Intravenous IgG Therapy Research Articles

Successful management of severe Kell alloimmunization in pregnancy with intravenous immune globulin

Hemolytic Disease of the Fetus and Newborn (HDFN) is a condition that affects 1 to 2 out of 1000 patients during pregnancy (1). When an alloantibody is present, it is essential to identify its nature in order to organize appropriate follow-up. Kell-mediated HDFN is rare; it occurs in about 5% of Kell alloimmunized pregnant women. It is important to note that in case of anti-Kell immunization, the severity of HDFN is not correlated with maternal antibody titers, and anemia tends to occur earlier and more severely. Therefore, early diagnosing and management of this condition is crucial. In the management of severe fetal anemia due to Kell immunization, available treatments include in utero transfusion (IUT), immunoglobulin therapy. Other alternative treatments exist, such as plasmapheresis. Intravenous immunoglobulin (IVIG), a noninvasive therapeutic approach, acts through multiple mechanisms. IVIG has been evaluated in cases of RhD immunization with high maternal antibody titers and a history of pregnancies involving early hydrops or intrauterine death. Regarding the potential benefits of intravenous IgG therapy, it may delay the need for early IUT, reduce the overall reliance on IUT, and have a positive impact on obstetric outcomes. This case of IV IgG therapy of anti-Kell immunization offers a thought-provoking avenue for future exploration.

Real-World Usage of Hyaluronidase-Facilitated Subcutaneous Immunoglobulin 10% in Patients with Multiple Myeloma Diagnosed with Secondary Immunodeficiency Disease

Introduction: In patients with mature B-cell malignancies such as multiple myeloma (MM), both the underlying disease and immunosuppressive therapy can result in patients developing secondary immunodeficiency disease (SID). SID often manifests in severe, recurrent, or persistent infections that can impair a patient's quality of life and are a major cause of morbidity and mortality in patients with MM, thereby resulting in significant clinical and socioeconomic burden. The use of immunoglobulin (Ig) replacement therapy is well established for the treatment of hypogammaglobulinemia in patients with primary immunodeficiency disease (PIDD). Hyaluronidase-facilitated subcutaneous immunoglobulin (fSCIG) 10% is a dual-vial unit of IgG 10% and recombinant human hyaluronidase (rHuPH20). fSCIG 10% is approved in the USA for the treatment of adults and children aged 2 years or older with PIDD, and in the EU for adults and children of any age with PIDD or SID. fSCIG 10% is not fully integrated into standard of care for patients with SID in many countries, in part owing to comparatively less familiarity of hematologists and oncologists with the evidence supporting the use of fSCIG 10% than for intravenous IgG therapy in patients with SID. Therefore, the HyMMy study (NCT05879757) aims to generate real-world evidence to gain a better understanding of patterns of fSCIG 10% use and outcomes in patients with MM and SID. Methods: HyMMy is a prospective, noninterventional, observational study that will be conducted at 30 centers in Europe and South America. It is planned for 100 patients to be enrolled and observed over a 12-month follow-up period. The overall duration of the study will be 38 months. Patients will be eligible for inclusion if they are aged 18 years or older, have a diagnosis of MM, and fulfill the diagnosis criteria for SID: patient suffers from severe, recurrent, or persistent infection despite appropriate anti-infective treatment, and has proven specific antibody failure or a serum IgG trough level of less than 4 g/L (excluding paraprotein). Patients may enter the study after being diagnosed with SID and no more than 30 days after fSCIG 10% treatment initiation, or with no more than 2 doses of fSCIG 10% already infused ( Figure). Patients newly starting fSCIG 10% have a 30-day window from enrollment to inclusion. Patients will be excluded if they have PIDD, have received Ig treatment or prophylaxis within 3 months of enrollment, have an ongoing serious infection requiring intravenous antimicrobial therapy, have a history of malignancy other than MM within 3 years of enrollment, or have had major surgery within 2 weeks of enrollment. The primary endpoint is to characterize the real-world infusion parameters of fSCIG 10% administration (dosing and administration characteristics; treatment interval; infusion volume, sites, rate, and duration; training visits; and fSCIG 10% discontinuation, interruption, or switch to other treatment). Secondary endpoints will examine the disease course and clinical management of MM (including overall survival and healthcare resource utilization) in patients treated with fSCIG 10%. The exploratory endpoints include: burden of infection; infection rate, type, duration, and severity; patient-reported outcomes pertaining to health-related quality of life, collected via questionnaire at the patients' visit; and overall physician assessment of fSCIG 10% utilization, tolerability, and effectiveness, as measured by a 5-point Likert scale questionnaire. Results: Patient recruitment started in June 2023. Interim analysis is planned after enrollment of the first 50 patients, and the estimated end of the study is 2026. Conclusions: HyMMy is the first prospective, observational study that focuses on the assessment of fSCIG 10% utilization patterns in adults with MM diagnosed with SID. The study will help to improve understanding of the treatment patterns and options available for patients with SID and hematologic malignancies, in particular MM. Takeda Development Center Americas, Inc. funded this study. Takeda Pharmaceuticals International AG funded writing support.

