Integrative molecular profiling of routine clinical prostate cancer specimens.
BACKGROUND
Comprehensive molecular profiling led to the recognition of multiple prostate cancer (PCa) molecular subtypes and driving alterations, but translating these findings to clinical practice is challenging.


METHODS
We developed a formalin-fixed paraffin-embedded (FFPE) tissue compatible integrative assay for PCa molecular subtyping and interrogation of relevant genetic/transcriptomic alterations (MiPC). We applied MiPC, which combines capture-based next generation sequencing and quantitative reverse transcription PCR (qRT-PCR), to 53 FFPE PCa specimens representing cases not well represented in frozen tissue cohorts, including 8 paired primary tumor and lymph node metastases. Results were validated using multiplexed PCR based NGS and Sanger sequencing.


RESULTS
We identified known and novel potential driving, somatic mutations and copy number alterations, including a novel BRAF T599_V600insHT mutation and CYP11B2 amplification in a patient treated with ketoconazole (a potent CYP11B2 inhibitor). qRT-PCR integration enabled comprehensive molecular subtyping and provided complementary information, such as androgen receptor (AR) target gene module assessment in advanced cases and SPINK1 over-expression. MiPC identified highly concordant profiles for all 8 tumor/lymph node metastasis pairs, consistent with limited heterogeneity amongst driving events. MiPC and exome sequencing were performed on separately isolated conventional acinar PCa and prostatic small cell carcinoma (SCC) components from the same FFPE resection specimen to enable direct comparison of histologically distinct components. While both components showed TMPRSS2:ERG fusions, the SCC component exclusively harbored complete TP53 inactivation (frameshift variant and copy loss) and two CREBBP mutations.


CONCLUSIONS
Our results demonstrate the feasibility of integrative profiling of routine PCa specimens, which may have utility for understanding disease biology and enabling personalized medicine applications.
Michigan Center for Translational Pathology, Department of Pathology; Department of Pathology, Oregon Health & Sciences University, Portland.
C S
CS
Grasso
Michigan Center for Translational Pathology, Department of Pathology.
A K
AK
Cani
Departments of Computational Medicine & Bioinformatics.
D H
DH
Hovelson
Michigan Center for Translational Pathology, Department of Pathology; Department of Pathology, Oregon Health & Sciences University, Portland.
M J
MJ
Quist
Anesthesiology.
N J
NJ
Douville
Michigan Center for Translational Pathology, Department of Pathology.
V
V
Yadati
Michigan Center for Translational Pathology, Department of Pathology.
A M
AM
Amin
Division of Human Biology; Division of Clinical Research, Fred Hutchinson Cancer Research Center, Seattle.
P S
PS
Nelson
Michigan Center for Translational Pathology, Department of Pathology.
B L
BL
Betz
Michigan Center for Translational Pathology, Department of Pathology.
C-J
CJ
Liu
Department of Cancer Biology; Departments of Urology; Radiation Oncology, Thomas Jefferson University, Philadelphia, USA.
K E
KE
Knudsen
Internal Medicine; Comprehensive Cancer Center, University of Michigan Medical School, Ann Arbor.
K A
KA
Cooney
Radiation Oncology; Comprehensive Cancer Center, University of Michigan Medical School, Ann Arbor.
F Y
FY
Feng
Michigan Center for Translational Pathology, Department of Pathology.
A S
AS
McDaniel
Michigan Center for Translational Pathology, Department of Pathology; Urology; Comprehensive Cancer Center, University of Michigan Medical School, Ann Arbor. Electronic address: tomlinss@umich.edu.
S A
SA
Tomlins
D014408:Biomarkers, Tumor / Q000235:genetics*; D001706:Biopsy; D056915:DNA Copy Number Variations; D004252:DNA Mutational Analysis; D005240:Feasibility Studies; D005404:Fixatives; D005557:Formaldehyde; D020869:Gene Expression Profiling / Q000379:methods*; D015972:Gene Expression Regulation, Neoplastic; D020022:Genetic Predisposition to Disease; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D008207:Lymphatic Metastasis; D008297:Male; D009154:Mutation; D016612:Paraffin Embedding; D010641:Phenotype; D020641:Polymorphism, Single Nucleotide; D057285:Precision Medicine; D011237:Predictive Value of Tests; D011379:Prognosis; D011471:Prostatic Neoplasms / Q000145:classification / Q000235:genetics* / Q000473:pathology / Q000628:therapy; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016707:Tissue Fixation
D014408:Biomarkers, Tumor; D005404:Fixatives; D005557:Formaldehyde
formalin-fixed paraffin-embedded tissue; molecular subtyping; next generation sequencing; qPCR; small-cell carcinoma