The testis cancer proteome


Testis cancer constitutes approximately 1% of cancer in males. Tumors of germ cell origin account for approximately 95% of all testis cancers. Pathology plays a key role in management of patients with testicular tumors by allowing for accurate classification of tumors to provide the prognostic parameters needed for optimizing decisions regarding treatment and follow-up.

Testis cancer is divided into two major categories: seminoma and non-seminomatous germ cell tumors. Seminomas account for approximately 45% of all germ cell tumors and are characterized histologically by evenly spaced and relatively large uniform tumor cells with distinct cell borders. Tumor cell nuclei are often centrally localized and show distinct nuclear membranes and one or two distinct nucleoli. The characteristic tumor stroma in seminoma is built up by a delicate fibrovascular network with thin collagenous septa containing variable amounts of small lymphocytes. Of the non-seminomatous tumors, embryonal carcinoma accounts for 15-30% and represents the second most frequent pure type of testicular cancer. Embryonal cancer exhibits an acinar, tubular, papillary or solid growth pattern with areas of necrosis, hemorrhage and fibrosis. Tumor cells are highly pleomorphic with large, irregular nuclei and indistinct cell borders.

Here, we explore the testis cancer proteome using TCGA transcriptomics data and antibody based protein data. 57 genes are suggested as prognostic based on transcriptomics data from 134 patients; 42 genes associated with unfavourable prognosis and 15 genes associated with favourable prognosis.

TCGA data analysis


In this metadata study we used data from TCGA where transcriptomics data was available from 134 males with seminoma or non-seminomatous germ cell tumors. Most of the patients (130 patients) were still alive at the time of data collection. The stage distribution was i) 55 patients ii) 12 patients iii) 14 patients is) 46 patients and 7 patients with missing stage information.

Unfavourable prognostic genes in testis cancer


For unfavourable genes, higher relative expression levels at diagnosis gives significantly lower overall survival for the patients. There are 42 genes associated with unfavourable prognosis in testis cancer. In Table 1, the top 20 most significant genes related to unfavourable prognosis are listed.

CLCN7 is a gene associated with unfavourable prognosis in testis cancer. The best separation is achieved by an expression cutoff at 8.6 fpkm which divides the patients into two groups with 87 % 5-year survival for patients with high expression versus 100 % for patients with low expression (p-value: 3.21e-4 ). Immunohistochemical using an antibody targeting CLCN7 (HPA043586) shows differential expression pattern in testis cancer samples.

CLCN7 - survival analysis p<0.001
CLCN7 - high expression
CLCN7 - low expression

GIMAP8 is another gene associated with unfavourable prognosis in testis cancer. The best separation is achieved by an expression cutoff at 6.2 fpkm which divides the patients into two groups with 88 % 5-year survival for patients with high expression versus 100 % for patients with low expression (p-value: 8.47e-4 ). Immunohistochemical staining using an antibody targeting GIMAP8 (HPA014474) shows differential expression pattern in testis cancer samples.

GIMAP8 - survival analysis p<0.001
GIMAP8 - high expression
GIMAP8 - low expression

Table 1. The 20 genes with highest significance associated with unfavourable prognosis in testis cancer.

Gene

Description

Predicted localization

mRNA (cancer)

p-value

CD300C CD300c molecule Membrane 1.6 8.53e-5
PTGIR prostaglandin I2 (prostacyclin) receptor (IP) Intracellular,Membrane,Secreted 1.1 1.74e-4
CLCN7 chloride channel, voltage-sensitive 7 Membrane 6.9 3.21e-4
GUCY1B3 guanylate cyclase 1, soluble, beta 3 Intracellular 2.8 3.23e-4
GUCY1A3 guanylate cyclase 1, soluble, alpha 3 Intracellular 1.8 3.59e-4
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Favourable prognostic genes in testis cancer


For favourable genes, higher relative expression levels at diagnosis gives significantly higher overall survival for the patients. There are 15 genes associated with favourable prognosis in testis cancer. In Table 2, the top 20 most significant genes related to favourable prognosis are listed.

