Shotgun LC/MS proteomics of breast cancer sera from African American women
Authors: Padma Tadi-Uppala, Sharon Lum, Carlos Garberoglio, Guru Uppala, David Kirchner, Richard Katenhusen, Kumar Kolli, Richard Mural, Michael Liebman
Journal: Cancer Research (2008)

3949   African American (AA) women are less likely than Caucasian women to have breast cancer but are more likely to die from it. Recent research has indicated that breast tumor biology in AA women is different from that in Caucasian women. AA women are more likely to be diagnosed with breast cancer at an earlier age and with more aggressive form of the disease, characterized by higher grade and negative estrogen and progesterone receptor status.  The  purpose  of our study was to profile the serum proteins from healthy and cancer patients in order to identify differentially expressed proteins between the two groups, and to complement the LC/MS data to the protein expression data obtained previously from the 2D-DIGE/MS analysis on similar type of samples. This pilot study included six AA breast cancer women and six healthy AA controls.  Hypothesis : We expected that changes taking place in the mammary gland prior to the appearance of breast cancer will be reflected in proteins that will appear in the blood.  Methods : Serum samples were processed on IgY12 (Beckman) antibody column to remove the top 12 proteins. The flow through fraction was digested using trypsin for LC/MS analysis. The protein digest serum samples were analyzed in triplicates on a high performance LTQFT( thermo Electron) mass spectrometer coupled to an online Surveyor LC system equipped with an autosmapler. Samples were loaded on to a trap column using sample pump. MS pump was used for eluting the peptides onto an analytical column for further separation and online nano-ESI LC/MS/MS analysis. The raw MS data files were imported into DeCyderMS (GE Healthcare) module and data was processed for peptide/protein expression analysis.  Results:  DeCyder/MS analysis revealed significant altered expression of 215 peptides between the two groups (t-test:  2 and presence of peptides in all replicates was >2.0). Most of these peptides were mapped to 28 proteins in the Swiss-Prot database. DeCyderMS analysis on all the replicates from each sample are underway to identify the peptide pattern differences between the healthy and cancer samples. In addition to high abundant proteins identified by 2-D DIGE/MS in our earlier study, we identified low abundant proteins such as Angiotensinogen precursor, Insulin-like growth factor-binding protein complex acid labil, Vitamin D -binding protein precursor and others. Some of these proteins have been implicated in breast cancer and either served as individual biomarkers or are under investigation as potential biomarkers for early detection of breast cancer.  Conclusions:  A panel of proteins differed in abundance in the sera of AA cancer patients vs. healthy controls; these could serve as biomarkers of early-stage breast cancer. Future efforts will focus on further validation of this panel of biomarkers to gain insight into their role(s) in the etiology of aggressive breast tumors in AA women. Supported by The Susan G. Komen Breast Cancer Foundation.