The prostate cancer proteome

Prostate cancer is one of the most common malignancies in men. In 2015, the incidence of prostate cancer in Sweden was more than 10 000, and in the US 160 000 new cases are estimated in 2017. The vast majority of primary prostate cancer cases are adenocarcinomas, and the cancers often show characteristics such as multifocality and differentiation pattern heterogeneity. The rate of prostate tumor growth varies and is often low. Since the median age at diagnosis is high (65-70 years), most patients, especially those with localized tumors, often die of other complications without ever having suffered significant disability from the cancer. Approximately 15% of patients with prostate cancer are in high-risk of disease-related symptoms and death. However, certain patients may have prolonged survival even after the cancer has metastasized to distant sites. Prostate cancer is often stimulated by hormones (androgens) and anti-hormonal therapies can be effective to limit and slow down tumor growth, while radical prostatectomy is an option for treating localized prostate cancer.

Both normal prostatic glands and prostate cancer cells express the protein PSA (encoded by the gene KLK3) and the detection of elevated levels of PSA in the blood is today heavily used as a screening tool to detect the growth of prostate cancer. Prostate cancer can be diagnosed using rectal examination (palpation), and histological examinations of tissue biopsies using the Gleason Grade scoring system, where dominant histological characteristics of the cancer cells are used to determine the cancer stage. Immunostaining of relevant molecular markers is also important.

Here, we explore the prostate cancer proteome using TCGA transcriptomics data and antibody-based protein data. 159 genes are suggested as prognostic based on transcriptomics data from 494 patients; 134 genes are associated with unfavorable prognosis and 25 genes are associated with favorable prognosis.

TCGA data analysis

In this metadata study, we used data from TCGA where transcriptomics data was available from 494 male patients with prostate adenocarcinoma in total. Most of the patients (484 patients) were still alive at the time of data collection. Information on stage distribution was missing.

Unfavorable prognostic genes in prostate cancer

For unfavorable genes, higher relative expression levels at diagnosis give significantly lower overall survival for the patients. There are 134 genes associated with an unfavorable prognosis in prostate cancer. In Table 1, the top 20 most significant genes related to an unfavorable prognosis are listed.

ODF2 is a gene associated with an unfavorable prognosis in prostate cancer. The best separation is achieved by an expression cutoff at 10.4 fpkm which divides the patients into two groups with 99% 5-year survival for patients with high expression versus 93% for patients with low expression, p-value: 1.72e-4. The Outer dense fiber protein 2 is encoded by the ODF2 gene and is a major structural component of the sperm tail. Previous reports has shown that ODF2 may be a crucial component of the microtubule organizing center. Defects in this gene have been linked to infertility and abnormal sperm morphology. Immunohistochemical staining using an antibody targeting ODF2 (HPA001874) shows a differential expression pattern in prostate cancer samples.

p<0.001
ODF2 - survival analysis
ODF2 - high expression
ODF2 - low expression

Table 1. The 20 genes with highest significance associated with an unfavorable prognosis in prostate cancer.

Gene	Description	Predicted location	mRNA (cancer)	p-value
MBD1	methyl-CpG binding domain protein 1	Intracellular	7.6	1.70e-6
TEX30	testis expressed 30	Intracellular	1.9	2.10e-6
ZNF330	zinc finger protein 330	Intracellular	11.7	3.66e-6
USP53	ubiquitin specific peptidase 53	Intracellular	2.4	7.54e-6
ZNF654	zinc finger protein 654	Intracellular	3.7	9.97e-6
FBXO45	F-box protein 45	Intracellular	3.8	1.16e-5
ARRB2	arrestin beta 2	Intracellular	5.4	2.61e-5
RPAIN	RPA interacting protein	Intracellular	4.4	4.41e-5
DVL3	dishevelled segment polarity protein 3	Intracellular	16.9	4.77e-5
RNF8	ring finger protein 8	Intracellular	2.6	4.99e-5
ZNF875	zinc finger protein 875	Intracellular	9.8	5.18e-5
DNAJC9	DnaJ heat shock protein family (Hsp40) member C9	Intracellular	3.4	7.07e-5
HOOK2	hook microtubule tethering protein 2	Intracellular	9.8	7.59e-5
RAN	RAN, member RAS oncogene family	Intracellular	41.1	7.86e-5
LMNB2	lamin B2	Intracellular	9.3	9.53e-5
CEP70	centrosomal protein 70	Intracellular	6.7	9.60e-5
MTF2	metal response element binding transcription factor 2	Intracellular	2.6	1.05e-4
ZNF587B	zinc finger protein 587B	Intracellular	3.7	1.33e-4
RBM8A	RNA binding motif protein 8A	Intracellular	17.8	1.55e-4
COPS3	COP9 signalosome subunit 3	Intracellular	17.4	1.56e-4
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Favorable prognostic genes in prostate cancer

For favorable genes, higher relative expression levels at diagnosis give significantly higher overall survival for the patients. There are 25 genes associated with a favorable prognosis in prostate cancer. In Table 2, the top 20 most significant genes related to a favorable prognosis are listed.

ATP6V1E1 is a gene associated with a favorable prognosis in prostate cancer. The best separation is achieved by an expression cutoff at 33.1 fpkm which divides the patients into two groups with 100% 5- year survival for patients with high expression versus 95% for patients with low expression, p-value: 4.18e-4. The ATP6V1E1 gene encodes a multi-subunit enzyme important for the acidification of organelles in eukaryotic cells. This process is deemed necessary for intracellular mechanisms such as protein sorting, zymogen activation and receptor-mediated endocytosis. Immunohistochemical staining using an antibody targeting ATP6V1E1 (HPA029196) shows a differential expression pattern in prostate cancer samples.

p<0.001
ATP6V1E1 - survival analysis
ATP6V1E1 - high expression
ATP6V1E1 - low expression

Table 2. The 20 genes with highest significance associated with a favorable prognosis in prostate cancer.

