Isotype selection for antibody-based cancer therapy

The clinical application of monoclonal antibodies (mAbs) has revolutionized the field of cancer therapy, as it has enabled the successful treatment of previously untreatable types of cancer. Different mechanisms play a role in the anti‐tumour effect of mAbs. These include blocking of tumour‐specific growth factor receptors or of immune modulatory molecules as well as complement and cell‐mediated tumour cell lysis. Thus, for many mAbs, Fc‐mediated effector functions critically contribute to the efficacy of treatment. As immunoglobulin (Ig) isotypes differ in their ability to bind to Fc receptors on immune cells as well as in their ability to activate complement, they differ in the immune responses they activate. Therefore, the choice of antibody isotype for therapeutic mAbs is dictated by its intended mechanism of action. Considering that clinical efficacy of many mAbs is currently achieved only in subsets of patients, optimal isotype selection and Fc optimization during antibody development may represent an important step towards improved patient outcome. Here, we discuss the current knowledge of the therapeutic effector functions of different isotypes and Fc‐engineering strategies to improve mAbs application

Effect of soil bacteriomes on mycorrhizal colonization by <i>Rhizophagus irregularis</i>:Interactive effects on maize (<i>Zea mays</i> L.) growth under salt stress

In this study, we investigated the interactive effects of the arbuscular mycorrhizal fungus (AMF) Rhizophagus irregularis and soil bacteriomes on maize growth under salt stress (100 mM NaCl) and also the effect of salt and bacteriomes on the mycorrhizal infection levels. We found that soil bacteriomes directly promoted the growth of maize and indirectly enhanced maize biomass by increasing mycorrhizal colonization levels, irrespective of salt stress. Although R. irregularis by itself had no maize growth-promoting effect even at a high mycorrhizal colonization level in roots, its benefits to maize were reflected in other aspects, evidenced by the significantly increased rate of arbuscule formation (a proxy for a functional plant-AMF nutritional exchange) under salinity. A negative correlation between arbuscule colonization and root biomass suggested R. irregularis expands the role of maize roots. Besides, the positive correlation between the overall AMF colonization level and shoot biomass supported the tenet of a positive contribution of R. irregularis to maize growth. Our findings suggest that soil bacteriomes interactively work with R. irregularis, modulating the growth of maize by affecting the colonization of AMF in roots

ras Oncogene Activation Does Not Induce Sensitivity to Natural Killer Cell&mdash;Mediated Lysis in Human Melanoma

An important phenomenon in tumor immunology that has come under recent attention is the impact of oncogene activation in tumor cells on the sensitivity to lysis by immune effector cells. Several studies suggested that transfer of an activated ras oncogene into cultured rodent fibroblasts induces susceptibility to natural killer cell (NK)-mediated lysis. Experiments using human tumor cells, however, have produced conflicting data on the effect of ras activation in this respect. In studying the activation of the oncogene c-myc, which is often overexpressed in human melanoma, we have found that in cell lines expressing high levels of Myc protein, the sensitivity to lysis by NK cells was dramatically increased due to reduced expression of Human Leukocyte Antigen B locus products. Since the N-ras oncogene was found to be activated in 15% of human melanomas, we examined the possibility that in melanoma, in analogy to the murine systems, the mutated ras oncogene may influence NK susceptibility of human melanoma cells. Two N-ras genes harboring frequently found mutations were cloned into an expression vector. Transfection of the IGR39D melanoma cell line with wildtype and mutant N-ras constructs yielded several ras-expressing clones that were tested for NK sensitivity. Neither high expression of the wildtype N-ras protein, nor expression of two mutant proteins (N61-arg, N61-lys) was shown to result in enhanced NK-mediated lysis. We conclude that activation of ras oncogenes does not lead to the induction of an NK-sensitive phenotype in human melanoma cells. J Invest Dermatol 103:117S&ndash;121S, 199

A novel efficient bispecific antibody format, combining a conventional antigen-binding fragment with a single domain antibody, avoids potential heavy-light chain mis-pairing

Due to the technical innovations in generating bispecific antibodies (BsAbs) in recent years, BsAbs have become important reagents for diagnostic and therapeutic applications. However, the difficulty of producing a heterodimer consisting of two different arms with high yield and purity constituted a major limitation for their application in academic and clinical settings. Here, we describe a novel Fc-containing BsAb format (Fab × sdAb-Fc) composed of a conventional antigen-binding fragment (Fab), and a single domain antibody (sdAb), which avoids heavy-light chain mis-pairing during antibody assembly. In this study, the Fab x sdAb-Fc BsAbs were efficiently produced by three widely used heavy-heavy chain heterodimerization methods: Knobs-into-holes (KIH), Charge-pairs (CP) and controlled Fab-arm exchange (cFAE), respectively. The novel Fab x sdAb-Fc format provided a rapid and efficient strategy to generate BsAb with high purity and a unique possibility to further purify desired BsAbs from undesired antibodies based on molecular weight (MW). Compared to conventional BsAb formats, the advantages of Fab x sdAb-Fc format may thus provide a straightforward opportunity to apply bispecific antibody principles to research and development of novel targets and pathways in diseases such as cancer and autoimmunity

PCR-based detection of mobile genetic elements in total community DNA

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Shortened hinge design of Fab x sdAb-Fc bispecific antibodies enhances redirected T-Cell killing of tumor cells

T cell engager (TCE) antibodies have emerged as promising cancer therapeutics that link cytotoxic T-cells to tumor cells by simultaneously binding to CD3E on T-cells and to a tumor-associated antigen (TAA) expressed by tumor cells. We previously reported a novel bispecific format, the IgG-like Fab x sdAb-Fc (also known as half-IG_VH-h-CH2-CH3), combining a conventional antigen-binding fragment (Fab) with a single domain antibody (sdAb). Here, we evaluated this Fab x sdAb-Fc format as a T-cell redirecting bispecific antibody (TbsAbs) by targeting mEGFR on tumor cells and mCD3E on T cells. We focused our attention specifically on the hinge design of the sdAb arm of the bispecific antibody. Our data show that a TbsAb with a shorter hinge of 23 amino acids (TbsAb.short) showed a significantly better T cell redirected tumor cell elimination than the TbsAb with a longer, classical antibody hinge of 39 amino acids (TbsAb.long). Moreover, the TbsAb.short form mediated better T cell-tumor cell aggregation and increased CD69 and CD25 expression levels on T cells more than the TbsAb.long form. Taken together, our results indicate that already minor changes in the hinge design of TbsAbs can have significant impact on the anti-tumor activity of TbsAbs and may provide a new means to improve their potency