Reduction In Antibody Titers Research Articles

Immune Tolerance Induced Using Plasma Exchange and Rituximab in an Infantile Pompe Disease Patient

Infantile Pompe disease, resulting from deficiency of lysosomal acid α-glucosidase, requires enzyme replacement therapy with recombinant human acid α-glucosidase. Most patients develop antirecombinant human acid α-glucosidase antibodies, leading to reduced response to enzyme therapy in a subgroup of them. Aiming to improve treatment response, several immune tolerance induction strategies have been explored. We describe a patient with life-threatening infusion-associated reactions presenting anti-recombinant human acid α-glucosidase antibodies. He was successfully treated with an immune tolerance induction protocol, consisting of plasma exchange combined with a single dose of rituximab. Immediate reduction of antibody titer was obtained and enzyme therapy was resumed without infusion-associated reactions. Twenty-two months later, immunoglobulin G titer remained below 1:100. In conclusion, we applied a short-course immune tolerance induction strategy in a patient with severe infusion-associated reactions and anti-recombinant human acid α-glucosidase antibodies, leading to early and persisting reduction of antibody titer, in the absence of significant adverse events.

Neonatal antibody responses are attenuated by interferon-γ produced by NK and T cells during RSV infection

Respiratory syncytial virus (RSV) infects most children in the first year of life and is a major single cause of hospitalization in infants and young children. There is no effective vaccine, and antibody generated by primary neonatal infection is poorly protective against reinfection even with antigenically homologous viral strains. Studying the immunological basis of these observations in neonatal mice, we found that antibody responses to infection were low and unaffected by CD4 depletion, in contrast with adult mice, which had stronger CD4-dependent antibody responses. Natural killer cell depletion or codepletion of CD4(+) and CD8(+) cells during neonatal RSV infection caused a striking increase in anti-RSV antibody titer. These cells are major sources of the cytokine IFN-γ, and blocking IFN-γ also enhanced RSV-specific antibody responses in neonates. In addition, infection with a recombinant RSV engineered to produce IFN-γ reduced antibody titer, confirming that IFN-γ plays a pivotal role in inhibition of antibody responses after neonatal infection. These unexpected findings show that the induction of a strong cellular immune response may limit antibody responses in early life and that vaccines that induce IFN-γ-secreting cells might, in some situations, be less protective than those that do not.

ABO-Incompatible kidney transplantation

HLA sensitization and ABO incompatibility continue to pose a significant barrier to expansion of living donation. In fact, either anti-blood or anti-donor HLA antibodies result in the occurrence of hyperacute rejection and graft loss. Reducing this early rejection risk by planned desensitization protocols has clearly improved the outcome of ABO-incompatible (ABOi) kidney transplantation. B-cell depletive therapy has replaced splenectomy, overcoming the disadvantages of the latter. Plasma exchange techniques have considerably reduced antibody titers, allowing better results. Thus, newer immunosuppressive protocols reduced early graft loss and early rejections episodes and, consequently, improved the long-term graft survival. Therefore, ABOi kidney transplantation can be more broadly practiced, especially to expand the pool donor and to reduce the waiting time for transplantation.

Alloimmunosensitization in Left Ventricular Assist Device Recipients and Impact on Posttransplantation Outcome

Left ventricular assist devices (LVADs) have become an established surgical therapy for patients with end-stage heart failure who require hemodynamic support as a bridge-to-transplant or destination therapy. However, the anatomic and physiologic consequences of long-term LVAD support have yet to be fully clarified. Despite the clinical success of these devices, it has been reported that many patients bridged to transplantation with mechanical support develop circulating antibodies with potential donor reactivity. Transplanting against existing or historic donor-specific antibodies is associated with increased risk of antibody-mediated rejection, graft dysfunction, and decreased survival. Safe transplantation of allosensitized patients is dependent on using prospective crossmatching and antibody titer reduction techniques (desensitization). Strict protocols requiring a negative prospective crossmatch before transplantation result in a decreased donor pool and a longer duration of support in sensitized LVAD recipients with increased inherent morbidity such as infections and thromboembolic complications. The aim of this review is to present the current state of knowledge of possible immunologic mechanisms involved in alloimmunization of LVAD recipients, outline new methods of antibody detection, compare various desensitization strategies, and present an overview of clinical data assessing the impact of sensitization on posttransplantation outcome.

