scholarly journals Design and Evaluation of a Multiplexed Assay to Assess Human Immunogenicity Against Humira®

2020 ◽  
Vol 22 (5) ◽  
Author(s):  
Matthew Alleyn ◽  
Kristin Closson ◽  
Adam Gentile ◽  
Nathan Gulbis ◽  
Christopher Taylor ◽  
...  
Keyword(s):  
Immune Responses ◽  
Drug Efficacy ◽  
Clinical Samples ◽  
Screening Assay ◽  
Specific Antibodies ◽  
Tiered Approach ◽  
Clinical Events ◽  
Industry Standards ◽  
Recommended Guidelines ◽  
Immunogenicity Assays

Abstract The use of biologic-based therapeutics has revolutionized our ability to treat complex diseases such as cancer- and autoimmune-related disorders. Biologic-based therapeutics are known to generate anti-drug immune responses or immunogenicity in clinical patients which can lead to altered pharmacokinetics, decreased drug efficacy, and unwanted adverse clinical events. Assays designed to detect and assess anti-drug immune responses are used to help monitor patients and improve drug safety. Utilizing a tiered approach, screening assays are developed first to identify patients that are potentially positive for anti-drug-specific antibodies. Patients that screen positive are subjected to additional tiers of testing that include a confirmation assay to confirm the presence of expected anti-drug-specific antibodies, a titer assay to assess relative levels of anti-drug-specific antibodies, and, depending on the drug’s mechanism of action or concerns of adverse clinical reactions, further characterization such as drug neutralization and anti-drug antibody isotyping. This tiered approach can prove to be detrimental to clinical samples from exposure to multiple cycles of testing, freeze thaws, and repeated handling by lab personnel. Multiplexing some of these assays together may streamline the characterization of anti-drug immune responses and help reduce the repeated usage of clinical samples. In this study, we combined a screening assay and anti-drug isotyping assays into one multiplexed assay using the Luminex® xMAP® Technology. The multiplexed assay was developed and validated to meet the FDA recommended guidelines for immunogenicity assessments. These results show that multiplexed assays perform comparably to industry standards. This study should encourage labs to explore the use of multiplexing immunogenicity assays to characterize anti-drug antibody responses quickly, with less repeat testing and reduced sample handling.

scholarly journals Development and validation of an assay for detection of Japanese Encephalitis virus specific antibody responses

2020 ◽  
Author(s):  
Pradeep Darshana Pushpakumara ◽  
Chandima Jeewandara ◽  
Laksiri Gomes ◽  
Yashodha Perera ◽  
Ananda Wijewickrama ◽  
...  
Keyword(s):  
Immune Responses ◽  
Japanese Encephalitis Virus ◽  
Disease Severity ◽  
Japanese Encephalitis ◽  
Specific Antibody ◽  
Encephalitis Virus ◽  
Antibody Responses ◽  
Specific Antibodies ◽  
Dengue Disease ◽  
Conserved Regions

