Enhanced phagocytic activity of HIV-specific antibodies correlates with natural production of immunoglobulins with skewed affinity for FcγR2a and FcγR2b.

While development of an HIV vaccine that can induce neutralizing antibodies remains a priority, decades of research have proven that this is a daunting task. However, accumulating evidence suggests that antibodies with the capacity to harness innate immunity may provide some protection. While significant research has focused on the cytolytic properties of antibodies in acquisition and control, less is known about the role of additional effector functions. In this study, we investigated antibody-dependent phagocytosis of HIV immune complexes, and we observed significant differences in the ability of antibodies from infected subjects to mediate this critical effector function. We observed both quantitative differences in the capacity of antibodies to drive phagocytosis and qualitative differences in their FcγR usage profile. We demonstrate that antibodies from controllers and untreated progressors exhibit increased phagocytic activity, altered Fc domain glycosylation, and skewed interactions with FcγR2a and FcγR2b in both bulk plasma and HIV-specific IgG. While increased phagocytic activity may directly influence immune activation via clearance of inflammatory immune complexes, it is also plausible that Fc receptor usage patterns may regulate the immune response by modulating downstream signals following phagocytosis--driving passive degradation of internalized virus, release of immune modulating cytokines and chemokines, or priming of a more effective adaptive immune response.

Evaluation of Delcath Systems' Generation 2 (GEN 2) melphalan hemofiltration system in a porcine model of percutaneous hepatic perfusion.

PURPOSE: A new melphalan hemoperfusion filter (GEN 2) was evaluated in a simulated-use porcine model of percutaneous hepatic perfusion (PHP). The current study evaluated melphalan filtration efficiency, the transfilter pressure gradient, and the removal of specific blood products. MATERIALS AND METHODS: A porcine PHP procedure using the GEN 2 filter was performed under Good Laboratory Practice conditions to model the 60-min clinical PHP procedure. RESULTS: The mean filter efficiency for removing melphalan in six filters was 99.0 ± 0.4 %. The transfilter pressure gradient across the filter averaged 20.9 mmHg for the 60-min procedure. Many blood components, including albumin and platelets, decreased on average from 3.55 to 2.02 g/dL and from 342 to 177 × 10.e3/µL, respectively, during the procedure. CONCLUSION: The increased melphalan extraction efficiency of the new filter is expected to decrease systemic melphalan exposure. In addition, the low transfilter pressure gradient resulted in low resistance to blood flow in the GEN 2 filter, and the changes to blood components are expected to be clinically manageable.

Determining the phagocytic activity of clinical antibody samples.

Antibody-driven phagocytosis is induced via the engagement of Fc receptors on professional phagocytes, and can contribute to both clearance as well as pathology of disease. While the properties of the variable domains of antibodies have long been considered critical to in vivo function, the ability of antibodies to recruit innate immune cells via their Fc domains has become increasingly appreciated as a major factor in their efficacy, both in the setting of recombinant monoclonal antibody therapy, as well as in the course of natural infection or vaccination(1-3). Importantly, despite its nomenclature as a constant domain, the antibody Fc domain does not have constant function, and is strongly modulated by IgG subclass (IgG1-4) and glycosylation at Asparagine 297(4-6). Thus, this method to study functional differences of antigen-specific antibodies in clinical samples will facilitate correlation of the phagocytic potential of antibodies to disease state, susceptibility to infection, progression, or clinical outcome. Furthermore, this effector function is particularly important in light of the documented ability of antibodies to enhance infection by providing pathogens access into host cells via Fc receptor-driven phagocytosis(7). Additionally, there is some evidence that phagocytic uptake of immune complexes can impact the Th1/Th2 polarization of the immune response(8). Here, we describe an assay designed to detect differences in antibody-induced phagocytosis, which may be caused by differential IgG subclass, glycan structure at Asn297, as well as the ability to form immune complexes of antigen-specific antibodies in a high-throughput fashion. To this end, 1 µm fluorescent beads are coated with antigen, then incubated with clinical antibody samples, generating fluorescent antigen specific immune complexes. These antibody-opsonized beads are then incubated with a monocytic cell line expressing multiple FcγRs, including both inhibitory and activating. Assay output can include phagocytic activity, cytokine secretion, and patterns of FcγRs usage, and are determined in a standardized manner, making this a highly useful system for parsing differences in this antibody-dependent effector function in both infection and vaccine-mediated protection(9).