Evidence that inflammation associated with Sarcocystis neurona infection plays a role in clinical signs associated with equine protozoal myeloencephalitis

Sarcocystis neurona is a protozoal pathogen that causes equine protozoal myeloencephalitis (EPM). The commonly recognized neurologic signs that EPM causes are due to the pathological effects of the disease and can be due to infections of somatic and neural tissues by the asexual stages of S. neurona that cause cell dysfunction and death. A second component of EPM is the inflammatory host response to the parasitic insult. Inflammation may be as deleterious to the host as the initial insult. Anticoccidial drugs are necessary to treat the infectious aspect of EPM. An objective evaluation of treatment effect on protozoa is measured by a reduction in SnSAG 1, 5, and 6 antibodies. Recurrences of neurologic signs in horses that have diminishing antibody titers was controlled by levamisole HCl and indicate inflammation is a significant component of disease. There are no specific markers of inflammation due to S. neurona, therefore clinical exam was used to determine effectiveness of treatment. In a clinical field trial horses with a presumptive diagnosis of EPM based on veterinary clinical exam and the presence of serum antibodies against recombinant SAG’s 1, 5, and 6 by ELISA (> 1:16) were treated with 0.5 mg/kg decoquinate and 1 mg/ kg levamisole HCl PO for 10 days. A post-treatment clinical examination and antibody titer was determined. Horses that improved and then experienced recurrence of neurologic signs were treated with levamisole HCl PO until the horse resolved the clinical signs. Ninety-four percent of the EPM suspect horses resolved neurological signs within the 10 day treatment period based on clinical exam. Eighty-nine percent showed a reduction in antibody titers at 4-5 weeks and 97% showed a reduction in antibody titers by 12 weeks post treatment. Eleven percent of horses responded incompletely to decoquinate/ levamisole or responded completely but experienced recurrence from 7 to 14 days following treatment. Horses that experienced recurrence of signs responded to levamisole HCl and maintained the improvement after

Distribution of ABO Blood Group Antibody Titers in Pediatric Patients Awaiting Renal Transplantation

Blood group-incompatible transplantation is one strategy used when a potential recipient does not have a compatible living donor. Current practice includes desensitization strategies to reduce antibody titers. However, when antibodies are low, in cardiac transplantation in neonates for example, no desensitization is required. This study is the first to examine the distribution of ABO blood group antibody titers in a population of pediatric patients on the deceased-donor renal transplantation waiting list. All patients from two pediatric nephrology centers active on the national deceased-donor waiting list had antibody titers (total immunoglobulin load) measured. A simulation modeling the effect of allocating blood group-incompatible deceased-donor kidneys to those patients with titers of 16 or lower was developed. Twenty-four children were screened; eight (33.3%) had titers of either anti-A or anti-B antibodies of 8 or lower. A further three (12.5%) had either an anti-A or anti-B antibody titer of 16. Blood group A or B patients had lower antibody levels than blood group O patients. In blood group O patients, levels of anti-A antibodies were higher than anti-B antibodies (Wilcoxon signed rank test, P=0.028). The simulation model showed that a change in organ allocation policy would increase pediatric transplant activity by 2.2% and reduce the median waiting time for a transplant. This allocation strategy may be of particular benefit to those pediatric patients who have been on the deceased-donor waiting list for a long time or those with a high calculated reaction frequency.

IgY stability in eggs stored at room temperature or at +4°C

1. The aim of this study was to investigate the stability of immunoglobulin-Y (IgY) in stored eggs from immunised hens. 2. Eggs from individual hens were randomised and stored for up to one month at room temperature, or for up to 6 months at +4°C. IgY was extracted from the egg yolks and the antibody activities were tested by ELISA. 3. There was no significant reduction in antibody titres with egg storage under these conditions. 4. Egg yolks of immunised chickens provide an inexpensive source of large amounts of polyclonal antibodies for use in immunotherapy and immunoassays. By collecting eggs from different immunised hens and pooling their yolks, it should be possible to reduce batch-to-batch variation.

B Cell Strategy to Maintain Remission in ANCA-associated Vasculitides?

