The bladder urothelial carcinoma proteome

Urothelial carcinoma, also termed transitional cell carcinoma or urinary bladder cancer, is a major cause of morbidity and mortality throughout the world. The highest frequency of cancer in the urinary bladder is found among urban Caucasians in Western Europe and in the United States of America. Urothelial cancer typically presents in patients over the age of 50 years and is approximately three times as common in males as in females. Smoking is considered an important risk factor.

Urothelial cancer can be divided into papillary and non-papillary tumors depending on the morphological appearance. Approximately 25% of all urothelial tumors are non-invasive papillary tumors. However, 10-15% of these patients will subsequently develop an invasive tumor.

Here, we explore the bladder urothelial carcinoma proteome using TCGA transcriptomics data and antibody-based protein data. 510 genes are suggested as prognostic based on transcriptomics data from 169 patients; 186 genes are associated with unfavorable prognosis and 324 genes are associated with favorable prognosis.

TCGA data analysis

In this metadata study, we used data from TCGA where transcriptomics data was available from 169 patients in total, where all patients were deceased at the time of sample collection. The dataset included 46 females and 123 males. The stage distribution was stage i) 0 patients, stage ii) 32 patients, stage iii) 55 patients, stage iv) 81 patients and 1 patient with missing stage information.

Unfavorable prognostic genes in bladder urothelial carcinoma

For unfavorable genes, higher relative expression levels at diagnosis give significantly lower overall survival for the patients. There are 186 genes associated with an unfavorable prognosis in bladder urothelial carcinoma. In Table 1, the top 20 most significant genes related to an unfavorable prognosis are listed.

PSMD2 is a gene associated with an unfavorable prognosis in bladder urothelial carcinoma. The best separation is achieved by an expression cutoff at 92 TPM which divides the patients into two groups with 1% 5-year survival for patients with high expression versus 8% for patients with low expression, p-value: 6.23e-6. Immunohistochemical staining using an antibody targeting PSMD2 (HPA045192) shows a differential expression pattern in bladder urothelial carcinoma samples.

p<0.001
PSMD2 - survival analysis
PSMD2 - high expression
PSMD2 - low expression

SERBP1 is a gene associated with an unfavorable prognosis in bladder urothelial carcinoma. The best separation is achieved by an expression cutoff at 72 TPM which divides the patients into two groups with 2% 5-year survival for patients with high expression versus 13% for patients with low expression, p-value: 5.78e-5. Immunohistochemical staining using an antibody targeting SERBP1 (CAB026297) shows a differential expression pattern in bladder urothelial carcinoma samples.

p=0.0011
SERBP1 - survival analysis
SERBP1 - high expression
SERBP1 - low expression

Table 1. The 20 genes with highest significance associated with an unfavorable prognosis in bladder urothelial carcinoma.

Gene	Description	Predicted location	mRNA (cancer)	p-value	Prognostic
GGT1	Gamma-glutamyltransferase 1	Membrane, Intracellular	6.1	6.50e-8	potential
CORO6	Coronin 6	Intracellular	1.8	3.17e-7	potential
PTGES2	Prostaglandin E synthase 2	Intracellular	51.6	5.27e-7	potential
H3C10	H3 clustered histone 10	Intracellular	8.7	6.77e-6	potential
GABRD	Gamma-aminobutyric acid type A receptor subunit delta	Membrane, Intracellular	1.7	7.19e-6	potential
CTSC	Cathepsin C	Membrane, Intracellular	230.9	1.04e-5	potential
RNF217	Ring finger protein 217	Membrane, Intracellular	3.0	1.14e-5	potential
HOXC9	Homeobox C9	Intracellular	2.7	1.57e-5	potential
PSMD2	Proteasome 26S subunit ubiquitin receptor, non-ATPase 2	Membrane, Intracellular	137.3	1.69e-5	potential
BRD9	Bromodomain containing 9	Intracellular	17.3	2.44e-5	potential
MAGEB2	MAGE family member B2	Intracellular	2.4	2.61e-5	potential
IGF2BP1	Insulin like growth factor 2 mRNA binding protein 1	Intracellular	2.7	2.81e-5	potential
SLIRP	SRA stem-loop interacting RNA binding protein	Intracellular	221.8	3.34e-5	potential
MKKS	MKKS centrosomal shuttling protein	Intracellular	10.5	4.85e-5	potential
PAX8	Paired box 8	Intracellular	3.3	5.27e-5	potential
PA2G4	Proliferation-associated 2G4	Intracellular	143.8	5.70e-5	potential
AGPAT4	1-acylglycerol-3-phosphate O-acyltransferase 4	Membrane, Intracellular	2.2	5.78e-5	potential
IL1RAP	Interleukin 1 receptor accessory protein	Secreted, Membrane, Intracellular	9.3	6.29e-5	potential
UCK2	Uridine-cytidine kinase 2	Intracellular	37.6	6.38e-5	potential
MRPL36	Mitochondrial ribosomal protein L36	Intracellular	84.6	6.54e-5	potential
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Favorable prognostic genes in bladder urothelial carcinoma

For favorable genes, higher relative expression levels at diagnosis give significantly higher overall survival for the patients. There are 324 genes associated with a favorable prognosis in bladder urothelial carcinoma. In Table 2, the top 20 most significant genes related to a favorable prognosis are listed.

