Integrated genome and transcriptome sequencing identifies a novel form of hybrid and aggressive prostate cancer

Abstract

Next-generation sequencing is making sequence-based molecular pathology and personalized oncology viable. We selected an individual initially diagnosed with conventional but aggressive prostate adenocarcinoma and sequenced the genome and transcriptome from primary and metastatic tissues collected prior to hormone therapy. The histology-pathology and copy number profiles were remarkably homogeneous, yet it was possible to propose the quadrant of the prostate tumour that likely seeded the metastatic diaspora. Despite a homogeneous cell type, our transcriptome analysis revealed signatures of both luminal and neuroendocrine cell types. Remarkably, the repertoire of expressed but apparently private gene fusions, including C15orf21:MYC, recapitulated this biology. We hypothesize that the amplification and over-expression of the stem cell gene MSI2 may have contributed to the stable hybrid cellular identity. This hybrid luminal-neuroendocrine tumour appears to represent a novel and highly aggressive case of prostate cancer with unique biological features and, conceivably, a propensity for rapid progression to castrate-resistance. Overall, this work highlights the importance of integrated analyses of genome, exome and transcriptome sequences for basic tumour biology, sequence-based molecular pathology and personalized oncology.

Figure 1

Histopathology and copy number analysis. Haematoxylin and eosin stains showing uniform adenocarcinoma with a Gleason score of 10 in patient 963's radical prostatectomy specimen (A, ×0.5; B, ×40) and a lymph node metastasis (C, ×20). No normal prostate structure can be discerned in (A, B). Lymphocytes can be seen in (C). Scale bars = 1 mm (A); 20 μm (B); 100 μm (C). (D) Differences between the samples, i.e. RP differs in signal intensity from LNmet at 2.9% of aCGH probe loci. The most and least similar pairs are emphasized. (E) Frequency plot showing the copy number (CN) aberrations in the primary tumour quadrants and the LNmet (green, gain; red, loss). A frequency of 100% indicates a CN aberration detected in all five samples; the majority of CN aberrations are detected in all samples. CN aberrant genes previously associated with PCa are annotated. Blue, fusion genes (the genomic breakpoints of 12 genes involved in fusion events coincided with aCGH segment breaks); pink, readthrough events. Note: Chromosomes X and Y demonstrated no aberrations and are not shown. (F, G) Chr11 CN profile in LT (F) and RP (G), indicating the marginal differences. (H, I) Chr17 CN profile in LP (H) and LNmet (I).

Figure 2

Fusion genes validated in the primary tumour and lymph node metastasis. The top three fusions involve either an androgen-related gene, a tumour suppressor or an oncogene. The bottom five fusions involve genes with a neuroendocrine function. Note that despite absence of RNA-Seq reads (ie expression) for some fusion genes, the underlying genomic breakpoints for each fusion were detected in both primary and metastatic tumours (see Supporting information, Figure S4).

Figure 3

Analysis of gene expression levels. (A) Heat map demonstrating expression of genes with a neural/endocrine function in the primary tumour (LP) and the lymph node metastatic tumour (LNmet) of patient 963 compared to adenocarcinoma cell lines and benign prostate. FP, filament protein; TF, transcription factor. (B–E) Antibody stains showing strong expression of AR (B, ×2; C, ×40) and CHGA (D, ×2; E, ×40) in the primary tumour. Bands of stromal cells are unstained. (F, G, I, J) Dual antibody stains of AR and CHGA (F, ×8; G, ×40; I, ×10; J, ×40) confirming co-expression in 100% of tumour cells. Note that in (I, J) benign prostate glands are visible, demonstrating that normal luminal cells are AR-positive only, while normal NE cells are CHGA-positive only. Scale bars = 1 mm (B, D); 200 μm (I); 100 μm (C, E, F); 20 μm (G, J). (H) Hexbin plot illustrating correlation of gene expression levels between LP and LNmet (log₂ scale). Red and green lines indicate a four- and 16-fold expression difference, respectively, between LP and LNmet.