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A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen.

Rudchenko S, Scanlan M, Kalantarov G, Yavelsky V, Levy C, Estabrook A, Old L, Chan GL, Lobel L, Trakht I - BMC Cancer (2008)

Bottom Line: Confocal microscopy revealed cell membrane and cytoplasmic localization of the target protein, which is consistent with previous studies of this protein.We have determined that GIPC1 is a novel breast cancer-associated immunogenic antigen that is overexpressed in breast cancer.Its role, however, in the initiation and/or progression of breast cancer remains unclear and needs further clarification.

View Article: PubMed Central - HTML - PubMed

Affiliation: College of Physicians and Surgeons, Columbia University, 630 W, 168 St,, New York, NY 10032, USA. RudchenkoS@HSS.EDU

ABSTRACT

Background: We have been studying the native autoimmune response to cancer through the isolation of human monoclonal antibodies that are cancer specific from cancer patients. To facilitate this work we previously developed a fusion partner cell line for human lymphocytes, MFP-2, that fuses efficiently with both human lymph node lymphocytes and peripheral blood lymphocytes. Using this unique trioma fusion partner cell line we isolated a panel of autologous human monoclonal antibodies, from both peripheral blood and lymph node lymphocytes, which are representative of the native repertoire of anti-cancer specific antibodies from breast cancer patients.

Methods: The current study employs immunocytochemistry, immunohistochemistry, Western blot analysis as well as Northern blots, Scatchard binding studies and finally SEREX analysis for target antigen identification.

Results: By application of an expression cloning technique known as SEREX, we determined that the target antigen for two monoclonal antibodies, 27.B1 and 27.F7, derived from lymph node B-cells of a breast cancer patient, is the PDZ domain-containing protein known as GIPC1. This protein is highly expressed not only in cultured human breast cancer cells, but also in primary and metastatic tumor tissues and its overexpression appears to be cancer cell specific. Confocal microscopy revealed cell membrane and cytoplasmic localization of the target protein, which is consistent with previous studies of this protein.

Conclusion: We have determined that GIPC1 is a novel breast cancer-associated immunogenic antigen that is overexpressed in breast cancer. Its role, however, in the initiation and/or progression of breast cancer remains unclear and needs further clarification.

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Related in: MedlinePlus

Human monoclonal 27.B1 stains the membrane and cytosol. Staining of SK-BR-3 cells and breast cancer tissue was performed with human monoclonal 27.B1. Staining of the SK-BR-3 breast cancer cell line was analyzed by confocal microscopy and indicates that the target antigen is present in the membrane and cytoplasm. Staining of human breast cancer tissue was analyzed by standard fluorescent microscopy.
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Figure 1: Human monoclonal 27.B1 stains the membrane and cytosol. Staining of SK-BR-3 cells and breast cancer tissue was performed with human monoclonal 27.B1. Staining of the SK-BR-3 breast cancer cell line was analyzed by confocal microscopy and indicates that the target antigen is present in the membrane and cytoplasm. Staining of human breast cancer tissue was analyzed by standard fluorescent microscopy.

Mentions: We previously described the construction of a unique fusion Partner cell line, MFP-2, and its use for the immortalization of both human peripheral blood and lymph node B-lymphocytes [24,25]. MFP-2 was employed for the generation of hybridoma cells from lymphocytes of breast cancer patients that produce autologous anti-breast cancer specific antibodies. The results of that study are described elsewhere [25]. One of these native human monoclonal antibodies, designated 27.B1 (IgM, k), was chosen for further study. It demonstrated an intensely positive reactivity with two human breast cancer cell lines, SK-BR-3 and MCF-7 and no reaction with normal diploid primary human fibroblasts as tested by cELISA (In cELISA whole cells are used in place of a purified antigen as in ELISA) [25]. Confocal microscopy with 27.B1 demonstrated the presence of the target antigen throughout the cytosol and in addition staining of the membrane was especially strong (see Figure 1). Furthermore, 27.B1 stained both primary and metastatic breast cancer with a high specificity and sensitivity [25]. These results along with a more detailed immunocytochemical and immunohistochemical analysis are described elsewhere [25].


A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen.

