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

<p>Abstract</p> <p>Background</p> <p>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.</p> <p>Methods</p> <p>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.</p> <p>Results</p> <p>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.</p> <p>Conclusion</p> <p>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.</p

DNA Methylation Profiling of Human Prefrontal Cortex Neurons in Heroin Users Shows Significant Difference between Genomic Contexts of Hyper- and Hypomethylation and a Younger Epigenetic Age.

We employed Illumina 450 K Infinium microarrays to profile DNA methylation (DNAm) in neuronal nuclei separated by fluorescence-activated sorting from the postmortem orbitofrontal cortex (OFC) of heroin users who died from heroin overdose (N = 37), suicide completers (N = 22) with no evidence of heroin use and from control subjects who did not abuse illicit drugs and died of non-suicide causes (N = 28). We identified 1298 differentially methylated CpG sites (DMSs) between heroin users and controls, and 454 DMSs between suicide completers and controls (p &lt; 0.001). DMSs and corresponding genes (DMGs) in heroin users showed significant differences in the preferential context of hyper and hypo DM. HyperDMSs were enriched in gene bodies and exons but depleted in promoters, whereas hypoDMSs were enriched in promoters and enhancers. In addition, hyperDMGs showed preference for genes expressed specifically by glutamatergic as opposed to GABAergic neurons and enrichment for axonogenesis- and synaptic-related gene ontology categories, whereas hypoDMGs were enriched for transcription factor activity- and gene expression regulation-related terms. Finally, we found that the DNAm-based "epigenetic age" of neurons from heroin users was younger than that in controls. Suicide-related results were more difficult to interpret. Collectively, these findings suggest that the observed DNAm differences could represent functionally significant marks of heroin-associated plasticity in the OFC

DNA Methylation Profiling of Human Prefrontal Cortex Neurons in Heroin Users Shows Significant Difference between Genomic Contexts of Hyper- and Hypomethylation and a Younger Epigenetic Age

We employed Illumina 450 K Infinium microarrays to profile DNA methylation (DNAm) in neuronal nuclei separated by fluorescence-activated sorting from the postmortem orbitofrontal cortex (OFC) of heroin users who died from heroin overdose (N = 37), suicide completers (N = 22) with no evidence of heroin use and from control subjects who did not abuse illicit drugs and died of non-suicide causes (N = 28). We identified 1298 differentially methylated CpG sites (DMSs) between heroin users and controls, and 454 DMSs between suicide completers and controls (p &lt; 0.001). DMSs and corresponding genes (DMGs) in heroin users showed significant differences in the preferential context of hyper and hypo DM. HyperDMSs were enriched in gene bodies and exons but depleted in promoters, whereas hypoDMSs were enriched in promoters and enhancers. In addition, hyperDMGs showed preference for genes expressed specifically by glutamatergic as opposed to GABAergic neurons and enrichment for axonogenesis- and synaptic-related gene ontology categories, whereas hypoDMGs were enriched for transcription factor activity- and gene expression regulation-related terms. Finally, we found that the DNAm-based “epigenetic age” of neurons from heroin users was younger than that in controls. Suicide-related results were more difficult to interpret. Collectively, these findings suggest that the observed DNAm differences could represent functionally significant marks of heroin-associated plasticity in the OFC

DNA Methylation Profiling of Human Prefrontal Cortex Neurons in Heroin Users Shows Significant Difference between Genomic Contexts of Hyper- and Hypomethylation and a Younger Epigenetic Age

We employed Illumina 450 K Infinium microarrays to profile DNA methylation (DNAm) in neuronal nuclei separated by fluorescence-activated sorting from the postmortem orbitofrontal cortex (OFC) of heroin users who died from heroin overdose (N = 37), suicide completers (N = 22) with no evidence of heroin use and from control subjects who did not abuse illicit drugs and died of non-suicide causes (N = 28). We identified 1298 differentially methylated CpG sites (DMSs) between heroin users and controls, and 454 DMSs between suicide completers and controls (p &lt; 0.001). DMSs and corresponding genes (DMGs) in heroin users showed significant differences in the preferential context of hyper and hypo DM. HyperDMSs were enriched in gene bodies and exons but depleted in promoters, whereas hypoDMSs were enriched in promoters and enhancers. In addition, hyperDMGs showed preference for genes expressed specifically by glutamatergic as opposed to GABAergic neurons and enrichment for axonogenesis- and synaptic-related gene ontology categories, whereas hypoDMGs were enriched for transcription factor activity- and gene expression regulation-related terms. Finally, we found that the DNAm-based “epigenetic age” of neurons from heroin users was younger than that in controls. Suicide-related results were more difficult to interpret. Collectively, these findings suggest that the observed DNAm differences could represent functionally significant marks of heroin-associated plasticity in the OFC

A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen-3

St epithelium cell line HBL100 and breast cancer cell lines MCF-7, T47D, SK-BR-3, MDA231, MDA157 and MDA453. A probe for the GAPDH gene was used to normalize expression. Panel B: Densitometry analysis of the Northern blot was performed to quantitate the mRNA expression. The data indicates that the GIPC1 gene is upregulated in breast cancer cell lines.<p><b>Copyright information:</b></p><p>Taken from "A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen"</p><p>http://www.biomedcentral.com/1471-2407/8/248</p><p>BMC Cancer 2008;8():248-248.</p><p>Published online 24 Aug 2008</p><p>PMCID:PMC2529336.</p><p></p

A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen-2

This suggests that two populations of GIPC1 molecules exist in these cells and may be related to the protein doublet identified by Western blot analysis in Figure 2.<p><b>Copyright information:</b></p><p>Taken from "A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen"</p><p>http://www.biomedcentral.com/1471-2407/8/248</p><p>BMC Cancer 2008;8():248-248.</p><p>Published online 24 Aug 2008</p><p>PMCID:PMC2529336.</p><p></p

A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen-1

displayed. Human Ig H and L chains are present in the tissue and are recognized by the secondary anti-human antiserum. The target antigen is detected as a doublet in both the breast cancer tissue and SK-BR-3 cell line but is not detected in normal breast tissue. Both bands in the doublet are present in all breast cancer cell lines analyzed by Western blot but their intensity is variable. B. Immunoblotting with 27.B1 antibody of total cell lysates prepared from SK-BR-3 cells. 27.B1 antibody was preincubated with recombinant GIPC1 protein prior to blotting (lane 1), and compared to non-preincubated control (lane 2).<p><b>Copyright information:</b></p><p>Taken from "A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen"</p><p>http://www.biomedcentral.com/1471-2407/8/248</p><p>BMC Cancer 2008;8():248-248.</p><p>Published online 24 Aug 2008</p><p>PMCID:PMC2529336.</p><p></p

A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen-0

S 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.<p><b>Copyright information:</b></p><p>Taken from "A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen"</p><p>http://www.biomedcentral.com/1471-2407/8/248</p><p>BMC Cancer 2008;8():248-248.</p><p>Published online 24 Aug 2008</p><p>PMCID:PMC2529336.</p><p></p

A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen-4

S 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.<p><b>Copyright information:</b></p><p>Taken from "A human monoclonal autoantibody to breast cancer identifies the PDZ domain containing protein GIPC1 as a novel breast cancer-associated antigen"</p><p>http://www.biomedcentral.com/1471-2407/8/248</p><p>BMC Cancer 2008;8():248-248.</p><p>Published online 24 Aug 2008</p><p>PMCID:PMC2529336.</p><p></p