During the past decades therapeutic regimens in the anti-neutrophil cytoplasmic autoantibodies (ANCA)-associated vasculitides (AAV) aiming at induction or maintenance of remission have undergone substantial changes. Glucocorticoids plus cyclophosphamide as standard therapy for induction have been challenged by monoclonal tumor necrosis factor blocking antibodies; and maintenance therapy with azathioprine has been expanded by methotrexate, leflunomide, and mycophenolate mofetil1,2. Regarding pathogenesis, numerous recent studies have suggested a central role of the ANCA3. In line with this hypothesis, plasma exchange aiming at a reduction of antibody titer showed positive results in life-threatening vasculitis4. Along this line, rituximab (RTX), a chimeric monoclonal anti-CD20 antibody primarily used for therapy of B cell lymphomas, proved to be highly effective in the induction of remission, successfully challenging cyclophosphamide5,6,7. With the US Food and Drug Administration approval of RTX for a single course treatment in both granulomatosis with polyangiitis (GPA; Wegener’s granulomatosis) and microscopic polyangiitis (MPA) in April of 2011, a novel induction regimen was established8. Notably, RTX is the only treatment approved so far. Disease relapses after induction — irrespective of the agent — … Address correspondence to Dr. Villiger. E-mail: Peter.Villiger{at}Insel.ch

Early prediction of instability of chinese hamster ovary cell lines expressing recombinant antibodies and antibody‐fusion proteins

One of the most important criteria for the successful manufacture of a therapeutic protein (e.g., an antibody) is to develop a mammalian cell line that maintains stability of production. Problems with process yield, lack of effective use of costly resources, and a possible delay in obtaining regulatory approval of the product may ensue otherwise. Therefore the stability of expression in a number of Chinese hamster ovary (CHO) derived production cell lines that were isolated using the glutamine synthetase (GS) selection system was investigated by defining a culture as unstable if the titer (which is a measure of productivity) of a cell line expressing an antibody or antibody-fusion protein declined by 20-30% or more as it underwent 55 population doublings. Using this criterion, a significant proportion of the GS-selected CHO production cell lines were observed to be unstable. Reduced antibody titers correlated with the gradual appearance of a secondary, less productive population of cells as detected with flow cytometric analysis of intracellular antibody content. Where tested, it was observed that the secondary population arose spontaneously from the parental population following multiple passages, which suggested inherent clonal instability. Moreover, the frequency of unstable clones decreased significantly if the host cell line from which the candidate production cell lines were derived was apoptotic-resistant. This data suggested that unstable cell lines were more prone to apoptosis, which was confirmed by the fact that unstable cell lines had higher levels of Annexin V and caspase 3 activities. This knowledge has been used to develop screening protocols that identify unstable CHO production cell lines at an early stage of the cell line development process, potentially reducing the cost of biotherapeutic development.

Response to hepatitis B vaccine differs by birthweight among neonates

ObjectivesTo assess the immunogenicity of same needle length of hepatitis B vaccine for infants with different birthweights. Previous studies have focused on obese adolescents and adults that have lower titers of antibody response to hepatitis B vaccine. The aim of this study was to compare antibody titers of macrosomic and other neonates for hepatitis B antibody with the same needle length. Methods96 healthy infants were vaccinated at birth, 1, and 6 months of age with hepatitis B vaccine, with follow-up to 7 months of age. Cord blood and seventh month hepatitis B antibody titer were repeated. Infants were divided into three groups, according to their birthweight, as macrosomic, appropriate for gestational age and small for gestational age. The same needle with 26G and 16mm length was used for all infants. ResultsMacrosomic infants with a birthweight over 4000g had lower antibody levels than small and appropriate for gestational age infants. ConclusionsThese data support the hypothesis that birthweight has an effect on response to HBV vaccine. Using the same needle length for immunization of macrosomic infants for hepatitis B reveals significantly reduced antibody titers, this can be related also to possible endogenous factors. Further studies are needed to assess appropriate needle length and effects of other factors on vaccine reactogenicity for macrosomic infants.