A Systematic Review and Meta-Regression Analysis on the Impact of Increasing IgG Trough Level on Infection Rates in Primary Immunodeficiency Patients on Intravenous IgG Therapy

To determine if increasing IgG trough levels was associated with decreased rates of infection in patients with primary immunodeficiency (PID) who were treated with intravenous immunoglobulin (IVIG).This systematic review included data from 1218 patients with PID who were treated with IVIG from 28 studies between 2001 and 2018. Both children and adults were included in the analysis.This was a systematic review of studies in PubMed and Cochrane and meta-regression analysis in which researchers evaluated immunoglobulin G (IgG) trough levels and their impact on clinical outcomes.The mean IgG trough levels in this analysis ranged from 660 mg/dL to 1280 mg/dL. The mean IVIG dose ranged from 387 mg/kg to 560 mg/kg dosed every 3-4 weeks (the majority were on every 4 week dosing). The most common PIDs reported in this study were common variable immunodeficiency and X-linked agammaglobulinemia (XLA). By using random-effects linear regression slope, IgG trough levels increased by 73 mg/dL for every IVIG dose increase of 100 mg/kg per 4 weeks. Increasing trough level up to a maximum trough of 1280 mg/dL significantly decreased hospitalization (P = .02) but did not significantly decrease infection rates (P = .21) or missed school days or workdays (P = .08). A break point IgG trough level of 960 mg/dL was identified to significantly decrease infection rates (P = .018), and, for >960 mg/dL, there was no significant decrease in infection rates.In patients with PID who were treated with IVIG, increasing IgG trough levels up to 960 mg/dL was associated with improved clinical outcomes and a decreased rate of infection.Goal IgG trough levels have not been clearly established for patients with PID treated with IVIG therapy. In this study, the authors found that increasing IgG trough levels up to a trough of 960 mg/dL decreases the rate of infection. Increasing trough levels >960 mg/dL is significantly associated with decreased hospitalization rates but not with decreased infections or missed school days or workdays. Given that there is a wide range of IVIG dosing used in the treatment of PID, this study is helpful and offers trough targets on the basis of desired effect (eg, reducing infection rates versus reducing hospitalization rates). One limitation of this study is the inability to differentiate between various disease states, such as XLA, which may necessitate a higher IgG trough. Additionally, all studies were cohort studies, which increases the heterogeneity of the population studied.

Maternal and fetal safety of intravenous immunoglobulin in women with reproductive failure.

Intravenous immunoglobulin G (IVIG) is an emerging regimen for women with reproductive failures (RF) during- or pre-pregnancy who have aberrant cellular immune reactions. Studies investigating teratogenicity of IVIG have been limited. Herein, we evaluated the fetal teratogenicity of IVIG and IVIG-related obstetric complications. Women who used IVIG during pregnancy due to RF with cellular immune aberrances were enrolled from four medical centers in Korea. The pregnancy outcomes were collected. A total of 370 RF women who used IVIG during their pregnancy were enrolled. Most of the patients started the IVIG therapy before 12weeks of gestation and 229 women continued IVIG treatment beyond 12weeks of gestation. The mean age of the subjects was 34.8 years and the mean total dosage of IVIG was 125.3g. A total of 307 women had livebirths and six of them were twins. Of 301 singleton livebirths, obstetric complications were developed as follows: preterm births (12.0%), gestational diabetes (7.0%), preeclampsia (4.0%), placental abruption (1.3%), placenta previa (4.3%), and placenta accrete (1.7%). Total six cases (1.99%) had major fetal anomalies in livebirths. The incidence of birth defects is similar to those of the general population in Korea and the previous report in infertile women. No IVIG -related viral contamination was noted. IVIG use during pregnancy did not increase obstetric complications and fetal teratogenicity. This study can be an evidence of maternal and fetal safety of IVIG administration during pregnancy.

High Dose Intravenous IgG Therapy Modulates Multiple NK Cell and T Cell Functions in Patients With Immune Dysregulation.

