Evaluation of pre-existent immunity in patients with primary breast cancer: molecular and cellular assays to quantify antigen-specific T lymphocytes in peripheral blood mononuclear cells.
OBJECTIVE
Breast cancers are known to frequently (over)express several well-characterized tumor-associated antigens (TAAs) such as carcinoembryonic antigen, MUC-1, Her-2/neu, and cancer/testis antigens such as NY-ESO-1, SSX-2, and members of the MAGE family. Whereas in melanoma patients, the detection of pre-existing T cell responses to tumor-associated differentiation antigens was a rationale to initiate several vaccination strategies, little is known thus far concerning tumor-specific immunity in breast cancer patients. The objectives of our study were (a) to modify and compare different immunodiagnostic T cell assays with regard to their suitability for clinical applications and (b) to determine endogenous TAA-specific T cell immunity of breast cancer patients at the time point of primary diagnosis.


METHODS
Using MUC-1- and Her-2/neu-derived HLA-A*0201-restricted peptides as model antigens, we analyzed antigen-dependent IFN-gamma release of T cells by enzyme-linked immunospot (ELISpot) assay, intracellular cytokine flow cytometry (CytoSpot), and quantitative real-time PCR. As an assay independent of T cell function, we performed tetramer staining.


RESULTS
In our hands, the quantitative real-time PCR method is most sensitive and a feasible screening test to perform an "immunological staging" of cancer patients. By doing this, we detected in 7 of 13 (54%) of HLA-A*0201(+) breast cancer patients a pre-existent specific cellular immune response to at least one of the investigated TAAs (MUC-1, Her-2/neu, carcinoembryonic antigen, NY-ESO-1, and SSX-2). Four of 21 patients (19%) were found to have a significant Her-2/neu-specific T cell response as defined by a stimulation index >/==" BORDER="0"> 2 (range, 10-88).


CONCLUSIONS
Although the clinical relevance of endogenous TAA-specific immunity remains unclear, our findings suggest that patients with primary breast cancer can mount a T cell immune response to their tumor that might be beneficially enhanced by TAA-dependent vaccination strategies in the adjuvant situation.
Rentzsch
Christine
C
Department of Obstetrics and Gynecology, University of Tübingen, 72076 Tübingen, Germany.
Kayser
Simone
S
Stumm
Susanne
S
Watermann
Iris
I
Walter
Steffen
S
Stevanoviç
Stefan
S
Wallwiener
Diethelm
D
Gückel
Brigitte
B
D000938:Antigen-Presenting Cells; D000951:Antigens, Neoplasm / Q000235:genetics / Q000276:immunology*; D001943:Breast Neoplasms / Q000276:immunology*; D002272:Carcinoembryonic Antigen / Q000235:genetics / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D015234:HLA-A Antigens / Q000276:immunology; D015789:HLA-A2 Antigen; D006801:Humans; D007111:Immunity, Cellular; D007371:Interferon-gamma / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000276:immunology; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology; D018396:Mucin-1 / Q000235:genetics / Q000276:immunology; D009363:Neoplasm Proteins / Q000235:genetics / Q000276:immunology; D009367:Neoplasm Staging; D010446:Peptide Fragments / Q000276:immunology; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics / Q000276:immunology; D012097:Repressor Proteins / Q000235:genetics / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013601:T-Lymphocytes / Q000276:immunology*
D000951:Antigens, Neoplasm; D002272:Carcinoembryonic Antigen; D015234:HLA-A Antigens; D015789:HLA-A2 Antigen; D007371:Interferon-gamma; D008565:Membrane Proteins; D018396:Mucin-1; D009363:Neoplasm Proteins; D010446:Peptide Fragments; D018719:Erb-b2 Receptor Tyrosine Kinases; D012097:Repressor Proteins; C104948:CTAG1B protein, human; C435939:HLA-A*02:01 antigen; C093962:synovial sarcoma X breakpoint proteins