AbstractBackgroundAlthough immune responses to the Japanese Encephalitis virus (JEV), and the dengue viruses (DENV) have a potential to modulate the immune responses to each other, this has been poorly investigated. Therefore, we developed an ELISA to identify JEV specific, DENV non cross-reactive antibody responses by identifying JEV specific, highly conserved regions of the virus and proceeded to investigate if the presence of JEV specific antibodies associate with dengue disease severity.Methodology/Principal findings20 JEV specific peptides were identified from highly conserved regions of the virus and the immunogenicity and specificity of these peptides were assessed in individuals who were non-immune to JEV and DENV (JEV-DENV-, N=30), those who were only immune to the JEV and not DENV (JEV+DENV-, N=30), those who were only immune to DENV(JEV-DENV+, N=30) and in those who were immune to both viruses (JEV+DENV+, N=30). 7/20 peptides were found to be highly immunogenic and specific and these 7 peptides were used as a pool to further evaluate JEV-specific responses. All 30/30 JEV+DENV-and 30/30 JEV+DENV+individuals, and only 3/30 (10%) JEV-DENV+individuals responded to this pool. We further evaluated this pool of 7 peptides in patients following primary and secondary dengue infection during the convalescent period and found that the JEV-specific peptides, were unlikely to cross react with DENV IgG antibodies. We further compared this in-house ELISA developed with the peptide pool with an existing commercial JEV IgG assay to identify JEV-specific IgG following vaccination, and our in-house ELISA was found to be more sensitive. We then proceeded to investigate if the presence of JEV-specific antibodies were associated with dengue disease severity, and we found that those who had past severe dengue (n=175) were significantly more likely (p<0.0001) to have JEV-specific antibodies than those with past non-severe dengue (n=175) (OR 5.3, 95% CI 3.3 to 8.3).Conclusions/SignificanceAs our data show that this assay is highly sensitive and specific for detection of JEV-specific antibody responses, it would be an important tool to determine how JEV seropositivity modulate dengue immunity and disease severity when undertaking dengue vaccine trials.Author summaryBoth Japanese Encephalitis virus (JEV), and the dengue viruses (DENV) co-circulate in the same geographical region and have a potential to modulate the immune responses to each other. However, due to the difficulty in identifying antibody responses specific to either virus due to the highly cross-reactive nature of virus-specific antibodies, this has been poorly investigated. Therefore, we developed an ELISA to identify JEV-specific, DENV non cross-reactive antibody responses by identifying JEV-specific, highly conserved regions of the virus and proceeded to investigate if the presence of JEV-specific antibodies associates with dengue disease severity. 20 JEV-specific peptides were identified from highly conserved regions of the virus and the immunogenicity and specificity of these peptides were assessed. We found that seven peptides were highly immunogenic and specific to the JEV and we further evaluated the usefulness of an ELISA developed using these pools of peptides. We found that our in-house ELISA was found to be significantly more sensitive some of the existing commercial assays. As this assay appears to be highly sensitive and specific for detection of JEV-specific antibody responses, it would be an important tool to determine how JEV seropositivity modulate dengue immunity and disease severity when undertaking dengue vaccine trials.

scholarly journals Differential antibody dynamics to SARS-CoV-2 infection and vaccination

2021 ◽  
Author(s):  
Yuezhou Chen ◽  
Pei Tong ◽  
Noah B. Whiteman ◽  
Ali Sanjari Moghaddam ◽  
Adam Zuiani ◽  
...  
Keyword(s):  
Immune Responses ◽  
B Cell ◽  
Somatic Mutation ◽  
Post Infection ◽  
Vaccine Strategy ◽  
Specific Antibodies ◽  
Antibody Dynamics ◽  
Induced Immunity ◽  
Original Virus

ABSTRACTOptimal immune responses furnish long-lasting (durable) antibodies protective across dynamically mutating viral variants (broad). To assess robustness of mRNA vaccine-induced immunity, we compared antibody durability and breadth after SARS-CoV-2 infection and vaccination. While vaccination delivered robust initial virus-specific antibodies with some cross-variant coverage, pre-variant SARS-CoV-2 infection-induced antibodies, while modest in magnitude, showed highly stable long-term antibody dynamics. Vaccination after infection induced maximal antibody magnitudes with enhanced longitudinal stability while infection-naïve vaccinee antibodies fell with time to post-infection-alone levels. The composition of antibody neutralizing activity to variant relative to original virus also differed between groups, with infection-induced antibodies demonstrating greater relative breadth. Differential antibody durability trajectories favored COVID-19-recovered subjects with dual memory B cell features of greater early antibody somatic mutation and cross-coronavirus reactivity. By illuminating an infection-mediated antibody breadth advantage and an anti-SARS-CoV-2 antibody durability-enhancing function conferred by recalled immunity, these findings may serve as guides for ongoing vaccine strategy improvement.

Rhamnose modified bovine serum albumin as a carrier protein promotes the immune response against sTn antigen

2020 ◽  
Vol 56 (90) ◽  
pp. 13959-13962
Author(s):  
Han Lin ◽  
Haofei Hong ◽  
Jinfeng Wang ◽  
Chen Li ◽  
Zhifang Zhou ◽  
...  
Keyword(s):  
Immune Response ◽  
Bovine Serum Albumin ◽  
Immune Responses ◽  
Serum Albumin ◽  
Cancer Vaccine ◽  
Bovine Serum ◽  
Vaccine Development ◽  
Carrier Protein ◽  
Antigen Uptake ◽  
Specific Antibodies

Rhamnose and sTn antigen were co-conjugated to bovine serum albumin (BSA) for cancer vaccine development. The immune responses against sTn have been significantly augmented with the involvement of Rha-specific antibodies to enhance antigen uptake.

