Targeting antigens through blood dendritic cell antigen 2 on plasmacytoid dendritic cells promotes immunologic tolerance.

The C-type lectin receptor blood dendritic cell Ag 2 (BDCA2) is expressed exclusively on human plasmacytoid DCs (pDCs) and plays a role in Ag capture, internalization, and presentation to T cells. We used transgenic mice that express human BDCA2 and anti-BDCA2 mAbs to deliver Ags directly to BDCA2 on pDCs in vivo. Targeting Ag to pDCs in this manner resulted in significant suppression of Ag-specific CD4(+) T cell and Ab responses upon secondary exposure to Ag in the presence of adjuvant.

T cell activation inhibitors reduce CD8+ T cell and pro-inflammatory macrophage accumulation in adipose tissue of obese mice.

Adipose tissue inflammation and specifically, pro-inflammatory macrophages are believed to contribute to insulin resistance (IR) in obesity in humans and animal models. Recent studies have invoked T cells in the recruitment of pro-inflammatory macrophages and the development of IR. To test the role of the T cell response in adipose tissue of mice fed an obesogenic diet, we used two agents (CTLA-4 Ig and anti-CD40L antibody) that block co-stimulation, which is essential for full T cell activation.

CD20-directed small modular immunopharmaceutical, TRU-015, depletes normal and malignant B cells.

Purpose: CD20-directed therapy with rituximab is effective in many patients with malignant lymphoma or follicular lymphoma. However, relapse frequently occurs within 1 year, and patients become increasingly refractory to retreatment. Our purpose was to produce a compact, single-chain CD20-targeting immunotherapeutic that could offer therapeutic advantages in the treatment of B-cell lymphoma.

Targeting CD37-positive lymphoid malignancies with a novel engineered small modular immunopharmaceutical.

CD37 is a lineage-specific B-cell antigen that to date has been neglected as an attractive therapeutic target. To exploit this, novel CD37-specific small modular immunopharmaceuticals (CD37-SMIP) that include variable regions linked to modified human IgG(1) hinge, CH(2), and CH(3) domains were designed. The lead CD37-SMIP molecule induces potent apoptosis in the presence of a cross-linker, and antibody-dependent cellular cytotoxicity against B-cell leukemia/lymphoma cell lines and primary chronic lymphocytic leukemia (CLL) cells superior to therapeutic antibodies used in these diseases.

Development of a CA125-mesothelin cell adhesion assay as a screening tool for biologics discovery.

Preventing peritoneal implantation of ovarian carcinoma cells could prolong patient remission and survival. CA125 is expressed on most ovarian cancer cells and was reported to be a ligand of mesothelin, a peritoneal protein. We developed a cell adhesion assay with CA125-expresser ovarian cancer cells and human mesothelin-transfected cells and we confirmed that CA125 and mesothelin mediate cell attachment.

The HE4 (WFDC2) protein is a biomarker for ovarian carcinoma.

The WFDC2 (HE4) gene is amplified in ovarian carcinomas, whereas its expression in normal tissues, including ovary, is low. Although the function of the HE4 protein is unknown,it is a member of a family of stable 4-disulfide core proteins that are secreted at high levels. We therefore performed experiments to explore whether quantitation of HE4 protein levels in serum can be used as a biomarker for ovarian carcinoma.

Expression and characterization of recombinant soluble human CD3 molecules: presentation of antigenic epitopes defined on the native TCR-CD3 complex.

The TCR-CD3 complex consists of the clonotypic disulfide-linked TCRalphabeta or TCRdeltagamma heterodimers, and the invariant CD3delta, epsilon, gamma and zeta chains. We generated plasmid constructs expressing the extracellular domains of the CD3delta, epsilon or gamma subunits fused to human IgG1 Fc. Recombinant fusion proteins consisting of individual CD3delta, epsilon or gamma subunits reacted poorly with anti-CD3 mAb including G19-4, BC3, OKT3 and 64.

Gene therapy for cancer using single-chain Fv fragments specific for 4-1BB.

Monoclonal antibodies against the T-cell activation molecule 4-1BB have been effective in the treatment of established mouse tumors. To create a vaccine that stimulates the immune system similarly to the efficacious monoclonal anti-4-1BB antibody, 1D8, we constructed a vector encoding cell-bound single-chain Fv fragments from 1D8. We transfected the vector into cells from the K1735 melanoma, selected because of its low immunogenicity and very low expression of major histocompatibility complex class I.

Cutting edge: CD83 regulates the development of cellular immunity.

We recently found that human CD83, a marker of mature dendritic cells, is an adhesion receptor that binds to resting monocytes and a subset of activated CD8(+) T cells. We injected CD83-Ig into mice transplanted with the immunogenic P815 mastocytoma and showed that it significantly enhanced the rate of tumor growth and inhibited the development of cytotoxic T cells. In contrast, mice immunized with CD83-transfected K1735 cells, a poorly immunogenic melanoma, could prevent the outgrowth of wild-type K1735 cells.

Tumor necrosis factor-alpha regulates the expression of inducible costimulator receptor ligand on CD34(+) progenitor cells during differentiation into antigen presenting cells.

The inducible costimulator receptor (ICOS) is a third member of the CD28 receptor family that regulates T cell activation and function. ICOS binds to a newly identified ligand on antigen presenting cells different from the CD152 ligands CD80 and CD86. We used soluble ICOSIg and a newly developed murine anti-human ICOS ligand (ICOSL) monoclonal antibody to further characterize the ICOSL during ontogeny of antigen presenting cells.

ICOS ligand costimulation is required for T-cell encephalitogenicity.

The interaction of ICOS with its ligand on APC provides a costimulatory signal to previously activated T-cells. In these studies, we blocked the ICOS:ICOS ligand interaction with ICOS-Ig during the in vitro activation of MBP-reactive transgenic CD4(+) T-cells. The presence of ICOS-Ig in these cultures inhibited the ability of the transgenic T-cells to transfer EAE, although they entered the brains of the recipient mice.

CD3-mediated activation of tumor-reactive lymphocytes from patients with advanced cancer.

Lymphocytes from blood or tumors of patients with advanced cancer did not proliferate and produced very low levels of tumor necrosis factor and IFN-gamma when cultured with autologous tumor cells. Proliferation and lymphokine production dramatically increased in the presence of beads conjugated with mAbs to CD3 plus mAbs to CD28 and/or CD40, and the lymphocytes destroyed the tumor cells. Expression density of CD3 concomitantly increased from low to normal levels.

CD83 is an I-type lectin adhesion receptor that binds monocytes and a subset of activated CD8+ T cells [corrected].

To help determine CD83 function, a cDNA encoding a soluble protein containing the CD83 extracellular domain was fused with a mutated human IgG1 constant region (CD83Ig) and expressed by stable transfection of Chinese hamster ovary cells. Purified CD83Ig bound to peripheral blood monocytes and a subset of activated CD3(+)CD8(+) lymphocytes but did not bind to FcR. Monocytes that had adhered to plastic lost their ability to bind to CD83Ig after 90 min of in vitro incubation.

Characterization of human inducible costimulator ligand expression and function.

The inducible costimulator (ICOS) is the newest member of the CD28/CD152 receptor family involved in regulating T cell activation. We constructed a soluble-Ig fusion protein of the extracellular domain of human ICOS and used it as a probe to characterize expression patterns of the ICOS ligand (ICOSL). ICOSIg did not bind to CD80- or CD86-transfected Chinese hamster ovary cell lines, demonstrating that ICOSL is distinct from those ligands identified for CD28/CD152.