Introduction
Review
Pipeline for biomarker development
Clinical conceptions on the biomarkers study design
Sample biobanking
Sample collection and treatment
Study design and evaluation of the analytical performance
Technical considerations regarding the analytical set up for biomarker development
A) Discovery of biomarkers
Gel-based proteomics platforms for biomarker discovery
Gel-free proteomics
Shotgun proteomics
CE-MS
Statistical data mining for proteomic biomarker discovery
B) Verification of biomarkers
Protein binding assays
Mass spectrometric quantitative approaches
Multiple reaction monitoring
Pre-treatment strategies
Data mining & statistical analysis
C) Bioinformatics platforms in clinical proteomics
Databases for protein/Peptide data repository | |
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Protein/Peptide Database | Website/Link |
UniProt/Swiss Prot | |
Proteomics Identifications Database | |
MEROPS | |
PepBank | |
PeptideAtlas | |
ProteinProspector | |
MassMatrix |
Most cited repositories for biological pathways | ||
---|---|---|
Pathway databases | Biological pathway | Website/Link |
Reactome KnowledgeBase | Signal Transduction Pathway | |
BioCarta Pathway Diagrams | Signal Transduction Pathway | |
Pathway Commons | Signal Transduction Pathway | |
Protein ANalysis THrough Evolutionary Relationships | Signal Transduction Pathway | |
Protein Lounge | Signal Transduction Pathway | |
WikiPathways | Signal Transduction Pathway | |
Transcription Factor encyclopedia | Regulatory Pathways | |
Transcription Regulatory Regions Database | Regulatory Pathways | |
A Public Database of Transcription Factor and Regulatory Sequence Annotation | Regulatory Pathways | |
Homo Sapiens Comprehensive Model Collection (HOCOMOCO) | Regulatory Pathways | |
Transcription Factor Database | Regulatory Pathways | |
Human Protein Reference Database | Protein-Protein Interactions | |
Human Annotated and Predicted Protein Interaction Database | Protein-Protein Interactions | |
Biomolecular Interaction Network Database | Protein-Protein Interactions | |
Molecular Interaction Database | Protein-Protein Interactions | |
Biological General Repository for Interaction Datasets | Protein-Protein Interactions | |
Search Tool for the Retrieval of Interacting Genes/Proteins | Protein-Protein Interactions |
Applications of systems biology – disease diagnosis and treatment
D) Validation of biomarker candidates
Application of proteomics approaches in BCa biomarker discovery
Biomarker identification; Biomarker candidate/panels
|
Verification/Validation
|
Regulation
|
Potential clinical value; Biomarker performance
|
Ref.
|
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Gel-based approaches
| ||||
2DE
, n=24
| [178] | |||
Tissue: |
Western Blot
| ↑ in both NMIBC and MIBC; | Predict cancer progression | |
6 normal urothelium, 9 NMIBC, 9 MIBC |
Immunohistochemistry, n=24
| ↑ phosphorylation level of cofilin in BCa tissue samples (most prominent in MIBC). | Lack of evaluation of biomarker performance. | |
Cofilin
|
For both experiments, the same material was used as in the discovery phase.
| |||
IMAC, 1-SDS-PAGE
, n=35
|
Western Blot
| [182] | ||
Urine: | Aminopeptidase N, n=108 | Aminopeptidase N | Biomarker for cancer aggressiveness | |
Two pools from NMIBC, n1=9, n2=7 | Myeloblastin, n=97 | ↑ in MIBC | ||
Two pools from MIBC, n3 = 9, n4=10 |
ELISA
| Myeloblastin, Profilin 1 | Lack of evaluation of biomarker performance. | |
Aminopeptidase N, Myeloblastin,
| Profilin-1, n=82 | ↓ in MIBC | ||
Profilin-1
| ||||
Western Blot, 8 BCa cell line models | ↑ in BCa cases, association with stage | [181] | ||
DIGE
, n=14
|
Tissue microarray, n=292
| Diagnosis, staging, outcome prognosis | ||
Urine: | Primary urothelial cell carcinoma |
Detection of BCa:
| ||
7 BCa (positive cytology), 7 controls (negative cytology) |
ELISA, n=80
| |||
Reg- 1
| Urine: | 81.3% sensitivity | ||
32 BCa (positive cytology), 48 Controls | 81.2% specificity | |||
(negative cytology) | ||||
Gel-free approaches
| ||||
ELISA, n = 166
| [179] | |||
Urine, | ||||
LC-MS/MS
, n=20
| ||||
Urine: | For H2B: n=147, | ↑ level of H2B with cancer stage in urine and tissue samples | Prediction of disease progression, discrimination of tumor stages | |
Benign (n=5), pTa, pT1 (n=10), pT2+ (n=5) | For NIF-1: n = 158 | |||
In both groups urine from benign, NIMB (Ta, Ta) and MIBC (T2+) were included.
| ||||
histone H2B, NIF-1
| ||||
↓ level of NIF-1 with cancer stages (not agreement with urinary level) | Lack of evaluation of biomarker performance. | |||
Immunohistochemistry, n=32
| ||||
pTa, pT1, n=23, pT2+ n=9 | ||||
iTRAQ
, n=12
|
Immunohistochemistry, n=303
| ↑ in 4/6 BCa samples in comparison to control (iTRAQ); | Prediction of disease progression | [180] |
Tissue: | ||||
6 bladder cancer tissues (4 NMIBC, 2 MIBC) and paired normal tissues; | ||||
Inverse correlation to stage and histological grade progression (immunohistochemistry) | Lack of evaluation of biomarker performance. | |||
DDX39
| ||||
CE-MS
, n=248
|
CE-MS, n=130
| ↓ regulated in MIBC in comparison to NMIBC |
Prediction of MIBC:
| [92] |
Urine: | Urine, | 81% sensitivity | ||
127 BCa patients, 121 Controls | Test set: 68 NMIBC and 62 MIBC | 57% specificity | ||
4 polypeptide panel
| ||||
CE-MS
, n=79
|
CE-MS, n=366
| Varied; 10 peptides ↑ in BCa; | [93] | |
Urine: | Urine, | 12 ↓ in BCa in comparison to control |
Detection of BCa:
| |
46 BCa patients, 33 Controls |
(Test set includes healthy controls, patients with non-malignant and malignant urological disorders)
| 100 % sensitivity | ||
22 polypeptides panel
| 73% specificity |