,28), the BRCA DNA repair pathway, including BRCA1 and ATM , were altered in 44% of lines in a predominantly mutually exclusive manner. This is similar to 40/127 (32%) of tumors from TCGA patients (4) which also exhibited BRCA pathway gene alterations that were predominantly mutually exclusive.…”

DNA Damage Response and Repair Gene Alterations Are Associated with Improved Survival in Patients with Platinum-Treated Advanced Urothelial Carcinoma

“…Tumors with mutation in at least one of these genes had significantly higher overall numbers of somatic mutations 25 and longer recurrence-free survival. Similarly, we observed that number of alterations in DDR genes was positively correlated with total number of mutations and copy number alterations.…”

Genomic DNA extraction methods using formalin-fixed paraffin-embedded tissue

“…Other methods less frequently usednot measured in our study include the Maxwell 16 method by Promega (Mannheim, Germany), which was shown to perform well against other automated Qiagen kits [9], and the Chelex bead method (Bio-Rad, Berkeley, California) which has been used with some success in manual or automated extractions from FFPE tissues [10]. A small number of studies have shown that the DNA obtained by a recent sonication-based method called adaptive focused acoustics (AFA) is more amenable to next generation sequencing than DNA from other methods, resulting in greater coverage of the genome and facilitating easier assembly and alignment [11][12][13]. To our knowledge, ours is the first study to compare AFA for use in aCGH.…”