Autoantibody Repertoire in APECED Patients Targets Two Distinct Subgroups of Proteins.

High titer autoantibodies produced by B lymphocytes are clinically important features of many common autoimmune diseases. APECED patients with deficient autoimmune regulator (AIRE) gene collectively display a broad repertoire of high titer autoantibodies, including some which are pathognomonic for major autoimmune diseases. AIRE deficiency severely reduces thymic expression of gene-products ordinarily restricted to discrete peripheral tissues, and developing T cells reactive to those gene-products are not inactivated during their development. However, the extent of the autoantibody repertoire in APECED and its relation to thymic expression of self-antigens are unclear. We here undertook a broad protein array approach to assess autoantibody repertoire in APECED patients. Our results show that in addition to shared autoantigen reactivities, APECED patients display high inter-individual variation in their autoantigen profiles, which collectively are enriched in evolutionarily conserved, cytosolic and nuclear phosphoproteins. The APECED autoantigens have two major origins; proteins expressed in thymic medullary epithelial cells and proteins expressed in lymphoid cells. These findings support the hypothesis that specific protein properties strongly contribute to the etiology of B cell autoimmunity.

Monoclonal antibody blockade of the human Eag1 potassium channel function exerts antitumor activity.

The potassium channel ether à go-go has been directly linked to cellular proliferation and transformation, although its physiologic role(s) are as of yet unknown. The specific blockade of human Eag1 (hEag1) may not only allow the dissection of the role of the channel in distinct physiologic processes, but because of the implication of hEag1 in tumor biology, it may also offer an opportunity for the treatment of cancer. However, members of the potassium channel superfamily are structurally very similar to one another, and it has been notoriously difficult to obtain specific blockers for any given channel. Here, we describe and validate the first rational design of a monoclonal antibody that selectively inhibits a potassium current in intact cells. Specifically blocking hEag1 function using this antibody inhibits tumor cell growth both in vitro and in vivo. Our data provide a proof of concept that enables the generation of functional antagonistic monoclonal antibodies against ion channels with therapeutic potential. The particular antibody described here, as well as the technique developed to make additional functional antibodies to Eag1, makes it possible to evaluate the potential of the channel as a target for cancer therapy.

Identification of MMP-15 as an anti-apoptotic factor in cancer cells.

We have performed an in vitro selection for an anti-apoptotic phenotype that resembles the selection process that pre-malignant cells undergo in the initial phase of carcinogenesis in vivo. Using the cervical carcinoma cell line HeLa S3 as a model system, the selection procedure yielded cell clones that displayed increased resistance to apoptosis induced by Fas, tumor necrosis factor-related apoptosis-inducing ligand, and serum starvation. Gene expression profiling using gene family focused cDNA arrays revealed numerous genes that are differentially expressed in HeLa S3 and the resistant subclones and therefore are potentially involved in the definition of sensitivity to apoptotic stimuli. From the genes identified in this functional genomics approach we validated the anti-apoptotic activity of the membrane-anchored matrix metalloproteinase 15 (MMP-15) by means of small interfering RNA-mediated knock-down and ectopic expression in parental HeLa S3 cells and, to confirm a more general significance of our findings, in other cancer cell lines. The in vivo relevance of these findings is supported by the overexpression of MMP-15 in human lung adenocarcinoma compared with normal lung. Because MMP-15 is known to promote invasion, our results suggest that this protease connects metastasis and apoptosis resistance by an unknown regulatory mechanism. Our findings therefore strongly suggest that cancer characteristics such as metastatic potential, which are thought to evolve late in cancer progression, could be manifested early on by selection for an anti-apoptotic phenotype.