Prophylactic and therapeutic activity of fully human monoclonal antibodies directed against Influenza A M2 protein

Influenza virus infection is a prevalent disease in humans. Antibodies against hemagglutinin have been shown to prevent infection and hence hemagglutinin is the major constituent of current vaccines. Antibodies directed against the highly conserved extracellular domain of M2 have also been shown to mediate protection against Influenza A infection in various animal models. Active vaccination is generally considered the best approach to combat viral diseases. However, passive immunization is an attractive alternative, particularly in acutely exposed or immune compromized individuals, young children and the elderly. We recently described a novel method for the rapid isolation of natural human antibodies by mammalian cell display. Here we used this approach to isolate human monoclonal antibodies directed against the highly conserved extracellular domain of the Influenza A M2 protein. The identified antibodies bound M2 peptide with high affinities, recognized native cell-surface expressed M2 and protected mice from a lethal influenza virus challenge. Moreover, therapeutic treatment up to 2 days after infection was effective, suggesting that M2-specific monoclonals have a great potential as immunotherapeutic agents against Influenza infection

Isolation of human monoclonal antibodies by mammalian cell display

Due to their low immunogenicity in patients, humanized or fully human mAbs are becoming increasingly important for the treatment of a growing number of diseases, including cancer, infections, and immune disorders. Here, we describe a technology allowing for the rapid isolation of fully human mAbs. In contrast to previously described methods, B cells specific for an antigen of interest are directly isolated from peripheral blood mononuclear cells (PBMC) of human donors. Recombinant, antigen-specific single-chain Fv (scFv) libraries are generated from this pool of B cells and screened by mammalian cell surface display by using a Sindbis virus expression system. This method allows isolating antigen-specific antibodies by a single round of FACS. The variable regions (VRs) of the heavy chains (HCs) and light chains (LCs) are isolated from positive clones and recombinant fully human antibodies produced as whole IgG or Fab fragments. In this manner, several hypermutated high-affinity antibodies binding the Qβ virus like particle (VLP), a model viral antigen, as well as antibodies specific for nicotine were isolated. All antibodies showed high expression levels in cell culture. The human nicotine-specific mAbs were validated preclinically in a mouse model. Thus, the technology presented here allows for rapid isolation of high-affinity, fully human antibodies with therapeutic potential from human volunteers

Secretory phospholipase A2-IID is an effector molecule of CD4+CD25+ regulatory T cells

Suppression by natural CD4+CD25+ regulatory T cells (Tregs) is one mechanism by which tolerance is maintained. However, the way in which Tregs mediate suppression is not well understood. Here, we show that secreted phospholipase A2 (sPLA2)-IID is selectively produced by Tregs. sPLA2-IID is a potent mediator of Treg function, because it strongly suppressed proliferation of CD4+ and CD8+ T cells in vitro and in vivo in a manner independent of its catalytic activity. Furthermore, sPLA2-IID promoted the differentiation of Tregs, presumably via attenuating signaling through the PI3K/Akt/mammalian target of rapamycin pathway. Importantly, administration of a sPLA2-IID-Fc fusion protein inhibited disease development in murine models of colitis and multiple sclerosis, suggesting that sPLA2-IID's immunosuppressive function might be exploited therapeutically