Analysis of Spontaneous Vs. Vaccine-Induced Antibody Responses Against Cancer-Testis Antigen MAGE-A3 in Cancer Patients

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 5087-5087
Author(s):  
Yanran Cao ◽  
Sacha Gnjatic ◽  
Vincent G. Brichard ◽  
Tim Luetkens ◽  
Sebastian Kobold ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
Cancer Patients ◽  
Peripheral Blood ◽  
Memory B Cells ◽  
Antibody Responses ◽  
Specific Antibodies ◽  
Fine Specificity ◽  
Specific Memory ◽  
Cancer Testis

Abstract Abstract 5087 Background: Cancer-testis antigens (CTA) are attractive targets for cancer immunotherapy based on their tumor-restricted expression and immunogenicity. A number of CTA, including Melanoma-associated antigen 3 (MAGE-A3), are already under clinical investigation and CTA have been shown to induce strong T cell and humoral immunity in cancer patients receiving active immunotherapy. However, little is known about the fine specificity and the function of vaccine-induced humoral immune responses and it is unclear how they relate to spontaneous CTA-specific immune responses occurring in a minority of patients. Methods: We have performed a longitudinal analysis of spontaneously occurring antibody responses against the CT antigen MAGE-A3 in sera (N=1537), which were collected from patients with multiple myeloma (N=355) over a period of 6 years. Antibody titers were determined by ELISA technique and a B cell ELISPOT assay was applied to estimate the number of MAGEA3-specific memory B cells in peripheral blood of the patients. Fine specificity of the antibody responses was examined using overlapping 20mer peptides spanning the whole sequence of MAGE-A3. The given IgG subtype was determined, and the quality of MAGE-A3-specific antibodies was analyzed using western blot as well as affinity assays. Results were compared to those obtained with MAGE-A3-specific antibody responses induced by vaccination with full-length MAGE-A3 protein and adjuvants AS02B or AS15 in patients with non-small cell lung cancer (NSCLC; N=15). Results: Out of 355 myeloma patients 4 (1.1%) evidenced spontaneous antibody responses against MAGE-A3 at least at one point during the course of their disease. Spontaneously occurring anti-MAGE-A3 humoral responses were usually of low titer. In contrast, all of the vaccinated patients showed high-titered and persisting antibody responses which usually appeared around week 6 after the first application of the vaccine. Accordingly, we found high frequencies of vaccine-induced MAGE-A3-specific memory B cells in the peripheral blood of NSCLC patients while they remained undetectable in most myeloma patients. Vaccine-induced antibody responses underwent affinity maturation reaching affinity levels of spontaneous immune responses after repeated cycles of treatment. MAGE-A3-specific antibodies consisted of IgG1 and IgG3>IgG2>IgG4 subtypes in vaccinated patients whereas spontaneously occurring antibodies were mainly of the IgG2 subtype. Spontaneous as well as vaccine-induced IgG antibodies both recognized the natural full-length protein. Analysis of the fine specificity of the antibody responses revealed that vaccine-induced antibodies recognized a much larger number of MAGE-A3 epitopes than spontaneously occurring antibodies. However, both, spontaneous as well as vaccine-induced responses, most frequently and strongly recognized a specific region within the MAGE-A3 protein corresponding to amino acids 51–70. Conclusions This study demonstrates for the first time important qualitative differences between spontaneously occurring and vaccine-induced antibody responses against the MAGE-A3 antigen in cancer patients. While the potential of both types of antibody responses to promote antigen uptake and induction of T cell responses by antigen-presenting cells might differ, they both recognized the same restricted region within the MAGE-A3 protein. The latter finding might be of importance for the design of future immunotherapies targeting MAGE-A3. Disclosures: No relevant conflicts of interest to declare.