Intravenous immunoglobulin (IVIG) is an effective immunomodulatory treatment for immune dysregulation diseases. However, the mechanisms by which it reduces systemic inflammation are not well understood. NK cell cytotoxicity is decreased by IVIG in women with reduced fertility, but IVIG effects on NK cells in immune dysregulation are less clear. We hypothesized that IVIG modulation of lymphocyte function, especially in NK cells, is important for resolution of inflammation. Our aim was to identify IVIG-induced changes in a cohort of patients with Kawasaki disease (KD) and those that occur broadly in pediatric patients with various immune dysregulatory diseases. Peripheral blood mononuclear cells (PBMCs) of patients with KD or autoimmune/inflammatory diseases were phenotyped pre and post high dose IVIG treatment by flow cytometry. In KD patients, after IVIG infusion Treg cell frequency and the proportion of activated CD25+ immunoregulatory CD56bright NK cells was increased, and multiple lymphocyte subsets showed increased expression of the lymphoid tissue homing receptor CD62L. Importantly, IVIG treatment decreased the frequency of cells expressing the degranulation marker CD107a among cytotoxic CD56dim NK cells, which was reflected in a significant reduction in target cell killing and in decreased production of multiple pro-inflammatory mediators. Interestingly, the activating receptor CD336 was expressed on a higher proportion of CD56bright NK cells after IVIG in both KD and autoimmune/inflammatory patients while other NK receptors were increased differentially in each cohort. In autoimmune/inflammatory patients IVIG induced the proliferation marker CD71 on a higher percentage of CD56dim NK cells, and in contrast to KD patients, CD107a+ cells were increased in this subset. Furthermore, when PBMCs were stimulated ex vivo with IL-2 or Candida antigen in autologous plasma, more of the CD4+ T cells of KD patients expressed CD25 after IVIG therapy but fewer cytotoxic T cells were degranulated based on CD107a expression. In summary, IVIG treatment in patients with immune dysregulation has multiple effects, especially on NK cell subsets and CD4+ T cells, which are compatible with promoting resolution of inflammation. These novel findings provide insight into the immunomodulatory actions of IVIG in autoimmune and inflammatory conditions.

Identifying Downregulation of Autophagy Markers in Kawasaki Disease.

Kawasaki disease (KD) is the most common cause of heart disease acquired in childhood. Even if treated with high-dose intravenous immunoglobulin G (IVIG) at the early stage; children are still at risk of developing coronary artery lesions. Accumulating evidence suggests that autophagy is enhanced in various heart diseases. Evaluating the pathogenic role of autophagy in KD and coronary artery lesions (CAL) may aid in identifying a potential therapeutic target for the treatment or prevention of the disease. Blood samples were obtained from 20 children with KD at the onset of disease and 21 days after IVIG therapy. Twenty children with other causes of febrile disease and 20 healthy children were included as controls. Total RNA was extracted from white blood cells; and autophagy-related gene mRNA expression levels were measured using real-time polymerase chain reaction. The patients with KD had downregulated levels of LC3B mRNA (0.50 ± 0.06 vs. 1.67 ± 0.15; p < 0.001), BECN 1 mRNA (0.70 ± 0.08 vs. 1.43 ± 0.23; p < 0.05), and ATG16L1 mRNA (0.28 ± 0.04 vs. 0.96 ± 0.16; p < 0.01) compared to the febrile control group. The values of these parameters all increased significantly 21 days after the IVIG therapy as follows: LC3B mRNA (1.77 ± 0.29 vs. 0.50 ± 0.06; p < 0.001), BECN 1 mRNA (1.67 ± 0.36 vs. 0.70 ± 0.08; p < 0.05), and ATG16L1 mRNA (2.96 ± 0.43 vs. 0.28 ± 0.04; p < 0.001), while the level of ATG16L1 mRNA persists low in KD patients with CAL. Our results showed the autophagy-related genes expressions in KD and their change after IVIG administration. This suggests that autophagy may have a protective effect on KD.

Dissecting the mechanism of action of intravenous immunoglobulin in human autoimmune disease: Lessons from therapeutic modalities targeting Fcγ receptors

Since the first description of the administration of high doses of pooled serum IgG, also referred to as intravenous IgG (IVIg) therapy, as being able to ameliorate various autoimmune diseases, researchers have been investigating which molecular and cellular pathways underlie IVIg activity. Apart from trying to understand the obvious conundrum that IgG can trigger both autoimmune pathology and resolution of inflammation, the rapidly expanding use of IVIg has led to a lack of availability of this primary blood product, providing a strong rationale for developing recombinant alternatives. During the last decade, a tremendous number of novel insights into IVIg activity brought the goal of replacing IVIg within reach, at least in select indications, and has led to the initiation of several clinical trials. At the forefront of this effort is the modulation of autoantibody half-life and blocking access of autoantibodies to fragment cystallizable γ receptors (Fcγ receptors). In this rostrum article, we will briefly discuss current models of IVIg activity, followed by a more specific focus on novel therapeutic avenues that are entering the clinic and may replace IVIg in the future.

Intravenous and subcutaneous immunoglobulin G replacement therapy.

