Targeted next-generation sequencing (NGS) of advanced prostate cancer (PCA) using formalin-fixed tissue.

Abstract

4649 Background: The genomic landscape of advanced PCA is not well characterized, partly due to limited availability of frozen metastatic tissue. This has created a gap in our knowledge of treatment related and potentially targetable genomic alterations. The purpose of this study was to demonstrate feasibility of performing NGS to molecularly characterize advanced PCA using formalin-fixed paraffin embedded (FFPE) tissue. Methods: 25 metastatic CRPC, 4 metastatic hormone naïve PCA, 3 primary localized PCA, and 18 benign matched prostates were evaluated (40mm FFPE tissue per case). High-density foci were captured and sequenced for 3230 exons of 182 cancer-related genes and 37 introns of 14 genes often rearranged in cancer to an average depth of >800x in a CLIA lab (Foundation Medicine). Recurrent mutations, copy number alterations, and fusions were validated using PCR and FISH. Results: In >90% of samples, there was sufficient DNA (≥50 ng) for analysis. Recurrent high confidence cancer alterations in CRPC included: TMPRSS2-ERG fusion (44%), PTEN loss (44%), TP53 mutation (40%), AR mutation (24%), AR gain (24%), RB mutation (28%), MYC gain (12%), and BRCA2 loss (12%). Overall 48% of CRPC harbored AR gene alterations. Additionally, there were mutations in CTNNB1 (12%), ATM (8%) and PIK3CA (4%). Copy number alterations not previously described in PCA included CCND1, CDK4/6 gains and CDKN2A/CDKN2B deletions. Hormone naïve metastatic and high risk localized PCA demonstrated similar frequency of TMPRSS2-ERG gene fusion, BRCA2 deletion, and TP53 mutations as CRPC, but AR alterations and MYC gain were not seen. Conclusions: This study demonstrates feasibility of in-depth, NGS based DNA analysis using FFPE tissue, even biopsy material. Frequent AR alterations in CRPC, mutations associated with disease progression, and potential drug targets were identified. Focused NGS has clinical potential to identify actionable genomic alterations in advanced PCA that can impact patient participation in trials as well as treatment and outcome. Treatment options include PARP inhibitors for patients with BRCA2 and ATM alterations (20% of cases) and PI3K/AKT inhibitors for PIK3CA mutations.