DNA microarrays have been widely applied to cancer transcriptome analysis; however, the majority of such data are not easily accessible or comparable. Furthermore, several important analytic approaches have been applied to microarray analysis; however, their application is often limited. To overcome these limitations, we have developed Oncomine, a bioinformatics initiative aimed at collecting, standardizing, analyzing, and delivering cancer transcriptome data to the biomedical research community. Our analysis has identified the genes, pathways, and networks deregulated across 18,000 cancer gene expression microarrays, spanning the majority of cancer types and subtypes. Here, we provide an update on the initiative, describe the database and analysis modules, and highlight several notable observations. Results from this comprehensive analysis are available at http://www.oncomine.org.

Multiple, complex molecular events characterize cancer development and progression1,2. Deciphering the molecular networks that distinguish organ-confined disease from metastatic disease may lead to the identification of critical biomarkers for cancer invasion and disease aggressiveness. Although gene and protein expression have been extensively profiled in human tumors, little is known about the global metabolomic alterations that characterize neoplastic progression. Using a combination of high throughput liquid and gas chromatography-based mass spectrometry, we profiled more than 1126 metabolites across 262 clinical samples related to prostate cancer (42 tissues and 110 each of urine and plasma). These unbiased metabolomic profiles were able to distinguish benign prostate, clinically localized prostate cancer, and metastatic disease. Sarcosine, an N-methyl derivative of the amino acid glycine, was identified as a differential metabolite that was highly elevated during prostate cancer progression to metastasis and can be detected non-invasively in urine. Sarcosine levels were also elevated in invasive prostate cancer cell lines relative to benign prostate epithelial cells. Knockdown of glycine-N-methyl transferase (GNMT), the enzyme that generates sarcosine from glycine, attenuated prostate cancer invasion. Addition of exogenous sarcosine or knockdown of the enzyme that leads to sarcosine degradation, sarcosine dehydrogenase (SARDH), induced an invasive phenotype in benign prostate epithelial cells. Androgen receptor and the ERG gene fusion product coordinately regulate components of the sarcosine pathway. Taken together, we profiled the metabolomic alterations of prostate cancer progression revealing sarcosine as a potentially important metabolic intermediary of cancer cell invasion and aggressivity.

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Despite efforts to profile prostate cancer, the genetic alterations and biological processes that correlate with the observed histological progression are unclear. Using laser-capture microdissection to isolate 101 cell populations, we have profiled prostate cancer progression from benign epithelium to metastatic disease. By analyzing expression signatures in the context of over 14,000 'molecular concepts', or sets of biologically connected genes, we generated an integrative model of progression. Molecular concepts that demarcate critical transitions in progression include protein biosynthesis, E26 transformation-specific (ETS) family transcriptional targets, androgen signaling and cell proliferation. Of note, relative to low-grade prostate cancer (Gleason pattern 3), high-grade cancer (Gleason pattern 4) shows an attenuated androgen signaling signature, similar to metastatic prostate cancer, which may reflect dedifferentiation and explain the clinical association of grade with prognosis. Taken together, these data show that analyzing gene expression signatures in the context of a compendium of molecular concepts is useful in understanding cancer biology.

Through a prospective clinical sequencing program for advanced cancers, four index cases were identified which harbor gene rearrangements of FGFR2 including patients with cholangiocarcinoma, breast cancer, and prostate cancer. After extending our assessment of FGFR rearrangements across multiple tumor cohorts, we identified additional FGFR gene fusions with intact kinase domains in lung squamous cell cancer, bladder cancer, thyroid cancer, oral cancer, glioblastoma, and head and neck squamous cell cancer. All FGFR fusion partners tested exhibit oligomerization capability, suggesting a shared mode of kinase activation. Overexpression of FGFR fusion proteins induced cell proliferation. Two bladder cancer cell lines that harbor FGFR3 fusion proteins exhibited enhanced susceptibility to pharmacologic inhibition in vitro and in vivo. Due to the combinatorial possibilities of FGFR family fusion to a variety of oligomerization partners, clinical sequencing efforts which incorporate transcriptome analysis for gene fusions are poised to identify rare, targetable FGFR fusions across diverse cancer types.

Recurrent gene fusions, typically associated with hematological malignancies and rare bone and soft tissue tumors1, have been recently described in common solid tumors2–9. Here we employ an integrative analysis of high-throughput long and short read transcriptome sequencing of cancer cells to discover novel gene fusions. As a proof of concept we successfully utilized integrative transcriptome sequencing to “re-discover” the BCR-ABL1 10 gene fusion in a chronic myelogenous leukemia cell line and the TMPRSS2-ERG 2,3 gene fusion in a prostate cancer cell line and tissues. Additionally, we nominated, and experimentally validated, novel gene fusions resulting in chimeric transcripts in cancer cell lines and tumors. Taken together, this study establishes a robust pipeline for the discovery of novel gene chimeras using high throughput sequencing, opening up an important class of cancer-related mutations for comprehensive characterization.

A 44-year old woman with recurrent solitary fibrous tumor (SFT)/hemangiopericytoma was enrolled in a clinical sequencing program including whole exome and transcriptome sequencing. A gene fusion of the transcriptional repressor NAB2 with the transcriptional activator STAT6 was detected. Transcriptome sequencing of 27 additional SFTs all revealed the presence of a NAB2-STAT6 gene fusion. Using RT-PCR and sequencing, we detected this fusion in 51 of 51 SFTs, indicating high levels of recurrence. Expression of NAB2-STAT6 fusion proteins was confirmed in SFT, and the predicted fusion products harbor the early growth response (EGR)-binding domain of NAB2 fused to the activation domain of STAT6. Overexpression of the NAB2-STAT6 gene fusion induced proliferation in cultured cells and activated EGR-responsive genes. These studies establish NAB2-STAT6 as the defining driver mutation of SFT and provide an example of how neoplasia can be initiated by converting a transcriptional repressor of mitogenic pathways into a transcriptional activator.

Individual cancers harbor a set of genetic aberrations that can be informative for identifying rational therapies currently available or in clinical trials. We implemented a pilot study to explore the practical challenges of applying high-throughput sequencing in clinical oncology. We enrolled patients with advanced or refractory cancer who were eligible for clinical trials. For each patient, we performed whole-genome sequencing of the tumor, targeted whole-exome sequencing of tumor and normal DNA, and transcriptome sequencing (RNA-Seq) of the tumor to identify potentially informative mutations in a clinically relevant time frame of 3 to 4 weeks. With this approach, we detected several classes of cancer mutations including structural rearrangements, copy number alterations, point mutations, and gene expression alterations. A multidisciplinary Sequencing Tumor Board (STB) deliberated on the clinical interpretation of the sequencing results obtained. We tested our sequencing strategy on human prostate cancer xenografts. Next, we enrolled two patients into the clinical protocol and were able to review the results at our STB within 24 days of biopsy. The first patient had metastatic colorectal cancer in which we identified somatic point mutations in NRAS, TP53, AURKA, FAS, and MYH11, plus amplification and overexpression of cyclin-dependent kinase 8 (CDK8). The second patient had malignant melanoma, in which we identified a somatic point mutation in HRAS and a structural rearrangement affecting CDKN2C. The STB identified the CDK8 amplification and Ras mutation as providing a rationale for clinical trials with CDK inhibitors or MEK (mitogenactivated or extracellular signal–regulated protein kinase kinase) and PI3K (phosphatidylinositol 3-kinase) inhibitors, respectively. Integrative high-throughput sequencing of patients with advanced cancer generates a comprehensive, individual mutational landscape to facilitate biomarker-driven clinical trials in oncology.