Human polyclonal immunoglobulin G (IgG) for therapeutic use has been available for decades. This drug was developed for treatment of antibody deficiency (replacement therapy), although its use has expanded into many anti-inflammatory and immunomodulatory applications in recent years. This review focuses on IgG prescribing for replacement therapy. IgG for replacement is most often administered via the intravenous IgG (IVIG) or subcutaneous IgG (SCIG) routes. IVIG is usually administered every 34 weeks, and SCIG is usually administered weekly, although variations may be considered in all cases. Recently, a new product became available that uses hyaluronidase to facilitate absorption of large doses of SCIG less frequently (every 34 weeks, as with IVIG). There are important differences between the pharmacokinetics of these three routes of administration. IVIG therapy leads to high peaks and low troughs between infusions. IgG concentration fluctuates much less over time with SCIG. Hyaluronidase-facilitated SCIG is intermediate. SCIG may have lower bioavailability in comparison with IVIG and may require higher doses over time; this is not true for hyaluronidase SCIG. However, there are large variations in IgG half-life among individuals and with different products. Therefore, individualization of therapy is essential. Mild systemic flu-like adverse effects may affect up to 2025% of patients who receive IVIG, smaller fractions may experience more-severe symptoms, whereas anaphylaxis is exceedingly rare. General flu-like systemic adverse effects are minimal with SCIG (intermediate with hyaluronidase SCIG), but transient (24 hours), mild, local inflammatory symptoms at infusion sites are relatively common with both forms. Additional rare but important complications of IgG therapy include thrombotic events and hemolysis that can be seen at high doses with any route of administration. Renal adverse effects may occur with IVIG as well. The variety of IgG products and routes of administration available today creates many opportunities for physicians to work with patients to find the optimal therapy for all.

Functional antibodies to Haemophilus influenzae type B, Neisseria meningitidis, and Streptococcus pneumoniae contained in intravenous immunoglobulin products

Intravenous immunoglobulin G (IVIG) replacement therapy is used to prevent invasive infections in patients with primary antibody deficiency (PAD). However, few studies have functionally evaluated specific antibodies against encapsulated bacteria that cause invasive infection in patients with PAD. In this study, functional antibodies against Haemophilus influenzae type b (Hib), Streptococcus pneumoniae (pneumococci), and Neisseria meningitidis (meningococci) in IVIG therapy were evaluated. Sixteen lots of IVIG products prepared by two Korean manufacturers (Products A and B) were evaluated. The functional antibodies were measured by serum bactericidal assay for Hib and four meningococcal serogroups and by multiplexed opsonophagocytic assay for 26 pneumococcal serotypes. The estimated trough levels of antibodies against Hib, pneumococcus, and meningococcus were calculated to determine whether the usual IVIG dose is appropriate for protecting patients with PAD. The functional antibody levels for Hib were similar in all of the IVIG products. In contrast, serum bacterial indices of meningococcal serogroups A and Y showed significant differences between products A and B. Opsonic indices to pneumococci varied depending on the serotype in each IVIG product. The estimated trough levels of antibodies against Hib, pneumococcus, and meningococcus exceeded the protective levels in most of the IVIG products except for the antibodies against two pneumococcal serotypes. Most of the tested commercial IVIG products had sufficient functional antibodies against Hib, pneumococcus, and meningococcus to protect patients with PAD receiving IVIG treatment. Regular and continuous evaluation of IVIG products is necessary to maintain an optimal therapeutic effect.

Inversion of the Vδ1 to Vδ2 γδ T cell ratio in CVID is not restored by IVIg and is associated with immune activation and exhaustion.

Common variable immunodeficiency (CVID) is defined by low levels of IgG and IgA, but perturbations in T cells are also commonly found. However, there is limited information on γδ T cells in CVID patients. Newly diagnosed CVID patients (n = 15) were enrolled before and after intravenous IgG (IVIg) replacement therapy. Cryopreserved peripheral blood mononuclear cells were then used to study γδ T cells and CVID patients were compared to healthy controls (n = 22). The frequency and absolute count of Vδ1 γδ T cells was found to be increased in CVID (median 0.60% vs 2.64%, P <0.01 and 7.5 vs 39, P <0.01 respectively), while they were decreased for Vδ2 γδ T cells (median, 2.36% vs 0.74%, P <0.01 and 37.8 vs 13.9, P <0.01 respectively) resulting in an inversion of the Vδ1 to Vδ2 ratio (0.24 vs 1.4, P <0.001). Markers of immune activation were elevated on all subsets of γδ T cells, and HLA-DR expression was associated with an expansion of Vδ1 γδ T cells (r = 0.73, P = 0.003). Elevated PD-1 expression was found only on Vδ2 γδ T cells (median 1.15% vs 3.08%, P <0.001) and was associated with the decrease of Vδ2 γδ T cells (r = -0.67, P = 0.007). IVIg had no effect on the frequency of Vδ1 and Vδ2 γδ T cells or HLA-DR expression, but alleviated CD38 expression on Vδ1 γδ T cells (median MFI 965 vs 736, P <0.05). These findings suggest that immunological perturbations of γδ T cells are a general feature associated with CVID and are only partially reversed by IVIg therapy.