Advances in basic and clinical immunology in 2010

Reports in basic and clinical immunology in 2010 reflected the use of state-of-the-art genetic and immunologic tools to characterize the pathogenesis of immunologic diseases and the development of novel therapies directed to these conditions. B-cell biology has been explained in greater detail, significantly with lessons from the genetic defects found in the humoral immunodeficiencies. Therapeutic mAbs are given for an increasing number of indications, such as anti-CD20 antibodies or rituximab, which was initially developed for non-Hodgkin lymphomas and is currently used in diverse autoimmune and inflammatory disorders. The report of an infant with severe combined immunodeficiency (SCID) in Massachusetts detected by means of newborn screening and successfully treated with hematopoietic stem cell transplantation validated recent efforts toward newborn screening for SCID. Improvement of survival outcomes for patients with primary immunodeficiencies treated with hematopoietic stem cell transplantation was demonstrated in a large European cohort, with significant appreciation of the type of donor graft, particularly the use of HLA-matched unrelated donors for patients with non-SCID. Progress in cellular mechanisms of drug hypersensitivity included the characterization of nitroso-modified drug metabolites as potent T-cell activators and the identification of the relocation of plasmacytoid dendritic cells from blood to skin as a potential risk factor for reactivation of viral disease.

Effect of vaccination of boars against porcine circovirus type 2 on ejaculate characteristics, serum antibody titers, viremia, and semen virus shedding1

The objective of this research was to determine the effect of vaccination against porcine circovirus type 2 (PCV2) on ejaculate characteristics, PCV2-specific antibody titers in serum, viremia, and viral shedding in the semen of PCV2-positive boars. Before vaccination, all boars were confirmed by PCR to be naturally infected with PCV2. The boars were vaccinated with a commercial killed vaccine against PCV2 (n = 5) or served as controls and received 2 mL of 0.9% saline (n = 5). Semen and blood samples were collected before vaccination at wk 0 and at 7-d intervals thereafter until wk 8. Sperm concentration and characteristics of sperm motility were assessed using a computer-assisted sperm analysis system, and sperm morphology was evaluated using light microscopy after staining. The PCV2 antibody titers were determined in serum using an ELISA, and the genomic copy numbers of PCV2 DNA in serum and semen were determined by real-time PCR. In general, there were no effects of treatment or treatment × week on semen or sperm characteristics (P > 0.10). An effect of treatment × week was detected for serum antibody titers (P < 0.01). Compared with controls, PCV2 antibody titers in vaccinated boars were less (P < 0.01) at wk 7 (1.01 ± 0.05 titer/mL vs. 1.23 ± 0.05 titer/mL) and tended (P = 0.07) to be less at wk 8 (1.05 ± 0.05 titer/mL vs. 1.17 ± 0.05 titer/mL). There were no effects of treatment or treatment × week for serum and semen genomic copy numbers of PCV2 DNA (P > 0.10). There was a tendency (P = 0.09) for an effect of week on serum viral load. It was evident that during this experiment, boars experienced reoccurring PCV2 infection, and the detection of an increased PCV2 DNA load in serum preceded that in semen; the duration of reoccurring infection appeared to be less in vaccinated boars compared with controls. In summary, vaccination against PCV2 can reduce antibody titers when given postinfection and has no dramatic effect on indicators of semen quality. Vaccination against PCV2 in naturally infected boars can also decrease the length of reoccurring infection and decrease the duration of viral shedding in semen.

Rituximab reduces anti-cardiolipin levels in patients with systemic lupus erythematosus

Patients with systemic lupus erythematosus (SLE), can produce antibodies directed against the complex phospholipds-β-glycoprotein 1. Rituximab is an antibody directed to most B-cells, except precursors and mature plasma cells. It can produce reductions in antibody titers. We have treated SLE patients refractory to conventional immunosuppressive therapy, with Rituximab and we searched for a reduction in anticardiolipin autoantibodies.

The kinetics of donor HLA class I-specific antibody absorption following a combined split liver and kidney transplant

Hyperacute rejection of a transplanted liver is rare even when the recipient has circulating donor-specific alloantibodies (DSA). There is also evidence that a transplanted liver may provide immunological protection for other organs transplanted from the same donor. We monitored the kinetics of circulating DSA in a highly sensitized recipient of a combined split liver and kidney transplant and demonstrated a reduction in antibody titres immediately after liver perfusion. The absorption of DSA was not compromised by the smaller liver mass transplanted. DSA titres remained low at 3 months post-transplant, and the recipient did not experience antibody-mediated rejection.