Gene	Description	Predicted location	mRNA (cancer)	p-value
CRACR2A	calcium release activated channel regulator 2A	Intracellular	1.6	2.85e-6
TMEM158	transmembrane protein 158	Membrane	6.7	6.43e-5
EPHX1	epoxide hydrolase 1	Intracellular	73.3	1.32e-4
TSKU	tsukushi, small leucine rich proteoglycan	Secreted	16.9	1.66e-4
AOC1	amine oxidase copper containing 1	Secreted	5.7	1.71e-4
TXN2	thioredoxin 2	Intracellular	47.1	2.04e-4
SESN1	sestrin 1	Intracellular	11.9	2.07e-4
AK5	adenylate kinase 5	Intracellular	2.3	3.88e-4
GPAT4	glycerol-3-phosphate acyltransferase 4	Membrane	11.0	4.15e-4
RFPL2	ret finger protein like 2	Intracellular	5.6	4.16e-4
ATP6V1E1	ATPase H+ transporting V1 subunit E1	Intracellular	35.5	4.18e-4
RNF122	ring finger protein 122	Membrane	3.5	5.37e-4
COMT	catechol-O-methyltransferase	Intracellular	27.8	5.67e-4
UBE2D4	ubiquitin conjugating enzyme E2 D4 (putative)	Intracellular	5.7	5.77e-4
HBEGF	heparin binding EGF like growth factor	Membrane,Secreted	7.0	6.00e-4
NTNG2	netrin G2	Intracellular	1.3	6.06e-4
SLC35B2	solute carrier family 35 member B2	Membrane	37.9	6.07e-4
FOXQ1	forkhead box Q1	Intracellular	2.3	6.68e-4
ARL2	ADP ribosylation factor like GTPase 2	Intracellular	21.3	7.70e-4
FOS	Fos proto-oncogene, AP-1 transcription factor subunit	Intracellular	225.2	7.81e-4
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The prostate cancer transcriptome

The transcriptome analysis shows that 69% (n=13902) of all human genes (n=20090) are expressed in prostate cancer. All genes were classified according to the prostate cancer-specific expression into one of five different categories, based on the ratio between mRNA levels in prostate cancer compared to the mRNA levels in the other 16 analyzed cancer tissues.

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

332 genes show some level of elevated expression in prostate cancer compared to other cancers (Figure 1). The elevated category is further subdivided into three categories as shown in Table 3.

Table 3. The number of genes in the subdivided categories of elevated expression in prostate cancer.

		Distribution in the 17 cancers
		Detected in single	Detected in some	Detected in many	Detected in all	Total
Specificity	Cancer enriched	17	14	27	20	78
	Group enriched	0	46	41	13	100
	Cancer enhanced	14	29	65	46	154
	Total	31	89	133	79	332

Additional information

Prostate carcinomas are classified using the Gleason grade (Modified Gleason Grading System). The Gleason grade is a system of characterizing prostate cancer tissue based on the microscopical pattern of growth. The Gleason grade ranges from 1 to 5. Grade 1 is not used in clinical practice as it is impossible to distinguish grade 1 from adenosis and it is considered to lack malignant potential. Gleason grades corresponding to the two dominating growth patterns in prostate cancer are combined to provide the Gleason score and consequently, Gleason scores range from 4 (2+2) to 10 (5+5). The Gleason score is correlated to the risk of tumor spread and a low Gleason score reflects a more well differentiated tumor, while a high Gleason score corresponds to more poorly differentiated tumors. A tumor with Gleason score of 4 is least likely, and a tumor with a score of 10 is most likely to spread beyond the prostate.

Grade 1 tumors are small, uniform, well-differentiated, closely packed, and consist of glands in essentially circumscribed masses. In Grade 2 there is a moderate variation in size and shape of glands and more atypia in individual cells. A cribriform pattern may be present - still essentially circumscribed - but more loosely arranged. Grade 3 tumors show marked irregularity in size and shape of glands, with tiny glands/individual cells invading into the stroma. Solid cords and masses (papillary or cribriform), with easily distinguishable glandular differentiation, vary in size and may be quite large, but the essential feature is a smooth and usually rounded edge around all the circumscribed masses of the tumor. In Grade 4, tumor cells grow in a diffuse pattern and may show gland formation, however, glands are not single and separate, but coalesce and branch. Grade 5 tumors are poorly differentiated, usually presenting solid masses or diffuse growth with little or no differentiation into glands or with a few tiny glands or signet ring cells.

Immunohistochemistry using antibodies towards PSA can be used to discriminate prostate cancer from other forms of cancer. Immunohistochemistry is also important to delineate the two cell types in benign prostatic ducts and acini. The superficial layer is the secretory epithelium, beneath which there is a layer of continuous basal cells in normal glands. Sparse interspersed endocrine-paracrine cells are also present. The basal cells are useful for histological diagnosis as their presence confirms that the acinus is benign. The basal cells stain positive for p63 and high-molecular weight keratins. Malignant prostatic acini do not display basal cells and are identified using other immunohistochemistry markers such asAMACR/P504S.

Relevant links and publications

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

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

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

O'Hurley G et al., Analysis of the Human Prostate-Specific Proteome Defined by Transcriptomics and Antibody-Based Profiling Identifies TMEM79 and ACOXL as Two Putative, Diagnostic Markers in Prostate Cancer. PLoS One. (2015)
PubMed: 26237329 DOI: 10.1371/journal.pone.0133449

Histology dictionary - Prostate cancer