Rudchenko S, Scanlan M, Kalantarov G, Yavelsky V, Levy C, Estabrook A, Old L, Chan GL, Lobel L, Trakht I - BMC Cancer (2008)

Human monoclonal 27.B1 stains the membrane and cytosol. Staining of SK-BR-3 cells and breast cancer tissue was performed with human monoclonal 27.B1. Staining of the SK-BR-3 breast cancer cell line was analyzed by confocal microscopy and indicates that the target antigen is present in the membrane and cytoplasm. Staining of human breast cancer tissue was analyzed by standard fluorescent microscopy.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2529336&req=5

Figure 1: Human monoclonal 27.B1 stains the membrane and cytosol. Staining of SK-BR-3 cells and breast cancer tissue was performed with human monoclonal 27.B1. Staining of the SK-BR-3 breast cancer cell line was analyzed by confocal microscopy and indicates that the target antigen is present in the membrane and cytoplasm. Staining of human breast cancer tissue was analyzed by standard fluorescent microscopy.
Mentions: We previously described the construction of a unique fusion Partner cell line, MFP-2, and its use for the immortalization of both human peripheral blood and lymph node B-lymphocytes [24,25]. MFP-2 was employed for the generation of hybridoma cells from lymphocytes of breast cancer patients that produce autologous anti-breast cancer specific antibodies. The results of that study are described elsewhere [25]. One of these native human monoclonal antibodies, designated 27.B1 (IgM, k), was chosen for further study. It demonstrated an intensely positive reactivity with two human breast cancer cell lines, SK-BR-3 and MCF-7 and no reaction with normal diploid primary human fibroblasts as tested by cELISA (In cELISA whole cells are used in place of a purified antigen as in ELISA) [25]. Confocal microscopy with 27.B1 demonstrated the presence of the target antigen throughout the cytosol and in addition staining of the membrane was especially strong (see Figure 1). Furthermore, 27.B1 stained both primary and metastatic breast cancer with a high specificity and sensitivity [25]. These results along with a more detailed immunocytochemical and immunohistochemical analysis are described elsewhere [25].

Bottom Line: Confocal microscopy revealed cell membrane and cytoplasmic localization of the target protein, which is consistent with previous studies of this protein.We have determined that GIPC1 is a novel breast cancer-associated immunogenic antigen that is overexpressed in breast cancer.Its role, however, in the initiation and/or progression of breast cancer remains unclear and needs further clarification.

View Article: PubMed Central - HTML - PubMed

Affiliation: College of Physicians and Surgeons, Columbia University, 630 W, 168 St,, New York, NY 10032, USA. RudchenkoS@HSS.EDU

ABSTRACT

Background: We have been studying the native autoimmune response to cancer through the isolation of human monoclonal antibodies that are cancer specific from cancer patients. To facilitate this work we previously developed a fusion partner cell line for human lymphocytes, MFP-2, that fuses efficiently with both human lymph node lymphocytes and peripheral blood lymphocytes. Using this unique trioma fusion partner cell line we isolated a panel of autologous human monoclonal antibodies, from both peripheral blood and lymph node lymphocytes, which are representative of the native repertoire of anti-cancer specific antibodies from breast cancer patients.

Methods: The current study employs immunocytochemistry, immunohistochemistry, Western blot analysis as well as Northern blots, Scatchard binding studies and finally SEREX analysis for target antigen identification.

Results: By application of an expression cloning technique known as SEREX, we determined that the target antigen for two monoclonal antibodies, 27.B1 and 27.F7, derived from lymph node B-cells of a breast cancer patient, is the PDZ domain-containing protein known as GIPC1. This protein is highly expressed not only in cultured human breast cancer cells, but also in primary and metastatic tumor tissues and its overexpression appears to be cancer cell specific. Confocal microscopy revealed cell membrane and cytoplasmic localization of the target protein, which is consistent with previous studies of this protein.

Conclusion: We have determined that GIPC1 is a novel breast cancer-associated immunogenic antigen that is overexpressed in breast cancer. Its role, however, in the initiation and/or progression of breast cancer remains unclear and needs further clarification.

Show MeSH
Related in: MedlinePlus