Role of pentamer complex-specific and IgG subclass 3 antibodies in HCMV hyperimmunoglobulin and standard intravenous IgG preparations.

BACKGROUND: HCMV hyperimmunoglobulin-preparations (HIG) contain high concentrations of HCMV-specific IgG. The reduced maternofetal-HCMV-transmission rate of IgG may be due to HCMV-specific neutralizing antibodies against the HCMV pentameric complex (PC). In contrast to HIG, standard intravenous immunoglobulin (IVIG) may have more neutralization (NT) capacity than HIG due to higher IgG subclass 3 levels (Planitzer et al., 2011). METHODS: We investigated the HCMV-specific NT-capacity of HIG Cytotect®, using a recombinant pentameric complex (gHgLUL128-131A) for specific antibody-depletion. We used a modified UL130-peptide (TANQNPSPPWSKLTYSKPH) based on original-sequence of Saccoccio et al. (Vaccine 29(15):2705-2711, 2011) (SWSTLTANQNPSPPWSKLTY) as neutralization target. Both UL130-peptides and the PC were bound via sixfold HisTag and anti-HisTag mAbs to magnetic beads to deplete HCMV-specific IgGs from HIG (Cytotect®). Modifying this depletion strategy, we analyzed the role of IgG subclass 3 in both HIG and IVIG. RESULTS: After CMV IgG-normalization of HIG and IVIG, we found a significant trend towards a decrease (16%) of neutralization-capacity for the UL130 TAN-peptide, but not for the original UL130 SWS-peptide. However, highly significant loss of NT-capacity could be only observed by PC depletion (42%). The IgG subclass 3 depletion revealed no significant reduction of NT-capacity in both HIG and IVIG. CONCLUSION: Via specific antibody depletion, we could demonstrate that pentameric complex-specific antibodies are present in HIG and bind to the recombinant PC resulting in a highly significant reduction of NT-capacity compared to the UL130 TAN-and SWS-peptides. We could not confirm the functional role of IgG subclass 3 neutralizing antibodies in IgG-preparations.

Comparison of cytomegalovirus (CMV)-specific neutralization capacity of hyperimmunoglobulin (HIG) versus standard intravenous immunoglobulin (IVIG) preparations: Impact of CMV IgG normalization.

BACKGROUND: Based on a non-randomized study of Nigro et al. (2005) the intravenous administration of hyperimmunoglobulins (HIGs) is applied frequently to women with primary CMV-infection as "off-label use" in Germany. OBJECTIVES: In order to describe their CMV-specific neutralization-capacity in vitro, we analyzed the HIG preparations Cytotect®, and Cytogam® as well as the standard intravenous immunoglobulins (IVIG) Octagam®, Gamunex®, Kiovig®. STUDY DESIGN: We performed short-term cell-free CMV neutralization assays (CFNT) and long-term cell-adapted neutralization-plaque-reduction assays (PRANT). Human retinal epithelial cells (ARPE-19) were used as target cells. A clinical CMV primary-isolate from amnion fluid propagated in epithelial cells without any initial fibroblast adaption was used. For calibration we previously generated serum-pools (N=100) from two cohorts of mothers at birth: seronegative and latently CMV-infected mothers. Biochemical analysis included total protein, albumin, Ig-class, and IgG-subclasses. Additionally, CMV antibody-reactivity was checked using recombinant immunoblotting. RESULTS: HIG and IVIG preparations showed differences in levels and patterns of protein, Ig-class and CMV-specific antibody concentrations. All IgG-preparations showed high in vitro NT-capacity and high IgG-avidity. The NT90-values for HIGs and IVIGs and our seropositive reference-pool showed similar NT-capacity at a dilution of (1:100) which corresponded well to 4.1 PEI-Units/ml. CONCLUSION: All HIG- and IVIG-preparations showed similar NT-capacity following CMV IgG-normalization. Our in vitro results are in strong contrast to former findings suggesting higher functional CMV NT titers in IVIG-preparations compared to HIGs.