However, while ADCC activity against CE1176

However, while ADCC activity against CE1176.A3 was significantly improved by IgG3, particularly for PGT151, 10E8v4, 3BNC117 and VRC01 (Supplementary Figure3F), this was not true of 246.F3.C10.2 or BJOX002000.03.2, where mixed isotype-driven effects were observed. To understand epitope effects, we assessed the effect BAY 293 BAY 293 of IgG3 variant for each bNAb pair, represented as fold change between IgG3 and IgG1 (Figure2C). significantly improved binding to FcRIIa which correlated with enhanced phagocytosis of all trimeric Env antigens. Differences in ADCC were epitope-specific, with IgG3 bNAbs to the MPER, CD4 binding site and gp120-gp41 interface showing increased ADCC. We also explored the pH dependence of IgG1 and IgG3 variants for FcRn binding, as this determines the half-life of antibodies. We observed reduced pH dependence, associated with shorter half-lives for IgG3 bNAbs, with -light chains. However, IgG3 bNAbs that use -light chains showed similar pH dependence to their IgG1 counterparts. This study supports the manipulation of the constant region to improve both the neutralizing and Fc effector activity of BAY 293 bNAbs, and suggests that Rabbit Polyclonal to NMDAR1 IgG3 versions of bNAbs may be preferable for passive immunity given their polyfunctionality. Keywords:broadly neutralizing antibodies (bnAbs), Fc effector function, IgG3, phagocytosis, ADCC (antibody dependent cellular cytotoxicity) == Introduction == Antibodies mediate pathogen neutralization through the binding of the Fab portion to antigen, and also elicit several effector functions through interaction of the Fc with a variety of receptors. Both neutralization and Fc effector function have been shown to be criticalin vivo. The importance of neutralization has been confirmed by passive transfer studies, where bNAbs provide sterilizing immunity in animal models (13) and robust antiviral activity in chronically infected humans (4,5). Despite this, combinations of bNAbs with high levels of potency and breadth will be needed to improve on the results of the antibody-mediated protection (AMP) passive immunization trial with VRC01 (6). Fc receptor engagement results in the recruitment of cytotoxic functions which have been shown in HIV infection to BAY 293 restrict the number of transmitted/founder viruses that establish infection, reduce viral load, and drive viral escape (710) and are associated with spontaneous HIV control (11) and slowed disease progression (12). Polyfunctional Fc effector function has also been associated with vaccine protection in humans and non-human primates (1319) and with the development of broadly neutralizing antibodies (bNAbs) during infection (20,21). Further, Fc receptor binding is required for several bNAbs to optimally protect from infection or clear infected cells in different animal models (2228). Thus, while the elicitation of HIV bNAbs is likely necessary for an efficacious vaccine, Fc effector function can complement this function to improve efficacy. Among the factors that contribute to modulating Fc effector function is antibody isotype (IgM, IgA, IgG and IgE) and subclass (IgG1-4 and IgA1-2), determined by sequence variation in the constant regions of the heavy chain (CH1-3) genes. The unique structures of each isotype result in differential binding to multiple Fc receptors, and this translates to diversity of Fc effector functions, varying half-lives and immune complex formation (29). Furthermore, there is substantial evidence that isotype can significantly alter antigen affinity and/or neutralization capacity of monoclonal antibodies (3035), indicating the importance of the isotype well beyond Fc receptor binding. Isotype therefore potentially represents an important factor to improve the function of bNAbs for passive immunization. Of the IgG subclasses, IgG3 antibodies are the most polyfunctional, owing to their increased affinity for Fc receptors (36). IgG3 is highly polymorphic with 29 reported alleles (37), and this variability is known to alter antibody activity and half-life (34,38,39). Structurally, IgG3 is distinct from other subclasses, with a long flexible hinge, enabling high rotational freedom about the Fc-Fab and Fab-Fab axes (40,41). In HIV infection, skewing towards IgG3 has been associated with reduced risk of infection in the RV144 and HVTN 505 vaccine trials (14,15,42) and in viral control (11). IgG3 specific bNAbs have also been shown to mediate greater antibody-dependent cellular phagocytosis (ADCP) compared to IgG1 (43,44), largely through BAY 293 their elongated hinge (34,45). In addition to better ADCP, we previously demonstrated that IgG3 variants of the V2-specific bNAb CAP256-VRC26.25 showed enhanced antibody-dependent cellular trogocytosis (ADCT) and significantly improved neutralization potency when compared to IgG1 (34). IgG3 is however not currently used for any therapeutic antibodies in a clinical setting. One of the major reasons for.