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Proteomic Analysis of Urine to Identify Breast Cancer Biomarker Candidates Using a Label-Free LC-MS/MS Approach.

Beretov J, Wasinger VC, Millar EK, Schwartz P, Graham PH, Li Y - PLoS ONE (2015)

Bottom Line: Thirty-six urinary proteins were exclusively found in specific breast cancer stages, with 24 increasing and 12 decreasing in their abundance.Our results indicate that urine is a useful non-invasive source of biomarkers and the profile patterns (biomarkers) identified, have potential for clinical use in the detection of BC.Validation with a larger independent cohort of patients is required in the following study.

View Article: PubMed Central - PubMed

Affiliation: Cancer Care Centre, St George Hospital, Kogarah, Australia.

ABSTRACT

Introduction: Breast cancer is a complex heterogeneous disease and is a leading cause of death in women. Early diagnosis and monitoring progression of breast cancer are important for improving prognosis. The aim of this study was to identify protein biomarkers in urine for early screening detection and monitoring invasive breast cancer progression.

Method: We performed a comparative proteomic analysis using ion count relative quantification label free LC-MS/MS analysis of urine from breast cancer patients (n = 20) and healthy control women (n = 20).

Results: Unbiased label free LC-MS/MS-based proteomics was used to provide a profile of abundant proteins in the biological system of breast cancer patients. Data analysis revealed 59 urinary proteins that were significantly different in breast cancer patients compared to the normal control subjects (p<0.05, fold change >3). Thirty-six urinary proteins were exclusively found in specific breast cancer stages, with 24 increasing and 12 decreasing in their abundance. Amongst the 59 significant urinary proteins identified, a list of 13 novel up-regulated proteins were revealed that may be used to detect breast cancer. These include stage specific markers associated with pre-invasive breast cancer in the ductal carcinoma in-situ (DCIS) samples (Leucine LRC36, MAST4 and Uncharacterized protein CI131), early invasive breast cancer (DYH8, HBA, PEPA, uncharacterized protein C4orf14 (CD014), filaggrin and MMRN2) and metastatic breast cancer (AGRIN, NEGR1, FIBA and Keratin KIC10). Preliminary validation of 3 potential markers (ECM1, MAST4 and filaggrin) identified was performed in breast cancer cell lines by Western blotting. One potential marker MAST4 was further validated in human breast cancer tissues as well as individual human breast cancer urine samples with immunohistochemistry and Western blotting, respectively.

Conclusions: Our results indicate that urine is a useful non-invasive source of biomarkers and the profile patterns (biomarkers) identified, have potential for clinical use in the detection of BC. Validation with a larger independent cohort of patients is required in the following study.

No MeSH data available.


Related in: MedlinePlus

Expression of MAST4 in the individual urine samples from BC patients and controls.Western blotting was performed on the remaining urine samples from BC patients and control subjects to confirm the expression of MAST4. A. Overexpression of MAST4 was shown in the individual DCIS urine samples (DCIS 1–5), weak expression in IBC1 urine sample and no expression in control urine samples. B. There was no MAST4 expression found in the BBD urine sample but medium expression in IBC (IBC 2–5) and MBC urine samples (MBC 1–3). ß-tubulin was chosen as a loading control.
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pone.0141876.g006: Expression of MAST4 in the individual urine samples from BC patients and controls.Western blotting was performed on the remaining urine samples from BC patients and control subjects to confirm the expression of MAST4. A. Overexpression of MAST4 was shown in the individual DCIS urine samples (DCIS 1–5), weak expression in IBC1 urine sample and no expression in control urine samples. B. There was no MAST4 expression found in the BBD urine sample but medium expression in IBC (IBC 2–5) and MBC urine samples (MBC 1–3). ß-tubulin was chosen as a loading control.

Mentions: In order to confirm MAST4 overexpression in the individual DCIS patients, the remaining BC urine samples available were re-examined using Western blotting. Our results clearly indicate that high levels of MAST4 expression were found in the individual DCIS urine samples and low levels in IBC and MBC urine samples, while no expression was seen in the samples from BBD patients and normal health control subjects (see Fig 6), further confirming that a strong link exists between MAST4 protein and the DCIS urine samples identified with the LC-MS/MS.