CD4+FOXP3+ regulatory T cells confer long-term regulation of factor VIII–specific immune responses in plasmid-mediated gene therapy–treated hemophilia mice

AbstractGene transfer of a factor VIII (FVIII) plasmid into hemophilia A (HemA) mice achieved supraphysiologic FVIII expression, but triggered production of high-titer FVIII-specific antibodies and loss of functional FVIII activity. To test whether FVIII-specific regulatory T cells (Tregs) can modulate immune responses against FVIII, we developed a HemA mouse model in which all T cells overexpressed Foxp3 (HemA/Foxp3-Tg). FVIII plasmid therapy did not induce antibody production in HemA/Foxp3-Tg mice. CD4+Foxp3+ T cells isolated from plasmid-treated HemA/Foxp3-Tg mice significantly suppressed proliferation of FVIII-stimulated CD4+ effector T cells. The percentage of CD4+ T cells expressing CD25, glucocorticoid-induced tumor necrosis factor receptor, and cytotoxic T lymphocyte antigen 4 increased significantly in spleen and peripheral blood for 9 weeks. Mice receiving adoptively transferred Tregs from FVIII-exposed HemA/Foxp3-Tg mice produced significantly reduced antibody titers compared with controls after initial challenge with FVIII plasmid and second challenge 16 weeks after first plasmid treatment. Adoptively transferred Tregs engrafted and distributed at 2% to 4% in the Treg compartment of blood, lymph nodes, and spleens of the recipient mice and induced activation of endogenous Tregs; the engraftment decreased to negligible levels over 8 to 12 weeks. Antigen-specific Tregs can provide long-lasting protection against immune responses in vivo and limit recall responses induced by a second challenge via infectious tolerance.

Antibody production: Low dose immunogen vs. low incorporation hapten using salmeterol as a model

Haptens are low molecular weight compounds that are non-immunogenic and so must be conjugated to carrier molecules to elicit an immune response. Doses of 50–1000 μg protein conjugate have been suggested for immunisation of rabbits with hapten–protein immunogens. Although larger doses may give a faster response, lower doses may result in higher affinity antibodies. The amount of hapten presented to the host's immune system can be controlled by varying either the amount of immunogen administered or the quantity coupled to a fixed amount of protein. This study compares the two approaches for the production of antibodies to the β-agonist salmeterol as a model. A range of salmeterol–HSA conjugates was prepared, with varying molar ratios of hapten:protein (80:1, 15:1, 7.5:1, 4:1), for use as immunogens. The 80:1 immunogen was administered to different animals at four concentrations (0.5, 0.2, 0.1 and 0.05 mg dose −1) while the remaining three immunogens were administered at 0.5 mg dose −1. The effects of immunogen dose and hapten incorporation on the titre and affinity of the antibodies produced to salmeterol were investigated. It was found that both approaches resulted in the production of more sensitive antibodies, although reducing the degree of hapten incorporation brought about a larger reduction in antibody titre. Reducing the degree of hapten incorporation also produced a more consistent pattern of results regarding antibody sensitivity (after the sixth immunisation in this study) which may make it easier to predict the most suitable time for antibody harvesting.

Immunogenicity of the outer domain of a HIV-1 clade C gp120

BackgroundThe possibility that a sub domain of a C clade HIV-1 gp120 could act as an effective immunogen was investigated. To do this, the outer domain (OD) of gp120CN54 was expressed and characterized in a construct marked by a re-introduced conformational epitope for MAb 2G12. The expressed sequence showed efficient epitope retention on the isolated ODCN54 suggesting authentic folding. To facilitate purification and subsequent immunogenicity ODCN54 was fused to the Fc domain of human IgG1. Mice were immunised with the resulting fusion proteins and also with gp120CN54-Fc and gp120 alone.ResultsFusion to Fc was found to stimulate antibody titre and Fc tagged ODCN54 was substantially more immunogenic than non-tagged gp120. Immunogenicity appeared the result of Fc facilitated antigen processing as immunisation with an Fc domain mutant that reduced binding to the FcR lead to a reduction in antibody titre when compared to the parental sequence. The breadth of the antibody response was assessed by serum reaction with five overlapping fragments of gp120CN54 expressed as GST fusion proteins in bacteria. A predominant anti-inner domain and anti-V3C3 response was observed following immunisation with gp120CN54-Fc and an anti-V3C3 response to the ODCN54-Fc fusion.ConclusionThe outer domain of gp120CN54 is correctly folded following expression as a C terminal fusion protein. Immunogenicity is substantial when targeted to antigen presenting cells but shows V3 dominance in the polyvalent response. The gp120 outer domain has potential as a candidate vaccine component.