Pathways Responsible for Human Autoantibody and Therapeutic Intravenous IgG Activity in Humanized Mice.

Immunoglobulin G (IgG) antibodies are major drivers of autoimmune pathology, but they are also used in the form of intravenous IgG (IVIg) therapy to suppress autoantibody activity. To identify the pathways underlying human autoantibody and IVIg activity, we established a humanized mouse model of an autoantibody-dependent autoimmune disease responding to treatment with IVIg preparations. We show that the human IgG subclass strongly impacts autoantibody activity and that the Fc-receptor genotype of the human donor immune system further modulates autoantibody activity. Human mononuclear phagocytes were responsible for autoantibody activity, and IVIg therapy was able to suppress disease pathology in an Fc-fragment-dependent manner. While highly sialylated IgG glycovariants were essential for IVIg activity, it was independent of the Fc-receptor genotype and did not result in a general block of activating or the neonatal Fc-receptor. These findings may help in the development of strategies to block autoantibody and enhance therapeutic IVIg activity in humans.

Potentiation of cytokine-induced proliferation of human Natural Killer cells by intravenous immunoglobulin G

Intravenous IgG (IVIG) therapy can be used for immunomodulation. IL-2 is an immunoregulatory cytokine. We evaluated IVIG modulation of human blood lymphocyte response to IL-2 and other cytokines. Neither IVIG nor low concentrations of IL-2 (3–30U/ml) induced lymphocyte proliferation, but in combination they synergistically enhanced proliferation of NK cells. The CD56bright cells expanded more than CD56dim NK cells, with 90% of NK cells dividing up to 8 generations by day 6, while <8% of T cells divided. IVIG also potentiated NK cell proliferation with IL-12, IL-15 and IL-18. The IVIG+cytokine-expanded NK cells were less cytotoxic for K562 cells, than NK cells with cytokine alone. IVIG also enhanced interferon-γ production with IL-2, IL-12 and IL-18. In conclusion, IVIG selectively potentiates NK cell proliferation and interferon-γ secretion with IL-2, IL-12, IL-15 and IL-18 in vitro. These findings warrant evaluation in vivo in relation to NK cells and the immunoregulatory actions of IVIG.

Molecular and immunological biomarkers to predict IVIg response

Some patients with autoimmune and inflammatory diseases treated with intravenous immunoglobulin G (IVIg) as a first line therapy are refractory. Identification of predictive biomarker(s) to segregate responders and non-responders to IVIg therapy remains critical. A number of biomarkers, particularly in Kawasaki disease, have shown potential for predicting response to IVIg.

Platelet Desialylation: A Novel Mechanism of Fc-Independent Platelet Clearance and a Potential Diagnostic Biomarker and Therapeutic Target in immune Thrombocytopenia