FUCA1 is a gene associated with a favorable prognosis in bladder urothelial carcinoma. The best separation is achieved by an expression cutoff at 35 TPM which divides the patients into two groups with 7% 5-year survival for patients with high expression versus 0% for patients with low expression, p-value: 2.90e-5. Immunohistochemical staining using an antibody targeting FUCA1 (HPA046542) shows a differential expression pattern in bladder urothelial carcinoma samples.

p<0.001
FUCA1 - survival analysis
FUCA1 - high expression
FUCA1 - low expression

MYO6 is a gene associated with a favorable prognosis in bladder urothelial carcinoma. The best separation is achieved by an expression cutoff at 6.6 TPM which divides the patients into two groups with 5% 5-year survival for patients with high expression versus 0% for patients with low expression, p-value: 2.15e-4. Immunohistochemical staining using an antibody targeting MYO6 (CAB010762) shows a differential expression pattern in bladder urothelial carcinoma samples.

p=0.0275
MYO6 - survival analysis
MYO6 - high expression
MYO6 - low expression

Table 2. The 20 genes with highest significance associated with a favorable prognosis in bladder urothelial carcinoma.

Gene	Description	Predicted location	mRNA (cancer)	p-value	Prognostic
MFSD6	Major facilitator superfamily domain containing 6	Membrane, Intracellular	22.2	6.55e-8	potential
ACOXL	Acyl-CoA oxidase like	Intracellular	6.2	5.97e-7	potential
ZNF681	Zinc finger protein 681	Intracellular	2.6	6.01e-7	potential
CEP57	Centrosomal protein 57	Intracellular	17.4	8.14e-7	potential
APOL3	Apolipoprotein L3	Intracellular	17.4	1.38e-6	potential
ZNF816	Zinc finger protein 816	Membrane, Intracellular	9.8	1.59e-6	potential
TCEAL3	Transcription elongation factor A like 3	Intracellular	40.4	1.62e-6	potential
ASAH1	N-acylsphingosine amidohydrolase 1	Membrane, Intracellular	123.3	1.66e-6	potential
ZSCAN9	Zinc finger and SCAN domain containing 9	Intracellular	12.9	1.70e-6	potential
SEPTIN2	Septin 2	Intracellular	158.2	2.96e-6	potential
BTG2	BTG anti-proliferation factor 2	Intracellular	153.2	5.99e-6	potential
LYPD5	LY6/PLAUR domain containing 5	Membrane, Intracellular	6.9	7.20e-6	potential
MYH14	Myosin heavy chain 14	Intracellular	106.8	8.74e-6	potential
RBM47	RNA binding motif protein 47	Intracellular	25.2	9.21e-6	potential
CD2AP	CD2 associated protein	Intracellular	31.4	9.26e-6	potential
MAL2	Mal, T cell differentiation protein 2	Membrane	139.5	1.00e-5	potential
SCP2	Sterol carrier protein 2	Intracellular	148.4	1.14e-5	potential
EPM2AIP1	EPM2A interacting protein 1	Intracellular	7.3	1.24e-5	potential
FOXA1	Forkhead box A1	Intracellular	30.8	1.41e-5	potential
ZNF83	Zinc finger protein 83	Intracellular	27.3	1.48e-5	potential
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The bladder urothelial carcinoma transcriptome

The transcriptome analysis shows that 73% (n=14785) of all human genes (n=20162) are expressed in bladder urothelial carcinoma. All genes were classified according to the bladder urothelial carcinoma-specific expression into one of five different categories, based on the ratio between mRNA levels in bladder urothelial carcinoma 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 bladder urothelial carcinoma as well as in all other cancer tissues.

186 genes show some level of elevated expression in bladder urothelial carcinoma 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 bladder urothelial carcinoma.

		Distribution in the 31 cancers
		Detected in single	Detected in some	Detected in many	Detected in all	Total
Specificity	Cancer enriched	4	6	10	0	20
	Group enriched	0	27	42	5	74
	Cancer enhanced	7	29	45	11	92
	Total	11	62	97	16	186

Additional information

The majority of patients with urothelial cancer present symptoms of hematuria (blood in the urine) and/or dysuria (painful urination). Urothelial cancer most commonly arises in the urinary bladder, but can develop in the renal pelvis, ureters or urethra.

Papillary tumors appear in cystoscopy with variable size, ranging from minute excrescences to large, cauliflower like tumors protruding into the urinary bladder lumen. Histologically papillary tumors are characterized by tall and branched papillae that are usually covered by several layers of urothelial tumor cells.

Sarcomatoid (sarcoma and carcinoma features) variants of urothelial carcinoma exist and squamous cell carcinomas and adenocarcinomas can develop in the urinary bladder. Squamous cell carcinoma in the urinary bladder accounts for approximately 5% of all cancers in the urinary bladder. However, in areas where schistosomiasis is endemic, squamous cell carcinomas account for approximately 75% of all bladder carcinomas.

Urothelial cancer is classified with respect to surgical stage at time for diagnosis and tumor grade based on histology. Staging of urothelial cancer defines a tumor as cancer in situ, non-invasive, invasive in the lamina propria or invasive into muscular bladder wall. The TNM-based classification provides important prognostic information and guides further treatment. In addition, important prognostic information can also be obtained through the grading of the tumor based on morphological features. Tumor cells vary in atypical appearance and are graded according to the degree of nuclear atypia into four different grades, including urothelial neoplasm of low malignant potential. The most malignant tumors are of grade 3, and exhibit at least focal areas of high-grade nuclear atypia and common mitotic figures. Urothelial carcinoma grade 3 represents a tumor with more aggressive behavior and increased risk for recurrence and metastatic spread.

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

Habuka M et al., The Urinary Bladder Transcriptome and Proteome Defined by Transcriptomics and Antibody-Based Profiling. PLoS One. (2015)
PubMed: 26694548 DOI: 10.1371/journal.pone.0145301

Histology dictionary - Urothelial cancer