Mucosal Delivery of a Pneumococcal Vaccine UsingLactococcus lactisAffords Protection against Respiratory Infection

Economical and effective vaccines against Streptococcus pneumoniae (pneumococcus) are needed for implementation in poorer countries where the disease burden is highest. Here, we evaluated Lactococcus lactis intracellularly producing the pneumococcal surface protein A (PspA) as a mucosal vaccine in conferring protection against pneumococcal disease. Mice were intranasally (inl) immunized with the lactococcal vaccine. Control groups were also immunized with similar amounts of recombinant PspA administered inl or subcutaneously with alum. PspA-specific antibodies in serum samples and lung lavage fluids were measured before challenge in intraperitoneal sepsis and inl respiratory-infection models of pneumococcal disease. The lactococcal vaccine afforded better protection against respiratory challenge with pneumococcus than did vaccination with purified antigen given inl or by injection with alum. This finding was associated with a shift toward a Th1-mediated immune response characterized by reduced antibody titers to the PspA antigen. In the sepsis model, the lactococcal vaccine afforded resistance to disease on a par with that obtained with the injected vaccine, demonstrating its efficacy against different forms of pneumococcal disease. Given the safety profile of L. lactis, there is considerable potential to develop a pneumococcal vaccine for use in humans and to broaden this approach to combat other major pathogens.

Effect of AHCC on the immune response to primary influenza infection in young and aged mice

The emergence of H5N1 avian influenza and the threat of new or adapted viruses in bioterrorism have created an urgent interest in identifying agents to enhance the immune response to primary virus infection. Influenza (FLU) virus is a public health concern, particularly among the elderly, and is well-studied in relation to changes that occur with age. Aged mice exhibit reduced antibody titers, impaired CTL responses, and increased virus burden after primary infection with FLU. Our data indicate that natural killer (NK) cell activity is also reduced by 50% in both the spleen and lungs of aged mice, while the number and percent of total lymphocytes do not differ with age in response to infection. Active hexose correlated compound (AHCC) is an agent reported to increase NK cell activity, survival, and bacterial clearance in young mice stressed by head-down tilt, although no data currently exist on the effects of AHCC on the response to FLU infection. C57Bl/6 mice were supplemented with 1g AHCC/kg body weight/day for 1 week prior to and throughout infection with influenza A (H1N1, PR8). Spleen and lung lymphocytes were assessed for NK cytotoxicity by 51Cr release assay and phenotype by flow cytometry, and lung viral clearance by MDCK infectivity. AHCC appears to increase NK activity in the spleen and enhance NK cell recruitment to the lung by 48 hours following FLU infection. Bolstering innate immunity with dietary bioactives may be one avenue for improving the immune response to primary FLU infection. Supported by Amino Up Chemical Company

Heart transplantation across the antibodies against HLA and ABO

We have intentionally performed heart transplantation in a 5-year-old child, despite the most unfavourable risk factors for patient survival; the presence of high level of antibodies against donor's human leucocyte antigen (HLA) class I/II and blood group antigens. Pretransplant treatment by mycophenolate mofetil, prednisolone, tacrolimus, intravenous immunoglobulin, rituximab, protein-A immunoadsorption (IA) and plasma exchange reduced antibody titres against the donor's lymphocytes from 128 to 16 and against the donor's blood group antigen from 256 to 0. The patient was urgently transplanted with a heart from an ABO incompatible donor (A(1) to O). A standard triple-drug immunosuppressive protocol was used. No hyperacute rejection was seen. Antibodies against the donor's HLA antigens remained at a low level despite three acute rejections. Rising anti-A(1) blood group antibodies preceded the second rejection and were reduced by two blood group-specific IAs and remained at a low level. The patient is doing well despite the persistence of donor-reactive antibodies.