Background:Immune thrombocytopenia (ITP) is a common bleeding disorder caused primarily by autoantibodies against platelet GPIIbIIIa (70-80%) and/or GPIb-complex (20-40%). Current theory suggests antibody-mediated platelet destruction occurs in the spleen, via macrophages through Fc-FcγR interactions. However, evidence from us and others demonstrated that anti-GPIbα, but not anti-GPIIbIIIa, can induce thrombocytopenia via an Fc-independent pathway, which is resistant to intravenous IgG (IVIG) therapy in murine ITP models (Blood 2006) and subsequent IVIG studies in human ITP patients, including our recent large patient cohort study (JTH 2014). This suggests that binding of anti-GPIbα antibodies may induce platelet clearance through a presently unidentified mechanism different than that of anti-GPIIbIIIa.Methods: We developed unique mouse anti-mouse monoclonal antibodies (mAbs) in GPIIIa-/- or GPIba-/- mice, which also recognize GPIbα and GPIIbIIIa of different species including human. Flow cytometry, immunofluorescence, and western blotting were used to evaluate whether these mAbs induced platelet activation, neuraminidase-1 translocation and desialylation of the heavily glycosylated GPIbα in the presence of sialidase inhibitor N-acetyl-2,3-dehydro-2-deoxy neuraminic acid (DANA). These experiments were repeated with human platelets and human ITP patient plasma.We further investigated the effects of anti-GPIbα antibodies on platelet activation, desialylation and clearance in vivo; BALB/c mice were injected with anti-GPIbα or anti-GPIIbIIIa mAbs and following, platelet activation and desialylation were measured by flow cytometry. Hepatocytic Ashwell-Morell receptor (AMR) mediated anti-GPIbα platelet clearance in the liver was examined using immunohistochemistry or blocking the AMR with asialofetuin in both wild-type and macrophage depleted mice. Therapeutic administration of DANA in a murine ITP model assessed the significance of Fc-independent anti-GPIbα mediated platelet clearance in ITP.Results and Discussion: We found that anti-GPIbα, but not anti-GPIIbIIIa antibodies, induced significant P-selectin expression, JON/A binding, neuraminidase-1 translocation and desialylation in murine platelets. Interestingly, certain human platelets were activated (P-selectin expression) and desialylated in the presence of both anti-GPIbα and anti-GPIIbIIIa mAbs or ITP patient plasma. However, we demonstrate that the anti-GPIIbIIIa antibody mediated platelet effects are dependent on the FcγRIIa present exclusively on human platelets as FcγRII blocker IV.3 completely attenuated the response. In contrast, IV.3 had little effect on anti-GPIbα mediated platelet activation or desialylation. Anti-GPIbα Fab fragments and platelet signal pathway inhibitors demonstrate that anti-GPIbα mediated platelet activation and desialylation are consequences of GPIbα cross linking and are reinforced by a positive feedback loop. In vivo, we found significant increases in P-selectin and desialylation in anti-GPIbα injected mice, independent of IgG subclass. A significant role for the hepatic AMR in the clearance of deglycosylated platelets was observed; particularly in macrophage depleted mice whereby, although anti-GPIIbIIIa mediated platelet clearance was completely attenuated, anti-GPIbα mediated platelets clearance still occurred, but was completely rescued with asialofetuin. Immunohistochemistry revealed significant co-localization of anti-GPIbα opsonized platelets with AMR. These suggest the AMR is the dominant Fc-independent anti-GPIbα mediated platelet clearance pathway in the absence of macrophages. Remarkably, sialidase inhibitor DANA ameliorated anti-GPIbα mediated thrombocytopenia in mice. Thus, we demonstrate for the first time that anti-GPIbα antibodies induce platelet activation leading to GPIbα desialyation and platelet clearance via a novel Fc-independent pathway: the hepatic AMR. Our data also suggested that some anti-GPIIbIIIa autoantibodies in human patients may also induce platelet activation and desialylation via the platelet FcR signaling pathway. These findings may lead to novel therapeutic regimens including designating desialylation as a potential diagnostic biomarker and therapeutic target in the treatment of both anti-GPIIbIIIa and anti-GPIbα mediated and/or refractory ITP. DisclosuresNo relevant conflicts of interest to declare.

Association of autoantibody specificity and response to intravenous immunoglobulin G therapy in immune thrombocytopenia: a multicenter cohort study

Immune thrombocytopenia (ITP) is a common autoimmune bleeding disorder, in which platelet glycoprotein (GP)IIb-IIIa and GPIb-IX are the two most frequently targeted autoantigens. Our previous studies in animal models of ITP demonstrated that intravenous immunoglobulinG (IVIG) could protect against anti-GPIIb-IIIa autoantibody-mediated thrombocytopenia but failed to ameliorate ITP induced by most anti-GPIb-IX autoantibodies. The objective of this human study was to evaluate the association between the specificity of antiplatelet autoantibodies and response to IVIG treatment. In this retrospective study, a cohort of 156 previously untreated adults with severe ITP who underwent IVIG therapy (0.4gkg(-1) day(-1) for 5days) was analyzed. The primary outcome was response defined as platelet counts of ≥30×10(9) L(-1) and a doubling of baseline counts within 7days of dosing, and an absence of bleeding. Among the 66 patients with anti-GPIb-IX autoantibodies, only 24 (36.4%) achieved a response, as compared with 72 of 90 patients (80.0%) who were negative for anti-GPIb-IX autoantibodies (relative risk 2.2; 95% confidence interval 1.6-3.1). This study found no difference in response between patients with anti-GPIIb-IIIa autoantibodies (61.7%) and those without anti-GPIIb-IIIa autoantibodies (61.3%). Logistic regressions, including main effects and the interaction between these two autoantibodies, showed no influence of anti-GPIIb-IIIa autoantibodies on the effects of anti-GPIb-IX autoantibodies with regard to their association with IVIG response. Thus, in adults with ITP, the presence of anti-GPIb-IX autoantibodies is a predictive factor for poor response to IVIG treatment. ClinicalTrials.gov NCT01666795.