IFT52 is a gene associated with favourable prognosis in testis cancer. The best separation is achieved by an expression cutoff at 13.2 fpkm which divides the patients into two groups with 100 % 5-year survival for patients with high expression versus 87 % for patients with low expression (p-value: 3.33e-4). Immunohistochemical staining using an antibody targeting IFT52 (HPA067423) shows differential expression in testis cancer samples.

IFT52 - survival analysis p<0.001
IFT52 - high expression
IFT52 - low expression

Table 2. The 15 genes with highest significance associated with favourable prognosis in testis cancer.

Gene

Description

Predicted localization

mRNA (cancer)

p-value

SNX31 sorting nexin 31 Intracellular 1.1 1.47e-4
PIN4 peptidylprolyl cis/trans isomerase, NIMA-interacting 4 Intracellular 9.3 2.58e-4
IFT52 intraflagellar transport 52 Intracellular 16.7 3.33e-4
DVL1 dishevelled segment polarity protein 1 Intracellular 21.3 3.33e-4
IQCC IQ motif containing C Intracellular 3.3 3.79e-4
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The testis cancer transcriptome


The transcriptome analysis shows that 74% (n=14540) of all human genes (n=19571) are expressed in testis cancer. All genes were classified according to the testis cancer-specific expression into one of five different categories, based on the ratio between mRNA levels in testis cancer compared to the mRNA levels in the other 16 analyzed cancer tissues. 467 genes show some level of elevated expression in testis cancer compared to other cancers (Figure 1). The elevated category is further subdivided into three categories as shown in Table 3.

Figure 1. The distribution of all genes across the five categories based on transcript abundance in testis cancer as well as in all other cancer tissues.

Table 3. Number of genes in the subdivided categories of elevated expression in testis cancer

Category

Number of genes

Description

Tissue enriched 193 At least five-fold higher mRNA levels in a particular cancer as compared to all other cancers
Group enriched 94 At least five-fold higher mRNA levels in a group of 2-7 cancers
Tissue enhanced 180 At least five-fold higher mRNA levels in a particular cancer as compared to average levels in all cancers
Total 467 Total number of elevated genes in testis cancer

Additional information


Non-seminomatous tumors are further classified as pure or mixed tumors. For mixed tumors, all included components must be defined and reported along with an estimation of the extent of each component. Tumors containing both seminomatous and non-seminomatous components are regarded as non-seminomatous germ cell tumors for treatment purposes.

Embryonal carcinoma is a relatively undifferentiated germ cell tumor from which the other more differentiated components are derived. These non-seminomatous components include i) endodermal sinus tumor (yolk sac tumor), typically characterized by the presence of Schiller-Duval bodies (resembling a central capillary lined by flattened layers of tumor cells), and ii) choriocarcinoma, representing extra-embryonic development, and iii) teratoma, representing embryonic development. Teratomas can in turn be composed of both immature and mature components representing cell types and structures from all embryonic germ layers. Schiller-Duval bodies are said to resemble a glomerulus. They have a mesodermal core with a central capillary, all lined by flattened layers of both visceral and parietal cells.

The distinction between different tumor types and components within a testicular tumor is based on microscopical examination and in addition to morphology, immunohistochemistry provides important information. Commonly used antibodies for the differential diagnostics of these tumors include D2-40, OCT 3/4, hCG and AFP in addition to CD30 and markers of intermediate filaments, e.g. cytokeratin and vimentin.

Relevant links and publications


Uhlen M et al, 2017. A pathology atlas of the human cancer transcriptome. Science.
PubMed: 28818916 DOI: 10.1126/science.aan2507

Cancer Genome Atlas Research Network et al, 2013. The Cancer Genome Atlas Pan-Cancer analysis project. Nat Genet.
PubMed: 24071849 DOI: 10.1038/ng.2764

UhlÚn M et al, 2015. Tissue-based map of the human proteome. Science
PubMed: 25613900 DOI: 10.1126/science.1260419

Djureinovic D et al, 2014. The human testis-specific proteome defined by transcriptomics and antibody-based profiling. Mol Hum Reprod.
PubMed: 24598113 DOI: 10.1093/molehr/gau018

Histology dictionary - Testis cancer