INDICES OF IMMUNE RESPONSES OF BULLS IN THE NECROBACILLOSIS VACCINATION OF THE CATTLE

2015 ◽  
Vol 10 (3) ◽  
pp. 93-97
Author(s):  
Михеева ◽  
Ekaterina Mikheeva ◽  
Бабинцева ◽  
Tatyana Babintseva ◽  
Макаев ◽  
...  
Keyword(s):  
T Lymphocytes ◽  
Immune Responses ◽  
Blood Serum ◽  
B Lymphocytes ◽  
Metabolic Disorders ◽  
Specific Antibodies ◽  
Biological Safety ◽  
Digestive Organs ◽  
Udmurt Republic ◽  
And Control

Metabolic disorders and dysfunctional state of the digestive organs lead to the development of distal extremities diseases. The greatest danger is neсrobacrillosis. Vaccination is one of the ways to prevent and control this disease. The article reflects the data on the effect of vaccination (“Nekovak” and formol-emulsion neсrobacrillosis vaccine of Federal Centre for toxicology, radiation and biological safety) on immunological and serological parameters of blood and blood serum of cattle in the Udmurt Republic. After immunization with “Nekovak” vaccine and formol-emulsion neсrobacrillosis vaccine, we determined an increase of the number of leukocytes to 7th day, and also an increase of T-lymphocytes to 14th day. The number of B-lymphocytes reaches a maximum on 21st day. The content of gamma globulins, including specific antibodies against the neсrobacrillosis pathogen, exceeded the control and reached a maximum on the 21st day after vaccination.

scholarly journals RecombinantLactococcus lactisExpressing Haemagglutinin from a Polish Avian H5N1 Isolate and Its Immunological Effect in Preliminary Animal Trials

2017 ◽  
Vol 2017 ◽  
pp. 1-8 ◽  
Author(s):  
Agnieszka K. Szczepankowska ◽  
Katarzyna Szatraj ◽  
Przemysław Sałański ◽  
Agnieszka Rózga ◽  
Roman K. Górecki ◽  
...  
Keyword(s):  
Immune Responses ◽  
Lactococcus Lactis ◽  
Expression System ◽  
Heterologous Proteins ◽  
Specific Antibodies ◽  
Recombinant Cells ◽  
Animal Trials ◽  
Immunological Effect ◽  
H5n1 Isolate ◽  
Different Origin

Lactic acid bacteria (LAB) are Gram-positive, nonpathogenic microorganisms that are gaining much interest as antigen producers for development of live vaccine vectors. Heterologous proteins of different origin have been successfully expressed in various LAB species, includingLactococcus lactis. RecombinantL. lactisstrains have been shown to induce specific local and systemic immune responses against various antigens. Our study aimed at constructing aL. lactisstrain expressing haemagglutinin of a Polish avian H5H1 influenza isolate and examining its effect on animals. Expression of the clonedH5gene was achieved using the nisin-controlled gene expression system. Detection of the intracellular H5 antigen produced inL. lactiswas performed by Western blot analysis and confirmed using mass spectrometry. The potential ofL. lactisrecombinant cells to induce an immune response was examined by setting up preliminary immunization trials on chickens and mice. Obtained sera were tested for specific antibodies by ELISA assays. The results of these studies are a promising step toward developing a vaccine against the bird flu usingLactococcus lactiscells as bioreactors for efficient antigen production and delivery to the mucosal surface.

scholarly journals Enhancement of Innate and Cell-Mediated Immunity by Antimycobacterial Antibodies

2005 ◽  
Vol 73 (10) ◽  
pp. 6711-6720 ◽  
Author(s):  
S. de Vallière ◽  
G. Abate ◽  
A. Blazevic ◽  
R. M. Heuertz ◽  
D. F. Hoft
Keyword(s):  
T Cells ◽  
Immune Responses ◽  
Fluorescent Protein ◽  
Cell Mediated Immunity ◽  
Phagocytic Cells ◽  
Surface Binding ◽  
Serum Samples ◽  
Specific Antibodies ◽  
Green Fluorescent ◽  
Mycobacterial Growth