Desialylation: A Novel Platelet Clearance Mechanism and a Potential New Therapeutic Target in Anti-GPIb Antibody Mediated Thrombocytopenia

AbstractAbstract 265 Background:Immune thrombocytopenia (ITP) is an autoimmune disease characterized by autoantibodies directed at patient’s own platelet antigens, primarily glycoprotein (GP)IIbIIIa-integrin (70–80%) and GPIb-complex (20–40%). Current paradigm suggests that clearance of opsonized platelets through the reticuloendothelial system via Fcγ-receptors results in thrombocytopenia and bleeding disorders. However, evidence from others and our group demonstrated that anti-GPIbα, but not anti-GPIIbIIIa, can induce thrombocytopenia via an Fc-independent pathway, which is resistant to intravenous IgG (IVIG) therapy in murine ITP-models (Blood 2006). These observations are consistent with subsequent IVIG studies in human ITP patients. Interestingly, human anti-GPIb-mediated ITP patients seem also resistant to steroid therapy in our recent retrospective study (American Journal of Hematology 2012). This suggests that binding of anti-GPIbα antibodies may induce platelet clearance through a different mechanism which is currently poorly understood. Methods:We developed unique mouse anti-mouse monoclonal antibodies (mAbs) in GPIIIa or GPIba deficient mice. Some of the mAbs have cross-reactivity to both mouse and human GPIIbIIIa and GPIba. Flow cytometry was used to evaluate whether these mAbs were able to induce platelet activation, apoptosis and desialylation. GPIbα is heavily glycosylated and the role of desialylation and exposure of underlying galactose and β-N-acetyl-D-glucosamine (βGN) residues on GPIbα in platelet clearance was assessed using the sialidase neuraminidase (NA) and it’s inhibitor N-acetyl-2,3-dehydro-2-deoxy neuraminic acid (DANA). Desialyation effects on platelet activation and apoptosis was measured by flow cytometry. We also repeated these experiments with human platelets and plasma from human ITP-patients.We also investigated the effects of anti-GPIbα antibodies on platelet activation, apoptosis and clearance in vivo. Briefly, BALB/c mice were injected with anti-GPIbαor anti-GPIIIa mAbs and 24 hrs later, platelet desialylation, activation and apoptosis were measured by flow cytometry. The effect of desialylation on platelet clearance was assessed with DANA. The possible roles of Ashwell-Morell and MAC-1 receptors in GPIbα-mediated platelet clearance in the liver were examined using immunohistochemistry (anti-CD11b) or blocking of the Ashwell-Morell receptor with asialofetuin. Results and Discussion:We found that anti-GPIbα, but not anti-GPIIbIIIa mAbs, induced significant P-selectin expression and phosphatidylserine (PS)-exposure, and increased inner membrane mitochondrial depolarization (ΔYm). Interestingly, platelets were desialylated in the presence of anti-GPIbα but not anti-GPIIbIIIa mAbs. Moreover, we found that desialylation of GPIbα lies directly upstream of platelet activation and apoptosis, as prior treatment with DANA diminished PS-exposure, and P-selectin expression. Most importantly, incubations of human platelets with ITP-patient plasma showed similar effects. In vivo, we found significant increases in PS-exposure and ΔYm induced by anti-GPIbα, but not by anti-GPIIIa mAbs, independent of IgG subclass. Interestingly, prior injection with DANA rescued platelets numbers in anti-GPIbα, but not in anti-GPIIIa injected mice. A significant role for the Ashwell-Morell and MAC-1 receptors in the clearance of deglycosylated platelets was observed; blocking of the Ashwell-Morell receptors by asialofetuin, decreased platelet clearance in anti-GPIbα, but not anti-GPIIbIIIa antibody injected mice, there was also increased staining for MAC-1 on Kupffer cells, exclusively in the presence of an anti-GPIbα mAb tested. Thus, we demonstrate for the first time that anti-GPIbα antibodies induce GPIbα desialyation, leading to platelet activation and apoptosis. Therefore, we identified novel Fc-independent platelet clearance pathways, more specifically, via Ashwell-Morell and MAC-1 receptors on hepatocytes and liver macrophages. These findings may lead to novel therapeutic regimens including the potential use of sialidase inhibitors as a solution for anti-GPIb-mediated ITP patients previously refractory to both steroid and IVIG therapies. Disclosures:No relevant conflicts of interest to declare.

Intravenous Immunoglobulin G Treatment in ABO Hemolytic Disease of the Newborn, is it Myth or Real?