Proteomic Analysis of Urine to Identify Breast Cancer Biomarker Candidates Using a Label-Free LC-MS/MS Approach.

Beretov J, Wasinger VC, Millar EK, Schwartz P, Graham PH, Li Y - PLoS ONE (2015)

Expression of MAST4 in the individual urine samples from BC patients and controls.Western blotting was performed on the remaining urine samples from BC patients and control subjects to confirm the expression of MAST4. A. Overexpression of MAST4 was shown in the individual DCIS urine samples (DCIS 1–5), weak expression in IBC1 urine sample and no expression in control urine samples. B. There was no MAST4 expression found in the BBD urine sample but medium expression in IBC (IBC 2–5) and MBC urine samples (MBC 1–3). ß-tubulin was chosen as a loading control.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4636393&req=5

pone.0141876.g006: Expression of MAST4 in the individual urine samples from BC patients and controls.Western blotting was performed on the remaining urine samples from BC patients and control subjects to confirm the expression of MAST4. A. Overexpression of MAST4 was shown in the individual DCIS urine samples (DCIS 1–5), weak expression in IBC1 urine sample and no expression in control urine samples. B. There was no MAST4 expression found in the BBD urine sample but medium expression in IBC (IBC 2–5) and MBC urine samples (MBC 1–3). ß-tubulin was chosen as a loading control.
Mentions: In order to confirm MAST4 overexpression in the individual DCIS patients, the remaining BC urine samples available were re-examined using Western blotting. Our results clearly indicate that high levels of MAST4 expression were found in the individual DCIS urine samples and low levels in IBC and MBC urine samples, while no expression was seen in the samples from BBD patients and normal health control subjects (see Fig 6), further confirming that a strong link exists between MAST4 protein and the DCIS urine samples identified with the LC-MS/MS.

Bottom Line: Thirty-six urinary proteins were exclusively found in specific breast cancer stages, with 24 increasing and 12 decreasing in their abundance.Our results indicate that urine is a useful non-invasive source of biomarkers and the profile patterns (biomarkers) identified, have potential for clinical use in the detection of BC.Validation with a larger independent cohort of patients is required in the following study.

View Article: PubMed Central - PubMed

Affiliation: Cancer Care Centre, St George Hospital, Kogarah, Australia.

ABSTRACT

Introduction: Breast cancer is a complex heterogeneous disease and is a leading cause of death in women. Early diagnosis and monitoring progression of breast cancer are important for improving prognosis. The aim of this study was to identify protein biomarkers in urine for early screening detection and monitoring invasive breast cancer progression.

Method: We performed a comparative proteomic analysis using ion count relative quantification label free LC-MS/MS analysis of urine from breast cancer patients (n = 20) and healthy control women (n = 20).

Results: Unbiased label free LC-MS/MS-based proteomics was used to provide a profile of abundant proteins in the biological system of breast cancer patients. Data analysis revealed 59 urinary proteins that were significantly different in breast cancer patients compared to the normal control subjects (p<0.05, fold change >3). Thirty-six urinary proteins were exclusively found in specific breast cancer stages, with 24 increasing and 12 decreasing in their abundance. Amongst the 59 significant urinary proteins identified, a list of 13 novel up-regulated proteins were revealed that may be used to detect breast cancer. These include stage specific markers associated with pre-invasive breast cancer in the ductal carcinoma in-situ (DCIS) samples (Leucine LRC36, MAST4 and Uncharacterized protein CI131), early invasive breast cancer (DYH8, HBA, PEPA, uncharacterized protein C4orf14 (CD014), filaggrin and MMRN2) and metastatic breast cancer (AGRIN, NEGR1, FIBA and Keratin KIC10). Preliminary validation of 3 potential markers (ECM1, MAST4 and filaggrin) identified was performed in breast cancer cell lines by Western blotting. One potential marker MAST4 was further validated in human breast cancer tissues as well as individual human breast cancer urine samples with immunohistochemistry and Western blotting, respectively.

Conclusions: Our results indicate that urine is a useful non-invasive source of biomarkers and the profile patterns (biomarkers) identified, have potential for clinical use in the detection of BC. Validation with a larger independent cohort of patients is required in the following study.

No MeSH data available.


Related in: MedlinePlus