ABSTRACT We investigated the ability of human antibodies induced by Mycobacterium bovis bacillus Calmette-Guérin (BCG) vaccination to protect against mycobacterial infections. Serum samples containing mycobacterium-specific antibodies were obtained from volunteers who had received two intradermal BCG vaccinations 6 months apart. Significant increases in lipoarabinomannan (LAM)-specific immunoglobulin G (IgG) were detected after both the primary and booster vaccinations. Effects of mycobacterium-specific antibodies on surface binding and internalization of BCG by neutrophils and monocytes/macrophages were studied, using green fluorescent protein (gfp)-expressing BCG. Surface-bound gfp-expressing BCG were distinguished from intracellular BCG by surface labeling with LAM-specific monoclonal antibody. Internalization of BCG by phagocytic cells was shown to be significantly enhanced in postvaccination serum samples. Furthermore, the inhibitory effects of neutrophils and monocytes/macrophages on mycobacterial growth were significantly enhanced by BCG-induced antibodies. The growth-inhibiting effects of postvaccination sera were reversed by preabsorption of IgG with Protein G. Finally, the helper effects of antimycobacterial antibodies for the induction of cell-mediated immune responses were investigated. BCG-induced antibodies significantly enhanced proliferation and gamma interferon production in mycobacterium-specific CD4+ and CD8+ T cells, as well as the proportion of proliferating and degranulating CD8+ T cells. We conclude that mycobacterium-specific antibodies are capable of enhancing both innate and cell-mediated immune responses to mycobacteria.

scholarly journals Supplemented nutrition increases immunity and drug efficacy in a natural host-helminth system

2019 ◽  
Author(s):  
Amy R. Sweeny ◽  
Melanie Clerc ◽  
Paulina A. Pontifes ◽  
Saudamini Venkatesan ◽  
Simon A. Babayan ◽  
...  
Keyword(s):  
Immune Responses ◽  
Drug Efficacy ◽  
Natural Host ◽  
Chronic Infections ◽  
Gastrointestinal Helminths ◽  
Transmission Potential ◽  
Wood Mice ◽  
Balanced Diet ◽  
Helminth Infections ◽  
The World

AbstractGastrointestinal helminths are common parasites of humans, wildlife, and livestock, leading to chronic infections in large parts of the world. In humans, there is also an overlap in the incidence of malnutrition and helminth infections which can predispose individuals to higher infection burdens and reduced anthelmintic efficacy due to compromised immunity. This relationship has been well-studied in laboratory models by testing for the impact of dramatic reductions of specific nutrients on infection outcomes. However, much less is known about the benefits of whole-diet supplementation in natural host-helminth systems. We experimentally supplemented the diet of wood mice (Apodemus sylvaticus) and measured resistance to the gastrointestinal nematode Heligmosomoides polygyrus and anthelmintic treatment efficacy in both natural and captive populations. Supplemented wood mice were more resistant to H. polygyrus infection, cleared worms more efficiently after treatment, produced stronger general and parasite-specific antibody responses, and maintained better body condition. In addition, supplemented nutrition in conjunction with anthelminthic treatment significantly reduced H. polygyrus transmission potential. These large-scale improvements in condition and immunity of supplemented nutrition found in controlled and wild environments show the rapid and extensive benefits of a well-balanced diet and have important implications for using diet interventions to improve disease control programmes.Author SummaryGastrointestinal helminths are ubiquitous parasites which cause chronic infections and debilitating symptoms worldwide for human and animal populations alike. Efforts to control helminth infections rely primarily on deworming drugs. However, despite high availability and low cost of these drugs, reinfection is common and helminths remain a substantial problem in many areas of the world. One factor which contributes to the persistence of helminth infections is co-occurrence of malnutrition. Extensive work in model laboratory systems has shown that deficiencies in both macro- and micronutrients can impair host immune responses to helminth infections, but how this relationship translates to natural host-helminth systems and drug efficacy is still poorly understood. Here, we used a combination of experimental nutrition supplementation and deworming treatment in paired wood mouse populations in both wild and captive environments to determine the role of a well-balanced diet for infection control in a natural host-helminth system. We found that mice on a supplemented diet had lower burden of infection and increased drug efficacy, body condition, and immune responses. We also found that supplementation lowered transmission potential of hosts. These broad and rapid effects of increased nutrition quality have important implications for the benefits of diet interventions alongside deworming drugs.

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