Intravenous Immunoglobulin G (IVIG) therapy has been used as a component of the treatment of hemolytic disease of the newborn. There is still no consensus on its use in ABO hemolytic disease of the newborn routinely. The aim of this study is to determine whether administration of IVIG to newborns with ABO incompatibility is necessary. One hundred and seventeen patients with ABO hemolytic disease and positive Coombs test were enrolled into the study. The subjects were healthy except jaundice. Infants were divided into two groups: Group I (n=71) received one dose of IVIG (1g/kg) and LED phototherapy whereas Group II (n=46) received only LED phototherapy. One patient received erythrocyte transfusion in Group I, no exchange transfusion was performed in both groups. Mean duration of phototherapy was 3.1±1.3days in Group I and 2.27±0.7days in Group II (p<0.05). Mean duration of hospital stay was 5.34±2.2days in Group I and 3.53±1.3days in Group II (p<0.05). Mean duration of phototherapy was 4.0±1.5days and 2.73±1.1days in double and single doses of IVIG respectively, and this was statistically significant (p<0.05). IVIG therapy didn't decrease neither phototherapy nor hospitalization duration in infants with ABO hemolytic disease. Meticulus follow-up of infants with ABO hemolytic disease and LED phototherapy decreases morbidity. IVIG failed to show preventing hemolysis in ABO hemolytic disease.

IVIg‐mediated amelioration of ITP in mice is dependent on sialic acid and SIGNR1

Intravenous immunoglobulin G (IVIg) therapy is widely used to treat autoimmune and inflammatory diseases. Recent evidence suggests that in mice, splenic resident cells might be important for the anti-inflammatory activity of IVIg in a model of serum transfer arthritis. Splenectomized human immunothrombocytopenia (ITP) patients, however, still respond to IVIg therapy. To investigate whether the requirement of the spleen is essential for mouse ITP, we used a passive model of induced ITP and demonstrated that IVIg activity was functional in splenectomized animals. Further analysis showed that the IVIg-mediated amelioration of platelet phagocytosis was fully dependent on terminal sialic acid residues in the IVIg preparation and could be blocked with a specific ICAM3 grabbing nonintegrin-related 1 (SIGNR1) specific antibody. These results suggest that, similar to the human system, a spleen-independent but sialic acid- and SIGNR1-dependent pathway is responsible for IVIg-mediated suppression of autoantibody-dependent platelet depletion in mice.

Choices in IgG replacement therapy for primary immune deficiency diseases

Subcutaneous IgG (SCIG) is becoming more popular, but there is still uncertainty about efficacy and optimal dosing. This review discusses recent pharmacokinetic studies and applications of SCIG therapy, and its efficacy in the context of emerging understanding of the relationship between dosing and efficacy of both intravenous IgG (IVIG) and SCIG replacement therapy for primary immunodeficiency diseases. Three preparations of IgG have been licensed in the US in the past year. Their bioavailabilities are 65-70% of that of IVIG. Pooled analyses show that the efficacy of SCIG in preventing infections is proportional to the steady-state levels achieved, and similar to that of IVIG. Pharmacokinetic studies allow estimation of doses that will yield desired serum levels with both IVIG and SCIG, and when switching from one route to another. Pooled analyses show that at equivalent total doses, weekly SCIG results in steady-state levels 10-20% higher than troughs on monthly IVIG. For most patients, the choice between routes should be based on individual preference, and the regimen should be individualized to achieve the desired outcomes.

881 In Infants with An Initial Diagnosis of Transient Hypogammaglobulinemia of Infancy (THI), is the Use of IV Immunoglobulin Warranted?

Introduction: THI is a defect in a child's endogenous immunoglobulin production. It presents early following the physiological catabolism of the remnants of maternal immunoglobulin, transferred trans-placentally during the course of pregnancy. Physiological hypogammaglobulinemia normalises by 2-4 months. However during prolonged THI; the infant is rendered susceptible to a range of bacterial infections. In severe cases intravenous immunoglobulin-G (IV-IgG) replacement therapy has been proposed as treatment. Objectives: To analyse current experimental literature in order to assess the suitability of IVIgG therapy for patients diagnosed with suspected THI. Assessment of therapeutic use of IV-IgG from retrospective case controls. Further qualification of when to use IV-IgG by assessment of additional factors: symptomatology, time to normalisation of immunoglobulins and comparison with other immunodeficiency syndromes. Methods: PubMed - searched with MeSH term “hypogammaglobulinemia”; “transient hypogammaglobulinemia infancy”, “hypogammaglobulinemia IVIg”, “paediatric hypogammaglobulinaemia”. Results: Antibiotics are first-line treatment for recurrent infections. IV-IgG replacement therapy is not routinely warranted for symptomatic THI. Cases where infections are severe/resistant to conventional management, IV-IgG therapy can be considered. Other primary immunodeficiencies requiring IVIG therapy present similarly to THI as recurrent infections. In contrast THI is self-resolving with a benign clinical progression if managed appropriately; Conclusion: Most children with THI do not require IV-IgG, with the exception of severe life-threatening infections or ongoing bacterial respiratory infection despite prophylactic antibiotic therapy. In these circumstances trials of IV-IgG for 6-12 months are indicated. There are no published studies that comprehensively assess the cost-effectiveness of IV-IgG replacement in any healthcare setting for THI.