The prevention and treatment of prevalent infectious diseases and tumors should benefit from improvements in the induction of antigen-specific T cell immunity. To assess the potential of antigen targeting to dendritic cells to improve immunity, we incorporated ovalbumin protein into a monoclonal antibody to the DEC-205 receptor, an endocytic receptor that is abundant on these cells in lymphoid tissues. Simultaneously, we injected agonistic α-CD40 antibody to mature the dendritic cells. We found that a single low dose of antibody-conjugated ovalbumin initiated immunity from the naive CD4+ and CD8+ T cell repertoire. Unexpectedly, the αDEC-205 antigen conjugates, given s.c., targeted to dendritic cells systemically and for long periods, and ovalbumin peptide was presented on MHC class I for 2 weeks. This was associated with stronger CD8+ T cell–mediated immunity relative to other forms of antigen delivery, even when the latter was given at a thousand times higher doses. In parallel, the mice showed enhanced resistance to an established rapidly growing tumor and to viral infection at a mucosal site. By better harnessing the immunizing functions of maturing dendritic cells, antibody-mediated antigen targeting via the DEC-205 receptor increases the efficiency of vaccination for T cell immunity, including systemic and mucosal resistance in disease models.

IntroductionMost antitumor chemotherapies kill tumor cells by the induction of apoptosis. However, in most (but not all) instances, chemotherapyinduced cell death does not lead to enhanced antitumor immunity. [1][2][3][4] Improved understanding of which specific agents lead to immunogenic cell death and the underlying mechanisms may facilitate the integration of chemotherapy and immunotherapy of cancer. Bortezomib is a specific inhibitor of 26S proteasome subunit that has shown promising clinical activity in several tumors. 5 Bortezomib induces apoptosis of human tumor cells by several mechanisms, including the inhibition of NFB and disruption of unfolded protein response. 5 We hypothesized that the mode of induction of tumor cell death may have a major impact on their immunogenicity. Here we show that the uptake of bortezomibkilled human myeloma cells by DCs leads to strong induction of antitumor immunity in culture without the need for any additional adjuvants and characterize the mechanism of enhanced immunogenicity. Patients, materials, and methods PatientsPeripheral blood and bone marrow samples from patients with multiple myeloma were obtained following informed consent, which was provided in accordance with the Declaration of Helsinki, approved by the institutional review boards of St Vincent's Cancer Center and the Rockefeller University. Isolation of primary tumor cellsMononuclear cells from blood or bone marrow were obtained by density gradient centrifugation using Ficoll Hypaque (GE HealthCare, Uppsala, Sweden). Bone marrow mononuclear cells were separated into tumor (CD138ϩ) and nontumor (CD138Ϫ) fractions using CD138 magnetic beads (Miltenyi Biotec, Auburn, CA). Tumor cells were cryopreserved in aliquots for use as source of antigen and targets. Cell linesMyeloma cell lines were obtained from American Type Culture Collection (U266, HLA-A2 positive; ATCC, Manassas, VA) or provided by J. Epstein, Arkansas Cancer Center, Little Rock, AR (cag, HLA-A2 negative). Other tumor cell lines used were NCEB1 (mantle cell lymphoma; gift of O. O'Connor, Memorial Sloan Kettering [MSKCC], New York, NY) and MCF-7 (breast cancer; gift of P. Livingston, MSKCC). Cell lines were grown in RPMI 1640 supplemented with 10% fetal calf serum (FCS)/glutamine/gentamicin. Generation of tumor-loaded DCsDCs were generated by culture of purified CD14 ϩ cells isolated from patients peripheral blood mononuclear cells or from buffy coats in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF) (Immunex, Thousand Oaks, CA) and interleukin-4 (IL-4) (R&D Systems, Minneapolis, MN) as described. 6 Tumor cells were killed by culture in the presence of bortezomib (100 nM) (Velcade, Millennium Pharmaceuticals, Cambridge, MA) or dexamethasone (1 M) for 24 hours or by ␥ irradiation (20 Gy). Extent of apoptosis was monitored by Submitted October 25, 2006; accepted February 7, 2007. Prepublished online as Blood First Edition Paper, February 13, 2007 DOI 10.1182 DOI 10. /blood-2006.The publication costs of this article were def...

We evaluated a possible association between the brain-derived neurotrophic factor (BDNF) gene and susceptibility to obsessive-compulsive disorder (OCD) by genotyping a number of single-nucleotide polymorphisms (SNPs) and one microsatellite marker from the extended BDNF locus in 164 triads with OCD. Extensive background linkage disequilibrium was observed at this locus. Single-locus transmission-distortion tests revealed significant evidence of association with the disease for all the BDNF gene markers tested, including a Val66Met variation affecting the sequence of the proBDNF protein. Analysis of multi-SNP haplotypes provided similar results. Haplotype transmission comparisons in this and previous studies point to a functionally distinct BDNF haplotype uniquely marked by the rare Met66 allele, which is undertransmitted and likely confers a protective effect in OCD and other psychiatric disorders.

SIGN-R1, a recently discovered C-type lectin expressed at high levels on macrophages within the marginal zone of the spleen, mediates the uptake of dextran polysaccharides by these phagocytes. We now find that encapsulated Streptococcus pneumoniae are rapidly cleared by these macrophages from the bloodstream, and that capture also takes place when different cell lines express SIGN-R1 after transfection. To assess the role of the capsular polysaccharide of S. pneumoniae (CPS) in the interaction of SIGN-R1 with pneumococci, we first studied binding and uptake of serotype 14 CPS in transfected cells. Binding was observed and was of a much higher avidity (3,000-fold) for CPS 14 than dextran. The CPSs from four different serotypes were also cleared by marginal zone macrophages in vivo. To establish a role for SIGN-R1 in this uptake, we selectively down-regulated expression of the lectin by pretreatment of the mice with SIGN-R1 antibodies, including a newly generated hamster monoclonal called 22D1. For several days after this transient knockout, the marginal zone macrophages were unable to take up either CPSs or dextrans. Therefore, marginal zone macrophages in mice have a receptor that interacts with capsular pneumococcal polysaccharides, setting the stage for further studies of the functional consequences of this interaction.T he spleen functions at several points in innate and adaptive immunity. A major innate function is exerted by macrophages that are abundant in vascular regions termed the splenic red pulp, whereas adaptive functions are carried out by B and T lymphocytes, typically located in white pulp nodules. At the junction of each white pulp nodule with the red pulp is a specialized region called the marginal zone, which is composed of several concentric regions (1). Innermost is a ring of macrophages termed marginal metallophils, expressing a hemagglutinin termed sialoadhesin or CD169 (2, 3). Then there is a vascular sinus that receives blood via penetrating small arterial vessels from the white pulp. Surrounding the marginal sinus is a zone composed of large macrophages as well as specialized B lymphocytes (4). Within and surrounding the marginal zone are also dendritic cells (5, 6), possibly in the process of migrating from the blood to the T cell regions of the white pulp.With respect to host defense, the spleen plays a special role during blood-borne infection with encapsulated microorganisms, particularly Streptococcus pneumoniae bacteria (7-12). A critical role of the spleen is the formation of antibodies by marginal zone B cells (13-15), particularly complement-fixing antibodies (16)(17)(18)(19)(20). The role of macrophages in the processes of microbial clearance and resistance and antibody formation to S. pneumoniae needs to be considered (21), particularly given recent data that marginal zone macrophages interact and retain B cells in this region (22). Here we show that marginal zone macrophages express a receptor called SIGN-R1 that is able to bind and internalize the capsular pneumococcal polys...